Primary Culture of Rat Aortic Vascular Smooth Muscle Cells: A New Method.

Abstract

BackgroundDeveloping a simple and efficient method of obtaining primary cultured VSMCs is necessary for basic cardiovascular research.Material/MethodsThe procedure of our new method mainly includes 6 steps: isolation of the aortic artery, removal of the fat tissue around the artery, separation of the media, cutting the media into small tissue blocks, transferring the tissue blocks to cell culture plates, and incubation until the cells reach confluence. The cells were identified as VSMCs by morphology and immunofluorescence. Then, VSMCs obtained by this new tissue explants method, the traditional tissue explants method, the enzyme digestion method, and A7r5 cell line were divided into 4 groups. The purity of cells was test by multiple fluorescent staining. Western blotting was used to investigate the phenotype of VSMCs obtained by different methods.ResultsCells began to grow out at about 8 days and became relatively confluent within 16 days. Compared with VSMCs from the traditional tissue explants method and enzyme digestion method or A7r5 cell line, VSMCs obtained by our method showed higher purity and manifested a more “contractile” phenotype characteristic.ConclusionsWe have conquered the disadvantages in the previous primary culture methods and established a simple and reliable way to isolate and culture rat aortic VSMCs with high purity and stability.