Rat aortic smooth muscle cells become pericytes during angiogenesis in vitro.

BACKGROUND

We previously reported that the intimal endothelium of the rat aorta switches to a microvascular phenotype during angiogenesis in vitro. The microvessels formed by the rat aortic endothelium are coated with pericytes. The purpose of this study was to evaluate the relation of the pericytes to the angiogenic process and to identify the site of origin of these cells in the aortic wall.

EXPERIMENTAL DESIGN

Rings of rat aorta were cultured in collagen gel under serum-free conditions. The formation of a pericyte coating around aorta-derived microvessels was evaluated by counting pericytes and microvessels in the living cultures. Pericytes and endothelial cells were studied by immunohistochemistry, lectin labeling, electron microscopy, 3H-thymidine labeling followed by autoradiography, and time-lapse video microscopy. The capacity of aortic smooth muscle cells to differentiate into pericytes was studied by coculturing intimal- or medial-derived smooth muscle cells with endothelial cells in a collagen gel overlay assay that induced reorganization of endothelial cells into microvessels.

RESULTS

Microvessels during the early stages of angiogenesis were composed primarily of endothelial cells. As vascular proliferation decreased, the microvessels became coated with pericytes. The pericytes migrated from the root to the tip of the microvessels using the endothelium as a surface for attachment, proliferation, and contact guidance. The pericytes were continuous with the myointimal endothelial cells of the cultured aorta. Pericytes and myointimal cells were positive for alpha-smooth muscle actin and vimentin and were actively engaged in DNA synthesis. Treatment of the cultures with heparin caused a marked reduction in the number of pericytes. Smooth muscle cells isolated from the intimal aspect of the rat aorta migrated toward the endothelium and differentiated into pericytes when cocultured with microvessels formed by isolated endothelial cells in a collagen gel overlay assay. Conversely, smooth muscle cells isolated from the deep layers of the media had no significant endothelial tropism and failed to differentiate into pericytes.

CONCLUSIONS

This study demonstrates that the rat aorta contains a subpopulation of intimal/subintimal smooth muscle cells that differentiate into pericytes during angiogenesis in vitro. These cells have a distinct endothelial tropism and respond to endothelial cues by contributing to the differentiation and maturation of microvessels. Smooth muscle cells of rat aortic intimal/subintimal origin can be used as a source of pericytes for the in vitro assembly of histotypic microvessels.