Feasibility of repairing skin defects by VEGF ₁₆₅ gene‐modified iPS‐HFSCs seeded on a 3D printed scaffold containing astragalus polysaccharide

Abstract

The preparation of biodegradable scaffolds loaded with cells and cytokine is a feature of tissue‐engineered skin. IPSCs‐based tissue‐engineered skin treatment for wound repair is worth exploring. Healthy human skin fibroblasts were collected and reprogrammed into iPSCs. After gene modification and induction, CK19 ⁺/Integrinβ1 ⁺/CD200 ⁺ VEGF ₁₆₅ gene‐modified iPS‐HFSCs ^GFP were obtained and identified by a combination of immunofluorescence and RT‐qPCR. Astragalus polysaccharide‐containing 3D printed degradable scaffolds were prepared and co‐cultured with VEGF ₁₆₅ gene‐modified iPS‐HFSCs ^GFP, and the biocompatibility and spatial structure of the tissue‐engineered skin was analysed by cell counting kit‐8 (CCK8) assay and scanning electron microscopy. Finally, the tissue‐engineered skin was transplanted onto the dorsal trauma of nude mice, and the effect of tissue‐engineered skin on the regenerative repair of total skin defects was evaluated by a combination of histology, immunohistochemistry, immunofluorescence, RT‐qPCR, and in vivo three‐dimensional reconstruction under two‐photon microscopy. CK19 ⁺/Integrinβ1 ⁺/CD200 ⁺ VEGF ₁₆₅ gene‐modified iPS‐HFSCs ^GFP, close to the morphology and phenotype of human‐derived hair follicle stem cells, were obtained. The surface of the prepared 3D printed degradable scaffold containing 200 μg/mL astragalus polysaccharide was enriched with honeycomb‐like meshwork, which was more conducive to the proliferation of the resulting cells. After tissue‐engineered skin transplantation, combined assays showed that it promoted early vascularization, collagen and hair follicle regeneration and accelerated wound repair. VEGF ₁₆₅ gene‐modified iPS‐HFSCs ^GFP compounded with 3D printed degradable scaffolds containing 200 μg/mL astragalus polysaccharide can directly and indirectly participate in vascular, collagen, and hair follicle regeneration in the skin, achieving more complete structural and functional skin regenerative repair.