A macrophage-targeting miRNA delivery biomimetic system, and a preparation method and application thereof

Abstract

This invention belongs to the field of biomedicine and nanodelivery technology, specifically relating to a biomimetic system for macrophage-targeted miRNA delivery, its preparation method, and its applications. To address the problems of poor nucleic acid stability, low loading efficiency, insufficient targeting, need for improved biocompatibility, and difficulty in effective enrichment within macrophages in existing miRNA nucleic acid delivery systems, this invention uses hollow mesoporous silica nanoparticles as the core delivery carrier. After amination modification, aminated hollow mesoporous silica nanoparticles are formed, which are loaded with miRNA-regulated nucleic acids and biomimeticly coated with macrophage-derived cell membrane vesicles, forming a macrophage-targeted miRNA delivery biomimetic system with a core-membrane structure. This system can achieve highly efficient loading of miRNA-regulated nucleic acids while possessing good biocompatibility, in vivo safety, and macrophage-targeted delivery capability, and can serve as a macrophage-targeted nucleic acid delivery platform.

Description

Technical Field

[0001] This invention belongs to the field of biomedicine and nanodelivery technology, specifically relating to a biomimetic system for macrophage-targeted miRNA delivery, its preparation method, and its application. Background Technology

[0002] Macrophages are important effector cells in the body's innate immune system, playing a crucial role in the occurrence, development, and outcome of infectious diseases. After infection, macrophages can recognize pathogen-associated molecular patterns through pattern recognition receptors, thereby mediating the phagocytosis and clearance of pathogens, the secretion of inflammatory and chemokine-like factors, the recruitment of other immune cells, and the regulation of the local immune microenvironment. Furthermore, macrophages participate in antigen processing and presentation, connecting innate and adaptive immune responses. Therefore, the functional state of macrophages directly affects pathogen clearance efficiency, the intensity of inflammatory responses, the degree of tissue damage, and infection outcomes, making them important target cells in immune interventions for infectious diseases.

[0003] MicroRNAs, or miRNAs for short, are a class of endogenous non-coding single-stranded RNA molecules approximately 22 nucleotides in length. miRNAs can regulate gene expression at the post-transcriptional level by binding to complementary sequences of target mRNAs, promoting their degradation or inhibiting their translation. Studies have shown that miRNAs are widely involved in immune processes such as macrophage polarization, activation, release of inflammatory factors, autophagy, apoptosis, phagocytosis, and pathogen clearance. Therefore, miRNA-based immunomodulatory strategies offer a new technological direction for the treatment of infectious diseases. However, miRNA-based nucleic acid drugs still face many limitations in application. On the one hand, the complex miRNA regulatory network and non-specific delivery may cause off-target effects and potential safety risks; on the other hand, miRNAs are easily degraded by nucleases, have poor cell membrane permeability, and exhibit low in vivo stability and target cell delivery efficiency, limiting their therapeutic effects. Furthermore, traditional nanocarriers may suffer from insufficient blood circulation stability, significant non-specific distribution, low target cell enrichment efficiency, and potential cytotoxicity during in vivo delivery.

[0004] Therefore, there is an urgent need to develop a biomimetic nanodelivery system that can simultaneously improve miRNA stability, loading efficiency, cellular uptake efficiency, and macrophage-targeted delivery capability, in order to promote the application of miRNA-regulated nucleic acids in infectious diseases and macrophage-related immune regulation. Summary of the Invention

[0005] To address the problems of poor nucleic acid stability, low loading efficiency, insufficient targeting, need for improved biocompatibility, and difficulty in effectively enriching miRNAs in macrophages in existing miRNA delivery systems, this invention provides a biomimetic system for macrophage-targeted miRNA delivery, its preparation method, and its applications.

[0006] To achieve the above objectives, the present invention employs the following technical solution:

[0007] In a first aspect, the present invention provides a biomimetic system for macrophage-targeted miRNA delivery, which uses hollow mesoporous silica nanoparticles (HMSN) as the core delivery carrier. After amination modification, the HMSN forms amination-modified hollow mesoporous silica nanoparticles, which are loaded with miRNA-regulated nucleic acids and biomimeticly coated with a macrophage-derived cell membrane to form a macrophage-targeted miRNA delivery biomimetic system with a nanocore-cell membrane shell structure.

[0008] Furthermore, the average particle size of the hollow mesoporous silica nanoparticles is 90–100 nm. Preferably, the average particle size is about 97.3 nm.

[0009] Furthermore, the amination modification is performed using γ-aminopropyltriethoxysilane to obtain amination-modified HMSN (HMSN-NH2, HN). After amination modification, amino groups are introduced onto the surface of HMSN, increasing its surface potential from negative to positive or near-positive, thereby enhancing its adsorption and loading capacity for negatively charged miRNA-regulated nucleic acids.

[0010] Furthermore, the miRNA regulatory nucleic acid is selected from one or more of miRNA inhibitory nucleic acids, miRNA mimic nucleic acids, and their chemical modifications; wherein, the miRNA inhibitory nucleic acid includes miRNA inhibitor, anti-miR, and antagomiR, and the miRNA mimic nucleic acid includes miRNA mimic and agomir.

[0011] Furthermore, the miRNA-regulated nucleic acid is antagomiR-101-3p.1, and HN is loaded with antagomiR-101-3p.1 to form HNA nanoparticles.

[0012] Furthermore, the macrophage-derived cell membrane is derived from RAW 264.7 cells, THP-1-derived macrophages, bone marrow-derived macrophages, peritoneal macrophages, or other mammalian-derived macrophages. RAW 264.7 cells are preferred.

[0013] Secondly, the present invention provides a method for preparing a biomimetic system for macrophage-targeted miRNA delivery, comprising the following steps:

[0014] Step 1: Prepare aminated hollow mesoporous silica nanoparticles;

[0015] Step 2: Mix and incubate the dispersion of aminated hollow mesoporous silica nanoparticles with miRNA-regulated nucleic acid solution to obtain miRNA-loaded nanoparticles.

[0016] Step 3: Extract the cell membrane from macrophages and prepare macrophage membrane vesicles;

[0017] Step 4: Mix the miRNA-loaded nanoparticles with macrophage membrane vesicles and perform membrane extrusion to obtain a biomimetic miRNA delivery system coated with macrophage membrane.

[0018] Furthermore, the dispersion of the aminated hollow mesoporous silica nanoparticles is mixed with the miRNA-regulated nucleic acid solution at a mass concentration ratio of 100–1000:1, preferably 500:1. Under these preferred conditions, the loading efficiency of antagomiR-101-3p.1 is approximately 87%. It should be noted that the above mass concentration ratio is particularly suitable for chemically modified miRNA-regulated nucleic acids such as antagomiR and / or agomir; antagomiR can serve as a miRNA-inhibiting nucleic acid, and agomir can serve as a miRNA-mimicking nucleic acid. For different types, modification methods, or chain structures of miRNA-regulated nucleic acids, the applicable mass concentration ratio can be appropriately adjusted and optimized based on the molecular weight, chain structure, charge characteristics, and chemical modification method of the nucleic acid molecule.

[0019] Furthermore, the mass ratio of the miRNA-loaded nanoparticles to macrophage membrane vesicles is 1 to 4:1, preferably 2:1. The macrophage membrane is coated onto the HNA surface by membrane extrusion to obtain a macrophage membrane-coated biomimetic miRNA delivery system (MM-HNA).

[0020] Thirdly, this invention provides an application of a macrophage-targeted miRNA delivery biomimetic system for preparing nucleic acid delivery vectors, macrophage function regulation drugs, and immunomodulatory drugs related to infectious diseases.

[0021] Compared with the prior art, the present invention has the following advantages:

[0022] First, this invention uses hollow mesoporous silica nanoparticles (HMSN) as the core carrier, utilizing their high specific surface area, hollow structure and mesoporous channels to achieve effective loading of miRNA-regulated nucleic acids.

[0023] Secondly, this invention introduces amino groups onto the surface of HMSNs through amination modification, changing the surface charge of the nanoparticles from negative to positive or near-positive. This enhances the electrostatic interaction between the nanoparticles and the negatively charged miRNA-regulated nucleic acids, thereby improving the nucleic acid loading efficiency. Experimental results show that the loading rate of antagomiR-101-3p.1 in HNA is approximately 87%, indicating that this system has a high miRNA loading capacity.

[0024] Third, this invention utilizes macrophage-derived cell membranes to biomimeticly coat HNA, constructing an MM-HNA biomimetic delivery system. This system can retain the main protein composition of the macrophage membrane and typical membrane marker proteins CD11b and F4 / 80, indicating that it has a good ability to retain membrane functional characteristics.

[0025] Fourth, the MM-HNA of the present invention has good morphology, uniform particle size, and good dispersibility. TEM results show that the MM-HNA has a typical core-shell structure with a film coating layer of about 8 nm thick on the surface; DLS results show that the MM-HNA has a concentrated particle size distribution and a PDI of about 0.2, indicating that it has good dispersibility.

[0026] Fifth, the MM-HNA of this invention exhibits good biocompatibility and in vivo safety. In vitro cell viability experiments showed that MM-HNA reduced the potential cytotoxicity of RAW 264.7 cells compared to uncoated HNA. In vivo experiments indicated that no significant abnormalities were observed in hematological parameters, liver and kidney function parameters, or the structure of major organs and tissues in mice after MM-HNA administration.

[0027] Sixth, the MM-HNA of this invention has the ability to be preferentially taken up and targeted by macrophages. Flow cytometry results showed that the uptake level of MM-HNA in mouse RAW 264.7 macrophages and human THP-1 macrophages was significantly higher than that in mouse fibroblasts L929, indicating that it has good potential for targeted delivery to macrophages.

[0028] In summary, the MM-HNA biomimetic miRNA delivery system constructed in this invention can achieve efficient loading of miRNA-regulated nucleic acids, and has good biocompatibility, in vivo safety and macrophage-targeted delivery capability. It can be used as a macrophage-targeted nucleic acid delivery platform for research on immune regulation related to infectious diseases and macrophage function regulation. Attached Figure Description

[0029] Figure 1 The images show transmission electron microscopy (TEM) images and average particle size statistics of HMSN, HNA, and MM-HNA in this invention.

[0030] Figure 2This is the standard curve diagram of antagomiR-101-3p.1 in this invention.

[0031] Figure 3 The results of load efficiency measurement of antagomiR-101-3p.1 in this invention are shown.

[0032] Figure 4 The results show the zeta potential detection results of HMSN, HN, HNA and MM-HNA in this invention.

[0033] Figure 5 The results show the hydration particle size, particle size distribution, and dispersibility of MM-HNA in this invention.

[0034] Figure 6 The X-ray diffraction patterns of HMSN, HN, and HNA in this invention are shown.

[0035] Figure 7 The results show the protein composition analysis of HNA, macrophages, macrophage membrane (MM), and MM-HNA in this invention.

[0036] Figure 8 These are the results of the expression detection of HNA, macrophages, macrophage membrane (MM), and MM-HNA macrophage marker proteins CD11b and F4 / 80 in this invention.

[0037] Figure 9 The results show the cell viability assays after RAW 264.7 cells were co-incubated with different concentrations of HNA and MM-HNA for 24 hours in this invention.

[0038] Figure 10 The results of hematoxylin-eosin (HE) staining of mouse heart, liver, spleen, lung and kidney tissues after HNA and MM-HNA treatment in this invention are shown.

[0039] Figure 11 The results show the detection results of serum biochemical indicators ALT, CR and UA, and hematological parameters RBCs, HGB, HCT, WBCs, MPV and PLT in mice after HNA and MM-HNA treatment in this invention.

[0040] Figure 12 The results are flow cytometry analysis and quantitative statistics of MM-HNA uptake in RAW 264.7, THP-1 and L929 cells in this invention.

[0041] Figure 13 This presents the quantitative statistical results of MM-HNA uptake in RAW 264.7, THP-1, and L929 cells in this invention. Detailed Implementation

[0042] To further illustrate the technical solution of the present invention, the present invention will be further described below through embodiments.

[0043] Example 1: Preparation and Characterization of HN Nanoparticles

[0044] 1.1 Preparation of HN nanoparticles

[0045] 12.5 mg of HMSN was dispersed in 12.5 mL of ethanol containing 12.5 μL of γ-aminopropyltriethoxysilane (APTES). After ultrasonic dispersion, the mixture was stirred at 45 °C and 300 rpm for 12 h. After the reaction was completed, the mixture was collected by centrifugation and washed twice with ethanol to remove unreacted APTES. The resulting precipitate was then vacuum dried for 12 h to obtain amino-modified HMSN, i.e., HN nanoparticles.

[0046] 1.2 Surface charge characterization of HN nanoparticles

[0047] The surface charge changes of HMSN and HN were detected using Zeta potential. The results are as follows: Figure 4 As shown, the Zeta potential of HMSN is approximately -20.9 mV, while the Zeta potential of HN increases to approximately +6.11 mV, indicating that amino groups were successfully introduced onto the surface of HMSN, resulting in HN nanoparticles with positive surface charge.

[0048] The above results indicate that this embodiment successfully prepared aminated hollow mesoporous silica nanoparticles (HN).

[0049] Example 2: Preparation and Characterization of HNA Nanoparticles

[0050] 2.1 Preparation of HNA nanoparticles

[0051] Weigh 2 mg of HN powder and add 4 mL of DNase / RNase-Free water to prepare an HN dispersion with a concentration of 0.5 mg / mL. Then, sonicate the HN powder to make it uniformly dispersed.

[0052] Take 6.23 μL of Alexa Fluor 647 fluorescently labeled antagomiR-101-3p.1 stock solution, dilute with DNase / RNase-free water to 1 mL, and prepare an antagomiR-101-3p.1 solution with a concentration of 1 μg / mL. At this point, the mass concentration ratio of HN dispersion to antagomiR-101-3p.1 solution is 500:1.

[0053] The above HN dispersion was mixed with antagomiR-101-3p.1 solution and incubated at 25 °C in the dark for 2 h with stirring, so that antagomiR-101-3p.1 could be adsorbed onto the surface and / or pore structure of HN through electrostatic interaction.

[0054] After incubation, the mixture was centrifuged at 10,000 rpm for 10 min, the precipitate was collected, and the supernatant was retained for the determination of the content of unloaded antagomiR-101-3p.1. The centrifuged precipitate was vacuum dried for 12 h to obtain antagomiR-101-3p.1-loaded HN nanoparticles, denoted as HNA.

[0055] 2.2 Antagomi R-101-3p.1 Load Efficiency Measurement

[0056] The total amount of anagomiR-101-3p.1 initially added to the system is denoted as C. input The amount of unloaded antagomiR-101-3p.1 in the supernatant after centrifugation is denoted as C. free .

[0057] C was detected using a fluorescence spectrophotometer. input and C free The corresponding fluorescence intensity was used to calculate the corresponding content based on the standard curve. The loading efficiency (E) of antagomiR-101-3p.1 was calculated using the following formula:

[0058] ;

[0059] The standard curve of antagomiR-101-3p.1 is as follows: Figure 2 As shown. The load efficiency results for the antagomiR-101-3p.1 are as follows. Figure 3 As shown, when HN and antagomiR-101-3p.1 are mixed at a preferred mass ratio of 500:1, the loading rate of antagomiR-101-3p.1 in HN is approximately 87%, indicating that HN can achieve efficient loading of antagomiR-101-3p.1.

[0060] 2.3 Morphology characterization of HNA nanoparticles

[0061] The morphology and particle size variations of HMSN and HNA were observed using transmission electron microscopy. The results are as follows: Figure 1As shown, HMSN exhibits a uniform spherical structure with good monodispersity and an average particle size of approximately 97.3 nm. HNA also maintains a good nanoparticle morphology with an average particle size of approximately 110 nm. Compared to HN, the particle size of HNA is slightly increased, indicating that antagomiR-101-3p.1 was successfully loaded onto HN nanoparticles.

[0062] 2.4 Zeta potential characterization of HNA nanoparticles

[0063] Zeta potential was used to detect changes in the surface charge of the HNA. The results are as follows: Figure 4 As shown, the Zeta potential of HNA is approximately -7.1 mV, which is significantly lower than the positive potential of HN, indicating that the negatively charged antagomiR-101-3p.1 was successfully loaded onto the surface and / or pore structure of HN nanoparticles.

[0064] The above results indicate that this embodiment successfully prepared HNA nanoparticles loaded with antagomiR-101-3p.1.

[0065] Example 3: Preparation and characterization of MM-HNA biomimetic nanoparticles (miRNA delivery system)

[0066] 3.1 Extraction of macrophage membranes

[0067] RAW 264.7 macrophages were cultured and collected after reaching a suitable growth stage. The cells were lysed under hypotonic conditions, subjected to repeated freeze-thaw cycles, and centrifuged using differential centrifugation to remove nuclei, organelles, and other impurities, resulting in the collection of macrophage membrane precipitate. The macrophage membrane precipitate was resuspended in PBS to obtain a macrophage membrane suspension. Subsequently, using a microliposome extruder, the macrophage membrane suspension was repeatedly extruded through polycarbonate porous membranes with pore sizes of 400 nm and 200 nm to prepare macrophage membrane vesicles (MMs) with relatively uniform particle size.

[0068] 3.2 Preparation of MM-HNA biomimetic nanoparticles

[0069] The HNA nanoparticles prepared in Example 2 were mixed with macrophage membrane vesicles at a mass ratio of 2:1. After thorough mixing, the mixture was repeatedly extruded 15 times using a micro liposome extruder to uniformly coat the macrophage membrane onto the surface of the HNA nanoparticles. After extrusion, excess free membrane vesicles were removed by centrifugation, and the precipitate was collected to obtain the macrophage membrane-coated biomimetic miRNA delivery system (MM-HNA).

[0070] 3.3 Morphology and Core-Shell Structure Characterization of MM-HNA

[0071] The morphology and membrane coating structure of MM-HNA were observed using transmission electron microscopy. The results are as follows: Figure 1 As shown, MM-HNA exhibits a typical core-shell structure with an average particle size of approximately 118 nm. Compared to HNA, an approximately 8 nm thick membrane coating layer can be observed on the surface of MM-HNA, suggesting that the macrophage membrane successfully coats the surface of the HNA nanoparticles.

[0072] 3.4 Zeta potential characterization of MM-HNA

[0073] The surface charge change of MM-HNA was detected using Zeta potential. The results are as follows: Figure 4 As shown, the Zeta potential of MM-HNA is approximately -31.8 mV, exhibiting a significantly negative charge compared to HNA, indicating a marked change in the surface charge characteristics of the nanoparticles after being coated with macrophage membranes. TEM morphology observation and the change in Zeta potential together confirm the successful construction of MM-HNA biomimetic nanoparticles.

[0074] 3.5 Hydrated particle size and dispersibility characterization of MM-HNA

[0075] Dynamic light scattering was used to detect the hydration particle size, particle size distribution, and dispersibility of MM-HNA. The results are as follows: Figure 5 As shown, the particle size distribution of MM-HNA is relatively concentrated, with a PDI of approximately 0.252, indicating that the system has good particle size uniformity and dispersion stability.

[0076] 3.6 XRD characterization of HMSN, HN and HNA

[0077] The structural features of HMSN, HN, and HNA were analyzed using X-ray diffraction. The results are as follows: Figure 6 As shown, HMSN, HN, and HNA all exhibit broad, diffuse peaks typical of amorphous SiO2, and their spectra are generally similar, indicating that the amination modification and antagomiR-101-3p.1 loading did not significantly disrupt the amorphous framework structure of HMSN.

[0078] 3.7 Analysis of membrane protein composition of MM-HNA

[0079] To verify the preservation of macrophage membrane protein composition, SDS-PAGE combined with Coomassie Brilliant Blue staining was used to analyze the protein profiles of HNA, macrophages, macrophage membrane (MM), and MM-HNA. The results are as follows: Figure 7 As shown, MM-HNA retains the major protein bands in the macrophage membrane, indicating that the membrane protein composition is well preserved after macrophage membrane coating.

[0080] The expression of macrophage membrane-associated marker proteins CD11b and F4 / 80 was further detected by Western blot. The results are as follows: Figure 8As shown, CD11b and F4 / 80 were significantly expressed in macrophages, macrophage membrane vesicles (MM), and MM-HNA, while no corresponding protein bands were observed or their expression was significantly lower in HNA, indicating that MM-HNA successfully preserved the typical protein markers and membrane functional characteristics of macrophage membrane.

[0081] The above results demonstrate that the MM-HNA biomimetic nanodelivery system was successfully prepared in this embodiment. This system has good core-shell structure, particle size distribution, dispersibility, and the functional integrity of macrophage cell membrane proteins.

[0082] Example 4: Biosafety Evaluation of MM-HNA

[0083] 4.1 In vitro cell compatibility evaluation

[0084] To evaluate the in vitro cell compatibility of MM-HNA, different concentrations of HNA and MM-HNA were co-incubated with RAW 264.7 cells for 24 h, and cell viability was then detected using the CCK-8 assay. The results are as follows: Figure 9 As shown, HNA at 400 μg / mL significantly reduced the viability of RAW 264.7 cells, while MM-HNA maintained high cell viability within the same concentration range. These results indicate that macrophage membrane coating can effectively improve the cytocompatibility of HNA nanoparticles and reduce their potential cytotoxicity to RAW 264.7 cells.

[0085] 4.2 In vivo safety evaluation

[0086] To evaluate the in vivo biosafety of MM-HNA, C57BL / 6 mice were used as experimental animals. MM-HNA was administered via tail vein injection every 6 days for a total of 5 administrations. After the last administration, blood samples were collected from the mice. One portion was used for routine blood tests, and the other portion was used to separate serum for the detection of liver and kidney function-related biochemical indicators. Simultaneously, major organs such as the heart, liver, spleen, lungs, and kidneys were collected from the mice for histopathological observation.

[0087] HE staining results are as follows Figure 10 As shown, the major organs of mice in each drug administration group, such as the heart, liver, spleen, lungs, and kidneys, remained intact, and no obvious inflammatory cell infiltration, tissue necrosis, or pathological morphological changes were observed.

[0088] Serum biochemical test and hematological analysis results as follows Figure 11As shown, compared with the Control group, no significant abnormalities were observed in the liver function indicators alanine aminotransferase (ALT) and kidney function indicators creatinine (CR) and uric acid (UA) in the MM-HNA-treated mice; no significant abnormalities were also observed in hematological parameters such as red blood cell count (RBCs), hemoglobin (HGB), hematocrit (HCT), white blood cell count (WBCs), mean platelet volume (MPV), and platelet count (PLT).

[0089] The above results indicate that MM-HNA has good in vitro cell compatibility and in vivo biosafety.

[0090] Example 5: Evaluation of macrophage targeting of MM-HNA

[0091] To evaluate the macrophage-targeted delivery capability of MM-HNA, MM-HNA was co-incubated with mouse RAW264.7 macrophages, human THP-1 macrophages, and mouse L929 fibroblasts for 12 h, 24 h, and 48 h, respectively, and the uptake level of MM-HNA in different cells was detected by flow cytometry.

[0092] Flow cytometry analysis and quantitative statistical results are as follows: Figure 12 and Figure 13 As shown, the uptake levels of MM-HNA in RAW 264.7 and THP-1 cells were significantly higher than those in L929 cells, indicating that MM-HNA is more easily taken up and enriched by macrophages.

[0093] The above results indicate that MM-HNA has good macrophage preferential uptake characteristics and targeted delivery capabilities, supporting its application potential as a macrophage-targeted nucleic acid delivery platform.

[0094] The present invention provides a relatively simple method for preparing a biomimetic system for macrophage-targeted miRNA delivery, with a clearly defined material composition. It enables efficient loading and macrophage-targeted delivery of miRNA-regulated nucleic acids. The system exhibits uniform particle size distribution, good biocompatibility and in vivo safety, and is preferentially taken up by macrophages. It can serve as a nucleic acid drug delivery platform, particularly suitable for macrophage-targeted delivery of miRNA-inhibitory nucleic acids and / or miRNA-mimicking nucleic acids.

[0095] In summary, this invention constructs a biomimetic system MM-HNA for macrophage-targeted miRNA delivery, with hollow mesoporous silica nanoparticles as the core, antagomiR-101-3p.1 as the representative miRNA regulating nucleic acids, and encapsulated by a macrophage-derived cell membrane. This system achieves highly efficient loading of antagomiR-101-3p.1 and possesses excellent morphology, dispersion stability, biocompatibility, in vivo safety, and macrophage-targeted delivery capability, providing a novel technological platform for nucleic acid delivery and immune regulation in macrophage-related infectious diseases.

[0096] The foregoing has shown and described the main features and advantages of the present invention. It will be apparent to those skilled in the art that the present invention is not limited to the details of the exemplary embodiments described above, and that the invention can be implemented in other specific forms without departing from its spirit or essential characteristics. Therefore, the embodiments should be considered exemplary and non-limiting in all respects, and the scope of the invention is defined by the appended claims rather than the foregoing description. Thus, it is intended that all variations falling within the meaning and scope of equivalents of the claims be included within the present invention.

[0097] Furthermore, it should be understood that although this specification describes embodiments, not every embodiment contains only one independent technical solution. This narrative style is merely for clarity. Those skilled in the art should consider the specification as a whole, and the technical solutions in each embodiment can also be appropriately combined to form other embodiments that can be understood by those skilled in the art.

Claims

1. A macrophage-targeting miRNA delivery biomimetic system, characterized in that, The biomimetic system uses hollow mesoporous silica nanoparticles as the core delivery carrier. After amination modification, aminated hollow mesoporous silica nanoparticles are formed, which are loaded with miRNA to regulate nucleic acids and biomimeticly coated with a macrophage-derived cell membrane to form a macrophage-targeted miRNA delivery biomimetic system with a nanocore-cell membrane shell structure.

2. The macrophage-targeting miRNA delivery biomimetic system of claim 1, wherein, The hollow mesoporous silica nanoparticles have an average particle size of 90–100 nm.

3. The biomimetic system for macrophage-targeted miRNA delivery according to claim 1, characterized in that, The amination modification was performed using γ-aminopropyltriethoxysilane.

4. The macrophage-targeting miRNA delivery biomimetic system of claim 1, wherein, The miRNA-regulated nucleic acid includes one or more of miRNA inhibitory nucleic acids, miRNA mimic nucleic acids, and their chemical modifications; the miRNA inhibitory nucleic acid includes miRNA inhibitor, antagomiR, and / or anti-miR, and the miRNA mimic nucleic acid includes miRNA mimic.

5. The macrophage-targeting miRNA delivery biomimetic system of claim 4, wherein the targeting moiety is a peptide comprising the amino acid sequence of SEQ ID NO:

1. The miRNA-regulated nucleic acid is antagomiR-101-3p.

1.

6. The macrophage-targeting miRNA delivery biomimetic system of claim 1, wherein, The macrophages are derived from RAW 264.7 cells, THP-1 macrophages, bone marrow macrophages, or peritoneal macrophages.

7. A method for preparing a macrophage-targeted miRNA delivery biomimetic system according to any one of claims 1 to 6, characterized in that, Includes the following steps: Step 1: Prepare aminated hollow mesoporous silica nanoparticles; Step 2: Mix and incubate the dispersion of aminated hollow mesoporous silica nanoparticles with miRNA-regulated nucleic acid solution to obtain miRNA-loaded nanoparticles. Step 3: Extract the cell membrane from macrophages and prepare macrophage membrane vesicles; Step 4: Mix the miRNA-loaded nanoparticles with macrophage membrane vesicles and perform membrane extrusion to obtain a biomimetic miRNA delivery system coated with macrophage membrane.

8. The method for preparing a biomimetic system for macrophage-targeted miRNA delivery according to claim 7, characterized in that, The mass concentration ratio of the dispersion of the aminated hollow mesoporous silica nanoparticles to the miRNA-regulated nucleic acid solution is 100–1000:

1.

9. The method for preparing a biomimetic system for macrophage-targeted miRNA delivery according to claim 7, characterized in that, The mass ratio of the miRNA-loaded nanoparticles to macrophage membrane vesicles is 1–4:

1.

10. Use of the macrophage-targeting miRNA delivery biomimetic system according to any one of claims 1 to 6 for the preparation of a nucleic acid delivery vehicle, characterized in that, The nucleic acid delivery vector is used for macrophage-targeted delivery of miRNA-repressive nucleic acids and / or miRNA-mimicking nucleic acids.

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