Direct comparison of a cultured composite skin substitute containing human keratinocytes and fibroblasts to an epidermal sheet graft containing human keratinocytes on athymic mice

Abstract

This study compares two techniques for making cultured skin substitutes: a composite graft made of human fibroblasts and keratinocytes on a collagen-glycosaminoglycan membrane (CG) and a cultured epidermal cell sheet graft (CEG), without a dermal component. The "take" and quality of these cultured skin substitutes were evaluated by placing them on full-thickness, excised wounds of athymic mice. These cultured skin substitutes were placed onto 2-X-2-cm wounds created on athymic mice. Mice were sacrificed at days 10, 20, and 42 with histologic sections obtained for light, electron, immunofluorescent, and immunohistochemical microscopy. "Take" was determined separately by a direct immunofluorescent stain for human leukocyte ABC antigens. There were ten mice of each graft type with at least two animals sacrificed at each time point. Results showed positive "take" for all animals. Grossly, there was little difference between the two graft types, with the CEG having occasional blister formation. By light microscopy, the CEG had a dissociation of dermis from epidermis until day 42, which was never apparent with the CG. By day 42, the CG had increased dermoepidermal interdigitations similar to rete ridges, with a mature epithelium. Neither of these findings were seen with the CEG. Immunofluorescent and immunohistochemical microscopy for type IV collagen and laminin, as well as electron microscopy, showed similar retardation of basement membrane formation with the CEG. Using this model, the composite graft had significant advantages over the epidermal sheet graft in the closure of full-thickness wounds.