all-trans-Retinoic acid preserves viability of fibroblasts and keratinocytes in full-thickness human skin and fibroblasts in isolated dermis in organ culture

Abstract

Human dermal fibroblast and human epidermal keratinocyte survival was examined under various conditions in organ culture. Using cell recovery from organ-cultured tissue as the criterion, it was observed that no keratinocytes and few fibroblasts survived incubation for 10-12 days in serum-free basal medium containing a low level (0.15 mM) of extracellular Ca2+. Increasing the extracellular Ca2+ concentration to 1.4 mM or treating the tissue with 3 microM retinoic acid (RA) under low Ca2+ conditions resulted in increased keratinocyte and fibroblast survival; the two treatments together were more effective than either treatment alone. The same treatments preserved fibroblast survival when pieces of isolated dermal tissue were incubated in organ culture and also supported fibroblast survival in monolayer culture. These findings indicate that recovery of keratinocytes and fibroblasts from skin after maintenance in organ culture provides a simple but definitive measure of the viability of the major cellular elements present in the tissue. These findings suggest that RA treatment enhances survival of both fibroblasts and keratinocytes and that these effects of RA can be seen at physiological Ca2+ concentrations as well as at suboptimal levels of extracellular Ca2+. Finally, these results indicate that the dermis is a direct target of RA.

All-trans Retinoic Acid-Induced Hyaluronan Production and Hyperplasia Are Partly Mediated by EGFR Signaling in Epidermal Keratinocytes

“…The effects of all-trans RA on proliferation, differentiation, and hyaluronan synthesis, similar in the epidermis in vivo (data not shown) and the organotypic epidermal keratinocyte cultures, indicate that these effects of all-trans RA are targeted directly to epidermal keratinocytes, without the contribution of dermal cells or signals (Varani et al, 1993(Varani et al, , 1994. The fact that the stimulation in hyaluronan production was also evident in monolayer cultures suggests that the signaling systems for all-trans RA-mediated hyaluronan upregulation are operating both in differentiating keratinocytes and in monolayers resembling the basal cell phenotype.…”

Heparin-Binding Epidermal-Growth-Factor-Like Growth Factor Activation of Keratinocyte ErbB Receptors Mediates Epidermal Hyperplasia, a Prominent Side-Effect of Retinoid Therapy

“…It is perhaps less well appreciated that epithelial±mesenchymal interactions are also important for ongoing homeostasis of mature epidermis and other epithelia. For example, when the dermis and epidermis of mature skin are separated and cultured in isolation, the dermis remains viable but the epidermis rapidly dies (Varani et al, 1994). We have shown that survival of organ-cultured human skin requires a medium Ca 2+ concentration that is optimal for ®broblast growth (e.g., 1.4 mM) rather than for growth of keratinocytes (e.g., 0.1 mM).…”

“…In addition to retinoid effects on keratinocytes via the c-erbB system, it is also important to consider the effects of topically applied retinoids on all the resident and migratory cell types of the skin. Studies from our laboratory have demonstrated that retinoid-induced epidermal changes result, at least under some conditions, from retinoid effects on dermal elements (Varani et al, 1993b(Varani et al, , 1994. Whether retinoid action in the stroma also involves c-erbB activation is not known.…”

Lipid functions in skin: Differential effects of n-3 polyunsaturated fatty acids on cutaneous ceramides, in a human skin organ culture model

“…The Ca 2 + (1.4 mM) was added in the form of CaCl 2 , directly to the media. Exogenous calcium has been shown to maintain keratinocyte and fibroblast survival in full-thickness human skin [41], [50], [51]. Fatty acid supplementation was performed by dissolving EPA or DHA in DMSO and adding this (2 μl per 10 ml medium, or an equal volume of vehicle (DMSO)) directly to the media to a final concentration of 50 μM.…”