Cultured human melanocytes provide an excellent model for the study of neural crest development. Discrete growth and differentiation signals can be delivered to homogeneous cell populations and the effects of these signals monitored by changes in morphology, growth rate, differentiated functions and gene expression. In this way, an in vitro model of melanocyte differentiation can be defined and ultimately correlated with characteristic developmental abnormalities of the pigmentary system. The goal of these studies is to critically examine the effect of local tissue environment and exogenous factors suspected to impact on human melanocyte differentiation during development utilizing a sophisticated tissue culture system. We will use known and potential modulators of melanocyte growth and differentiation to examine the effect of these factors on the expression of specific genes and protein products potentially involved in the differentiated function of melanocytes. The effect of nerve growth factor, laminin, fibronectin and vitamins A and D, and medium conditioned by cultured keratinocytes on melanocyte morphology (dendricity), proliferative capacity, melanin synthesis and the expression of the receptor for nerve growth factor will be determined using the appropriate techniques (immunofluoresence, immunobloting, northern blotting). We will subsequently use phorbol esters, known to affect melanocyte morphology, to examine the induction of genes potentially involved in melanocyte growth and differentiation. We will ultimately attempt to transfect the human nerve growth receptor gene into cultured human melanocytes in an effort to explore the tissue specific expression of this gene. In this way, we hope to construct a model of normal human melanocyte differentiation in vitro.
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