Composite hydrogel loaded with herba violae exosome as well as preparation method and application of composite hydrogel

Abstract

The invention belongs to the technical field of biomedical materials, and discloses a viola philippica exosome-loaded composite hydrogel as well as a preparation method and application thereof, the composite hydrogel is prepared by mixing methacrylated polylysine (PLMA), oxidized dextran (ODex), viola philippica exosome (V-E), a photoinitiator and a physiologically compatible solvent and performing illumination crosslinking; the preparation method comprises the following steps: respectively preparing PLMA and ODex solutions, extracting and purifying V-E, adding a photo-crosslinking agent (LAP), mixing the components to obtain a precursor liquid, and carrying out in-situ curing molding through ultraviolet-visible light irradiation. The composite hydrogel provided by the invention solves the problems that the existing wound dressing is difficult to fit with irregular wounds, is antibacterial and easy to resist drugs, and lacks active anti-inflammatory and efficient healing promotion capabilities, and is suitable for repair treatment of infectious open wounds such as burns, trauma, diabetic foot ulcer and postoperative infected incisions.

Description

Technical Field

[0001] This invention relates to the field of biomedical materials technology, specifically to a composite hydrogel loaded with Viola yedoensis exosomes, its preparation method, and its application. Background Technology

[0002] Open wounds, such as burns, traumatic injuries, and surgical incisions, often result in large, irregularly shaped wounds. These wounds are highly susceptible to bacterial infection, leading to excessive and persistent inflammatory responses and oxidative stress, severely hindering the normal healing process. For open wounds, clinical treatment commonly uses dressings (such as gauze, films, and sponges) and some novel antibacterial dressings (such as silver-containing dressings and antibiotic-loaded dressings). However, existing dressings and novel antibacterial dressings all have shortcomings, mainly including: 1) poor adhesion, making it difficult to tightly adhere to and effectively fill irregular wounds, easily creating dead space and leading to secondary infection; 2) limited antibacterial properties, or the risk of inducing bacterial resistance; 3) passive function, lacking the ability to actively regulate the excessively inflammatory microenvironment of the wound; 4) insufficient repair-promoting capacity, with limited effect on promoting tissue regeneration.

[0003] Injectable photocurable hydrogels, due to their ability to be injected in situ and cured by light, can perfectly conform to wounds of any shape, making them a promising dressing matrix material. In constructing hydrogels, ε-polylysine (ε-PL), a natural cationic antimicrobial peptide, possesses broad-spectrum antimicrobial activity, but its poor mechanical strength limits its application alone. By modifying ε-PL with methacrylamide, methacrylated polylysine (PLMA) is obtained, retaining its antimicrobial activity while introducing photopolymerizable double bonds, thereby enhancing the crosslinkability and mechanical properties of the hydrogel material. Oxyglucan (ODex) undergoes a dynamic Schiff base reaction with the amino groups of PLMA through its aldehyde group, forming a dual-network structure with the photocrosslinking network of PLMA. This structure not only significantly improves the mechanical strength of the hydrogel but also endows it with self-healing properties, improving the stability and lifespan of the dressing in complex environments.

[0004] Wound healing is a complex biological process that requires the simultaneous and effective control of infection and regulation of inflammation. In recent years, plant-derived exosomes have attracted widespread attention due to their low immunogenicity, high biocompatibility, and natural targeted delivery capabilities. Viola yedoensis, a traditional Chinese medicine, has extracts that have been shown to possess good anti-inflammatory and antioxidant activities. Viola yedoensis exosomes (VE), one of its active ingredients, are natural nanoscale vesicles. Studies have shown that they have potential application value in regulating immune cells (such as promoting macrophage polarization towards an anti-inflammatory phenotype) and inhibiting the excessive release of inflammatory factors, providing a new strategy for proactively intervening in the wound inflammatory microenvironment.

[0005] However, in the prior art, there is no technology that combines PLMA / ODex dual-network photocurable hydrogel with Viola yedoensis exosomes (VE) to achieve multiple functions such as in-situ adaptation to irregular wounds, long-lasting antibacterial activity, active anti-inflammatory effects, and efficient wound healing promotion. Therefore, this invention provides a composite hydrogel loaded with Viola yedoensis exosomes, which is of great significance for improving the clinical treatment effect of open wounds, especially complex infected wounds. Summary of the Invention

[0006] The present invention aims to provide a composite hydrogel loaded with Viola yedoensis exosomes, its preparation method and application, in order to solve the problems of existing wound dressings that are difficult to adhere to irregular wounds, are prone to drug resistance, and lack active anti-inflammatory and efficient healing promotion capabilities.

[0007] To achieve the above objectives, the present invention provides the following technical solution:

[0008] A composite hydrogel loaded with Viola yedoensis exosomes, by weight percentage, consists of 10-25% methacrylamide polylysine, 5% oxidized dextran, 1-20% Viola yedoensis exosomes, 0.25% photoinitiator, and 49.75-83.75% physiologically compatible solvent.

[0009] Furthermore, by mass percentage, it consists of 12% methacrylamide polylysine, 5% oxidized dextran, 10% Viola yedoensis exosomes, 0.25% photoinitiator, and 72.75% physiologically compatible solvent.

[0010] Furthermore, the photoinitiator is lithium phenyl-2,4,6-trimethylbenzoylphosphinic acid.

[0011] Furthermore, the physiologically compatible solvent is phosphate buffer.

[0012] A method for preparing a composite hydrogel loaded with Viola yedoensis exosomes includes the following steps:

[0013] S1. Prepare a solution of methacrylamide polylysine and an oxidized dextran solution;

[0014] S2. Extract exosomes from Viola yedoensis;

[0015] S3. Mix the methacrylamide polylysine solution, oxidized dextran solution, photoinitiator and Viola yedoensis exosomes in a certain mass ratio to obtain a precursor mixture;

[0016] S4. Place the precursor mixture prepared in step S3 into the target area and perform photocrosslinking to form a hydrogel.

[0017] Further, in S1, methacrylated polylysine is prepared by reacting ε-polylysine with methacrylic anhydride, and the methacrylated polylysine is dissolved in a physiologically compatible solvent to obtain a methacrylated polylysine solution; oxidized dextran is prepared by oxidizing dextran with sodium periodate, and the oxidized dextran is dissolved in a physiologically compatible solvent to obtain an oxidized dextran solution.

[0018] Furthermore, in S2, Viola yedoensis exosomes were extracted from fresh Viola yedoensis whole plant by ultra-high speed centrifugation and resuspended in a physiologically compatible solvent.

[0019] Furthermore, in the photocrosslinking of S4, the wavelength of the light is 365~405 nm, and the light intensity is 5~50 mW / cm². 2 The irradiation time is 10-60 seconds.

[0020] The application of a composite hydrogel loaded with Viola yedoensis exosomes in the preparation of medical devices or dressings for the prevention and / or treatment of infected wounds.

[0021] Furthermore, the infected wound includes burn wounds, traumatic open wounds, diabetic foot ulcers, or postoperative infected incisions.

[0022] The beneficial effects of the technical solution are:

[0023] 1. This invention provides a composite hydrogel loaded with Viola yedoensis exosomes, which is injectable and photocurable, allowing it to be injected into irregular wounds of arbitrary shape and depth. Under gentle light, it rapidly molds in situ, forming a solid dressing that closely conforms to the wound contour, effectively eliminating dead space and providing ideal three-dimensional spatial support for healing. The synergistic effect of the photocrosslinking network of PLMA and the dynamic Schiff base crosslinking network between PLMA and ODex significantly enhances the mechanical strength and toughness of the hydrogel, enabling it to withstand mechanical stress at the wound site. Simultaneously, the dynamic covalent bonds endow the hydrogel with self-healing capabilities, allowing it to repair itself if damaged during use, maintaining the integrity and long-lasting protective function of the dressing. Furthermore, PLMA inherits the cationic antimicrobial peptide properties of ε-polylysine, exerting its effects by physically disrupting bacterial cell membranes. It has a broad antimicrobial spectrum and is less likely to induce bacterial resistance, providing a reliable guarantee for controlling wound infection. The loaded vitamin E, as a highly efficient natural active ingredient delivery system, can actively regulate the polarization of wound macrophages towards an anti-inflammatory phenotype, inhibiting excessive release of inflammatory factors, thereby effectively alleviating excessive inflammatory response and oxidative stress in the wound and creating a favorable microenvironment for tissue repair. The bioactivity of vitamin E itself, combined with the constructed three-dimensional hydrogel matrix, can promote angiogenesis, epithelialization, and collagen deposition, thereby accelerating the full-thickness healing process of the wound. Moreover, the main components of this composite hydrogel, PLMA, ODex, and vitamin E, are all biologically derived materials with inherent good biocompatibility and low immunogenicity. The hydrogel network can be gradually degraded and absorbed in the physiological environment after completing its therapeutic function, eliminating the need for secondary surgery and effectively reducing patient suffering.

[0024] 2. This invention provides a method for preparing a composite hydrogel loaded with Viola yedoensis exosomes. The raw material preparation methods are mature, such as PLMA being prepared by reacting ε-polylysine with methacrylic anhydride, ODex being prepared by oxidizing dextran, and VE being extracted by ultra-high speed centrifugation. These operation steps are easy to operate and repeat. Moreover, the preparation conditions are mild and efficient. The extraction and purification methods of VE can better preserve its biological activity. The preparation of the precursor mixture is simple, requiring only uniform mixing. The photocuring process uses low-intensity visible / near-ultraviolet light (365~405 nm, 5~50 mW / cm²) for a short time, which does not cause thermal damage to tissues and has a fast curing speed, making it suitable for immediate clinical use. Furthermore, the mechanical properties, drug release rate, and biological activity of the hydrogel can be flexibly adjusted by controlling the mass ratio of each component to meet different clinical needs.

[0025] 3. The present invention provides an application of a composite hydrogel loaded with Viola yedoensis exosomes for the preparation of medical devices, dressings, or drugs for the prevention and / or treatment of infected wounds. It is particularly suitable for complex and difficult-to-heal wounds such as burns, traumatic open wounds, diabetic foot ulcers, and postoperative infected incisions, and has significant clinical translational potential. Compared with traditional dressings, it not only provides a physical barrier but also actively regulates the wound microenvironment through a multi-effect synergistic effect of antibacterial, anti-inflammatory, and healing-promoting properties, significantly accelerating the healing process and reducing the risk of infection and complications. Furthermore, the injectable precursor can be directly infused into the wound and rapidly solidifies in situ, simplifying dressing application. It is especially suitable for pre-hospital emergency care, intraoperative and outpatient treatment, effectively improving treatment success rates and patient compliance. Attached Figure Description

[0026] Figure 1 This is a flowchart illustrating the preparation of the PLMA / ODex@VE composite hydrogel in Example 2 of the present invention;

[0027] Figure 2 This is a transmission electron microscope (TEM) image of VE in Embodiment 2 of the present invention;

[0028] Figure 3 This is a particle size distribution diagram of VE in Embodiment 2 of the present invention;

[0029] Figure 4 These are scanning electron microscope (SEM) images of different groups of hydrogels in Example 3 of the present invention;

[0030] Figure 5 This is a porosity diagram of different groups of hydrogels in Example 3 of the present invention;

[0031] Figure 6 This is a diagram showing the pore size of different groups of hydrogels in Example 3 of the present invention;

[0032] Figure 7 This is a staining image of live / dead bacteria of Escherichia coli (S. aureus), Staphylococcus aureus (E. coli), and methicillin-resistant Staphylococcus aureus (MRSA) in Example 4 of the present invention;

[0033] Figure 8 This is an immunofluorescence staining image of RAW264.7 cells expressing iNOS after different treatment groups in Example 5 of the present invention;

[0034] Figure 9 This is a quantitative analysis diagram of iNOS fluorescence intensity in Example 5 of the present invention;

[0035] Figure 10 This is an immunofluorescence staining image of RAW264.7 cells expressing Arg-1 after different treatments in Example 5 of the present invention;

[0036] Figure 11 This is a quantitative analysis diagram of Arg-1 fluorescence intensity in Example 5 of the present invention;

[0037] Figure 12 This is a ROS staining image of HUVEC cells from different groups in Example 5 of the present invention;

[0038] Figure 13 This is a quantitative analysis diagram of ROS staining fluorescence intensity in Example 5 of the present invention;

[0039] Figure 14 The H&E staining and MASSON image of rat skin tissue in Example 6 of this invention; Detailed Implementation

[0040] The present invention will now be described in further detail with reference to the accompanying drawings and embodiments:

[0041] Example 1: A composite hydrogel loaded with Viola yedoensis exosomes, comprising, by weight percentage, 10-25% methacrylamide polylysine, 5% oxidized dextran, 1-20% Viola yedoensis exosomes, 0.25% photoinitiator, and 49.75-83.75% physiologically compatible solvent. Specifically, by weight percentage, it comprises 12% methacrylamide polylysine, 5% oxidized dextran, 10% Viola yedoensis exosomes, 0.25% photoinitiator, and 72.75% physiologically compatible solvent.

[0042] Example 2: As Figure 1 As shown, a method for preparing a composite hydrogel loaded with Viola yedoensis exosomes (PLMA / ODex@VE) includes the following steps:

[0043] S1. Preparation of PLMA: Weigh 5 g of ε-polylysine (ε-PL) and add it to 30 mL of phosphate buffered saline (PBS). Stir in a 50°C water bath for 1 h until completely dissolved. Add methacrylic anhydride (MA) dropwise to the ε-PL solution at a rate of 0.5 mL / min, with a volume ratio of 1:15. After the addition is complete, stir at 50°C for 6 h to obtain the PLMA solution. Dialyze the PLMA solution in a dialysis bag with a molecular weight cutoff of 3500 Da for 3 days. After freeze-drying, store it in a 4°C refrigerator for later use.

[0044] S2. Preparation of ODex: Dissolve 7.5 g of dextran in 100 mL of deionized water and heat to 90 °C with continuous stirring. After the solution becomes colorless and transparent, adjust the reaction temperature to 30 °C. Then, dissolve 7.5 g of NaIO4 in 25 mL of deionized water and add it dropwise to the dextran solution at room temperature in the dark. After reacting for 6 h, quickly add 25 mL of diethylene glycol to terminate the reaction. Dialyze the mixture with deionized water for 3 days. Finally, freeze and freeze-dry the solution for 48 h to obtain powdered ODex.

[0045] S3. Extraction of Vitamin E: Viola yedoensis was initially washed with tap water at room temperature, then rinsed three times with PBS solution. The washed Viola yedoensis was soaked in an appropriate amount of PBS, crushed in a high-speed blender for 3 minutes, filtered through a mesh screen to remove residue, and the juice was collected. At 4°C, the filtered Viola yedoensis juice was centrifuged at 3000 g for 30 minutes, and the supernatant was collected. The collected supernatant was then centrifuged at 5000 g for 40 minutes, and the supernatant was collected again. The collected supernatant was then centrifuged at 15000 g for 30 minutes. The supernatant was filtered through a 0.22 μm filter membrane to remove insoluble residue. The filtered supernatant was centrifuged at 150000 g for 90 minutes at 4°C to obtain a precipitate. The precipitate was resuspended in sterile PBS solution, and the pH was adjusted to neutral to obtain Viola yedoensis exosomes (VE), which were stored at -80°C. The structural characterization and particle size distribution of VE were determined using transmission electron microscopy (TEM) and a nanoparticle size and zeta potential analyzer (DLS), such as... Figure 2 and Figure 3 As shown, TEM reveals spherical nanoparticles with a unique cup-shaped morphology for VE, and nanoparticle size analysis shows that most VE have a diameter of approximately 131 nm.

[0046] S4. Preparation of precursor mixture: First, dissolve PLMA and ODex in PBS solution to prepare PLMA and ODex solutions of different concentrations; then mix the two in equal volumes, add appropriate concentrations of VE and photoinitiator (LAP) and mix evenly to prepare injectable precursor mixture.

[0047] S5. Photocuring: Inject or coat the precursor mixture into the target area and irradiate it for 10 to 60 seconds under light with a wavelength of 365~405 nm and a light intensity of 5~50 mW / cm² to form a stable solid PLMA / ODex@VE composite hydrogel in situ.

[0048] Example 3: Physicochemical characterization of PLMA / ODex@VE composite hydrogel

[0049] The hydrogels containing only PLMA and the hydrogels containing PLMA / ODex were characterized in comparison with the PLMA / ODex@VE composite hydrogel prepared in Example 2:

[0050] Microstructure: The surface morphology of different groups of hydrogels was observed using scanning electron microscopy (SEM), and porosity and pore size were analyzed. Figure 4 , 5 As shown, all groups of hydrogels exhibited typical cross-linked porous network structures with relatively uniform distribution. The porosity of each group of hydrogels was greater than 60%, indicating that the loading of VE on the hydrogel had minimal impact on its cross-linking density. The porous structure can promote the transport of oxygen and nutrients, rapidly absorb exudate, and keep the wound moist. Figure 6 As shown, the pore size of each group of hydrogels is greater than 100 μm, which is beneficial to cell proliferation and migration.

[0051] Example 4: Evaluation of in vitro biosafety and antibacterial properties

[0052] Cell safety test: Human umbilical vein endothelial cells (HUVECs) were co-cultured with the hydrogel material prepared in Example 2. After 1, 2 and 3 days of culture, cytotoxicity was detected by CCK-8 assay, and cell survival was observed by live and dead cell staining.

[0053] Hemolysis rate test: The experimental group was divided into three groups: PLMA group, PLMA / ODex group, and PLMA / ODex@VE group. 1 g of hydrogel was added to each test tube, followed by 5 mL of sodium chloride injection. 5 mL of sodium chloride injection was added to each tube of the negative control group. 5 mL of distilled water was added to each tube of the positive control group. Three tubes were prepared in parallel for each group.

[0054] Place all test tubes in a constant temperature water bath at (37±1)℃ for 30 min. Add diluted rabbit blood to each test tube at a ratio of 0.2 mL diluted rabbit blood to 5 mL test solution, and mix gently. After placing the test tubes at (37±1)℃ for 60 min, centrifuge at 3500 rpm for 5 min. Then, measure the absorbance of the supernatant using spectrophotometry at a wavelength of 540 nm, and calculate the hemolysis percentage based on the absorbance.

[0055] Bacterial co-culture: *Escherichia coli*, *Staphylococcus aureus*, and methicillin-resistant *Staphylococcus aureus* (MRSA) were inoculated into 24-well plates at a density of 1 × 10⁶ CFU per well, resulting in four groups: Control, PLMA, PLMA / ODex, and PLMA / ODex@VE. The Control group used PBS buffer. The bacteria and corresponding materials were co-incubated at 37°C for 12 h. After 12 h of incubation, the OD value of the bacterial suspension in each well was measured using a microplate reader. The bacterial suspension was then diluted 10³ times, and 100 μL was spread onto agar plates. After incubation at 37°C for 24 h, bacterial colonies in each group were observed and counted.

[0056] Fluorescent staining of live and dead bacteria: *S. aureus*, *E. coli*, and *MRSA* were inoculated into 24-well plates at a density of 1 × 10⁶ CFU per well, resulting in four groups: Control, PLMA, PLMA / ODex, and PLMA / ODex@VE. The Control group was incubated with PBS buffer. The bacteria and corresponding materials were co-incubated at 37°C for 12 h. After 12 h, the bacterial suspensions in each group were centrifuged at 3500 rpm for 5 min, the supernatant was discarded, and the suspensions were resuspended in the prepared live / dead bacterial staining working solution. The suspensions were incubated at room temperature in the dark for 15 min. Finally, 5 μL of the stained bacterial suspension was transferred to a glass slide, covered with a coverslip, and the images were observed and acquired using a fluorescence inverted microscope. Figure 7 As shown, compared with the Control group, the PLMA group, PLMA / ODex group, and PLMA / ODex@VE group all showed that most bacteria died (red fluorescence), and only a few bacteria survived (green fluorescence). In particular, the PLMA / ODex@VE group had the fewest surviving bacteria, indicating that the PLMA / ODex@VE composite hydrogel has strong antibacterial activity and comparable antibacterial performance against different bacteria.

[0057] Example 5: Evaluation of in vitro anti-inflammatory and antioxidant properties

[0058] Immunofluorescence staining (IF): Mouse RAW 264.7 cells were co-cultured for 24 h with the Control group, PLMA group, PLMA / ODex group, and PLMA / ODex@VE group from Example 4. Cells were then incubated for 24 h in complete medium containing 200 mg / mL LPS. After the incubation period, the cells were washed with PBS. Subsequently, 4% paraformaldehyde was added for fixation for 30 min. The paraformaldehyde was washed away with PBS, and the cells were permeabilized with 0.5% Triton X-100 for 30 min. The Triton X-100 was then discarded, and the cells were blocked with goat serum for 1 h. After blocking, inducible nitric oxide synthase (iNOS) and arginase (Arg-1) primary antibodies diluted with antigen dilution buffer were added, and the cells were incubated overnight at 4°C. The next day, the primary antibodies were recovered, and the cells were incubated with diluted secondary antibodies in a light-protected incubator for 30 min. The secondary antibodies were washed away with PBS, and DAPI-containing anti-quenching agent was added. The anti-inflammatory properties of the hydrogel were observed using an inverted fluorescence microscope, and the fluorescence intensity was analyzed. Figure 8 , 9 As shown, the PLMA / ODex@VE group exhibited the lowest fluorescence intensity for the pro-inflammatory factor iNOS, and its expression level was also significantly lower than that of other groups in the fluorescence intensity quantification analysis; for example... Figure 10 , 11 As shown, the PLMA / ODex@VE group exhibited the strongest fluorescence intensity against the anti-inflammatory factor Arg-1, and its expression level was also significantly higher than that of other groups in the fluorescence intensity quantitative analysis. These results indicate that the PLMA / ODex@VE composite hydrogel can effectively promote the polarization of the macrophage M2 phenotype and inhibit the transformation of the macrophage M1 phenotype, thus exerting significant anti-inflammatory activity.

[0059] Western blot (WB) assay: RAW264.7 cells were co-cultured with hydrogels of different groups under LPS induction for 24 h. Total protein was extracted from the cells, and the protein concentration was measured. The proteins were then subjected to SDS-PAGE electrophoresis, and the electrophoretic bands were transferred to a PVDF membrane. After washing and blocking, the membrane was incubated overnight at 4°C with an internal control and primary antibody against NF-κB-NLRP3 pathway-related proteins. The next day, after washing, the membrane was incubated at room temperature for 2 h with the corresponding secondary antibody. Finally, the protein bands were observed using ECL chemiluminescence assay, and the expression of related proteins was quantitatively analyzed.

[0060] ROS staining: HUVEC cells were seeded in 24-well plates and cultured overnight to allow cell adhesion. Different groups of materials were co-cultured with cells for 24 h, followed by induction with 3% H2O2 for 30 min. Afterward, cells were stained with ROS using a ROS staining kit for 30 min. ROS expression in the cells was then imaged under a fluorescence inverted microscope and quantitatively analyzed. Figure 12 , 13As shown, the ROS staining fluorescence intensity of the PLMA / ODex@VE group was significantly lower than that of the other groups, and the quantitative analysis of fluorescence intensity was also significantly lower than that of the other groups. The results indicate that the PLMA / ODex@VE composite hydrogel has significant antioxidant activity.

[0061] Example 6: Evaluation of in vivo wound repair efficacy

[0062] SD rats were randomly divided into four groups: Control group, PLMA group, PLMA / ODex group, and PLMA / ODex@VE group. After anesthesia, a 10 mm diameter wound was created on the rat's back, and Staphylococcus aureus was then applied to form an infected wound. Subsequently, the hydrogel prepared in Example 2 was applied to the infected wound, and the gross healing of the wound was observed at different time points (3, 7, 10, and 14 days).

[0063] In vivo antibacterial properties: On day 3, the scabs on the rat wounds were removed, and wound secretions were collected with sterile cotton swabs, diluted in physiological saline, and spread on agar plates. After incubation at 37°C for 24 h, bacterial colonies in each group were observed and counted.

[0064] Histological analysis of rat wounds: Rats were sacrificed at different time points (7 and 14 days), and tissue samples were collected and prepared into paraffin sections. Hematoxylin and eosin (H&E) staining was used to observe the inflammatory response near the injury, the degradation of the hydrogel, and wound healing. Masson staining was used to observe collagen growth and deposition in the wound. Immunohistochemical staining and immunofluorescence staining were used to observe the inflammatory response and the anti-infection effect of the hydrogel. Proteins were extracted from the wound tissue, and Western blotting was used to further verify the expression of inflammation-related proteins and analyze the in vivo anti-inflammatory mechanism. Figure 14 As shown in the H&E staining results, the PLMA / ODex@VE group had the smallest wound defect, while the control group showed a large number of neutrophil infiltrations, as well as severe inflammatory reactions and signs of tissue infection caused by *S. aureus*, on days 7 and 14. The PLMA / ODex@VE group had fewer inflammatory cells in its skin tissue. In the MASSON staining results, the PLMA / ODex@VE group showed the most collagen deposition on days 7 and 14, with a significantly higher density than the control group. These results indicate that the PLMA / ODex@VE composite hydrogel, while effectively fighting infection, can promote collagen fiber formation and accelerate the healing of infected wounds.

[0065] The above descriptions are merely embodiments of the present invention, and common knowledge regarding specific technical solutions or characteristics is not elaborated upon here. It should be noted that those skilled in the art can make various modifications and improvements without departing from the technical solutions of the present invention, and these should also be considered within the scope of protection of the present invention. These modifications and improvements will not affect the effectiveness of the implementation of the present invention or the practicality of the patent. The scope of protection claimed in this application should be determined by the content of its claims, and the specific embodiments described in the specification can be used to interpret the content of the claims.

Claims

1. A composite hydrogel loaded with Viola yedoensis exosomes, characterized in that, By weight percentage, it consists of 10-25% methacrylamide polylysine, 5% oxidized dextran, 1-20% Viola yedoensis exosomes, 0.25% photoinitiator, and 49.75-83.75% physiologically compatible solvent.

2. The composite hydrogel loaded with Viola yedoensis exosomes according to claim 1, characterized in that, By weight percentage, it consists of 12% methacrylamide polylysine, 5% oxidized dextran, 10% Viola yedoensis exosomes, 0.25% photoinitiator and 72.75% physiologically compatible solvent.

3. A composite hydrogel loaded with Viola yedoensis exosomes according to claim 1 or 2, characterized in that, The photoinitiator is lithium phenyl-2,4,6-trimethylbenzoylphosphinate.

4. A composite hydrogel loaded with Viola yedoensis exosomes according to claim 1 or 2, characterized in that, The physiologically compatible solvent is phosphate buffer.

5. The method for preparing a composite hydrogel loaded with Viola yedoensis exosomes according to claim 1, characterized in that, Includes the following steps: S1. Prepare a solution of methacrylamide polylysine and an oxidized dextran solution; S2. Extract exosomes from Viola yedoensis; S3. Mix the methacrylamide polylysine solution, oxidized dextran solution, photoinitiator and Viola yedoensis exosomes in a certain mass ratio to obtain a precursor mixture; S4. Place the precursor mixture prepared in step S3 into the target area and perform photocrosslinking to form a hydrogel.

6. The method for preparing a composite hydrogel loaded with Viola yedoensis exosomes according to claim 5, characterized in that, In S1, methacrylated polylysine is prepared by reacting ε-polylysine with methacrylic anhydride, and the methacrylated polylysine is dissolved in a physiologically compatible solvent to obtain a methacrylated polylysine solution; oxidized dextran is prepared by oxidizing dextran with sodium periodate, and the oxidized dextran is dissolved in a physiologically compatible solvent to obtain an oxidized dextran solution.

7. The method for preparing a composite hydrogel loaded with Viola yedoensis exosomes according to claim 5, characterized in that, In S2, Viola yedoensis exosomes were extracted from fresh Viola yedoensis whole plant by ultra-high speed centrifugation and resuspended in a physiologically compatible solvent.

8. The method for preparing a composite hydrogel loaded with Viola yedoensis exosomes according to claim 5, characterized in that, In the photocrosslinking of S4, the wavelength of the light is 365~405 nm, the light intensity is 5~50 mW / cm², and the irradiation time is 10~60 seconds.

9. The use of a composite hydrogel loaded with Viola yedoensis exosomes according to claim 1 or 2 in the preparation of medical devices or dressings for the prevention and / or treatment of infected wounds.

10. The application of the composite hydrogel loaded with Viola yedoensis exosomes according to claim 9 in the preparation of medical devices or dressings for the prevention and / or treatment of infected wounds, characterized in that, The infected wounds include burn wounds, traumatic open wounds, diabetic foot ulcers, or postoperative infected incisions.

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