Development of an endothelial-smooth muscle cell coculture model using phenotype-controlled smooth muscle cells.

A coculture of endothelial cells (ECs) and smooth muscle cells (SMCs), which mimics cellular interactions appearing in vivo, has been performed in studies on the relationship between atherogenesis and fluid shear stress conditions. Although healthy arteries in vivo consist of contractile phenotype SMCs, cultured cells used in many studies normally exhibit a synthetic phenotype. Here, we developed an EC-SMC coculture model to investigate the interactions between ECs and contractile SMCs, and examined the effect of shear stress applied to ECs on SMC phenotypes. Cultured human umbilical artery SMCs were differentiated into contractile states by arresting cell growth using a serum-free medium. Western blotting confirmed that SMC expression of contractile protein markers, α-smooth muscle actin (SMA) and calponin, increased to levels similar to those observed in arterial cells. After coculturing contractile SMCs with ECs separated by a collagen gel layer, the expression of α-SMA decreased under static conditions, indicating that the SMC phenotype tended to be synthetic by coculturing with ECs, but shear stress applied to cocultured ECs maintained the level of α-SMA expression in SMCs. The coculture model constructed in the present study will be a useful tool to investigate interactions between ECs and contractile SMCs under shear conditions.