pH-responsive bilayer hydrogel with sequential release of morin-based nanoparticles and bFGF for the treatment of the “ice and fire” wounds

When the skin suffers from burns or frostbite, its normal structure and function are severely damaged, leading to problems such as inflammation, oxidative stress and difficulties in vascularization, which result in difficult wound healing. Currently, the existing treatment options have limitations in efficacy. The pH value of the burns and frostbite (B/F) wound microenvironment shows a dynamic change: it is acidic in the initial stage, gradually alkalizes as the repair process progresses, and then returns to a slightly acidic state in the later stage. Based on this, we designed a pH-responsive bilayer hydrogel with sequential release of morin-based nanoparticles and basic fibroblast growth factor (bFGF) for the treatment of B/F. 4-hydroxyphenylboronic acid pinacol ester (PAPE) modified fucoidan (Fu), chitosan (Cs) and morin self-assembled to form nanoparticles (CFMNPs), which exhibited excellent pH sensitivity. Then, using methacrylated hyaluronic acid (MeHA) as the main component, taking advantage of the concentration difference and the property that MeHA is easily degraded in an acidic environment, a 0.8% MeHA lower layer hydrogel loaded with CFMNPs and a 2% MeHA upper layer hydrogel loaded with bFGF were prepared. In vitro experiments confirmed the pH responsiveness and cytocompatibility of the bilayer hydrogel. In vivo experiments on mouse B/F wounds verified that the bilayer hydrogel can achieve sequential release of active ingredients according to the pH changes of the B/F wound, effectively inhibit inflammation, resist oxidative stress, promote new blood vessel formation, and ultimately accelerate wound healing. This provides a highly potential and efficient new strategy for the treatment of B/F. In this study, a pH-responsive bilayer hydrogel was designed and fabricated by integrating an antioxidant flavonoid with a pro-angiogenic growth factor for the treatment of burn and frostbite (B/F) wounds. The bilayer hydrogel (bFGF-CFMN) comprises a lower methacrylated hyaluronic acid (MeHA, 0.8%) layer loaded with morin self-assembled nanoparticles (CFMNPs) and an upper MeHA (1.5%) layer loaded with basic fibroblast growth factor (bFGF). By leveraging the concentration dependent, pH-responsive degradation of MeHA and the pH-triggered release characteristics of CFMNPs, bFGF-CFMN bilayer hydrogel enables staged, precise modulation of the wound microenvironment. During the early acidic phase of wound healing, the lower layer degrades rapidly to release CFMNPs, which subsequently release morin as pH shifts toward alkaline, thereby suppressing inflammation and mitigating oxidative stress. As the wound progresses into the proliferative and remodeling phases, the upper layer gradually degrades under alkaline conditions, sustaining bFGF release to promote endothelial cell proliferation and migration, enhance angiogenesis, and support collagen deposition and re-epithelialization. Animal experiments have demonstrated that the bFGF-CFMN bilayer hydrogel can effectively suppress inflammation, resist oxidative stress, and facilitate the formation of new blood vessels, ultimately accelerating wound healing. In summary, the bFGF-CFMN bilayer hydrogel holds great potential for the treatment of B/F wounds.