Self-Healing Polymeric Puerarin Hydrogel Dressing Promotes Diabetic Wound Healing Through Synergistic Immunomodulation and Tissue-Regenerative Remodeling

Abstract

Chronic wound healing is a significant challenge in diabetes. Puerarin is an active compound extracted from the traditional Chinese medicine Pueraria lobata. Puerarin has been used in the treatment of diabetes and derives benefits from its antioxidant, anti-inflammatory, antibacterial, and pro-angiogenesis properties, but its efficacy is hampered by poor water solubility and bioavailability. In this study, we designed a polyvinyl alcohol (PVA)-borax-puerarin (BP) hydrogel system that self-assembled via boronic ester bonds. The BP hydrogel exhibited exceptional physical characteristics, including adaptability, injectability, plasticity, self-healing capabilities, and robust compressive strength, as well as good biocompatibility. In the chronic wound diabetic rats model, the BP hydrogel significantly accelerated wound healing, as evidenced by hematoxylin and eosin (HE) staining, as well as Masson and picrosirius red (PSR) staining. RNA-sequencing and multiple immunohistochemistry (mIHC) analyses revealed that the BP hydrogel exerts a therapeutic effect by modulating macrophage polarization, promoting angiogenesis, and regulating collagen remodeling. Our findings suggest that the BP hydrogel represents a promising wound dressing and holds great potential for clinical applications in acute and chronic wound management.

Keywords: biocompatibility; hydrogel; polymeric biomaterials; puerarin; tissue engineering; tissue regeneration; wound dressings; wound healing.

Figure 1

Schematic diagram of raw materials, production process, and reaction principle required for BP hydrogel. Note: (A) Raw materials for preparing puerarin–borax–PVA hydrogel; (B) The apparent characteristics of puerarin–borax solution and BP hydrogel; (C) SEM structure of PB–PVA hydrogel; (D) FTIR spectra of PVA, borax–PVA, borax–puerarin–PVA; (E) FTIR spectra of puerarin, borax, and puerarin–borax.

Figure 2

Evaluation of physical properties of BP hydrogel. Note: (A) G′ and G′′ of PB hydrogel at different concentrations under strain amplitude; (B) G′ and G′′ of BP hydrogel under alternate scanning; (C) Gravity adaptation of hydrogel (it can flow out of the syringe under the action of gravity), injectability and plasticity (various letters and patterns can be made from the injected hydrogel); (D) Self-adaptation and self-healing properties of hydrogels (the hydrogel pieces can approach each other through adaptability and re-fuse into a new hydrogel); (E) Ductility (the hydrogel can stretch the strip to adapt to the external pressure to better protect the wound).

Figure 3

Biocompatibility of BP hydrogel. Note: (A,B) Live and dead staining of L929 when co-culture with BP hydrogel; (C) CCK–8 test of BP hydrogel (n = 3); (D) Detection of BP hydrogel hemolysis test and comparison of hemolysis rate; (E) Photo of BP hydrogel inhibiting S.a.; (F) The comparison of S.a. count of BP hydrogel (n = 3); (G) The antibacterial rate of BP hydrogel (n = 3). * p < 0.05.

Figure 4

The effect of BP hydrogel for promoting the healing of chronic wound healing in diabetes. Note: (A) Design of the experiment; (B,C) Comparison pictures of wound healing at different time points; (D) The comparison of wound healing rate at Day 3 (n = 12); (E) The comparison of wound healing rate at Day 7 (n = 12); (F) The comparison of wound healing rate at Day 10 (n = 12); (G) The HE staining of wound tissue at Day 7 and 10; (H) The Masson staining of wound tissue at Day 7 and 10; (I–L) The comparison of wound widths and thickness at Day 7 and 10 (n = 3). * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, ns: no significance.

Figure 5

RNA–Seq analysis of BP hydrogel in promoting wound healing. Note: (A) The volcano diagram of DEGs between model and control group; (B) The volcano diagram of DEGs between BP and model group; (C) The volcano diagram of DEGs between P and model group; (D) The volcano diagram of DEGs between B and model group; (E) The GO enrichment terms about inflammatory of DEGs between BP and model group; (F) The GO enrichment terms about wound healing of DEGs between BP and model group; (G) The KEGG enrichment of DEGs between four groups; (H) The GSEA analysis of JAK–STAT signaling pathway between BP and model group; (I) The DEGs of JAK–STAT signaling pathway between four groups (n = 3). * p-adjust < 0.05, ** p-adjust < 0.01, *** p-adjust < 0.001, **** p-adjust < 0.0001, ns: no significance.

Figure 6

Investigation of the mechanisms underlying the wound healing promotion by BP hydrogel based on mIHC. Note: (A) The expression of CD206 and iNOS in wound area (n = 3); (B) The expression of VEGFA and CD31 in wound area (n = 3); (C) The expression of COL–1 and COL–3 in wound area (n = 3); (D) The IOD of CD206 and iNOS in wound area (n = 3); (E) The IOD of VEGFA and CD31 in wound area (n = 2–3); (F) The IOD of COL–1 and COL–3 in wound area (n = 3); (G) The mechanism diagram of BP hydrogel promoting wound healing. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, ns: no significance.