Hypoxia activates NADPH oxidase to increase [ROS]i and [Ca2+]i through the mitochondrial ROS-PKCepsilon signaling axis in pulmonary artery smooth muscle cells.

Abstract

The importance of NADPH oxidase (Nox) in hypoxic responses in hypoxia-sensing cells, including pulmonary artery smooth muscle cells (PASMCs), remains uncertain. In this study, using Western blot analysis we found that the major Nox subunits Nox1, Nox4, p22(phox), p47(phox), and p67(phox) were equivalently expressed in mouse pulmonary and systemic (mesenteric) arteries. However, acute hypoxia significantly increased Nox activity and translocation of p47(phox) protein to the plasma membrane in pulmonary, but not mesenteric, arteries. The Nox inhibitor apocynin and p47(phox) gene deletion attenuated the hypoxic increase in intracellular concentrations of reactive oxygen species and Ca(2+) ([ROS](i) and [Ca(2+)](i)), as well as contractions in mouse PASMCs, and abolished the hypoxic activation of Nox in pulmonary arteries. The conventional/novel protein kinase C (PKC) inhibitor chelerythrine, specific PKCepsilon translocation peptide inhibitor, and PKCepsilon gene deletion, but not the conventional PKC inhibitor GO6976, prevented the hypoxic increase in Nox activity in pulmonary arteries and [ROS](i) in PASMCs. The PKC activator phorbol 12-myristate 13-acetate could increase Nox activity in pulmonary and mesenteric arteries. Inhibition of mitochondrial ROS generation with rotenone or myxothiazol prevented hypoxic activation of Nox. Glutathione peroxidase-1 (Gpx1) gene overexpression to enhance H(2)O(2) removal significantly inhibited the hypoxic activation of Nox, whereas Gpx1 gene deletion had the opposite effect. Exogenous H(2)O(2) increased Nox activity in pulmonary and mesenteric arteries. These findings suggest that acute hypoxia may distinctively activate Nox to increase [ROS](i) through the mitochondrial ROS-PKCepsilon signaling axis, providing a positive feedback mechanism to contribute to the hypoxic increase in [ROS](i) and [Ca(2+)](i) as well as contraction in PASMCs.

• Authors: Rakesh Rathore, Yun-Min Zheng, Chun-Feng Niu, Qing-Hua Liu, Amit Korde, Ye-Shih Ho, Yong-Xiao Wang
• Journal: Free radical biology & medicine (Free Radic Biol Med) Vol. 45 Issue 9 Pg. 1223-31 (Nov 01 2008) ISSN: 0891-5849 [Print] United States
• PMID: 18638544 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)

Chemical References

• Enzyme Inhibitors
• Reactive Oxygen Species
• NADPH Oxidases
• neutrophil cytosolic factor 1
• Protein Kinase C-epsilon
• Calcium

Topics

• Animals
• Calcium (metabolism)
• Enzyme Inhibitors (pharmacology)
• Gene Expression Regulation, Enzymologic
• Hypoxia
• Mice
• Mice, Transgenic
• Mitochondria (metabolism)
• Models, Biological
• Myocytes, Smooth Muscle (metabolism)
• NADPH Oxidases (metabolism)
• Protein Kinase C-epsilon (metabolism, physiology)
• Pulmonary Artery (metabolism)
• Reactive Oxygen Species
• Signal Transduction