Preparation method and application of hydrogel adhesive bandage for preventing recurrence and promoting tissue repair after melanoma operation

Abstract

The invention discloses a preparation method of a hydrogel adhesive bandage for preventing relapse after melanoma operation and promoting tissue repair, and the preparation method comprises the following steps: preparing mesoporous bioactive glass rich in cerium and neodymium through an organic template method, ammonifying the surface of the mesoporous bioactive glass, and grafting methotrexate on the bioactive surface through an amido bond. And dispersing the obtained nanoparticles in a solution containing sulfobetaine methacrylate, acrylamide and chitosan, adding ammonium persulfate, tetramethylethylenediamine and N, N '-methylene bisacrylamide, fully mixing, injecting into a glass mold separated by polytetrafluoroethylene, and incubating at 40-60 DEG C for 30 minutes to obtain the hydrogel adhesive bandage. The hydrogel adhesive bandage effectively combines the photo-thermal performance of neodymium, the cell proliferation promoting capacity of cerium and the targeting and cancer inhibition effects of methotrexate, targeted therapy of nanoparticles can be achieved, and multiple cancer cells are killed in a photo-thermal and chemotherapy combined mode.

Description

Technical Field

[0001] The invention relates to a method for preparing a hydrogel wound dressing for tissue repair after melanoma resection, and belongs to the technical field of biomaterials. Background Art

[0002] Skin melanoma has become a major health problem worldwide, especially in my country, where the incidence of melanoma is increasing rapidly, with high malignancy and mortality. In particular, acral lentiginous melanoma (ALM), which is prone to occur on the fingers and soles of the feet, is a common subtype with a very high incidence in my country. The gold standard for the treatment of skin melanoma is surgical resection and chemotherapy, but it is limited by the inability to completely eradicate the tumor; the rapid transformation of the tumor into vertical growth is highly invasive; and the side effect of slow tissue healing after surgical resection eventually leads to tumor recurrence. In addition, existing dressings have problems with poor comfort, intensive joint movement, and easy shedding when treating ALM, which makes management difficult.

[0003] Therefore, there is an urgent need to develop a material that can not only take into account different subtypes such as common melanoma and acral lentiginous melanoma, but also match the entire dynamic process of tumor elimination and tissue repair to advance the treatment of melanoma. Summary of the invention

[0004] The technical problem to be solved by the present invention is to provide a method for preparing a hydrogel wound dressing in view of the deficiencies of the above-mentioned prior art. The hydrogel wound dressing of the present invention is obtained by copolymerization of sulfobetaine methacrylate (SBMA), acrylamide (AM), chitosan (CYCS) and nanoparticles, the nanoparticles are double-doped and surface-modified bioactive glass, the mesoporous bioactive glass is double-doped with cerium and neodymium ions to give it tissue repair and photothermal effects, and then the surface is aminated and grafted with the chemotherapy drug methotrexate, which has multiple cancer cell killing effects in the form of photothermal combined with chemotherapy.

[0005] The implementation process of the present invention is as follows: A method for preparing a hydrogel wound dressing for preventing recurrence of melanoma after surgery and promoting tissue repair comprises the following steps: (1) Preparation of methotrexate-modified bioactive glass (A) dissolving hexadecyltrimethylammonium bromide, anhydrous ethanol and ammonia water in deionized water to obtain a mixed solution, then sequentially adding triethyl phosphate, tetraethyl orthosilicate, calcium nitrate tetrahydrate, cerium nitrate hexahydrate and neodymium nitrate hexahydrate to react fully, collecting the precipitate and calcining it to obtain cerium and neodymium dual-doped mesoporous bioactive glass nanospheres; (B) dispersing the prepared cerium and neodymium dual-doped mesoporous bioactive glass nanospheres in anhydrous ethanol containing 3-aminopropyltriethoxysilane, reacting them fully, and collecting the precipitate to obtain surface-aminated mesoporous bioactive glass nanospheres; (C) dispersing the prepared surface aminated mesoporous bioactive glass nanospheres in deionized water containing methotrexate, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide, stirring thoroughly, dialyzing and freeze-drying to obtain methotrexate-grafted bioactive glass nanospheres; (2) Preparation of hydrogel wound dressings encapsulating bioactive glass modified with methotrexate [2-(Methacryloyloxy)ethyl]dimethyl-(3-sulfonatepropyl)ammonium hydroxide (SBMA), acrylamide (AM), and carboxylated chitosan (Cycs) were dissolved in deionized water, and methotrexate-grafted bioactive glass nanospheres, ammonium persulfate (APS), tetramethylethylenediamine (TMEDA) and N, N'-methylenebisacrylamide (MBAA) were added. After being fully mixed, the mixture was injected into a mold and incubated at 40-60°C to obtain a hydrogel wound dressing.

[0006] The above-mentioned hydrogel wound dressing preparation method, in step (A), the hexadecyltrimethylammonium bromide is dissolved in deionized water with a mass fraction of 2-5 wt%; the volume ratio of deionized water to anhydrous ethanol and ammonia water is 1: (0.5-0.8): (0.4-0.7); the volume ratio of ammonia water to triethyl phosphate and tetraethyl orthosilicate is 1: (0.3-0.6): (0.1-0.4); the molar ratio of triethyl phosphate to calcium nitrate tetrahydrate, cerium nitrate hexahydrate and neodymium nitrate hexahydrate is 1: (0.5-1.5): (0.05-0.2): (0.05-0.2); the concentration of ammonia water is 15-25%; the calcination temperature is 500-800°C, and the calcination time is 3-5 h.

[0007] In the above-mentioned method for preparing the hydrogel wound dressing, in step (B), the volume ratio of the 3-aminopropyltriethoxysilane to anhydrous ethanol is 1:(20-100); the dual-doped mesoporous bioactive glass nanospheres are dispersed in anhydrous ethanol to obtain a nanoparticle solution with a concentration of 0.1-0.7%.

[0008] In the above-mentioned method for preparing the hydrogel wound dressing, in step (C), the volume of the deionized water is 150 to 220 times the mass of the aminated mesoporous bioactive glass nanospheres; the mass ratio of the methotrexate to the aminated mesoporous bioactive glass nanospheres is 1:1.5 to 2.2; the molar ratio of methotrexate, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide is 1:(1 to 1.6):(1 to 2); the mass unit of the aminated mesoporous bioactive glass nanospheres is g, and the volume unit of the deionized water is mL.

[0009] In the above-mentioned method for preparing a hydrogel wound dressing, in step (2), the mass fraction of the [2-(methacryloyloxy)ethyl]dimethyl-(3-sulfonic acid propyl)ammonium hydroxide is 8-14 wt %; the mass fraction of acrylamide is 6-12 wt %; the mass fraction of carboxylated chitosan (Cycs) is 1-7 wt%; the mass fraction of the bioactive glass nanospheres modified by methotrexate grafting is 1-1.7wt%; the molar ratio of the [2-(methacryloyloxy)ethyl]dimethyl-(3-sulfonic acid propyl) ammonium hydroxide to ammonium persulfate (APS) and N,N'-methylenebisacrylamide (MBAA) is 1:(0.03-0.046):(0.004-0.025); the tetramethylethylenediamine (TMEDA) accounts for 0.1%-0.7% of the total volume; the mass unit of the [2-(methacryloyloxy)ethyl]dimethyl-(3-sulfonic acid propyl) ammonium hydroxide, ammonium persulfate, N,N'-methylenebisacrylamide and the bioactive glass nanospheres modified by methotrexate grafting is g, and the volume unit of the deionized water and tetramethylethylenediamine is mL.

[0010] The hydrogel wound dressing prepared by the above method is used in the preparation of a wound dressing for repairing tissue defects after melanoma surgery and inhibiting cancer recurrence.

[0011] Compared with the prior art, the present invention has the following advantages: (1) The hydrogel wound dressing of the present invention combines the angiogenesis-promoting ability of bioactive glass; the high biocompatibility of cerium and its ability to promote cell proliferation; the photothermal effect of neodymium; and the targeted chemotherapy properties of methotrexate. It can target chemotherapy and photothermal therapy to kill residual tumor cells; and when there is no melanoma, it does not release chemotherapy drugs but releases ions to promote wound healing, so it can kill cancer cells while promoting tissue repair; (2) The hydrogel wound dressing of the present invention is composed of SBMA, AM and Cycs with high biocompatibility, wherein Cycs provides antibacterial properties. The prepared hydrogel wound dressing can be strongly adhered to the skin surface in an ultra-thin state and adapt to joint movement; (3) The method for preparing the hydrogel wound dressing of the present invention has cheap and readily available raw materials, is environmentally friendly, is easy to scale up, and has good market application value. BRIEF DESCRIPTION OF THE DRAWINGS

[0012] Figure 1 This is a SEM image of the mesoporous bioactive glass nanospheres modified by methotrexate grafting in Example 1; Figure 2 TEM image of the mesoporous bioactive glass nanospheres modified by methotrexate grafting in Example 1; Figure 3 is a Fourier transform infrared spectrum (FT-IR) of the hydrogel wound dressing of Example 1; Figure 4 This is a SEM image of the hydrogel wound dressing of Example 1; Figure 5 This is a photothermal performance diagram of the hydrogel wound dressing of Example 1; Figure 6 This is a cytological experiment diagram of the hydrogel wound dressing of Example 1. DETAILED DESCRIPTION

[0013] Unless otherwise specified, the experimental methods used in the following examples are all conventional methods; the materials and reagents used, unless otherwise specified, can be obtained from commercial sources. The technical solution of the present invention is further described in detail below in conjunction with the accompanying drawings and embodiments. Example 1

[0014] This embodiment provides a method for preparing a hydrogel wound dressing for preventing recurrence of melanoma after surgery and promoting tissue repair, comprising: Step 1: providing methotrexate-modified bioactive glass, specifically comprising: Step 101, under the stirring condition of 500 rpm, dissolve 2 g of hexadecyltrimethylammonium bromide in 40 mL of deionized water, mix well, add 20 mL of anhydrous ethanol and 16 mL of ammonia water, stir for 10-30 min, then add 1.6 mL of triethyl phosphate, 4.8 mL of tetraethyl orthosilicate, 3.12 g of calcium nitrate tetrahydrate, 0.2 g of cerium nitrate hexahydrate and 0.2 g of neodymium nitrate hexahydrate in sequence, mix well for 3 h, wash three times with anhydrous ethanol by centrifugation, dry, and calcine at 800°C for 3 h to obtain dual-doped mesoporous bioactive glass nanospheres; Step 102, dispersing 0.4 g of the collected dual-doped mesoporous bioactive glass nanospheres in anhydrous ethanol containing 4 mL of 3-aminopropyltriethoxysilane dispersion, the volume of anhydrous ethanol being 400 mL, stirring and reacting for 24 hours, washing by centrifugation with anhydrous ethanol three times, and drying to obtain surface-ammonified mesoporous bioactive glass nanospheres; Step 103, dissolve 0.16 g of methotrexate, 0.068 g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 0.041 g of N-hydroxysuccinimide in 36 mL of deionized water, stir for 5 h, add 0.24 g of the aminated mesoporous bioactive glass nanospheres synthesized in step 3, stir well at room temperature for 24 h, and collect the methotrexate-grafted mesoporous bioactive glass nanospheres by dialysis (MWXO = 3500 Da) and freeze-drying.

[0015] Step 201, dissolve 0.4 g [2-(methacryloyloxy)ethyl]dimethyl-(3-sulfonic acid propyl)ammonium hydroxide (SBMA), 0.3 g acrylamide (AM), and 0.05 g carboxylated chitosan (Cycs) in 5 mL deionized water, and after they are fully dissolved, add 0.05 g methotrexate-grafted modified bioactive glass nanospheres, 0.01 g ammonium persulfate (APS), 0.005 mL tetramethylethylenediamine (TMEDA) and 0.001 g N, N'-methylenebisacrylamide (MBAA), and after they are fully mixed, inject them into a glass mold separated by polytetrafluoroethylene, and incubate at 40°C for 30 min to obtain a hydrogel wound dressing.

[0016] Test and performance evaluation results:

[0017] The SEM image of the mesoporous bioactive glass nanospheres modified by methotrexate grafting in Example 1 is as follows: Figure 1 As shown. Figure 1 It can be seen that the nanospheres are about 125 nm, and the surface of the spheres is rough due to the grafting of drugs.

[0018] The TEM image of the mesoporous bioactive glass nanospheres modified by methotrexate grafting in Example 1 is as follows: Figure 2 As shown in Figure 2, corresponding to the SEM image, the coated drug can be clearly observed on the surface of the sphere.

[0019] Example 1 Fourier transform infrared spectrum (FT-IR) of hydrogel wound dressing Figure 3 As shown in Figure 3, FT-IR shows that SBMA, AM and CYCS exist in the hydrogel and interact with each other through hydrogen bonds without forming chemical bonds.

[0020] Example 1 SEM image of hydrogel wound dressing Figure 4 As shown in the figure, it can be seen that the SAC surface has interconnected pores, and the pores are relatively smooth. When nanoparticles are incorporated into the hydrogel, its microstructure changes significantly, the pores are no longer smooth, and spheres are attached to the hydrogel surface, which proves the successful loading of nanoparticles.

[0021] Example 1 Photothermal performance of hydrogel wound dressing Figure 5 Under 808 nm laser irradiation, obvious temperature changes can be observed, and the temperature increases with the increase of nanoparticle concentration.

[0022] Example 1 Cytological Experiment of Hydrogel Wound Dressing Figure 6 As shown, it is based on the mesoporous bioactive glass nanosphere hydrogel wound dressing modified by methotrexate grafting as the research object to investigate its efficacy in inhibiting or killing cancer. The test method includes: (1) Preparation of 1640 extract of mesoporous bioactive glass and hydrogel wound dressing modified with methotrexate ①1640 cell complete culture medium: add 10% fetal bovine serum, 100 μg / ml streptomycin, and 100 U / ml penicillin to 500 mL of 1640 basal culture medium, mix well, and place in a 4 °C refrigerator for later use; ② Preparation of PBS solution: Weigh 0.01 g potassium chloride, 0.40 g sodium chloride, 0.01 g dipotassium hydrogen phosphate, and 0.17 g disodium hydrogen phosphate, respectively, add 1000 ml double distilled water, dissolve by ultrasonic, adjust the pH to 7.40, sterilize at 121°C for 30 min, and store in a refrigerator at 4°C for later use; ③ Preparation of 0.25% pancreatic enzyme: weigh 100 mg of pancreatic enzyme, add 40 ml PBS, filter with a 0.22 μm filter membrane, and store in a 4°C refrigerator for later use; ④ Preparation of 1640 extract of mesoporous bioactive glass and hydrogel wound dressing grafted with methotrexate: Weigh the grafted modified mesoporous bioactive glass and hydrogel wound dressing in Example 1 and soak them in 1640 complete cell culture medium. Place them in a 37°C incubator for three days, then filter them with a 0.22 μm filter membrane to obtain the extract of the grafted modified mesoporous bioactive glass. Seal the extract and place it in a 4°C refrigerator for use.

[0023] (2) Cell culture B16F10 cells were selected as model cells of cancer cells and cultured with the extracts of grafted modified mesoporous bioactive glass and hydrogel wound dressing, respectively, to investigate the effects of the extracts on the growth of model cells.

[0024] ①Cell passaging Transfer B16F10 cells into a culture dish containing 1640 culture medium and culture in a 37°C, 5% CO2 incubator for 12 h. Discard the culture medium, add 3 ml PBS to wash the cells, add 2 ml trypsin, and place in an incubator for digestion for 1.5 min. Take out the culture dish and add 3 ml culture medium to terminate digestion. Use a pipette to blow the cell culture medium several times, then transfer the culture medium containing B16F10 cells into 15 ml centrifuge tubes, centrifuge at 1000 rpm for 5 min, discard the supernatant, add 2 ml culture medium to each centrifuge tube, blow evenly, divide into two portions, add to new culture dishes containing 9 ml 1640 culture medium, shake crosswise to mix well, and place flat in a 37°C, 5% CO2 incubator for 24 h.

[0025] ②Cell layout Take the cells cultured in step ① out of the incubator, discard the culture medium, add 3 ml PBS to wash the cells, add 2 ml trypsin, and put them into the incubator for digestion for 1.5 min. After digestion is completed, blow the cell culture medium several times with a pipette, add 5-8 ml culture medium and blow the cells several times, then pipette 50 μl of each cell sample into the cell counting plate, count, dilute the cells according to a certain ratio, and then plate the cells in a 96-well plate. The cell density is about 1×10 4 The cells were placed in a 37°C, 5% CO2 incubator and cultured for another 24 h.

[0026] ③Cell culture in the extract of grafted modified mesoporous bioactive glass The 96-well plate containing cells cultured in ② was taken out, the culture medium was discarded, and the grafted modified mesoporous bioactive glass extract (experimental group), hydrogel wound dressing extract (experimental group) and an equal volume of 1640 culture medium (control group) prepared in this embodiment (1) were added thereto respectively, and after laser irradiation, they were placed in an incubator for culture, and 5 replicate wells were set for each concentration. After 24 hours of culture, 50 μL of MTT was added to each well, and after further culture for 2-4 hours, 150 μL of DMSO was added to each well, and the absorbance (OD) value corresponding to each well was detected at 490 nm using an ELISA reader, and the cell survival rate was calculated based on the OD value.

[0027] The experimental results showed that the viability of B16F10 cells was reduced after being treated with hydrogel bandage, while after 808nm laser irradiation (M 3%Nd / 3%Ce @M SAC + NIR), the cell survival rate of B16F10 was significantly reduced. This means that the combined effect of PTT and chemotherapy has a higher cancer cell killing effect, that is, the nanospheres have the ability to effectively inhibit and kill tumor cells.

[0028] Cytological experiments have shown that the hydrogel wound dressing prepared by the present invention for postoperative treatment of melanoma can effectively repair tissue defects after melanoma surgery and inhibit cancer recurrence. Example 2

[0029] This embodiment provides a method for preparing a hydrogel wound dressing for preventing recurrence of melanoma after surgery and promoting tissue repair, comprising: Step 1: providing methotrexate-modified bioactive glass, specifically comprising: Step 101, under the stirring condition of 550 rpm, dissolve 2.025 g of hexadecyltrimethylammonium bromide in 45 mL of deionized water, mix well, add 20 mL of anhydrous ethanol and 20.25 mL of ammonia water, stir for 10 to 30 min, then add 3.04 mL of triethyl phosphate, 7.1 mL of tetraethyl orthosilicate, 3.94 g of calcium nitrate tetrahydrate, 0.43 g of cerium nitrate hexahydrate and 0.43 g of neodymium nitrate hexahydrate in sequence, mix thoroughly for 2 h, wash three times with anhydrous ethanol by centrifugation, dry, and calcine at 750° C. for 3 h to obtain dual-doped mesoporous bioactive glass nanospheres; Step 102, dispersing 0.84 g of the collected dual-doped mesoporous bioactive glass nanospheres in anhydrous ethanol containing 10.5 mL of 3-aminopropyltriethoxysilane, the volume of anhydrous ethanol being 420 mL, stirring and reacting for 24 h, washing by centrifugation with anhydrous ethanol three times, and drying to obtain surface-ammonified mesoporous bioactive glass nanospheres; Step 103, dissolve 0.16 g of methotrexate, 0.074 g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 0.045 g of N-hydroxysuccinimide in 41 mL of deionized water, stir for 5 h, add 0.25 g of the aminated mesoporous bioactive glass nanospheres synthesized in step 3, stir well at room temperature for 24 h, and collect the methotrexate-grafted mesoporous bioactive glass nanospheres by dialysis (MWXO = 3500 Da) and freeze-drying.

[0030] Step 201, dissolve 0.45 g of [2-(methacryloyloxy)ethyl]dimethyl-(3-sulfonic acid propyl)ammonium hydroxide (SBMA), 0.35 g of acrylamide (AM), and 0.1 g of carboxylated chitosan (Cycs) in 5 mL of deionized water, and after they are fully dissolved, add 0.055 g of methotrexate-grafted bioactive glass nanospheres, 0.016 g of ammonium persulfate (APS), 0.008 mL of tetramethylethylenediamine (TMEDA) and 0.002 g of N, N'-methylenebisacrylamide (MBAA), and after they are fully mixed, inject them into a glass mold separated by polytetrafluoroethylene, and incubate at 40°C for 30 min to obtain a hydrogel wound dressing. Example 3

[0031] This embodiment provides a method for preparing a hydrogel wound dressing for preventing recurrence of melanoma after surgery and promoting tissue repair, comprising: Step 1: providing methotrexate-modified bioactive glass, specifically comprising: Step 101, under the stirring condition of 500 rpm, dissolve 2 g of hexadecyltrimethylammonium bromide in 50 mL of deionized water, mix well, add 30 mL of anhydrous ethanol and 25 mL of ammonia water, stir for 10 to 30 min, then add 5 mL of triethyl phosphate, 10 mL of tetraethyl orthosilicate, 5.83 g of calcium nitrate tetrahydrate, 0.83 g of cerium nitrate hexahydrate and 0.83 g of neodymium nitrate hexahydrate in sequence, mix well for 4 h, wash three times with anhydrous ethanol by centrifugation, dry, and calcine at 700° C. for 3 h to obtain dual-doped mesoporous bioactive glass nanospheres; Step 102, dispersing 1.32 g of the collected dual-doped mesoporous bioactive glass nanospheres in anhydrous ethanol containing 13.2 mL of 3-aminopropyltriethoxysilane, the volume of anhydrous ethanol being 440 mL, stirring and reacting for 24 h, washing by centrifugation with anhydrous ethanol three times, and drying to obtain surface-ammonified mesoporous bioactive glass nanospheres; Step 103, dissolve 0.16 g of methotrexate, 0.081 g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 0.049 g of N-hydroxysuccinimide in 46 mL of deionized water, stir for 5 h, add 0.27 g of the aminated mesoporous bioactive glass nanospheres synthesized in step 3, stir well at room temperature for 24 h, and collect the methotrexate-grafted mesoporous bioactive glass nanospheres by dialysis (MWXO = 3500 Da) and freeze-drying.

[0032] Step 201, dissolve 0.5 g of [2-(methacryloyloxy)ethyl]dimethyl-(3-sulfonic acid propyl)ammonium hydroxide (SBMA), 0.4 g of acrylamide (AM), and 0.15 g of carboxylated chitosan (Cycs) in 5 mL of deionized water, and after they are fully dissolved, add 0.06 g of methotrexate-grafted bioactive glass nanospheres, 0.018 g of ammonium persulfate (APS), 0.01 mL of tetramethylethylenediamine (TMEDA) and 0.003 g of N, N'-methylenebisacrylamide (MBAA), and after they are fully mixed, inject them into a glass mold separated by polytetrafluoroethylene, and incubate at 40°C for 30 min to obtain a hydrogel wound dressing. Example 4

[0033] This embodiment provides a method for preparing a hydrogel wound dressing for preventing recurrence of melanoma after surgery and promoting tissue repair, comprising: Step 1: providing methotrexate-modified bioactive glass, specifically comprising: Step 101, under the stirring condition of 300 rpm, dissolve 1.925 g of hexadecyltrimethylammonium bromide in 55 mL of deionized water, mix well, add 35.75 mL of anhydrous ethanol and 30.25 mL of ammonia water, stir for 10 to 30 min, add 7.6 mL of triethyl phosphate, 13.6 mL of tetraethyl orthosilicate, 7.84 g of calcium nitrate tetrahydrate, 1.44 g of cerium nitrate hexahydrate and 1.45 g of neodymium nitrate hexahydrate in sequence, mix well for 5 h, wash three times with anhydrous ethanol by centrifugation, dry, and calcine at 700° C. for 3 h to obtain dual-doped mesoporous bioactive glass nanospheres; Step 102, dispersing 1.84 g of the collected dual-doped mesoporous bioactive glass nanospheres in anhydrous ethanol containing 16.1 mL of 3-aminopropyltriethoxysilane, the volume of anhydrous ethanol being 460 mL, stirring and reacting for 24 h, washing by centrifugation with anhydrous ethanol three times, and drying to obtain surface-ammonified mesoporous bioactive glass nanospheres; Step 103, dissolve 0.16 g of methotrexate, 0.087 g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 0.058 g of N-hydroxysuccinimide in 52 mL of deionized water, stir for 5 h, add 0.29 g of the aminated mesoporous bioactive glass nanospheres synthesized in step 3, stir well at room temperature for 24 h, and collect the methotrexate-grafted mesoporous bioactive glass nanospheres by dialysis (MWXO = 3500 Da) and freeze-drying.

[0034] Step 201, dissolve 0.55 g of [2-(methacryloyloxy)ethyl]dimethyl-(3-sulfonic acid propyl)ammonium hydroxide (SBMA), 0.45 g of acrylamide (AM), and 0.2 g of carboxylated chitosan (Cycs) in 5 mL of deionized water, and after they are fully dissolved, add 0.065 g of methotrexate-grafted modified bioactive glass nanospheres, 0.02 g of ammonium persulfate (APS), 0.012 mL of tetramethylethylenediamine (TMEDA) and 0.004 g of N, N'-methylenebisacrylamide (MBAA), and after they are fully mixed, inject them into a glass mold separated by polytetrafluoroethylene, and incubate at 40°C for 30 min to obtain a hydrogel wound dressing. Example 5

[0035] This embodiment provides a method for preparing a hydrogel wound dressing for preventing recurrence of melanoma after surgery and promoting tissue repair, comprising: Step 1: providing methotrexate-modified bioactive glass, specifically comprising: Step 101, under the stirring condition of 350 rpm, dissolve 1.8 g of hexadecyltrimethylammonium bromide in 60 mL of deionized water, mix well, add 42 mL of anhydrous ethanol and 36 mL of ammonia water, stir for 10-30 min, then add 10.8 mL of triethyl phosphate, 18 mL of tetraethyl orthosilicate, 9.8 g of calcium nitrate tetrahydrate, 2.31 g of cerium nitrate hexahydrate and 2.33 g of neodymium nitrate hexahydrate in sequence, mix well for 6 h, wash three times with anhydrous ethanol by centrifugation, dry, and calcine at 600° C. for 3 h to obtain dual-doped mesoporous bioactive glass nanospheres; Step 102, dispersing 2.35 g of the collected dual-doped mesoporous bioactive glass nanospheres in anhydrous ethanol containing 18.8 mL of 3-aminopropyltriethoxysilane, the volume of anhydrous ethanol being 470 mL, stirring and reacting for 24 h, washing by centrifugation with anhydrous ethanol three times, and drying to obtain surface-ammonified mesoporous bioactive glass nanospheres; Step 103, dissolve 0.16 g of methotrexate, 0.095 g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 0.065 g of N-hydroxysuccinimide in 58 mL of deionized water, stir for 5 h, add 0.304 g of the aminated mesoporous bioactive glass nanospheres synthesized in step 3, stir well at room temperature for 24 h, and collect the methotrexate-grafted mesoporous bioactive glass nanospheres by dialysis (MWXO = 3500 Da) and freeze-drying.

[0036] Step 201, dissolve 0.6 g [2-(methacryloyloxy)ethyl]dimethyl-(3-sulfonic acid propyl)ammonium hydroxide (SBMA), 0.5 g acrylamide (AM), and 0.25 g carboxylated chitosan (Cycs) in 5 mL deionized water, and after they are fully dissolved, add 0.07 g methotrexate-grafted bioactive glass nanospheres, 0.022 g ammonium persulfate (APS), 0.014 mL tetramethylethylenediamine (TMEDA) and 0.005 g N, N'-methylenebisacrylamide (MBAA), and after they are fully mixed, inject them into a glass mold separated by polytetrafluoroethylene, and incubate at 40°C for 30 min to obtain a hydrogel wound dressing. Example 6

[0037] This embodiment provides a method for preparing a hydrogel wound dressing for preventing recurrence of melanoma after surgery and promoting tissue repair, comprising: Step 1: providing methotrexate-modified bioactive glass, specifically comprising: Step 101, under the stirring condition of 380 rpm, dissolve 1.625 g of hexadecyltrimethylammonium bromide in 65 mL of deionized water, mix well, add 48.75 mL of anhydrous ethanol and 42.25 mL of ammonia water, stir for 10 to 30 min, then add 14.78 mL of triethyl phosphate, 23.23 mL of tetraethyl orthosilicate, 11.5 g of calcium nitrate tetrahydrate, 3.52 g of cerium nitrate hexahydrate and 3.55 g of neodymium nitrate hexahydrate in sequence, mix well for 6 h, wash three times with anhydrous ethanol by centrifugation, dry, and calcine at 600° C. for 3 h to obtain dual-doped mesoporous bioactive glass nanospheres; Step 102, dispersing 2.88 g of the collected dual-doped mesoporous bioactive glass nanospheres in anhydrous ethanol containing 21.6 mL of 3-aminopropyltriethoxysilane, the volume of anhydrous ethanol being 480 mL, stirring and reacting for 24 h, washing by centrifugation with anhydrous ethanol three times, and drying to obtain surface-ammonified mesoporous bioactive glass nanospheres; Step 103, dissolve 0.16 g of methotrexate, 0.10 g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 0.073 g of N-hydroxysuccinimide in 64 mL of deionized water, stir for 5 h, add 0.32 g of the aminated mesoporous bioactive glass nanospheres synthesized in step 3, stir well at room temperature for 24 h, and collect the methotrexate-grafted mesoporous bioactive glass nanospheres by dialysis (MWXO = 3500 Da) and freeze-drying.

[0038] Step 201, dissolve 0.65 g of [2-(methacryloyloxy)ethyl]dimethyl-(3-sulfonic acid propyl)ammonium hydroxide (SBMA), 0.55 g of acrylamide (AM), and 0.3 g of carboxylated chitosan (Cycs) in 5 mL of deionized water, and after they are fully dissolved, add 0.075 g of methotrexate-grafted modified bioactive glass nanospheres, 0.024 g of ammonium persulfate (APS), 0.016 mL of tetramethylethylenediamine (TMEDA) and 0.006 g of N, N'-methylenebisacrylamide (MBAA), and after they are fully mixed, inject them into a glass mold separated by polytetrafluoroethylene, and incubate at 40°C for 30 min to obtain a hydrogel wound dressing. Example 7

[0039] This embodiment provides a method for preparing a hydrogel wound dressing for preventing recurrence of melanoma after surgery and promoting tissue repair, comprising: Step 1: providing methotrexate-modified bioactive glass, specifically comprising: Step 101, under the stirring condition of 400 rpm, dissolve 1.4 g of hexadecyltrimethylammonium bromide in 70 mL of deionized water, mix well, add 56 mL of anhydrous ethanol and 49 mL of ammonia water, stir for 10 to 30 min, then add 19.6 mL of triethyl phosphate, 29.4 mL of tetraethyl orthosilicate, 12.7 g of calcium nitrate tetrahydrate, 9 g of cerium nitrate hexahydrate and 9 g of neodymium nitrate hexahydrate in sequence, mix well for 6 h, wash three times by centrifugation with anhydrous ethanol, dry, and calcine at 500° C. for 5 h to obtain dual-doped mesoporous bioactive glass nanospheres; Step 102, dispersing 3.5 g of the collected dual-doped mesoporous bioactive glass nanospheres in anhydrous ethanol containing 25 mL of 3-aminopropyltriethoxysilane dispersion, the volume of anhydrous ethanol being 500 mL, stirring and reacting for 24 hours, washing by centrifugation with anhydrous ethanol three times, and drying to obtain surface-ammonified mesoporous bioactive glass nanospheres; Step 103, dissolve 0.16 g of methotrexate, 0.11 g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 0.076 g of N-hydroxysuccinimide in 74 mL of deionized water, stir for 5 h, add 0.34 g of the aminated mesoporous bioactive glass nanospheres synthesized in step 3, stir well at room temperature for 24 h, and collect the methotrexate-grafted mesoporous bioactive glass nanospheres by dialysis (MWXO = 3500 Da) and freeze-drying.

[0040] Step 201, dissolve 0.7 g [2-(methacryloyloxy)ethyl]dimethyl-(3-sulfonic acid propyl)ammonium hydroxide (SBMA), 0.6 g acrylamide (AM), and 0.35 g carboxylated chitosan (Cycs) in 5 mL deionized water, and after they are fully dissolved, add 0.08 g methotrexate-grafted modified bioactive glass nanospheres, 0.026 g ammonium persulfate (APS), 0.026 mL tetramethylethylenediamine (TMEDA) and 0.007 g N, N'-methylenebisacrylamide (MBAA), and after they are fully mixed, inject them into a glass mold separated by polytetrafluoroethylene, and incubate at 40°C for 30 min to obtain a hydrogel wound dressing.

[0041] The performance evaluation results of Examples 2-7 are similar to those of Example 1.

[0042] Finally, it should be noted that although the present invention has been described above in conjunction with the embodiments, the present invention is not limited to the above embodiments and application fields, and the above schemes are only for illustration and guidance purposes, rather than restrictive. Any simple modification, change and equivalent structural change made to the above embodiments by ordinary technicians in this field under the inspiration of this specification and without departing from the scope of protection of the claims of the present invention still falls within the protection scope of the technical solution of the present invention.

Claims

1. A method for preparing a hydrogel wound dressing for preventing recurrence of melanoma after surgery and promoting tissue repair, characterized in that The following steps are involved: (1) Preparation of methotrexate-modified bioactive glass (A) dissolving hexadecyltrimethylammonium bromide, anhydrous ethanol and ammonia water in deionized water to obtain a mixed solution, then sequentially adding triethyl phosphate, tetraethyl orthosilicate, calcium nitrate tetrahydrate, cerium nitrate hexahydrate and neodymium nitrate hexahydrate to react fully, collecting the precipitate and calcining it to obtain cerium and neodymium dual-doped mesoporous bioactive glass nanospheres; (B) dispersing the prepared cerium and neodymium dual-doped mesoporous bioactive glass nanospheres in anhydrous ethanol containing 3-aminopropyltriethoxysilane, reacting them fully, and collecting the precipitate to obtain surface-aminated mesoporous bioactive glass nanospheres; (C) dispersing the prepared surface aminated mesoporous bioactive glass nanospheres in deionized water containing methotrexate, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide, stirring thoroughly, dialyzing and freeze-drying to obtain methotrexate-grafted bioactive glass nanospheres; (2) Preparation of hydrogel wound dressings encapsulating bioactive glass modified with methotrexate [2-(Methacryloyloxy)ethyl]dimethyl-(3-sulfonatepropyl)ammonium hydroxide (SBMA), acrylamide (AM), and carboxylated chitosan (Cycs) were dissolved in deionized water, and methotrexate-grafted bioactive glass nanospheres, ammonium persulfate (APS), tetramethylethylenediamine (TMEDA) and N, N'-methylenebisacrylamide (MBAA) were added. After being fully mixed, the mixture was injected into a mold and incubated at 40-60°C to obtain a hydrogel wound dressing.

2. The method for preparing the hydrogel wound dressing according to claim 1, characterized in that: In step (A), the hexadecyltrimethylammonium bromide is dissolved in deionized water with a mass fraction of 2-5 wt%; the volume ratio of deionized water to anhydrous ethanol and ammonia water is 1: (0.5-0.8): (0.4-0.7); the volume ratio of ammonia water to triethyl phosphate and tetraethyl orthosilicate is 1: (0.3-0.6): (0.1-0.4); the molar ratio of triethyl phosphate to calcium nitrate tetrahydrate, cerium nitrate hexahydrate and neodymium nitrate hexahydrate is 1: (0.5-1.5): (0.05-0.2): (0.05-0.2), and the concentration of ammonia water is 15-25%.

3. The method for preparing the hydrogel wound dressing according to claim 1, characterized in that: In step (A), the calcination temperature is 500-800° C., and the calcination time is 3-5 h.

4. The method for preparing the hydrogel wound dressing according to claim 1, characterized in that: In step (B), the volume ratio of 3-aminopropyltriethoxysilane to anhydrous ethanol is 1:(20-100); the dual-doped mesoporous bioactive glass nanospheres are dispersed in anhydrous ethanol to obtain a nanoparticle solution with a concentration of 0.1-0.7%.

5. The method for preparing the hydrogel wound dressing according to claim 1, characterized in that: In step (C), the volume of deionized water is 150 to 220 times the mass of the aminated mesoporous bioactive glass nanospheres; the mass ratio of methotrexate to the aminated mesoporous bioactive glass nanospheres is 1:1.5 to 2.2; and the molar ratio of methotrexate, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide is 1:(1 to 1.6):(1 to 2).

6. The method for preparing the hydrogel wound dressing according to claim 1, characterized in that: In step (2), the mass fraction of the [2-(methacryloyloxy)ethyl]dimethyl-(3-sulfonic acid propyl) ammonium hydroxide is 8-14 wt %, the mass fraction of acrylamide is 6-12 wt %, the mass fraction of carboxylated chitosan (Cycs) is 1-7 wt %, and the mass fraction of the bioactive glass nanospheres grafted and modified by methotrexate is 1-1.7 wt %; the molar ratio of the [2-(methacryloyloxy)ethyl]dimethyl-(3-sulfonic acid propyl) ammonium hydroxide to ammonium persulfate (APS) and N,N'-methylenebisacrylamide (MBAA) is 1:(0.03-0.046):(0.004-0.025); and the tetramethylethylenediamine (TMEDA) accounts for 0.1%-0.7% of the total volume.

7. The hydrogel wound dressing prepared by the method according to any one of claims 1 to 6.

8. Use of the hydrogel wound dressing according to claim 7 in the preparation of medicines for repairing tissue defects after melanoma surgery and inhibiting cancer recurrence.

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