Osteonectin (SPARC) Expression in Vascular Calcification: In Vitro and Ex Vivo Studies

Osteonectin, also termed SPARC, is a noncollagenous protein of bone matrix. Since there are controversial results regarding its role during the process of vascular calcification, we investigated osteonectin expression in our in vitro calcification model. Rat vascular smooth muscle cells (VSMCs) were challenged with high phosphate (5 mmol/L Pi) and analyzed quantifying calcium levels, through immunohistochemical studies, and studying gene expression. We detected a peak of osteonectin expression at day 7 in cell treated with high phosphate. The time course of calcium deposition, reflected the expression of osteonectin, resulting extensively present at day 7. On the contrary, the expression of the mitotic marker Ki-67 had a peak at day 4, showing no correlation between osteonectin and cell proliferation. Moreover, 7 days was the time point in which Cbf alpha 1/RUNX-2 had its maximal expression. Furthermore, ascorbic acid increased osteonectin expression, supporting a procalcifying role for this protein. Next we decided to study osteonectin expression ex vivo in fetal, adult not calcified, and adult calcific vessels. Immunohistochemical studies demonstrated a spread and strong reactivity in VSMCs of a 20-week fetus, confirming that osteonectin may have a potential role in regulation of mitosis and in cell differentiation. In adult not calcified arteries, osteonectin was constitutively expressed and its levels increased in atherosclerotic and in calcified plaques, where it could have a regulatory role in the calcification process. Our in vitro and ex vivo data show osteonectin expression during the calcification process and suggest its potential role as procalcifying factor.