This study seeks to identify the master regulatory genes responsible for the differentiation of vascular smooth muscle cells. A cultured, immortalized, mouse neural crest cell line (Monc-l) will be employed. These cells will be induced to differentiate along the smooth muscle cell lineage. Cell-type specific markers will be used to confirm expression of the smooth muscle cell phenotype at various stages. Mouse genetic microarrays will be used to probe the cells during differentiation to identify candidate regulatory genes responsible for smooth muscle cell determination and differentiation, with an emphasis on early transcription factors. Assays of expression levels and kinetics will be performed in the presence of stimulating and inhibitory substances, with the goal of identifying the earliest critical factors during differentiation. Selected genes identified during this process will then be isolated and cloned in order to further characterize them. These genes may be reinserted into the neural crest cells to induce differentiation by overexpression. The primary objective will be to reconstruct the sequence of transcriptional controls responsible for the conversion of neural crest cells to vascular smooth muscle cells. This model will help to elucidate the process of vasculogenesis, and give insights into the origins of atherosclerosis.
