Hydrogel for promoting healing of diabetic wound as well as preparation method and application of hydrogel

Abstract

The invention relates to hydrogel for promoting healing of diabetic wounds as well as a preparation method and application of the hydrogel, and belongs to the technical field of hydrogel dressings. The invention discloses an exosome hydrogel, which is characterized in that exosome is loaded in crosslinked aldehyde hyaluronic acid and carboxymethyl chitosan, the aldehyde hyaluronic acid is grafted with L-arginine, and the exosome is derived from bone marrow mesenchymal stem cells. L-arginine can release nitric oxide under the action of a diabetic wound surface or an inflammation microenvironment to achieve the effects of resisting bacteria and promoting wound surface recovery, and the exosome successfully avoids the risks of cell survival defects, vascular embolism, immunological rejection and the like while retaining powerful bioactive components. The product has excellent antibacterial activity, degradability, tissue adhesion, injectable performance and self-repairing performance, and can be suitable for wounds in different shapes.

Description

Technical Field

[0001] This invention belongs to the field of hydrogel dressing technology, specifically relating to a hydrogel that promotes the healing of diabetic wounds, its preparation method, and its application. Background Technology

[0002] The information disclosed in this background section is intended only to enhance understanding of the overall background of the invention and is not necessarily to be construed as an admission or in any way implying that such information constitutes prior art known to those skilled in the art.

[0003] Diabetic foot ulcer (DFU) is a devastating complication of diabetes, characterized by recurrent infections, impaired angiogenesis, and chronic inflammation. These wounds are not only difficult to heal, manage, and costly to treat, but also have high recurrence rates and high rates of disability and mortality, imposing a heavy psychological and physical burden on patients. Globally, approximately 15% of diabetic patients suffer from DFU, leading to over 80% of non-traumatic lower limb amputations, resulting in high mortality rates and posing a significant public health challenge.

[0004] The main reason for the difficulty in healing DFU wounds lies in their complex microenvironment, which contains a triple barrier. First, the excessive proliferation of pathogenic microorganisms (especially Staphylococcus aureus and Escherichia coli colonizing the biofilm) constitutes the primary treatment barrier. These structured microbial communities significantly weaken treatment efficacy and increase the risk of serious infection by forming physical barriers, enzymatically degrading drugs, and developing inherent drug resistance through metabolic dormancy. Second, abnormally elevated levels of pro-inflammatory cytokines drive persistent chronic inflammation, disrupting the normal wound healing cascade and delaying epithelialization and granulation tissue formation. Finally, endothelial dysfunction caused by diabetes fundamentally damages angiogenesis signaling pathways, leading to insufficient tissue perfusion and chronic hypoxia in the wound. This angiogenesis disorder, intertwined with bacterial colonization and persistent inflammation from the first two barriers, significantly exacerbates the pathological complexity of the wound, forming a vicious cycle that hinders wound closure and exacerbates tissue damage.

[0005] Currently, the core treatments for diabetic foot ulcers (DFU) encompass wound debridement (surgery / enzymatic hydrolysis / autolysis, etc.) to remove necrotic tissue, targeted anti-infective therapy (local / systemic antibiotics, osteomyelitis treatment), strict wound decompression (e.g., full-contact casts, pressure-relieving footwear, activity restriction) to eliminate mechanical pressure, moist environment dressing management (foam, hydrocolloid, silver-containing dressings, etc.) to promote granulation tissue growth and manage exudate, and intensive glycemic control as a foundation. However, these treatments generally have significant limitations: DFU has complex and diverse causes, with neuropathy, peripheral artery disease, and persistent hyperglycemia intertwined, resulting in limited effectiveness of single therapies and necessitating multidisciplinary collaboration. Deep infections (especially osteomyelitis) are difficult to diagnose, and biofilm formation and multidrug-resistant bacteria often significantly reduce the effectiveness of antibiotics. Current therapies such as debridement, antibiotics, and growth factor therapy often fail to effectively address the multifactorial pathogenesis of DFU, highlighting the urgent need for novel, multifunctional treatment strategies. Summary of the Invention

[0006] To address the shortcomings of existing technologies, the present invention aims to provide a hydrogel that promotes wound healing in diabetic patients, its preparation method, and its application. The invention involves grafting the natural nitric oxide donor molecule L-arginine onto the hyaluronic acid molecular backbone, further mixing it with carboxymethyl chitosan, and simultaneously encapsulating exosomes derived from bone marrow mesenchymal stem cells. This results in an injectable hydrogel with antibacterial, anti-inflammatory, and angiogenesis-promoting functions.

[0007] To achieve the above objectives, the technical solution of the present invention is as follows: In a first aspect, a hydrogel for promoting diabetic wound healing comprises cross-linked aldehyde-modified hyaluronic acid and carboxymethyl chitosan, wherein exosomes are loaded thereon; the aldehyde-modified hyaluronic acid is grafted with L-arginine, and the exosomes are derived from bone marrow mesenchymal stem cells.

[0008] Secondly, the preparation method of the aforementioned hydrogel for promoting diabetic wound healing includes: S1. L-arginine was mixed with aldehyde-modified hyaluronic acid and incubated to obtain L-arginine-grafted aldehyde-modified hyaluronic acid. S2. Load exosomes into a solution of L-arginine-grafted aldehyde-modified hyaluronic acid to obtain a mixture; S3. Mix the carboxymethyl chitosan solution with the mixture to obtain a hydrogel that promotes the healing of diabetic wounds.

[0009] Thirdly, the applications of the aforementioned hydrogels that promote wound healing in diabetic patients include: Application in the preparation of products that promote wound healing in diabetic patients.

[0010] The beneficial effects of this invention include: The L-arginine in this invention can release nitric oxide under the influence of diabetic wounds or inflammatory microenvironments, achieving antibacterial and wound-healing effects. Nitric oxide has a dual mechanism: it induces oxidative damage through the generation of peroxynitrite to achieve a broad-spectrum antibacterial effect; simultaneously, it promotes angiogenesis by enhancing endothelial cell proliferation and vascular endothelial growth factor expression. Bone marrow mesenchymal stem cells have the potential to overcome cell proliferation defects, angiogenesis disorders, and immune dysregulation; compared with stem cell transplantation, exosomes retain potent bioactive components while successfully avoiding risks such as cell survival defects, vascular embolism, and immune rejection. The exosome composite hydrogel prepared through Schiff base reaction has excellent antibacterial activity, degradability, tissue adhesion, injectability, and self-healing properties, making it suitable for wounds of different shapes. During DFU wound treatment, it can significantly accelerate wound healing by promoting granulation tissue formation and collagen deposition. Attached Figure Description

[0011] The accompanying drawings, which form part of this invention, are used to provide a further understanding of the invention. The illustrative embodiments of the invention and their descriptions are used to explain the invention and do not constitute an improper limitation of the invention.

[0012] Figure 1 These are morphological images of bone marrow mesenchymal stem cells in Example 1 under a phase contrast microscope; where (a) is a morphological image of bone marrow mesenchymal stem cells, and (b) is a magnified view of (a).

[0013] Figure 2 This is a flowchart of the exosome extraction method in Example 1.

[0014] Figure 3 These are morphological images of exosomes in Example 1 under a transmission electron microscope; where (a) is a morphological image of exosomes and (b) is a magnified view of a portion of (a).

[0015] Figure 4 These are photographs of the actual objects in Example 1; where (a) is a photograph of a double-barreled syringe, (b) is a photograph of the hydrogel product, and (c) is a photograph of the assembled fragments.

[0016] Figure 5 The second photograph is a physical photograph of Example 1; wherein, (a) is a photograph of the unstained hydrogel, (b) is a micrograph of the freeze-dried hydrogel, and (c) is a photograph of the dispersion of exosomes in the hydrogel.

[0017] Figure 6 The figure shows the results of the in vitro antibacterial test in Example 2; where (a) is the control group, i represents the colony of Escherichia coli, and ii represents the colony of Staphylococcus aureus; (b) is the experimental group, i represents the colony of Escherichia coli, and ii represents the colony of Staphylococcus aureus.

[0018] Figure 7 This is a graph showing the degradation performance test results in Example 2.

[0019] Figure 8 These are adhesion photographs of the hydrogel; (a) is an adhesion photograph on the surface of a tissue simulator in Example 2, (b) is an adhesion photograph under torsional force, and (c) is an adhesion photograph on the surface of a wound in Example 3.

[0020] Figure 9 These are wound photographs taken at different times during Example 3.

[0021] Figure 10 This is a fitting diagram of the wound shape in Example 3.

[0022] Figure 11 These are the results of staining wound tissue sections in Example 3; where (a) is a diagram of granulation tissue thickness and (b) is a diagram of collagen deposition results. Detailed Implementation

[0023] It should be noted that the following detailed descriptions are exemplary and intended to provide further illustration of the invention. Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

[0024] It should be noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the scope of exemplary embodiments according to the invention. As used herein, the singular form is intended to include the plural form as well, unless the context clearly indicates otherwise. Furthermore, it should be understood that when the terms "comprising" and / or "including" are used in this specification, they indicate the presence of features, steps, operations, devices, components, and / or combinations thereof.

[0025] Unless otherwise specified, the experimental methods described in the following examples are generally performed under standard conditions. All raw materials and reagents used in the following examples are commercially available unless otherwise indicated.

[0026] The unit "%w / v" refers to the number of grams of solute or dispersed phase contained in 100 mL of liquid.

[0027] One or more embodiments of the present invention provide a hydrogel for promoting diabetic wound healing, comprising cross-linked aldehyde-modified hyaluronic acid and carboxymethyl chitosan, wherein exosomes are loaded, the aldehyde-modified hyaluronic acid being grafted with L-arginine, and the exosomes being derived from bone marrow mesenchymal stem cells.

[0028] L-arginine is used to release nitric oxide at the wound site, achieving antibacterial and wound healing effects. Exosomes are used to overcome the challenges of cell proliferation defects, angiogenesis disorders, and immune dysregulation. The exosomes can interact extensively with the immune system, adjusting the pro-inflammatory state to an anti-inflammatory and repair state, and promoting cell proliferation and angiogenesis. The hydrogel generated by the Schiff base reaction of aldehyde-modified hyaluronic acid and carboxymethyl chitosan has excellent tissue adhesion, injectability, and self-healing properties. As a wound dressing, it can sustainably release exosomes and nitric oxide at the wound site, achieving the multimodal therapeutic effect of DFU.

[0029] Optionally, the degree of aldehyde modification of the aldehyde-modified hyaluronic acid is 15-25%. Adding aldehyde groups to the hyaluronic acid makes it an aldehyde donor in the Schiff base reaction, which crosslinks carboxymethyl chitosan into a three-dimensional network structure, giving it tissue adhesion, injectability, and self-healing properties.

[0030] Optionally, the exosomes are derived from bone marrow mesenchymal stem cells and can interact extensively with the immune system, modulating a pro-inflammatory state to an anti-inflammatory and repair state, and promoting cell proliferation and angiogenesis.

[0031] Optionally, the grafting rate of L-arginine is 10-20%. L-arginine can react with the high level of H2O2 in diabetic wounds or inflammatory areas to continuously release nitric oxide.

[0032] Optionally, the mass ratio of the L-arginine-grafted aldehyde-modified hyaluronic acid, carboxymethyl chitosan, and exosomes is (1.5~3):(1.5~3):(0.005~0.02).

[0033] Optionally, the carboxymethyl chitosan is carboxymethyl chitosan; the Schiff base reaction is a reaction between an aldehyde donor and a primary amino donor. Carboxymethyl chitosan, as a crosslinking matrix, can provide a primary amino group, which reacts with the aldehyde group provided by aldehyde-modified hyaluronic acid to form a Schiff base containing an imine group (C=N).

[0034] One or more embodiments of the present invention provide a method for preparing the above-mentioned hydrogel for promoting diabetic wound healing, comprising the steps of: S1. L-arginine was mixed with aldehyde-modified hyaluronic acid and incubated to obtain L-arginine-grafted aldehyde-modified hyaluronic acid. S2. Load exosomes into a solution of L-arginine-grafted aldehyde-modified hyaluronic acid to obtain a mixture; S3. Mix the carboxymethyl chitosan solution and the mixture to obtain a hydrogel that promotes the healing of diabetic wounds.

[0035] Through the above process, L-arginine is first grafted onto aldehyde-modified hyaluronic acid, then exosomes are loaded into the solution of the grafted aldehyde-modified hyaluronic acid, and finally a hydrogel is generated through a Schiff base reaction. L-arginine can be uniformly distributed in the hydrogel, and the exosomes are uniformly encapsulated in the hydrogel, which can slowly release exosomes and nitric oxide at the wound site.

[0036] Optionally, in S1, L-arginine and aldehyde-modified hyaluronic acid are mixed in PBS buffer and incubated for 24-36 h to ensure that L-arginine is evenly distributed in the generated hydrogel during the subsequent Schiff base reaction.

[0037] Optionally, in S2, the loading method involves adding exosomes to an L-arginine-grafted aldehyde-modified hyaluronic acid solution and vortexing to mix.

[0038] Optionally, in step S2, the concentration of L-arginine-grafted aldehyde-modified hyaluronic acid in the mixture is 3-6% w / v (containing 3-6 g of L-arginine-grafted aldehyde-modified hyaluronic acid per 100 mL of the mixture), and the concentration of exosomes is 0.01-0.04% w / v (containing 0.01-0.04 g of exosomes per 100 mL of the mixture), which serves as the reaction raw material for step S3 to generate a hydrogel with tissue adhesion, injectability, and self-healing properties.

[0039] Optionally, in S3, the concentration of the carboxymethyl chitosan solution is 3-6% w / v (containing 3-6 g of carboxymethyl chitosan per 100 mL of solution).

[0040] Optionally, in S3, the volume ratio of the carboxymethyl chitosan solution to the mixture is 1:(0.5~1.5).

[0041] One or more embodiments of the present invention provide the application of the above-described hydrogel for promoting diabetic wound healing, including its application in the preparation of products for promoting diabetic wound healing.

[0042] The present invention will be further described below with reference to specific embodiments.

[0043] Example 1 A hydrogel that promotes wound healing in diabetic patients includes L-arginine-grafted aldehyde-modified hyaluronic acid, carboxymethyl chitosan, and exosomes derived from bone marrow mesenchymal stem cells.

[0044] Preparation methods include: S0. Under light-protected conditions, a hyaluronic acid solution with a molecular weight of 200-400 kDa was reacted with sodium periodate for 12 hours to generate aldehyde-modified hyaluronic acid with a smaller molecular weight. Then, an excess of 10% v / v ethylene glycol solution was added to the reaction system to quench the reaction, and a mixture containing aldehyde-modified hyaluronic acid was obtained. The mixture was then dialyzed to obtain an aqueous solution of aldehyde-modified hyaluronic acid, which was then lyophilized for later use. The degree of aldehyde modification was approximately 20%.

[0045] The degree of aldehyde oxidation, also known as the degree of oxidation, refers to the average number of aldehyde groups introduced into each sugar unit (or disaccharide unit) in an oxidized polysaccharide molecule. It is calculated as: (number of moles of aldehyde groups / number of moles of sugar units) × 100%.

[0046] S1. Mix L-arginine and aldehyde-modified hyaluronic acid in PBS buffer and react for 24 h. The reaction product is obtained by dialysis, which is the L-arginine-grafted aldehyde-modified hyaluronic acid solution. It is then lyophilized for later use. The grafting rate is about 10%.

[0047] The grafting rate refers to the ratio of the mass of L-arginine successfully grafted onto the hyaluronic acid backbone to the mass of aldehyde-modified hyaluronic acid.

[0048] S2. Bone marrow mesenchymal stem cells were cultured in exosome-free medium for 48 h to avoid exogenous exosome contamination. Cell morphology under a phase-contrast microscope was as follows: Figure 1 As shown, Figure 1 Image (a) in the image is a morphological diagram of bone marrow mesenchymal stem cells. Figure 1 Image (b) is a magnified view of a portion of the sample. The cell suspension was then centrifuged at 10,000 g for 30 min at 4°C, and the supernatant was collected to remove cells. The collected supernatant was then centrifuged at 100,000 g for 70 min at 4°C, and the supernatant was discarded to remove cell debris. The collected precipitate was the exosome, and at this point, the exosome concentration was 100%, which was then used in subsequent mixing steps. The number of particles per milligram of exosome was approximately 1 × 10⁻⁶. 11 Each exocrine sample was resuspended in PBS buffer and stored at -80°C for later use. The appropriate processing method is as follows: Figure 2 As shown; the morphology of the resuspended exosomes is as follows Figure 3 As shown, Figure 3 (a) in the image is a morphological diagram of exosomes. Figure 3 (b) in the image is a magnified view of (a), with a diameter of approximately 100 nm.

[0049] Exosomes were added to a solution of L-arginine-grafted aldehyde-modified hyaluronic acid and then vortexed thoroughly to obtain a mixture, wherein the concentration of L-arginine-grafted aldehyde-modified hyaluronic acid was 4% w / v and the concentration of exosomes was 0.02% w / v.

[0050] S3. Take carboxymethyl chitosan with a molecular weight of 20-30 kDa and prepare a 4% w / v solution. Take the mixture obtained in S2 and fill the two liquids separately into containers as shown in the figure. Figure 4 In the double-barreled syringe shown in (a), the two liquids were mixed in a 1:1 volume ratio to prepare a hydrogel that promotes the healing of diabetic wounds.

[0051] like Figure 4 As shown in (b), mixing the two liquids in a double-barreled syringe and injecting them into molds of different shapes can produce hydrogel products of various shapes, indicating that the obtained hydrogel that promotes the healing of diabetic wounds can effectively adhere to the wound.

[0052] like Figure 4 As shown in (c), the red hydrogel product stained with Rhodamine B and the transparent hydrogel product not stained with Rhodamine B were cut into fragments and spliced ​​together. The fragments could be combined into one, indicating that it has a self-healing function.

[0053] The obtained unstained hydrogel products, such as Figure 5 As shown in (a) above; the results of microscopic observation after freeze-drying are as follows. Figure 5 As shown in (b) above, it exhibits a loose, porous structure; and as Figure 5 As shown in (c), the exosomes indicated by the white arrow are uniformly dispersed in the hydrogel.

[0054] The above results demonstrate that the hydrogel product successfully prepared in this embodiment possesses tissue adhesion, injectability, and self-healing properties, making it suitable for wounds of different shapes.

[0055] Example 2 The in vitro antibacterial performance test method includes: culturing Escherichia coli and Staphylococcus aureus to the logarithmic growth phase, and adjusting the bacterial suspension concentration to 1×10⁻⁶. 6 CFU / mL was co-cultured with the hydrogel or PBS buffer prepared in Example 1 at 37°C for 24 h. After incubation, the bacterial suspension was spread onto the surface of a solid culture medium using a tenfold serial dilution method, and incubated at 37°C for 24 h before colony counting. The results are as follows: Figure 6 As shown, Figure 6 (a) in the text refers to the culture results in PBS buffer. Figure 6 (b) shows the culture results in the hydrogel prepared in Example 1; where i represents the colony of Escherichia coli and ii represents the colony of Staphylococcus aureus; it can be seen that the hydrogel in Example 1 exhibits significant in vitro antibacterial activity and can effectively inhibit the proliferation of Escherichia coli and Staphylococcus aureus.

[0056] The degradation performance test method includes: immersing 500 mg of hydrogel in excess PBS buffer at 37℃; every other day, removing the hydrogel, gently wiping off the surface liquid with filter paper, and weighing it; obtaining the degradation rate change curve over time as shown in the figure. Figure 7 As shown, it has excellent biodegradability, with a degradation rate of over 90% within 9 days.

[0057] The adhesion performance testing method includes: preparing the hydrogel from Example 1 on the surface of a tissue simulator, which exhibits strong adhesion performance, such as... Figure 8 As shown in (a) above; the hydrogel does not detach under torsional force, as... Figure 8 As shown in (b) of the diagram.

[0058] Example 3 The application of hydrogels that promote wound healing in diabetic patients in DFU treatment includes the following methods: Diabetes was induced in 8-10 week old male C57BL / 6 mice by intraperitoneal injection of streptozotocin (50 mg / kg) for 5 consecutive days; a diabetes model was considered successfully established when the blood glucose level reached 16 mmol / L. Skin wounds with a diameter of 8 mm were created on the dorsal side of the mice. In the experimental group, the wounds were treated with 100 μL of the hydrogel prepared in Example 1 every 3 days; in the control group, the wounds were treated with 100 μL of PBS buffer every 3 days. Figure 8 As shown in (c), the hydrogel in the experimental group adhered firmly to the skin surface and formed a film at wound sites of different shapes.

[0059] Mice in each group were fed continuously for 14 days. Wound photographs on day 0, day 7, and day 14 are shown below. Figure 9 As shown; the wound shape fitting results are as follows Figure 10 As shown, on day 14, the experimental group showed significantly better treatment results compared to the control group, with wound size reduced by more than 90%.

[0060] The obtained wound tissue was then fixed in 4% paraformaldehyde and embedded in paraffin to prepare 4-micron sections. Granulation tissue thickness was assessed using hematoxylin and eosin (H&E) staining, and the results are as follows: Figure 11 As shown in (a), Masson staining was used to quantitatively analyze collagen deposition, and the results are as follows. Figure 11 As shown in (b), the hydrogel prepared in Example 1 can be seen to significantly accelerate wound healing by promoting granulation tissue formation and collagen deposition. Hyaluronic acid hydrogels are frequently used in wound dressings due to their excellent biocompatibility and ease of availability. However, their inherent antibacterial and anti-biofilm capabilities are insufficient, making it difficult to effectively combat complex deep infections and drug-resistant bacteria. Furthermore, conventional hyaluronic acid hydrogels struggle to intelligently and dynamically adapt to varying wound environments (such as fluctuations in exudate volume, pH, and enzyme activity), often exhibiting poor performance in adhesion, degradation rate, and exudation management. In addition, the relatively simple properties of conventional hyaluronic acid hydrogels, coupled with the complex processes involved in DFU healing (diuretic oxidase), such as inflammatory response control, cardiovascular formation, and tissue regeneration, place higher demands on dressings. These limitations severely restrict the clinical application and promotion of hyaluronic acid hydrogels. Nitric oxide possesses a dual therapeutic mechanism; however, its clinical translation faces multiple constraints: a short half-life, dose-dependent systemic toxicity, and poor biodistribution efficacy. In this specific embodiment, the L-arginine in the hydrogel can generate nitric oxide under the influence of high levels of H2O2 and other inflammatory factors in diabetic wounds. This allows for precise delivery of nitric oxide to the target tissue and spatiotemporally controlled release. Compared to direct stem cell transplantation, exosomes retain potent bioactive components while successfully avoiding risks such as cell survival defects, vascular embolism, and immune rejection. By using aldehyde-modified hyaluronic acid and carboxymethyl chitosan with set molecular weights, a hydrogel with tissue adhesion, injectability, and self-healing properties is prepared, providing a comprehensive therapy that integrates effective antibacterial, precise anti-inflammatory regulation, and highly efficient regenerative capabilities.

[0061] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention. Various modifications and variations can be made to the present invention by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. A hydrogel that promotes wound healing in diabetic patients, characterized in that, It includes cross-linked aldehyde-modified hyaluronic acid and carboxymethyl chitosan, wherein exosomes are loaded, the aldehyde-modified hyaluronic acid is grafted with L-arginine, and the exosomes are derived from bone marrow mesenchymal stem cells.

2. The hydrogel for promoting diabetic wound healing as described in claim 1, characterized in that, The degree of aldehyde oxidation of the aldehyde-modified hyaluronic acid is 15-25%.

3. The hydrogel for promoting diabetic wound healing as described in claim 1, characterized in that, The grafting rate of L-arginine is 10-20%.

4. The hydrogel for promoting diabetic wound healing as described in claim 1, characterized in that, The mass ratio of the L-arginine-grafted aldehyde-modified hyaluronic acid, carboxymethyl chitosan, and exosomes is (1.5~3):(1.5~3):(0.005~0.02).

5. A method for preparing a hydrogel for promoting diabetic wound healing as described in any one of claims 1-4, characterized in that, Including the following steps: S1. L-arginine was mixed with aldehyde-modified hyaluronic acid and incubated to obtain L-arginine-grafted aldehyde-modified hyaluronic acid. S2. Load exosomes into a solution of L-arginine-grafted aldehyde-modified hyaluronic acid to obtain a mixture; S3. Mix the carboxymethyl chitosan solution and the mixture to obtain a hydrogel that promotes the healing of diabetic wounds.

6. The method for preparing the hydrogel for promoting diabetic wound healing as described in claim 5, characterized in that, In S1, L-arginine and aldehyde-modified hyaluronic acid were mixed in PBS buffer and incubated for 24–36 h.

7. The method for preparing the hydrogel for promoting diabetic wound healing as described in claim 5, characterized in that, In S2, the loading method involves adding exosomes to an L-arginine-grafted aldehyde-modified hyaluronic acid solution and vortexing to mix.

8. The method for preparing the hydrogel for promoting diabetic wound healing as described in claim 5, characterized in that, In S2, the concentration of L-arginine-grafted aldehyde-modified hyaluronic acid in the mixture is 3-6% w / v, and the concentration of exosomes is 0.005-0.02% w / v.

9. The method for preparing the hydrogel for promoting diabetic wound healing as described in claim 5, characterized in that, In S3, the concentration of the carboxymethyl chitosan solution is 3-6% w / v; In S3, the volume ratio of the carboxymethyl chitosan solution to the mixture is 1:(0.5~1.5).

10. The application of a hydrogel for promoting diabetic wound healing as described in any one of claims 1-4, characterized in that, include: Application in the preparation of products that promote wound healing in diabetic patients.

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