Bilayered skin substitute incorporating rutin nanoparticles for antioxidant, anti-inflammatory, and anti-fibrotic effect

Hypertrophic scarring in large burns and delayed healing in chronic wounds are consequences of prolonged and aggravated inflammation, sustained infiltration of immune cells, free radical generation, and abundance of inflammatory mediators. Therefore, it is imperative to curb hyperinflammation to expedite wound healing. In this study, rutin nanoparticles (RNPs) were synthesized without an encapsulant and incorporated into eggshell membrane powder-crosslinked gelatin-chitosan cryogels to impart antioxidant and anti-inflammatory properties for treating hyperinflammation. The resultant nanoparticles were found to be 17.53 ± 4.03 nm in size and were stable at room temperature for a month with no visible sedimentation. RNPs were found to be non-cytotoxic and exhibited anti-inflammatory (by increasing IL-10 levels) and antioxidant properties (by controlling the generation of reactive oxygen species and enhancing catalase production in human macrophages). Additionally, RNPs were found to reduce α-SMA expression in fibroblasts, thereby demonstrating their anti-scarring effect. In vivo studies with a bilayered skin substitute constituting an RNP-incorporated cryogel proved that it is biocompatible, does not induce renal toxicity, aids wound healing, and induces better re-epithelialization than the control groups at the initial stages. Thus, RNP-incorporated cryogels containing bilayered skin substitutes are an advanced and novel alternative to commercial dermo-epidermal substitutes that lack anti-inflammatory or anti-scarring properties.