Phenotype related changes of intimal smooth muscle cells from human aorta in primary culture

To study the functional characteristics of smooth muscle cell (SMC) phenotypes, we have investigated myosin expression, cell proliferation, collagen production and low-density lipoprotein (LDL) receptor activity in intimal SMCs of normal human aorta during their growth in primary culture. By staining with rabbit antibodies to smooth muscle myosin (ASMM) 3 cell types could be distinguished in culture: homogeneously stained cells, cells with discontinuous myosin fibrils and myosin-negative cells. The ratio of cell types greatly changed with culture growth: on days 5, 7 and 14 it was 82:1:17%, 70:5:25% and 10:30:60%, respectively. After 5-6 days of culture intimal SMCs began to proliferate and DNA-synthesizing nuclei were seen 1.5-4.3 times more frequently in myosin-negative cells than in cells with homogeneous myosin distribution. At that time the number of cells reacted with monoclonal antibody (MAb) to an epitope shared collagen types I and III started to increase. By double immunofluorescence staining it was shown that the cultured cells containing both ASMM and MAb markers were found 2.0-4.8 times more rarely than MAb-positive staining in myosin-negative cells. During the first 5 days in culture LDL binding and uptake were diminished in intimal cells with intercellular lipid inclusions independently of their myosin staining pattern, but their activity increased with culture growth. Thus, SMCs from human aortic intima change their phenotype on days 6 and 7 in primary culture as manifested by alteration of myosin expression, increased cell proliferation, collagen production and LDL receptor activity. Changes in myosin expression, however, are not an essential prerequisite for cell proliferation and collagen production.