Hydrophilicity-Graded Trilayered Nanofiber Patch With Keratin-Activated Hemostatic Interface for Sequential...

We have developed a trilayered nanofiber composite (KTP) via successive electrospinning, engineered with hydrophilicity gradients supporting rapid hemostasis and wound healing. The innermost hydrophilic keratin-PVA layer initiated clotting, middle TEOS-PVA coaxial layer enhanced hemostatic efficacy, and outer PVDF layer provided protective barrier. KTP showed robust mechanical strength, cyto- and hemocompatibility, and supported cell migration. It triggered both intrinsic and extrinsic coagulation pathways and enabled platelet- and RBC-independent plasma clotting. Consistent with this, KTP upregulates tissue factor (TF) and modulates factor XII (FXII) levels in contact with plasma, indicating coordinated activation of extrinsic and contact pathways. Fibrinogen adsorption and fibrin clot stabilization occurred within 2 min, surpassing commercial dressings. Molecular docking revealed keratin's binding with coagulation proteins, which supports its mechanistic role in blood clotting. FE-SEM confirmed platelet adhesion and RBC entrapment; immunofluorescence imaging showed strong CD62P and vinculin expression, indicating active platelet engagement. In vivo injury models demonstrated rapid and stable clot formation within 35-40 s. KTP also accelerated tissue repair fully closing wounds by day 10, with complete re-epithelialization and hair follicle regeneration by day 12. Overall, this functionally graded nanofiber patch demonstrates strong potential for rapid hemorrhage control and accelerated wound healing for trauma care.