Abstract: BACKGROUND: As a new type of dressing developed based on the theory of moist healing, hydrogel provides a suitable humidity environment for the wound surface, sufficient space and mechanical support for cell migration and tissue regeneration, and can also be used as a carrier of drugs, growth factors, cells, and nanoparticles. It is widely used in the biomedical field. Puerarin has antioxidant, anti-inflammatory, and angiogenic effects, but its lipid solubility and water solubility are poor, which limits its clinical application.
OBJECTIVE: To prepare a puerarin-loaded double-network hydrogel and evaluate its effects on cellular behavior and wound healing in animal models.
METHODS: (1) Puerarin solution (0.5 mg/mL) was added to carboxymethyl chitosan-phenylboronic acid solution to prepare single-network sodium alginate-furan/bismaleimide hydrogel and carboxymethyl chitosan-phenylboronic acid-epigallocatechin gallate hydrogel, respectively. The two single-network hydrogels were mixed according to different volume ratios of the raw materials to prepare double-network hydrogel loaded with puerarin. In the H0 hydrogel without puerarin, the volume ratio of sodium alginate-furan, carboxymethyl chitosan-phenylboronic acid, epigallocatechin gallate and maleimide-maleimide was 25:40:8:3. In the puerarin-loaded H1, H2, H3, H4, and H5 hydrogels, the volume ratios of sodium alginate-furan, carboxymethyl chitosan-phenylboronic acid (containing puerarin), epigallocatechin gallate and maleimide-maleimide were 25:30:6:3, 25:40:8:3, 25:50:10:3, 25:60:12:3, and 25:70:14:3, respectively. The micromorphology, mechanical properties, adhesion, swelling properties, degradation properties, and drug release properties of the hydrogels were characterized, and the inhibitory effect of the hydrogels on Escherichia coli and the effects of the hydrogels on the proliferation and migration of L929 cells were detected. (2) A circular full-thickness skin defect with a diameter of 1 cm on the back of 9 SD rats was made. Three wounds were made on each rat. The wounds of the control group (n=3) were injected with normal saline; the wounds of the H0 group (n=3) were injected with H0 hydrogel, and the wounds of the H3 group (n=3) were injected with H3 hydrogel. The wound healing was observed. The wounds were collected 3 and 7 days after the wound modeling. Hematoxylin-eosin staining, Masson staining, and CD31 immunohistochemical staining were performed respectively.
RESULTS AND CONCLUSION: (1) The double-network hydrogel loaded with puerarin could gel. Scanning electron microscopy showed that the hydrogel had dense micropores, a large number of pores, and a small pore size. It had injectability, tissue adhesion and mechanical properties. The double-network hydrogel loaded with puerarin had strong water absorption capacity, slow degradation rate, could maintain the stability of morphology and volume, was responsive to pH and glucose, and could stably release the puerarin loaded therein. The double-network hydrogel loaded with puerarin could promote the proliferation and migration of L929 cells and inhibit the growth and reproduction of Escherichia coli. (2) Compared with the control group and H0 group, the wound healing rate of the H3 group was faster, and the wound healing rate was as high as 70% 7 days after modeling. The results of hematoxylin-eosin staining, Masson staining, and CD31 immunohistochemical staining 7 days after modeling showed that the wounds of the H3 group had the most granulation tissue growth, collagen accumulation and angiogenesis compared with the control group and H0 group. (3) The results showed that the double-network hydrogel loaded with puerarin had good biocompatibility and drug release properties, which could promote the growth and migration of L929 cells and the healing of full-thickness skin defects in rats.

Key words: wound repair, three-dimensional porous structure, double-network hydrogel, puerarin, drug release, biological dressing, small molecules of traditional Chinese medicine;, click chemistry