Influence of endothelial cells on structure, biochemistry and functionality of epidermis reconstructed on synthetic porous membrane

The model of keratinocytes cultured on a synthetic porous membrane at the a ir-liquid interface leads to the formation of a pluristratified and cornifi ed epidermis with histological and biochemical characteristics near those o bserved in vivo. In the present study, we evaluated the effect of prolifera tive endothelial cells on epidermalization. Keratinocytes were grown in thr ee culture conditions: in defined medium (DM; control), in medium previousl y conditioned by proliferative endothelial cells (CM) and in medium with pr oliferative endothelial cells (pEC). The structures of reconstructed epider mis were analyzed by electron microscopy, their biochemistry by DNA, protei n and cytokine analyses and finally their functionality was evaluated by es tradiol and water absorption testing. Ultrastructural analysis showed a wel l-developed and cornified epidermis for each culture condition. In addition , living epidermis was thinner in the presence of endothelial cells, reveal ing faster epidermal differentiation. DNA and protein analyses were in acco rdance with these results. Secreted soluble factors varied according to cul ture conditions. At 37 degrees C, the permeability of reconstructed epiderm is in DM, in CM or with pEC was 5- to 10-fold higher than that of native hu man epidermis with both tracers. Laminin coating of the inserts led to simi lar absorption results except for the DM where the barrier function to estr adiol was decreased 2-fold. At 32 degrees C, reconstructed and native epide rmis were, respectively, 1.5- and 2-fold less permeable to estradiol compar ed to 37 degrees C. In conclusion, this model is adequate for fundamental a nd pharmacological studies since it allows the study of interactions betwee n two cell types without their direct contact as well as percutaneous absor ption tests directly performed in the modified culture chamber.