Calcifying Subpopulation of Bovine Aortic Smooth Muscle Cells Is Responsive to 17β-Estradiol

Abstract

Arterial calcification, common in atherosclerosis, is associated with an increased risk of clinical events such as myocardial infarction. We previously identified a subpopulation of bovine aortic medial cells, calcifying vascular cells (CVCs), that have osteoblastic characteristics and form bone mineral in vitro in the form of calcified nodules. To assess whether estrogen modulates arterial calcification as well as bone calcification, we tested CVCs for estrogen receptors and for the effect of 17 beta-estradiol on formation of calcified nodules, calcium content, alkaline phosphatase activity, and osteocalcin concentration in the culture medium. Estrogen receptor immunoreactivity was identified in the cytoplasm and the perinuclear region of CVCs by immunocytochemistry. CVCs were treated with 17 beta-estradiol at concentrations of 0, 5, and 10 nmol/L. Twenty-one days of 17 beta-estradiol treatment resulted in a significantly increased number of calcified nodules, visualized by von Kossa staining, as well as increased calcium content of the cultures. Increases in alkaline phosphatase activity, a marker for early osteoblastic differentiation, and secreted osteocalcin, a marker for late osteoblastic differentiation, were enhanced in cells treated with 17 beta-estradiol compared with control cells. These results suggest that 17 beta-estradiol promotes osteoblastic differentiation and calcification in vascular cells and that estrogen may play a regulatory role in arterial calcification.