IN-VITRO PRODUCTION AND TRANSPLANTATION OF IMMUNOLOGICALLY ACTIVE SKIN EQUIVALENTS

In this study, we produced in vitro epidermal equivalents (EE) and ski n equivalents (SE) with and without spleen lymphocytes. These skin sub stitutes were used for in vitro and in vivo (after isograft) histologi c studies of cell and extracellular matrix organization and for protei n synthesis. Then, using spleen lymphocytes in syngeneic and allogenei c SE, we assessed the immunogenicity of these skin substitutes after t ransplantation. In vitro histologic analyses showed that the epidermal organization of EE was comparable to that of SE. Fibroblasts and sple en lymphocytes were present in the extracellular matrix, as is the cas e in normal skin. Comparative immunohistologic studies after EE and SE isografting showed that the newly generated cutaneous tissues were we ll structured and vascularized. There were indications of physiologica lly active skin. The dermal component in these regenerated skins was, however, more organized after SE than after EE isografting, which indi cates the importance of the dermis. Lastly, allografting of SE with an d without spleen lymphocytes showed interesting results. Indeed, 10 da ys after allografting, all SE allowed skin regeneration comparable to isografts. Moreover, leukocyte infiltration in allografts was observed as early as 10 days and increased during the postgrafting period. Als o, the presence of allogeneic spleen lymphocytes alone in syngeneic SE initiated recipient immune activation and induced leukocyte infiltrat ion and graft rejection. The density of infiltrating leukocytes was hi gher in the complete allograft (allogeneic keratinocytes, fibroblasts, and spleen lymphocytes) compared with the partial allograft (only spl een lymphocytes were allogeneic), with the allograft (allogeneic kerat inocytes and fibroblasts), and with the partial isograft (presence of syngeneic lymphocytes with allogeneic keratinocytes and fibroblasts). Mac-1(+) and CD8(+) cells were present in these leukocyte infiltration s, which indicates recipient immune system activation and allograft re jection. CD4-positive cells were not, however, seen in these leukocyte infiltrations. These results suggest that the incorporation of spleen lymphocytes in SE enhanced their immunogenicity as judged by leukocyt e infiltration and the presence of CD8(+) cells in the implants.