Keratinocytes are involved in regulating the developmental changes in the proliferative activity of mouse epidermal melanoblasts in serum-free culture

Abstract

When epidermal cell suspensions derived from 0.5-, 2.5-, and 4.5-day-old mice were plated onto uncoated polystyrene dishes and cultured with serum-free medium supplemented with dibutyryl cyclic adenosine 3',5'-monophosphate and basic fibroblast growth factor, melanoblasts proliferated dramatically around keratinocyte colonies and after 12-14 days pure and enriched cultures of melanoblasts (ca. 75%) and melanocytes (ca. 25%) were obtained. In contrast, when epidermal cell suspensions derived from 7.5-, 20.5-, and 60.5-day-old mice were cultured similarly, keratinocytes failed to attach to the dish and melanoblasts did not proliferate at all. However, when epidermal cell suspensions of older mice were plated onto type I collagen-coated dishes and cultured similarly, keratinocytes attached well to the dish and melanoblasts proliferated dramatically around keratinocyte colonies. Moreover, pure melanoblasts and melanocytes derived from primary cultures of young and old mice could be subcultured on collagen-coated dishes with the medium in the presence of secondary keratinocytes that were subcultured from a pure population of primary keratinocytes. No differences were observed in the proliferative activity of secondary melanoblasts between young and old mice. These results suggest that keratinocytes are involved in regulating the proliferation of mouse epidermal melanoblasts and that the developmental changes in the proliferative activity of epidermal melanoblasts in culture are due to the developmental changes in the substrate attachment and proliferation of keratinocytes, rather than to intrinsic changes in melanoblasts.