Signaling Mechanisms for Hypoxic Pulmonary Vasoconstriction

描述(申请人提供)：缺氧性肺血管收缩(HPV)是维持肺内局部肺泡通气量和肺灌注量的充分匹配的重要生理过程，但也可能导致肺高压。肺动脉平滑肌细胞(PASMCs)内[Ca~(2+)]i升高是HPV发病的关键因素。[Ca~(2+)]i的缺氧性升高可能是由于细胞内钙离子通过兰尼定受体/钙释放通道(RyRs)从肌浆网释放出来，而RyRs被线粒体的ROS信号和NADPH氧化酶(NOx)激活。依赖NOx的缺氧性ROS的产生仅次于线粒体ROS。然而，低氧产生ROS信号的分子基础仍不清楚。在这里，我们提出了一个创新的中心假说，即线粒体复合体III中的Rieske是一个关键的初始低氧敏感分子，它介导了低氧ROS的产生(在复合体III中，线粒体，然后是胞浆)，RyR激活，PASMCs[Ca2+]i增加和收缩，导致肺血管收缩和高血压。此外，Rieske的表达可能会有所不同，以确定阻力和导管肺以及体动脉SMC的低氧反应的异质性。为了验证这一新的假说，我们将解决以下基本问题(特定目的)：(1)线粒体复合体III中的Rieske是否是介导缺氧性ROS产生、RyR激活、PASMCs[Ca~(2+)]i增加和收缩的关键初始分子~(2)Rieske介导的缺氧性ROS信号和伴随的RyR依赖的Ca~(2+)信号对缺氧性肺动脉高压很重要~(3)Rieske的表达变化是否介导了缺氧性ROS信号的异质性，RyR介导的信号在肺动脉和体动脉SMC中的异质性？这些具体目标将在分子、复合体、线粒体、细胞、组织和生物体水平上使用互补的分子、生化、生理和遗传方法来实现。这些发现将扩大我们对PASMCs低氧钙和收缩反应的机制的理解，以及对肺动脉和体动脉SMC低氧反应的异质性的理解。我们的数据也可能有助于确定肺高压和其他相关肺部疾病的新治疗靶点。

DESCRIPTION (provided by applicant): Hypoxic pulmonary vasoconstriction (HPV) is an important physiological process that maintains the sufficient matching of regional alveolar ventilation and pulmonary perfusion in the lungs, but may also lead to pulmonary hypertension. An increase in [Ca2+]i in pulmonary artery smooth muscle cells (PASMCs) is a key factor in HPV. The hypoxic increase of [Ca2+]i may occur due to intracellular Ca2+ release from the sarcoplasmic reticulum via ryanodine receptors/Ca2+ release channels (RyRs), which are activated by reactive oxygen species (ROS) signaling from mitochondria and NADPH oxidase (NOX). NOX-dependent hypoxic ROS generation is secondary to mitochondrial ROS. However, the molecular basis by which hypoxia generates ROS signaling still remains unclear. Here, we propose an innovative central hypothesis that rieske in the mitochondrial complex III is a key, initial hypoxic sensing molecule, which mediates the hypoxic ROS generation (in the complex III, mitochondria and then cytosol), RyR activation, [Ca2+]i increase and contraction in PASMCs, leading to pulmonary vasoconstriction and hypertension. In addition, rieske expression may vary to determine the heterogeneity of hypoxic responses in resistance and conduit pulmonary as well as systemic artery SMCs. To test this novel hypothesis, we will address the following fundamental questions (Specific Aims): (1) is rieske in the mitochondrial complex III a key, initial molecule that mediates the hypoxic ROS production, RyR activation, [Ca2+]i increase and contraction in PASMCs~ (2) are rieske-mediated hypoxic ROS signaling and attendant RyR-dependent Ca2+ signaling important for hypoxic pulmonary hypertension~ and (3) does rieske vary in expression to mediate the heterogeneity of hypoxic ROS signaling, RyR-mediated signaling in pulmonary and systemic artery SMCs? These specific aims will be implemented using complementary molecular, biochemical, physiological, and genetic approaches at the molecular, complex, mitochondrial, cellular, tissue and organism levels. The findings will extend our understanding of the mechanisms for hypoxic Ca2+ and contractile responses in PASMCs and also the heterogeneity of hypoxic responses in pulmonary and systemic artery SMCs. Our data may also help to identify novel therapeutic targets for pulmonary hypertension and other related lung diseases.