Composite Grafts of Human Keratinocytes Grown on a Polyglactin Mesh-Cultured Fibroblast Dermal Substitute Function as a Bilayer Skin Replacement in Full-Thickness Wounds on Athymic Mice

Abstract

We have developed and tested in athymic mice a new, cultured, dermal-epidermal graft composed of two human cell types coupled with a biodegradable dermal scaffold. Cultured, proliferating human keratinocytes (HK) were applied to the surface of a living dermal tissue replacement that is composed of human fibroblasts cultured on a polyglactin mesh. After 4 to 6 days of coculture, proliferating HKs achieved confluency on the surface of the living dermal tissue replacement. Grafts were then transferred to full-thickness wounds on the dorsum of athymic mice. Sixteen animals were grafted, and the mean percentage of graft take (original wound area covered) on day 20 after grafting was 51.25%. Staining with antibody specific for human involucrin confirmed the presence of HKs on closed wounds, and staining with antibody specific for human laminin revealed a continuous layer of laminin at the dermal-epidermal junction on day 20. Animals closed with living dermal tissue replacement alone markedly contracted, whereas application of living dermal tissue replacement-HK grafts appeared to retard contraction. Because polyglactin mesh fibers are absorbed by hydrolysis rather than by enzymatic degradation, this living composite graft may be more resistant to destruction when placed on excised human wounds than are composite grafts, which are composed of a collagen matrix. The inclusion of the living dermal substitute may ultimately provide better skin quality than is achieved from the use of cultured keratinocytes alone. Fragility of the epidermal layer is probably due to the short-term culture of HKs on the living dermal tissue replacement, and further efforts to develop a thicker epithelial layer may improve graft durability.