Important Role of Sarcoplasmic Reticulum Ca Release Ryanodine Receptor-2 Channel in Hypoxia-Induced Rieske Iron-Sulfur Protein-Mediated Mitochondrial Reactive Oxygen Species Generation in Pulmonary Artery Smooth Muscle Cells.

It is known that mitochondrial reactive oxygen species generation ([ROS]) causes the release of Ca ryanodine receptor-2 (RyR2) on the sarcoplasmic reticulum (SR) in pulmonary artery smooth muscle cells (PASMCs), playing an essential role in hypoxic pulmonary vasoconstriction (HPV). In this study, we sought to determine whether hypoxia-induced RyR2-mediated Ca release may in turn promote [ROS] in PASMCs and the underlying signaling mechanism. Our data reveal that application of caffeine or norepinephrine to induce Ca release increased [ROS] in PASMCs. Likewise, exogenous Ca augmented ROS generation in isolated mitochondria and at complex III from PASMCs. Inhibition of mitochondrial Ca uniporter (MCU) with Ru360 attenuated agonist-induced [ROS]. Ru360 produced a similar inhibitory effect on hypoxia-induced [ROS]. Rieske iron-sulfur protein (RISP) gene knockdown inhibited Ca- and caffeine-induced [ROS]. Inhibition of RyR2 by tetracaine or RyR2 gene knockout suppressed hypoxia-induced [ROS] as well. In this article, we present convincing evidence that Ca release following hypoxia or RyR simulation causes a significant increase in MCU, and the increased MCU subsequently RISP-dependent [ROS], which provides a positive feedback mechanism to enhance hypoxia-initiated [ROS] in PASMCs. Our findings demonstrate that hypoxia-induced mitochondrial ROS-dependent SR RyR2-mediated Ca release increases MCU and then RISP-dependent [ROS] in PASMCs, which may make significant contributions to HPV and associated pulmonary hypertension.