Hypoxia Inducible Factor-1 Protects Against Nitrate Tolerance and Stunning in Rabbit Cardiac Myocytes

Abstract

We tested whether upregulation of hypoxia inducible factor-1 (HIF-1) would restore the blunted effects of natriuretic peptides and nitric oxide caused by chronic nitrate exposure and stunning in cardiac myocytes. HIF-1alpha was increased with deferoxamine (150 mg/kg for 2 days). Nitrate tolerance was induced by a chronic nitroglycerin patch (0.3 mg/h for 5 days). We used freshly isolated rabbit ventricular myocytes. Half the myocytes were subjected to simulated ischemia [15 min 95% N(2)-5% CO(2)] and reperfusion [reoxygenation] to produce stunning. Cell function was measured utilizing a video-edge detector. Shortening was examined at baseline and after brain natriuretic peptide (BNP, 10(-8), 10(-7) M) or S-nitroso-N-acetyl-penicillamine (SNAP, 10(-6), 10(-5) M) followed by KT5823 (cyclic GMP protein kinase inhibitor, 10(-6) M). We also measured cyclic GMP protein kinase protein levels and kinase activity. In control, BNP (-29%) reduced percent shortening, while KT5823 partially restored function. Deferoxamine treated control myocytes responded similarly. In patched nonstunned myocytes, BNP (-12%) reduced shortening less and KT5823 did not increase function. However, deferoxamine restored the blunted effects of BNP (-21%) and KT5823. In stunned myocytes, BNP (-11%) reduced shortening less and KT5823 did not affect function. Deferoxamine increased the effects of BNP (-27%) and KT5823 in stunning. Patch combined with stunning also similarly blunted the effects of BNP (-12%) and KT5823. Deferoxamine improved the effects of BNP (-22%) and KT5823. Similar results were observed after SNAP. Stunning reduced cyclic GMP protein kinase activity and deferoxamine restored activity. Deferoxamine had no effect on kinase activity in nitrate tolerance. We found that upregulation of HIF-1 could protect isolated cardiac myocytes against nitrate tolerance through a cyclic GMP protein kinase-independent mechanism and through a kinase-dependent mechanism in stunning.