Hypoxia inducible factor-1 protects against nitrate tolerance and stunning in rabbit cardiac myocytes

Purpose: 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. Methods: HIF-1α 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% N₂-5% 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. Results: 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. Conclusion: 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.