Development and evaluation of a new composite Laserskin graft

BACKGROUND: Tremendous effort has been made to improve the graft take rate of cultured epidermal autograph. The purpose of this study is to develop and evaluate a new composite Laserskin graft (CLSG) as a human skin substitute for wound resurfacing. METHODS: The seeding efficacy of cultured keratinocytes on plain Laserskin was compared with the 3T3 cell-seeded Laserskin and allogenic fibroblast-populated Laserskin. Three different types of CLSG, 2 cm in diameter each, were prepared and tested in rats. Type A CLSG consisted of proliferative allogenic rat fibroblasts on both sides of the Laserskin with autologous keratinocytes also on the upper side. Fibroblasts and keratinocytes were seeded only on the upper side of the Laserskin in type B CLSG. Keratinocytes alone were seeded on plain Laserskin in type C CLSG. Type B CLSG consisting of autologous keratinocytes and autologous dermal fibroblasts was tested on five selected wounds (5x5 cm each) of a patient with full-thickness burn. In another burn patient, type B CLSG consisting of autologous keratinocytes and allogenic dermal fibroblasts was grafted onto three wounds (5x5 cm each). RESULTS: The seeding efficacy of human keratinocytes on plain Laserskin increased from 75% to 95% when proliferative allogenic fibroblasts were grown as a feeder layer on the Laserskin. The seeding efficacy of rat keratinocytes increased from 36% to 88% in the presence of a proliferative allogenic fibroblast feeder layer, whereas human/rat keratinocytes had respective seeding efficacy of 98%/91% on Laserskin preseeded with mitomycin C-treated 3T3 cells. Skin biopsies of grafted type A CLSG on day 14 after grafting showed complete epithelialization without severe inflammation in 16 of 20 (80%) grafted surgical wounds in rats. There were eight (40%) and seven (35%) "takes" of the CLSG in types B and C, respectively. The infection rate in type B CLSG was two (10%). There was one (5%) infection in types A and C. The respective take rates on the two patients grafted with type B CLSG were 60% and 100%. CONCLUSION: The animal experiment and the preliminary clinical data showed that the CSLGs consisting of autologous keratinocytes and of autologous/allogenic fibroblasts are good human skin substitutes in terms of durability, biocompatibility, high seeding efficacy for keratinocytes, high graft take rate, and low infection rate.