Recombinant Keratin-Chitosan Cryogel Decorated with Gallic Acid-Reduced Silver Nanoparticles for Wound Healing

Nanan Miao,1,* Tao Jiang,1,* Yuanchao Li,1 Sihong Xue,1 Shilei Hao,2 Chunli Zhou,1 Yujie Gu,1 Ran Li,1 Bo Yu,1 Xiaoqu Duan,1 Wenchao Xu,1 Rupeng Wang,1 Lei Ran1 1Department of Rheumatology and Dermatology, The Second Affiliated Hospital of Army Medical University, Chongqing, People’s Republic of China; 2College of Bioengineering, University of Chongqing, Chongqing, People’s Republic of China*These authors contributed equally to this workCorrespondence: Rupeng Wang; Lei Ran, Department of Rheumatology and Dermatology, The Second Affiliated Hospital of Army Medical University, Chongqing, 400037, People’s Republic of China, Tel +86 23 68755739, Email [email protected]; [email protected]: Wound healing is a complex physiological process that can be roughly divided into four stages: hemostasis, inflammation, proliferation, and remodeling. Conventional wound dressings often fail to meet the diverse needs of these healing stages due to their limited functionality. Cryogels, however, possess several attractive properties, such as large, interconnected pores, good mechanical strength, and ease of modification, making them suitable for developing advanced dressings with multiple functions. In this study, we developed a multifunctional cryogel dressing, with biocompatible polysaccharides as the main component, designed to provide a breathable, moist, and antibacterial microenvironment for chronic infected wounds, thereby promoting wound healing.Methods: Recombinant keratin 31 (RK31) was combined with chitosan (CS) to produce a CS/RK31 cryogel, referred to as CK. Gallic acid-reduced silver nanoparticles (GA/Ag NPs) were incorporated as the active antibacterial component to create the CS/K31@GA/Ag cryogel, known as CKGA. The cryogel was characterized using scanning electron microscopy (SEM) and a universal testing machine, and its biocompatibility was assessed in vitro. The dynamic hemostatic performance of the cryogel was evaluated with a rat tail amputation bleeding model. Additionally, the antibacterial effects of the cryogel against Staphylococcus aureus and Escherichia coli were tested using agar diffusion assays and turbidimetry. The antioxidant capacity of the CKGA cryogel was also measured in vitro. Finally, the cryogel’s ability to promote wound healing was tested in an SD rat model of infected wounds.Results: Characterization results showed that the CKGA cryogel features an interpenetrating porous network structure and exhibits excellent mechanical properties, with a swelling rate of up to 1800%. Both in vitro and in vivo experiments confirmed that the cryogel has good biocompatibility, effectively absorbs exudates, and rapidly stops bleeding. The addition of GA/Ag NPs provided significant antibacterial effects, achieving an inhibition rate of over 99.9% against both S. aureus and E. coli. Furthermore, CKGA cryogels demonstrated a strong scavenging capacity for ROS in a dose-dependent manner. Studies using the SD rat infected wound model showed that the cryogel effectively inhibited bacterial proliferation on wound surfaces, reduced local tissue inflammation, and promoted the healing of infected wounds.Conclusion: The multifunctional cryogel, with its rapid hemostatic, antibacterial, and antioxidant properties, as well as its ability to promote cell proliferation, could be widely used as a wound dressing for the healing of bacterial infections. Keywords: nanocomposite cryogel, hemostasis, antioxidant, antibacterial, wound healing