Hypoxia Upregulates AT1 Receptor Expression and Ang II Response to Osteopontin and Proliferation of Cultured Vascular Smooth Muscle (VSM) Cells

Abstract

13 Hypertension is often associated with arterial vasoconstriction which, if severe, may cause local hypoxia. Hypoxia, in turn, stimulates osteopontin (OPN, an adhesion molecule) synthesis and proliferation of VSM cells, which are the key events in the development of atherosclerosis. In the present study, we examined the effect of hypoxia on the expression of Ang II receptor AT1 in cultured VSM cells, and determined whether Ang II-induced regulation of AT1, OPN, and proliferation of VSM cells are altered under hypoxic conditions. Rat aortic VSM cells in culture were rendered quiescent and exposed in a serum-free medium either to hypoxia (3% O 2 ) or normoxia (18% O 2 ) for 2-24 hours in the absence or presence of 1 uM Ang II. Northern blot analysis shows 40-90% increases in AT1 mRNA levels by hypoxia. Consistent with the upregulation of AT1 mRNA levels, hypoxia induced a 125% increase in [ 125 I]Ang II binding, suggesting an increase in AT1 receptor density. Parallel to the increases in AT1 expression, cells exposed to hypoxia produced 40-50% stimulation of OPN mRNA and protein levels as assessed by northern and western blot analysis. Treatment with Ang II for 24 hours under normoxic conditions resulted in a 235% and 45% increases in OPN and [ 3 H]-thymidine incorporation, respectively. However, under hypoxic conditions Ang II-induced increases in OPN and [ 3 H]-thymidine incorporation were enhanced to 387% and 173%, respectively. Incubation with Ang II for 24 hours under normoxic conditions resulted in a 62% inhibition of AT1 mRNA levels. By contrast, the downregulation of AT1 expression by Ang II was blunted under conditions of hypoxia. In conclusion, hypoxia upregulates the expression of AT1 receptors in cultured VSM cells and blocks the downregulation of AT1 expression expected with excess of Ang II. This hypoxia-induced increase in AT1 receptors may, in part, explain the enhancement of OPN synthesis and proliferation of VSM cells observed with Ang II in an environment of hypoxia associated with local vasoconstriction.