NTRK2 signaling-mediated alveolar capillary injury and repair

Abstract

Disclosed herein are methods and systems of studying and modeling Neurotrophic Receptor Tyrosine Kinase 2 (NTRK2) signaling-mediated alveolar capillary injury and repair. Embodiments of the disclosure include generating a NTRK2 vessel organoid having hyperoxic injury (NTRK2 HIVO) comprising increased NTRK2-truncated isoform 1 (NTRK2-T1) expression compared to a control NTRK2 vessel organoid (NTRK2 VO) cultured in a normoxic culture condition. The disclosure also includes methods of treating alveolar capillary injury by transfected NTRK2 full length (FL) isoforms, for example, via lipid nanoparticle delivery.

Description

NTRK2 SIGNALING-MEDIATED ALVEOLAR CAPILLARY INJURY ANDREPAIRCROSS REFERENCE

[0001] This application claims priority to U.S. Provisional Patent Application No. 63 / 586,557, filed September 29, 2023, entitled “NTRK2 SIGNALING MEDIATED ALVEOLAR CAPILLARY INJURY AND REPAIR,” incorporated herein by reference in its entirety.BACKGROUND

[0002] The Neurotrophic Receptor Tyrosine Kinase 2 (NTRK2), a receptor for Brain-Derived Neurotrophic Factor (BDNF), plays a crucial role not only in neuronal function but also in the proper development of the lung. Mice lacking Ntrk2 in PDGFRa fibroblasts failed to recover from the acid-induced acute lung injury. Recently, decreased Ntrk2 was observed in alveolar capillary endothelial cells (ECs) from a hyperoxia-induced lung injury murine model resembling the alveolar simplification seen in human Bronchopulmonary Dysplasia (BPD), a common respiratory morbidity in preterm infants affecting -10,000 newborns per year in the US with no cure.SUMMARY

[0003] Embodiments of the present disclosure include the following numbered embodiments.

[0004] Embodiment 1 : A method of producing a Neurotrophic Receptor Tyrosine Kinase 2 (NTRK2)+vessel organoid comprising a hyperoxic injury (NTRK2+HIVO), the method comprising the steps of: a) activating a vascular endothelial growth factor (VEGF) signaling pathway, a fibroblast growth factor (FGF) signaling pathway, and a brain- derived neurotrophic factor (BDNF) signaling pathway of an angioblast spheroid in a hyperoxic culture condition with fetal bovine serum (FBS) for a first period of time sufficient to form a spheroid comprising acute hyperoxic injury (AHI); and b) activating a VEGF signaling pathway, an FGF signaling pathway, and a BDNF signaling pathway of the spheroid comprising AHI in a normoxic culture condition with FBS for a second period of timesufficient to form the NTRK2+HIVO, wherein the NTRK2+HIVO comprises increased NTRK2-truncated isoform 1 (NTRK2-T1) expression compared to a control NTRK2+vessel organoid (NTRK2+VO) cultured in a normoxic culture condition.

[0005] Embodiment 2: The method of embodiment 1, wherein the method further comprises producing the angioblast spheroid, by a method comprising: a) culturing induced pluripotent stem cells (iPSCs) to form an embryoid body; b) activating a WNT signaling pathway and a BMP signaling pathway of the embryoid body for a period of time sufficient to form a spheroid comprising mesoderm cells, optionally for about 2 days to about 4 days; and c) activating a VEGF signaling pathway of the spheroid comprising mesoderm cells with forskolin for a period of time sufficient to form the angioblast spheroid, optionally for about 1 day to about 3 days.

[0006] Embodiment 3 : The method of embodiment 1 or 2, wherein the angioblast spheroid comprises PECAM1, Ki67, and NTRK2-full length isoform (NTRK2-FL) expression.

[0007] Embodiment 4: The method of any one of embodiments 1-3, wherein the spheroid comprising AHI comprises reduced PECAM1 and Ki67 expression compared to a spheroid cultured in a normoxic culture condition.

[0008] Embodiment 5: The method of any one of embodiments 1-4, wherein the spheroid comprising AHI comprises decreased NTRK2-FL expression compared to a spheroid cultured in a normoxic culture condition.

[0009] Embodiment 6: The method of any one of embodiments 1-5, wherein the NTRK2+HIVO comprises alveolar capillary endothelial cells (ECs), pericytes, a basement membrane, and a lumen.

[0010] Embodiment 7: The method of embodiment 6, wherein the lumen is about 7-13 pm in diameter.

[0011] Embodiment 8: The method of any one of embodiments 1-7, wherein the NTRK2+HIVO comprises reduced PECAM1 and Ki67 expression compared to a control NTRK2+vessel organoid (NTRK2+VO) cultured in a normoxic culture condition.

[0012] Embodiment 9: The method of any one of embodiments 1-8, wherein the NTRK2+HIVO comprises reduced tube formation and cell proliferation compared to a control NTRK2+vessel organoid (NTRK2+VO) cultured in a normoxic culture condition.

[0013] Embodiment 10: The method of embodiment 1 or embodiment 2, further comprising the step of overexpressing one or more proteins selected from RBFOX2, H0XA5, and MEIS1 in the angioblast spheroid.

[0014] Embodiment 11 : The method of embodiment 1 or embodiment 2, further comprising the step of knocking down RBFOX2 expression in the angioblast spheroid.

[0015] Embodiment 12: The method of embodiment 1 or embodiment 2, wherein the iPSCs comprise a deletion of one or more genes selected from H0XA5 and MEIS1.

[0016] Embodiment 13: The method of any one of embodiments 1-12, wherein the iPSCs are derived from a patient with a lung disorder, optionally bronchopulmonary dysplasia (BPD).

[0017] Embodiment 14: The method of any one of embodiments 1-13, wherein the iPSCs are human iPSCs.

[0018] Embodiment 15: The method of any one of embodiments 1-14, whereinBDNF is provided at a concentration of about 10 to 40 pg / mL.

[0019] Embodiment 16: The method of any one of embodiments 1-15, wherein the hyperoxic culture condition comprises about 35% O2 to about 80% O2.

[0020] Embodiment 17: The method of any one of embodiments 1-16, wherein the hyperoxic culture condition comprises about 50% O2 to about 80% O2, optionally 75% O2

[0021] Embodiment 18: The method of any one of embodiments 1-17, wherein the normoxic culture condition comprises about 18% O2 to about 24% O2

[0022] Embodiment 19: The method of any one of embodiments 1-18, wherein the normoxic culture condition comprises about 20% O2 to about 23% O2, optionally 21% O2.

[0023] Embodiment 20: The method of any one of embodiments 1-19, wherein the first period of time is about 4 days to about 6 days.

[0024] Embodiment 21: The method of any one of embodiments 1-20, wherein the second period of time is about 4 days to about 6 days.

[0025] Embodiment 22: The method of any of the preceding embodiments, further comprising: modeling a treatment for hyperoxic injury using the NTRK2+ HIVO as a model.

[0026] Embodiment 23 : A NTRK2+HIVO produced by the method of any one of embodiments 1-21.

[0027] Embodiment 24: A Neurotrophic Receptor Tyrosine Kinase 2 (NTRK2)+vessel organoid comprising a hyperoxic injury (NTRK2+HIVO) comprising increased NTRK2 -truncated isoform 1 (NTRK2-T1) expression compared to a control NTRK2+ vessel organoid (NTRK2+ VO) cultured in a normoxic culture condition.

[0028] Embodiment 25: The NTRK2+HIVO of embodiment 23 or 24, further comprising alveolar capillary endothelial cells (ECs), pericytes, a basement membrane, and a lumen.

[0029] Embodiment 26: The NTRK2+HIVO of embodiment 25, wherein the lumen is about 7 to 13 pm in diameter.

[0030] Embodiment 27: The NTRK2+HIVO of any one of embodiments 23-26, wherein the NTRK2+HIVO comprises reduced PEC AMI and Ki67 expression compared to a control NTRK2+VO cultured in a normoxic culture condition.

[0031] Embodiment 28: The NTRK2+HIVO of any one of embodiments 23-27, wherein the NTRK21HIVO comprises reduced tube formation and cell proliferation compared to a control NTRK2+VO cultured in a normoxic culture condition.

[0032] Embodiment 29: The NTRK2+HIVO of any one of embodiments 23-28, wherein the NTRK2+HIVO comprises reduced NTRK2-FL expression compared to a control NTRK2+VO cultured in a normoxic culture condition.

[0033] Embodiment 30: The NTRK2+HIVO of any one of embodiments 23-29, wherein the NTRK2+HIVO comprises increased expression of RBFOX2.

[0034] Embodiment 31 : The NTRK2+HIVO of any one of embodiments 23-30, wherein the NTRK2+HIVO comprises reduced expression of HOXA5 and MEIS1.

[0035] Embodiment 32: The NTRK2+HIVO of any one of embodiments 23-31, wherein the NTRK2+HIVO comprises increased expression of HOXA5 and MEIS1.

[0036] Embodiment 33: The NTRK2+HIVO of any one of embodiments 23-32, wherein the NTRK2+HIVO is configured to model a treatment for hyperoxic injury.

[0037] Embodiment 34: The NTRK2+HIVO of any one of embodiments 23-33, wherein the NTRK2+HIVO is derived from human induced pluripotent stem cells (iPSCs).

[0038] Embodiment 35: The NTRK2+HIVO of any one of embodiments 23-34, wherein the NTRK2+HIVO is derived from a patient with a lung disorder, optionally bronchopulmonary dysplasia (BPD).

[0039] Embodiment 36: A method of screening a compound for improving alveolar regeneration comprising contacting the NTRK2+HIVO of any one of embodiments 23-35 with the compound and measuring a response to the compound.

[0040] Embodiment 37: A method of producing a lung organoid comprising lungspecific endothelial cells, the method comprising adding lung-specific endothelial cells derived from induced pluripotent stem cells (iPSCs) to a lung epithelial cell organoid derived from iPSCs for about 6 days to about 8 days.

[0041] Embodiment 38: The method of embodiment 37, wherein the lung organoid comprising lung-specific endothelial cells comprises AGER+ATI cells, HTII-280+AT2 cells, and CD31+lung-specific endothelial cells.

[0042] Embodiment 39: A lung organoid comprising lung-specific endothelial cells produced by the method of embodiment 37 or embodiment 38.

[0043] Embodiment 40: A lung organoid comprising lung-specific endothelial cells, further comprising AGER1ATI cells, HTII-2801AT2 cells, and CD311lung-specific endothelial cells.

[0044] Embodiment 41: A method to produce a lung epithelial -vessel assembloid, the method comprising contacting a vessel organoid (VO) with a lung epithelial organoid.

[0045] Embodiment 42: The method of embodiment 41, wherein the lung epitheli al -vessel assembloid comprises CD31+lung-specific endothelial cells and NKX2.1+lung epithelial progenitor cells.

[0046] Embodiment 43: A lung epithelial -vessel assembloid produced by the method of embodiment 41 or embodiment 42.

[0047] Embodiment 44: A lung epithelial-vessel assembloid comprising CD31+endothelial cells and NKX2.1+lung epithelial progenitor cells.

[0048] Embodiment 45: A vascularized alveolar organoid comprising: lung epithelial progenitor cells; lung-specific endothelial cells; and lung-specific mesenchymal cells.

[0049] Embodiment 46: The vascularized alveolar organoid of embodiment 45, wherein the lung epithelial progenitor cells comprises NKX2.1+.

[0050] Embodiment 47: The vascularized alveolar organoid of embodiment 45 or 46, wherein the endothelial cells express CD31.

[0051] Embodiment 48: The vascularized alveolar organoid of any one of embodiments 45-47, wherein the endothelial cells express FOXF1.

[0052] Embodiment 49: The vascularized alveolar organoid of any one of embodiments 45-48, wherein the endothelial cells express FENDRR.

[0053] Embodiment 50: The vascularized alveolar organoid of any one of embodiments 45-49, wherein the endothelial cells express CDH5.

[0054] Embodiment 51 : The vascularized alveolar organoid of any one of embodiments 45-50, wherein the mesenchymal cells express FOXFE

[0055] Embodiment 52: The vascularized alveolar organoid of any one of embodiments 45-51, wherein the mesenchymal cells express TBX4.

[0056] Embodiment 53 : A method of increasing endothelial cell proliferation and angiogenesis in pulmonary microvascular endothelial cells (PMVECs), the method comprising: a) transfecting PMVECs with a nucleic acid encoding NTRK2-FL; and b) activating a NTRK2 signaling pathway with BDNF in PMVECs following a post-transfection period.

[0057] Embodiment 54: The method of embodiment 53, wherein transfecting PMVECs with a nucleic acid encoding NTRK2-FL comprises transfecting PMVECs with a lipid nanoparticle comprising NTRK2-FL mRNA.

[0058] Embodiment 55: The method of embodiment 53 or 54, wherein activating NTRK2 signaling pathway of the PMVECs comprising contacting the PMVECs with BDNF for a period of time sufficient to activate the NTRK2 signaling pathway.

[0059] Embodiment 56: The method of embodiment 55, wherein the period of time sufficient to activate NTRK2 signaling following the post-transfection period is about 20- 40 minutes.

[0060] Embodiment 57: The method of embodiment 55 or 56, wherein the period of time sufficient to activate NTRK2 signaling following the post-transfection period is about 25-35 minutes.

[0061] Embodiment 58: The method of any one of embodiments 53-57, wherein the post-transfection period is about 35-60 hours.

[0062] Embodiment 59: The method of any one of embodiments 53-58, wherein the post-transfection period is about 40-55 hours.

[0063] Embodiment 60: The method of any one of embodiments 53-59, wherein the nucleic acid encoding NTRK2-FL or the lipid nanoparticle comprising NTRK2-FL mRNA is at a concentration of about 0.5 - 2 pg / ml.

[0064] Embodiment 61: The method of any one of embodiments 53-60, wherein activating NTRK2 signaling pathway of the PMVECs comprising contacting the PMVECs with BDNF for a period of time sufficient to activate the NTRK2 signaling pathway, wherein the BDNF is at a concentration of about 15-35 ng / ml.

[0065] Embodiment 62: The method of embodiment 61, wherein the BDNF is at a concentration of about 20-30 ng / ml.

[0066] Embodiment 63: A method of reducing endothelial cell proliferation and impairing angiogenesis in pulmonary microvascular endothelial cells (PMVECs), the method comprising: a) transfecting PMVECs with a nucleic acid encoding NTRK2-T1; and b) activating a NTRK2 signaling pathway of the PMVECs following a post-transfection period.

[0067] Embodiment 64: The method of embodiment 63, wherein transfecting PMVECs with a nucleic acid encoding NTRK2-T1 comprises transfecting PMVECs with a lipid nanoparticle comprising NTRK2-T1 mRNA.

[0068] Embodiment 65: The method of embodiment 63 or 64, wherein activating NTRK2 signaling pathway of the PMVECs comprising contacting the PMVECs with BDNF for a period of time sufficient to activate the NTRK2 signaling pathway.

[0069] Embodiment 66: The method of embodiment 65, wherein the period of time sufficient to activate NTRK2 signaling following the post-transfection period is about 20- 40 minutes.

[0070] Embodiment 67: The method of embodiment 65 or 66, wherein the period of time sufficient to activate NTRK2 signaling following the post-transfection period is about 25-35 minutes.

[0071] Embodiment 68: The method of any one of embodiments 63-67, wherein the post-transfection period is about 35-60 hours.

[0072] Embodiment 69: The method of any one of embodiments 63-68, wherein the post-transfection period is about 40-55 hours.

[0073] Embodiment 70: The method of any one of embodiments 63-69, wherein the nucleic acid encoding NTRK2-T1 or the NTRK2-T1 mRNA is at a concentration of about 0.5-2 pg / ml.

[0074] Embodiment 71 : The method of any one of embodiments 63-70, wherein activating the NTRK2 signaling pathway of the PMVECs comprising contacting the PMVECs with BDNF for a period of time sufficient to activate the NTRK2 signaling pathway, wherein the BDNF is at a concentration of about 15-35 ng / ml.

[0075] Embodiment 72: The method of embodiment 71, wherein the BDNF is at a concentration of about 20-30 ng / ml.

[0076] Embodiment 73: A method of treating an in vitro hyperoxic injury, the method comprising the steps of a) transfecting a spheroid comprising acute hyperoxic injury (AHI) with a nucleic acid encoding NTRK2-FL; b) activating a VEGF signaling pathway, an FGF signaling pathway, and a BDNF signaling pathway of the transfected spheroid comprising AHI in a normoxic culture condition with FBS for a second period of time sufficient to form a treated NTRK2+HIVO.

[0077] Embodiment 74: The method of embodiment 73, wherein transfecting the spheroid comprising AHI with a nucleic acid encoding NTRK2-FL comprises transfecting the spheroid comprising AHI with a lipid nanoparticle comprising NTRK2-FL mRNA.

[0078] Embodiment 75: The method of embodiment 73 or 74, wherein the second period of time sufficient to form the treated NTRK2+HIVO is about 4 days to about 6 days.

[0079] Embodiment 76: The method of any one of embodiments 73-75, wherein the nucleic acid encoding NTRK2-FL or the NTRK2-FL mRNA is at a concentration of about 0.5 - 2 pg / ml.

[0080] Embodiment 77: The method of any one of embodiments 73-76, wherein the BDNF signaling pathway is activated by culturing the transfected spheroid comprising AHI with BDNF, optionally wherein the BDNF is at a concentration of about 15 - 35 ng / ml.

[0081] Embodiment 78: The method of embodiment 77, wherein the BDNF is at a concentration of about 20-30 ng / ml.

[0082] Embodiment 79: The method of any one of embodiments 73-78, wherein the spheroid comprising AHI is derived from a method comprising: activating a VEGF signaling pathway, an FGF signaling pathway, and a BDNF signaling pathway in an angioblastspheroid, optionally derived from induced pluripotent stem cells (iPSCs), in a hyperoxic culture condition with FBS for a period of time sufficient to form a spheroid comprising acute hyperoxic injury (AHI).

[0083] Embodiment 80: The method of embodiment 79, wherein the period of time sufficient to form the spheroid comprising AHI is about 4 days to about 6 days.

[0084] Embodiment 81 : The method of embodiment 79 or 80, wherein the hyperoxic culture condition comprises about 35% to about 80% O2, optionally about 75% O2.

[0085] Embodiment 82: The method of any one of embodiments 73-81, wherein the treated NTRK2+HIVO comprises increased PEC AMI and Ki67 expression compared to a control NTRK2+HIVO.

[0086] Embodiment 83: The method of any one of embodiments 73-82, wherein the treated NTRK2+HIVO comprises increased tube formation and cell proliferation compared to a control NTRK2+HIVO.

[0087] Embodiment 84: A method of treating an in vivo hyperoxic injury, comprising increasing expression of NTRK2-FL in cells of the subject.

[0088] Embodiment 85: The method of embodiment 84, wherein increasing expression of NTRK2-FL in cells of the subject comprises transfecting the cells with a nucleic acid encoding NTRK2-FL.

[0089] Embodiment 86: The method of embodiment 85, wherein transfecting the cells with the nucleic acid encoding NTRK2-FL comprises injecting lipid nanoparticles comprising NTRK2-FL mRNA intravenously into the subject.

[0090] Embodiment 87: The method of any one of embodiments 84-86, wherein the subject is a mouse.

[0091] Embodiment 88: The method of any one of embodiments 84-87, wherein the in vivo hyperoxic injury is induced by a method comprising the steps: a) exposing the subject to a hyperoxic condition for a first period of time, optionally about 6 days to about 14 days; and b) returning the subject to a normoxic condition for a second period of time, optionally about 20 days to about 22 days.

[0092] Embodiment 89: The method of embodiment 88, wherein the hyperoxic culture condition comprises about 35% O2 to about 80% O2, optionally 75% O2 and the normoxic culture condition comprises about 18% O2 to about 24% O2, optionally 21% O2.

[0093] Embodiment 90: The method of embodiment 88 or embodiment 89, wherein the increasing expression of NTRK2-FL in cells of the subject occurs prior to, at the start of, during or after the first period of time.

[0094] Embodiment 91 : The method of any one of embodiments 84-90, further comprising: promoting vascular regeneration and / or alveologenesis.

[0095] Embodiment 92: The method of any one of the preceding embodiments, wherein the lipid nanoparticle targets the pulmonary microvascular ECs, optionally wherein the lipid nanoparticle is a polyethyleneimine (PEI)-based nanoparticle.

[0096] Embodiment 93: The method of any one of the preceding embodiments, wherein activating the VEGF signaling pathway of the angioblast spheroid, of the spheroid comprising AHI, of the spheroid comprising mesoderm cells, or of the transfected spheroid comprising AHI comprises contacting the angioblast spheroid, the spheroid comprising AHI, the spheroid comprising mesoderm cells, or the transfected spheroid comprising AHI, respectively, with VEGF.

[0097] Embodiment 94: The method of any one of the preceding embodiments, wherein activating the FGF signaling pathway of the angioblast spheroid, of the spheroid comprising AHI, or of the transfected spheroid comprising AHI comprises contacting the angioblast spheroid, the spheroid comprising AHI, or the transfected spheroid comprising AHI, respectively, with FGF2.

[0098] Embodiment 95: The method of any one of the preceding embodiments, wherein activating the BDNF signaling pathway of the angioblast spheroid, of the spheroid comprising AHI, or of the transfected spheroid comprising AHI comprises contacting the angioblast spheroid, the spheroid comprising AHI, or the transfected spheroid comprising AHI, respectively, with BDNF.

[0099] Embodiment 96: The method of any one of the preceding embodiments, wherein activating the WNT signaling pathway of the embryoid body comprises contacting the embryoid body with CHIR99021.[000100] Embodiment 97: The method of any one of the preceding embodiments, wherein activating the BMP signaling pathway of the embryoid body comprises contacting the embryoid body with BMP4.BRIEF DESCRIPTION OF THE DRAWINGS[000101] In addition to the features described herein, additional features and variations will be readily apparent from the following descriptions of the drawings and exemplary embodiments. It is to be understood that these drawings depict embodiments and are not intended to be limiting in scope.[000102] Figs. 1A-1B are schematic representations of NTRK2 isoforms and downstream signaling, according to example embodiments of the present disclosure. For example, Fig. 1A is a schematic representation of the genomic structure and protein domains of the NTRK2 splice variants. Exons are shown as boxes and introns are shown as lines. Color of exons indicate the domains of the NTRK2 protein shown in upper panel. Fig. IB shows intracellular signaling pathways downstream of NTRK2-Full length (NTRK2-FL) and NTRK2-Truncated isoform (NTRK2-T1). These pathways were identified in neurons and astrocytes. However, the NTRK2-T1 regulatory network in alveolar capillary ECs remains unknown. BDNF: brain-derived neurotrophic factor.[000103] Figs. 2A-2C are schematic representations of a dynamic expression of NTRK2 in general capillary endothelial (gCap) cells and lung-specific mesenchymal alveolar niche cells (MANC) during lung development in human and mouse, according to example embodiments of the present disclosure. For example, Fig. 2A is a set of graphs showing the expression of Ntrk2 in gCap and MANC cells in Drop-seq of mouse lung development. The graph on the left is a Dotplot of Ntrk2 average expression level (dot color) and expression percentage (dot size) in each cell type in each Dropseq sample. The graph on the right is a boxplot of average expression (log transformed; top panels) and expression percentage (bottom panels) of Ntrk2 in gCap or MANC cells in Drop-seq samples from each time point. Drop-seq data were from LungMAP mouse lung CellRef (Guo et al., BioRxiv 2022); PND: postnatal day. Fig. 2B is a set of graphs showing the expression of NTRK2 in gCap and MANC cells in single nucleus RNA-seq (snRNA-seq) of human lung at saccular (n=3;PNDl, 29-31 weeks of gestational age [wks GA]), early alveolar (n=2; PND1, 38-39 wks GA), and late alveolar (n=3; 3 years old) phases. snRNA-seq of human lung were from our new sequencing.[000104] Figs. 3A-3B are a set of graphs and images showing the dynamic changes of NTRK2 splice variants in human and mouse lungs exposed to hyperoxic injury, according to example embodiments of the present disclosure. For example, in Fig. 3A, the upper graphshows the results of RNAScope of NTRK2-Shared (NTRK2-Sh, green dots) and NTRK2-TK (red dots) in human peripheral lung samples from control and BPD patients collected at different stages. The lower graph of Fig. 3A shows the quantification of NTRK2 isoforms. NTRK-T1+ cells (green arrow) were defined as cells positive with NTRK2-Sh (green dots) but not NTRK2-TK (red dots); NTRK2-FL+ cells (red arrow) were defined as cells positive with both NTRK2-Sh and NTRK2-TK. n=3 or 4 individuals / group. Fig. 3B is a set if graphs and images showing the results when newborn mice were exposed to 90% hyperoxia (HO) for 7 days followed by 21 days of room air (RA). The upper graph of Fig. 3B is a representative H&E staining and RNAScope of Ntrk2-Shared (Ntrk2-Sh, red dots) and Ntrk2-TK (green dots) in mouse peripheral lung tissues from postnatal day 4 (P4) to P28 with either recovered alveoli structure, or impaired recovery. The lower graph of Fig. 3B shows a quantification of these results - the red arrows indicate Ntrk2-Tl+ cells, the green arrows indicate Ntrk2-FL+ cells (the colors were opposite in human and mouse). n=3 or 4 mice / group. Data shown as the mean ± SEM. *p<0.05, **p<0.01, ***p<0.001, two-way ANOVA followed by Bonferroni’s test.[000105] Figs. 4A-4F are a set of graphs showing the multi omic analysis of enriched ECs from control and BPD patient lung samples, according to example embodiments of the present disclosure. For example, Fig. 4A is a schematic workflow for Multiome of human distal lungs from three BPD cases and two age-matched controls. Endothelial population was enriched with a CD31 antibody by magnetic-activated cell sorting. Fig. 4B is a UMAP visualization of the identified cell types. gCap: general Capillary; gCap C1 / C2: gCap subcluster 1 / 2. aCap: aerocyte; AF: Alveolar Fibroblasts; ASMC: Airway Smooth Muscle Cells; VSMC: Vascular Smooth Muscle Cells. Fig. 4C is a UMAP visualization of highlighted gCap-C2 (green) cells in two controls and three BPD cases. Fig. 4D is a feature plot of the MKI67 expression in human lung Multiome. Fig. 4E is a feature plot of the NTRK2 expression in Multiome profiling of three BPD and two control distal lung samples. In Fig. 4F, the left graph is a USCS browser track showing NTRK2 sequencing reads distribution based on the Multiome profiling BPD and Control lung ECs; and the right graph shows quantifications of mapping reads for each domain. Two-way ANOVA with Bonferroni’s post-test was performed. *p<0.05 BPD vs. Control.[000106] Figs. 5A-5C are a set of graphs and images showing the signaling and functions of NTRK2-FL and NTRK2-T1 in hPMVECs, according to example embodiments ofthe present disclosure. hPMVECs were transfected with GFP, NTRK2-FL, or NTRK2-T1 mRNA (lug / ml) respectively using the jetMESSENGER nanoparticle delivery system 24h after seeding, according to example embodiments of the present disclosure. 48h after transfection, BDNF (25ng / ml) was added to activate NTRK2 signaling. After 30 mins of treatment, cells were collected for western blot (n=3) (e.g., as shown in Fig. 5A), RT-qPCR (n=3) (e.g., as shown in Fig. 5B, left graph), or seeded in 96-well plate for BrdU assay (n=5) (e.g., as shown in Fig. 5B, right graph). Fig. 5C is a set of graphs and images showing the results when hPMVECs were treated as described above, and seeded on growth factor-reduced Matrigel with or without BDNF (25ng / ml). 12h after incubation, the cells were stained with Calcein (2uM). The images were quantified using FastTrack Al tube formation analysis pack (n=5). Data shown as mean±S.D., *p<0.05, **p<0.01, ***p<0.001, unpaired t-test for Fig. 5A and Fig. 5B; two-way ANOVA followed by Bonferroni’s test for Fig. 5C.[000107] Figs. 6A-6C are a set of graphs and images showing the changes in EC function and NTRK2 expression during capillary formation under hyperoxia in iPSC-VOs, according to example embodiments of the present disclosure. For example, Fig. 6A is a schematic showing how hiPSCs are differentiated to angioblasts from Day 0 to Day 5 under room air (RA). On day 5, angioblasts were divided into two groups: one was exposed to hyperoxia (HO, 75% oxygen) for five days followed by 5 days of RA, and in the other group, angioblasts were maintained in RA for 10 days. BDNF (25ng / ml) was added from Day 5 to Day 15 to mimic the alveolar microenvironment. Fig. 6B is a set of images showing representative immunostaining of endothelial marker PECAM1, proliferation marker MKI67, and RNAScope staining of NTRK2- Shared (NTRK2-Sh, green) and NTRK2-TK (red) of VOs with or without hyperoxia injury. Fig. 6C is a set of graphs showing the quantification of the aforementioned images shown in Fig. 6B. Tube length was quantified as described in Fig. 5. NTRK2-FL+ cells (red arrows) were defined as cells positive for both NTRK2-Sh and NTRK2-TK, and NTRK2-T1+ cells (green arrows) represented cells positive for NTRK2-Sh but not NTRK2-TK. n=3 biological repeats. Data shown as mean ± SEM. *p<0.05, **p<0.01, ***p<0.001. Unpaired t-test in Fig. 6A, one-way ANOVA followed by Tukey’s test in Fig. 6B, and two-way ANOVA followed by Bonferroni’s test in Fig. 6C. HO: Hyperoxia, RA: Room Air.[000108] Figs. 7A-7C are a set of graphs and images showing how the overexpression of NTRK2-FL in VOs resulted in improved tube formation and cell proliferation after hyperoxia exposure, according to example embodiments of the present disclosure. For example, Fig. 7A is a schematic showing how iPSC-VOs were transfected with GFP, NTRK2- FL and NTRK2-T1 mRNAs (lug / ml) using the jetMESSENGER nanoparticle delivery system on Day 10 after five days of hyperoxia injury (75% oxygen). BDNF (25ng / ml) was added from Day 5 to Day 15. Fig. 7B is a set of images showing representative whole-mount staining of PEC AMI (white) and immunostaining of MKI67 (red) in D15 VOs. Fig. 7C is a set of graphs showing quantifications of total tube length (Left graph) and MKI67 percentage (Right graph). n=3-5 VOs / group. Data shown as the mean ± SEM. *p<0.05, **p<0.01, ***p<0.001, one-way ANOVA followed by Tukey’s test. HO: Hyperoxia, RA: Room Air.[000109] Figs. 8A-8C are a set of graphs and images showing the dynamic changes of Ntrk2 isoforms in mouse lungs exposed to mild versus severe hyperoxic injuries, according to example embodiments of the present disclosure. For example, Fig. 8A is a set of representative H&E images and RNAScope of Ntrk2-shared region (Ntrk2-Sh, red) and Ntrk2- TK region (Ntrk2-TK, green) on P28 mouse lungs after mild (75% 02 from P0-P7, room air P7-28) and severe (90% 02 from P0-P7, room air P7-28) hyperoxic injuries. Ntrk2-FL+ cells (pink dashed-circle): Ntrk2-Sh+ and Ntrk2-TK+; Ntrk2-Tl+ cells (yellow dashed-circle): Ntrk2-Sh+ only. Fig. 8B is a set of graphs showing the quantification of Mean Linear Intercept (MLI) and Ntrk2+ cells from Fig. 8A. n=3-4 for MLI, n=6 for RNAScope. Data shown as the mean SEM. **p<0.01, ***p<0.001. One-way ANOVA followed by Tukey’s test for MLI analysis, Two-way ANOVA followed ± by Tukey’s test for Ntrk2 isoform analysis. Fig. 8C shows the results when newborn mice were exposed to 75% or 90% 02 for 7 days. gCap cells (CD31+, KIT+) were sorted on day 10 by FACS for western blot analysis. Protein samples were obtained from six mice per condition.[000110] Figs. 9A-9C are a set of graphs and images showing the changes in EC function and NTRK2 isoforms under mild and severe hyperoxic injuries in iPSC-VOs, according to example embodiments of the present disclosure. For example, Fig. 9A is a graph showing how hiPSCs were differentiated to angioblasts from Day 0 to Day 5 under room air (RA). On day 5, angioblasts were divided into three groups: 1) maintained under RA for 10 days; 2) subjected to mild hyperoxia (HO, 60% 02) for 3 days followed by 7 days of RA; 3)subjected to severe hyperoxia (HO, 75% 02) for 5 days followed by 5 days of RA. BDNF (25ng / ml) was added from Day 5 to Day 15 to mimic the alveolar microenvironment. Fig. 9B is a set of representative immunostaining images and quantification of endothelial marker PECAM1 and proliferation marker MKI67. Fig. 9C is a set of representative immunostaining images and quantification of NTRK2-FL (NTRK2-Total+ and NTRK2-T1-, red circles), and NTRK2-T1 (NTRK2-Total+ and NTRK2-T1-, yellow circles) under the conditions decribed in (A). n=3 biological repeats. Data shown as mean ± SEM. *p<0.05, **p<0.01, ***p<0.001. One-way ANOVA followed by Tukey’s test in Fig. 9B, and two-way ANOVA followed by Bonferroni’s test in Fig. 9C.[000111] Fig. 10 is a graph showing the dynamic changes of NTRK2 isoforms in response to hyperoxia-induced lung injury, according to example embodiments of the present disclosure. The human results were summarized based on iPSC-VO studies and multiomic analysis of control and BPD lung samples. Mouse results were summarized based on hyperoxic mouse model with variable degree of recovery.[000112] Figs. 11A-11B are a set of graphs and images showing how the spatial transcriptomic (ST) revealed localization of different cell types in human distal lung, according to example embodiments of the present disclosure. For example, Fig. 11A is a schematic workflow of lOx Visium ST analysis of human BPD distal lung samples and age-matched controls. Cell21ocation was used for the cell type deconvolution analysis using two single cell references: the Multiome profiling (see Figure 2, 15 cell types) and the LungMAP human lung CellRef (48 cell types). In Fig. 11B, the left panel shows H&E staining with histological annotation (top) and illustration (bottom) of a healthy human distal lung tissue; and the right panel shows the visualization of Cell21ocation estimation of cell type abundance in each spatial spot. Cell identities were learned using the scRNA-seq data of the LungMAP CellRef. ATI : Alveolar Type 1 cells; AT2: Alveolar Type 2 cells; EC: Endothelial Cells; gCap: general Capillary; aCap: aerocyte; AF1 : Alveolar fibroblast type 1, AF2: Alveolar fibroblast type 2.[000113] Figs. 12A-12B are a set of graphs and images showing the expression of BDNF and NTRK2 in the 10X Visium ST of BPD vs. Control lungs, according to example embodiments of the present disclosure. For example, Fig. 12A is a spatial visualization of BDNF (top) and NTRK2 (bottom) expression in each 10X Visium sample. Data normalizationwas performed using SCTransformation in Seurat 4. Fig. 12B is a dotplot visualization of average expression and frequencies (Percent) of BDNF and NTRK2 in each sample.[000114] Figs. 13A-13C are a set of graphs showing the analyses of cell-cell interactions based on ST, according to example embodiments of the present disclosure. For example, Figs. 13 A and 13B show the spatial visualization of deconvoluted cell types in BPD lung sample. Fig. 13C is a graph showing cell-cell communication pairs determined in either BPD gCap-C2 spots (n=86), or in Control gCap-Cl spots (n=243). stLearn was used with Cell21ocation to infer ligand-receptor pairs significant for a specific cell-cell pair co-occurrent within the same spot or in immediate neighboring spots. Node color represents the number of ST spots in which a L-R pair was significant (p<0.05) for a source-target cell pair inferred by stLearn. AF1: Alveolar Fibroblasts 1.[000115] Figs. 14A-14C are a set of images showing how 3D cellular models can be used to study cell-cell interaction, according to example embodiments of the present disclosure. For example, Fig. 14A is an image showing the immunostaining of ATI (AGER), AT2 (HTIL280), and AT1 / AT2 (CLDN4) in an iPSC -derived epithelial organoid. ATI was induced by 10% of human serum. Fig. 14B is an immunostaining image showing the results when iPSC-ECs were added to the epithelial organoids for 7 days. The dashed line indicates the area with both ATI and ECs (CD31+). Fig. 14C is an image showing the whole-mount staining of the VO-epithelial assembloids. NKX2.1: lung epithelial progenitors. Fig. 14D is a set of images showing the immunostaining of lung ECs (e.g., based on markers CD31, FOXF1, FENDRR, CDH5) and lung-specific mesenchymal cells (e.g., based on markers FOXF1, TBX4) in human iPSC-derived vascularized alveolar organoid (40 days of differentiation).[000116] Figs. 15A-15B are a set of graphs showing the transcriptional regulation of NTRK2 in control vs. BPD gCap cells, according to example embodiments of the present disclosure. For example, in Fig. 15A, the top graph shows the multiome AT AC peaks in the promoter region of the NTRK2 genome from gCap-Cl and gCap-C2. The NTRK2 gene expression (violin plot) was altered corresponding to the ATAC-peak magnitude. The bottom graph of Fig. 15A shows the enriched transcription factor binding motifs in the NTRK2 promoter region. Fig. 15B shows violin plots showing H0XA5 and MEIS1 expressions in gCap-Cl and gCap-C2 from BPD and control lung tissues. Wilcoxon rank sum test was performed.[000117] Figs. 16A-16C are a set of graphs and images showing the increase of RBFOX2 in BPD lungs, according to example embodiments of the present disclosure. For example, Fig. 16A is a feature plot of RBFOX2 expression based on the multiomic data presented in Fig. 4A. Fig. 16B is a violin plot of RBFOX2 expression within gCap-C2 defined in Fig. 4A. Fig. 16C is a set of representative immunostaining images of RBFOX2 and PECAM1 (Upper panel), and graphs showing the percentage of RBFOX2+ECs in total ECs (Lower) in control and BPD lung samples (active evolving and established stages). n=3 / group. Data shown as the mean ± SEM. *p<0.05, **p<0.01, ***p<0.001, unpaired 2-tailed t-test. AF: alveolar fibroblasts.[000118] Fig. 17 is a set of graphs showing the optimal transfection efficiency of GFP in ECs, according to example embodiments of the present disclosure. GFP positive cell percentage (bar) and GFP median fluorescence intensity (dot-line) for size-controlled jetMESSENGER or PEIpro particles with different dosage of mRNAs. n=3.[000119] Figs. 18A-18E are a set of graphs and images showing how lipid nanoparticle delivery of Ntrk2-FL mRNA to lung ECs in the mouse rescued the alveoli simplification following hyperoxic injury, according to example embodiments of the present disclosure. For example, Fig. 18A is a pictorial demonstration of the lipid nanoparticles (LNPs) with lung EC specificity. Fig. 18B is a set of images showing the results after neonatal mice were administered LNPs containing luciferase mRNAs via retro-orbital injection, and tissue samples were collected 24 hours post-injection. The empty LNPs (first row) were used as a negative control. Fig. 18C is a set of graphs showing the results after neonatal Ail4 mice received transorbital injections of Cre-mRNA LNPs (0.75 mg / kg) at P4 and P6. Lungs were collected at P8 and dissociated into a single-cell suspension for subsequent flow cytometry analysis. Fig. 18D is a set of schematics and images showing how neonatal mice were exposed to room air or hyperoxia (90% 02) for 7 days. Within the hyperoxia group, mice received LNPs containing GFP mRNA or Ntrk2-FL mRNA (0.75 pg / g) every 48 hours from day 2 to day 10. H&E staining was performed on P28 to assess lung histology. Fig. 18E is a set of images showing the immunostaining of EC markers in distal lungs from P28 mice. n=5-6 mice / group. Fig. 18F is a graph showing how mice receiving LNPs containing the Ntrk2-FL mRNA exhibited significant improvement in alveologenesis and increase in alveolar ECscompared to contro. Data shown as mean ± SEM. ***p<0.001, one-way ANOVA followed by Tukey’s test. HO: Hyperoxia.DETAILED DESCRIPTION[000120] In the following detailed description, reference is made to the accompanying drawings, which form a part hereof. In the drawings, similar symbols typically identify similar components, unless context dictates otherwise. The illustrative embodiments described in the detailed description, drawings, and claims are not meant to be limiting. Other embodiments may be utilized, and other changes may be made, without departing from the spirit or scope of the subject matter presented herein. It will be readily understood that the aspects of the present disclosure, as generally described herein, and illustrated in the Figures, can be arranged, substituted, combined, separated, and designed in a wide variety of different configurations, all of which are explicitly contemplated herein.[000121] The following description of various embodiments is exemplary and explanatory only and is not to be construed as limiting or restrictive in any way. Other embodiments, features, objects, and advantages of the present teachings will be apparent from the description and accompanying drawings, and from the claims.[000122] The disclosure herein uses affirmative language to describe the numerous embodiments. The disclosure also includes embodiments in which subject matter is excluded, in full or in part, such as substances or materials, method steps and conditions, protocols, or procedures.[000123] It should be understood that any use of subheadings herein are for organizational purposes, and should not be read to limit the application of those subheaded features to the various embodiments herein. Each and every feature described herein is applicable and usable in all the various embodiments discussed herein and that all features described herein can be used in any contemplated combination, regardless of the specific example embodiments that are described herein. It should further be noted that exemplary description of specific features are used, largely for informational purposes, and not in any way to limit the design, subfeature, and functionality of the specifically described feature.Definitions of Terms[000124] Unless otherwise noted, terms are to be understood according to conventional usage by those of ordinary skill in the relevant art. For purposes of the present disclosure, the following terms are explained below.[000125] As used herein, “a” or “an” may mean one or more. As used herein in the claim(s), when used in conjunction with the word “comprising,” the words “a” or “an” may mean one or more than one. Some embodiments of the disclosure may consist of or consist essentially of one or more elements, method steps, and / or methods of the disclosure. It is contemplated that any method or composition described herein can be implemented with respect to any other method or composition described herein and that different embodiments may be combined.[000126] The use of the term “or” in the claims is used to mean “and / or” unless explicitly indicated to refer to alternatives only or the alternatives are mutually exclusive, although the disclosure supports a definition that refers to only alternatives and “and / or.” For example, “x, y, and / or z” can refer to “x” alone, “y” alone, “z” alone, “x, y, and z,” “(x and y) or z,” “x or (y and z),” or “x or y or z.” It is specifically contemplated that x, y, or z may be specifically excluded from an embodiment. As used herein “another” may mean at least a second or more.[000127] The term “ones” means more than one.[000128] As used herein, the term “plurality” may be 2, 3, 4, 5, 6, 7, 8, 9, 10, or more.[000129] As used herein, the term “set of’ means one or more. For example, a set of items includes one or more items.[000130] As used herein, the phrase “at least one of,” when used with a list of items, means different combinations of one or more of the listed items may be used and only one of the items in the list may be needed. The item may be a particular object, thing, step, operation, process, or category. In other words, “at least one of’ means any combination of items or number of items may be used from the list, but not all of the items in the list may be required. For example, without limitation, “at least one of item A, item B, or item C” means item A; item A and item B; item B; item A, item B, and item C; item B and item C; or item A and C. In some cases, “at least one of item A, item B, or item C” means, but is not limited to, two of item A, one of item B, and ten of item C; four of item B and seven of item C; or some other suitable combination.[000131] As used herein, “substantially” means sufficient to work for the intended purpose. The term “substantially” thus allows for minor, insignificant variations from an absolute or perfect state, dimension, measurement, result, or the like such as would be expected by a person of ordinary skill in the field but that do not appreciably affect overall performance. When used with respect to numerical values or parameters or characteristics that can be expressed as numerical values, “substantially” means within ten percent.[000132] Throughout this specification, unless the context requires otherwise, the words “comprise”, “comprises” and “comprising” will be understood to imply the inclusion of a stated step or element or group of steps or elements but not the exclusion of any other step or element or group of steps or elements. By “consisting of’ is meant including, and limited to, whatever follows the phrase “consisting of.” Thus, the phrase “consisting of’ indicates that the listed elements are required or mandatory, and that no other elements may be present. By “consisting essentially of’ is meant including any elements listed after the phrase, and limited to other elements that do not interfere with or contribute to the activity or action specified in the disclosure for the listed elements. Thus, the phrase “consisting essentially of’ indicates that the listed elements are required or mandatory, but that no other elements are optional and may or may not be present depending upon whether or not they affect the activity or action of the listed elements.[000133] Reference throughout this specification to “one embodiment,” “an embodiment,” “a particular embodiment,” “a related embodiment,” “a certain embodiment,” “an additional embodiment,” or “a further embodiment” or combinations thereof means that a particular feature, structure or characteristic described in connection with the embodiment is included in at least one embodiment of the present disclosure. Thus, the appearances of the foregoing phrases in various places throughout this specification are not necessarily all referring to the same embodiment. Furthermore, the particular features, structures, or characteristics may be combined in any suitable manner in various embodiments.[000134] As used herein, the terms “treatment,” “treating,” “treat,” and the like, with respect to a disease or condition, can refer to obtaining a desired pharmacologic and / or physiologic effect. The effect can be prophylactic in terms of completely or partially preventing a disease or symptom thereof and / or can be therapeutic in terms of a partial or complete cure for a disease and / or adverse effect attributable to the disease. For example, atreatment can include executing a protocol, which may include administering one or more drugs to a patient, in an effort to alleviate signs or symptoms of the disease. Desirable effects of treatment include decreasing the rate of disease progression, ameliorating or palliating the disease state, and remission or improved prognosis. Alleviation can occur prior to signs or symptoms of the disease or condition appearing, as well as after their appearance. Thus, “treating” or “treatment” may include “preventing” or “prevention” of disease or undesirable condition. In addition, “treating” or “treatment” does not require complete alleviation of signs or symptoms, does not require a cure, and specifically includes protocols that have only a marginal effect on the patient.[000135] “Treatment,” as used herein, thus can cover any treatment of a disease in a subject, particularly in a human, and includes: (a) preventing the disease from occurring in a subject which may be predisposed to the disease but has not yet been diagnosed as having it; (b) inhibiting the disease, i.e., arresting its development; and (c) relieving the disease, i.e., causing regression of the disease and / or relieving one or more disease symptoms. “Treatment” can also encompass delivery of an agent or administration of a therapy in order to provide for a pharmacologic effect, even in the absence of a disease or condition.[000136] The term “therapeutically effective” or “therapeutically effective amount” as used throughout this application can refer to an amount effective to achieve a desired and / or beneficial effect, and / or anything that promotes or enhances the well-being of the subject with respect to the medical treatment of a condition. This includes, but is not limited to, a reduction in the frequency or severity of one or more signs or symptoms of a disease. An effective amount can be administered in one or more administrations. In the methods, a therapeutically effective amount is an amount appropriate to treat an indication. By treating an indication is meant achieving any desirable effect, such as one or more of palliate, ameliorate, stabilize, reverse, slow, or delay disease progression, increase the quality of life, or to prolong life. Such achievement can be measured by any suitable method, such as measurement of tumor size or blood cell count, or any other suitable measurement.[000137] The terms “effective amount” or “effective dose” as used herein have their plain and ordinary meaning as understood in light of the specification, and can refer to that amount of a recited composition or compound that, results in an observable effect. Actual dosage levels of active ingredients in an active composition of the presently disclosed subjectmatter can be varied so as to administer an amount of the active composition or compound that, is effective to achieve the desired response for a particular subject and / or application. The selected dosage level wall depend upon a variety of factors including, but not limited to, the activity of the composition, formulation, route of administration, combination with other drugs or treatments, severity of the condition being treated, and the physical condition and prior medical history of the subject being treated. In some embodiments, a minimal dose is administered, and dose is escalated in the absence of dose-limiting toxicity to a minimally effective amount. Determination and adjustment of an effective dose, as well as evaluation of when and how to make such adjustments, are contemplated herein.[000138] The term “disease state” as used herein, can generally refer to a condition that affects the structure or function of an organism. Disease states can include, for example, stages of a disease progression.[000139] As used herein, a “lung-specific” cell, such as a “lung-specific endothelial cell” or a “lung-specific mesenchymal cell” may refer to a cell having one or more characteristics that are uniquely found or particularly commonly found in the lung. Thus, a “lung-specific endothelial cell” may refer to a certain endothelial cell, or type of endothelial cell, having one or more characteristics uniquely found or particularly commonly found in lungs, and a “lung-specific mesenchymal cell” may refer to a certain mesenchymal cell, or type of mesenchymal cell, having one or more characteristics uniquely found or particularly commonly found in lungs.[000140] As used herein, the term “assessing” can include any form of measurement, and includes determining if an element is present or not. The terms “determining,” “measuring,” “evaluating,” “assessing” and “assaying” can be used interchangeably and can include quantitative and / or qualitative determinations.[000141] As used herein, the terms “modulated” or “modulation,” or “regulated” or “regulation” and “differentially regulated” can refer to both up regulation (i.e., activation or stimulation, e.g., by agonizing or potentiating) and down regulation (i.e., inhibition or suppression, e.g., by antagonizing, decreasing or inhibiting), unless otherwise specified or clear from the context of a specific usage.[000142] As used herein, the term “subject” can refer to any member of the animal kingdom. In some embodiments, a subject is a human patient.[000143] As used herein, the term “marker” or “biomarker” can refer to any measurable substance taken as a sample from a subject whose presence is indicative of some phenomenon. Non-limiting examples of such phenomenon can include a disease state, a condition, or exposure to a compound or environmental condition. In various embodiments described herein, biomarkers may be used for diagnostic purposes (e.g., to diagnose a disease state, a health state, an asymptomatic state, a symptomatic state, etc.). The term “biomarker” may be used interchangeably with the term “marker”. The term “marker” or “biomarker” can include a biological molecule, such as, for example, a nucleic acid, peptide, protein, hormone, and the like, whose presence or concentration can be detected and correlated with a known condition, such as a disease state. It can also be used to refer to a differentially expressed gene whose expression pattern can be utilized as part of a predictive, prognostic or diagnostic process in healthy conditions or a disease state, or which, alternatively, can be used in methods for identifying a useful treatment or prevention therapy.[000144] As used herein, the term “cellular phenotype” can refer to any determinable, observable, and / or measurable characteristic associated with a cell population.[000145] As used herein, a “model” can include one or more in vitro or in vivo disease models; a model can also include algorithms, one or more mathematical techniques, one or more machine learning algorithms, or a combination thereof. A model can be used in a process and / or applied to an assay, in accordance with various embodiments as disclosed herein.[000146] As used herein, a “process” can include one or more steps involving one or more features of one or more model as disclosed herein.[000147] The terms “function” and “functional” as used herein have their plain and ordinary meaning as understood in light of the specification, and can refer to a biological, enzymatic, or therapeutic function.[000148] The term “inhibit” as used herein has its plain and ordinary meaning as understood in light of the specification, and can refer to the reduction or prevention of a biological activity. The reduction can be by a percentage that is, is about, is at least, is at least about, is not more than, or is not more than about, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%, or an amount that is within a range defined by any two of the aforementioned values. As used herein, the term “delay” has its plain and ordinary meaning as understood in light of the specification, and refers to a slowing, postponement, or defermentof a biological event, to a time which is later than would otherwise be expected. The delay can be a delay of a percentage that is, is about, is at least, is at least about, is not more than, or is not more than about, 0%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, or an amount within a range defined by any two of the aforementioned values. The terms inhibit and delay may not necessarily indicate a 100% inhibition or delay, A partial inhibition or delay may be realized.[000149] As used herein, the term “isolated” has its plain and ordinary meaning as understood in light of the specification, and can refer to a substance and / or entity that has been (1) separated from at least some of the components with which it was associated when initially produced (whether in nature and / or in an experimental setting), and / or (2) produced, prepared, and / or manufactured by the hand of man. Isolated substances and / or entities may be separated from equal to, about, at least, at least about, not more than, or not more than about, 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, about 95%, about 98%, about 99%, substantially 100%, or 100% of the other components with which they were initially associated (or ranges including and / or spanning the aforementioned values). In some embodiments, isolated agents are, are about, are at least, are at least about, are not more than, or are not more than about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, substantially 100%, or 100% pure (or ranges including and / or spanning the aforementioned values). As used herein, a substance that is “isolated” may be “pure” (e g., substantially free of other components). As used herein, the term “isolated cell” can refer to a cell not contained in a multi -cellular organism or tissue.[000150] As used herein, “ / / / vivo" is given its plain and ordinary meaning as understood in light of the specification and can refer to the performance of a method inside living organisms, usually animals, mammals, including humans, and plants, as opposed to a tissue extract or dead organism.[000151] As used herein, “ex vivo" is given its plain and ordinary meaning as understood in light of the specification and can refer to the performance of a method outside a living organism with little alteration of natural conditions.[000152] As used herein, “ / / / vitro” is given its plain and ordinary' meaning as understood in light of the specification and can refer to the performance of a method outside of biological conditions, e.g., in a petri dish or test tube.[000153] The terms “nucleic acid” or “nucleic acid molecule” as used herein have their plain and ordinary meaning as understood in light of the specification, and can refer to polynucleotides, such as deoxyribonucleic acid (DNA) or ribonucleic acid (RNA), oligonucleotides, those that appear in a cell naturally, fragments generated by the polymerase chain reaction (PCR), and fragments generated by any of ligation, scission, endonuclease action, and exonuclease action. Nucleic acid molecules can be composed of monomers that are naturally-occurring nucleotides (such as DNA and RNA), or analogs of naturally-occurring nucleotides (e.g., enantiomeric forms of naturally-occurring nucleotides), or a combination of both. Modified nucleotides can have alterations in sugar moieties and / or in pyrimidine or purine base moieties. Sugar modifications include, for example, replacement of one or more hydroxyl groups with halogens, alkyl groups, amines, and azido groups, or sugars can be functionalized as ethers or esters. Moreover, the entire sugar moiety can be replaced with statically and electronically similar structures, such as aza- sugars and carbocyclic sugar analogs. Examples of modifications in a base moiety include alkylated purines and pyrimidines, acylated purines or pyrimidines, or other well-known heterocyclic substitutes. Nucleic acid monomers can be linked by phosphodiester bonds or analogs of such linkages. Analogs of phosphodi ester linkages include phosphorothioate, phosphorodithioate, phosphoroselenoate, phosphorodi selenoate, phosphoroanilothioate, phosphoramlidate, or phosphoramidate. The term “nucleic acid molecule” also includes so-called “peptide nucleic acids,” which comprise naturally-occurring or modified nucleic acid bases attached to a polyamide backbone. Nucleic acids can be either single stranded or double stranded. “Oligonucleotide” can be used interchangeable with nucleic acid and can refer to either double stranded or single stranded DNA or RNA. A nucleic acid or nucleic acids can be contained in a nucleic acid vector or nucleic acid construct (e.g. plasmid, virus, retrovirus, lentivirus, bacteriophage, cosmid, fosmid, phagemid, bacterial artificial chromosome (BAG), yeast artificial chromosome (YAC), or human artificial chromosome (HAG)) that can be used for amplification and / or expression of the nucleic acid or nucleic acids in various biological systems. Typically, the vector or construct will also contain elements including but not limitedto promoters, enhancers, terminators, inducers, ribosome binding sites, translation initiation sites, start codons, stop codons, polyadenylation signals, origins of replication, cloning sites, multiple cloning sites, restriction enzyme sites, epitopes, reporter genes, selection markers, antibiotic selection markers, targeting sequences, peptide purification tags, or accessory genes, or any combination thereof.[000154] A nucleic acid or nucleic acid molecule can comprise one or more sequences encoding different peptides, polypeptides, or proteins. These one or more sequences can be joined in the same nucleic acid or nucleic acid molecule adjacently, or with extra nucleic acids in between, e.g. linkers, repeats or restriction enzyme sites, or any other sequence that is, is about, is at least, is at least about, is not more than, or is not more than about, I, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 150, 200, or 300 bases long, or any length in a range defined by any two of the aforementioned lengths. The term “downstream” on a nucleic acid as used herein has its plain and ordinary meaning as understood in light of the specification and refers to a sequence being after the 3 ’-end of a previous sequence, on the strand containing the encoding sequence (sense strand) if the nucleic acid is double stranded. The term “upstream” on a nucleic acid as used herein has its plain and ordinary meaning as understood in light of the specification and refers to a sequence being before the 5’- end of a subsequent sequence, on the strand containing the encoding sequence (sense strand) if the nucleic acid is double stranded. The term “grouped” on a nucleic acid as used herein has its plain and ordinary meaning as understood in light of the specification and refers to two or more sequences that occur in proximity either directly or with extra nucleic acids in between, e.g. linkers, repeats, or restriction enzyme sites, or any other sequence that is, is about, is at least, is at least about, is not more than, or is not more than about, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 150, 200, or 300 bases long, or any length in a range defined by any two of the aforementioned lengths, but generally not with a sequence in between that encodes for a functioning or catalytic polypeptide, protein, or protein domain.[000155] The nucleic acids described herein comprise nucleobases. Primary, canonical, natural, or unmodified bases are adenine, cytosine, guanine, thymine, and uracil. Other nucleobases include but are not limited to purines, pyrimidines, modified nucleobases, 5- methylcytosine, pseudouridine, dihydrouridine, inosine, 7-methylguanosine, hypoxanthine,xanthine, 5, 6-di hydrouracil, 5-hydroxymethylcytosine, 5-bromouracil, isoguanine, isocytosine, aminoallyl bases, dye-labeled bases, fluorescent bases, or biotin-labeled bases.[000156] The terms “peptide”, “polypeptide”, and “protein” as used herein have their plain and ordinary meaning as understood in light of the specification and refer to macromolecules comprised of amino acids linked by peptide bonds. The numerous functions of peptides, polypeptides, and proteins are known in the art, and include but are not limited to enzymes, structure, transport, defense, hormones, or signaling. Peptides, polypeptides, and proteins are often, but not always, produced biologically by a ribosomal complex using a nucleic acid template, although chemical syntheses are also available. By manipulating the nucleic acid template, peptide, polypeptide, and protein mutations such as substitutions, deletions, truncations, additions, duplications, or fusions of more than one peptide, polypeptide, or protein can be performed. These fusions of more than one peptide, polypeptide, or protein can be joined in the same molecule adjacently, or with extra amino acids in between, e.g. linkers, repeats, epitopes, or tags, or any other sequence that is, is about, is at least, is at least about, is not more than, or is not more than about, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 150, 200, or 300 bases long, or any length in a range defined by any two of the aforementioned lengths. The term “downstream” on a polypeptide as used herein has its plain and ordinary meaning as understood in light of the specification and refers to a sequence being after the C- terminus of a previous sequence. The term “upstream” on a polypeptide as used herein has its plain and ordinary meaning as understood in light of the specification and refers to a sequence being before the N- terminus of a subsequent sequence.[000157] The term “purity” of any given substance, compound, or material as used herein has its plain and ordinary meaning as understood in light of the specification and can refer to the actual abundance of the substance, compound, or material relative to the expected abundance. For example, the substance, compound, or material may be at least 80, 85, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% pure, including all decimals in between. Purity may be affected by unwanted impurities, including but not limited to nucleic acids, DNA, RNA, nucleotides, proteins, polypeptides, peptides, amino acids, lipids, cell membrane, cell debris, small molecules, degradation products, solvent, carrier, vehicle, or contaminants, or any combination thereof. In some embodiments, the substance, compound, or material issubstantially free of host cell proteins, host cell nucleic acids, plasmid DNA, contaminating viruses, proteasomes, host cell culture components, process related components, mycoplasma, pyrogens, bacterial endotoxins, and adventitious agents. Purity' can be measured using technologies including but not limited to electrophoresis, SDS-PAGE, capillary electrophoresis, PCR, rtPCR, qPCR, chromatography, liquid chromatography, gas chromatography, thin layer chromatography, enzyme-linked immunosorbent assay (ELISA), spectroscopy, UV-visible spectrometry, infrared spectrometry, mass spectrometry, nuclear magnetic resonance, gravimetry, or titration, or any combination thereof.[000158] The term “yield” of any given substance, compound, or material as used herein has its plain and ordinary meaning as understood in light of the specification and can refer to the actual overall amount of the substance, compound, or material relative to the expected overall amount. For example, the yield of the substance, compound, or material is, is about, is at least, is at least about, is not more than, or is not more than about 80, 85, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% of the expected overall amount, including ail decimals in between. Yield may be affected by the efficiency of a reaction or process, unwanted side reactions, degradation, quality of the input substances, compounds, or materials, or loss of the desired substance, compound, or material during any step of the production.[000159] The term “% w / w” or “% wt / wt” as used herein has its plain and ordinary meaning as understood in light of the specification and refers to a percentage expressed in terms of the weight of the ingredient or agent over the total weight of the composition multiplied by 100. The term “% v / v” or “% vol / vol” as used herein has its plain and ordinary meaning as understood in the light of the specification and refers to a percentage expressed in terms of the liquid volume of the compound, substance, ingredient, or agent over the total liquid volume of the composition multiplied by 100.[000160] The term “basement membrane matrix” or “extracellular matrix” as used herein has its plain and ordinary meaning m light of the specification and refers to any biological or synthetic compound, substance, or composition that enhances cell attachment and / or growth. Any extracellular matrix, as well as any mimetic or derivative thereof, known in the art can be used for the methods disclosed herein. Some examples of extracellular matrices, or numerics or derivative thereof, include but are not limited to cell-based feeder layers, polymers, proteins, polypeptides, nucleic acids, sugars, lipids, poly-lysine, poly-ornithine, collagen, gelatin, fibronectin, vitronectin, laminin, elastin, tenascin, heparan sulfate, entactin, osteopontin, basement membrane, Matrigel, hydrogel, PEI, WGA, or hyaluronic acid, or any combination thereof.[000161] The terms “passage” and “passaging” as used herein have their plain and ordinary meaning as understood in light of the specification, and refer to the conventional approaches performed in biological cell culture methods to maintain a viable population of cells for prolonged periods of time. As cells are generally proliferative in cell culture, they undergo multiple cycles of mitosis until occupying the available space, which is typically a surface of a cell culture container (e.g., a plate, dish, or flask) submerged under culture medium. For example, the cells may grow out as a monolayer on a cell culture container surface. If the growing cells occupy the entire available space of surface, they cannot proliferate further and may exhibit senescent behavior. In order to continue growth of the cells, which may be performed to maintain the viability and proliferative nature of the cells and / or to expand the number of cells for downstream purposes, the cells may be passaged by taking a fraction of the cells and seeding this fraction onto a fresh surface (e.g., of a cell culture container) in culture medium. This fraction of the cells will continue to proliferate and multiply until they occupy the available space of the new surface, upon which this passaging can be repeated successively.[000162] The term “organoid” as used herein refers to a population of cells differentiated in vitro to form self-organizing structures, which generally are three-dimensional (3D), and include one or more functional cell types. As such, organoids are man-made, artificial constructs which undergo controlled differentiation in order to provide a specific desired cell population or combination of cell populations. Organoids can have one or more cell types and / or structural and / or functional characteristics which recapitulate one or more such feature (e.g. cell type, structural characteristic, and / or functional characteristic) of a specific type of tissue or organ. Many types of organoids have been developed to date and are known in the art and can recapitulate features of, for example, intestinal, colonic, gastric, esophageal, liver, pancreatic, kidney, and / or other types of tissues and / or organs. As organoids are differentiated in vitro, by human direction (e.g. through carefully selected promotion and / or inhibition of one or more signaling pathways, addition / removal of growth factors, culture conditions, etc.), they are generally differ from naturally occurring tissue in a number of ways,such as in overall maturity, cell types present, ratios of cells types, structural features, functional characteristics, shape, size, and / or other such qualities.[000163] The term “assembloid” as used herein refers to a construct including one or more organoids, in combination with one or more additional cells, or cell types, in addition to those of the one or more organoids. In some embodiments, the one or more additional cells, or cell types, of the assembloid construct are external to, and / or cultured separately from, the organoid(s) of the assembloid construct prior to being combined. In some embodiments, the one or more additional cells, or cell types, of the assembloid construct can be found in the organoid(s) of the assembloid construct. In some embodiments, the one or more additional cells, or cell types, of the assembloid construct are not present in the organoid(s) of the assembloid construct. The assembloid construct can be an engineered construct.[000164] The term “engineered” as used herein refers to an entity that is generated by the hand of man, including a cell, combination of cells, organoid, nucleic acid, polypeptide, vector, and so forth. In at least some cases, an engineered entity is synthetic and comprises elements that are not naturally present or configured in the manner in which it is utilized in the disclosure. In certain embodiments, a construct and / or vector is engineered through recombinant nucleic acid technologies, and a cell is engineered through transfection or transduction of an engineered vector. Cells may be engineered to express heterologous proteins that are not naturally expressed by the cells, either because the heterologous proteins are recombinant or synthetic or because the cells do not naturally express the proteins.Overview[000165] Impaired regeneration of the alveolar capillaries after hyperoxia and / or ventilation-induced lung injury is commonly observed in patients with bronchopulmonary dysplasia (BPD). As described herein, various embodiments of the present disclosure involve cutting-edge 3D multi-cellular in vitro systems (e.g., human induced pluripotent stem cell (iPSC)-derived vessel organoids and vascularized lung organoids) and employ state-of-art multi omic measures of human lung tissues (e.g., spatial transcriptomics and multi ome) to study impaired regeneration of alveolar capillaries after hyperoxia and / or ventilation-induced lung injury. The present disclosure also elucidates the molecular mechanism by which truncated NTRK2 contributes to the dysfunction of alveolar capillary endothelial cells (ECs), anddetermines the therapeutic potential of NTRK2 activation in human organoid and murine models of hyperoxia-induced injury.Elucidation of the mechanism by which truncated NTRK2 contributes to the abnormal regeneration of alveolar capillaries following hyperoxic lung injury.[000166] NTRK2 (Neurotrophic Receptor Tyrosine Kinase 2) encodes a member of the neurotrophic tyrosine receptor kinase (NTRK, also known as Tropomyosin receptor kinase B, TrkB) family. Upon ligand binding, NTRK2 phosphorylates itself and members of the MAPK pathway. NTRK2 signaling is critical to the development of the central and peripheral nervous system. Its strong pro-survival function on neurons has attracted interest as a potential therapeutic target for neurodegenerative disorders such as Parkinson’s and Alzheimer’s disease. In addition to its known roles in brain development and neurological diseases, there is increasing recognition that NTRK2 is also expressed in peripheral tissues including the lung, where it contributes to both normal lung development and responses to lung injury including neonatal and adult asthma, pulmonary hypertension, acid-induced lung injury, viral injury, and lung cancer.[000167] The NTRK2 gene, in addition to the full-length receptor, encodes a number of isoforms (e.g., as shown in Fig. 1A). The full-length transmembrane NTRK2 receptor (NTRK2-FL) contains an intracellular tyrosine kinase domain that is essential for activating downstream signaling such as MAPK-ERK, PI3K / Akt, and NF-kB, all of which can modulate cell survival, proliferation, and angiogenesis. The most widely expressed truncated isoform is NTRK2-T1, which lacks a tyrosine kinase domain, but possesses alternative cytoplasmic domains that signal through less extensively characterized mechanisms. In the brain, NTRK2- T1 sequesters its ligand brain-derived neurotrophic factor (BDNF) to prevent signaling. In addition to its role as a dominant-negative receptor, NTRK2-T1 stimulates calcium signaling, regulates Rho GTPase signaling, and modifies cytoskeletal structures (Fig. IB). However, the role of NTRK2 isoforms in human alveolar capillary development and disease remains heretofore unknown.[000168] As described herein, a multi omic experiment was conducted, showing that the expression of NTRK2-T1, but not other isoforms, was significantly increased in alveolar capillary ECs from patients with BPD, and in murine lungs with impaired alveolar regenerationafter hyperoxic injury. The present disclosure identifies upstream regulators and downstream signaling of NTRK2-T1 in abnormal ECs lacking regenerative capacity in response to hyperoxia-induced lung injury.Analysis of hyperoxia-induced abnormal lung-specific endothelial cell regeneration in BPD at single cell resolution.[000169] Although the etiology of BPD is multifactorial, the most common causes of BPD in preterm infants are the exposure to hyperoxia and mechanical ventilation. BPD affects nearly 15,000 infants annually in the US. It also increases the risks of chronic respiratory and cardiovascular diseases, growth failure, and neurodevelopmental delay in preterm infants. While advances in neonatal care have resulted in improved survival rates of premature infants, limited progress has been made in reducing rates of BPD due to poor understanding of the pathogenesis. BPD is characterized by simplified alveoli, fewer and dysmorphic vessels, and hypertensive vascular remodeling. Accumulating evidence indicates that disrupted pulmonary angiogenesis following hyperoxia injury is the key to the pathogenesis of BPD. Pathologic examination of lung tissue from infants dying from the BPD, or long-term ventilated preterm infants found evidence of either decreased and abnormally distributed pulmonary microvessels, or simplified and immature capillaries.[000170] The alveolar capillary network is composed of two intermingled lungspecific endothelial cell (EC) subtypes: (1) aerocyte capillary (aCap) cells and general capillary (gCap) cells. The aCap cell is unique to the lung and specialized for gas exchange, whereas the gCap cell functions as a progenitor cell in capillary homeostasis and repair after injury. However, the alveolar capillary EC subtype-specific pathology in BPD patients remains unknown. The present disclosure demonstrates that the abundance of a gCap cell subcluster, gCap-C2, was drastically increased in lung tissue from patients with BPD. The existing infrastructure of the NIH funded LungMAP2 consortium was leveraged to collect human BPD lung tissue and performed multiomic sequencing. By combining these resources with organoid and murine models of hyperoxia-induced vascular injury, the gCap cell-mediated mechanisms underlying impaired regeneration of alveolar capillary beds are investigated in response to hyperoxic injury. The present disclosure also assesses how abnormal gCap cells affect surrounding cell types at various stages of lung regeneration and repair.Targeting NTRK2 signaling for alveolar regeneration and repair in BPD and similar lung disorders.[000171] Current therapies for BPD are primarily supportive and lacking effective therapies promoting repair and regeneration of the damaged alveoli and pulmonary vasculature. There is an unmet need for developing therapies that enhance regeneration of arrested alveolar and vasculature in injured lungs. Enhancing angiogenesis by augmenting VEGF (vascular endothelial growth factor) signaling improved alveolarization in the presence of injury; however, severe side effects of VEGF therapy hinder its use in neonates with BPD. NTRK2 activation attenuates pro-atherosclerotic factors-induced EC hyperpermeability, and promotes neovascularization, EC migration, vascular relaxation, and coronary vessel development.[000172] The present disclosure demonstrates that overexpressing NTRK2-FL in human iPSC-derived vessel organoids (VOs) significantly improves capillary formation and cell proliferation in response to hyperoxia-induced injury. Some embodiments of the present disclosure further describe and document the efficacy of a novel and nonobvious therapeutic techniques for lipid nanoparticle delivery of the NTRK2-FL in restoring the alveolar capillary development and function in hyperoxia-induced BPDlike mouse and organoid models. In some embodiments, lipid nanoparticles specifically targeting neonatal pulmonary capillary ECs are utilized, which reduces potential off-target effects of small molecule activators or synthetic peptides. Since the change of NTRK2 expression level is not only observed in BPD but also in asthma, inflammation, and lung cancer, therapies that activate NTRK2 signaling can be used as therapies for variety of lung disorders. Accordingly, various embodiments of the disclosure relate to the use of therapies that activate NTRK2 signaling for treating lung disorders.Applying human iPSC-derived organoid systems to investigate vascular defects and their impact on neighboring cells in hyperoxia-induced lung injury.[000173] While analysis of lung tissues from deceased BPD patients has provided valuable insights related to the advanced stages of the disease, this approach does not provide insight into the early pathological changes in BPD, or the cellular abnormality directly caused by hyperoxic exposure. To investigate the impaired response to hyperoxia in human vascularcell s, human iPSCs-derived VOs were generated and subjected to hyperoxia. These VOs comprise well-formed and functional vascular networks composed of ECs, pericytes, basement membrane, and a lumen of about 7 -13 pm in diameter (e.g., about 10pm in diameter) resembling a human capillary network. The data demonstrates that tube formation and cell proliferation were significantly reduced in VOs exposed to hyperoxia, consistent with the arrested vascular growth observed in lungs from BPD patients. The present disclosure also describes a complex lung organoid system that consists of lung-specific endothelium, mesenchyme, and epithelium. These iPSC-derived organoid models provide a unique complement to the existing hyperoxia-induced BPD-like mouse model, enabling the study of lung-specific endothelial cell phenotypes and aberrant cell-cell crosstalk specific to hyperoxia- induced lung injury within a human context.Exemplary process for generating iPSC-derived vessel organoids, such as NTRK2 vessel organoid comprising a hyperoxic injury (NTRK2+ HIVO)[000174] Fig. 2C is a schematic showing an example process for generating iPSC- derived vessel organoids, according to example embodiments of the present disclosure. In various embodiments, the iPSC-derived vessel organoids may include an NTRK2 vessel organoid comprising a hyperoxic injury (NTRK2+ HIVO) and a control NTRK2 vessel organoid developed under normoxic conditions. As shown in FIG. 2C, the example process may involve producing an angioblast spheroid from iPSCs, and using the angioblast spheroids to produce iPSC-derived vessel organoids (e.g., the NTRK2+ HIVO and the control NTRK2 vessel organoid developed under normoxic conditions).[000175] An example method for producing the angioblast spheroids can include: a) culturing iPSCs to form an embryoid body; and b) activating a WNT signaling pathway and a BMP signaling pathway of the embryoid body for a period of time sufficient to form a spheroid comprising mesoderm cells. For example, such period of time may be about 2 days to about 4 days. One skilled in the art will appreciate that various specific components can be used, alone or in combination, in order to achieve activation (or inhibition) of a signaling pathway.[000176] In some embodiments, activating the WNT signaling pathway may be performed by contacting the embryoid body with an activator of the WNT signaling pathway. In some embodiments, the WNT signaling pathway activator may be Wntl, Wnt2, Wnt2b,Wnt3, Wnt3a, Wnt4, Wnt5a, Wnt5b, Wnt6, Wnt7a, Wnt7b, Wnt8a, Wnt8b, Wnt9a, Wnt9b, WntlOa, WntlOb, Wntl l, or Wntl6, or any combination thereof. In some embodiments, the Wnt signaling pathway activator comprises a glycogen synthase kinase-3 (GSK3) inhibitor, which acts as a Wnt signaling pathway activator. In some embodiments, the GSK3 inhibitor is CHIR99021. Furthermore, activating the BMP signaling pathway may be performed by contacting the embryoid body with an activator of the BMP signaling pathway, such as but not limited to BMP1, BMP2, BMP3, BMP4, BMP5, BMP6, BMP7, BMP8a, BMP8b, BMP 10, BMP 11, BMP 15, IDE1, or IDE2.[000177] The example method may further comprise c) activating a VEGF signaling pathway of the spheroid comprising mesoderm cells with forskolin for a period of time sufficient to form the angioblast spheroid. The period of time may be about 1 day to about 3 days. The VEGF signaling pathway may be activated by culturing the contents (e g., the spheroid comprising mesoderm cells) with VEGF or one or more other VEGF signaling pathway activators (e.g., type V receptor tyrosine kinases (RTKs), VEGFR-1, -2 and -3, coreceptors for the VEGF signaling pathway, such as neuropilins and heparan sulfate proteoglycans (HSPG)).[000178] In various embodiments, as shown in Fig. 2C, the example process for producing the iPSC-derived vessel organoids may include: a) activating a vascular endothelial growth factor (VEGF) signaling pathway, a fibroblast growth factor (FGF) signaling pathway and a brain-derived neurotrophic factor (BDNF) signaling pathway of an angioblast spheroid in fetal bovine serum (FBS) to form a spheroid. In order to produce a control NTRK2+ vessel organoid, step a) may be performed in normoxic conditions.[000179] The example process for generating the iPSC-derived vessel organoids may further include: b) activating a VEGF signaling pathway, a FGF signaling pathway, and a BDNF signaling pathway of the spheroid in a normoxic culture condition with FBS. The normoxic culture condition is, is about, is at least about, or is not more than about 10%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, or 35% O2 , or a range defined by any two of the preceding values, such as about 18% to about 24% O2, 20% O2 to about 23% O2, or optionally about 21% O2.[000180] In some embodiments, in order to produce NTRK2+ HIVO, the aforementioned signaling pathways of the angioblast spheroid may be activated in a hyperoxicculture condition for a first period of time sufficient to form a spheroid comprising acute hyperoxic injury (AHI). The hyperoxic culture condition is, is about, is at least about, or is not more than, 30%, 35%, 40%, 50%, 60%, 70%, 80%, 90% O2 , or a range defined by any two of the preceding values, such as about 30% to about 80% O2, 50% O2 to about 80% O2, or optionally about 75% O2. For example, as shown in FIG. 2C, the hyperoxic condition may be about 40% O2. The first period of time may be about 4 days to about 6 days. Furthermore, as the aforementioned spheroid in such embodiments may comprise a spheroid comprising AHI, the example process to produce NTRK2+ HIVO can subsequently include: b) activating a VEGF signaling pathway, a FGF signaling pathway, and a BDNF signaling pathway of the spheroid comprising AHI in a normoxic culture condition with FBS for a second period of time sufficient to form the NTRK2+HIVO. The second period of time may be about 4 days to about 6 days. The NTRK2+HIVO thus produced may comprise increased NTRK2 -truncated isoform 1 (NTRK2-T1) expression compared to the control NTRK2+vessel organoid (NTRK2+VO) cultured in normoxic culture condition.[000181] In some embodiments, activating a VEGF signaling pathway (e.g., of the angioblast spheroid, the spheroid comprising acute hyperoxic injury (AHI), etc.) may comprise culturing with VEGF or other activator or co-receptor of the VEGF signaling pathway. In some embodiments, activating a FGF signaling pathway (e.g., of the angioblast spheroid, the spheroid comprising acute hyperoxic injury (AHI), etc.) may comprise culturing with FGF2 or other activator or co-receptor of the FGF signaling pathway. In some embodiments, activating a BDNF signaling pathway (e.g., of the angioblast spheroid, the spheroid comprising acute hyperoxic injury (AHI), etc.) may comprise culturing with BDNF or other activator or co- receptor of the BDNF signaling pathway.Elucidation of the role of NTRK2 isoforms in alveolar capillary injury and repair.[000182] The pro-survival role of the BDNF-NTRK2 signaling has been extensively studied in neuronal systems. However, increasing evidence indicates that NTRK2 is also highly expressed in peripheral tissues including the heart, lung, and pancreas. At present, little is known about the functions and downstream signaling of NTRK2-FL and NTRK2-T1 in alveolar cells. Although altered expression of NTRK2 was associated with a variety of lung disorders, the specific NTRK2 isoforms have heretofore not been examined. A recent studydemonstrated that Ntrk2 is expressed in mouse lung-specific Mesenchymal Alveolar Niche Cells (MANCs). When Ntrk2 was deleted in PDGFRa+ fibroblasts, mice exhibited nonresolving damage to the alveoli in an acid-induced lung injury model in which mice usually recover. This result shows a role of Ntrk2 in alveolar regeneration after injury in mice. The initial data demonstrated that NTRK2 is most enriched in gCap cells compared with other lung cell types including MANCs in both human and mouse. The present disclosure further elucidates the roles of NTRK2 isoforms in human lung gCap cells, and explores the potential utility of targeting NTRK2 as a therapeutic approach for BPD-like lung injuries.Simultaneous profiling of single-nucleus transcriptome and chromatin accessibility in BPD human lung tissue.[000183] Although single cell (sc)RNA-seq has been performed in hyperoxia induced BPD-like mouse models, little has heretofore been known about the molecular basis of the BPD pathology in humans. Utilizing the Chromium Single Cell Multiome (combined single nucleus (sn)RNA and ATAC-seq) platform, RNAs and open chromatin regions were simultaneously profded within the same cell, enabling highly-resolved cell state classification, identification of drivers of differential gene expression in control versus BPD, and identification of cells with similar transcriptional profiles, but with functionally distinct chromatin footprints.[000184] This technique provided accurate and comprehensive information generated from the same cell, avoiding computational challenges in defining cell types by aligning snRNA-seq and snATAC-seq datasets generated separately. The integrative analysis of gene expression profiles and chromatin accessibility enable identification of upstream regulators of NTRK2 in a cellsubtype-specific manner, and gain comprehensive understanding of the NTRK2 gene regulatory network in human alveolar gCap cells.Spatial transcriptomic (ST) profiling of human lung to define local cell-cell communication.[000185] Traditional scRNA-seq analysis lacks spatial information important for studying disease mechanisms. ST enables the investigation of spatially-resolved whole transcriptome mRNA expression at a resolution of 5000 55pm spots across the tissue section,while capturing histological and protein information from the same region by H&E staining or immunostaining. Thus, gene expression profiles can be mapped back to their original anatomical location, which is not possible in standard approaches. Although the resolution is not sufficient to detect transcripts within a single cell, especially in regions with simplified alveoli structure, additional deconvolution steps using human lung scRNA-seq datasets and Multi ome as references for cell type annotation made it possible to quantitatively estimate the cellular composition within each spot. Using ST followed by data deconvolution, cells that reside in close proximity to abnormal gCap-C2 cells in BPD human lung tissue were identified. Subsequent ligand-receptor binding analysis demonstrated altered signaling leading to the disrupted EC-epithelial cell and EC-fibroblasts crosstalk in damaged alveoli.NTRK2 is highly expressed in gCap cells during alveologenesis in human and mouse lungs.[000186] Although Ntrk2 expression is typically enriched in MANCs (e.g., in mouse lung), the expression in other lung cell types has heretofore not been examined. Leveraging the lungMAP scRNA-seq datasets of mouse lungs, the present disclosure describes an assessment c Ntrk2 expression patterns during the saccular, early alveolar, and late alveolar stages of lung development (e.g., as shown in Fig. 2A).[000187] The expression of Ntrk2 peaked in gCap cells at the early alveolar stage, indicating its role in capillary formation during alveologenesis in mice. In contrast to the mouse, human alveologenesis begins in utero before birth and continues in childhood. Consistent with the findings in mice as described herein, NTRK2 was found to be highly expressed in gCap cells during the early alveolar stage, and decreased when the alveologenesis was completed (e.g., as shown in Fig. 2B). Notably, in both human and mouse, NTRK2 was found to be expressed at higher levels in gCap cells compared with MANCs. These data indicate a pivotal role for NTRK2 in capillary formation and alveologenesis.[000188] Further technical details are provided in the Examples and elsewhere herein.Methods of Producing a Neurotrophic Receptor Tyrosine Kinase 2 (NTRK2) Vessel Organoids Comprising a Hyperoxic Injury (NTRK2+HIVO)[000189] Generating various types of organoids from pluripotent stem cells, and precursors thereof, such as definitive endoderm, gut endoderm, foregut endoderm, mid / hindgut endoderm, lateral plate mesoderm, splanchnic mesoderm, splanchnic mesenchyme, vascularized mesoderm, vascularized mesenchyme, and the like, are generally known in the art. Exemplary methods may be found in PCT publications WO 2011 / 140441, WO 2015 / 183920, WO 2016 / 061464, WO 2017 / 192997, WO 2018 / 085622, WO 2018 / 085623, WO 2018 / 106628, WO 2018 / 200481, WO 2018 / 226267, WO 2019 / 074793, WO 2020 / 023245, WO 2020 / 160371, WO 2020 / 243633, WO 2021 / 030373, WO 2021 / 041443, WO 2022 / 072553, WO 2023 / 102133, WO 2023 / 023180, WO 2023 / 137467, and WO 2023 / 278676, each of which is hereby expressly incorporated by reference in its entirety. Any methods for producing definitive endoderm, gut endoderm, foregut endoderm, mid / hindgut endoderm, lateral plate mesoderm, splanchnic mesoderm, splanchnic mesenchyme, vascularized mesoderm, vascularized mesenchyme, and the like, from pluripotent stem cells disclosed herein or otherwise known in the art are applicable to the methods described herein.[000190] In some embodiments, vessel organoids (VOs), such as a Neurotrophic Receptor Tyrosine Kinase 2 (NTRK2)+vessel organoid comprising a hyperoxic injury (NTRK2+HIVO), can be differentiated through the culture of precursor cells, such as definitive endoderm cells, or from a spheroid, such as an angioblast spheroid. In some embodiments, spheroids used to generate the VOs, such as the NTRK2+HIVO, can be generated from precursor cells, such as definitive endoderm.[000191] In some embodiments, progenitor cells, such as induced pluripotent stem cells (iPSCs), can be cultured to form an embryoid body. In some embodiments, the embryoid body can be differentiated by activating and / or inhibiting one or more pathway in the spheroid, such as a WNT signaling pathway and / or a BMP signaling pathway. In some embodiments, WNT and BMP signaling pathways are activated in order to differentiate the embryoid body into a spheroid comprising mesoderm cells. In some embodiments, the differentiation is for at least about 2, 3, 4, or 5 days, or from about 4 days to about 6 days.[000192] In some embodiments, a spheroid comprising mesoderm cells can be cultured to form an angioblast spheroid. In some embodiments, the embryoid body can be differentiated by activating and / or inhibiting one or more pathway in the spheroid, such as a vascular endothelial growth factor (VEGF) signaling pathway, in order to differentiate theembryoid body into an angioblast spheroid. In some embodiments, the differentiation is for at least about 2, 3, 4, or 5 days, or from about 4 days to about 6 days.[000193] In some embodiments, the spheroid, such as an angioblast spheroid, can be differentiated by activating and / or inhibiting one or more pathway in the spheroid, such as a vascular endothelial growth factor (VEGF) signaling pathway, fibroblast growth factor (FGF) signaling pathway, and / or brain-derived neurotrophic factor (BDNF) signaling pathway. In some embodiments, VEGF, FGF, and BDNF signaling pathways are activated in order to differentiate the spheroid. In some embodiments, the differentiation can be carried out in a hyperoxic culture condition in order to provide a spheroid comprising acute hyperoxic injury (AHI). In some embodiments, the differentiation is in the presence of fetal bovine serum (FBS). In some embodiments, the differentiation is for at least about 2, 3, 4, or 5 days, or from about 4 days to about 6 days.[000194] In some embodiments, the spheroid comprising AHI can be differentiated by activating and / or inhibiting one or more pathway in the spheroid, such as a VEGF, FGF, and BDNF signaling pathway. In some embodiments, VEGF, FGF, and BDNF signaling pathways are activated in order to differentiate the spheroid comprising AHI into a NTRK2+HIVO. In some embodiments, the differentiation can be carried out in a normoxic culture condition in order to provide the NTRK2+HIVO. In some embodiments, the differentiation is in the presence of fetal bovine serum (FBS). In some embodiments, the differentiation is for at least about 2, 3, 4, or 5 days, or from about 4 days to about 6 days. In some embodiments, the NTRK2+HIVO can have increased NTRK2-truncated isoform 1 (NTRK2-T1) expression compared to a control NTRK2+vessel organoid (NTRK2+VO) cultured in a normoxic culture condition.[000195] In some embodiments, activating and / or inhibiting includes contacting the cells with the Nodal, Activin, and / or BMP subgroups of the TGF[3 superfamily of growth factors. In some embodiments, the pluripotent stem cells are contacted with Nodal, Activin A, Activin B, BMP4, or any combination thereof, to differentiate the pluripotent stem cells to definitive endoderm. In some embodiments, the pluripotent stem cells can be contacted with Activin A to differentiate the pluripotent stem cells to definitive endoderm.[000196] In some embodiments, the Wnt signaling pathway activator comprises Wntl, Wnt2, Wnt2b, Wnt3, Wnt3a, Wnt4, Wnt5a, Wnt5b, Wnt6, Wnt7a, Wnt7b, Wnt8a,Wnt8b, Wnt9a, Wnt9b, WntlOa, WntlOb, Wntl 1, or Wntl6, or any combination thereof. In some embodiments, the Wnt signaling pathway activator is Wnt3a. In some embodiments, the Wnt signaling pathway activator comprises a glycogen synthase kinase-3 (GSK3) inhibitor, which acts as a Wnt signaling pathway activator. In some embodiments, the GSK3 inhibitor is CHIR99021. In some embodiments, the FGF signaling pathway activator comprises FGF1, FGF2, FGF3, FGF4, FGF4, FGF5, FGF6, FGF7, FGF8, FGF8, FGF9, FGF10, FGF11, FGF12, FGF 13, FGF 14, FGF 15 (FGF 19, FGF15 / FGF19), FGF 16, FGF 17, FGF 18, FGF20, FGF21, FGF22, FGF23, or any combination thereof. In some embodiments, the FGF signaling pathway activator is FGF2. In some embodiments, the BMP signaling pathway activator comprises BMP1, BMP2, BMP3, BMP4, BMP5, BMP6, BMP7, BMP8a, BMP8b, BMP10, BMP11, BMP 15, IDE1, or IDE2, or any combination thereof. In some embodiments, the BMP signaling pathway activator comprises BMP2 or BMP4. In some embodiments, the BMP signaling pathway inhibitor comprises Noggin, RepSox, LY364947, LDN193189, or SB431542, or any combination thereof. In some embodiments, the BMP signaling pathway inhibitor comprises Noggin. In some embodiments, the VEGF signaling pathway inhibitor comprises VEGF. In some embodiments, the BDNF signaling pathway inhibitor comprises BDNF.Stem Cells[000197] The term “totipotent stem cells” (also known as omnipotent stem cells) as used herein has its plain and ordinary meaning as understood in light of the specification and are stem cells that can differentiate into embryonic and extra-embryonic cell types. Such cells can construct a complete, viable organism. These cells are produced from the fusion of an egg and sperm cell. Cells produced by the first few divisions of the fertilized egg are also totipotent.[000198] The term “embryonic stem cells (ESCs),” also commonly abbreviated as ES cells, as used herein has its plain and ordinary meaning as understood in light of the specification and refers to cells that are pluripotent and derived from the inner cell mass of the blastocyst, an early- stage embryo. For purpose of the present disclosure, the term "ESCs" is used broadly sometimes to encompass the embryonic germ cells as well.[000199] The term “pluripotent stem cells (PSCs)” as used herein has its plain and ordinary' meaning as understood in light of the specification and encompasses any cells that can differentiate into nearly all cell types of the body, i.e., cells derived from any of the three germ layers (germinal epithelium), including endoderm (interior stomach lining,gastrointestinal tract, the lungs), mesoderm (muscle, bone, blood, urogenital), and ectoderm (epidermal tissues and nervous system), PSCs can be the descendants of inner cell mass cells of the preimplantation blastocyst or obtained through induction of a non-pluripotent cell, such as an adult somatic cell, by forcing the expression of certain genes. Pluripotent stem cells can be derived from any suitable source. Examples of sources of pluripotent stem cells include mammalian sources, including human, rodent, porcine, and bovine.[000200] The term “induced pluripotent stem cells (iPSCs),” also commonly abbreviated as iPS cells, as used herein has its plain and ordinary meaning as understood in light of the specification and refers to a type of pluripotent stem cells artificially derived from a normally non-pluripotent cell, such as an adult somatic cell, by inducing a "forced" expression of certain genes. hiPSC refers to human iPSCs. In some methods known in the art, iPSCs may be derived by transfection of certain stem cell-associated genes into non-pluripotent cells, such as adult fibroblasts. Transfection may be achieved through viral transduction using viruses such as retroviruses or lentiviruses. Transfected genes may include the master transcriptional regulators Oct-3 / 4 (PUU5F1) and Sox2, although other genes may enhance the efficiency of induction. After 3-4 weeks, small numbers of transfected cells begin to become morphologically and biochemically similar to pluripotent stem cells, and are typically isolated through morphological selection, doubling time, or through a reporter gene and antibiotic selection. As used herein, iPSCs include first generation iPSCs, second generation iPSCs in mice, and human induced pluripotent stem cells. In some methods, a retroviral system is used to transform human fibroblasts into pluripotent stem cells using four pivotal genes: Oct3 / 4, Sox2, Klf4, and c-Myc. In other methods, a lentiviral system is used to transform somatic cells with GCT4, SOX2, NANOG, and LIN28. Genes whose expression are induced in iPSCs include but are not limited to Oct-3 / 4 (POU5F1); certain members of the Sox gene family (e.g., Soxl, Sox2, Sox3, and Soxl5); certain members of the Klf family (e.g., Klfl, Klf2, Klf4, and Klf5), certain members of the Mye family (e.g., C-myc, L-myc, and N- myc), Nanog, LIN28, Tert, Fbxl5, ERas, EC ATI 5-1, ECAT15-2, Tell, b-Catenm, EC ATI, Esgi, Dnmt3L, EC ATS, Gdf3, Fthll7, Sall4, Rexl, UTF1, Stella, Stat3, Grb2, Prdml4, Nr5al, Nr5a2, or E-cadherin, or any combination thereof.[000201] The term “precursor cell” as used herein has its plain and ordinary meaning as understood in light of the specification and encompasses any cells that can be used inmethods described herein, through which one or more precursor cells acquire the ability to renew itself or differentiate into one or more specialized cell types. In some embodiments, a precursor cell is pluripotent or has the capacity to becoming pluripotent. In some embodiments, the precursor cells are subjected to the treatment of external factors (e.g., growth factors) to acquire pluripotency. In some embodiments, a precursor cell can be a totipotent (or omnipotent) stem cell; a pluripotent stem cell (induced or non-induced); a multipotent stem cell; an oligopotent stem cells and a unipotent stem cell. In some embodiments, a precursor cell can be from an embryo, an infant, a child, or an adult. In some embodiments, a precursor cell can be a somatic cell subject to treatment such that pluripotency is conferred via genetic manipulation or protein / peptide treatment. Precursor cells include embryonic stem cells (ESC), embryonic carcinoma cells (ECs), and epiblast stem cells (EpiSC).[000202] In some embodiments, one step can include obtaining stem cells that are pluripotent or can be induced to become pluripotent. In some embodiments, pluripotent stem cells are derived from embryonic stem cells, which are in turn derived from totipotent cells of the early mammalian embryo and are capable of unlimited, undifferentiated proliferation in vitro. Embryonic stem cells are pluripotent stem cells derived from the inner cell mass of the blastocyst, an early-stage embryo. Methods for deriving embryonic stem cells from blastocytes are well known in the art. It would be understood by one of skill in the art that the methods and systems described herein are applicable to any stem cells.[000203] Additional stem cells that can be used in embodiments in accordance with the present disclosure include but are not limited to those provided by or described in the database hosted by the National Stem Cell Bank (NSCB), Human Embryonic Stem Cell Research Center at the University of California, San Francisco (UCSF); WISC cell Bank at the Wi Cell Research Institute; the University of Wisconsin Stem Cell and Regenerative Medicine Center (IJW- SCRMC); Novocell, Inc. (San Diego, Calif.); Cellartis AB (Goteborg, Sweden); ES Cell International Pte Ltd (Singapore); Techmon at the Israel Institute of Technology (Haifa, Israel); and the Stem Cell Database hosted by Princeton University and the University of Pennsylvania. Exemplary embryonic stem cells that can be used in embodiments in accordance with the present disclosure include but are not limited to SA01 (SA001); SA02 (SA002); ESDI (HES-1); ES02 (HES-2); ES03 (HES-3); ES04 (HES-4); ES05 (HES-5); ES06 (HES-6); BG01 (BGN-01); BG02 (BGN-02); BG03 (BGN-03); TE03 (13); TE04 (14); TE06(16); UC01 (HSF1); UC06 (HSF6); WA01 (HI); WA07 (H7); WA09 (H9); WA13 (HI 3); WA14 (HI 4). Exemplary human pluripotent cell lines include but are not limited to TkDA3- 4, 1231 A3, 317-D6, 317-A4, CDH1, 5-T-3, 3-34- 1, NAFLD27, NAFLD77, NAFLD150, WD90, WD91, WD92, 1.20012. C213, 1383D6, FF, or 317-12 cells.[000204] In developmental biology, cellular differentiation is the process by which a less specialized cell becomes a more specialized cell type. As used herein, the term “directed differentiation” describes a process through which a less specialized cell becomes a particular specialized target cell type. The particularity of the specialized target cell type can be determined by any applicable methods that can be used to define or alter the destiny of the initial cell Exemplary methods include but are not limited to genetic manipulation, chemical treatment, protein treatment, and nucleic acid treatment.[000205] In some embodiments, an adenovirus can be used to transport the requisite four genes, resulting in iPSCs substantially identical to embryonic stem cells. Since the adenovirus does not combine any of its own genes with the targeted host, the danger of creating tumors is eliminated, in some embodiments, non-viral based technologies are employed to generate iPSCs. In some embodiments, reprogramming can be accomplished via plasmid without any virus transfection system at all, although at very low efficiencies. In other embodiments, direct deliver}' of proteins is used to generate iPSCs, thus eliminating the need for viruses or genetic modification. In some embodiment, generation of mouse iPSCs is possible using a similar methodology: a repeated treatment of the cells with certain proteins channeled into the cells via poly-arginine anchors was sufficient to induce pluripotency. In some embodiments, the expression of pluripotency induction genes can also be increased by treating somatic cells with FGF2 under low oxygen conditions.[000206] The term “feeder cell” as used herein has its plain and ordinary meaning as understood in light of the specification and refers to cells that support the growth of pluripotent stem cells, such as by secreting growth factors into the medium or displaying on the cell surface. Feeder cells are generally adherent cells and may be growth arrested. For example, feeder cells are growth-arrested by irradiation (e.g. gamma rays), mitomycin-C treatment, electric pulses, or mild chemical fixation (e.g. with formaldehyde or glutaraldehyde). However, feeder cells do not necessarily have to be growth arrested. Feeder cells may serve purposes such as secreting growth factors, displaying growth factors on the cell surface,detoxifying the culture medium, or synthesizing extracellular matrix proteins. In some embodiments, the feeder cells are allogeneic or xenogeneic to the supported target stem cell, which may have implications m downstream applications. In some embodiments, the feeder cells are mouse cells. In some embodiments, the feeder cells are human cells. In some embodiments, the feeder cells are mouse fibroblasts, mouse embryonic fibroblasts, mouse STO cells, mouse 3T3 cells, mouse SNL 76 / 7 cells, human fibroblasts, human foreskin fibroblasts, human dermal fibroblasts, human adipose mesenchymal cells, human bone marrow mesenchymal cells, human amniotic mesenchymal cells, human amniotic epithelial cells, human umbilical cord mesenchymal cells, human fetal muscle cells, human fetal fibroblasts, or human adult fallopian tube epithelial cells. In some embodiments, conditioned medium prepared from feeder cells is used in lieu of feeder cell co-culture or in combination with feeder cell co-culture. In some embodiments, feeder cells are not used during the proliferation of the target stem cells.Differentiation of PSCs[000207] Known methods for producing definitive endoderm from pluripotent cells (e.g., iPSCs or ESCs) are applicable to the methods described herein. In some embodiments, pluripotent cells are derived from a morula. In some embodiments, pluripotent stem cells are stem cells. Stem cells used in these methods can include, but are not limited to, embryonic stem cells or induced pluripotent stem cells. Embryonic stem cells can be derived from the embryonic inner cell mass or from the embryonic gonadal ridges. Embryonic stem cells or germ cells can originate from a variety of animal species including, but not limited to, various mammalian species including humans. In some embodiments, human embryonic stem cells are used to produce definitive endoderm. In some embodiments, human embryonic germ cells are used to produce definitive endoderm. In some embodiments, iPSCs are used to produce definitive endoderm, in some embodiments, human iPSCs (hiPSCs) are used to produce definitive endoderm.[000208] In some embodiments, PSCs, such as ESCs and iPSCs, undergo directed differentiation into embryonic germ layer cells, organ tissue progenitor cells, and then into tissue such as GI tissue or any other biological tissue. In some embodiments, the directed differentiation is done in a stepwise manner to obtain each of the differentiated cell types where molecules (e.g. growth factors, ligands, agonists, antagonists) are added sequentially asdifferentiation progresses. In some embodiments, the directed differentiation is done in a nonstepwise manner where molecules (e.g. growth factors, ligands, agonists, antagonists) are added at the same time. In some embodiments, directed differentiation is achieved by selectively activating certain signaling pathways in the PSCs or any downstream cells.[000209] In some embodiments, the embryonic stem cells or germ cells or iPSCs are treated with one or more small molecule compounds, activators, inhibitors, or growth factors for a time that is, is about, is at least, is at least about, is not more than, or is not more than about, 6 hours, 12 hours, 18 hours, 24 hours, 36 hours, 48 hours, 60 hours, 72 hours, 84 hours, 96 hours, 120 hours, 150 hours, 180 hours, 240 hours, 300 hours or any time within a range defined by any two of the aforementioned times, for example 6 hours to 300 hours, 24 hours to 120 hours, 48 hours to 96 hours, 6 hours to 72 hours, or 24 hours to 300 hours, in some embodiments, more than one small molecule compounds, activators, inhibitors, or growth factors are added. In these cases, the more than one small molecule compounds, activators, inhibitors, or growth factors can be added simultaneously or separately.[000210] In some embodiments, the embryonic stem cells or germ cells or iPSCs are treated with one or more small molecule compounds, activators, inhibitors, or growth factors at a concentration that is, is about, is at least, is at least about, is not more than, or is not more than about, 10 ng / mL, 20 ng / mL, 50 ng / mL, 75 ng / mL, 100 ng / mL, 120 ng / mL, 150 ng / mL, 200 ng / mL, 500 ng / mL, 1000 ng / mL, 1200 ng / mL, 1500 ng / mL, 2000 ng / mL, 5000 ng / mL, 7000 ng / niL, 10000 ng / mL, or 15000 ng / mL, or any concentration that is within a range defined by any two of the aforementioned concentrations, for example, 10 ng / mL to 15000 ng / mL, 100 ng / mL to 5000 ng / mL, 500 ng / mL to 2000 ng / mL, 10 ng / mL to 2000 ng / mL, or 1000 ng / mL to 15000 ng / mL. In some embodiments, concentration of the one or more small molecule compounds, activators, inhibitors, or growth factors is maintained at a constant level throughout the treatment. In some embodiments, concentration of the one or more small molecule compounds, activators, inhibitors, or growth factors is varied during the course of the treatment. In some embodiments, more than one small molecule compounds, activators, inhibitors, or growth factors are added. In these cases, the more than one small molecule compounds, activators, inhibitors, or growth factors can differ in concentrations.[000211] In some embodiments, the ESCs or iPSCs, or the ESCs, germ cells, or iPSCs are cultured in growth media that supports the growth of stem cells. In some embodiments, theESCs or iPSCs, or the ESCs, germ cells, or iPSCs, are cultured in stem cell growth media. In some embodiments, the stem cell growth media is RPMI 1640, DMEM, DMEM / F12, or Advanced DMEM / F12. In some embodiments, the stem cell growth media comprises fetal bovine serum (FBS). In some embodiments, the stem cell growth media comprises FBS at a concentration that is, is about, is at least, is at least about, is not more than, or is not more than about, 0%, 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0,6%, 0.7%, 0.8%, 0,9%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, or 20%, or any percentage within a range defined by any two of the aforementioned concentrations, for example 0% to 20%, 0.2% to 10%, 2% to 5%, 0% to 5%, or 2% to 20%. In some embodiments, the stem cell growth media does not contain xenogeneic components. In some embodiments, the growth media comprises one or more small molecule compounds, activators, inhibitors, or growth factors.[000212] In some embodiments, populations of cells enriched in definitive endoderm cells are used. In some embodiments, the definitive endoderm cells are isolated or substantially purified. In some embodiments, the isolated or substantially purified definitive endoderm cells express one or more (e.g. at least 1, 3) of SOX17, FOXA2, or CXRC4 markers to a greater extent than one or more (e.g. at least 1, 3, 5) of GCT4, AFP, I'M, SPARC, or SGX7 markers.[000213] In some embodiments, pluripotent stem cells are prepared from somatic cells. In some embodiments, pluripotent stem cells are prepared from biological tissue obtained from a biopsy. In some embodiments, the pluripotent stem cells are cryopreserved. In some embodiments, the somatic cells are cryopreserved. In some embodiments, pluripotent stem cells are prepared from PBMCs. In some embodiments, human PSCs are prepared from human PBMCs. In some embodiments, pluripotent stem cells are prepared from cryopreserved PBMCs. In some embodiments, PBMCs are grown on a feeder cell substrate. In some embodiments, PBMCs are grown on a mouse embryonic fibroblast (MEF) feeder cell substrate. In some embodiments, PBMCs are grown on an irradiated MEF feeder cell substrate.[000214] In some embodiments, stem cells are treated with one or more growth factors to differentiate to definitive endoderm cells. Such growth factors can include growth factors from the TGF-beta superfamily. In some embodiments, the one or more growth factors comprise the Nodal / Activin and / or the BMP subgroups of the TGF-beta superfamily of growth factors. In some embodiments, the one or more growth factors are selected from the groupconsisting of Nodal, Activin A, Activin B, BMP4, Wnt3a or combinations of any of these growth factors. In some embodiments, the stem cells are contacted with Activin A. In some embodiments, the stem cells are contacted with Activin A and BMP4.[000215] It will be understood by one of skill in the art that altering the concentration, expression or function of one or more Wnt signaling proteins in combination with altering the concentration, expression, or function of one or more FGF proteins can give rise to directed differentiation in accordance with the present disclosure. In some embodiments, cellular constituents associated with the FGF, Wnt, EGF, or retinoic acid (RA) signaling pathways, or with the FGF, Wnt, EGF, BMP, or retinoic acid (RA) signaling pathways, for example, natural inhibitors, antagonists, activators, or agonists of the pathways can be used to result in inhibition or activation of the FGF, Wnt, or retinoic acid signaling pathways, or of the FGF, Wnt, EGF, BMP, or retinoic acid signaling pathways. In some embodiments, siRNA and / or shRNA targeting cellular constituents associated with the FGF, Wnt, or retinoic acid signaling pathways, or the FGF, Wnt, EGF, BMP, or retinoic acid signaling pathways, are used to inhibit or activate these pathways.[000216] In some embodiments, pluripotent stem cells, definitive endoderm, spheroids, or downstream cell types are contacted with a Wnt signaling pathway activator or Wnt signaling pathway inhibitor. In some embodiments, the Wnt signaling pathway activator comprises a Wnt protein, in some embodiments, the Wnt protein comprises a recombinant Wnt protein. In some embodiments, the Wnt signaling pathway activator comprises Wntl, Wnt2, Wnt2b, Wnt3, Wnt3a, Wnt4, WntSa, WntSb, Wnt6, Wnt7a, Wnt7b, Wnt8a, WntSb, Wnt9a, Wnt9b, WntlOa, WntlOb, Wntl 1 Wntl 6, BML 284, IQ-1, WAY 262611, or any combination thereof. In some embodiments, the Wnt signaling pathway activator comprises a GSK3 signaling pathway inhibitor. In some embodiments, the Wnt signaling pathway activator comprises CHIR99Q21, CfflR 98014, AZD2858, BIO, AR-A014418, SB 216763, SB 415286, aloisine, indirubin, alsterpaullone, kenpauilone, lithium chloride, TDZD 8, or TWS119, or any combination thereof. In some embodiments, the Wnt signaling pathway inhibitor comprises C59, PNU 74654, KY-02111, PRI-724, FH-535, DIF-1, or XAV939, or any combination thereof. In some embodiments, the cells are not treated with a Wnt signaling pathway activator or Wnt signaling pathway inhibitor. The Wnt signaling pathway activator or Wnt signalingpathway inhibitor provided herein may be used in combination with any of the other growth factors, signaling pathway activators, or signaling pathway inhibitors provided herein.[000217] In some embodiments, pluripotent stem cells, definitive endoderm, spheroids, or downstream cell types are contacted with an FGF signaling pathway activator. In some embodiments, the FGF signaling pathway activator comprises an FGF protein. In some embodiments, the FGF protein comprises a recombinant FGF protein. In some embodiments, the FGF signaling pathway activator comprises one or more of FGF 1 , FGF2, FGF3, FGF4, FGF4, FGF 5, FGF6, FGF7, FGF 8, FGF8, FGF9, FGF 10, FGF11, FGF 12, FGF 13, FGF 14, FGF 15 (FGF 19, FGF15 / FGF19), FGF 16, FGF 17, FGF 18, FGF20, FGF21, FGF22, or FGF23. In some embodiments, the cells are not treated with an FGF signaling pathway activator. The FGF signaling pathway activator provided herein may be used in combination with any of the other growth factors, signaling pathway activators, or signaling pathway inhibitors provided herein.[000218] In some embodiments, pluripotent stem cells, definitive endoderm, spheroids, or downstream cell types are contacted with a VEGF pathway activator. In some embodiments, the VEGF pathway activator or inhibitor comprises VEGF-A. In some embodiments, the cells are not treated with a VEGF pathway activator or inhibitor. VEGF pathway activation and / or inhibition as used with embodiments provided herein may be used in combination with any of the other growth factors, pathway activators, or pathway inhibitors provided herein. The VEGF pathway activator provided herein may be used in combination with any of the other growth factors, pathway activators, or pathway inhibitors provided herein.[000219] In some embodiments, pluripotent stem cells, definitive endoderm, spheroids, or downstream cell types are contacted with a BDNF pathway activator. In some embodiments, the BDNF pathway activator or inhibitor comprises BDNF. In some embodiments, the cells are not treated with a BDNF pathway activator or inhibitor. BDNF pathway activation and / or inhibition as used with embodiments provided herein may be used in combination with any of the other growth factors, pathway activators, or pathway inhibitors provided herein. The BDNF pathway activator provided herein may be used in combination with any of the other growth factors, pathway activators, or pathway inhibitors provided herein.[000220] In some embodiments, pluripotent stem cells, definitive endoderm, spheroids, or downstream cell types are contacted with a BMP pathway activator or inhibitor.In some embodiments, the BMP pathway activator comprises a BMP protein. In some embodiments, the BMP protein is a recombinant BMP protein. In some embodiments, the BMP pathway activator comprises BMP1, BMP2, BMP3, BMP4, BMP5, BMP6, BMP7, BMP8a, BMP8b, BMP10, BMP11, BMP15, IDE1, or IDE2, or any combination thereof. In some embodiments, the BMP pathway inhibitor comprises Noggin, Dorsomorphin, RepSox, LY364947, LDN193189, SB431542, or any combination thereof. In some embodiments, the cells are not treated with a BMP pathway activator or BMP pathway inhibitor. BMP pathway activation and / or inhibition as used with embodiments provided herein may be used in combination with any of the other growth factors, pathway activators, or pathway inhibitors provided herein. The BMP pathway activator or BMP pathway inhibitor provided herein may be used in combination with any of the other growth factors, pathway activators, or pathway inhibitors provided herein.[000221] In some embodiments, pluripotent stem cells, definitive endoderm, spheroids, or downstream cell types are contacted with an EGF signaling pathway activator. In some embodiments, the EGF pathway activator is EGF. In some embodiments, the cells are not treated with an EGF pathway activator or inhibitor. EGF pathway activation and / or inhibition as used with embodiments provided herein may be used in combination with any of the other growth factors, pathway activators, or pathway inhibitors provided herein. The EGF pathway activator provided herein may be used in combination with any of the other growth factors, pathway activators, or pathway inhibitors provided herein.[000222] In some embodiments, pluripotent stem cells, definitive endoderm, spheroids, or downstream cell types are contacted with a retinoic acid signaling pathway activator or retinoic acid signaling pathway inhibitor. In some embodiments, the retinoic acid signaling pathway activator comprises retinoic acid, all-trans retinoic acid, 9-cis retinoic acid, CD437, EC23, BS 493, TTNPB, or AM580, or any combination thereof, in some embodiments, the retinoic acid signaling pathway inhibitor comprises guggul sterone. In some embodiments, the cells are not treated with a retinoic acid signaling pathway activator or retinoic acid signaling pathway inhibitor. The retinoic acid signaling pathway activator or retinoic acid signaling pathway inhibitor provided herein may be used in combination with any of the other growth factors, signaling pathway activators, or signaling pathway inhibitors provided herein.[000223] In some embodiments, iPSCs are expanded in cell culture. In some embodiments, pluripotent stem cells are expanded in a basement membrane matrix. In some embodiments, iPSCs are expanded in Matrigel, In some embodiments, the iPSCs are expanded in cell culture comprising a ROCK inhibitor (e.g. Y-27632). In some embodiments, the iPSCs are differentiated into definitive endoderm cells. In the iPSCs are differentiated into definitive endoderm cells by- contacting the iPSCs with Activin A, BMP4, or both. In some embodiments, the iPSCs are contacted with a concentration of Activin A that is, is about, is at least, is at least about, is not more than, or is not more than about, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, or 200 ng / mL, or any concentration of Activin A within a range defined by any two of the aforementioned concentrations, for example, 10 to 200 ng / mL, 10 to 100 ng / mL, 100 to 200 ng / mL, or 50 to 150 ng / mL. In some embodiments, the pluripotent stem cells are contacted with Activin A at a concentration of 100 ng / mL or about 100 ng / mL. In some embodiments, the iPSCs are contacted with a concentration of BMP4 that is, is about, is at least, is at least about, is not more than, or is not more than about, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, or 200 ng / mL, or any concentration of BMP4 within a range defined by any two of the aforementioned concentrations, for example, 1 to 200 ng / mL, 1 to 100 ng / mL, 25 to 200 ng / mL, 1 to 80 ng / mL, or 25 to 100 ng / mL, In some embodiments, the pluripotent stem cells are contacted with BMP4 at a concentration of 50 ng / mL or about 50 ng / mL.[000224] In some embodiments, the PSCs are differentiated into definitive endoderm cells. In some embodiments, the PSCs are differentiated into spheroids, such as angioblast spheroids. In some embodiments, the PSCs are differentiated into a vessel organoid.[000225] In some embodiments, any of the cells disclosed herein may be cryopreserved for later use. The cells can be cryopreserved according to methods generally known in the art, optionally including one or more cryoprotectants.[000226] Cryoprotectants are cell composition additives to improve efficiency and yield of low temperature cryopreservation by preventing formation of large ice crystals. Cryoprotectants include but are not limited to DMSO, ethylene glycol, glycerol, propylene glycol, trehalose, formamide, methyl-formamide, dimethyl-formamide, glycerol 3 -phosphate, proline, sorbitol, diethyl glycol, sucrose, triethylene glycol, polyvinyl alcohol, polyethyleneglycol, or hydroxy ethyl starch. Cryoprotectants can be used as part of a cry opreservation medium, which include other components such as nutrients (e g. albumin, serum, bovine serum, fetal calf serum (FCS)) to enhance post-thawing survivability of the cells, in these cry opreservation media, at least one cryoprotectant may be found at a concentration that is, is about, is at least, is at least about, is not more than, or is not more than about, 0.01%, 0.05%, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90%, or any percentage within a range defined by any two of the aforementioned numbers.Gene Editing[000227] Embodiments of the disclosure can include PSCs, iPSCs, definitive endoderm cells, spheroids, or organoids which have been or which can be genetically modified or edited according to methods known in the art. For example, gene editing using CRISPR nucleases such as Cas9 are explored in PCT Publications WO 2013 / 176772, WO 2014 / 093595, WO 2014 / 093622, WO 2014 / 093655, WO 2014 / 093712, WO 2014 / 093661, WO 2014 / 204728, WO 2014 / 204729, WO 2015 / 071474, WO 2016 / 115326, WO 2016 / 141224, WO 2017 / 023803, and WO 2017 / 070633, each of which is hereby expressly incorporated by reference in its entirety.Pharmaceutical Compositions[000228] Embodiments of the disclosure can include pharmaceutical compositions. Such pharmaceutical compositions can include one or more additional pharmaceutically acceptable components, which can include carriers, excipients, and / or stabilizers that are nontoxic to the cell or mammal being exposed thereto at the dosages and concentrations employed or that have an acceptable level of toxicity. A “pharmaceutically acceptable” “diluent,” “excipient,” and / or “carrier” as used herein have their plain and ordinary meaning as understood in light of the specification and are intended to include any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like, compatible with administration to humans, cats, dogs, or other vertebrate hosts. Typically, a pharmaceutically acceptable diluent, excipient, and / or carrier is a diluent, excipient, and / or earner approved by a regulatory agency of a Federal, a state government, or other regulatory agency, or listed in the LJ.S. Pharmacopeia or other generally recognized pharmacopeia for use in animals, including humans as well as non-humanmammals, such as cats and dogs. The term diluent, excipient, and / or “carrier” can refer to a diluent, adjuvant, excipient, or vehicle with which the pharmaceutical composition is administered. Such pharmaceutical diluent, excipient, and / or earners can be sterile liquids, such as water and oils, including those of petroleum, animal, vegetable or synthetic origin. Water, saline solutions and aqueous dextrose and glycerol solutions can be employed as liquid diluents, excipients, and / or carriers, particularly for injectable solutions. Suitable pharmaceutical diluents and / or excipients include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, dried skim milk, glycerol, propylene, glycol, water, ethanol and the like. A non-limiting example of a physiologically acceptable carrier is an aqueous pH buffered solution. The physiologically acceptable carrier may also comprise one or more of the following: antioxidants, such as ascorbic acid, low molecular weight (less than about 10 residues) polypeptides, proteins, such as serum albumin, gelatin, immunoglobulins, hydrophilic polymers such as polyvinylpyrrolidone, ammo acids, carbohydrates such as glucose, mannose, or dextrins, chelating agents such as EDTA, sugar alcohols such as mannitol or sorbitol, saltforming counterions such as sodium, and nonionic surfactants such as TWEEN®, polyethylene glycol (PEG), and PLURONICS®. The composition, if desired, can also contain minor amounts of wetting, bulking, emulsifying agents, or pH buffering agents. These compositions can take the form of solutions, suspensions, emulsion, sustained release formulations and the like. The formulation should suit the mode of administration.[000229] Additional excipients with desirable properties include but are not limited to preservatives, adjuvants, stabilizers, solvents, buffers, diluents, solubilizing agents, detergents, surfactants, chelating agents, antioxidants, alcohols, ketones, aldehydes, ethylenediaminetetraacetic acid (EDTA), citric acid, salts, sodium chloride, sodium bicarbonate, sodium phosphate, sodium borate, sodium citrate, potassium chloride, potassium phosphate, magnesium sulfate sugars, dextrose, fructose, mannose, lactose, galactose, sucrose, sorbitol, cellulose, serum, amino acids, polysorbate 20, polysorbate 80, sodium deoxycholate, sodium taurodeoxy cholate, magnesium stearate, octylphenol ethoxylate, benzethonium chloride, thimerosal, gelatin, esters, ethers, 2-phenoxyethanol, urea, or vitamins, or any combination thereof. Some excipients may be in residual amounts or contaminants from the process of manufacturing, including but not limited to serum, albumin, ovalbumin, antibiotics,inactivating agents, formaldehyde, glutaraldehyde, b-propiolactone, gelatin, cell debris, nucleic acids, peptides, ammo acids, or growth medium components or any combination thereof. The amount of the excipient may be found in composition at a percentage that is, is about, is at least, is at least about, is not more than, or is not more than about, 0%, 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 100% w / w or any percentage by weight in a range defined by any two of the aforementioned numbers.[000230] Pharmaceutical compositions can include one or more “pharmaceutically acceptable salts”, which can include relatively non-toxic, inorganic and organic acid, or base addition salts of compositions or excipients, including without limitation, analgesic agents, therapeutic agents, other materials, and the like. Examples of pharmaceutically acceptable salts include those derived from mineral acids, such as hydrochloric acid and sulfuric acid, and those derived from organic acids, such as ethanesulfonic acid, benzenesulfonic acid, p- toluenesulfonic acid, and the like. Examples of suitable inorganic bases for the formation of salts include the hydroxides, carbonates, and bicarbonates of ammonia, sodium, lithium, potassium, calcium, magnesium, aluminum, zinc, and the like. Salts may also be formed with suitable organic bases, including those that are non-toxic and strong enough to form such salts. For example, the class of such organic bases may include but are not limited to mono-, di-, and trialkylamines, including methylamine, dimethylamine, and triethylamine; mono-, di-, or trihydroxyalkylamines including mono-, di-, and triethanolamine; ammo acids, including glycine, arginine and lysine; guanidine; N-methylglucosamine; N-methylglucamine; L- glutamine; N-methylpiperazine; morpholine; ethylenediamine; N-benzylphenethylamine; trihydroxymethyl ammoethane.[000231] Proper formulation is dependent upon the route of administration chosen. Techniques for formulation and administration of the compounds described herein are known to those skilled in the art. Multiple techniques of administering a compound exist in the art including, but not limited to, enteral, oral, rectal, topical, sublingual, buccal, intraaural, epidural, epicutaneous, aerosol, parenteral delivery, including intramuscular, subcutaneous, intra-arterial, intravenous, intraportal, intra-articular, intradermal, peritoneal, intramedullary injections, intrathecal, direct intraventricular, intraperitoneal, intranasal or intraocularinjections. Pharmaceutical compositions will generally be tailored to the specific intended route of administration.[000232] As used herein, a “carrier” has its plain and ordinary meaning as understood in light of the specification and can refer to a compound, particle, solid, semi-solid, liquid, or diluent that facilitates the passage, delivery and / or incorporation of a compound to cells, tissues and / or bodily organs.[000233] As used herein, a “diluent” has its plain and ordinary meaning as understood in light of the specification and can refer to an ingredient in a pharmaceutical composition that lacks pharmacological activity but may be pharmaceutically necessary or desirable. For example, a diluent may be used to increase the bulk of a potent drug whose mass is too small for manufacture and / or administration. It may also be a liquid for the dissolution of a drug to be administered by injection, ingestion or inhalation. A common form of diluent in the art is a buffered aqueous solution such as, without limitation, phosphate buffered saline that mimics the composition of human blood.Dosage and Administration Routes[000234] Embodiments of the disclosure can include methods of administering or treating an animal, which can involve administering an amount of at least one treatment, that is effective to treat the disease, condition, or disorder that the organism has, or is suspected of having, or is susceptible to, or to bring about a desired physiological effect. In some embodiments, the disease, condition, or disorder can involve a hyperoxic injury-related disease or disorder. In some embodiments, the disease, condition, or disorder can involve bronchopulmonary dysplasia (BPD).[000235] In some embodiments, at least one treatment can include a composition or pharmaceutical composition, which can be administered to an animal (e.g., mammals, primates, monkeys, or humans) in an amount of about 0.005 to about 50 mg / kg body weight, about 0.01 to about 15 mg / kg body weight, about 0.1 to about 10 mg / kg body weight, about 0.5 to about 7 mg / kg body weight, about 0.005 mg / kg, about 0.01 mg / kg, about 0.05 mg / kg, about 0.1 mg / kg, about 0.5 mg / kg, about 1 mg / kg, about 3 mg / kg, about 5 mg / kg, about 5.5 mg / kg, about 6 mg / kg, about 6.5 mg / kg, about 7 mg / kg, about 7.5 mg / kg, about 8 mg / kg, about 10 mg / kg, about 12 mg / kg, or about 15 mg / kg. In regard to some conditions, the dosage can be about 0.5 mg / kg human body weight or about 6.5 mg / kg human body weight. In someinstances, some subjects (e ., mammals, mice, rabbits, feline, porcine, or canine) can be administered a dosage of about 0.005 to about 50 mg / kg body weight, about 0.01 to about 15 mg / kg body weight, about 0.1 to about 10 mg / kg body weight, about 0.5 to about 7 mg / kg body weight, about 0.005 mg / kg, about 0.01 mg / kg, about 0.05 mg / kg, about 0.1 mg / kg, about 1 mg / kg, about 5 mg / kg, about 10 mg / kg, about 20 mg / kg, about 30 mg / kg, about 40 mg / kg, about 50 mg / kg, about 80 mg / kg, about 100 mg / kg, or about 150 mg / kg. Of course, those skilled in the art will appreciate that it is possible to employ many concentrations in the methods of the present disclosure, and using, in part, the guidance provided herein, will be able to adjust and test any number of concentrations in order to find one that achieves the desired result in a given circumstance. In some embodiments, a dose or a therapeutically effective dose of a compound disclosed herein will be that which is sufficient to achieve a plasma concentration of the compound or its active metabolite(s) within a range set forth herein, e.g., about 1-10 nM, 10-100 nM, 0.1-1 pM, 1-10 pM, 10-100 pM, 100-200 pM, 200-500 pM, or even 500-1000 pM, preferably about 1-10 nM, 10-100 nM, or 0.1-1 pM.[000236] In other embodiments, a treatment can be administered in combination with one or more other therapeutic agents for a given disease, condition, or disorder.[000237] The compounds and pharmaceutical compositions are preferably prepared and administered in dose units. Solid dose units are tablets, capsules and suppositories. For treatment of a subject, depending on activity of the compound, manner of administration, nature and severity of the disease or disorder, age and body weight of the subject, different daily doses can be used.[000238] Under certain circumstances, however, higher or lower daily doses can be appropriate. The administration of the daily dose can be carried out both by single administration in the form of an individual dose unit or else several smaller dose units and also by multiple administrations of subdivided doses at specific intervals.[000239] A treatment can be administered locally or systemically in a therapeutically effective dose. Amounts effective for this use will, of course, depend on the severity of the disease or disorder and the weight and general state of the subject. Typically, dosages used in vitro can provide useful guidance in the amounts useful for in situ administration of the pharmaceutical composition, and animal models can be used to determine effective dosages for treatment of particular disorders.[000240] Various considerations are described, e. g. , in Langer, 1990, Science, 249: 1527; Goodman and Gilman's (eds.), 1990, Id., each of which is herein incorporated by reference and for all purposes. Dosages for parenteral administration of active pharmaceutical agents can be converted into corresponding dosages for oral administration by multiplying parenteral dosages by appropriate conversion factors. As to general applications, the parenteral dosage in mg / mL times 1.8 = the corresponding oral dosage in milligrams (“mg”). As to oncology applications, the parenteral dosage in mg / mL times 1.6 = the corresponding oral dosage in mg. An average adult weighs about 70 kg. See e.g., Miller-Keane, 1992, Encyclopedia & Dictionary of Medicine, Nursing & Allied Health, 5th Ed., (W. B. Saunders Co.), pp.1708 and 1651.[000241 ] It will be understood, however, that the specific dose level for any particular patient will depend upon a variety of factors including the activity of the specific compound employed, the age, body weight, general health, sex, diet, time of administration, route of administration, rate of excretion, drug combination and the severity of the particular disease undergoing therapy.[000242] In some embodiments, the administration can include a unit dose of one or more treatments in combination with a pharmaceutically acceptable carrier and, in addition, can include other medicinal agents, pharmaceutical agents, carriers, adjuvants, diluents, and excipients. In certain embodiments, the carrier, vehicle or excipient can facilitate administration, delivery and / or improve preservation of the composition. In other embodiments, the one or more carriers, include but are not limited to, saline solutions such as normal saline, Ringer's solution, PBS (phosphate-buffered saline), and generally mixtures of various salts including potassium and phosphate salts with or without sugar additives such as glucose. Carriers can include aqueous and non-aqueous sterile injection solutions that can contain antioxidants, buffers, bacteriostats, bactericidal antibiotics, and solutes that render the formulation isotonic with the bodily fluids of the intended recipient; and aqueous and nonaqueous sterile suspensions, which can include suspending agents and thickening agents. In other embodiments, the one or more excipients can include, but are not limited to water, saline, dextrose, glycerol, ethanol, or the like, and combinations thereof. Nontoxic auxiliary substances, such as wetting agents, buffers, or emulsifiers may also be added to the composition. Oral formulations can include such normally employed excipients as, forexample, pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharine, cellulose, and magnesium carbonate. The quantity of active component in a unit dose preparation can be varied or adjusted from 0. 1 mg to 10000 mg, more typically 1.0 mg to 1000 mg, most typically 10 mg to 500 mg, according to the particular application and the potency of the active component. The composition can, if desired, also contain other compatible therapeutic agents.[000243] A treatment can be administered to subjects by any number of suitable administration routes or formulations. The treatment, such as an immunotherapy, can also be used to treat subjects for a variety of diseases. Subjects include but are not limited to mammals, primates, monkeys (e.g., macaque, rhesus macaque, or pig tail macaque), humans, canine, feline, bovine, porcine, avian (e.g., chicken), mice, rabbits, and rats. In particular embodiments described herein, the subject is a human.[000244] The route of administration of the compounds of the treatments described herein can be of any suitable route. Administration routes can be, but are not limited to the oral route, the parenteral route, the cutaneous route, the nasal route, the rectal route, the vaginal route, and the ocular route. In other embodiments, administration routes can be parenteral administration, a mucosal administration, intravenous administration, subcutaneous administration, topical administration, intradermal administration, oral administration, sublingual administration, intranasal administration, or intramuscular administration. The choice of administration route can depend on the compound identity (e.g., the physical and chemical properties of the compound) as well as the age and weight of the animal, the particular disease (e.g., type of cancer), and the severity of the disease (e.g., stage or severity of cancer). Of course, combinations of administration routes can be administered, as desired.[000245] Some embodiments of the disclosure include a method for providing a subject with a treatment which comprises one or more administrations of one or more compositions; the compositions may be the same or different if there is more than one administration.Toxicity[000246] The ratio between toxicity and therapeutic effect for a particular treatment is its therapeutic index and can be expressed as the ratio between LD50 (the amount of compound lethal in 50% of the population) and ED50 (the amount of compound effective in50% of the population). Compounds that exhibit high therapeutic indices are preferred. Therapeutic index data obtained from in vitro assays, cell culture assays and / or animal studies can be used in formulating a range of dosages for use in humans. The dosage of such compounds preferably lies within a range of plasma concentrations that include the ED50 with little or no toxicity. The dosage can vary within this range depending upon the dosage form employed and the route of administration utilized. See, e.g. Fingl et al., In: THE PHARMACOLOGICAL BASIS OF THERAPEUTICS, Ch.l, p.l, 1975. The exact formulation, route of administration, and dosage can be chosen by the individual practitioner in view of the patient’s condition and the particular method in which the compound is used. For in vitro formulations, the exact formulation and dosage can be chosen by the individual practitioner in view of the patient’ s condition and the particular method in which the compound is used.[000247] Having described various aspects of the disclosure in detail, it will be apparent that modifications, variations, and equivalent embodiments are possible without departing from the scope of the disclosure defined in the appended claims. Furthermore, it should be appreciated that all examples in the present disclosure are provided as non-limiting examples.EXAMPLES[000248] The following non-limiting examples are provided to further illustrate embodiments of the disclosure herein. It should be appreciated by those of skill in the art that the techniques disclosed in the examples that follow represent approaches that have been found to function well in the practice of the disclosure, and thus can be considered to constitute examples of modes for its practice. However, those of skill in the art should, in light of the present disclosure, appreciate that many changes can be made in the specific embodiments that are disclosed and still obtain a like or similar result without departing from the spirit and scope of the disclosure.Example 1NTRK2 splice variants change dynamically in human and mouse lungs exposed to hyperoxic injury[000249] NTRK2 splice variants were found to change dynamically in human and mouse lungs exposed to hyperoxic injury. While Ntrk2 was one of the most downregulated genes in lungs from mice exposed to seven days of hyperoxia, the isoforms of NTRK2 were not yet identified.[000250] To further understand the roles of NTRK2 isoforms in neonatal lung injury, the existing infrastructure of the NIH funded LungMAP2 consortium and the Biorepository for Investigations of Diseases of the Lung (BRINDL) at the University of Rochester were leveraged to collect OCT embedded lung tissue and dissociated cells from 16 BPD patients and 18 age-matched controls. Out of the 16 BPD patients, 10 had pathological changes in pulmonary arteries, indicating severe damage of the vasculature. BPD patients were stratified into four categories: “acute developing (1 day to 3.8 months, n=4)”, “active evolving (8-13 months, n=4)”, “established (15-21 months, n=4)”, and “Healed (3-5 years, n=4)”. ‘Healed’ cases were defined based on their previous BPD history (under ClinPathDx), age of death (>20 months), and the cause of death was not directly associated with BPD or its consequences. In situ hybridization (ISH) was performed with RNAScope probes recognizing either shared or tyrosine kinase (TK) region of NTRK2 to identify the isoforms in control vs. BPD samples.[000251] A significant reduction of NTRK2-FL was observed in “acute developing” BPD cases compared with either preterm control or age-matched fullterm control. At the “active evolving” and “established” stages of BPD, the expression of NTRK2-T1 was increased. Expression of both isoforms was low in resolving “healed” BPD (Fig. 3A), consistent with our previous finding that NTRK2 expression decreased in late alveolar stage of lung development.[000252] In neonatal mice exposed to 7 days of 90% oxygen, the expression of Ntrk2- FL was decreased during the acute injury phase on postnatal day 7 (P7). Interestingly, the responses to hyperoxic injury varied among different mice (e.g., as shown via the H&E staining in Fig. 3B) during P10 to P28. Mice with improved or partially recovered alveolar structure after injury, as determined by the mean linear intercept, exhibited a significant increase of Ntrk2-FL expression. On the other hand, in alveoli with impaired regeneration (“Impaired Recovery”), the expression of the alternative isoform, Ntrk2-Tl, was elevated (e.g., as shown in Fig. 3B) These data demonstrate that NTRK2-FL plays a protective role in alveolarregeneration and repair following hyperoxic injury, whereas NTRK2-T1 leads to impaired alveologenesis in both human and mouse.Example 2Increased NTRK2-T1 in gCap-C2 cells from BPD human lung samples[000253] To elucidate the gene regulatory network related to NTRK2 in gCap cells during alveolar regeneration and repair, single nucleus (sn) multiomic analysis was performed on enriched EC populations from three BPD patients (active evolving and established stages) and two age-matched controls (e.g., as shown in Fig. 4A). The integrated snRNA-seq and snATAC-seq analyses identified 7 EC subtypes as well as immune and mesenchymal cell populations (e.g., as shown in Fig. 4B). Notably, one of the gCap subpopulation, gCap-C2, was significantly increased in BPD (e.g., as shown in Fig. 4C). Because gCap cells are known to be progenitor cells in alveolus, proliferation genes were examined in gCap-Cl and gCap- C2. The expression oiMK167 (encodes Ki67) was significantly lower in gCap-C2 compared with gCap-Cl (e.g., as shown in Fig. 4D), perhaps contributing to the arrested vascular growth in BPD. Next, differentially expressed genes (DEGs) were systematically analyzed in each EC subcluster comparing control vs. BPD. One of the most enriched genes in gCap-C2 was NTRK2 (e.g., as shown in Fig. 4E). Consistent with the findings shown in Fig. 3, it was found that the transcripts mapping to the shared domain (lacking tyrosine kinase domains) were significantly increased in BPD gCap-C2 (Fig. 4F), indicating an increased expression of NTRK2-T1 in BPD. These data demonstrate that increased NTRK2-T1 may contribute to the reduced gCap-C2 proliferation in alveoli undergoing regeneration following hyperoxic injury.Example 3Distinct downstream signaling and functional roles of NTRK2-FL and NTRK2-T1 in human pulmonary microvascular ECs (hPMVECs)[000254] To elucidate the specific contributions of NTRK2 isoforms in human peripheral lung endothelial cells (ECs), human pulmonary microvascular endothelial cells (hPMVECs) were transfected with NTRK2-FL or NTRK2-T1 mRNAs using the jetMESSENGER nanoparticle delivery system. Upon stimulation with BDNF, both the AKT and ERK pathways were activated in the GFP control group (expressing endogenous NTRK2-FF) as well as in the NTRK2-FL overexpression group. In contrast, no activation of these pathways was observed in cells overexpressing NTRK2-T1 (Fig. 5A). These findings suggest an inhibitory effect of NTRK2-T1 on endogenous NTRK2-FL. Furthermore, the overexpression of NTRK2-FL increased EC proliferation (Fig. 5B) and enhanced angiogenesis (Fig. 5C) in the presence of BDNF. In contrast, overexpression ofNTRK2-Tl suppressed these cellular functions. These in vitro studies show the regenerative potential of NTRK2-FL and highlight the detrimental role of NTRK2-T1 in hPMVECs.Example 4Decreased cell proliferation, impaired tube formation, and increased NTRK2-T1 in human iPSC-derived VOs after hyperoxia exposure[000255] To further understand the role of NTRK2 isoforms during the process of capillary formation and in response to hyperoxic injury in 3D, vessel organoids (VOs) were generated from human induced pluripotent stem cells (iPSCs). After 5 days of differentiation, iPSCs-derived angioblasts were either subjected to 75% oxygen exposure from day 5 to day 10 to induce injury during capillary formation, or cultured under normal room air (21% oxygen) as a control. After day 10, VOs were maintained under room air for an additional 5 days to establish the phenotype (Fig 6A). It is contemplated that, in some embodiments, hyperoxic conditions may range from 35% O2 to about 80% O2,, 50% O2 to about 80% O2, BDNF was added from day 5 to day 15 to activate NTRK2 signaling. In day 5 angioblasts, high levels of PECAM1, Ki67, and NTRK2-FL staining were observed, indicating active EC proliferation and capillary formation (Fig. 6B). After 5 days of hyperoxia exposure, capillary formation was significantly disrupted, as indicated by reduced PECAM1 staining and decreased cell proliferation judged by Ki67 staining in both day 10 and day 15 VOs (Fig. 6B&C) Consistent with the findings of the present disclosure, decreased NTRK2-FL expression was observed during the acute phase of hyperoxia injury (day 10), and increased NTRK2-T1 expression was observed at a later stage (day 15) (Fig. 6B&C). However, no recovery events, such as improved cellular function or increased NTRK2-FL expression, were observed in VOs exposed to five days of 75% oxygen.[000256] The role of NTRK2 isoforms in capillary regeneration and repair is further described herein. Various embodiments of the present disclosure thus demonstrate the potentialof iPSC-derived VOs as a valuable model for studying NTRK2 signaling in response to hyperoxia, in which can be used to develop patient-specific platforms to identify optimal drugs for improving alveolar regeneration following hyperoxic injury.Example 5Overexpression of NTRK2-FL in VOs improved tube formation and cell proliferation after hyperoxia exposure[000257] In order to recapitulate the dynamic changes observed in human and mouse lungs during alveolar regeneration after hyperoxia injury, a lipid nanoparticle mRNA delivery system was employed to overexpress NTRK2-FL and NTRK2-T1 in VOs after five days of hyperoxia exposure (Fig. 7A). Intriguingly, the overexpression of NTRK2-FL significantly improved tube formation, as demonstrated by increased staining of the capillary ECs (PECAM1+), and increased cell proliferation judged by Ki67 staining (Fig. 7B & 7C). The overexpression of NTRK2-T1 did not exacerbate EC injury under hyperoxia condition (Fig. 7B&C) These results further support the finding that activation of the NTRK2 pathway can promote repair and regeneration of alveolar capillaries in response to hyperoxia-induced injury.Example 6Comparing the effects of mild versus severe hyperoxic injuries on NTRK2 isoforms and EC function[000258] On studying the effect of mild versus severe hyperoxic injurites on NTRK2 isoforms, dynamic differences in mice were found. In particular, NTRK2-FL was found to be increased in mild conditions, whereas NTRK2-T1 was increased in severe conditions.[000259] In order to investigate whether NTRK2 plays a role in BPD cases that recover from injury, for which human tissues are unavailable, a hyperoxia-induced lung injury mouse model was used. This model may be utilized to induce alveolar simplification, a hallmark of human BPD pathology. Neonatal mice were exposed to 7 days of 75% 02 to create a mild injury model which recovered from hyperoxic injury, or to 90% 02 to recapitulate the severe and non-recovery BPD pathology (e.g., as shown in Figs. 8A & 8B). In some embodiments, to induce hyperoxic injury, a mice or other subject (e.g., vessel organoid model)may be subjected to hyperoxic conditions for about 6 to 14 days, preferably about 6 to 8 days. It was found that NTRK2-FL increased in response to mild injury, whereas Ntrk2-Tl increased under severe hyperoxic injury (e.g., as shown in Figs. 8A & 8B). To further determine gCap- specific changes, gCap cells (CD31+, KIT+) were isolated from mouse lungs exposed to 75% 02 or 90% 02 and western blot analysis was performed (e.g., as shown in Fig. 8C). Compared with the mild injury group, severe injury resulted in a decrease in NTRK2-FL and an increase in NTRK2-T1, potentially contributing to the non-resolving alveolar pathology. These data suggest that NTRK2-FL is associated with alveolar regeneration, whereas an elevated NTRK2- T1 to NTRK2-FL ratio may contribute to persistent alveolar simplification in response to hyperoxic injury.[000260] It was then found that human iPSC-derived vessel organoids (VOs) can be used to recapitulate endothelial defects and NTRK2 changes observed in human and mouse models. In particular, the role of NTRK2 isoforms in mild versus severe hyperoxic injuries in humans was studied by generating VOs from human iPSCs based on techniques described herein.[000261] Angioblasts derived from iPSCs were cultured under three conditions: 60% 02 for three days to induce “mild injury”, 75% 02 for five days to induce “severe injury”, or in room air for control (e.g., as shown in Fig. 9A). After injury, VOs were maintained in room air for another 5-7 days for recovery. Immunostaining of day 15 VOs revealed significant reduction of capillary formation and cell proliferation (Ki67) in the severe injury group compared with room air and mild injury groups (e.g., as shown in Fig. 9B). Consistent with findings in mice, decreased NTRK2-FL and increased NTRK2-T1 were observed in the severe injury group compared with mild injury (Figs. 9B & 9C).[000262] These results demonstrate the utility of human iPSC-derived VOs as an in vitro model for investigating NTRK2 signaling in response to varying degrees of hyperoxic injuries. These findings have applications in developing patient-specific platforms to identify optimal drugs for improving capillary regeneration following hyperoxic injury.Example 7Differences in expression of NTRK2 isoforms during lung development[000263] As shown in Fig. 10, NTRK2-FL is typically highly expressed in gCap cells, reaching its peak during the early alveolar stage of lung development in both human and mouse. The expression of NTRK2-FL significantly decreased during acute injury phase following exposure to hyperoxia. During the recovery stage, NTRK2-FL enhanced capillary regeneration by promoting angiogenesis and cell proliferation. In contrast, NTRK2-T1 impeded the repair process, leading to persistent alveolar simplification, which resembles the pathological features observed in severe BPD patients.Example 8Downstream signaling and functional roles of NTRK2 isoforms in human lung[000264] In mice with influenza-induced acute lung injury, a subpopulation of highly proliferative ECs contributed to the alveolar revascularization after injury, indicating the importance of EC proliferation in alveolar regeneration. As described herein, NTRK2-FL was found to activate AKT and ERK pathways, promoting cell proliferation and angiogenesis in hPMVECs. Conversely, NTRK2-T1 inhibited these pathways, resulting in abnormal EC functions. In the brain, beyond an inhibitory role on NTRK2-FL-mediated canonical signaling, NTRK2-T1 activated calcium signaling, which may contribute to increased capillary permeability.[000265] The consequences of abnormal capillary ECs with disrupted NTRK2 signaling on neighboring cells during alveolar regeneration and repair was determined. The increased NTRK2-T1 was found to inhibit the protective signaling mediated by NTKR2-FL, resulting in gCap dysfunction and disrupted cell-cell crosstalk, ultimately impairing alveolar regeneration following hyperoxia-induced injury.Example 9Determining the activation status of NTRK2 signaling in human and mouse lungs exposed to hyperoxia[000266] pAKT and pERK signaling, EC proliferation, and tube formation in hPMVECs transfected with NTRK2-FL were then assessed and compared to hPMVECstransfected with NTRK2-T1 . To better understand the functional roles of the NTRK2 isoforms in gCap cells and their response to hyperoxia, gCap cells from commercially available hPMVECs can be enriched via magnetic beads sorting with antibody recognizing the gCap cell surface marker KIT / CD177. Similar to the experimental design described in Fig. 7A, gCap cells can be exposed to hyperoxia for five days, and transfected with either NTRK2-T1 or NTRK2-FL mRNAs using the jetMESSENGER nanoparticle delivery system.[000267] Upon BDNF stimulation, the phosphorylation of pAKT, pERK, and pNTRK2 during the recovery phase following hyperoxia injury can be assessed by western blot. The activation of calcium signaling can be determined by time-lapse live imaging of the calcium spikes with the calcium indicator, Fura-2. These cells can then be imaged using a confocal widefield system equipped with a microfluidic pump and a temperature- and gas- controlled chamber. In addition to the in vitro studies described herein, the experiment can further tested the aforementioned signaling pathways in mouse lungs following hyperoxia- induced injury, as well as human lung tissue from control and BPD cases representing different stages of BPD progression by western blots.Example 10Effects of NTRK2 isoforms on EC functions including cell survived and calcium signaling- mediated EC barrier function in hPMVECs and vessel organoids[000268] Cell proliferation and tube formation can be determined using methods described in Fig. 5. Capillary permeability can be measured in two ways - endothelial layer permeability assay and transendothelial electrical resistance (TEER) measurement in vitro . Secondly, an XPerT permeability assay can be conducted using gCap cells grown on a biotin- labeled collagen substrate. FITC-labeled avidin was added to allow visualization and quantification of intercellular gaps. Immunostaining was used to assess the expression of endothelial junctional proteins, such as ZO-1, VE-Cadherin and Claudin-5, to visualize the barrier integrity of the gCap cells.Example 11Determining the hyperoxia conditions for modeling capillary regeneration following hyperoxic injury in iPSC-VOs[000269] As previously discussed five days of 75% oxygen resulted in severe capillary damage, and the iPSC-VOs were not able to recover from the hyperoxic injury on day 15. Therefore, there were no observations of an increased NTRK2-FL expression in VOs following hyperoxia injury.[000270] To establish an in vitro VO model that recapitulates the spectrum of response to hyperoxia from “impaired recovery” to “recovery”, protocols were revised by reducing the percentage of oxygen, shortening the duration of hyperoxia exposure, or extending the recovery period under room air, in order to determine the appropriate hyperoxia conditions allowing regeneration of tubular structures within the VOs. The regeneration of capillaries may be assessed using whole-mount staining of endothelial marker CD31 and quantification of Ki67, a marker of cell proliferation. Alterations in endogenous NTRK2-FL and NTRK2-T1 expression within the VOs can be determined using RNAScope and western blot. The present disclosure also explores the role of NTRK2-T1 overexpression in VOs under room air conditions, for example, to determine if NTRK2-T1 alone is sufficient to induce capillary injury without the challenge of hyperoxia.Example 12Impact of aberrant NTRK2 signaling in gCap cells on surrounding cell types[000271] To identify the spatial location of the NTRK2+ gCap cells in human lung post hyperoxia injury, and to uncover their neighboring cell types and the ligand-receptor (L- R) pairs mediating the cell-cell interactions, a Special Transcriptomic (ST) analysis was performed on human peripheral lung tissue from three BPD patients (active evolving and established stages) and three age-matched controls using the lOx Visium platform. OCT / PBS inflated frozen human lung tissues were collected through the BRINDL network, and two regions of the peripheral lung representing alveoli and respiratory bronchioles were selected for ST analyses. Cell21ocation was used for the cell type deconvolution based on two single cell reference maps: the Multi ome dataset and the LungMAP human lung CellRef (Fig. 11A). After deconvolution, the spatial distributions of different lung cell types were identified (Fig. 11B) This computational approach further improved the resolution of the current lOx Visium platform. Notably, the locations of the cell types correlated closely with the lung histology. Airway epithelial cells aligned with the respiratory bronchiole, while distal epithelial cells andcapillary ECs occupied the entire distal alveoli region; arterial ECs were spatially distributed along bronchial structures, while lymphatic and venous ECs were enriched in the interlobular septum. Pleural regions were enriched with airway fibroblasts. The correlation of cell types and their anatomical location demonstrated the robustness of the ST analysis suitable for downstream cell-cell communication analysis.[000272] To better understand the impact of the abnormal gCap-C2 cells with aberrant NTRK2 signaling on surrounding cell types, cell-cell interaction analysis was performed based on ST datasets. Notably, ST analyses of three controls and three BPD lung samples also revealed an increase o NTRK2 in BPD. The expression level of its ligand, BDNF, remained unchanged (Fig. 12), indicating that the abnormal downstream signaling is more likely due to the abnormal expression of the receptor rather than the ligand. Using both LungMAP CellRef and Multiome datasets as references, the ST data was deconvoluted, and the cell type composition within each spot was determined. Spots containing gCap cells were selected (Figs. 13A & 13B) and ligand-receptor (L-R) binding analysis was performed using the stLearn algorithm. The L-R pairs were either detected within the same spot, or from two neighboring spots (Fig. 13C). Several signaling pathways important for alveologenesis, such as PDGF, BMP, and Wnt, were lost in BPD, whereas increased CX3CL1 signaling was observed in gCap-C2 - ATI (Alveolar Type 1 cell) interaction. These L-R pairs in control and BPD human lung samples were then examined using RNAScope.[000273] To determine the direct impact of abnormal gCap cells on neighboring cells, an endothelial-epithelial co-culture system was established by either incorporating iPSC- derived endothelial cells (iPSCECs) into iPSC-derived lung organoids (Fig. 14A & 14B), or by assembling VOs and epithelial organoids (Fig. 14C). Using this system, NTRK2-T1 was overexpressed in purified gCap cells and co-cultured with the lung epithelium, to identify the changes in signaling between gCap-C2 and ATI. Human iPSC-derived lung-specific mesenchyme was also incorporated to study the interaction between gCap-C2 and alveolar fibroblasts. Alternatively, a newly developed multicellular alveolar organoid with vasculature was used, in which mesodermal and endodermal lineages were co-developed (Fig. 14D). This organoid model contains lung-specific ECs (FOXF1+, FENDRR+, CDH5+, Fig. 14D) and lung-specific mesenchymal cells (FOXF1+, TBX4+, Fig. 14D), providing a unique platform to understand cell-cell interaction under hyperoxia-induced injury.[000274] Inactivation of pAKT and pERK was observed in mouse lung samples with persistent alveoli simplification post hyperoxic injury, and in human lung tissues from BPD patients with respiratory failure. If NTRK2 signals through a completely novel pathway in gCap cells, an unbiased RNA-seq analysis can be performed in gCap cells overexpressing NTRK2 isoforms and downstream signaling pathways can be identified. Overexpression of NTRK2-T1 will impair endothelial barrier function and increase capillary permeability. Alternatively, in some embodiments, whether NTRK2 overexpression regulates gCap to aerocytes transdifferentiation can be tested. Since gCap cells may interact with other cell types in addition to those described in Fig. 13C, the previously described cell deconvolution method may be revised, and a ST platform with higher resolution may be used to reveal more precise L-R pairs such as Curio Seeker.[000275] The aforementioned experiment of the present disclosure thus elucidate the dynamic changes of NTRK2 isoforms in gCap cells in response to hyperoxia injury, and determine the roles of NTRK2-FL and NTRK2-T1 in regulating alveolar capillary regeneration and repair. The impact of abnormal gCap cells on surrounding cells was also identified.Example 13Upstream regulators ofNTRK2-Tl in gCap cells with impaired regenerative capacity[000276] Experiments of the present disclosure further demonstrate the critical roles of NTRK2 in alveolar capillary regeneration following hyperoxia injury. The present disclosure also determines previously unknown upstream regulators of NTRK2-FL and NTRK2-T1 in human lung gCap cells. In particular, the present disclosure describes a multiomic analysis that identified increased chromatin accessibility at H0XA5 and MEIS1 binding sites within the promoter region of the NTRK2-T1 in BPD human lung tissues. Sustained H0XA5 expression in ECs inhibited angiogenesis and reduced EC proliferation. MEISI, a HOX co-factor, is highly enriched in lung capillary ECs, and was detected in BPD patient lung tissues by gene array.[000277] Interestingly, the multimoic analysis described herein also identified upregulation of the RNA-binding FOX protein 2 (RBFOX2) in BPD gCap cells, which may contribute to increased alternative splicing of NTRK2. Based on the analysis, the presentdisclosure finds that the increase of NTRK2-T1 in abnormal gCap cells is controlled by the upregulation of H0XA5 and / or MEISI, and augmented RNA splicing mediated by RBFOX2.Example 14Roles of H0XA5 and MEIS1 in regulating NTRK2 expression in human and mouse lungs exposed to hyperoxia[000278] To determine the upstream regulator(s) that contribute to the increased NTRK2-T1 in response to hyperoxia injury in gCap cells with reduced regenerative capacity, an experiment is disclosed that integrated snRNA-seq and snATAC-seq datasets generated through Multiome and identified increased chromatin accessibility at two Transcription Factor (TF) binding sites, H0XA5 and MEIS 1, within the promotor region of the NTRK2 (Fig. 15A). snRNAseq data showed increased H0XA5 and MEIS I gene expression levels (log2FC>0.25) in BPD gCap- C2 cells (Fig. 15B). The experiment assessed changes in H0XA5 and MEIS1 in control and BPD human lung samples, and in human iPSC-VOs exposed to hyperoxia injury (same condition as described in Fig. 6A). Similar studies were performed in mouse lungs exposed to hyperoxia (same condition as Fig. 3B) using qPCR, RNAScope, and immunostaining. In particular, it was found that the expression of H0XA5 and MEIS1 are reduced during acute injury phase of hyperoxia, whereas expressions of H0XA5 and MEIS1 are increased during the recovery phase.[000279] To assess the binding of HOXA5 and MEIS1 to the promoter region of NTRK2 in response to hyperoxic injury, a chromatin immunoprecipitation (ChlP)-qPCR was performed with H0XA5 and MEIS1 antibodies in human and mouse lung samples and in VOs exposed hyperoxia. qPCR primers were designed based on the sequences in the shared region and tyrosine kinase region of the NTRK2.Example 15Testing the roles ofHOXAS and MEIS1 in regulating NTRK2 expression, signaling, and functions in response to hyperoxia injury[000280] The human-specific iPSC-VO model can be used to test the roles of H0XA5 and MEIS1 in regulating NTRK2 expression, signaling, and functions in response to hyperoxia injury. Utilizing the human-specific iPSC-VO system, H0XA5 and MEIS1 genescan be deleted in VOs using CRISPR, and then the VOs can be subjected to five days of 75% hyperoxia. After another five days of room air recovery, changes in NTRK2 isoforms and their downstream signaling pathways can be determined by qPCR, RNAScope, and western blot (NTRK2-FL vs. NTRK2-T1 can be distinguished by different protein sizes on the blot). The functions of ECs, including capillary formation and cell proliferation can be evaluated, as described in Figs 6 and 7.[000281] HOXA5 and MEIS1 were overexpressed in VOs cultured under room air condition. Thus, the experiment can then determine if these TFs induce the expression of NTRK2 isoforms. The same set of experiments can be performed in human gCap cells purified from hPMVECs.Example 16Role of RBF OX2-mediated RNA splicing of NTRK2 in lung ECs exposed to hyperoxia[000282] The role of RBFOX2-mediated RNA splicing of NTRK2 in lung ECs exposed to hyperoxia was then evaluated. In order to understand the molecular mechanism underlying the increased RNA splicing in gCap cells exposed to hyperoxia injury, differentially expressed genes were analyzed in control versus BPD gCap cells based on the human lung multiomic datasets.[000283] Interestingly, it was found that RBFOX2, a regulator of RNA alternative splicing, was significantly enriched in EC population and upregulated in BPD gCap-C2 cells compared with controls (Fig. 16A & 16B). This result was also validated in control and BPD human lung samples (Fig. 16C). RBFOX2 is the only RBFOX family member that was expressed in arterial ECs. The overexpression of Rbfox2 increased expression of the Ntrk2-Tl isoform in primary hippocampal neurons. In embryonic mouse hearts, RBFOX2 conditional knockout caused profound defects in yolk sac vasculogenesis. Next, the experiment 1) determined changes in RBFOX2 in gCap cells and VOs exposed to hyperoxia; 2) overexpressed RBFOX2 in gCap cells and VOs, and examined the change of NTRK2 isoforms by qPCR and western blot; 3) performed knockdown of RBFOX2 in gCap cells and VOs, subjected the cells to hyperoxia injury, and assessed the change of NTRK2 isoforms; and 4) studied the RBFOX2-NTRK2 interaction using Ribonucleoprotein Immunoprecipitation (RIP)-qPCR.[000284] Hyperoxia was found to upregulate H0XA5, MEIS1, and RBFOX2 levels. H0XA5 and MEIS1 were found to synergistically regulate NTRK2 gene expression in gCap cells exposed to hyperoxia injury, as both TFs have been implicated in regulating alveologenesis. H0XA5 can play a predominant role in regulating gCap proliferation and angiogenesis given its high expression level in quiescent ECs compared with active angiogenic ECs. The present disclosure contemplates that, if none of the predicted TFs and RNA binding protein (RBP) are directly involved in capillary regeneration in response to hyperoxia injury, unbiased RNA-seq can be performed in gCap cells and VOs exposed to hyperoxia injury, and data analysis can be focused on the identification of TFs and RBPs.[000285] The aforementioned experiments elucidate the upstream regulators of NTRK2-T1 in gCap cells that exhibit impaired regeneration after hyperoxia. Comprehensive understanding of the NTRK2 regulatory network can provide novel drug targets for promoting alveolar capillary regeneration and repair.Example 17Pro-regenerative role of NTRK2 activation in hyperoxia-induced lung injury models[000286] To date, no curative therapy has been developed for improving alveolar regeneration following hyperoxia-induced lung injury, such as BPD. Previous studies have shown that NTRK2 agonist improved alveolar repair and regeneration following acid or influenza-induced lung injury in mice. The present disclosure describe experiments that tests and finds that restoring the balance of NTRK2-FL and NTRK2-T1 in gCap cells can improve vascular function and promote alveologenesis in response to hyperoxia. Although there are certain differences between human and mouse alveologenesis, newborn mice exposed to chronic hyperoxia exhibit lung and vascular developmental arrest, similar to pathology in premature infants with BPD.[000287] Therefore, the present disclosure describes an exemplary experiment delivering NTRK2-FL to mouse lungs exposed to hyperoxia using a polyethyleneimine (PEI)- based nanoparticle system. This system has been specifically designed to target pulmonary microvascular ECs, and provides protection against nucleic acid degradation while facilitating cellular uptake of the genetic material.Example 18Evaluating the effect of NTRK2 agonist or overexpression of NTRK2-FL in hPMVECs and VOs in promoting capillary regeneration during hyperoxia in vitro[000288] The effect of NTRK2 agonist or overexpression of NTRK2-FL in hPMVECs and VOs in promoting capillary regeneration during hyperoxia in vitro were then evaluated. To further revise the transfection efficiency of the lipid nanoparticle delivery system, the experiment involves tuning the size of the particles and dosage of the mRNA. In human umbilical vein ECs (HUVECs), the optimal GFP transfection efficiency was achieved at 2 pg / mL for 400-nm and 900-nm PEIpro particles (Fig. 17). The mRNA delivery system can be revised to achieve a high transfection efficiency of NTRK2-FL mRNA specifically in gCap cells purified from hPMVECs, and in iPSC-VOs.[000289] In order to test whether lipid nanoparticles at a smaller size with multiple dosing can improve transfection efficiency, considering the small diameter of the alveolar capillary bed, the optimal condition can be determined by high level of NTRK2-FL measured by RNAScope, a low level of cell toxicity measured by Caspase3 / 7 staining, and capillary formation in VOs following hyperoxic injury. The activation of downstream signaling such as pAKT and pERK is examined by western blot upon BDNF or NTRK2 agonist 7,8- dihydroxyflavone (7,8-DHF) treatment. The additive effect of NTRK2-FL overexpression and NTRK2 agonist can then be tested.Example 19Therapeutic potential of lipid nanoparticle delivery of NTRK2-FL in hyperoxia mouse model[000290] After in vitro optimization, the therapeutic efficacy of Ntrk2-FL lipid nanoparticle delivery in improving alveolar regeneration and repair in response to hyperoxia induced injury can be evaluated. Briefly, newborn mice were exposed to 75% O2 for 7 days and returned to a room air environment. In some embodiments, in order to induce hyperoxic injury, a mice or other subjects (e.g., vessel organoid model) may be subjected to hyperoxic conditions for about 6 to 14 days, preferably about 6 to 8 days. On postnatal day 2 (P2), lipid nanoparticles containing Ntrk2-FL mRNAs were injected intravenously. Lung tissues fromthree groups (room air control, hyperoxia with GFP control, and hyperoxia with Ntrk2-FL treatment, n=14 mice per group) were harvested at P28. The specificity of the lipid nanoparticle delivery to the lung microvascular ECs is examined by RNAScope. The effects of the Ntrk2- FL gene therapy on lung structure and functions are evaluated by H&E and immunostaining (CD31 and gCap marker genes), and the total lung capacity is determined by flexiVent smallanimal ventilator. The activation status of the Ntrk2 signaling was determined by western blot. Mouse lung gCap cells from these groups are isolated and relevant EC functions are tested as described in previous aims. Multiple dosing of Ntrk2-FL from P2 to P28 were examined. The experiment further included determining if there were synergistic effects of coadministration of Ntrk2-FL and BDNF or the Ntrk2 agonist 7,8-DHF.Example 20Lipid Nanoparticle delivery of NTRK2-FL mRNA to mouse lung ECs in vivo rescued alveolar simplification caused by hyperoxic injury[000291] An example experiment is thus described that finds that lipid nanoparticle delivery of NTRK2-FL mRNA to mouse lung ECs in vivo rescued alveolar simplification caused by hyperoxic injury. To determine the therapeutic potential of NTRK2-FL in mice exposed to hyperoxia, the experiment established a lipid nanoparticle (LNP) delivery system with high specificity for targeting lung ECs (e.g., as shown in Fig. 18A).[000292] To determine the organ-specificity, the LNPs were loaded with luciferase mRNAs. A distinct and pronounced lung-targeted luciferase signal was identified in comparison to other organs (Fig. 18B). To further determine the cell-type specificity of the LNPs, LNPs encasulating Cre mRNA were delivered to Ail4 mice, which express tdTomato upon Cre recombination. Flow cytometry revealed that -94% of tdTomato-positive cells were ECs, with approximately 57% of total ECs were targeted by LNPs (tdTomato+, Fig. 18C).[000293] Next, LNPs encapsulating Ntrk2 -FL mRNA were injected in mice subjected to 90% 02. Intriguingly, a significant improvement of alveologenesis compared to control mice receiving GFP mRNA (Fig. 18D) was observed. The number of alveolar ECs was also drastically increased upon Ntrk2-mRNA therapy (Fig. 18E). These findings further reinforce the beneficial effect of NTRK2-FL on the regeneration and repair of alveolar capillariesfollowing hyperoxia-induced lung injury, highlighting its substantial clinical translational implications.[000294] The overexpression of the NTRK2-FL in gCap cells, iPSC-VOs, and mouse lungs promoted capillary regeneration and alveologenesis in response to hyperoxia-induced lung injury. The specificity of the current nanoparticle system in targeting lung microvascular ECs may have limitations. To address this potential issue and reduce off-target effects, a lipid system can be synthesized that incorporates a ligand-head specifically targeting the receptors APLNR and / or SLC6A4 on gCap cells, in order to improve the specificity of the lipid nanoparticle system and enhance its efficiency in targeting the desired gCap cells in alveoli. Alternatively, a degradable polymer-lipid nanoparticle system (P-amino esters, PBAE) may be utilized that primarily transfect the lung endothelium in mice. The experiment and its findings improved the understanding of alveolar capillary regeneration and repair following hyperoxia- induced lung injury, and determined the therapeutic efficacy of activating NTRK2 signaling for alveolar capillary regeneration and repair in hyperoxic lung injury.Additional comments regarding scientific methodology[000295] Human lung samples from 16 BPD (7 male and 9 female) and 18 age- and gendermatched controls (10 male and 8 female) were collected for the aforementioned experiments through the BRINDL network, which was sufficient to perform statistical analyses. Although healthy control lung from preterm infant was extremely rare, one control lung from a preterm baby born at 29 weeks of gestation was obtained. All VOs were generated from at least three different human iPSC lines, and the differentiation was repeated at least 3 times to increase reproducibility. Variability based on sex was considered and thus the same differentiation was performed with both male and female iPSCs. Each experiment was performed in triplicate and rigorous statistical tests were used to compare different groups. Power calculations were performed to determine that 6-8 mice would be needed per experiment to detect at least a 50% change in an outcome with 20% variability in the control group with an alpha of 0.05 and power of 80%. Equal numbers of female and male mice for each experimental group were used.[000296] The various methods and techniques described above provide a number of ways to carry out the disclosure. Of course, it is to be understood that not necessarily all objectives or advantages described can be achieved in accordance with any particular embodiment described herein. Thus, for example, those skilled in the art will recognize that the methods can be performed in a manner that achieves or optimizes one advantage or group of advantages as taught herein without necessarily achieving other objectives or advantages as taught or suggested herein. A variety of alternatives are mentioned herein. It is to be understood that some preferred embodiments specifically include one, another, or several features, while others specifically exclude one, another, or several features, while still others mitigate a particular feature by inclusion of one, another, or several advantageous features.[000297] Furthermore, the skilled artisan will recognize the applicability of various features from different embodiments. Similarly, the various elements, features and steps discussed above, as well as other known equivalents for each such element, feature or step, can be employed in various combinations by one of ordinary skill in this art to perform methods in accordance with the principles described herein. Among the various elements, features, and steps some will be specifically included and others specifically excluded in diverse embodiments.[000298] Although the application has been disclosed in the context of certain embodiments and examples, it will be understood by those skilled in the art that the embodiments of the disclosure extend beyond the specifically disclosed embodiments to other alternative embodiments and / or uses and modifications and equivalents thereof.[000299] In some embodiments, the numbers expressing quantities of ingredients, properties such as molecular weight, reaction conditions, and so forth, used to describe and claim certain embodiments of the application are to be understood as being modified in some instances by the term “about.” Accordingly, in some embodiments, the numerical parameters set forth in the written description and attached claims are approximations that can vary depending upon the desired properties sought to be obtained by a particular embodiment. In some embodiments, the numerical parameters should be construed in light of the number of reported significant digits and by applying ordinary rounding techniques. Notwithstanding that the numerical ranges and parameters setting forth the broad scope of some embodimentsof the application are approximations, the numerical values set forth in the specific examples are reported as precisely as practicable.[000300] In some embodiments, the terms “a” and “an” and “the” and similar references used in the context of describing a particular embodiment of the application (especially in the context of certain of the following claims) can be construed to cover both the singular and the plural. The recitation of ranges of values herein is merely intended to serve as a shorthand method of referring individually to each separate value falling within the range. Unless otherwise indicated herein, each individual value is incorporated into the specification as if it were individually recited herein. All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or exemplary language (for example, “such as”) provided with respect to certain embodiments herein is intended merely to better illuminate the application and does not pose a limitation on the scope of the application otherwise claimed. No language in the specification should be construed as indicating any non-claimed element essential to the practice of the application.[000301] Preferred embodiments of this application are described herein. Variations on those preferred embodiments will become apparent to those of ordinary skill in the art upon reading the foregoing description. It is contemplated that skilled artisans can employ such variations as appropriate, and the application can be practiced otherwise than specifically described herein. Accordingly, many embodiments of this application include all modifications and equivalents of the subject matter recited in the claims appended hereto as permitted by applicable law. Moreover, any combination of the above-described elements in all possible variations thereof is encompassed by the application unless otherwise indicated herein or otherwise clearly contradicted by context.[000302] All patents, patent applications, publications of patent applications, and other material, such as articles, books, specifications, publications, documents, things, and / or the like, referenced herein are hereby incorporated herein by this reference in their entirety for all purposes, excepting any prosecution file history associated with same, any of same that is inconsistent with or in conflict with the present document, or any of same that may have a limiting affect as to the broadest scope of the claims now or later associated with the present document. By way of example, should there be any inconsistency or conflict between thedescription, definition, and / or the use of a term associated with any of the incorporated material and that associated with the present document, the description, definition, and / or the use of the term in the present document shall prevail.[000303] In closing, it is to be understood that the embodiments of the application disclosed herein are illustrative of the principles of the embodiments of the disclosure. Other modifications that can be employed can be within the scope of the application. Thus, by way of example, but not of limitation, alternative configurations of the embodiments of the application can be utilized in accordance with the teachings herein. Accordingly, embodiments of the present application are not limited to that precisely as shown and described.

Claims

WHAT IS CLAIMED IS:

1. A method of producing a Neurotrophic Receptor Tyrosine Kinase 2 (NTRK2)+vessel organoid comprising a hyperoxic injury (NTRK2+HIVO), the method comprising the steps of: a) activating a vascular endothelial growth factor (VEGF) signaling pathway, a fibroblast growth factor (FGF) signaling pathway, and a brain-derived neurotrophic factor (BDNF) signaling pathway of an angioblast spheroid in a hyperoxic culture condition with fetal bovine serum (FBS) for a first period of time sufficient to form a spheroid comprising acute hyperoxic injury (AHI); and b) activating a VEGF signaling pathway, an FGF signaling pathway, and a BDNF signaling pathway of the spheroid comprising AHI in a normoxic culture condition with FBS for a second period of time sufficient to form the NTRK2+HIVO, wherein the NTRK2+HIVO comprises increased NTRK2 -truncated isoform 1 (NTRK2-T1) expression compared to a control NTRK2+vessel organoid (NTRK2+VO) cultured in a normoxic culture condition.

2. The method of claim 1, wherein the method further comprises producing the angioblast spheroid, by a method comprising: a) culturing induced pluripotent stem cells (iPSCs) to form an embryoid body; b) activating a WNT signaling pathway and a BMP signaling pathway of the embryoid body for a period of time sufficient to form a spheroid comprising mesoderm cells, optionally for about 2 days to about 4 days; and c) activating a VEGF signaling pathway of the spheroid comprising mesoderm cells with forskolin for a period of time sufficient to form the angioblast spheroid, optionally for about 1 day to about 3 days.

3. The method of claim 1 or 2, wherein the angioblast spheroid comprises PECAM1, Ki67, and NTRK2-full length isoform (NTRK2-FL) expression.

4. The method of any one of claims 1-3, wherein the spheroid comprising AHI comprises reduced PEC AMI and Ki67 expression compared to a spheroid cultured in a normoxic culture condition.

5. The method of any one of claims 1 -4, wherein the spheroid comprising AHI comprises decreased NTRK2-FL expression compared to a spheroid cultured in a normoxic culture condition.

6. The method of any one of claims 1-5, wherein the NTRK2+HIVO comprises alveolar capillary endothelial cells (ECs), pericytes, a basement membrane, and a lumen.

7. The method of claim 6, wherein the lumen is about 7-13 pm in diameter.

8. The method of any one of claims 1-7, wherein the NTRK2+HIVO comprises reduced PEC AMI and Ki67 expression compared to a control NTRK2+vessel organoid (NTRK2+VO) cultured in a normoxic culture condition.

9. The method of any one of claims 1-8, wherein the NTRK21HIVO comprises reduced tube formation and cell proliferation compared to a control NTRK2+vessel organoid (NTRK2+VO) cultured in a normoxic culture condition.

10. The method of claim 1 or claim 2, further comprising the step of overexpressing one or more proteins selected from RBFOX2, HOXA5, and MEIS1 in the angioblast spheroid.

11. The method of claim 1 or claim 2, further comprising the step of knocking down RBFOX2 expression in the angioblast spheroid.

12. The method of claim 1 or claim 2, wherein the iPSCs comprise a deletion of one or more genes selected from HOXA5 and MEIS1.

13. The method of any one of claims 1-12, wherein the iPSCs are derived from a patient with a lung disorder, optionally bronchopulmonary dysplasia (BPD).

14. The method of any one of claims 1-13, wherein the iPSCs are human iPSCs.

15. The method of any one of claims 1-14, wherein BDNF is provided at a concentration of about 10 to 40 ug / mL.

16. The method of any one of claims 1-15, wherein the hyperoxic culture condition comprises about 35% O2 to about 80% O2.

17. The method of any one of claims 1-16, wherein the hyperoxic culture condition comprises about 50% O2 to about 80% O2, optionally 75% O2.

18. The method of any one of claims 1-17, wherein the normoxic culture condition comprises about 18% O2 to about 24% O2.

19. The method of any one of claims 1-18, wherein the normoxic culture condition comprises about 20% O2 to about 23% O2, optionally 21% O2.

20. The method of any one of claims 1-19, wherein the first period of time is about 4 days to about 6 days.

21. The method of any one of claims 1-20, wherein the second period of time is about 4 days to about 6 days.

22. The method of any of the preceding claims, further comprising: modeling a treatment for hyperoxic injury using the NTRK2+ HIVO as a model.

23. A NTRK2+HIVO produced by the method of any one of claims 1-21.

24. A Neurotrophic Receptor Tyrosine Kinase 2 (NTRK2)+vessel organoid comprising a hyperoxic injury (NTRK2+HIVO) comprising increased NTRK2-truncated isoform 1 (NTRK2-T1) expression compared to a control NTRK2+ vessel organoid (NTRK2+ VO) cultured in a normoxic culture condition.

25. TheNTRK2 HIVO of claim 23 or 24, further comprising alveolar capillary endothelial cells (ECs), pericytes, a basement membrane, and a lumen.

26. The NTRK2+HIVO of claim 25, wherein the lumen is about 7 to 13 pm in diameter.

27. The NTRK2 HIVO of any one of claims 23-26, wherein the NTRK2+HIVO comprises reduced PECAM1 and Ki67 expression compared to a control NTRK2+VO cultured in a normoxic culture condition.

28. The NTRK2 HIVO of any one of claims 23-27, wherein the NTRK2+HIVO comprises reduced tube formation and cell proliferation compared to a control NTRK2+VO cultured in a normoxic culture condition.

29. The NTRK2 HIVO of any one of claims 23-28, wherein the NTRK21HIVO comprises reduced NTRK2-FL expression compared to a control NTRK2+VO cultured in a normoxic culture condition.

30. The NTRK2 HIVO of any one of claims 23-29, wherein the NTRK2+HIVO comprises increased expression of RBFOX2.

31. The NTRK2 HIVO of any one of claims 23-30, wherein the NTRK2+HIVO comprises reduced expression of HOXA5 and MEIS1.

32. The NTRK2 HIVO of any one of claims 23-31, wherein the NTRK2+HIVO comprises increased expression of HOXA5 and MEIS1.

33. The NTRK2+HIVO of any one of claims 23-32, wherein the NTRK2+HIVO is configured to model a treatment for hyperoxic injury.

34. The NTRK2 HIVO of any one of claims 23-33, wherein the NTRK2+HIVO is derived from human induced pluripotent stem cells (iPSCs).

35. The NTRK2 HIVO of any one of claims 23-34, wherein the NTRK2+HIVO is derived from a patient with a lung disorder, optionally bronchopulmonary dysplasia (BPD).

36. A method of screening a compound for improving alveolar regeneration comprising contacting the NTRK2+HIVO of any one of claims 23-35 with the compound and measuring a response to the compound.

37. A method of producing a lung organoid comprising lung-specific endothelial cells, the method comprising adding lung-specific endothelial cells derived from induced pluripotent stem cells (iPSCs) to a lung epithelial cell organoid derived from iPSCs for about 6 days to about 8 days.

38. The method of claim 37, wherein the lung organoid comprising lung-specific endothelial cells comprises AGER’ ATI cells, HTII-280+AT2 cells, and CD31+lung-specific endothelial cells.

39. A lung organoid comprising lung-specific endothelial cells produced by the method of claim 37 or claim 38.

40. A lung organoid comprising lung-specific endothelial cells, further comprising AGER+ATI cells, HTII-280+AT2 cells, and CD31+lung-specific endothelial cells.

41. A method to produce a lung epithelial -vessel assembloid, the method comprising contacting a vessel organoid (VO) with a lung epithelial organoid.

42. The method of claim 41, wherein the lung epithelial-vessel assembloid comprises CD31+lung-specific endothelial cells and NKX2.1’ lung epithelial progenitor cells.

43. A lung epithelial-vessel assembloid produced by the method of claim 41 or claim 42.

44. A lung epithelial-vessel assembloid comprising CD31+endothelial cells and NKX2.1 lung epithelial progenitor cells.

45. A vascularized alveolar organoid comprising: lung epithelial progenitor cells; lungspecific endothelial cells; and lung-specific mesenchymal cells.

46. The vascularized alveolar organoid of claim 45, wherein the lung epithelial progenitor cells comprises NKX2.1A47. The vascularized alveolar organoid of claim 45 or 46, wherein the endothelial cells express CD31.

48. The vascularized alveolar organoid of any one of claims 45-47, wherein the endothelial cells express FOXF1.

49. The vascularized alveolar organoid of any one of claims 45-48, wherein the endothelial cells express FENDRR.

50. The vascularized alveolar organoid of any one of claims 45-49, wherein the endothelial cells express CDH5.

51. The vascularized alveolar organoid of any one of claims 45-50, wherein the mesenchymal cells express FOXF1.

52. The vascularized alveolar organoid of any one of claims 45-51, wherein the mesenchymal cells express TBX4.

53. A method of increasing endothelial cell proliferation and angiogenesis in pulmonary microvascular endothelial cells (PMVECs), the method comprising: a) transfecting PMVECs with a nucleic acid encoding NTRK2-FL; andb) activating a NTRK2 signaling pathway with BDNF in PMVECs following a posttransfection period.

54. The method of claim 53, wherein transfecting PMVECs with a nucleic acid encoding NTRK2-FL comprises transfecting PMVECs with a lipid nanoparticle comprising NTRK2-FL mRNA.

55. The method of claim 53 or 54, wherein activating NTRK2 signaling pathway of the PMVECs comprising contacting the PMVECs with BDNF for a period of time sufficient to activate the NTRK2 signaling pathway.

56. The method of claim 55, wherein the period of time sufficient to activate NTRK2 signaling following the post-transfection period is about 20-40 minutes.

57. The method of claim 55 or 56, wherein the period of time sufficient to activate NTRK2 signaling following the post-transfection period is about 25-35 minutes.

58. The method of any one of claims 53-57, wherein the post-transfection period is about 35-60 hours.

59. The method of any one of claims 53-58, wherein the post-transfection period is about 40-55 hours.

60. The method of any one of claims 53-59, wherein the nucleic acid encoding NTRK2- FL or the lipid nanoparticle comprising NTRK2-FL mRNA is at a concentration of about 0.5 - 2 pg / ml.

61. The method of any one of claims 53-60, wherein activating NTRK2 signaling pathway of the PMVECs comprising contacting the PMVECs with BDNF for a period of time sufficient to activate the NTRK2 signaling pathway, wherein the BDNF is at a concentration of about 15-35 ng / ml.

62. The method of claim 61, wherein the BDNF is at a concentration of about 20-30 ng / ml.

63. A method of reducing endothelial cell proliferation and impairing angiogenesis in pulmonary microvascular endothelial cells (PMVECs), the method comprising: a) transfecting PMVECs with a nucleic acid encoding NTRK2-T1; and b) activating a NTRK2 signaling pathway of the PMVECs following a posttransfection period.

64. The method of claim 63, wherein transfecting PMVECs with a nucleic acid encoding NTRK2-T1 comprises transfecting PMVECs with a lipid nanoparticle comprising NTRK2-T1 mRNA.

65. The method of claim 63 or 64, wherein activating NTRK2 signaling pathway of the PMVECs comprising contacting the PMVECs with BDNF for a period of time sufficient to activate the NTRK2 signaling pathway.

66. The method of claim 65, wherein the period of time sufficient to activate NTRK2 signaling following the post-transfection period is about 20-40 minutes.

67. The method of claim 65 or 66, wherein the period of time sufficient to activate NTRK2 signaling following the post-transfection period is about 25-35 minutes.

68. The method of any one of claims 63-67, wherein the post-transfection period is about 35-60 hours.

69. The method of any one of claims 63-68, wherein the post-transfection period is about 40-55 hours.

70. The method of any one of claims 63-69, wherein the nucleic acid encoding NTRK2- T1 or the NTRK2-T1 mRNA is at a concentration of about 0.5-2 pg / ml.

71. The method of any one of claims 63-70, wherein activating the NTRK2 signaling pathway of the PMVECs comprising contacting the PMVECs with BDNF for a period of time sufficient to activate the NTRK2 signaling pathway, wherein the BDNF is at a concentration of about 15-35 ng / ml.

72. The method of claim 71, wherein the BDNF is at a concentration of about 20-30 ng / ml.

73. A method of treating an in vitro hyperoxic injury, the method comprising the steps of a) transfecting a spheroid comprising acute hyperoxic injury (AHI) with a nucleic acid encoding NTRK2-FL; b) activating a VEGF signaling pathway, an FGF signaling pathway, and a BDNF signaling pathway of the transfected spheroid comprising AHI in a normoxic culture condition with FBS for a second period of time sufficient to form a treated NTRK21HIVO.

74. The method of claim 73, wherein transfecting the spheroid comprising AHI with a nucleic acid encoding NTRK2-FL comprises transfecting the spheroid comprising AHI with a lipid nanoparticle comprising NTRK2-FL mRNA.

75. The method of claim 73 or 74, wherein the second period of time sufficient to form the treated NTRK2+HIVO is about 4 days to about 6 days.

76. The method of any one of claims 73-75, wherein the nucleic acid encoding NTRK2- FL or the NTRK2-FL mRNAis at a concentration of about 0.5 - 2 pg / ml.

77. The method of any one of claims 73-76, wherein the BDNF signaling pathway is activated by culturing the transfected spheroid comprising AHI with BDNF, optionally wherein the BDNF is at a concentration of about 15 - 35 ng / ml.

78. The method of claim 77, wherein the BDNF is at a concentration of about 20-30 ng / ml.

79. The method of any one of claims 73-78, wherein the spheroid comprising AHT is derived from a method comprising: activating a VEGF signaling pathway, an FGF signaling pathway, and a BDNF signaling pathway in an angioblast spheroid, optionally derived from induced pluripotent stem cells (iPSCs), in a hyperoxic culture condition with FBS for a period of time sufficient to form a spheroid comprising acute hyperoxic injury (AHI).

80. The method of claim 79, wherein the period of time sufficient to form the spheroid comprising AHI is about 4 days to about 6 days.

81. The method of claim 79 or 80, wherein the hyperoxic culture condition comprises about 35% to about 80% O2, optionally about 75% O2.

82. The method of any one of claims 73-81, wherein the treated NTRK21HIVO comprises increased PEC AMI and Ki67 expression compared to a control NTRK2+HIVO.

83. The method of any one of claims 73-82, wherein the treated NTRK2+HIVO comprises increased tube formation and cell proliferation compared to a control NTRK2+HIVO.

84. A method of treating an in vivo hyperoxic injury, comprising increasing expression of NTRK2-FL in cells of the subject.

85. The method of claim 84, wherein increasing expression of NTRK2-FL in cells of the subject comprises transfecting the cells with a nucleic acid encoding NTRK2-FL.

86. The method of claim 85, wherein transfecting the cells with the nucleic acid encoding NTRK2-FL comprises injecting lipid nanoparticles comprising NTRK2-FL mRNA intravenously into the subject.

87. The method of any one of claims 84-86, wherein the subject is a mouse.

88. The method of any one of claims 84-87, wherein the in vivo hyperoxic injury is induced by a method comprising the steps: a) exposing the subject to a hyperoxic condition for a first period of time, optionally about 6 days to about 14 days; and b) returning the subject to a normoxic condition for a second period of time, optionally about 20 days to about 22 days.

89. The method of claim 88, wherein the hyperoxic culture condition comprises about 35% O2 to about 80% O2, optionally 75% O2 and the normoxic culture condition comprises about 18% O2 to about 24% O2, optionally 21% O2.

90. The method of claim 88 or claim 89, wherein the increasing expression of NTRK2-FL in cells of the subject occurs prior to, at the start of, during or after the first period of time.

91. The method of any one of claims 84-90, further comprising: promoting vascular regeneration and / or alveologenesis.

92. The method of any one of the preceding claims, wherein the lipid nanoparticle targets the pulmonary microvascular ECs, optionally wherein the lipid nanoparticle is a polyethyleneimine (PEI)-based nanoparticle.

93. The method of any one of the preceding claims, wherein activating the VEGF signaling pathway of the angioblast spheroid, of the spheroid comprising AHI, of the spheroid comprising mesoderm cells, or of the transfected spheroid comprising AHI comprises contacting the angioblast spheroid, the spheroid comprising AHI, the spheroid comprising mesoderm cells, or the transfected spheroid comprising AHI, respectively, with VEGF.

94. The method of any one of the preceding claims, wherein activating the FGF signaling pathway of the angioblast spheroid, of the spheroid comprising AHI, or of the transfected spheroid comprising AHI comprises contacting the angioblast spheroid, the spheroid comprising AHI, or the transfected spheroid comprising AHI, respectively, with FGF2.

95. The method of any one of the preceding claims, wherein activating the BDNF signaling pathway of the angioblast spheroid, of the spheroid comprising AHI, or of the transfected spheroid comprising AHI comprises contacting the angioblast spheroid, the spheroid comprising AHI, or the transfected spheroid comprising AHI, respectively, with BDNF.

96. The method of any one of the preceding claims, wherein activating the WNT signaling pathway of the embryoid body comprises contacting the embryoid body with CHIR99021.

97. The method of any one of the preceding claims, wherein activating the BMP signaling pathway of the embryoid body comprises contacting the embryoid body with BMP4.

Images