ULTRASTRUCTURAL FEATURES OF COMPOSITE SKIN CULTURES GRAFTED ONTO ATHYMIC MICE

Skin substitutes composed of cultured keratinocytes with or without a dermal substrate are now being used in the treatment of burns and othe r cutaneous wounds. Composite skin cultures (Graftskin, LSE), consisti ng of epidermal keratinocytes seeded on a fibroblast-containing collag en matrix and maintained at the air-liquid interface, develop a well d ifferentiated epidermis in vitro with many of the morphological and bi ochemical features of intact skin. Basement membrane-associated antige ns, developing hemidesmosomes and short segments of lamina densa are p resent at the dermal-epidermal junction in vitro, although the LSE lac ks a continuous basement membrane. As epidermal differentiation procee ds, the culture develops a stratum corneum composed of electron-dense corneocytes surrounded by extracellular lipid. However, the intercorne ocyte lipid lameIlae do not exhibit the repeating pattern of broad and narrow electron lucent bands observed in electron micrographs of the stratum corneum of intact skin. In this study, LSE were grafted onto f ull thickness wounds in athymic mice. Animals were killed 6, 15, 30 an d 60 d after surgery for examination by light and electron microscopy to identify any ultrastructural changes which occurred in the culture in response to the host environment. The grafted LSE integrated well i nto the host tissue and remained intact throughout the 60 d study peri od. At the dermal-epidermal junction, a continuous basement membrane w ith a well defined lamina densa was established by 15 d after surgery. An extensive network of anchoring fibrils was present by 30 d after s urgery. Collagen fibrils within the dermal matrix condensed by 6 d aft er surgery and began organising into loosely packed bundles by 15 d af ter surgery. Tightly packed bundles of collagen fibrils with a Circula r cross-section were observed at 30 d after surgery. Landmann unit rep eats were identified in the intercorneocyte lipid lamellae at 30 d aft er surgery. The ultrastructural analysis of the grafted LSE demonstrat es that the culture, highly differentiated in vitro, not only persiste d after grafting, but responded to the biochemical features of the in vivo environment, rapidly developing additional morphological features of intact skin which may be critical to the establishment of a stable and durable skin replacement.