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The objective of this study was to determine whether intermediate filament expression, including desmin and vimentin, in cultured smooth muscle cells (SMCs) is related to cyto-differentiation or proliferation. Using antibodies to desmin and vimentin, we studied by immunoperoxidase technique the distribution of these proteins in subcultured SMCs derived from porcine aorta and coronary artery. In addition, the proliferative potentiality of the cells was estimated by the incorporation of [3H]thymidine into DNA. The frequency of desmin-positive cells in coronary arterial SMCs of 3 and 6 population doubling levels was significantly higher as compared to findings with the aortic SMCs and depended on the plating time. No difference was evident at the 12 population doubling level. In contrast, vimentin was present in the majority of both aortic and coronary arterial SMCs. With regard to the localization of vimentin, two cell types were observed, one had reaction products to vimentin in both perinuclear and cell-peripheral areas (type-I cell), the other only in the cell-peripheral region (type-II cell). The relative proportion of the type-I and -II cells varied with the period of culture. Most of the SMCs showed the type-I cell on the first day and the number of type-II cells was increased on the sixth day. Quiescent SMCs in serum-free media had the same percentage of desmin-positive cells and frequency distribution of type-I and -II cells as did the proliferating SMCs incubated in media containing 5% serum. These results suggest that intermediate filament expression, including desmin and vimentin in cultured SMCs, is related to cell origin and/or plating time, but not to the proliferating activity, per se.

In an attempt to better define the relationship between collagen matrices and vascular smooth muscle cells in vitro, proliferation of smooth muscle cells was observed in the early stages of culture. Cells spread on collagen gels had a longer doubling time and less incorporation of [3H]thymidine into DNA on the first day of culture than did cells grown on a plastic substrate. Cells on collagen gels were more elongated than were those on the plastic substrate and showed a "hills and valleys" arrangement from the first day in culture on the collagen type III gel. All cells were identified as smooth muscle having definite microfilaments, dense bodies, and pinocytotic vesicles. They had a distinct actin filament running from end to end when labeled with nitrobenzoxadiazole-phallacidin. Cells on the collagen gels had a larger number of actin filaments traveling parallel to the direction of the major axis of their cytoplasm than did those on the glass substrate. Therefore, cultured smooth muscle cells in the more physiological environment for cells in vitro, i.e., on collagen gels, show a suppression of cellular proliferation and an enhancement of differentiation in the early stages of culture. The effects of collagens on the differentiation of cells vary with the collagen phenotype.