Megakaryocyte-derived extracellular vesicle biomarkers

Abstract

Disclosed herein are compositions and methods relating to megakaryocyte-derived extracellular vesicles derived from human pluripotent stem cells, where the megakaryocyte-derived extracellular vesicles display unique biomarkers.

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of priority to U.S. Provisional Patent Application Nos. 63 / 278,378, filed November 11, 2021, 63 / 335,181, filed April 26, 2022, and 63 / 380,576, filed October 23, 2022, all of which are incorporated by reference in their entireties.

[0002] The present disclosure relates to biomarkers for megakaryocyte-derived extracellular vesicles and methods of using same. [Background technology]

[0003] To avoid the cytotoxicity and side effects and / or rapid clearance of drugs, various nanovesicles have been developed to administer therapeutic agents.

[0004] The largest category of clinically approved nanoparticles is liposomes, which consist of a simple lipid bilayer surrounding an aqueous compartment. However, because liposomes are exogenous synthetic entities with limited cell or tissue targeting mechanisms, they can cause adverse effects in patients, including immune responses and cytotoxicity, in addition to non-target specificity and inefficient unloading of therapeutic drugs. Adenoviral, retroviral, AAV, and lentiviral vectors are currently the most popular viral vectors used in gene therapy. However, traditional viral vector production methods, which use adherent cell lines and transient transfection in the presence of serum, are not scalable.

[0005] Alternatively, nanovesicles, such as extracellular vesicles, would benefit from a better understanding of their biological and compositional properties.

[0006] Thus, there is a need for compositions and methods that increase the efficiency and are well characterized for cargo packaging and delivery to cells to allow for predictable isolation and standardization. Summary of the Invention

[0007] Disclosed herein are compositions and methods relating to manufactured megakaryocyte-derived extracellular vesicles. Specifically, among other things, the megakaryocyte-derived extracellular vesicles exhibit a unique biomarker profile that makes them well-suited for use in therapeutic delivery and the treatment of various diseases or disorders. In various embodiments, the compositions and methods disclosed herein can be utilized for drug delivery and the treatment of one or more genetic disorders. In several embodiments, the compositions and methods disclosed herein can be utilized for drug delivery and the treatment of infectious diseases. In several embodiments, the compositions and methods disclosed herein can be utilized for drug delivery and the treatment of hematopoietic diseases or disorders, such as thrombocytopenia / anemia. In several embodiments, the compositions and methods disclosed herein can be utilized for drug delivery and the treatment of hemoglobinopathies. The methods disclosed herein can be in vivo or ex vivo and can be used, for example, for gene replacement therapy and gene editing.

[0008] In one aspect, the present invention relates to a composition comprising a plurality of substantially purified megakaryocyte-derived extracellular vesicles derived from human pluripotent stem cells, wherein the composition comprises one or more proteins that are unique to or preferentially present in the megakaryocyte-derived extracellular vesicles compared to other types of extracellular vesicles.

[0009] In one aspect, the present invention relates to a composition comprising a plurality of substantially purified megakaryocyte-derived extracellular vesicles derived from human pluripotent stem cells, the composition comprising one or more nucleic acids encoding one or more proteins that are unique to or preferentially present in the megakaryocyte-derived extracellular vesicles relative to other types of extracellular vesicles.

[0010] In one aspect, the present invention relates to a composition comprising a plurality of substantially purified megakaryocyte-derived extracellular vesicles derived from human pluripotent stem cells, the composition comprising a lipid bilayer membrane surrounding an inner lumen, the inner lumen of the megakaryocyte-derived extracellular vesicles comprising one or more megakaryocyte-derived nucleic acid molecules selected from mRNA, tRNA, rRNA, siRNA, microRNA, regulatory RNA, and non-coding RNA and coding RNA, the lipid bilayer membrane comprising one or more proteins associated with or embedded therein, the one or more proteins being unique to or preferentially present in the megakaryocyte-derived extracellular vesicles compared to other types of extracellular vesicles.

[0011] In one aspect, the present invention relates to a composition comprising a plurality of substantially purified megakaryocyte-derived extracellular vesicles derived from human pluripotent stem cells, the composition comprising a lipid bilayer membrane surrounding a lumen, the lumen of the megakaryocyte-derived extracellular vesicles comprising one or more megakaryocyte-derived nucleic acid molecules selected from mRNA, tRNA, rRNA, siRNA, microRNA, regulatory RNA, and non-coding RNA and coding RNA, the lipid bilayer membrane comprising one or more nucleic acids that encode one or more proteins associated with or embedded within the lipid bilayer membrane, and that are unique to or preferentially present in the megakaryocyte-derived extracellular vesicles compared to other types of extracellular vesicles.

[0012] In one aspect, the present invention relates to a composition comprising a plurality of substantially purified megakaryocyte-derived extracellular vesicles derived from human pluripotent stem cells, wherein the composition comprises one or more nucleic acids that are unique to or preferentially present in the megakaryocyte-derived extracellular vesicles compared to other types of extracellular vesicles.

[0013] In one aspect, the present invention relates to a composition comprising a plurality of substantially purified megakaryocyte-derived extracellular vesicles derived from human pluripotent stem cells, wherein the composition comprises one or more proteins encoded by one or more nucleic acids that are unique to or preferentially present in the megakaryocyte-derived extracellular vesicles compared to other types of extracellular vesicles.

[0014] In another aspect, the invention relates to a pharmaceutical composition comprising a composition disclosed herein and a pharmaceutically acceptable excipient or carrier.

[0015] In another aspect, the present invention relates to a method for delivering a deliverable therapeutic agent, the method comprising: (a) obtaining megakaryocyte-derived extracellular vesicles of a composition disclosed herein; (b) incubating the megakaryocyte-derived extracellular vesicles with a therapeutic agent to import the therapeutic agent into the lumen of the megakaryocyte-derived extracellular vesicles and / or associate with the surface of the megakaryocyte-derived extracellular vesicles, thereby producing a deliverable therapeutic agent; and (c) administering the deliverable therapeutic agent to a patient, or contacting the deliverable therapeutic agent with biological cells in vitro and administering the contacted biological cells to a patient.

[0016] In another aspect, the present invention relates to a method for producing megakaryocyte-derived extracellular vesicles of the compositions disclosed herein, the method comprising: (a) obtaining human pluripotent stem cells, wherein the human pluripotent stem cells are primary CD34+ hematopoietic stem cells sourced from peripheral blood or umbilical cord blood; (b) differentiating the human pluripotent stem cells into megakaryocytes in the absence of erythropoietin and in the presence of thrombopoietin; and (c) isolating megakaryocyte-derived extracellular vesicles from the megakaryocytes.

[0017] In various embodiments, the compositions and methods disclosed herein can be utilized for drug delivery and treatment of one or more genetic disorders.

[0018] In another aspect, the present invention relates to a method for treating or preventing an infectious disease, comprising administering an effective amount of a composition disclosed herein.

[0019] In another aspect, the present invention relates to a method for treating or preventing an infectious disease, comprising administering an effective amount of a composition comprising cells contacted in vitro with a composition disclosed herein.

[0020] In another aspect, the present invention relates to a method for treating a hematopoietic disease or disorder. In one aspect, the present invention relates to a method for treating thrombocytopenia, comprising administering an effective amount of a composition disclosed herein, wherein the composition comprises megakaryocyte-derived extracellular vesicles comprising a nucleic acid encoding a functional thrombocytopenia-associated gene or its protein product, or a nucleic acid encoding a gene-editing protein capable of generating a functional thrombocytopenia-associated gene or its protein product.

[0021] In another aspect, the invention relates to a method for treating thrombocytopenia, comprising administering an effective amount of a composition comprising cells contacted in vitro with a composition disclosed herein, wherein the composition comprises megakaryocyte-derived extracellular vesicles comprising a nucleic acid encoding a functional thrombocytopenia-associated gene or its protein product, or a nucleic acid encoding a gene-editing protein capable of generating a functional thrombocytopenia-associated gene or its protein product.

[0022] In another aspect, the present invention relates to a method for treating a hemoglobinopathy, comprising administering an effective amount of a composition disclosed herein, wherein the composition comprises megakaryocyte-derived extracellular vesicles comprising a nucleic acid encoding a functional hemoglobinopathy-associated gene or its protein product, or a nucleic acid encoding a gene-editing protein capable of generating a functional hemoglobinopathy-associated gene or its protein product.

[0023] In another aspect, the present invention relates to a method for treating a hemoglobinopathy, comprising administering an effective amount of a composition comprising cells contacted in vitro with a composition disclosed herein, wherein the composition comprises megakaryocyte-derived extracellular vesicles comprising a nucleic acid encoding a functional hemoglobinopathy-associated gene or its protein product, or a nucleic acid encoding a gene-editing protein capable of generating a functional hemoglobinopathy-associated gene or its protein product.

[0024] The Abstract, as well as the following Detailed Description, will be better understood when read in conjunction with the accompanying drawings. For the purpose of illustrating the methods of the present disclosure, there are shown in the drawings exemplary embodiments of the methods, but these should not be limited to the particular embodiments disclosed. [Brief explanation of the drawings]

[0025] [Figure 1A] Figure 1A shows a series of pie charts depicting the overlap between the MkEV proteome of the present disclosure and the top 100 exosome marker proteins identified in the database ExoCarta (Keerthikumar et al. J Mol Biol. 2016;428(4):688-692) (World Wide Web exocarta.org). Of these proteins, 96 were identified in the MkEV proteome of the present disclosure, while 4 were not (Figure 1A). As shown in Figure 1B, of the top 100 EV marker proteins defined by the VesiclePedia database (Pathan et al. Nucleic Acids Res. 2019 Jan 8;47:D516-D519) (World Wide Web microvesicles.org), 91 of these proteins were identified in the MkEV proteome of the present disclosure, while 9 were not. In summary, the MkEV of the present disclosure primarily contains common extracellular vesicle and exosome markers. [Figure 1B]Figure 1A shows a series of pie charts depicting the overlap between the MkEV proteome of the present disclosure and the top 100 exosome marker proteins identified in the database ExoCarta (Keerthikumar et al. J Mol Biol. 2016;428(4):688-692) (World Wide Web exocarta.org). Of these proteins, 96 were identified in the MkEV proteome of the present disclosure, while 4 were not (Figure 1A). As shown in Figure 1B, of the top 100 EV marker proteins defined by the VesiclePedia database (Pathan et al. Nucleic Acids Res. 2019 Jan 8;47:D516-D519) (World Wide Web microvesicles.org), 91 of these proteins were identified in the MkEV proteome of the present disclosure, while 9 were not. In summary, the MkEV of the present disclosure primarily contains common extracellular vesicle and exosome markers. [Figure 2A] A series of Venn diagrams showing the overlap of the MkEV proteome of this disclosure with the entire VesiclePedia and ExoCarta databases of human EV proteomes reveals 147 proteins unique to the MkEV of this disclosure (Figure 2A). The overlap of the cell surface proteins of the MkEV of this disclosure with those captured by the VesiclePedia database revealed that 13 cell surface and membrane proteins are unique to the MkEV of this disclosure (Figure 2B). Surface and membrane proteins were defined according to Bausch-Fluck et al. (Bausch-Fluck et al. PLoS One. 10:e0121314) and the Human Protein Atlas (Thul PJ, Åkesson L., Science. 2017. 365(6340)). [Figure 2B]A series of Venn diagrams showing the overlap of the MkEV proteome of this disclosure with the entire VesiclePedia and ExoCarta databases of human EV proteomes reveals 147 proteins unique to the MkEV of this disclosure (Figure 2A). The overlap of the cell surface proteins of the MkEV of this disclosure with those captured by the VesiclePedia database revealed that 13 cell surface and membrane proteins are unique to the MkEV of this disclosure (Figure 2B). Surface and membrane proteins were defined according to Bausch-Fluck et al. (Bausch-Fluck et al. PLoS One. 10:e0121314) and the Human Protein Atlas (Thul PJ, Åkesson L., Science. 2017. 365(6340)). [Figure 3] 1 is a histogram showing KEGG pathway enrichment in unique MkEV proteins of the present disclosure. Shown is a list of significantly enriched KEGG pathways (p-value < 0.05) in unique MkEV proteins of the present disclosure, including pathways involved in aminoacyl-tRNA biosynthesis and neutrophil extracellular trap formation. [Figure 4] Figure 1 shows the direct protein-protein interaction network of unique MkEV proteins. The network was constructed to include curated direct protein-protein interactions (interaction score >0.4). The contour highlighting represents the associated interaction score (confidence of the interaction). Clusters with more than three nodes are highlighted and numbered in descending order. Non-interacting nodes were excluded. Number 1 represents the histone cluster, number 2 represents the tRNA aminoacylation of protein translation cluster, number 3 represents the cluster of septin proteins, number 4 represents the mitochondria-associated cluster, and number 5 represents the cellular biogenesis process (transcription and translation). [Figure 5]Figure 1 shows the functional and physical protein-protein interaction network of unique MkEV proteins. This protein network was constructed using functional associations and direct protein-protein interactions. Experimental and database evidence was included (interaction score >0.4). Clusters with more than three nodes were highlighted and numbered in descending order. Non-interacting nodes were excluded. Number 1 represents the histone cluster, number 2 represents the tRNA aminoacylation of protein translation cluster, number 3 represents the cluster of septin proteins, number 4 represents the cell biosynthetic process (transcription and translation), number 5 represents the hematopoietic cluster, and number 6 represents the mitochondria-related cluster. [Figure 6] Venn diagram showing the overlap of protein identification between the two MkEV batches. Of the 2332 proteins, 973 were shared between both batches, 1226 were identified in the first batch, and 133 were identified in the second MkEV batch. [Figure 7] Heatmap of protein intensities of 973 overlapping MkEV proteins (sample loading normalized), highlighting the strong consistency of protein abundance across MkEV batches. [Figure 8] Venn diagram visualizing the overlap between the MkEV proteome and a comprehensive list of plasma EV proteins (created by combining four publicly available sources and a newly generated dataset). This comparison revealed 508 proteins uniquely identified in MkEV. [Figure 9A] A series of Venn diagrams revealing a set of MkEV proteins not previously identified in CD61+EVs from healthy donors (Figure 9A). There is strong overlap between CD61+EVs and plasma EVs (Figure 2B). A total of 2,109 proteins were identified in MkEVs but absent from CD61+EVs. [Figure 9B]A series of Venn diagrams revealing a set of MkEV proteins not previously identified in CD61+EVs from healthy donors (Figure 9A). There is strong overlap between CD61+EVs and plasma EVs (Figure 2B). A total of 2,109 proteins were identified in MkEVs but absent from CD61+EVs. [Figure 10]

[0023] Figure 1 is a Venn diagram showing protein-coding transcripts in the MkEV of the present disclosure. Overlap between MkEV protein-coding genes (RNA-seq) and transcripts identified across the VesiclePedia database reveals 2701 mRNA transcripts unique to the MkEV of the present disclosure. [Figure 11] 1 is a histogram showing KEGG pathways of unique protein-coding transcripts in MkEV of the present disclosure. KEGG pathway enrichment of all unique MkEV protein-coding transcripts (2701 genes total) is shown. A manually curated KEGG pathway set is displayed. [Figure 12] 1 is a table listing the MkEV-specific genes (RNA-seq) disclosed herein associated with selected KEGG pathways. [Figure 13] Confocal microscopy images of HSPCs (lineage-depleted CD150+CD48- mouse bone marrow cells) co-cultured with MkEV loaded with GFP-tagged Cas9 ribonucleoprotein (RNP) are shown. Cells co-cultured with cargo-loaded MkEV were GFP-positive, indicating cellular uptake of GFP-tagged Cas9-loaded MkEV. In contrast, control samples containing cells alone and control samples containing cells co-cultured with MkEV+RNP (MKEV was mixed with RNP but not loaded by electroporation (no EP) before co-culture with cells) did not show GFP positivity. These data demonstrate successful delivery of RNP-loaded MkEV into HSPCs. [Figure 14A]Preferential targeting of MkEV to hematopoietic stem and progenitor cells ex vivo is shown. MkEV loaded with either GFP-tagged Cas9 protein (Figure 14A) or labeled with the lipophilic fluorescent dye DiD (Figure 14B) was cocultured with primary whole bone marrow from wild-type mice. After 24 hours of coculture, cells were analyzed by flow cytometry for the percentage of cells that were GFP+ or DiD+ (i.e., MkEV+). In addition, the percentages of lineage-positive (Lin+), lineage-negative (Lin-), and lineage-negative / c-Kit+ / Sca-1+ cells were simultaneously determined using fluorescently labeled antibodies against the lineage-positive markers, Sca-1, and c-Kit cell surface protein. The percentages of each subtype of cells in the heterogeneous whole bone marrow population are shown in Figure 14A. In the case of cells cocultured with MkEV loaded with GFP-tagged Cas9 (shown by the bar graph in Figure 14A), despite the majority (95%) of cells in culture being Lin+ cells (differentiated cells), only up to 23% of these cells were positive for MkEV. In contrast, although less than 5% were hematopoietic stem and progenitor cells (Lin- cells), nearly 50% of these cells were positive for MkEV at a cell dose of 300 EVs. Finally, in the case of LSK cells, the rarest and most pluripotent hematopoietic stem cells evaluated in these cultures, which comprised only 0.25% of the population, nearly 40% of this population was positive for MkEV. These data indicate preferential ex vivo targeting of bone marrow-derived hematopoietic stem and progenitor cells. Similarly, as shown in Figure 14B, in the case of whole bone marrow cells cocultured with DiD-labeled MkEV, only 20% of Lin+ cells were positive for MkEV. In contrast, 30% of the rarer Lin- cell population was positive for MkEV. Finally, in the case of the rarest and most multipotent hematopoietic stem cells evaluated in these cultures (LSK), up to 48% of this population was positive for MKEV. There was no significant change in the percentage of total Lin+ and Lin- cells in whole bone marrow cultures across all MKEV coculture conditions compared with controls (Fig. 14C). [Figure 14B]Preferential targeting of MkEV to hematopoietic stem and progenitor cells ex vivo is shown. MkEV loaded with either GFP-tagged Cas9 protein (Figure 14A) or labeled with the lipophilic fluorescent dye DiD (Figure 14B) was cocultured with primary whole bone marrow from wild-type mice. After 24 hours of coculture, cells were analyzed by flow cytometry for the percentage of cells that were GFP+ or DiD+ (i.e., MkEV+). In addition, the percentages of lineage-positive (Lin+), lineage-negative (Lin-), and lineage-negative / c-Kit+ / Sca-1+ cells were simultaneously determined using fluorescently labeled antibodies against the lineage-positive markers, Sca-1, and c-Kit cell surface protein. The percentages of each subtype of cells in the heterogeneous whole bone marrow population are shown in Figure 14A. In the case of cells cocultured with MkEV loaded with GFP-tagged Cas9 (shown by the bar graph in Figure 14A), despite the majority (95%) of cells in culture being Lin+ cells (differentiated cells), only up to 23% of these cells were positive for MkEV. In contrast, although less than 5% were hematopoietic stem and progenitor cells (Lin- cells), nearly 50% of these cells were positive for MkEV at a cell dose of 300 EVs. Finally, in the case of LSK cells, the rarest and most pluripotent hematopoietic stem cells evaluated in these cultures, which comprised only 0.25% of the population, nearly 40% of this population was positive for MkEV. These data indicate preferential ex vivo targeting of bone marrow-derived hematopoietic stem and progenitor cells. Similarly, as shown in Figure 14B, in the case of whole bone marrow cells cocultured with DiD-labeled MkEV, only 20% of Lin+ cells were positive for MkEV. In contrast, 30% of the rarer Lin- cell population was positive for MkEV. Finally, in the case of the rarest and most multipotent hematopoietic stem cells evaluated in these cultures (LSK), up to 48% of this population was positive for MKEV. There was no significant change in the percentage of total Lin+ and Lin- cells in whole bone marrow cultures across all MKEV coculture conditions compared with controls (Fig. 14C). [Figure 14C]Preferential targeting of MkEV to hematopoietic stem and progenitor cells ex vivo is shown. MkEV loaded with either GFP-tagged Cas9 protein (Figure 14A) or labeled with the lipophilic fluorescent dye DiD (Figure 14B) was cocultured with primary whole bone marrow from wild-type mice. After 24 hours of coculture, cells were analyzed by flow cytometry for the percentage of cells that were GFP+ or DiD+ (i.e., MkEV+). In addition, the percentages of lineage-positive (Lin+), lineage-negative (Lin-), and lineage-negative / c-Kit+ / Sca-1+ cells were simultaneously determined using fluorescently labeled antibodies against the lineage-positive markers, Sca-1, and c-Kit cell surface protein. The percentages of each subtype of cells in the heterogeneous whole bone marrow population are shown in Figure 14A. In the case of cells cocultured with MkEV loaded with GFP-tagged Cas9 (shown by the bar graph in Figure 14A), despite the majority (95%) of cells in culture being Lin+ cells (differentiated cells), only up to 23% of these cells were positive for MkEV. In contrast, although less than 5% were hematopoietic stem and progenitor cells (Lin- cells), nearly 50% of these cells were positive for MkEV at a cell dose of 300 EVs. Finally, in the case of LSK cells, the rarest and most pluripotent hematopoietic stem cells evaluated in these cultures, which comprised only 0.25% of the population, nearly 40% of this population was positive for MkEV. These data indicate preferential ex vivo targeting of bone marrow-derived hematopoietic stem and progenitor cells. Similarly, as shown in Figure 14B, in the case of whole bone marrow cells cocultured with DiD-labeled MkEV, only 20% of Lin+ cells were positive for MkEV. In contrast, 30% of the rarer Lin- cell population was positive for MkEV. Finally, in the case of the rarest and most multipotent hematopoietic stem cells evaluated in these cultures (LSK), up to 48% of this population was positive for MKEV. There was no significant change in the percentage of total Lin+ and Lin- cells in whole bone marrow cultures across all MKEV coculture conditions compared with controls (Fig. 14C). [Figure 15-1]Figure 15A shows the in vivo biodistribution of fluorescently labeled MkEV after in vivo delivery to wild-type mice. The experimental design is shown in Figure 15A (n = 3-5 mice / group). Fluorescently labeled MkEV was intravenously injected into wild-type mice via the tail vein, and tissues were harvested and analyzed for fluorescence 16 hours after injection. Figure 15B shows the fluorescent signal detected by IVIS in femurs dissected from mice. n = 5 mice / group. Fluorescence in each homogenized tissue was assayed by plate reader and normalized by tissue weight (Figure 15C). In Figure 15C, the data points from left to right for each tissue are saline, dye only, and MkEV, respectively. Figures 15D and 15E show graphs of experimental data from bone marrow cells stained with antibodies against CD45, lineage markers, CD150, CD201, and CD48, analyzed by flow cytometry to determine the percentage of hematopoietic (CD45+ cells, Figure 15D) and highly primitive long-term hematopoietic stem cells (CD45+ / Lin- / CD150+ / CD201+ / CD48- cells, Figure 15E) that were positive for MkEV. MkEV preferentially targets hematopoietic cells within the bone marrow and its compartments (Figure 15D), targeting very rare (less than 0.03% of bone marrow cells) long-term hematopoietic stem cells (Figure 15E). There were no changes in peripheral white blood cell counts (Fig. 15F), hemoglobin (Fig. 15G), platelet counts (Fig. 15H), WBC differentials (Fig. 15I), or the % of CD45+ and CD45- cells (Fig. 11J), or the % of long-term hematopoietic stem cells (Fig. 15K) in the bone marrow 16 hours after MkEV injection, indicating a lack of hematopoietic toxicity after in vivo injection. [Figure 15-2]Figure 15A shows the in vivo biodistribution of fluorescently labeled MkEV after in vivo delivery to wild-type mice. The experimental design is shown in Figure 15A (n = 3-5 mice / group). Fluorescently labeled MkEV was intravenously injected into wild-type mice via the tail vein, and tissues were harvested and analyzed for fluorescence 16 hours after injection. Figure 15B shows the fluorescent signal detected by IVIS in femurs dissected from mice. n = 5 mice / group. Fluorescence in each homogenized tissue was assayed by plate reader and normalized by tissue weight (Figure 15C). In Figure 15C, the data points from left to right for each tissue are saline, dye only, and MkEV, respectively. Figures 15D and 15E show graphs of experimental data from bone marrow cells stained with antibodies against CD45, lineage markers, CD150, CD201, and CD48, analyzed by flow cytometry to determine the percentage of hematopoietic (CD45+ cells, Figure 15D) and highly primitive long-term hematopoietic stem cells (CD45+ / Lin- / CD150+ / CD201+ / CD48- cells, Figure 15E) that were positive for MkEV. MkEV preferentially targets hematopoietic cells within the bone marrow and its compartments (Figure 15D), targeting very rare (less than 0.03% of bone marrow cells) long-term hematopoietic stem cells (Figure 15E). There were no changes in peripheral white blood cell counts (Fig. 15F), hemoglobin (Fig. 15G), platelet counts (Fig. 15H), WBC differentials (Fig. 15I), or the % of CD45+ and CD45- cells (Fig. 11J), or the % of long-term hematopoietic stem cells (Fig. 15K) in the bone marrow 16 hours after MkEV injection, indicating a lack of hematopoietic toxicity after in vivo injection. [Figure 15-3]Figure 15A shows the in vivo biodistribution of fluorescently labeled MkEV after in vivo delivery to wild-type mice. The experimental design is shown in Figure 15A (n = 3-5 mice / group). Fluorescently labeled MkEV was intravenously injected into wild-type mice via the tail vein, and tissues were harvested and analyzed for fluorescence 16 hours after injection. Figure 15B shows the fluorescent signal detected by IVIS in femurs dissected from mice. n = 5 mice / group. Fluorescence in each homogenized tissue was assayed by plate reader and normalized by tissue weight (Figure 15C). In Figure 15C, the data points from left to right for each tissue are saline, dye only, and MkEV, respectively. Figures 15D and 15E show graphs of experimental data from bone marrow cells stained with antibodies against CD45, lineage markers, CD150, CD201, and CD48, analyzed by flow cytometry to determine the percentage of hematopoietic (CD45+ cells, Figure 15D) and highly primitive long-term hematopoietic stem cells (CD45+ / Lin- / CD150+ / CD201+ / CD48- cells, Figure 15E) that were positive for MkEV. MkEV preferentially targets hematopoietic cells within the bone marrow and its compartments (Figure 15D), targeting very rare (less than 0.03% of bone marrow cells) long-term hematopoietic stem cells (Figure 15E). There were no changes in peripheral white blood cell counts (Fig. 15F), hemoglobin (Fig. 15G), platelet counts (Fig. 15H), WBC differentials (Fig. 15I), or the % of CD45+ and CD45- cells (Fig. 11J), or the % of long-term hematopoietic stem cells (Fig. 15K) in the bone marrow 16 hours after MkEV injection, indicating a lack of hematopoietic toxicity after in vivo injection. [Figure 16]This figure shows a graph of experimental data demonstrating the loading of Cas9 into MkEVs. MkEVs were electroporated with Cas9, treated with proteinase K to remove any non-internalized cargo (free cargo and vesicle surface-associated cargo), or treated by filtration to remove any free cargo, and then analyzed by Western blotting to quantify Cas9. Controls included MkEVs + Cas9 without electroporation ± proteinase K ± filtration. Cas9 was present in electroporated MkEVs but not in control, non-electroporated MkEVs after filtration, indicating successful vesicle association and / or internalization of the protein cargo. Cas9 was present in electroporated MkEVs but not in control, non-electroporated MkEVs after proteinase K digestion, indicating successful internalization and protection of the protein cargo after electroporation. [Figure 17A] The in vivo biodistribution of pDNA-loaded EVs was demonstrated and analyzed. A non-limiting schematic of this method is shown in Figure 17A. Reporter pDNA (CMV-driven GFP) was loaded into MkEVs by electroporation, labeled with DiD, and then injected into wild-type mice via the tail vein. Tissues were isolated 16 hours after injection and analyzed for EV association by flow cytometry. Vehicle-only injection (saline control) served as a negative control. As shown in Figure 17B, cargo-loaded EVs targeted to the bone marrow after intravenous injection and showed preferential uptake in hematopoietic stem and progenitor cells within the bone marrow compartment. Bars represent the mean DiD+ cell percentage ± standard deviation, n = 1 mouse per group. [Figure 17B]The in vivo biodistribution of pDNA-loaded EVs was demonstrated and analyzed. A non-limiting schematic of this method is shown in Figure 17A. Reporter pDNA (CMV-driven GFP) was loaded into MkEVs by electroporation, labeled with DiD, and then injected into wild-type mice via the tail vein. Tissues were isolated 16 hours after injection and analyzed for EV association by flow cytometry. Vehicle-only injection (saline control) served as a negative control. As shown in Figure 17B, cargo-loaded EVs targeted to the bone marrow after intravenous injection and showed preferential uptake in hematopoietic stem and progenitor cells within the bone marrow compartment. Bars represent the mean DiD+ cell percentage ± standard deviation, n = 1 mouse per group. [Figure 18A] We demonstrate that pDNA-loaded MkEVs deliver pDNA cargo to hematopoietic stem and progenitor cells (HSPCs) in vivo. A non-limiting schematic of this method is shown in Figure 18A. Reporter pDNA (pDNA encoding CMV-driven GFP) was loaded into MkEVs by electroporation, labeled with DiD, and then injected into wild-type mice via the tail vein. 16 hours after injection, HSPCs (lineage-negative / c-Kit+ / Sca-1+ cells) were isolated and analyzed for pDNA delivery by qPCR. Two representative experiments are shown in Figures 18B-18C. In Figure 18B, the bars represent the relative pDNA levels (fold change relative to the saline control) in HSPCs injected with vehicle alone (saline control), 100 ng pDNA alone (pDNA-only control), and pDNA-loaded EVs. n = 1 mouse per group. As shown, pDNA was successfully delivered to HSPCs after intravenous injection. In Figure 18C, the bars represent the relative pDNA levels (fold change relative to the saline control) in HSPCs in mice injected with vehicle alone (saline control) and pDNA-loaded EVs. As shown, pDNA was successfully delivered to HSPCs after intravenous injection. Furthermore, pDNA cargo was recovered only from HSPCs that were positive for EVs (DiD+) after intravenous injection. In contrast, minimal, if any, pDNA was recovered from HSPCs that did not take up EVs after intravenous injection (DiD-). n = 1 mouse per group. [Figure 18B] We demonstrate that pDNA-loaded MkEVs deliver pDNA cargo to hematopoietic stem and progenitor cells (HSPCs) in vivo. A non-limiting schematic of this method is shown in Figure 18A. Reporter pDNA (pDNA encoding CMV-driven GFP) was loaded into MkEVs by electroporation, labeled with DiD, and then injected into wild-type mice via the tail vein. 16 hours after injection, HSPCs (lineage-negative / c-Kit+ / Sca-1+ cells) were isolated and analyzed for pDNA delivery by qPCR. Two representative experiments are shown in Figures 18B-18C. In Figure 18B, the bars represent the relative pDNA levels (fold change relative to the saline control) in HSPCs injected with vehicle alone (saline control), 100 ng pDNA alone (pDNA-only control), and pDNA-loaded EVs. n = 1 mouse per group. As shown, pDNA was successfully delivered to HSPCs after intravenous injection. In Figure 18C, the bars represent the relative pDNA levels (fold change relative to the saline control) in HSPCs in mice injected with vehicle alone (saline control) and pDNA-loaded EVs. As shown, pDNA was successfully delivered to HSPCs after intravenous injection. Furthermore, pDNA cargo was recovered only from HSPCs that were positive for EVs (DiD+) after intravenous injection. In contrast, minimal, if any, pDNA was recovered from HSPCs that did not take up EVs after intravenous injection (DiD-). n = 1 mouse per group. [Figure 18C]We demonstrate that pDNA-loaded MkEVs deliver pDNA cargo to hematopoietic stem and progenitor cells (HSPCs) in vivo. A non-limiting schematic of this method is shown in Figure 18A. Reporter pDNA (pDNA encoding CMV-driven GFP) was loaded into MkEVs by electroporation, labeled with DiD, and then injected into wild-type mice via the tail vein. 16 hours after injection, HSPCs (lineage-negative / c-Kit+ / Sca-1+ cells) were isolated and analyzed for pDNA delivery by qPCR. Two representative experiments are shown in Figures 18B-18C. In Figure 18B, the bars represent the relative pDNA levels (fold change relative to the saline control) in HSPCs injected with vehicle alone (saline control), 100 ng pDNA alone (pDNA-only control), and pDNA-loaded EVs. n = 1 mouse per group. As shown, pDNA was successfully delivered to HSPCs after intravenous injection. In Figure 18C, the bars represent the relative pDNA levels (fold change relative to the saline control) in HSPCs in mice injected with vehicle alone (saline control) and pDNA-loaded EVs. As shown, pDNA was successfully delivered to HSPCs after intravenous injection. Furthermore, pDNA cargo was recovered only from HSPCs that were positive for EVs (DiD+) after intravenous injection. In contrast, minimal, if any, pDNA was recovered from HSPCs that did not take up EVs after intravenous injection (DiD-). n = 1 mouse per group. DETAILED DESCRIPTION OF THE INVENTION

[0026] The present invention is based, in part, on the discovery of compositions of substantially purified megakaryocyte-derived extracellular vesicles characterized by a specific set of physical characteristics, such as biomarker composition (e.g., the presence, absence, or amount of protein biomarker(s) or gene biomarker(s)) and size, and capable of carrying cargo within their lumen for use in delivering therapeutic agents, as well as methods for making or using the same. In embodiments, the megakaryocyte-derived extracellular vesicles of the present disclosure are distinct from naturally occurring products collected from whole blood (platelet-free plasma) or derived from activated platelets (platelet EVs). Thus, in aspects, the present invention provides compositions of megakaryocyte-derived extracellular vesicles, as well as methods for obtaining and using the same, that are consistently produced to have desirable properties and carry specific cargo, increasing the likelihood of their successful therapeutic use.

[0027] Megakaryocyte-derived extracellular vesicles, which are relatively immune-silent, can be administered repeatedly, a clear advantage over immunogenic viral vectors. In some embodiments, megakaryocyte-derived extracellular vesicles are useful for in vivo genomic medicine without the need for conditioning therapy, allowing people to receive megakaryocyte-derived extracellular vesicles in outpatient settings. This platform represents a significant paradigm shift in gene therapy from ex vivo delivery to in vivo delivery, democratizing gene therapy by shortening time to treatment and reducing costs.

[0028] In one aspect, the present invention relates to a composition comprising a plurality of substantially purified megakaryocyte-derived extracellular vesicles derived from human pluripotent stem cells, wherein the composition comprises one or more proteins that are unique to or preferentially present in the megakaryocyte-derived extracellular vesicles compared to other types of extracellular vesicles.

[0029] In one aspect, the present invention relates to a composition comprising a plurality of substantially purified megakaryocyte-derived extracellular vesicles derived from human pluripotent stem cells, the composition comprising one or more nucleic acids encoding one or more proteins that are unique to or preferentially present in the megakaryocyte-derived extracellular vesicles compared to other types of extracellular vesicles. In some embodiments, the one or more proteins or the one or more nucleic acids encoding the one or more proteins are involved in aminoacyl-tRNA biosynthesis and / or neutrophil extracellular trap formation.

[0030] In one aspect, the present invention relates to a composition comprising a plurality of substantially purified megakaryocyte-derived extracellular vesicles derived from human pluripotent stem cells, the composition comprising a lipid bilayer membrane surrounding an inner lumen, the inner lumen of the megakaryocyte-derived extracellular vesicles comprising one or more megakaryocyte-derived nucleic acid molecules selected from mRNA, tRNA, rRNA, siRNA, microRNA, regulatory RNA, and non-coding RNA and coding RNA, the lipid bilayer membrane comprising one or more proteins associated with or embedded therein, the one or more proteins being unique to or preferentially present in the megakaryocyte-derived extracellular vesicles compared to other types of extracellular vesicles.

[0031] In one aspect, the present invention relates to a composition comprising a plurality of substantially purified megakaryocyte-derived extracellular vesicles derived from human pluripotent stem cells, the composition comprising a lipid bilayer membrane surrounding a lumen, the lumen of the megakaryocyte-derived extracellular vesicles comprising one or more megakaryocyte-derived nucleic acid molecules selected from mRNA, tRNA, rRNA, siRNA, microRNA, regulatory RNA, and non-coding RNA and coding RNA, the lipid bilayer membrane comprising one or more nucleic acids encoding one or more proteins associated with or embedded within the lipid bilayer membrane, and which are unique to or preferentially present in the megakaryocyte-derived extracellular vesicles compared to other types of extracellular vesicles. In some embodiments, the nucleic acid molecules are derived from an exogenous source.

[0032] In one aspect, the present invention relates to a composition comprising a plurality of substantially purified megakaryocyte-derived extracellular vesicles derived from human pluripotent stem cells, wherein the composition comprises one or more nucleic acids that are unique to or preferentially present in the megakaryocyte-derived extracellular vesicles compared to other types of extracellular vesicles.

[0033] In one aspect, the invention relates to a composition comprising a plurality of substantially purified megakaryocyte-derived extracellular vesicles derived from human pluripotent stem cells, the composition comprising one or more proteins encoded by one or more nucleic acids that are unique to or preferentially present in the megakaryocyte-derived extracellular vesicles compared to other types of extracellular vesicles. In some embodiments, the one or more nucleic acids, or the one or more proteins encoded by the one or more nucleic acids, are involved in neutrophil extracellular trap formation, hematopoietic cell lineage, cell adhesion, ATP-binding cassette (ABC) transport, chemokine signaling pathway, phagocytosis, and / or aminoacyl-tRNA biosynthesis.

[0034] In some embodiments, megakaryocyte-derived extracellular vesicles are suitable for loading cargo into the lumen of megakaryocyte-derived extracellular vesicles and / or loading cargo associated with the surface of megakaryocyte-derived extracellular vesicles. In some embodiments, in addition to or as an alternative to cargo located within the lumen of megakaryocyte-derived extracellular vesicles, cargo is loaded into megakaryocytes and packaged into the extracellular vesicles. In some embodiments, in addition to or as an alternative to cargo located within the lumen of megakaryocyte-derived extracellular vesicles, cargo is loaded directly into megakaryocyte-derived extracellular vesicles. In some embodiments, in addition to or as an alternative to cargo located within the lumen of megakaryocyte-derived extracellular vesicles, cargo is loaded into megakaryocytes and packaged into the extracellular vesicles. In some embodiments, in addition to or as an alternative to cargo located within the lumen of megakaryocyte-derived extracellular vesicles, cargo is loaded directly into megakaryocyte-derived extracellular vesicles.

[0035] In some embodiments, the cargo is one or more therapeutic agents, including therapeutic agents described herein. In some embodiments, the cargo comprises one or more megakaryocyte-derived nucleic acid molecules selected from mRNA, tRNA, rRNA, siRNA, microRNA, regulatory RNA, and non-coding RNA and coding RNA. In some embodiments, in addition to or as an alternative to cargo located within the lumen of megakaryocyte-derived extracellular vesicles, cargo is loaded into megakaryocytes and packaged into extracellular vesicles. In some embodiments, in addition to or as an alternative to cargo located within the lumen of megakaryocyte-derived extracellular vesicles, cargo is directly loaded into megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles are suitable for loading cargo associated with the surface of megakaryocyte-derived extracellular vesicles.

[0036] In another aspect, the invention relates to a pharmaceutical composition comprising a composition disclosed herein and a pharmaceutically acceptable excipient or carrier.

[0037] In another aspect, the present invention relates to a method for delivering a deliverable therapeutic agent, the method comprising: (a) obtaining megakaryocyte-derived extracellular vesicles of a composition disclosed herein; (b) incubating the megakaryocyte-derived extracellular vesicles with a therapeutic agent to allow the therapeutic agent to migrate into the lumen of the megakaryocyte-derived extracellular vesicles and produce a deliverable therapeutic agent; and (c) administering the deliverable therapeutic agent to a patient, or contacting the deliverable therapeutic agent with biological cells in vitro and administering the contacted biological cells to a patient.

[0038] In another aspect, the present invention relates to methods for treating various diseases or disorders with the present megakaryocyte-derived extracellular vesicles.

[0039] Biomarker Profile or Fingerprint In various embodiments, the megakaryocyte-derived extracellular vesicles are characterized by a unique biomarker profile or fingerprint that distinguishes them from, for example, naturally occurring megakaryocyte-derived extracellular vesicles and / or vesicles or extracellular vesicles derived from platelets. In various embodiments, the megakaryocyte-derived extracellular vesicles are characterized by such a biomarker profile or fingerprint, including identity (e.g., presence or absence) or quantity (e.g., substantial presence or substantial absence of a biomarker in a population of megakaryocyte-derived extracellular vesicles, or the presence or absence of a majority of megakaryocyte-derived extracellular vesicles in a population, or the percentage of megakaryocyte-derived extracellular vesicles bearing a biomarker).

[0040] Protein Biomarkers In embodiments, the compositions comprise substantially purified megakaryocyte-derived extracellular vesicles derived from human pluripotent stem cells, comprising a lipid bilayer membrane surrounding a lumen, the lipid bilayer membrane comprising one or more proteins (also referred to as biomarkers) associated with or embedded therein.

[0041] In embodiments, the compositions comprise substantially purified megakaryocyte-derived extracellular vesicles comprising one or more nucleic acids encoding one or more proteins that are unique to or preferentially present in megakaryocyte-derived extracellular vesicles compared to other types of extracellular vesicles.

[0042] In embodiments, direct protein-protein interactions exist between unique MkEV proteins of the present disclosure. In embodiments, the network comprises curated direct protein-protein interactions (interaction score greater than 0.4), where cluster number 1 represents a histone cluster, cluster number 2 represents a tRNA aminoacylation protein translation cluster, cluster number 3 represents a cluster of septin proteins, cluster number 4 represents a mitochondria-associated cluster, and cluster number 5 represents a cellular biogenesis process (both transcription and translation).

[0043] In embodiments, one or more proteins (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) or a gene encoding one or more proteins (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) One or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) involved in aminoacyl-tRNA biosynthesis are optionally selected from YARS1, AARS1, GARS1, LARS1, EPRS1, TARS1, DARS1, WARS1, VARS1, NARS1, RARS1, SARS1, KARS1, IARS1, CARS1, QARS, and HARS1.

[0044] In embodiments, one or more proteins (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) or one or more proteins (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or The one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) encoding a neutrophil extracellular trap (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) are involved in neutrophil extracellular trap formation and are optionally selected from H4C1, H2AC20, H3C1, H2BC12, H3C15, MACROH2A1, H2AX, H2AZ2, H3-3A, H2AC14, and MACROH2A2.

[0045] In some embodiments, one or more proteins (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) are encoded by one or more proteins (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175). The one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) to be loaded are selected from the group consisting of H4C16, H2AC17, H3C13, RPL9P9, PHB1, SETSIP, H2AZ1, XP32, COPS9, SEC61A2, MARS1, MRE11, ATP5F1C, H4C1, H2AC20, H3C1, H2BC12, FCSK, CAVIN2, YARS1, H3C15, M ACROH2A1, H2AX, H3-2, SELENOF, MPIG6B, Septin 7, AARS1, H1-5, H1-2, H2AZ2, GARS1, LARS1, THPO, RACK1, H3-3A, CCDC191, H2BC19P, Septin 2, EPRS1, TARS1, ATP5F1B, ERBIN, DARS1, WARS1, VARS1, NIBAN2, Septin 6, NARS1, RARS1, Septin 11, Septin 5, SARS1, NIBAN1, SNRPGP15, PIP4P2, CYRIB, CARMIL1, KARS1, IARS1, Septin 9, H1-4, ARHGAP45, H1-0, CARS1, GCN1, FADS2, TBC1D13, GET3, RO60, LAMTOR5, ELOC, H2AC14, SCARF1, RNF24, GCSAML, NRDC, ECPAS, MACROH2A2, ATP5 F1A, DMTN, TANGO2, CSF2RB, WASHC5, KCNA3, QARS1, MINDY1, PTPA, EXOC3L2, PRUNE1, PLPBP, THUMPD1, WASHC4, NECTIN2, GFUS, ADGRE2, AKAP8L, FAM234A,ADSS2, ANKRD13D, KCT2, NT5C3A, PIP4P1, CCDC9, ELOB, HPGDS, MEAK7, TOMM70, SMIM1, HARS1, ATP5PD, OGA, CHSY1, SQLE, S USD6, IGKV1-27, PEDS1-UBE2V1, CEP44, MYG1, NRROS, IL21R, GRK2, MCEMP1, ELAPOR2, SDAD1, CERT1, UBE2F, CALHM5, H1-10 , EIPR1, PBDC1, ARMH3, VIPAS39, MESD, PROSER2, RABL6, FYB1, C17orf49, RMDN3, KYAT3, TTMP, ERO1A, CD244, CZIB, PLPP3, BABAM2, H1-3, NAXE, ENSA, PALS2, GUCY1B1, UMAD1, MIX23, PRXL2B, SNU13, RTRAF, KXD1, VSIR, EPOR, or MARCHF2.

[0046] In embodiments, greater than about 1%, or greater than about 5%, or greater than about 10%, or greater than about 15%, or greater than about 20%, or greater than about 25%, or greater than about 30%, or greater than about 35%, or greater than about 40%, or greater than about 45%, or greater than about 50%, or greater than about 55%, or greater than about 60%, or greater than about 65%, or greater than about 70%, or greater than about 75%, or greater than about 80%, or greater than about 85%, or greater than about 90%, or greater than about 95% of the megakaryocyte-derived extracellular vesicles comprise one or more proteins or one or more nucleic acids encoding one or more proteins.

[0047] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of more than about 1%, more than about 2%, more than about 3%, more than about 4%, more than about 5%, more than about 6%, more than about 7%, more than about 8%, more than about 9%, more than about 10%, more than about 11%, more than about 12%, more than about 13%, more than about 14%, more than about 15%, more than about 16%, more than about 17%, more than about 18%, more than about 19%, more than about 20%, more than about 21%, more than about 22%, more than about 23%, more than about 24%, more than about 25%, more than about 26%, more than about 27%, more than about 28%, more than about 29%, more than about 30%, more than about 31%, more than about 32%, more than about 33%, more than about 34%, more than about 35%, more than about 36%, more than about 37%, more than about 38%, more than about 39%, more than about 40%, more than about 41%, more than about 42%, more than about 43%, more than about 44%, more than about 45%, more than about 46%, more than about 47%, more than about 48%, more than about 49%, more than about 50%, more than about 51%, more than about 52%, more than about 53%, more than about 54%, more than about 55%, more than about 56%, more than about 57%, more than about 58%, more than about 59%, more than about 60%, more than about 61%, more than about 62%, more than about 63%, more than about 64%, more than about 65%, more than about 66%, more than about 67%, more than about 68%, more than about 69%, more than about 70%, more than about 71%, More than 6%, more than about 27%, more than about 28%, more than about 29%, more than about 30%, more than about 31%, more than about 32%, more than about 33%, more than about 34%, more than about 35%, more than about 36%, more than about 37%, more than about 38%, more than about 39%, More than about 40%, more than about 41%, more than about 42%, more than about 43%, more than about 44%, more than about 45%, more than about 46%, more than about 47%, more than about 48%, more than about 49%, more than about 50%, more than about 51%, more than about 52%, about 53% Super, more than about 54%, more than about 55%, more than about 56%, more than about 57%, more than about 58%, more than about 59%, more than about 60%, more than about 61%, more than about 62%, more than about 63%, more than about 64%, more than about 65%, more than about 66%, about 6 More than 7%, more than about 68%, more than about 69%, more than about 70%, more than about 71%, more than about 72%, more than about 73%, more than about 74%, more than about 75%, more than about 76%, more than about 77%, more than about 78%, more than about 79%, more than about 80%, More than about 81%, more than about 82%, more than about 83%, more than about 84%, more than about 85%, more than about 86%, more than about 87%, more than about 88%, more than about 89%, more than about 90%, more than about 91%, more than about 92%, more than about 93%, more than about 94%, more than about 95%, more than about 96%, more than about 97%, more than about 98%, more than about 99%, or more than about 99% comprise one or more proteins or one or more nucleic acids encoding one or more proteins.

[0048] In several embodiments, less than about 95%, or less than about 90%, or less than about 85%, or less than about 80%, or less than about 75%, or less than about 70%, or less than about 65%, or less than about 60%, or less than about 55%, or less than about 50%, or less than about 45%, or less than about 40%, or less than about 35%, or less than about 30%, or less than about 25%, or less than about 20%, or less than about 15%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles comprise one or more proteins or one or more nucleic acids encoding one or more proteins.

[0049] In embodiments, less than about 99%, or less than about 98%, or less than about 97%, or less than about 96%, or less than about 95%, or less than about 94%, or less than about 93%, or less than about 92%, or less than about 91%, or less than about 90%, or less than about 89%, or less than about 88%, or less than about 87%, or less than about 86%, less than, less than about 85%, or less than about 84%, or less than about 83%, or less than about 82%, or less than about 81%, or less than about 80%, or less than about 79%, or less than about 78%, or less than about 77%, or less than about 99%, or less than about 98%, or less than about 98%, or less than about 99% of the megakaryocyte-derived extracellular vesicles. or less than about 76%, or less than about 75%, or less than about 74%, or less than about 73%, or less than about 72%, or less than about 71%, or less than about 70%, or less than about 69%, or less than about 68%, or less than about 67%, or less than about 66%, or less than about 65%, or less than about 64%, or less than about 63%, or less than about 62%, or less than about 61%, or less than about 60%, or less than about 59%, or less than about 58%, or less than about 57%, or less than about 56%, or less than about 55%, or less than about 54%, or less than about 53%, or less than about 52%, or or less than about 51%, or less than about 50%, or less than about 49%, or less than about 48%, or less than about 47%, or less than about 46%, or less than about 45%, or less than about 44%, or less than about 43%, or less than about 42%, or less than about 41%, or less than about 40%, or less than about 39%, or less than about 38%, or less than about 37%, or less than about 36%, or less, or less than about 35%, or less than about 34%, or less than about 33%, or less than about 32%, or less than about 31%, or less than about 30%, or less than about 29%, or less than about 28%, or less than about 27%, or less than about 26%, or less than about 25%, or less than about 24%, or less than about 23%, or less than about 22%, or less than about 21%, or less than about 20%, or less than about 19%, or less than about 18%, or less than about 17%, or less than about 16%, or less than about 15%, or less than about 14%, or less than about 13%, or less than about 12%, or less than about 11%, or less than about 10%, or less than about 9%, or less than about 8%, or less than about 7%, or less than about 6%, or less than about 5%, or less than about 4%, or less than about 3%, or less than about 2%, or less than about 1%It includes one or more proteins or one or more nucleic acids encoding one or more proteins.

[0050] In several embodiments, the lipid bilayer membrane comprises one or more proteins (e.g., at least three, or at least five, or at least ten) or one or more nucleic acids (e.g., at least three, or at least five, or at least ten) encoding one or more proteins (e.g., at least three, or at least five, or at least ten) selected from CAVIN2, MPIG6B, ERBIN, ELOC, CSF2RB, KCNA3, NECTIN2, IL21R, MCEMP1, PROSER2, FYB1, CD244, and EPOR. In embodiments, greater than about 1%, or greater than about 5%, or greater than about 10%, or greater than about 15%, or greater than about 20%, or greater than about 25%, or greater than about 30%, or greater than about 35%, or greater than about 40%, or greater than about 45%, or greater than about 50%, or greater than about 55%, or greater than about 60%, or greater than about 65%, or greater than about 70%, or greater than about 75%, or greater than about 80%, or greater than about 85%, or greater than about 90%, or greater than about 95% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising one or more proteins or one or more nucleic acids encoding one or more proteins.

[0051] In several embodiments, the megakaryocyte-derived extracellular vesicles are greater than about 1%, greater than about 2%, greater than about 3%, greater than about 4%, greater than about 5%, greater than about 6%, greater than about 7%, greater than about 8%, greater than about 9%, greater than about 10%, greater than about 11%, greater than about 12%, greater than about 13%, greater than about 14%, greater than about 15%, greater than about 16%, greater than about 17%, greater than about 18%, greater than about 19%, greater than about 20%, greater than about 21%, greater than about 22%, greater than about 23%, greater than about 24%, greater than about 25%, greater than about 26%, or greater than about 28%. Very, more than about 27%, more than about 28%, more than about 29%, more than about 30%, more than about 31%, more than about 32%, more than about 33%, more than about 34%, more than about 35%, more than about 36%, more than about 37%, more than about 38%, more than about 39%, about 40 More than %, more than about 41%, more than about 42%, more than about 43%, more than about 44%, more than about 45%, more than about 46%, more than about 47%, more than about 48%, more than about 49%, more than about 50%, more than about 51%, more than about 52%, more than about 53%, about 5 More than 4%, more than about 55%, more than about 56%, more than about 57%, more than about 58%, more than about 59%, more than about 60%, more than about 61%, more than about 62%, more than about 63%, more than about 64%, more than about 65%, more than about 66%, more than about 67%, about More than 68%, more than about 69%, more than about 70%, more than about 71%, more than about 72%, more than about 73%, more than about 74%, more than about 75%, more than about 76%, more than about 77%, more than about 78%, more than about 79%, more than about 80%, more than about 81%, More than about 82%, more than about 83%, more than about 84%, more than about 85%, more than about 86%, more than about 87%, more than about 88%, more than about 89%, more than about 90%, more than about 91%, more than about 92%, more than about 93%, more than about 94%, more than about 95%, more than about 96%, more than about 97%, more than about 98%, more than about 99%, or more than about 99% comprise lipid bilayer membranes comprising one or more proteins or one or more nucleic acids encoding one or more proteins.

[0052] In several embodiments, less than about 95%, or less than about 90%, or less than about 85%, or less than about 80%, or less than about 75%, or less than about 70%, or less than about 65%, or less than about 60%, or less than about 55%, or less than about 50%, or less than about 45%, or less than about 40%, or less than about 35%, or less than about 30%, or less than about 25%, or less than about 20%, or less than about 15%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising one or more proteins or one or more nucleic acids encoding one or more proteins.

[0053] In embodiments, less than about 99%, or less than about 98%, or less than about 97%, or less than about 96%, or less than about 95%, or less than about 94%, or less than about 93%, or less than about 92%, or less than about 91%, or less than about 90%, or less than about 89%, or less than about 88%, or less than about 87%, or less than about 86%, less than, less than about 85%, or less than about 84%, or less than about 83%, or less than about 82%, or less than about 81%, or less than about 80%, or less than about 79%, or less than about 78%, or less than about 77%, or less than about 99%, or less than about 98%, or less than about 98%, or less than about 99% of the megakaryocyte-derived extracellular vesicles. or less than about 76%, or less than about 75%, or less than about 74%, or less than about 73%, or less than about 72%, or less than about 71%, or less than about 70%, or less than about 69%, or less than about 68%, or less than about 67%, or less than about 66%, or less than about 65%, or less than about 64%, or less than about 63%, or less than about 62%, or less than about 61%, or less than about 60%, or less than about 59%, or less than about 58%, or less than about 57%, or less than about 56%, or less than about 55%, or less than about 54%, or less than about 53%, or less than about 52%, or or less than about 51%, or less than about 50%, or less than about 49%, or less than about 48%, or less than about 47%, or less than about 46%, or less than about 45%, or less than about 44%, or less than about 43%, or less than about 42%, or less than about 41%, or less than about 40%, or less than about 39%, or less than about 38%, or less than about 37%, or less than about 36%, or less, or less than about 35%, or less than about 34%, or less than about 33%, or less than about 32%, or less than about 31%, or less than about 30%, or less than about 29%, or less than about 28%, or less than about 27%, or less than about 26%, or less than about 25%, or less than about 24%, or less than about 23%, or less than about 22%, or less than about 21%, or less than about 20%, or less than about 19%, or less than about 18%, or less than about 17%, or less than about 16%, or less than about 15%, or less than about 14%, or less than about 13%, or less than about 12%, or less than about 11%, or less than about 10%, or less than about 9%, or less than about 8%, or less than about 7%, or less than about 6%, or less than about 5%, or less than about 4%, or less than about 3%, or less than about 2%, or less than about 1%A lipid bilayer membrane containing one or more proteins or one or more nucleic acids encoding one or more proteins.

[0054] In embodiments, the lipid bilayer membrane comprises one or more proteins (e.g., at least three, or at least five, or at least ten) or one or more nucleic acids (e.g., at least three, or at least five, or at least ten) encoding one or more proteins (e.g., at least three, or at least five, or at least ten) selected from CD54, CD18, CD43, CD11b, CD62P, CD41, CD61, CD21, CD51, phosphatidylserine (PS), CLEC-2, LAMP-1 (CD107a), CD63, CD42b, CD9, CD31, CD47, CD147, CD32a, and GPVI.

[0055] In embodiments, the lipid bilayer membrane comprises phosphatidylserine, for example but not limited to, Annexin V testing.

[0056] In embodiments, the lipid bilayer membrane comprises one or more proteins selected from CD62P, CD41, and CD61.

[0057] In several embodiments, greater than about 40%, greater than about 50%, greater than about 60%, greater than about 70%, greater than about 80%, greater than about 90%, greater than about 95%, or greater than about 99% of megakaryocyte-derived extracellular vesicles comprising a lipid bilayer membrane containing CD41 also contain CD61 in the lipid bilayer membrane.

[0058] In some embodiments, the megakaryocyte-derived extracellular vesicles are characterized by the expression and / or presence of one or more of CD54, CD18, CD43, CD11b, CD62P, CD41, CD61, CD21, CD51, and CLEC-2. In some embodiments, the megakaryocyte-derived extracellular vesicles are characterized by the expression and / or presence of one or more of PS, CD62P, LAMP-1 (CD107a), CD42b, CD9, CD43, CD31, and CD11b. In some embodiments, the megakaryocyte-derived extracellular vesicles are characterized by the expression and / or presence of one or more of PS, CD61, CD62P, LAMP-1 (CD107a), CLEC-2, and CD63. In some embodiments, the megakaryocyte-derived extracellular vesicles are characterized by the expression and / or presence of one or more of PS, CD62P, CLEC-2, CD9, CD31, CD147, CD32a, and GPVI. In some embodiments, the megakaryocyte-derived extracellular vesicles are characterized by the expression and / or presence of one or more of PS, CD62P, LAMP-1 (CD107a), CLEC-2, CD9, and CD31. In some embodiments, the megakaryocyte-derived extracellular vesicles are characterized by the expression and / or presence of one or more of CD62P, CD41, and CD61. In some embodiments, the megakaryocyte-derived extracellular vesicles are characterized by substantial expression and / or presence of one or more of CD54, CD18, CD43, CD11b, CD62P, CD41, CD61, CD21, CD51, and CLEC-2. In some embodiments, the megakaryocyte-derived extracellular vesicles are characterized by substantial expression and / or presence of one or more of CD62P, CD41, and CD61. In some embodiments, the megakaryocyte-derived extracellular vesicles are characterized by not expressing and / or not containing significant amounts of DRAQ5. In some embodiments, the megakaryocyte-derived extracellular vesicles are characterized by being substantially free of DRAQ5.

[0059] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, greater than about 99% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P.

[0060] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, about 40% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, about 90% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, about 95% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, about 99% or less of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P.

[0061] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P. In some embodiments, less than about 1% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD62P.

[0062] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD62P. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD62P. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD62P. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD62P. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD62P. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD62P. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD62P. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD62P. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD62P. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD62P.

[0063] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 4%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51% 1%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 100%, about 101%, about 102%, about 103%, about 104%, about 105%, about 106%, about 107%, about 108%, about 109%, about 110%, about 111%, about 112%, about 113%, about 114%, about 115%, about 116%, about 117%, about 118%, about 119%, about 120%, about 121%, about 122%, about 123%, about 124%, about 125%, about 126%, about 127%, about 128%, about 129%, about 130%, about 131%, about 132%, about 133%, about 134%, about 135%, about 136%, about 137%, about 138%, about 139%, about 140%, about 141%, about 142%, about 143%, about 14 8%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane containing CD62P.

[0064] In several embodiments, the megakaryocyte-derived extracellular vesicles are free or substantially free of CD62P.

[0065] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CD62P than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold greater amount of CD62P than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0066] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of CD62P than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CD62P than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0067] In some embodiments, megakaryocyte-derived extracellular vesicles have about 4 to about 32 times less, or about 8 to about 16 times less, of CD62P than platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 15 times less, or about 16 times less, of CD62P than platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 32 to about 128 times less, about 50 to about 75 times less, or about 60 to about 70 times less, of CD62P than platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have about 60 times less, about 64 times less, or about 70 times less, of CD62P than platelet-derived extracellular vesicles (PLT EVs).

[0068] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, greater than about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41.

[0069] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, about 40% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, about 90% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, about 95% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, about 99% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41.

[0070] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, less than about 1% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41.

[0071] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD41. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD41. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD41.

[0072] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 4%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51% 1%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 100%, about 101%, about 102%, about 103%, about 104%, about 105%, about 106%, about 107%, about 108%, about 109%, about 110%, about 111%, about 112%, about 113%, about 114%, about 115%, about 116%, about 117%, about 118%, about 119%, about 120%, about 121%, about 122%, about 123%, about 124%, about 125%, about 126%, about 127%, about 128%, about 129%, about 130%, about 131%, about 132%, about 133%, about 134%, about 135%, about 136%, about 137%, about 138%, about 139%, about 140%, about 141%, about 142%, about 143%, about 14 8%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane containing CD41.

[0073] In several embodiments, the megakaryocyte-derived extracellular vesicles comprise CD41.

[0074] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CD41 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold greater amounts of CD41 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0075] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower CD41 expression and / or presence than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CD41 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0076] In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold to about 8-fold or about 2-fold to about 4-fold the amount of CD41 / CD61 as platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, about 3-fold, or about 4-fold the amount of CD41 / CD61 as platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.1-fold to about 2-fold the amount of CD41 / CD61 as platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.1-fold or about 1.2-fold the amount of CD41 / CD61 as platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have substantially the same amount of CD41 / CD61 as platelet-derived extracellular vesicles (PLT EVs).

[0077] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, greater than about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61.

[0078] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, less than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, about 95% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, about 99% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61.

[0079] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, less than about 1% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61.

[0080] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD61. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD61. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD61. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD61.

[0081] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 4%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51% 1%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 100%, about 101%, about 102%, about 103%, about 104%, about 105%, about 106%, about 107%, about 108%, about 109%, about 110%, about 111%, about 112%, about 113%, about 114%, about 115%, about 116%, about 117%, about 118%, about 119%, about 120%, about 121%, about 122%, about 123%, about 124%, about 125%, about 126%, about 127%, about 128%, about 129%, about 130%, about 131%, about 132%, about 133%, about 134%, about 135%, about 136%, about 137%, about 138%, about 139%, about 140%, about 141%, about 142%, about 143%, about 14 8%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane containing CD61.

[0082] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CD61 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold greater amount of CD61 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0083] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of CD61 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CD61 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0084] In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold to about 8-fold or about 2-fold to about 4-fold the amount of CD61 as platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, about 3-fold, or about 4-fold the amount of CD61 as platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.1-fold to about 1 / 2-fold the amount of CD61 as platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.1-fold or about 1.2-fold the amount of CD61 as platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have substantially the same amount of CD61 as platelet-derived extracellular vesicles (PLT EVs).

[0085] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD4. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, greater than about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54.

[0086] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, about 40% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, about 90% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, about 95% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, about 99% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54.

[0087] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, less than about 1% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54.

[0088] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD54.

[0089] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 4%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51% 1%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 100%, about 101%, about 102%, about 103%, about 104%, about 105%, about 106%, about 107%, about 108%, about 109%, about 110%, about 111%, about 112%, about 113%, about 114%, about 115%, about 116%, about 117%, about 118%, about 119%, about 120%, about 121%, about 122%, about 123%, about 124%, about 125%, about 126%, about 127%, about 128%, about 129%, about 130%, about 131%, about 132%, about 133%, about 134%, about 135%, about 136%, about 137%, about 138%, about 139%, about 140%, about 141%, about 142%, about 143%, about 14 8%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane containing CD54.

[0090] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CD54 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold greater amount of CD54 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0091] In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold to about 10-fold or about 2-fold to about 4-fold the amount of CD54 as platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 3-fold the amount of CD54 as platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.1-fold to about 4-fold or about 1.1-fold to about 2-fold the amount of CD54 as platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.5-fold the amount of CD54 as platelet-derived extracellular vesicles (PLT EVs).

[0092] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of CD54 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CD54 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0093] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, greater than about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18.

[0094] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, about 40% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, about 90% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, about 95% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, about 99% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18.

[0095] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, less than about 1% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18.

[0096] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD18. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD 18. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD 18. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD 18.

[0097] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 4%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51% 1%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 100%, about 101%, about 102%, about 103%, about 104%, about 105%, about 106%, about 107%, about 108%, about 109%, about 110%, about 111%, about 112%, about 113%, about 114%, about 115%, about 116%, about 117%, about 118%, about 119%, about 120%, about 121%, about 122%, about 123%, about 124%, about 125%, about 126%, about 127%, about 128%, about 129%, about 130%, about 131%, about 132%, about 133%, about 134%, about 135%, about 136%, about 137%, about 138%, about 139%, about 140%, about 141%, about 142%, about 143%, about 14 8%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane comprising CD18.

[0098] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CD18 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold higher amounts of CD18 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0099] In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold to about 10-fold, 8-fold to about 64-fold, or about 16-fold to about 32-fold, or about 16-fold to about 24-fold the amount of CD18 as compared to platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 20-fold the amount of CD18 as compared to platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.1-fold to about 4-fold, or about 1.1-fold to about 2-fold the amount of CD18 as compared to platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.5-fold the amount of CD18 as compared to platelet-derived extracellular vesicles (PLT EVs).

[0100] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of CD18 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CD18 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0101] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, greater than about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43.

[0102] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, about 40% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, about 90% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, about 95% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, about 99% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43.

[0103] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, less than about 1% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43.

[0104] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD43. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD43. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD43. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD43.

[0105] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 4%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51% 1%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 100%, about 101%, about 102%, about 103%, about 104%, about 105%, about 106%, about 107%, about 108%, about 109%, about 110%, about 111%, about 112%, about 113%, about 114%, about 115%, about 116%, about 117%, about 118%, about 119%, about 120%, about 121%, about 122%, about 123%, about 124%, about 125%, about 126%, about 127%, about 128%, about 129%, about 130%, about 131%, about 132%, about 133%, about 134%, about 135%, about 136%, about 137%, about 138%, about 139%, about 140%, about 141%, about 142%, about 143%, about 14 8%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane containing CD43.

[0106] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CD43 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold more CD43 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0107] In some embodiments, megakaryocyte-derived extracellular vesicles have about 4-fold to about 64-fold, or about 8-fold to about 32-fold, or about 8-fold to about 16-fold the amount of CD43 as compared to platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 10-fold or about 12-fold the amount of CD43 as compared to platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.5-fold to about 8-fold, or about 2-fold to about 4-fold the amount of CD43 as compared to platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have about 3-fold or about 4-fold the amount of CD43 as compared to platelet-derived extracellular vesicles (PLT EVs).

[0108] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of CD43 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CD43 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0109] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, greater than about 99% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b.

[0110] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, about 40% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, about 90% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, about 95% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, about 99% or less of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b.

[0111] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, less than about 1% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b.

[0112] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD11b. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD11b. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD11b. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD11b.

[0113] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 4%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51% 1%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 100%, about 101%, about 102%, about 103%, about 104%, about 105%, about 106%, about 107%, about 108%, about 109%, about 110%, about 111%, about 112%, about 113%, about 114%, about 115%, about 116%, about 117%, about 118%, about 119%, about 120%, about 121%, about 122%, about 123%, about 124%, about 125%, about 126%, about 127%, about 128%, about 129%, about 130%, about 131%, about 132%, about 133%, about 134%, about 135%, about 136%, about 137%, about 138%, about 139%, about 140%, about 141%, about 142%, about 143%, about 14 8%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane containing CD11b.

[0114] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CD11b than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold greater amounts of CD11b than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0115] In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold to about 8-fold or about 2-fold to about 4-fold the amount of CD11b as platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 3-fold the amount of CD11b as platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.1-fold to about 4-fold or about 1.1-fold to about 2-fold the amount of CD11b as platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.5-fold the amount of CD11b as platelet-derived extracellular vesicles (PLT EVs).

[0116] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CD11b than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold greater amounts of CD11b than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0117] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of CD11b than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CD11b than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0118] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, greater than about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21.

[0119] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, about 40% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, about 90% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, about 95% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, about 99% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21.

[0120] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, less than about 1% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21.

[0121] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD21.

[0122] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 4%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51% 1%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 100%, about 101%, about 102%, about 103%, about 104%, about 105%, about 106%, about 107%, about 108%, about 109%, about 110%, about 111%, about 112%, about 113%, about 114%, about 115%, about 116%, about 117%, about 118%, about 119%, about 120%, about 121%, about 122%, about 123%, about 124%, about 125%, about 126%, about 127%, about 128%, about 129%, about 130%, about 131%, about 132%, about 133%, about 134%, about 135%, about 136%, about 137%, about 138%, about 139%, about 140%, about 141%, about 142%, about 143%, about 14 8%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane containing CD21.

[0123] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CD21 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, the megakaryocyte-derived extracellular vesicles have an amount that is about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold greater than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0124] In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold to about 64-fold, about 4-fold to about 32-fold, or about 8-fold to about 16-fold the amount of CD21 as compared to platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 10-fold or about 12-fold the amount of CD21 as compared to platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold to about 8-fold, or about 4-fold to about 8-fold the amount of CD21 as compared to platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have about 4-fold or about 5-fold the amount of CD21 as compared to platelet-derived extracellular vesicles (PLT EVs).

[0125] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of CD21 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CD21 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0126] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, greater than about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51.

[0127] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, about 40% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, about 90% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, about 95% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, about 99% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51.

[0128] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, less than about 1% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51.

[0129] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD51. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD51. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD51. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD51.

[0130] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 4%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51% 1%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 100%, about 101%, about 102%, about 103%, about 104%, about 105%, about 106%, about 107%, about 108%, about 109%, about 110%, about 111%, about 112%, about 113%, about 114%, about 115%, about 116%, about 117%, about 118%, about 119%, about 120%, about 121%, about 122%, about 123%, about 124%, about 125%, about 126%, about 127%, about 128%, about 129%, about 130%, about 131%, about 132%, about 133%, about 134%, about 135%, about 136%, about 137%, about 138%, about 139%, about 140%, about 141%, about 142%, about 143%, about 14 8%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane containing CD51.

[0131] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CD51 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold greater amounts of CD51 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0132] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of CD51 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CD51 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0133] In several embodiments, megakaryocyte-derived extracellular vesicles have about 1.1 to about 1 / 4 or about 1.1 to about 1 / 2 the amount of CD51 as compared to platelet-free plasma (PFP) MkEVs. In several embodiments, megakaryocyte-derived extracellular vesicles have about 1.5 times the amount of CD51 as compared to platelet-free plasma (PFP) MkEVs. In several embodiments, megakaryocyte-derived extracellular vesicles have about 1.1 to about 1 / 4 or about 1.1 to about 1 / 2 the amount of CD51 as compared to platelet-derived extracellular vesicles (PLT EVs). In several embodiments, megakaryocyte-derived extracellular vesicles have about 1.5 times the amount of CD51 as compared to platelet-derived extracellular vesicles (PLT EVs).

[0134] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In embodiments, greater than about 99% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2.

[0135] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, about 40% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, about 90% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, about 95% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, about 99% or less of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2.

[0136] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, less than about 1% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2.

[0137] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CLEC-2. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CLEC-2. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CLEC-2. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CLEC-2. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CLEC-2. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CLEC-2. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CLEC-2. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CLEC-2.

[0138] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 4%, approximately 25%, approximately 26%, approximately 27%, approximately 28%, approximately 29%, approximately 30%, approximately 31%, approximately 32%, approximately 33%, approximately 34%, approximately 35%, approximately 36%, approximately 37%, approximately 38%, approximately 39%, approximately 40%, approximately 41%, approximately 42%, approximately 43%, approximately 44%, approximately 45%, approximately 46%, approximately 47%, approximately 48%, approximately 49%, approximately 50%, approximately 51 %,approximately 52%,approximately 53%,approximately 54%,approximately 55%,approximately 56%,approximately 57%,approximately 58%,approximately 59%,approximately 60%,approximately 61%,approximately 62%,approximately 63%,approximately 64%,approximately 65%,approximately 66%,approximately 67%,approximately 68%,approximately 69%,approximately 70%,approximately 71%,approximately 72%,approximately 73%,approximately 74%,approximately 75%,approximately 76%,approximately 77%,approximately 78 %, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane comprising CLEC-2.

[0139] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CLEC-2 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold greater amounts of CLEC-2 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0140] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of CLEC-2 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CLEC-2 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0141] In several embodiments, megakaryocyte-derived extracellular vesicles have about 2 to about 16 times less, or about 4 to about 8 times less, of CLEC-2 than platelet-free plasma (PFP) MkEVs. In several embodiments, megakaryocyte-derived extracellular vesicles have about 4 to about 1 / 5 times less, or about 1 / 8 to about 1 / 16 times less, of CLEC-2 than platelet-derived extracellular vesicles (PLT EVs). In several embodiments, megakaryocyte-derived extracellular vesicles have about 1 / 4 to about 32 times less, or about 1 / 8 to about 1 / 16 times less, of CLEC-2 than platelet-derived extracellular vesicles (PLT EVs). In several embodiments, megakaryocyte-derived extracellular vesicles have about 1 / 10 or about 1 / 12 times less, or about 1 / 12 times less, of CLEC-2 than platelet-derived extracellular vesicles (PLT EVs).

[0142] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, greater than about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A).

[0143] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, about 40% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, about 90% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, about 95% or less of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, about 99% or less of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A).

[0144] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A). In some embodiments, less than about 1% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A).

[0145] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing LAMP-1 (CD107A). In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing LAMP-1 (CD107A). In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing LAMP-1 (CD107A). In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing LAMP-1 (CD107A). In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing LAMP-1 (CD107A). In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing LAMP-1 (CD107A). In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing LAMP-1 (CD107A). In several embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing LAMP-1 (CD107A). In several embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing LAMP-1 (CD107A). In several embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing LAMP-1 (CD107A).

[0146] In some embodiments, less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, or about 25% of megakaryocyte-derived extracellular vesicles. ,approximately 25%,approximately 26%,approximately 27%,approximately 28%,approximately 29%,approximately 30%,approximately 31%,approximately 32%,approximately 33%,approximately 34%,approximately 35%,approximately 36%,approximately 37%,approximately 38%,approximately 39%,approximately 40%,approximately 41%,approximately 42%,approximately 43%,approximately 44%,approximately 45%,approximately 46%,approximately 47%,approximately 48%,approximately 49%,approximately 50%,approximately 51%,approximately 52%,approximately 53%,approximately 54%,approximately 55%,approximately 56%,approximately 57%,approximately 58%,approximately 59%,approximately 60%,approximately 61%,approximately 62%,approximately 63%,approximately 64%,approximately 65%,approximately 66%,approximately 67%,approximately 68%,approximately 69%,approximately 70%,approximately 71%,approximately 72%,approximately 73%,approximately 74%,approximately 75%,approximately 76%,approximately 77%,approximately 78%,approximately 79%,approximately 80%,approximately 81%,approximately 82%,approximately 83%,approximately 84%,approximately 85%,approximately 86%,approximately 87%,approximately 2%, approximately 53%, approximately 54%, approximately 55%, approximately 56%, approximately 57%, approximately 58%, approximately 59%, approximately 60%, approximately 61%, approximately 62%, approximately 63%, approximately 64%, approximately 65%, approximately 66%, approximately 67%, approximately 68%, approximately 69%, approximately 70%, approximately 71%, approximately 72%, approximately 73%, approximately 74%, approximately 75%, approximately 76%, approximately 77%, approximately 78%, approximately 79% , about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane containing LAMP-1 (CD107A).

[0147] In several embodiments, the megakaryocyte-derived extracellular vesicles are free or substantially free of LAMP-1 (CD107A).

[0148] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of LAMP-1 (CD107A) than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold the amount of LAMP-1 (CD107A) than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0149] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of LAMP-1 (CD107A) than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less LAMP-1 (CD107A) than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0150] In some embodiments, megakaryocyte-derived extracellular vesicles have about one-half to about one-half the amount of LAMP-1 (CD107A) found in platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have substantially the same amount of LAMP-1 (CD107A) found in platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about one-half to about one-eighth, or about one-half to about one-quarter the amount of LAMP-1 (CD107A) found in platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have about one-third or about one-quarter the amount of LAMP-1 (CD107A) found in platelet-derived extracellular vesicles (PLT EVs).

[0151] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, greater than about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63.

[0152] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, about 40% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, about 90% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, about 95% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, about 99% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63.

[0153] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, less than about 1% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63.

[0154] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD63. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD63. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD63. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD63.

[0155] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 4%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51% 1%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 100%, about 101%, about 102%, about 103%, about 104%, about 105%, about 106%, about 107%, about 108%, about 109%, about 110%, about 111%, about 112%, about 113%, about 114%, about 115%, about 116%, about 117%, about 118%, about 119%, about 120%, about 121%, about 122%, about 123%, about 124%, about 125%, about 126%, about 127%, about 128%, about 129%, about 130%, about 131%, about 132%, about 133%, about 134%, about 135%, about 136%, about 137%, about 138%, about 139%, about 140%, about 141%, about 142%, about 143%, about 14 8%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane containing CD63.

[0156] In some embodiments, about 1% to about 30% of megakaryocyte-derived extracellular vesicles contain a lipid bilayer membrane containing CD63. In some embodiments, about 5% to about 25% of megakaryocyte-derived extracellular vesicles contain a lipid bilayer membrane containing CD63. In some embodiments, about 10% to about 20% of megakaryocyte-derived extracellular vesicles contain a lipid bilayer membrane containing CD63. In some embodiments, about 13% to about 19% of megakaryocyte-derived extracellular vesicles contain a lipid bilayer membrane containing CD63.

[0157] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CD63 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold greater amount of CD63 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0158] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of CD63 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CD63 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0159] In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold to about 8-fold or about 2-fold to about 4-fold the amount of CD63 as platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold or about 3-fold the amount of CD63 as platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.1-fold to about 1 / 2-fold the amount of CD63 as platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.1-fold or about 1.2-fold the amount of CD63 as platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have substantially the same amount of CD63 as platelet-derived extracellular vesicles (PLT EVs).

[0160] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, greater than about 99% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b.

[0161] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, about 40% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, about 90% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, about 95% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, about 99% or less of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b.

[0162] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b, and in some embodiments, less than about 1% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b.

[0163] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD42b. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD42b. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD42b. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD42b.

[0164] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 4%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51% 1%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 100%, about 101%, about 102%, about 103%, about 104%, about 105%, about 106%, about 107%, about 108%, about 109%, about 110%, about 111%, about 112%, about 113%, about 114%, about 115%, about 116%, about 117%, about 118%, about 119%, about 120%, about 121%, about 122%, about 123%, about 124%, about 125%, about 126%, about 127%, about 128%, about 129%, about 130%, about 131%, about 132%, about 133%, about 134%, about 135%, about 136%, about 137%, about 138%, about 139%, about 140%, about 141%, about 142%, about 143%, about 14 8%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane comprising CD42b.

[0165] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CD42b than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold greater amount of CD42b than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0166] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of CD42b than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CD42b than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0167] In some embodiments, megakaryocyte-derived extracellular vesicles have about 8 to about 32 times less CD42b than platelet-free plasma (PFP) MkEVs, or about 10 to about 20 times less CD42b than platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 16 to about 20 times less CD42b than platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 64 to about 128 times less CD42b than platelet-derived extracellular vesicles (PLT EVs), or about 50 to about 75 times less CD42b than platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have about 64 to about 70 times less CD42b than platelet-derived extracellular vesicles (PLT EVs).

[0168] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, greater than about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9.

[0169] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, about 40% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, about 90% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, about 95% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, about 99% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9.

[0170] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, less than about 1% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9.

[0171] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9.

[0172] In some embodiments, the megakaryocyte-derived extracellular vesicles are less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, approximately 25%, approximately 26%, approximately 27%, approximately 28%, approximately 29%, approximately 30%, approximately 31%, approximately 32%, approximately 33%, approximately 34%, approximately 35%, approximately 36%, approximately 37%, approximately 38%, approximately 39%, approximately 40%, approximately 41%, approximately 42%, approximately 43%, approximately 44%, approximately 45%, approximately 46%, approximately 47%, approximately 48%, approximately 49%, approximately 50%, approximately 51%, approximately 52%, approximately 53%, approximately 54%, approximately 55%, approximately 56%, approximately 57%, approximately 58%, approximately 59%, approximately 60%, approximately 61%, approximately 62%, approximately 63%, approximately 64%, approximately 65%, approximately 66%, approximately 67%, approximately 68%, approximately 69%, approximately 70%, approximately 71%, approximately 72%, approximately 73%, approximately 74%, approximately 75%, approximately 76%, approximately 77%, approximately 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane comprising CD9.

[0173] In some embodiments, about 50% to about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, about 60% to about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, about 62% to about 68% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9. In some embodiments, about 65% to about 66% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD9.

[0174] In several embodiments, about 50% to about 99% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD9.

[0175] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by a higher expression and / or presence of CD9 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold greater amount of CD9 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0176] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of CD9 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CD9 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0177] In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.5 to about 4 times or about 2 to about 4 times the amount of CD9 as platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about twice the amount of CD9 as platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.1 to about 1 / 2 the amount of CD9 as platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.1 or about 1.2 times the amount of CD9 as platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have substantially the same amount of CD9 as platelet-derived extracellular vesicles (PLT EVs).

[0178] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, greater than about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31.

[0179] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, about 40% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, about 90% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, about 95% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, about 99% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31.

[0180] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, less than about 1% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31.

[0181] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31.

[0182] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 4%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51% 1%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 100%, about 101%, about 102%, about 103%, about 104%, about 105%, about 106%, about 107%, about 108%, about 109%, about 110%, about 111%, about 112%, about 113%, about 114%, about 115%, about 116%, about 117%, about 118%, about 119%, about 120%, about 121%, about 122%, about 123%, about 124%, about 125%, about 126%, about 127%, about 128%, about 129%, about 130%, about 131%, about 132%, about 133%, about 134%, about 135%, about 136%, about 137%, about 138%, about 139%, about 140%, about 141%, about 142%, about 143%, about 14 8%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane containing CD31.

[0183] In some embodiments, about 5% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, about 10% to about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, about 10% to about 35% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31. In some embodiments, about 13% to about 31% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD31.

[0184] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CD31 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold greater amounts of CD31 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0185] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of CD31 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CD31 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0186] In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.1 to about 1 / 4 or about 1.1 to about 1 / 2 the amount of CD31 as found in platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.5 times the amount of CD31 as found in platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 1 / 2 to about 1 / 4 the amount of CD31 as found in platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have about 1 / 2 or about 1 / 3 the amount of CD31 as found in platelet-derived extracellular vesicles (PLT EVs).

[0187] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, greater than about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47.

[0188] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, about 40% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, about 90% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, about 95% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, about 99% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47.

[0189] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, less than about 1% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47.

[0190] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD47. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD47. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD47.

[0191] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 4%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51% 1%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 100%, about 101%, about 102%, about 103%, about 104%, about 105%, about 106%, about 107%, about 108%, about 109%, about 110%, about 111%, about 112%, about 113%, about 114%, about 115%, about 116%, about 117%, about 118%, about 119%, about 120%, about 121%, about 122%, about 123%, about 124%, about 125%, about 126%, about 127%, about 128%, about 129%, about 130%, about 131%, about 132%, about 133%, about 134%, about 135%, about 136%, about 137%, about 138%, about 139%, about 140%, about 141%, about 142%, about 143%, about 14 8%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane containing CD47.

[0192] In some embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD47. In some embodiments, about 10% to about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD47. In some embodiments, about 20% to about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD47. In some embodiments, about 25% to about 35% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD47.

[0193] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CD47 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold greater amount of CD47 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0194] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of CD47 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CD47 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0195] In some embodiments, megakaryocyte-derived extracellular vesicles have about 128 to about 512 times, or about 256 to about 512 times, or about 250 to about 300 times the amount of CD47 as platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 256 or about 300 times the amount of CD47 as platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.1 to about 1 / 2 the amount of CD47 as platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have about 1.1 or about 1.5 times the amount of CD47 as platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have substantially the same amount of CD47 as platelet-derived extracellular vesicles (PLT EVs).

[0196] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, greater than about 99% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147.

[0197] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, about 40% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, about 90% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, about 95% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, about 99% or less of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147.

[0198] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD 147. In some embodiments, less than about 1% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD 147.

[0199] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD147.

[0200] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 4%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51% 1%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 100%, about 101%, about 102%, about 103%, about 104%, about 105%, about 106%, about 107%, about 108%, about 109%, about 110%, about 111%, about 112%, about 113%, about 114%, about 115%, about 116%, about 117%, about 118%, about 119%, about 120%, about 121%, about 122%, about 123%, about 124%, about 125%, about 126%, about 127%, about 128%, about 129%, about 130%, about 131%, about 132%, about 133%, about 134%, about 135%, about 136%, about 137%, about 138%, about 139%, about 140%, about 141%, about 142%, about 143%, about 14 8%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane containing CD147.

[0201] In some embodiments, about 1% to about 15% of megakaryocyte-derived extracellular vesicles contain a lipid bilayer membrane comprising CD147. In some embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles contain a lipid bilayer membrane comprising CD147. In some embodiments, about 3% to about 8% of megakaryocyte-derived extracellular vesicles contain a lipid bilayer membrane comprising CD147. In some embodiments, about 4% to about 7% of megakaryocyte-derived extracellular vesicles contain a lipid bilayer membrane comprising CD147.

[0202] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CD147 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold higher amounts of CD147 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0203] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of CD147 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CD147 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0204] In several embodiments, megakaryocyte-derived extracellular vesicles have about half to about eighth, or about half to about quarter, the amount of CD147 found in platelet-free plasma (PFP) MkEVs. In several embodiments, megakaryocyte-derived extracellular vesicles have about half or about one-third the amount of CD147 found in platelet-free plasma (PFP) MkEVs. In several embodiments, megakaryocyte-derived extracellular vesicles have about 1.1 to about half the amount of CD147 found in platelet-derived extracellular vesicles (PLT EVs). In several embodiments, megakaryocyte-derived extracellular vesicles have about 1.1 or about 1.2 times less CD147 found in platelet-derived extracellular vesicles (PLT EVs). In several embodiments, megakaryocyte-derived extracellular vesicles have substantially the same amount of CD147 as platelet-derived extracellular vesicles (PLT EVs).

[0205] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, greater than about 99% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a.

[0206] In some embodiments, about 50% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, about 40% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, about 60% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, about 70% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, about 80% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, about 90% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, about 95% or less of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, about 99% or less of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a.

[0207] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a, and in some embodiments, less than about 1% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a.

[0208] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD32a. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD32a. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD32a. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing CD32a.

[0209] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 4%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51% 1%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 100%, about 101%, about 102%, about 103%, about 104%, about 105%, about 106%, about 107%, about 108%, about 109%, about 110%, about 111%, about 112%, about 113%, about 114%, about 115%, about 116%, about 117%, about 118%, about 119%, about 120%, about 121%, about 122%, about 123%, about 124%, about 125%, about 126%, about 127%, about 128%, about 129%, about 130%, about 131%, about 132%, about 133%, about 134%, about 135%, about 136%, about 137%, about 138%, about 139%, about 140%, about 141%, about 142%, about 143%, about 14 8%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane containing CD32a.

[0210] In several embodiments, the megakaryocyte-derived extracellular vesicles are free of or substantially free of CD32a.

[0211] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CD32a than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold greater amounts of CD32a than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0212] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of CD32a than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CD32a than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0213] In some embodiments, megakaryocyte-derived extracellular vesicles have about 50- to about 100-fold less, 128- to about 512-fold less, or about 256- to about 512-fold less, or about 250- to about 300-fold less CD32a than platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 250- or about 256-fold less CD32a than platelet-free plasma (PFP) MkEVs. In some embodiments, megakaryocyte-derived extracellular vesicles have about 250- to about 400- or about 256- to about 512-fold less CD32a than platelet-derived extracellular vesicles (PLT EVs). In some embodiments, megakaryocyte-derived extracellular vesicles have about 256- or about 300-fold less CD32a than platelet-derived extracellular vesicles (PLT EVs).

[0214] In some embodiments, greater than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GPVI. In some embodiments, greater than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GPVI. In some embodiments, greater than about 60% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GPVI. In some embodiments, greater than about 70% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GPVIa. In some embodiments, greater than about 80% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GPVI. In some embodiments, greater than about 90% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GPVI. In some embodiments, greater than about 95% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GVPI. In some embodiments, greater than about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GVPI.

[0215] In some embodiments, about 50% or less of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising GVPI. In some embodiments, about 40% or less of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising GVPI. In some embodiments, about 60% or less of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising GVPI. In some embodiments, about 70% or less of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising GVPI. In some embodiments, about 80% or less of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising GVPI. In some embodiments, about 90% or less of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising GVPI. In some embodiments, about 95% or less of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising GVPI. In some embodiments, about 99% or less of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising GVPI.

[0216] In some embodiments, less than about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising GVPI. In some embodiments, less than about 40% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising GVPI. In some embodiments, less than about 30% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising GVPI. In some embodiments, less than about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising GVPI. In some embodiments, less than about 20% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising GVPI. In some embodiments, less than about 15% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising GVPI. In some embodiments, less than about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising GVPI. In some embodiments, less than about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising GVPI. In some embodiments, less than about 1% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising GVPI.

[0217] In several embodiments, about 1% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GVPI. In several embodiments, about 1% to about 50% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GVPI. In several embodiments, about 1% to about 25% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GVPI. In several embodiments, about 1% to about 10% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GVPI. In several embodiments, about 1% to about 5% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GVPI. In several embodiments, about 1% to about 2% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GVPI. In several embodiments, about 50% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GVPI. In some embodiments, about 75% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GVPI. In some embodiments, about 90% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GVPI. In some embodiments, about 95% to about 99% of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane containing GVPI.

[0218] In several embodiments, the megakaryocyte-derived extracellular vesicles are present in an amount of less than about 1%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 4%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51% 1%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 100%, about 101%, about 102%, about 103%, about 104%, about 105%, about 106%, about 107%, about 108%, about 109%, about 110%, about 111%, about 112%, about 113%, about 114%, about 115%, about 116%, about 117%, about 118%, about 119%, about 120%, about 121%, about 122%, about 123%, about 124%, about 125%, about 126%, about 127%, about 128%, about 129%, about 130%, about 131%, about 132%, about 133%, about 134%, about 135%, about 136%, about 137%, about 138%, about 139%, about 140%, about 141%, about 142%, about 143%, about 14 8%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% comprise a lipid bilayer membrane containing GVPI.

[0219] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by a higher expression and / or presence of GPVI than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold the amount of GPVI of naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0220] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of GPVI than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less GPVI than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0221] In some embodiments, megakaryocyte-derived extracellular vesicles have about 8 to about 64 times, or about 16 to about 32 times, the amount of GPVI of platelet-free plasma (PFP) MkEV. In some embodiments, megakaryocyte-derived extracellular vesicles have about 30 times, or about 32 times, the amount of GPVI of platelet-free plasma (PFP) MkEV. In some embodiments, megakaryocyte-derived extracellular vesicles have about 1 / 2 to about 1 / 16 times, or about 1 / 4 to about 1 / 8 times the amount of GPVI of platelet-derived extracellular vesicles (PLT EV). In some embodiments, megakaryocyte-derived extracellular vesicles have about 1 / 4 or about 1 / 5 times the amount of GPVI of platelet-derived extracellular vesicles (PLT EV).

[0222] In some embodiments, megakaryocyte-derived extracellular vesicles are free or substantially free of LAMP-1 (CD107A), hi some embodiments, megakaryocyte-derived extracellular vesicles have less LAMP-1 (CD107A) than naturally occurring megakaryocyte-derived extracellular vesicles and / or vesicles or extracellular vesicles derived from platelets.

[0223] In several embodiments, less than about 20%, or less than about 15%, or less than about 10%, or less than about 5% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A).

[0224] In several embodiments, the megakaryocyte-derived extracellular vesicles are characterized by having CD62P and being free of, or substantially free of, LAMP-1 (CD107A).

[0225] In several embodiments, the megakaryocyte-derived extracellular vesicles are characterized by a megakaryocyte-derived extracellular vesicle population in which less than about 20%, or less than about 15%, or less than about 10%, or less than about 5% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising LAMP-1 (CD107A), and more than about 40%, or more than about 50%, or more than about 60%, or more than about 70%, or more than about 80%, or more than about 90%, or more than about 95%, or more than about 99% comprise a lipid bilayer membrane comprising CD62P.

[0226] In several embodiments, less than about 70%, or less than about 60%, or less than about 50%, less than about 40%, less than about 30%, less than about 20%, less than about 15%, less than about 10%, less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising phosphatidylserine (PS).

[0227] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of phosphatidylserine (PS) than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less phosphatidylserine (PS) than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles.

[0228] In several embodiments, the megakaryocyte-derived extracellular vesicles are characterized as being free or substantially free of phosphatidylserine (PS).

[0229] In several embodiments, the megakaryocyte-derived extracellular vesicles are characterized as a megakaryocyte-derived extracellular vesicle population in which less than about 50%, less than about 40%, less than about 30%, less than about 20%, less than about 15%, less than about 10%, less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising phosphatidylserine (PS), and more than about 40%, more than about 50%, more than about 60%, more than about 70%, more than about 80%, more than about 90%, more than about 95%, or more than about 99% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD47.

[0230] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by higher expression and / or presence of CD41 than naturally occurring megakaryocyte-derived extracellular vesicles and / or vesicles or extracellular vesicles derived from platelets. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold more CD41 than naturally occurring megakaryocyte-derived extracellular vesicles and / or vesicles or extracellular vesicles derived from platelets.

[0231] In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by lower expression and / or presence of CD41 than naturally occurring megakaryocyte-derived extracellular vesicles and / or vesicles or extracellular vesicles derived from platelets. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, or about 10-fold, or about 50-fold, or about 100-fold, or about 300-fold, or about 500-fold, or about 1000-fold less CD41 than naturally occurring megakaryocyte-derived extracellular vesicles and / or vesicles or extracellular vesicles derived from platelets.

[0232] In several embodiments, the megakaryocyte-derived extracellular vesicles comprise full-length filamin A.

[0233] In some embodiments, the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising phosphatidylserine. In some embodiments, the megakaryocyte-derived extracellular vesicles are characterized by a population of megakaryocyte-derived extracellular vesicles, of which more than about 40%, more than about 50%, more than about 60%, more than about 70%, more than about 80%, more than about 90%, more than about 95%, or more than about 99% comprise a lipid bilayer membrane comprising phosphatidylserine.

[0234] In some embodiments, megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane that is positive for annexin V. For example, annexin V, which interacts with phosphatidylserine (PS), can be used as a surrogate for the expression and / or presence or absence of phosphatidylserine. In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by a megakaryocyte-derived extracellular vesicle population in which more than about 40%, more than about 50%, more than about 60%, more than about 70%, more than about 80%, more than about 90%, or more than about 95% are positive for PS.

[0235] In some embodiments, substantially all of the megakaryocyte-derived extracellular vesicles in the population comprise two, three, four, five, six, seven, or eight of phosphatidylserine (PS), CD62P, LAMP-1 (CD107a), CD42b, CD9, CD43, CD31, and CD11b. In some embodiments, substantially all of the megakaryocyte-derived extracellular vesicles in the population comprise two, three, or four of PS, CD62P, CD9, and CD11b. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, 10-fold, 50-fold, 100-fold, 300-fold, 500-fold, or 1000-fold greater amounts of one or more of phosphatidylserine (PS), CD62P, LAMP-1 (CD107a), CD42b, CD9, CD43, CD31, and CD11b than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by not expressing significant amounts of DRAQ5. In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by being substantially free of DRAQ5.

[0236] In some embodiments, substantially all of the megakaryocyte-derived extracellular vesicles in the population comprise two, three, four, five, or six of phosphatidylserine (PS), CD61, CD62P, LAMP-1 (CD107a), CLEC-2, and CD63. In some embodiments, substantially all of the megakaryocyte-derived extracellular vesicles in the population comprise two or three of PS, CD61, and CD63. In some embodiments, substantially all of the megakaryocyte-derived extracellular vesicles in the population comprise phosphatidylserine (PS) and CD61. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, 10-fold, 50-fold, 100-fold, 300-fold, 500-fold, or 1000-fold greater amounts of one or more of phosphatidylserine (PS), CD61, CD62P, LAMP-1 (CD107a), CLEC-2, and CD63 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by not expressing significant amounts of DRAQ5. In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by being substantially free of DRAQ5.

[0237] In some embodiments, substantially all of the megakaryocyte-derived extracellular vesicles in the population comprise two, three, four, five, six, seven, or eight of phosphatidylserine (PS), CD62P, CLEC-2, CD9, CD31, CD147, CD32a, and GPVI. In some embodiments, substantially all of the megakaryocyte-derived extracellular vesicles in the population comprise two, three, or four of phosphatidylserine (PS), CD9, CD31, and CD147. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, 10-fold, 50-fold, 100-fold, 300-fold, 500-fold, or 1000-fold greater amounts of one or more of phosphatidylserine (PS), CD62P, CLEC-2, CD9, CD31, CD147, CD32a, and GPVI than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by not expressing significant amounts of DRAQ5. In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by being substantially free of DRAQ5.

[0238] In some embodiments, substantially all of the megakaryocyte-derived extracellular vesicles in the population comprise two, three, four, five, or six of phosphatidylserine (PS), CD62P, LAMP-1 (CD107a), CLEC-2, CD9, and CD31. In some embodiments, substantially all of the megakaryocyte-derived extracellular vesicles in the population comprise two or three of phosphatidylserine (PS), CD62P, and CD9. In some embodiments, substantially all of the megakaryocyte-derived extracellular vesicles in the population comprise PS and CD9. In some embodiments, megakaryocyte-derived extracellular vesicles have about 2-fold, 10-fold, 50-fold, 100-fold, 300-fold, 500-fold, or 1000-fold greater amounts of one or more of phosphatidylserine (PS), CD62P, LAMP-1 (CD107a), CLEC-2, CD9, and CD31 than naturally occurring megakaryocyte-derived extracellular vesicles, vesicles or extracellular vesicles derived from platelets, such as platelet-derived extracellular vesicles (PLT-EVs), and / or platelet-free plasma (PPF) megakaryocyte-derived extracellular vesicles. In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by not expressing significant amounts of DRAQ5. In some embodiments, megakaryocyte-derived extracellular vesicles are characterized by being substantially free of DRAQ5.

[0239] In some embodiments, the megakaryocyte-derived extracellular vesicles and / or the plurality of megakaryocyte-derived extracellular vesicles and / or populations of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane; less than about 50%, or less than about 40%, or less than about 30%, or less than about 20%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles contain lipid bilayer membranes that contain CD54; and / or less than about 50%, or less than about 40%, or less than about 30%, or less than about 20%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles contain lipid bilayer membranes that contain CD18; and / or less than about 50%, or less than about 40%, or less than about 30%, or less than about 20%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles contain a lipid bilayer membrane that contains CD43; and / or less than about 50%, or less than about 40%, or less than about 30%, or less than about 20%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles contain lipid bilayer membranes that contain CD11b; and / or less than about 50%, or less than about 40%, or less than about 30%, or less than about 20%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles contain lipid bilayer membranes that contain CD62P; and / or greater than about 40%, or greater than about 50%, or greater than about 60%, or greater than about 70%, or greater than about 80%, or greater than about 90%, or greater than about 95%, or greater than about 99% of the megakaryocyte-derived extracellular vesicles contain lipid bilayer membranes that contain CD41; and / or less than about 50%, or less than about 40%, or less than about 30%, or less than about 20%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles contain a lipid bilayer membrane that contains CD21; and / or less than about 50%, or less than about 40%, or less than about 30%, or less than about 20%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles contain lipid bilayer membranes that contain CD51; and / or greater than about 40%, or greater than about 50%, or greater than about 60%, or greater than about 70%, or greater than about 80%, or greater than about 90%, or greater than about 95%, or greater than about 99% of the megakaryocyte-derived extracellular vesicles contain lipid bilayer membranes that contain CD61; and / or less than about 50%, or less than about 40%, or less than about 30%, or less than about 20%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles contain a lipid bilayer membrane that contains CD147; and / or less than about 50%, or less than about 40%, or less than about 30%, or less than about 20%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles contain a lipid bilayer membrane that contains CD31; and / or less than about 50%, or less than about 40%, or less than about 30%, or less than about 20%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles contain a lipid bilayer membrane that contains CD47; and / or less than about 50%, or less than about 40%, or less than about 30%, or less than about 20%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles contain lipid bilayer membranes that contain CD32a; and / or greater than about 40%, or greater than about 50%, or greater than about 60%, or greater than about 70%, or greater than about 80%, or greater than about 90%, or greater than about 95%, or greater than about 99% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane that contains CD9; and / or less than about 50%, or less than about 40%, or less than about 30%, or less than about 20%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles contain a lipid bilayer membrane that contains CLEC-2; and / or less than about 50%, or less than about 40%, or less than about 30%, or less than about 20%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles contain lipid bilayer membranes containing LAMP-1 (CD107a); and / or less than about 50%, or less than about 40%, or less than about 30%, or less than about 20%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles contain lipid bilayer membranes that contain CD24b; and / or less than about 50%, or less than about 40%, or less than about 30%, or less than about 20%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles contain lipid bilayer membranes containing GVPI; and / or less than about 50%, or less than about 40%, or less than about 30%, or less than about 20%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles contain a lipid bilayer membrane that contains CD63; and / or Less than about 50%, or less than about 40%, or less than about 30%, or less than about 20%, or less than about 10%, or less than about 5%, or less than about 1% of the megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising phosphatidylserine (PS). In several embodiments, more than about 40%, more than about 50%, or more than about 60%, or more than about 70%, or more than about 80%, or more than about 90%, more than about 95%, or more than about 99% of the megakaryocyte-derived extracellular vesicles and / or a population of megakaryocyte-derived extracellular vesicles comprise a lipid bilayer membrane comprising CD41.

[0240] Nucleic Acid Biomarkers In embodiments, the compositions contain one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175, or at least 500, or at least 1000, or at least 1500, or at least 2000, or at least 2500) that are unique to or preferentially present in megakaryocyte-derived extracellular vesicles compared to other types of extracellular vesicles, or one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least or at least 100, or at least 150, or at least 175, or at least 500, or at least 1000, or at least 1500, or at least 2000, or at least 2500), wherein one or more nucleic acids are listed in Table 6 below. [Table 1-1] [Table 1-2] [Table 1-3] [Table 1-4] [Table 1-5]

Table 1-6

Table 1-7

Table 1-8

Table 1-9

[0241] In several embodiments, the megakaryocyte-derived extracellular vesicles of the present disclosure contain one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) that are unique to or preferentially present in the megakaryocyte-derived extracellular vesicles of the present disclosure compared to other types of extracellular vesicles. One or more proteins (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) encoded by the nucleotide sequence (e.g., at least 25, or at least 50, or at least 100, or at least 150, or at least 175) are involved in neutrophil extracellular trap formation, and optionally, H3-3A, H4C12, H2AC18, H2AC13, H2AC19 , H3-3B, H2BC12, H4C3, H3C10, H2AC14, H2BC4, H2AC20, H3C13, H4C14, H3C2, H2AC8, H2BC3, H3C7, H2AC17, H3C4, H2BC11, SELP, H3C11, H4C2, H2AC15, H 2AC12, H3C1, H4C4, H2AC4, H4C13, H2BC5, H2AC7, H2AC16, H2BC17, H2AJ, H3C3, H2BC10, H2AC21, H3C14, H3C15, H2AC11, H3C12, FPR1, H2BC13, H4C1, MAC selected from ROH2A1, H2BC14, H2AX, H2BC6, NCF4, H4C6, H2BC15, H2BC21, H2BC7, H2BC8, CYBB, FCGR1A, H2AZ2, H4C5, AQP9, H2BC9, CLCN4, FPR2, H4C9, HDAC9, ITGAL, FCGR3B, ITGAM, H3C8, TLR2, NCF1, NCF2, H3C6, CAMP, MPO, ELANE, FCGR3A, AZU1, H2AW, SIGLEC9, CLEC7A, PLCB4, CASP1, H2BU1, TLR4, and TLR8.

[0242] In several embodiments, the megakaryocyte-derived extracellular vesicles of the present disclosure contain one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) that are unique to or preferentially present in megakaryocyte-derived extracellular vesicles of the present disclosure compared to other types of extracellular vesicles, or one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least One or more proteins (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) encoded by the neutrophil extracellular traps (e.g., at least 50, or at least 100, or at least 150, or at least 175) are involved in neutrophil extracellular trap formation, hematopoietic cell lineage, cell adhesion, ATP-binding cassette (ABC) transport, chemokine signaling pathway, phagocytosis, and / or aminoacyl-tRNA biosynthesis.

[0243] In embodiments, one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175), or one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, if One or more proteins (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) encoded by the nucleotide sequences (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) are involved in neutrophil extracellular trap formation, and optionally include H3-3A, H4C12, H2AC18, H2AC13, H2AC19, H3-3B, H2BC12, H4C3, H3C10, H2AC14, H 2BC4, H2AC20, H3C13, H4C14, H3C2, H2AC8, H2BC3, H3C7, H2AC17, H3C4, H2BC11, SELP, H3C11, H4C2, H2AC15, H2AC12, H3C1, H4C4, H2AC4, H4C 13, H2BC5, H2AC7, H2AC16, H2BC17, H2AJ, H3C3, H2BC10, H2AC21, H3C14, H3C15, H2AC11, H3C12, FPR1, H2BC13, H4C1, MACROH2A1, H2BC14, H2 AX, H2BC6, NCF4, H4C6, H2BC15, H2BC21, H2BC7, H2BC8, CYBB, FCGR1A, H2AZ2, H4C5, AQP9, H2BC9, CLCN4, FPR2, H4C9, HDAC9, ITGAL, FCGR3B, ITGAM, H3C8, TLR2, NCF1, NCF2, H3C6, CAMP, MPO, ELANE, FCGR3A, AZU1, H2AW, SIGLEC9, CLEC7A, PLCB4, CASP1, H2BU1, TLR4, and TLR8.

[0244] In embodiments, one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) or one or more proteins (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) encoded by one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) are used. or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) are involved in hematopoietic cell lineages and are optionally selected from CD36, IL9R, FCGR1A, CSF3R, KIT, GP5, CSF1, ITGAM, CD33, IL1B, IL7R, IL6R, HLA-DPB1, CD24, CD22, GYPA, MME, ITGA1, CD1C, IL3RA, HLA-DPA1, IL1R2, CD34, HLA-DRA, HLA-DQA1, and CD7.

[0245] In embodiments, one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) or one or more (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) encoded by one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) proteins involved in cell adhesion and are optionally selected from CD226, SELP, PECAM1, VSIR, PTPRC, NRXN1, NECTIN2, ITGAL, ITGAM, SELL, CD28, HLA-DPB1, CNTNAP2, CLDN5, JAM2, CD40LG, LRRC4, CD274, CD40, CD276, CD22, ITGA9, NECTIN3, NEGR1, HLA-DPA1, CNTN1, ITGB8, CD34, HLA-DRA, SIGLEC1, and HLA-DQA1.

[0246] In embodiments, one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) or one or more proteins (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) encoded by the one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) are involved in ATP-binding cassette (ABC) transport and are optionally selected from CFTR, ABCD4, ABCA5, ABCA9, ABCC1, ABCB8, ABCA8, ABCB9, ABCC9, ABCA1, ABCA10, and ABCC8.

[0247] In embodiments, one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) or one or more proteins (e.g., at least 3, or at least 5, or at least 10, or at least 150, or at least 175) encoded by one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) are used. at least 25, or at least 50, or at least 100, or at least 150, or at least 175) are involved in the Rap1 signaling pathway and are optionally selected from F2R, LCP2, FYB1, FPR1, F2RL3, P2RY1, RAPGEF2, KIT, EGF, CSF1, ANGPT1, PDGFC, ITGAL, VAV1, VAV3, PDGFRA, ITGAM, RAPGEF5, MAGI2, FARP2, CTNND1, FGF23, FLT1, FGF1, DRD2, AFDN, PLCB4, APBB1IP, PLCE1, MAGI3, KDR, LPAR3, GRIN2B, ADCY1, FGF7, ANGPT4, and TEK.

[0248] In embodiments, one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) or one or more proteins (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) encoded by one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) are used. or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) are involved in the chemokine signaling pathway and are optionally selected from PF4, CCL5, CXCL2, CXCL8, ELMO1, GRK2, GNG8, CCR4, PF4V1, VAV1, VAV3, PIK3R6, PIK3CG, SHC4, GRK3, NCF1, HCK, GRK4, ITK, GNG2, CXCR2, CCR1, CXCL5, CCL3, PLCB4, CXCL6, CCL22, CCL8, CCL4L2, CXCL12, GNGT1, CXCL11, ADCY1, and XCL1.

[0249] In embodiments, one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) or one or more proteins (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) encoded by one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) are used. or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) are involved in phagocytosis and are optionally selected from TUBA8, NCF4, CD36, CYBB, FCGR1A, CTSS, FCGR3B, PIKFYVE, ITGAM, TLR2, NCF1, NCF2, HLA-DPB1, MPO, EEA1, MSR1, FCGR3A, FCAR, DYNC1I1, CLEC7A, FCGR2B, HLA-DPA1, ATP6V0D2, MRC2, TLR4, TLR6, HLA-DRA, MARCO, and HLA-DQA1.

[0250] In embodiments, one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) or encoded by one or more nucleic acids (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) One or more proteins (e.g., at least 3, or at least 5, or at least 10, or at least 25, or at least 50, or at least 100, or at least 150, or at least 175) involved in various pathways, optionally including H3-3A, H4C12, H2AC18, H2AC13, H2AC19, H3-3B, H2BC12, H4C3, H3C10, H2AC14, H2BC4, H2AC20, H3C13, H4C14, H3C2, H2AC8, H2BC3, H3C7, H2AC 17, H3C4, H2BC11, SELP, H3C11, H4C2, H2AC15, H2AC12, H3C1, H4C4, H2AC4, H4C13, H2BC5, H2AC7, H2AC16, H2BC17, H2AJ, H3C3, H2BC10, H2AC21, H3C14, H 3C15, H2AC11, H3C12, FPR1, H2BC13, H4C1, MACROH2A1, H2BC14, H2AX, H2BC6, NCF4, H4C6, H2BC15, H2BC21, H2BC7, H2BC8, CYBB, FCGR1A, H2AZ2, H4C5, AQP 9, H2BC9, CLCN4, FPR2, H4C9, HDAC9, ITGAL, FCGR3B, ITGAM, H3C8, TLR2, NCF1, NCF2, H3C6, CAMP, MPO, ELANE, FCGR3A, AZU1, H2AW, SIGLEC9, CLEC7A, PLCB 4, CASP1, H2BU1, TLR4, TLR8, CD36, IL9R, FCGR1A, CSF3R, KIT, GP5, CSF1, ITGAM, CD33, IL1B, IL7R, IL6R, HLA-DPB1, CD24, CD22, GYPA, MME, ITGA1, CD1C,IL3RA、HLA-DPA1、IL1R2、CD34、HLA-DRA、HLA-DQA1、CD7、CD226、SELP、PECAM1、VSIR、PTPRC、NRXN1、NECTIN2、ITGAL、ITGAM、SELL、CD28、HLA-DPB1、CN TNAP2、CLDN5、JAM2、CD40LG、LRRC4、CD274、CD40、CD276、CD22、ITGA9、NECTIN3、NEGR1、HLA-DPA1、CNTN1、ITGB8、CD34、HLA-DRA、SIGLEC1、HLA-DQA1、C FTR、ABCD4、ABCA5、ABCA9、ABCC1、ABCB8、ABCA8、ABCB9、ABCC9、ABCA1、ABCA10、ABCC8、F2R、LCP2、FYB1、FPR1、F2RL3、P2RY1、RAPGEF2、KIT、EGF、CSF1、 ANGPT1、PDGFC、ITGAL、VAV1、VAV3、PDGFRA、ITGAM、RAPGEF5、MAGI2、FARP2、CTNND1、FGF23、FLT1、FGF1、DRD2、AFDN、PLCB4、APBB1IP、PLCE1、MAGI3、KDR 、LPAR3、GRIN2B、ADCY1、FGF7、ANGPT4、TEK、PF4、CCL5、CXCL2、CXCL8、ELMO1、GRK2、GNG8、CCR4、PF4V1、VAV1、VAV3、PIK3R6、PIK3CG、SHC4、GRK3、NCF1、 HCK、GRK4、ITK、GNG2、CXCR2、CCR1、CXCL5、CCL3、PLCB4、CXCL6、CCL22、CCL8、CCL4L2、CXCL12、GNGT1、CXCL11、ADCY1、XCL1、TUBA8、NCF4、CD36、CYBB、FC GR1A, CTSS, FCGR3B, PIKFYVE, ITGAM, TLR2, NCF1, NCF2, HLA-DPB1, MPO, EEA1, MSR1, FCGR3A, FCAR, DYNC1I1, CLEC7A, FCGR2B, HLA-DPA1, ATP6V0D2, MRC2, TLR4, TLR6, HLA-DRA, MARCO, HLA-...

Claims

1. A composition comprising: a plurality of substantially purified megakaryocyte-derived extracellular vesicles derived from human pluripotent stem cells, the composition comprising one or more proteins, or one or more nucleic acids encoding the one or more proteins, that are unique to or preferentially present in the megakaryocyte-derived extracellular vesicles compared to other types of extracellular vesicles; the one or more proteins or the one or more nucleic acids involved in aminoacyl-tRNA biosynthesis and selected from the group consisting of YARS1, AARS1, GARS1, LARS1, EPRS1, TARS1, DARS1, WARS1, VARS1, NARS1, RARS1, SARS1, KARS1, IARS1, CARS1, QRS, and HARS1; involved in neutrophil extracellular trap formation and selected from the group consisting of H4C1, H2AC20, H3C1, H2BC12, H3C15, MACROH2A1, H2AX, H2AZ2, H3-3A, H2AC14, and MACROH2A2; or H4C16、H2AC17、H3C13、RPL9P9、PHB1、 SETSIP、H2AZ1、XP32、COPS9、SEC61A2 、MARS1、MRE11、ATP5F1C、H4C1、H2AC2 0、H3C1、H2BC12、FCSK、CAVIN2、YARS1 、㼨㼓㼣㼑㼕、㼭㼡㼣㼲㼯㼨㼒㼡㼑、㼨㼒㼡㼸、㼨㼓㼍㼒、㼳㼥㼬㼥㼮㼯㼦、㼭㼰㼩㼧㼖㼢、セプチン㼗、㼡㼡㼲㼳㼑、㼨㼑㼍、㼨㼑㼍2、H2AZ2、GARS1、LARS1、THPO、RACK1、H3-3A、CCDC191、H2BC19P、プチン2、EPRS 1、TARS1、ATP5F1B、ERBIN、DARS1、WAR S1、VARS1、NIBAN2、セプチン6、NARS1、RAR S1、セプチン11、セプチン5、SARS1、NIBAN1、SN RPGP15、PIP4P2、CYRIB、CARMIL1、KARS 1、IARS1、セプチン9、H1-4、ARHGAP45、H1- 0、CARS1、GCN1、FADS2、TBC1D13、GET3 、RO60、LAMTOR5、ELOC、H2AC14、SCARF 1、RNF24、GCSAML、NRDC、ECPAS、MACROH 2A2、ATP5F1A、DMTN、TANGO2、CSF2RB、 WASHC5、KCNA3、QARS1、MINDY1、PTPA、 EXOC3L2、PRUNE1、PLPBP、THUMPD1、WA SHC4、NECTIN2、GFUS、ADGRE2、AKAP8L 、FAM234A、ADSS2、ANKRD13D、KCT2、NT 5C3A、PIP4P1、CCDC9、ELOB、HPGDS、ME AK7、TOMM70、SMIM1、HARS1、ATP5PD、OGA、CHSY1、SQLE、SUSD6、IGKV1-27、PED S1-UBE2V1、CEP44、MYG1、NRROS、IL21 R、GRK2、MCEMP1、ELAPOR2、SDAD1、CER T1、UBE2F、CALHM5、H1-10、EIPR1、PBD C1、ARMH3、VIPAS39、MESD、PROSER2、RA BL6, FYB1, C17R49, RMDN3, KYAT3, TTMP, EROO1A, CD244, CZIB, BLAP3, BAA M2、H1-3、NAXE、ENSA、PALS2、GUCY1B1、UMAD1、MIX23、PRXL2B、SNU13、RTRAF、The composition is selected from the group consisting of KXD1, VSIR, EPOR, and MARCHHF2.

2. The composition described in claim 1, wherein the one or more proteins or the one or more nucleic acids encoding the one or more proteins are selected from the group consisting of CAVIN2, MPIG6B, ERBIN, ELOC, CSF2RB, KCNA3, NECTIN2, IL21R, MCEMP1, PROSER2, FYB1, CD244 and EPOR.

3. Each megakaryocyte-derived extracellular vesicle comprises a lipid bilayer membrane surrounding a lumen, the lumen comprises one or more megakaryocyte-derived nucleic acid molecules selected from the group consisting of mRNA, tRNA, rRNA, siRNA, microRNA, regulatory RNA, and non-coding and coding RNA; The lipid bilayer membrane contains one or more proteins that are unique to megakaryocyte-derived extracellular vesicles or that are preferentially present in megakaryocyte-derived extracellular vesicles compared to other types of extracellular vesicles. The composition according to claim 1 or claim 2.

4. The composition described in claim 1 or claim 2, wherein the megakaryocyte-derived extracellular vesicles have a diameter in the range of 100 nm to 600 nm.

5. The composition described in claim 1 or claim 2, wherein the megakaryocyte-derived extracellular vesicles do not contain autologous DNA, megakaryocytes and / or platelets.

6. A composition described in claim 1 or claim 2, wherein the megakaryocyte-derived extracellular vesicles are suitable for loading cargo into the lumen and / or loading cargo associated with the surface of the megakaryocyte-derived extracellular vesicles.

7. The composition described in claim 6, wherein the cargo is one or more therapeutic agents.

8. The composition of claim 7, wherein the therapeutic agent is a nucleic acid therapeutic agent selected from one or more non-self nucleic acid constructs and / or recombinant nucleic acid constructs selected from mRNA, tRNA, rRNA, siRNA, microRNA, regulatory RNA, non-coding RNA and coding RNA, linear DNA, DNA fragments, or DNA plasmids.

9. The composition described in claim 7, wherein the therapeutic agent is a biological therapeutic agent.

10. The composition of claim 9, wherein the biological therapeutic agent is a recombinant protein.

11. The composition described in claim 7, wherein the therapeutic agent is a small molecule therapeutic agent.

12. A therapeutic agent comprising a plurality of substantially purified megakaryocyte-derived extracellular vesicles for use in treating a patient in need thereof, said treatment comprising: (a) obtaining megakaryocyte-derived extracellular vesicles according to claim 7; (b) incubating the megakaryocyte-derived extracellular vesicles with a therapeutic agent to allow the therapeutic agent to be imported into the lumen of the megakaryocyte-derived extracellular vesicles and / or associated with the surface of the megakaryocyte-derived extracellular vesicles, thereby producing a deliverable therapeutic agent; and (c) using the deliverable therapeutic agent to treat a patient, or contacting the deliverable therapeutic agent with biological cells in vitro and using the contacted biological cells to treat a patient; A therapeutic agent comprising:

13. A composition comprising a plurality of substantially purified megakaryocyte-derived extracellular vesicles derived from human pluripotent stem cells, comprising: the composition comprises one or more proteins encoded by one or more nucleic acids that are unique to or preferentially present in the megakaryocyte-derived extracellular vesicles compared to other types of extracellular vesicles; 10. A composition wherein the one or more proteins are selected from the group set forth in Table 6.

14. The one or more nucleic acids, Involved in neutrophil extracellular trap formation, H3-3A, H4C12, H2AC18, H2AC13, H2AC19, H3-3B, H2BC12, H4C3 , H3C10, H2AC14, H2BC4, H2AC20, H3C13, H4C14, H3C2, H2AC8, H2BC3, H3C7, H2AC17 , H3C4, H2BC11, SELP, H3C11, H4C2, H2AC15, H2AC12, H3C1, H4C4, H2AC4, H4C13, H2 BC5, H2AC7, H2AC16, H2BC17, H2AJ, H3C3, H2BC10, H2AC21, H3C14, H3C15, H2AC11, H 3C12, FPR1, H2BC13, H4C1, MACROH2A1, H2BC14, H2AX, H2BC6, NCF4, H4C6, H2BC15, H2BC21, H2BC7, H2BC8, CYBB, FCGR1A, H2AZ2, H4C5, AQP9, H2BC9, CLCN4, FPR2, H4C 9, HDAC9, ITGAL, FCGR3B, ITGAM, H3C8, TLR2, NCF1, NCF2, H3C6, CAMP, MPO, ELANE, FCGR3A, AZU1, H2AW, SIGLEC9, CLEC7A, PLCB4, CASP1, H2BU1, TLR4, and TLR8; committed to the hematopoietic cell lineage and selected from CD36, IL9R, FCGR1A, CSF3R, KIT, GP5, CSF1, ITGAM, CD33, IL1B, IL7R, IL6R, HLA-DPB1, CD24, CD22, GYPA, MME, ITGA1, CD1C, IL3RA, HLA-DPA1, IL1R2, CD34, HLA-DRA, HLA-DQA1 and CD7; involved in cell adhesion and selected from CD226, SELP, PECAM1, VSIR, PTPRC, NRXN1, NECTIN2, ITGAL, ITGAM, SELL, CD28, HLA-DPB1, CNTNAP2, CLDN5, JAM2, CD40LG, LRRC4, CD274, CD40, CD276, CD22, ITGA9, and NECTIN3; involved in ATP-binding cassette (ABC) transport and selected from CFTR, ABCD4, ABCA5, ABCA9, ABCC1, ABCB8, ABCA8, ABCB9, ABCC9, ABCA1, ABCA10, and ABCC8; involved in the Rap1 signaling pathway and selected from F2R, LCP2, FYB1, FPR1, F2RL3, P2RY1, RAPGEF2, KIT, EGF, CSF1, ANGPT1, PDGFC, ITGAL, VAV1, VAV3, PDGFRA, ITGAM, RAPGEF5, MAGI2, FARP2, CTNND1, FGF23, FLT1, FGF1, DRD2, AFDN, PLCB4, APBB1IP, PLCE1, MAGI3, KDR, LPAR3, GRIN2B, ADCY1, FGF7, ANGPT4 and TEK; involved in the chemokine signaling pathway and selected from PF4, CCL5, CXCL2, CXCL8, ELMO1, GRK2, GNG8, CCR4, PF4V1, VAV1, VAV3, PIK3R6, PIK3CG, SHC4, GRK3, NCF1, HCK, GRK4, ITK, GNG2, CXCR2, CCR1, CXCL5, CCL3, PLCB4, CXCL6, CCL22, CCL8, CCL4L2, CXCL12, GNGT1, CXCL11, ADCY1 and XCL1; involved in phagocytosis and selected from TUBA8, NCF4, CD36, CYBB, FCGR1A, CTSS, FCGR3B, PIKFYVE, ITGAM, TLR2, NCF1, NCF2, HLA-DPB1, MPO, EEA1, MSR1, FCGR3A, FCAR, DYNC1I1, CLEC7A, FCGR2B, HLA-DPA1, ATP6V0D2, MRC2, TLR4, TLR6, HLA-DRA, MARCO and HLA-DQA1; involved in aminoacyl-tRNA biosynthesis and selected from RARS1, DARS1, KARS1, SARS1, AARS1, TARS1, NARS1, WARS1, GATB, CARS1, EPRS1, TARS3, EARS2, IARS1, QRSL1 and WARS2; or H4C16、H2AC17、H3C13、RPL9P9、PHB1、 SETSIP、H2AZ1、XP32、COPS9、SEC61A2 、MARS1、MRE11、ATP5F1C、H4C1、H2AC2 0、H3C1、H2BC12、FCSK、CAVIN2、YARS1 、㼨㼓㼣㼑㼕、㼭㼡㼣㼲㼯㼨㼒㼡㼑、㼨㼒㼡㼸、㼨㼓㼍㼒、㼳㼥㼬㼥㼮㼯㼦、㼭㼰㼩㼧㼖㼢、セプチン㼗、㼡㼡㼲㼳㼑、㼨㼑㼍、㼨㼑㼍2、H2AZ2、GARS1、LARS1、THPO、RACK1、H3-3A、CCDC191、H2BC19P、プチン2、EPRS 1、TARS1、ATP5F1B、ERBIN、DARS1、WAR S1、VARS1、NIBAN2、セプチン6、NARS1、RAR S1、セプチン11、セプチン5、SARS1、NIBAN1、SN RPGP15、PIP4P2、CYRIB、CARMIL1、KARS 1、IARS1、セプチン9、H1-4、ARHGAP45、H1- 0、CARS1、GCN1、FADS2、TBC1D13、GET3 、RO60、LAMTOR5、ELOC、H2AC14、SCARF 1、RNF24、GCSAML、NRDC、ECPAS、MACROH 2A2、ATP5F1A、DMTN、TANGO2、CSF2RB、 WASHC5、KCNA3、QARS1、MINDY1、PTPA、 EXOC3L2、PRUNE1、PLPBP、THUMPD1、WA SHC4、NECTIN2、GFUS、ADGRE2、AKAP8L 、FAM234A、ADSS2、ANKRD13D、KCT2、NT 5C3A、PIP4P1、CCDC9、ELOB、HPGDS、ME AK7、TOMM70、SMIM1、HARS1、ATP5PD、OGA、CHSY1、SQLE、SUSD6、IGKV1-27、PED S1-UBE2V1、CEP44、MYG1、NRROS、IL21 R、GRK2、MCEMP1、ELAPOR2、SDAD1、CER T1、UBE2F、CALHM5、H1-10、EIPR1、PBD C1、ARMH3、VIPAS39、MESD、PROSER2、RA BL6, FYB1, C17R49, RMDN3, KYAT3, TTMP, EROO1A, CD244, CZIB, BLAP3, BAA M2、H1-3、NAXE、ENSA、PALS2、GUCY1B1、UMAD1、MIX23、PRXL2B、SNU13、RTRAF、The composition of claim 13, wherein the IL-16 is selected from KXD1, VSIR, EPOR, and MARCHHF2.

15. The composition described in claim 13 or claim 14, wherein the megakaryocyte-derived extracellular vesicles have a diameter in the range of 100 nm to 600 nm.

16. The composition described in claim 13 or claim 14, wherein the megakaryocyte-derived extracellular vesicles do not contain autologous DNA, megakaryocytes and / or platelets.

17. The composition described in claim 13 or claim 14, wherein the megakaryocyte-derived extracellular vesicles are suitable for loading cargo into the lumen and / or loading cargo associated with the surface of the megakaryocyte-derived extracellular vesicles.

18. The composition described in claim 17, wherein the cargo is one or more therapeutic agents.

19. The composition of claim 18, wherein the therapeutic agent is a nucleic acid therapeutic agent selected from one or more non-self nucleic acid constructs and / or recombinant nucleic acid constructs selected from mRNA, tRNA, rRNA, siRNA, microRNA, regulatory RNA, non-coding RNA and coding RNA, linear DNA, DNA fragments, or DNA plasmids.

20. The composition of claim 18, wherein the therapeutic agent is a biological therapeutic agent.

21. The composition of claim 20, wherein the biological therapeutic agent is a recombinant protein.

22. The composition described in claim 18, wherein the therapeutic agent is a small molecule therapeutic agent.

23. A therapeutic agent comprising a plurality of substantially purified megakaryocyte-derived extracellular vesicles for use in treating a patient in need thereof, said treatment comprising: (a) obtaining megakaryocyte-derived extracellular vesicles according to claim 13 or claim 14; (b) incubating the megakaryocyte-derived extracellular vesicles with a therapeutic agent to allow the therapeutic agent to be imported into the lumen of the megakaryocyte-derived extracellular vesicles and / or associated with the surface of the megakaryocyte-derived extracellular vesicles, thereby producing a deliverable therapeutic agent; and (c) using the deliverable therapeutic agent to treat a patient, or contacting the deliverable therapeutic agent with biological cells in vitro and using the contacted biological cells to treat a patient; A therapeutic agent comprising:

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