An aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles and its preparation and application

Abstract

This invention belongs to the field of pharmaceutical technology, specifically disclosing an aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles and its preparation and application. The material comprises an aerogel and turmeric-derived extracellular vesicle nanoparticles loaded on the aerogel. The loading amount of extracellular vesicle nanoparticles in each 10 mg of aerogel ranges from 10 to 6000 micrograms. The aerogel material possesses advantages such as a wide range of adjustable loading amounts of turmeric-derived extracellular vesicle nanoparticles, high release efficiency, good permeability, good biocompatibility, lightweight, and room-temperature storage. It is a biomaterial with practical application potential and can be made into medical wound dressings for application in wound repair, especially in diabetic ulcers, effectively solving the problem of poor wound healing caused by the inflammatory and oxidative stress microenvironment of diabetic wounds.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, and in particular to an aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles, its preparation and application. Background Technology

[0002] Diabetes mellitus is an endocrine disorder characterized by chronic hyperglycemia, abnormal lipid metabolism, and insulin deficiency or resistance, and its incidence is increasing year by year. Diabetic ulcers are a common complication of diabetes, with a healing rate far slower than in healthy individuals, easily leading to recurrent infections and severely reducing patients' quality of life. The healing process in diabetic wounds involves multiple factors, including mononuclear macrophages, fibroblasts, extracellular matrix, and cytokines, all of which interact and influence each other. Currently, it is widely believed that inflammatory cell infiltration caused by hyperglycemia and ischemic hypoxia, a strong oxidative stress microenvironment, and cellular dysfunction are key reasons for the formation of difficult-to-heal diabetic wounds. Therefore, improving the inflammatory environment of the wound and promoting cell proliferation is an effective strategy for promoting the healing of diabetic wounds.

[0003] Extracellular vesicles (EVs) are cell-secreted nanovesicles with a lipid bilayer structure. Due to their rich content of biomolecules, they possess functional diversity and high biocompatibility, making them promising candidates for wound repair. Current research indicates that EVs derived from animal cells, such as mesenchymal stem cells, macrophages, and fibroblasts, can regulate the wound microenvironment in diabetic patients and promote wound healing. However, the relatively low yield of animal cell-derived EVs severely limits their translational applications.

[0004] In recent years, studies have shown that plant-derived extracellular vesicle nanoparticles (PDNPs) have broad therapeutic effects on various diseases, and their wide availability and high yield give them a significant advantage over animal-derived EVs. For example, wheat-derived PDNPs can promote the proliferation and migration of endothelial cells and dermal fibroblasts; ginseng-derived PDNPs can downregulate β-galactosidase and melanin-related proteins; dendrobium-derived PDNPs can reduce the activity levels of melanin and tyrosine kinases; and cabbage and red cabbage-derived PDNPs can promote mammalian cell proliferation and inhibit apoptosis of immune cells and fibroblasts. These findings all demonstrate the strong biological potential of plant-derived PDNPs in the treatment and prevention of skin-related diseases.

[0005] In the field of diabetic wound repair, endothelial growth factors (EVs) are often combined with functional materials to prepare bio-dressings for better repair and healing effects. Currently, some bio-dressings based on plant-derived plant-derived PDNPs have been developed, such as PDNP hybrid hydrogels and PDNP nanoemulsions. However, due to the unique structure of PDNPs, they have high requirements for storage conditions, often needing to be stored at -80 degrees Celsius. These stringent storage conditions limit the selection of hydrogel types and prevent the daily, universal application of these dressings. Therefore, developing diabetic wound dressings that can improve the inflammatory environment of wounds, promote wound healing, and are easy to store is a key bottleneck in current research. Furthermore, for load-bearing dressings, their loading capacity and release efficiency are crucial in the wound healing process. Parameters such as wound type, size, location, presence of inflammation and exudation impose strict requirements on the PDNP loading capacity, replacement frequency, and breathability of the dressing. However, in the current research field, most dressings have low loading capacity, poor release efficiency, and insufficient breathability, which greatly reduces the actual effect of PDNPs and does not meet the requirements of wound care such as breathability and timely removal of fluid accumulation.

[0006] Aerogels are a novel type of material with a porous network structure, possessing properties such as tunable material properties, extreme lightness, high porosity (80%–99%), and structural stability. These characteristics make them excellent in terms of air permeability, load-bearing capacity, and biocompatibility, making them a promising candidate for next-generation medical wound dressings. Therefore, bio-dressings based on aerogels and plant PDNPs have significant research and translational value in promoting the healing of diabetic wounds. However, the development of related technologies is still in its early stages. Summary of the Invention

[0007] In view of the shortcomings of the prior art described above, the purpose of this invention is to provide an aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles and its preparation and application. Based on aerogel and plant PDNPs, this invention develops a novel biomaterial with a wide range of adjustable PDNP loading content, high release efficiency, good air permeability, good biocompatibility, lightweight, room temperature storage, and practical transformation. It can be made into medical wound dressings, providing a new strategy for promoting wound healing, especially for the repair of diabetic wounds.

[0008] To achieve the above and other related objectives, the first aspect of the present invention provides an aerogel material loaded with turmeric-derived nanoparticles (TDNPs), comprising an aerogel and turmeric-derived TDNPs loaded on the aerogel, wherein the loading amount of TDNPs in each 10 mg aerogel is 10-6000 micrograms.

[0009] In some embodiments, the loading of extracellular vesicle nanoparticles in each 10 mg aerogel is 50-6000 micrograms, preferably 100-6000 micrograms, more preferably 100-1000 micrograms, for example 10, 50, 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1500, 2000, 2500, 3000, 3500, 4000, 4500, 5000, 5500, 6000 micrograms.

[0010] In some embodiments, the aerogel is a medical aerogel.

[0011] In some embodiments, the medical aerogel includes cellulose aerogel, which is selected from at least one of nanocellulose aerogel, regenerated cellulose aerogel, and cellulose derivative aerogel.

[0012] In some embodiments, the aerogel is obtained by cross-linking and drying multiple framework materials; preferably, the framework materials include sodium alginate and nanocellulose, and the cross-linking agent is selected from calcium carbonate.

[0013] In some embodiments, the drying method is selected from freeze drying.

[0014] In some embodiments, the aerogel has a loose, porous network structure; preferably, the turmeric-derived extracellular vesicle nanoparticles are uniformly loaded on the surface and / or inside the aerogel.

[0015] In some embodiments, the water absorption expansion coefficient of the aerogel is 4000-6000 times, preferably 5000-6000 times, for example 5000, 5050, 5100, 5150, 5200, 5250, 5300, 5350, 5400, 5450, 5500, 5550, 5600, 5650, 5700, 5750, 5800, 5850, 5900, 5900, 5950.

[0016] In some embodiments, the turmeric-derived extracellular vesicle nanoparticles are lyophilized powders.

[0017] In some embodiments, after the aerogel loaded with turmeric-derived extracellular vesicle nanoparticles covers the wound surface, the release rate of the loaded extracellular vesicle nanoparticles within 2 days is not less than 70%, preferably not less than 75%, more preferably 75%-85%, and most preferably 78%-82%, for example 78%, 78.5%, 79%, 79.5%, 80%, 80.5%, 81%, 81.5%, and 82%.

[0018] A second aspect of the present invention provides a medical wound dressing comprising the aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles as described in the first aspect, wherein the aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles is used as an active ingredient in the medical wound dressing.

[0019] A third aspect of this invention provides a method for preparing an aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles as described in the first aspect, comprising the following steps:

[0020] Extracellular vesicle nanoparticles derived from turmeric were loaded onto an aerogel to prepare the aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles.

[0021] In some embodiments, the method for preparing the aerogel material is selected from any one of the following methods (I) or (II):

[0022] (I) A solution or suspension containing turmeric-derived extracellular vesicle nanoparticles is mixed evenly with an aerogel, frozen and then dried to obtain the aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles.

[0023] (II) Place the sheet-like or block-like aerogel into a solution or suspension containing extracellular vesicle nanoparticles derived from turmeric, freeze and then dry to obtain the aerogel material loaded with extracellular vesicle nanoparticles derived from turmeric.

[0024] In some embodiments, in methods (I) and (II), the freezing temperature is -80°C to 4°C, for example 4°C, -20°C and -80°C.

[0025] In some embodiments, in methods (I) and (II), the freezing time is not less than 0.5 hours, preferably 0.5 to 24 hours, more preferably 0.5 to 10 hours, and most preferably 1 to 2 hours.

[0026] In some embodiments, in methods (I) and (II), the drying method is selected from vacuum drying.

[0027] In some embodiments, the method for preparing the aerogel includes the following steps:

[0028] The aerogel is prepared by cross-linking multiple skeleton materials and then drying them.

[0029] In some embodiments, the skeleton material includes sodium alginate and nanocellulose, wherein the mass ratio of sodium alginate to nanocellulose is 1:(30-60), preferably 1:(35-50), and more preferably 1:(40-45).

[0030] In some embodiments, the crosslinking agent used in preparing the aerogel is selected from calcium carbonate.

[0031] In some embodiments, the method for preparing sheet-like / or block-like aerogels in method (II) includes the following steps:

[0032] The aerogel stock solution (i.e., aerogel preparation solution) is poured into a mold, frozen and molded, demolded, and dried to obtain sheet or block aerogel.

[0033] In some embodiments, the preparation method of the aerogel stock solution includes the following steps;

[0034] Multiple skeleton raw materials are prepared into solutions, mixed and subjected to cross-linking reaction to obtain the aerogel stock solution.

[0035] In some embodiments, the freeze-forming temperature is -80°C to 4°C, for example 4°C, -20°C and -80°C.

[0036] In some embodiments, the demolding method includes the following steps: after freeze molding, the mold is immersed in an acetone solution containing acetic acid, and after demolding, the acetic acid is removed by washing with the acetone solution; preferably, the volume concentration of acetic acid in the acetone solution containing acetic acid is 10-20%, and the immersion time of the mold in the acetone solution containing acetic acid is 1-3 hours, preferably 1.5-2.5 hours.

[0037] In some embodiments, the drying method for obtaining sheet-like / block-like aerogels is selected from natural air drying or low-temperature drying.

[0038] In some embodiments, the thickness of the sheet / or block aerogel is 0.05-0.6 mm, preferably 0.1-0.6 mm, more preferably 0.1-0.5 mm, and most preferably 0.2-0.3 mm, for example 0.1 mm, 0.15 mm, 0.2 mm, 0.25 mm, 0.3 mm, 0.35 mm, 0.4 mm, 0.45 mm, and 0.5 mm.

[0039] In some embodiments, a solution or suspension containing curcumin-derived extracellular vesicle nanoparticles is prepared using a buffer solution, water, or an organic solvent.

[0040] In some embodiments, the concentration of the solution or suspension containing turmeric-derived extracellular vesicle nanoparticles is ≤10 mg / mL.

[0041] In some embodiments, extracellular vesicle-like nanoparticles are extracted from turmeric using density gradient ultracentrifugation to obtain extracellular vesicle-like nanoparticles derived from turmeric.

[0042] In some embodiments, the extraction method of extracellular vesicle nanoparticles derived from turmeric includes the following steps:

[0043] After removing impurities by gradient ultracentrifugation of the turmeric stock solution, the supernatant was collected, ultracentrifuged again, and the supernatant was discarded. The precipitate was resuspended in buffer solution and ultracentrifuged again to collect the precipitate. The precipitate was then suspended in buffer solution and ultracentrifuged again in a sucrose gradient to obtain extracellular vesicle nanoparticles derived from turmeric.

[0044] In some embodiments, the extraction method of extracellular vesicle nanoparticles derived from turmeric includes the following steps:

[0045] The turmeric extract was centrifuged at 3000g for 30 minutes, 10000g for 30 minutes, and 70000g for 1 hour to remove impurities. The supernatant was then centrifuged at 135000g for 70 minutes and discarded. The precipitate was resuspended in buffer solution and centrifuged again at 135000g for 70 minutes. The precipitate was collected, suspended in buffer solution, and centrifuged again at 150,000g at 4°C for 16 hours in a sucrose gradient, which included buffer solutions with sucrose concentrations of 5%, 10%, 20%, 30%, and 40%. Extracellular vesicle nanoparticles in the 20%-40% layer were collected.

[0046] In some embodiments, the method for obtaining the turmeric stock solution includes: soaking turmeric in a buffer solution and stirring at high speed to obtain a crude turmeric extract mixture; allowing the crude turmeric extract mixture to stand, taking the supernatant, and filtering to obtain the turmeric stock solution.

[0047] The fourth aspect of the present invention provides the use of aerogel materials as described in the first aspect or aerogel materials prepared by the method as described in the third aspect in the preparation or as wound repair drugs.

[0048] In some embodiments, the wound is a complication of diabetes, including diabetic ulcers.

[0049] As described above, the aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles of the present invention, its preparation and application, have the following beneficial effects:

[0050] Based on the characteristic that the inflammatory and oxidative stress microenvironment of diabetic wounds makes wound healing difficult, this invention prepares an aerogel material loaded with TDNPs and develops an aerogel dressing loaded with TDNPs.

[0051] The TDNPs in this invention are extracted using density gradient ultracentrifugation. Biological experiments show that the TDNPs extracted by this method can, on the one hand, transform the inflammatory phenotype of macrophages into an anti-inflammatory phenotype, providing a good healing environment for wounds; on the other hand, TDNPs can activate the endogenous antioxidant system of fibroblasts, resist the stress and survival pressure caused by the high glucose environment of diabetes, and promote their proliferation and wound repair.

[0052] This invention primarily uses sodium alginate and nanocellulose as the framework raw materials, and calcium carbonate particles as the cross-linking agent. A loose, porous aerogel is prepared via a low-temperature freezing method. Further, TDNPs are loaded onto the aerogel, and excess moisture is removed through freeze-drying to obtain an aerogel dressing loaded with lyophilized TDNP powder. Biological experiments show that the aerogel dressing prepared by this method exhibits good adhesion to the skin and provides sustained release of the loaded TDNPs under moist conditions, achieving excellent therapeutic effects in wound repair in a diabetic mouse model.

[0053] More importantly, the aerogel dressing of this invention can easily control the loading content of TDNPs, and the loading range is extremely wide (10-6000 micrograms of TDNPs / 10 milligrams of aerogel), allowing for precise control of TDNP dosage based on wound healing progress. Furthermore, due to its loose and porous structure, the aerogel dressing of this invention has extremely high air permeability (1967±193 g / (m³)). 2 / 24h), standard ≥500g / (m 2 / 24h), human body 240~1800g / (m 2 The TDNP load of this invention is greater than that of most current wound dressings, and it can release 80% of the load within two days, which is very much in line with the frequency of wound dressing changes in clinical practice, greatly improving its practicality.

[0054] Furthermore, the aerogel dressing of this invention can be stored for more than 49 days under normal temperature and freezing conditions, providing a good direction for the long-term preservation and transformation application of TDNPs dressings. Attached Figure Description

[0055] Figure 1 The diagram shows the density gradient centrifugation plot (A), transmission electron microscopy imaging (B), dynamic light scattering analysis results (C), and Zeta potential analysis plot (D) of TDNPs in this embodiment of the invention.

[0056] Figure 2This invention demonstrates that TDNPs can induce macrophages to transform from an inflammatory phenotype to an anti-inflammatory phenotype. Figures A and B show confocal microscopy detection of macrophage uptake of TDNPs and quantitative fluorescence statistics; Figures C and D show flow cytometry detection of polarization phenotype and cytokine gene expression levels after co-incubation of TDNPs and macrophages; Figures E and F show fluorescence imaging and flow cytometry detection after co-incubation of TDNPs and macrophages with an inflammatory phenotype; Figures G and H show quantitative statistics of cytokine transcription and protein levels after co-incubation of TDNPs and macrophages with an inflammatory phenotype; Figure I shows the immunoblotting detection of the TLR4-MyD88 pathway in macrophages after TDNP treatment.

[0057] Figure 3 This invention demonstrates that TDNPs can promote fibroblast proliferation and migration. Figures A, B, and C show the cell viability assay, CCK8 activity assay, and flow cytometry assay of fibroblasts treated with different concentrations of TDNPs. Figures D and E show EdU imaging and quantitative fluorescence statistics of fibroblasts after TDNP incubation. Figure F shows the cell viability analysis of low-density fibroblasts incubated with TDNPs for different time periods. Figures G, H, and I show the scratch assay results, migration rate analysis, and Transwell assay results after TDNP incubation of fibroblasts.

[0058] Figure 4 This diagram illustrates the enhancement of fibroblast antioxidant capacity and its mechanism by which TDNPs enhance the antioxidant capacity of fibroblasts in embodiments of the present invention. Figure A shows fluorescence imaging of fibroblast uptake of TDNPs; Figures B and C show fluorescence imaging and flow cytometry analysis of ROS levels in H2O2-treated fibroblasts after TDNP incubation using the DCFH-DA probe; Figures D and E show flow cytometry detection and cell viability assays of H2O2-treated fibroblasts after TDNP incubation; Figures F, G, and H show differentially expressed gene analysis, enrichment pathways, and antioxidant gene analysis of fibroblasts treated with TDNPs; Figure I shows a protein-protein interaction analysis of the antioxidant gene Nqo1; and Figures J and K show immunoblotting detection of antioxidant-related pathway proteins and apoptosis pathway proteins in fibroblasts after TDNP incubation.

[0059] Figure 5The figures show the characterization of the aerogel dressing in this embodiment of the invention and its good biocompatibility. Figure A shows the scanning electron microscope and elemental analysis of the aerogel dressing; Figure B shows the micro-component scan of the aerogel dressing; Figure C shows the quantitative expansion rate of the aerogel dressing; Figure D compares the water absorption and expansion coefficients of the aerogel dressing (AG), cotton (Cotton), and medical gauze (Gauze); Figures E and F show the CCK8 assay and fluorescence imaging analysis of the aerogel dressing's effect on fibroblast activity; Figure G shows the hematoxylin-eosin staining experiment of mouse skin tissue after 14 days of continuous treatment with the aerogel dressing; Figure I shows the hemolysis experiment of the aerogel dressing; and Figure F shows the good skin adhesion ability of the aerogel dressing.

[0060] Figure 6 The figures show the characterization of the properties of the TDNPs dressing in this embodiment of the invention. Figure A shows the light and airy state of the TDNPs dressing when placed on a flower; Figure B is a scanning electron microscope image of the TDNPs dressing; Figure C is a 3D fluorescence image of the dressing loaded with DiD dye-labeled TDNPs; Figure D shows the activity of the TDNPs dressing under different conditions; Figure E is a statistical graph of the time-release rate of the TDNPs dressing in mouse skin defects; and Figure F is an in vivo fluorescence image of the DiD dye-labeled TDNPs dressing.

[0061] Figure 7 This invention demonstrates how TDNPs dressings promote wound healing in diabetic mice. Figure A shows a flowchart of diabetic wound treatment; Figures B and D show representative wound healing diagrams and simulated wound healing diagrams for each group; Figure C shows the wound healing rate statistics for each group; Figure E shows H&E and Masson's trichrome staining images of representative tissues from each group after day 7 and day 14; Figure F shows DHE staining images of representative tissues from diabetic wounds in each group; and Figure G shows immunohistochemical staining images of representative tissues from diabetic wounds in each group. Detailed Implementation

[0062] The following specific examples illustrate the implementation of the present invention. Those skilled in the art can easily understand other advantages and effects of the present invention from the content disclosed in this specification. The present invention can also be implemented or applied through other different specific embodiments, and various details in this specification can also be modified or changed based on different viewpoints and applications without departing from the spirit of the present invention.

[0063] Example 1

[0064] 1. Experimental Materials and Methods

[0065] 1.1 Extraction and purification of turmeric-derived extracellular vesicle nanoparticles (TDNPs)

[0066] Turmeric was purchased from Baise, Guangxi. The turmeric was washed thoroughly with running water and soaked in phosphate-buffered saline (PBS). The mixture was stirred at high speed to obtain a crude turmeric extract mixture. The mixture was allowed to stand for 10 minutes, and the supernatant was collected and filtered through a coarse sieve to obtain the turmeric stock solution. The turmeric stock solution was centrifuged at 3000g for 30 minutes, 10,000g for 30 minutes, and 70,000g for 1 hour to remove impurities such as fibers. The supernatant was then ultracentrifuged at 135,000g for 70 minutes and discarded. The precipitate was resuspended in PBS and ultracentrifuged again at 135,000g for 70 minutes to collect the precipitate. The precipitate was resuspended in PBS and centrifuged at 150,000g at 4°C for 16 hours in a sucrose gradient (5%, 10%, 20%, 40%, 60% sucrose in 20mM Tris-HCl, pH 7.2) to collect TDNPs from the 20 / 40% layer.

[0067] 1.2 Characterization of TDNPs

[0068] (1) Take 10 μL of TDNPs and add it to a copper grid. Let it stand at room temperature for 10 minutes. After absorbing the liquid on the surface of the copper grid with filter paper, add 20 μL of 3% phosphotungstic acid solution and stain for 10 minutes. Absorb the liquid on the surface of the copper grid with filter paper again and let it air dry for 30 minutes. Observe and record the morphology and particle size of TDNPs by transmission electron microscopy (TEM).

[0069] (2) TDNPs were diluted with PBS, and the particle size and potential of EVs were measured by Zetasizer Nano ZS (Malvern) at a dilution ratio of 1:1000.

[0070] 1.3 Cellular uptake of TDNPs experiment

[0071] TDNPs (200 μg) were labeled with PKH67 dye (1 μL PKH67 + 250 μL dilution C) and incubated in the dark for 3 minutes. Next, an equal volume of 1% BSA in PBS buffer was added for 5 minutes, followed by ultrafiltration (100 kDa) five times to remove unbound dye and BSA. The labeled TDNPs were then added to L929 / RAW264.7 cells and incubated for 4 hours. The cells were then washed with PBS and further stained with DAPI, and observed under a laser scanning confocal microscope (CLSM, Nikon).

[0072] 1.4 TDNPs induce macrophage polarization

[0073] Mouse macrophages RAW264.7 were cultured in glass culture dishes for 24 hours, and then incubated with TDNPs (5 or 10 μg / mL) for another 24 hours. Cells were washed with PBS and labeled with CD86-APC and CD206-PE antibodies (Biolegend) for flow cytometry analysis. The expression of cytokines (IL-10 and TGF-β) was further detected by qPCR.

[0074] 1.5 Biological experiments on inflammatory macrophages using TDNPs

[0075] Mouse bone marrow cells were obtained from 6-8 week old female C57BL / 6J mice. Bone marrow was dissociated into single cells and filtered through a 70 μm cell strainer, followed by treatment with red blood cell (RBC) lysis buffer to remove RBCs. The mixture was centrifuged, and the pellet was cultured in DMEM / FBS medium containing 20 ng / mL mouse GM-CSF (Peprotech). After 5 days of culture, adherent cells (BMDM) were harvested. To verify the effect of TDNPs on M1 macrophages, BMDM cells were co-cultured with LPS (100 ng / mL) for 12 hours to induce M1 polarization. M1 macrophages were then incubated with TDNPs (5 or 10 μg / mL) for another 12 hours. Subsequently, cells were collected and stained with CD86-APC and CD206-PE antibodies, followed by flow cytometry analysis. The expression of cytokines (IL-1β, IL-6, TNF-α, and INF-γ) was further detected by real-time quantitative PCR. Furthermore, fluorescent images were captured using a confocal microscope, and the cells were fixed, permeabilized, and then labeled with CD86 and CD206 antibodies (Proteintech), a second fluorescent antibody (labeled with FITC and Alexa Fluor 647), and Hoechst 33342 before observation.

[0076] 1.6 Immunoblotting assay

[0077] Samples were lysed in RIPA buffer in the presence of protease and phosphatase inhibitors. Proteins were resolved on 5%–15% SDS-polyacrylamide gels and transferred to PVDF membranes. After blocking with 5% BSA, the membranes were incubated overnight at 4°C with appropriate primary antibodies. After washing with TBS-0.1% Tween-20 for 5 min, the membranes were immersed in HRP-labeled secondary antibodies (1:5000 dilution) for 2 h. The membranes were washed again before visualization. The Fdbio-Dura ECL kit (Biostep, Germany) was used for protein band visualization.

[0078] 1.7 TDNPs Toxicity Test

[0079] L929 cells were seeded in 6-well or 96-well plates and cultured for 24 hours. Different concentrations of TDNPs were added to the cells, and incubation was performed for 24 or 48 hours. Cells were washed with PBS and subjected to live / dead fluorescence assays and CCK8 assays. After TDNP incubation, apoptosis in L929 cells was assessed by double staining with Annexin V (FITC) and propidium iodide (PI) via flow cytometry (BD). Reagent preparation was performed according to the manufacturer's instructions.

[0080] 1.8 EdU detection of cell proliferation

[0081] L929 cells were seeded into 6-well plates and cultured overnight. TDNPs (10 μg / mL) were added to the cells, and the cells were incubated for another 24 hours. The cells were then washed with PBS and stained with EdU solution (10 μM) for 2 hours. Next, L929 cells were fixed with 4% paraformaldehyde, treated with Triton X-100 (0.3%), and incubated in the dark with the detection reagent for 30 minutes. Nucleic acids in the cells were stained with Hoechst 33342 for 10 minutes. Images were taken using a fluorescence microscope (excitation wavelengths of Hoechst 33342 and EdU were 350 nm and 488 nm, respectively).

[0082] 1.9 CCK8 assay for cell proliferation

[0083] L929 cells were seeded into 96-well plates and cultured overnight. TDNPs (10 μg / mL) were added to the cells and the cells were cultured for 12, 24, 36, and 48 hours, respectively. L929 cells were washed with PBS and replenished with 90 μL of DMEM containing 10 μL of CCK8 assay reagent. After incubation at 37°C for 2 hours, the absorbance at 450 nm was measured using a microplate reader. Results are expressed as mean ± SD.

[0084] 1.10 Scratch Test

[0085] L929 cells were cultured in 6-well plates and then scraped using a pipette tip. Cells were washed with PBS and replenished with serum-free DMEM with or without TDNPs (10 μg / mL) for 48 hours. Scratch images were captured using a microscope, and the wound closure distance was analyzed using ImageJ software.

[0086] 1.11 Transwell assay

[0087] L929 cells were seeded into 6-well plates and cultured overnight. Cells were washed and cultured for 24 hours in serum-free DMEM containing TDNPs (10 μg / mL). Cells were then collected and placed in the upper chamber of a 24-well plate. The transwell chamber (Corning) was used for transwell assays. The lower chamber was typically filled with DMEM containing 10% FBS. L929 cells were cultured for an additional 36 hours, and non-transferred cells were removed with swabs. Next, the transferred cells were fixed with 4% paraformaldehyde and stained with 0.1% crystal violet.

[0088] 1.12 Intracellular ROS Detection

[0089] ROS production in L929 cells was measured using the oxidation-sensitive fluorescent probe DCFH-DA. After treatment with TDNPs (10 μg / mL) for 12 h, cells were washed with PBS and incubated with DCFH-DA (10 μmol / L) for 20 min. Subsequently, H2O2 (1 mM) was added to the cells for stimulation for 30 min. Finally, the cells were washed with PBS and measured using fluorescence microscopy and flow cytometry.

[0090] 1.13 Biological Experiments on the Enhancement of Antioxidant Capacity by TDNPs

[0091] H2O2 induced oxidative stress in L929 cells. When the cell density reached 50%, PBS or TDNPs (10 μg / mL) were added and incubated for 12 hours. Subsequently, L929 cells were washed with PBS and treated with fresh DMEM medium containing H2O2 (0.5 and 1 mM) for 4 hours. Cell viability was determined by CCK8 assay and apoptosis detection.

[0092] 1.14 RNA sequencing (RNA-seq) and bioinformatics analysis

[0093] RNA sequencing was performed by Novogene Biotech Co., Ltd. to analyze the genetic material of L929 cells. Total RNA was extracted using the RNAeasy Mini Kit (Qiagen) and its quality was assessed using the RNA Nano 6000 assay kit on an Agilent Technologies Bioanalyzer 2100 system (CA, USA). Briefly, mRNA was purified from RNA samples using magnetic beads attached to poly-T oligonucleotides. Library fragments were purified using the AMPure XP system (Beckman Coulter, Beverly, USA) to select cDNA fragments with a preferred length range of 370–420 bp. PCR products were purified (AMPure XP system) and library quality was assessed on an Agilent 2100 bioanalyzer. Indexed samples were clustered on a cBot clustering system using the TruSeq PE Cluster Kit v3-cBot-HS (Illumia) according to the manufacturer's instructions. After cluster generation, the library preparations were sequenced on the Illumina Novaseq platform, generating 150 bp paired-end reads. Differential expression analysis was performed using the DESeq2R package (1.20.0). DESeq2 provides statistical routines for identifying differential expression in digital gene expression data using a negative binomial distribution-based model. The obtained p-values ​​were adjusted using the Benjamini and Hochberg method to control for false discovery rates. Genes with adjusted p-values ​​<= 0.05 identified by DESeq2 were designated as differentially expressed. Gene ontology (GO) enrichment analysis of differentially expressed genes was performed using the Cluster Profiler R package, correcting for gene length bias. GO terms with corrected p-values ​​< 0.05 were considered to have significantly enriched differentially expressed genes.

[0094] 1.15 Preparation of TDNPs dressing

[0095] (1) Preparation of aerogel dressing: A cellulose nanofiber solution (2%, 120 mL) and an alginate solution (0.6%, 10 mL) were mixed and diluted with ultrapure water to 232.4 mL. Then, a Na₂CO₃ solution (0.25 M, 25 mL) was added dropwise to a CaCl₂ solution (0.25 M, 25 mL), and the mixture was vigorously stirred on an ice bath for 10 minutes to obtain a CaCO₃ particle suspension. Next, the CaCO₃ particle suspension (34.16 mL) was injected into the mixture of cellulose nanofibers and alginate, and stirred for 5 minutes. The resulting sample was centrifuged at 800 g for 5 minutes to remove air bubbles. The mixture was poured into a cylindrical mold with a diameter of 2.5 cm and a height of 0.5 cm, and the lid was sealed. The mold was placed at 4 °C for one hour, and then transferred to a refrigerator (-20 °C) overnight. The frozen hydrogel was placed in an acetone solution containing 10% acetic acid and left at room temperature for 2 hours. The gel was then immersed in the acetone solution for solvent exchange. All gels were allowed to air dry naturally to obtain an aerogel dressing. The sample was cylindrical, approximately 2 cm in diameter, 0.2-0.3 mm in height, and weighed approximately 10 mg.

[0096] (2) Preparation of TDNPs loaded aerogel dressing: 200 μL of 1 mg / mL TDNPs solution was loaded into the aerogel dressing sample and frozen at 4℃, -20℃ and -80℃ for 1 h respectively. The PBS buffer in the TDNPs was removed by vacuum drying to obtain the TDNPs dressing.

[0097] 1.16 TDNPs Dressing Sustained-Release TDNPs Efficiency Testing

[0098] The release pattern of TDNPs in TDNP aerogel dressings was determined in vivo. C57BL / 6J mice (6-8 weeks old) were anesthetized by intraperitoneal injection of 2% sodium pentobarbital (50 mg / kg). The mice were shaved, and 1 cm × 1 cm perforated biopsy specimens were excised from their backs. The wounds were treated with lyophilized TDNP aerogel dressings, with 300 μL of physiological saline added. The remaining TDNPs were measured using the BCA protein assay at 3, 6, 9, 12, 24, 36, and 48 hours. The TDNP release rate was calculated as the ratio of remaining protein in the TDNP aerogel dressing to the total protein in the TDNP aerogel dressing.

[0099] 1.17 TDNPs dressing assessment for promoting diabetic wound healing

[0100] (1) Establishment of a full-thickness skin defect model in diabetic mice: C57BL / 6J mice (6-8 weeks old, male) were fasted for 12 hours and induced into a diabetic model by intraperitoneal injection of streptozotocin (150 mg / kg, citrate-sodium citrate buffer, pH 4.2-4.5). One week later, mice with a non-fasting blood glucose level exceeding 16.7 mmol / L were considered diabetic mice. Diabetic mice were anesthetized by intraperitoneal injection of 2% sodium pentobarbital (50 mg / kg), and a full-thickness wound was created on the back of the mice using a disposable biopsy puncturist. Subsequently, the wounds were treated with PBS, TDNP, AG, and TAG, respectively. The same treatment was performed on day 5. For wound area monitoring, digital photographs were recorded on days 1, 4, 7, 10, and 13. The wound area was quantified using ImageJ software, and the wound healing rate was calculated using the following formula: (wound area on day 0 - wound area on day X) / wound area on day 0 × 100%. Mice were sacrificed on day 14, and tissue samples were collected.

[0101] 1.18 Histological Analysis

[0102] Tissue specimens were fixed in 10% formalin and then embedded in paraffin. The tissue samples were cut into 5 μm thick sections, dewaxed and rehydrated, and then stained with hematoxylin and eosin (H&E) and Massen's trichrome. The stained sections were photographed under a microscope. Immunohistochemical staining was performed according to standard protocols; for immunofluorescence analysis, the tissue was frozen in a compound at the optimal cutting temperature and then cut into 5 μm thick sections at -20°C. The sections were blocked with PBS containing 5% BSA at -20°C for 1 hour. The slides were incubated with primary antibody overnight at 4°C. After several washing steps, the sections were incubated with secondary antibody conjugated to the fluorescent dye and DAPI for 60 minutes. Images were acquired using a confocal microscope and analyzed using ImageJ software.

[0103] 2. Experimental Results

[0104] 2.1 Isolation and Characterization of TDNPs

[0105] Curcuma-derived extracellular vesicle nanoparticles (TDNPs) were isolated from homogeneous curcuma using differential and density gradient centrifugation. Most TDNPs were found to accumulate at the 5% / 10% and 20% / 40% interfaces of the sucrose gradient (bands 1 and 2, respectively). Figure 1A). The morphology and size of TDNPs in both bands were characterized using transmission electron microscopy (TEM) and nanoparticle tracking analysis (NTA). TEM results showed that the products in band 1 (TDNPs 1) were a mixture of lipid vesicles, membrane fragments, and protein-like particles, while the nanoparticles in band 2 (TDNPs 2) exhibited a uniform membrane-closed vesicle structure with relatively low contamination from cell debris and lipoproteins. Figure 1 B). TDNPs 1 and TDNPs 2 have similar size distributions, ranging from 35.0 nm to 415.0 nm, with median diameters of 129.5 nm and 112.5 nm, respectively. Figure 1 C). The Zeta potentials of TDNP 1 and TDNP 2 are -23.7 mV and -32.7 mV, respectively. Figure 1 D).

[0106] 2.2 TDNPs regulate macrophage polarization

[0107] Confocal laser scanning microscopy (CLSM) images showed that TDNPs could also be taken up by RAW 264.7, and that TDNP uptake was correlated with TDNP concentration. Figure 2 (A and B). After 24 hours of incubation, TDNPs polarized RAW 264.7 cells into M2 macrophages, and the M2 macrophage maker CD206 increased from 0.96% to 19.6% (5 μg / mL) and 60.4% (10 μg / mL). Figure 2 C), the expression of anti-inflammatory cytokines IL-10 and TGF-β was increased. Figure 2 D). Furthermore, TDNPs reversed the LPS-induced reduction in the proportion of M1 macrophages, decreasing the M1 macrophage marker CD86 from 49.7% to 15.6%. Simultaneously, the proportion of M2 macrophages increased from 5.55% to 40.8%, indicating that TDNPs can reprogram M1 macrophages. Confocal microscopy imaging also showed that TDNP treatment decreased the number of M1 macrophages (CD86+), while the proportion of M2 macrophages (CD206+) increased significantly in a dose-dependent manner. Figure 2 F). Furthermore, TDNPs can downregulate the gene expression levels of inflammatory cytokines IL-1β, IL-6, TNF-α, and INF-γ. Figure 2 G), and regulates the expression of cytokines IL-1β and IL-10 (G). Figure 2The results showed that TDNPs could act as regulators of inflammatory responses. Further investigation revealed that TDNP-induced macrophage polarization was associated with the classical TLR4-MyD88 pathway. Western blot analysis showed that, compared to the LPS group, TDNPs significantly downregulated the expression of TLR4, MyD88, IL-6, and IL-1β in macrophages. Figure 2 I).

[0108] 2.3 TDNPs promote fibroblast proliferation and migration

[0109] The biocompatibility of TDNPs was first assessed, and the results showed that TDNPs at concentrations up to 20 μg / mL did not impair cell viability. Figure 3 A and B) or induce fibroblast apoptosis ( Figure 3 C). Simultaneously, TDNPs can promote fibroblast proliferation (C). Figure 3 (D, E, and F). Furthermore, in vitro migration assays showed that co-culturing with TDNP increased the migration rate of fibroblasts towards the scratch region. Figure 3 G). Quantitative results showed that the migration rates of cells treated with TDNPs were approximately 3.25 and 1.97 times faster than those of control cells at 24 and 48 hours, respectively. Figure 3 H). Furthermore, the results of Transwell experiments also support this conclusion, with more migrating cells observed in the TDNPs-treated fibroblast group. Figure 3 I). These results confirm that TDNPs have a positive regulatory effect on fibroblast proliferation and migration.

[0110] 2.4 TDNPs enhance the antioxidant capacity of fibroblasts and inhibit apoptosis.

[0111] TDNPs (labeled with the membrane dye PKH67) can be internalized by fibroblasts within 4 hours after incubation. Figure 4 A). Furthermore, TDNP pretreatment of fibroblasts for 12 hours can reduce H2O2-induced ROS accumulation ( Figure 4 B), flow cytometry results showed that the proportion of ROS-excess cells in the TNDPs group decreased from 62.4% to 30.1%. Figure 4 C), while H2O2 stimulation induced apoptosis in fibroblasts, decreasing from 74.2% to 48.0% (C). Figure 4 D). Even when exposed to high concentrations of H2O2 (1 mM), TDNPs can restore the overall viability of fibroblasts. Figure 4 E). These results indicate that TDNPs are involved in antioxidant pathways in fibroblasts to clear ROS accumulation.

[0112] RNA sequencing was performed on TDNP-treated cells using untreated fibroblasts as a control. This study identified 666 genes that showed differential expression between the control and TDNP-treated groups. Volcano plot analysis of differentially expressed genes (DEG, p < 0.05) showed that 288 genes were upregulated and 378 genes were downregulated in TDNP-treated cells. Figure 4 F). Gene Ontology (GO) enrichment analysis confirmed that DEG is involved in multiple biological processes, including antioxidant activity, glutathione binding, glutathione transferase activity, and lipid oxidation. Figure 4 G). This embodiment also investigated the effects of TDNPs on the expression of genes associated with oxidative stress. As expected, the heatmap showed that 18 of the 23 genes were upregulated in cells treated with TDNPs (p < 0.01), many of which are known transcriptional targets of the cellular antioxidant system, including Acox2, Nqo1, Cd36, CAT, etc. Figure 4 H). To further elucidate the role of DEG in cellular oxidation state, this embodiment constructed a protein-protein interaction network using Nqo1, an upregulated gene that plays an important role in redox regulation. Figure 4 I). Therefore, Western blot experiments showed that TDNP treatment significantly increased the expression of phosphorylated Nrf2 (p-Nrf2), NQO1, and HO-1 in a concentration-dependent manner. Figure 4 J). Furthermore, it was confirmed that TDNPs reduced BAX expression in fibroblasts and increased the expression of p-Nrf2, HO-1, NQO1, and Bcl-2 under oxidative stress. Figure 4 The above results indicate that TDNPs enhance the ability of fibroblasts to resist excessive ROS and reduce apoptosis by activating endogenous antioxidant pathways.

[0113] 2.5 Preparation and characterization of turmeric-derived extracellular vesicle nanoparticle dressing (TDNPs@aerogel, TDNPs@AG)

[0114] Aerogel dressings were prepared by a freezing method. Elemental analysis and scanning electron microscopy revealed that the prepared aerogels possessed a porous network structure. Figure 5 A and B), compared to cotton balls and gauze, have a water absorption and swelling coefficient nearly 6000 times higher. Figure 5 C and D). CCK8 assay and cell viability testing showed that the aerogel has good biocompatibility. Figure 5 E and F), no allergic reaction ( Figure 5 G) and hemolytic effect ( Figure 5 H), which has a good skin-adhesive effect and is suitable for epidermal wound dressings. Figure 5 I).

[0115] Dressings with TDNPs are lightweight ( Figure 6 A), scanning electron microscopy and confocal microscopy three-dimensional imaging show that TDNPs are uniformly distributed and have a size at the nanometer level. Figure 6 B and C) can maintain high activity within 49 days under different storage conditions (room temperature, 4°C, -20°C, and -80°C). Figure 6 D). Furthermore, it can gradually release TDNPs at the animal's epidermal wound site, achieving a long-lasting therapeutic effect. Figure 6 E and F).

[0116] 2.6 Turmeric-derived extracellular vesicle nanoparticle dressing (TDNPs@AG) promotes wound healing in diabetic patients.

[0117] The wound healing potential of TDNPs@AG (TAG) was evaluated in vivo using a full-thickness skin defect model in diabetic mice. Figure 7 A). Mice were divided into four groups and treated with PBS, AG, TDNPs, and TAG on day 0 and day 5 after wound formation, respectively. Figure 7 As shown in B, compared with AG and the control group, the TAG and TDNP treatment groups significantly accelerated the wound healing process and showed a higher wound contraction rate than the other two groups. Figure 7 C and D). Interestingly, mice treated with TAG showed better healing efficiency than the TDNPs group (C and D). Figure 7 (B) This may be attributed to the superiority of aerogel dressings and the long-term modulatory effects of TDNPs. H&E and Masson staining showed that in the groups using TAG and TDNPs, the severity of inflammatory infiltration in the wound tissue was reduced, the wound width was decreased, and collagen deposition was increased. Figure 7 E). By day 14, the wounds in the TAG and TDNPs groups had completely healed, with the microstructure continuously rearranging. In contrast, the control and AG groups showed significant defects and were still in the early stages of regeneration. Accordingly, DHE histological staining showed elevated ROS levels in the AG and control groups compared to the TDNPs and TAG groups (E). Figure 7 F). Furthermore, wounds treated with formulations containing TDNPs exhibited enhanced proliferative activity (Ki67), angiogenesis (CD31), and collagen production (COL1A1), demonstrating the effective cross-boundary regulation of natural plant drugs in mammalian tissues. Figure 7 G).

[0118] In summary, this embodiment first extracts turmeric-derived extracellular vesicle nanoparticles using density gradient ultracentrifugation; then, using sodium alginate and nanocellulose as the framework raw materials and calcium carbonate particles as the crosslinking agent, a loose and porous aerogel is prepared by low-temperature freezing. Further, TDNPs are loaded onto the aerogel, and excess water is removed by freeze-drying to obtain an aerogel dressing loaded with TDNPs freeze-dried powder.

[0119] Biological experiments showed that TDNPs can, on the one hand, convert the inflammatory phenotype of macrophages to an anti-inflammatory phenotype, providing a favorable environment for wound healing; on the other hand, TDNPs can activate the endogenous antioxidant system of fibroblasts, resisting the stress and survival pressure caused by the high glucose environment of diabetes, and promoting their proliferation and wound repair. Meanwhile, the turmeric-derived extracellular vesicle nanoparticle dressing prepared by the method of this invention exhibits good adhesion to the skin and slowly releases the loaded TDNPs under moist conditions, achieving excellent therapeutic effects in wound repair in a diabetic mouse model. Furthermore, the TDNP dressing prepared by the method of this invention can be stored for more than 49 days under both room temperature and frozen conditions, providing a promising direction for the long-term preservation and transformation application of EV dressings.

[0120] Example 2

[0121] 1. Preparation of TDNPs dressings with different TDNPs loading contents (TDNPs@AG)

[0122] According to the TDNPs dressing preparation method disclosed in section 1.7 of Example 1, a 10 μg / mL TDNPs solution (obtained by dilution with PBS) was added to the aerogel dressing sample to prepare TDNPs dressings with different TDNPs loading contents (10, 100, 1000, 2000, 3000, 4000, 5000, 6000 μg / sample).

[0123] 2. Performance comparison of different TDNPs dressings (TDNPs@AG)

[0124] As shown in Table 1, compared with other types of EV dressings, the turmeric-derived extracellular vesicle nanoparticle aerogel dressing of the present invention can easily control the loading content of TDNPs, and the loading range is extremely wide (10-6000 micrograms of TDNPs / 10 milligrams of aerogel). This allows for precise control of TDNP dosage based on wound healing progress. Furthermore, due to its loose and porous structure, the aerogel dressing sample prepared by the present invention has extremely high air permeability (the water vapor permeability of this dressing was 1967±193 g / (m²) as tested according to YY / T 0148-2006 General Requirements for Medical Tape). 2 The standard requires that the water vapor permeability of medical dressings be ≥500g / (m² / 24h).2 The normal human skin water vapor evaporation rate is 240–1800 g / (m²) / 24h. 2 The effect of this invention on wound dressings exceeds that of most current wound dressings; moreover, the dressing of this invention can release 89.5% of the loaded TDNPs within two days. Figure 6 E) is very much in line with the frequency of wound dressing changes in clinical practice, which greatly improves its practicality.

[0125] Table 1. Performance parameters of the extracellular vesicle dressings currently under research and the turmeric-derived extracellular vesicle nanoparticle aerogel dressing of the present invention.

[0126]

[0127]

[0128]

[0129]

[0130] The above embodiments are merely illustrative of the principles and effects of the present invention and are not intended to limit the invention. Any person skilled in the art can modify or alter the above embodiments without departing from the spirit and scope of the present invention. Therefore, all equivalent modifications or alterations made by those skilled in the art without departing from the spirit and technical concept disclosed in the present invention should still be covered by the claims of the present invention.

Claims

1. An aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles, characterized in that, include: It includes an aerogel and turmeric-derived extracellular vesicle nanoparticles loaded on the aerogel, with the loading of extracellular vesicle nanoparticles in each 10 mg aerogel being 10-6000 micrograms. The aerogel is obtained by cross-linking and drying multiple framework materials, including sodium alginate and nanocellulose, and the cross-linking agent is selected from calcium carbonate. The extraction method of the extracellular vesicle nanoparticles derived from turmeric includes the following steps: The turmeric stock solution was centrifuged at 3000 g for 30 minutes, 10000 g for 30 minutes, and 70000 g for 1 hour to remove impurities. The supernatant was then ultracentrifuged at 135000 g for 70 minutes and discarded. The precipitate was resuspended in buffer solution and ultracentrifuged again at 135000 g for 70 minutes. The precipitate was collected, suspended in buffer solution, and then centrifuged again at 150,000 g at 4°C for 16 hours in a sucrose gradient, which included buffer solutions with sucrose concentrations of 5%, 10%, 20%, 30%, and 40%. Extracellular vesicle nanoparticles were collected from the 20%-40% layer.

2. The aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles according to claim 1, characterized in that: The aerogel is a medical aerogel; And / or, the extracellular vesicle-like nanoparticles are lyophilized powders.

3. The aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles according to claim 2, characterized in that: The medical aerogel includes cellulose aerogel.

4. The aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles according to claim 1, characterized in that: The aerogel has a water absorption and swelling coefficient of 40-60 times; And / or, after the aerogel loaded with turmeric-derived extracellular vesicle nanoparticles covers the wound surface, the release rate of the loaded extracellular vesicle nanoparticles is not less than 70% within 2 days.

5. A medical wound dressing, characterized in that: Including the aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles according to any one of claims 1 to 4.

6. A method for preparing an aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles according to any one of claims 1 to 4, characterized in that, Includes the following steps: Extracellular vesicle nanoparticles derived from turmeric were loaded onto an aerogel to prepare the aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles.

7. The preparation method according to claim 6, characterized in that, The preparation method of the aerogel material is selected from any one of the following methods (I) or (II): (I) A solution or suspension containing turmeric-derived extracellular vesicle nanoparticles is mixed evenly with an aerogel, frozen and then dried to obtain the aerogel material loaded with turmeric-derived extracellular vesicle nanoparticles. (II) Place the sheet-like or block-like aerogel into a solution or suspension containing extracellular vesicle nanoparticles derived from turmeric, freeze and then dry to obtain the aerogel material loaded with extracellular vesicle nanoparticles derived from turmeric.

8. The preparation method according to claim 7, characterized in that: The preparation method of sheet / or block aerogel in method (II) includes the following steps: pouring the aerogel stock solution into a mold, freezing and molding, demolding, and drying to obtain sheet or block aerogel.

9. The preparation method according to claim 7, characterized in that: The thickness of the sheet / or block aerogel is 0.05~0.6cm.

10. The use of the aerogel material according to any one of claims 1 to 4 or the aerogel material prepared by the method according to any one of claims 6 to 9 in the preparation of wound repair drugs.

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