Development of an optimized protocol for primary culture of smooth muscle cells from rat thoracic aortas.

Primary culture of smooth muscle cells has been widely used as a valuable tool to study the molecular mechanisms underlying atherosclerosis and restenosis. Currently, tissue explants and enzymatic digestion methods are frequently applied to produce smooth muscle cells. Explants method is time consuming, usually taking several weeks. The enzymatic digestion method requires large amounts of proteolytic enzymes to generate enough cells for cardiovascular research. The present study reports an optimized method by combining both techniques to obtain high purity smooth muscle cells. The cultured cells exhibited the characteristic "hills and valleys" growth pattern as observed by phase contrast microscopy and showed alpha-SM-actin positive staining by indirect immunocytochemistry and immunofluorescence. Purity of the cells is guaranteed by the lack of von Willebrand Factor immunoreactivity. Finally, the cultured cells well proliferate on oxidized-LDL stimulation, suggesting the practical utility of this new method.