Reactive Oxygen Species and Vascular O2 Sensing

描述（由申请人提供）：该提案侧重于定义Nox氧化酶衍生的过氧化氢，细胞质NADPH和NADH氧化还原系统在控制信号机制的过程中的作用，这些信号机制调节血管平滑肌对PO2生理变化的反应。本应用的研究调查了我们的新证据的重要性，即细胞质内NADPH和可能的NADH水平控制着Nox氧化酶衍生的过氧化氢介导的牛肺动脉松弛的基线，这种松弛被缺氧所消除，导致收缩。另一方面，牛冠状动脉维持较低水平的NADPH和NADPH生成戊糖磷酸途径（PPP）酶葡萄糖-6-磷酸脱氢酶。我们假设，冠状动脉中PPP功能的差异导致缺氧引起细胞质内NADPH的氧化，从而通过我们已经证明由PPP控制的机制激活松弛。Aim 1的研究将探讨PO2如何调节涉及nox衍生过氧化氢的血管反应的表达。目的2侧重于了解PO2如何调节细胞质内NADH和NADPH氧化还原相关的信号机制，这些信号机制有助于控制力的产生。Aim 3的研究旨在确定氮氧化物氧化酶活性和胞质NAD(P)H氧化还原控制差异的起源，这些差异有助于在肺动脉和冠状动脉中观察到缺氧引起的收缩和放松反应。将研究分离的内皮去除的冠状动脉和肺动脉导管以及阻力动脉，通过扰动改变Nox活性和表达，以及胞浆内NADH和NADPH氧化还原来确定这些过程调节的信号系统如何在po2诱导的反应中发挥作用。PO2对胞质内NAD(P)H氧化还原和氧化剂产生的控制将通过代谢测量、组织荧光和化学发光技术以及细胞荧光成像方法进行检查，这些方法旨在表征所涉及的信号机制的ROS和氧化还原方面。还将对大鼠动脉进行检查，以确定Nox和细胞质NAD(P)H氧化还原控制的po2诱导反应机制的异同。这些研究应该有助于确定肺动脉和冠状动脉对氧张力变化的生理反应差异的起源。

DESCRIPTION (provided by applicant): This proposal focuses on defining the role of Nox oxidase-derived hydrogen peroxide, and cytosolic NADPH and NADH redox systems in processes that control signaling mechanisms regulating vascular smooth muscle force responses to physiological changes in PO2. Studies in this application investigate the importance of our new evidence that cytosolic NADPH and possibly NADH levels control a baseline Nox oxidase-derived hydrogen peroxide-mediated relaxation of bovine pulmonary arteries which is removed by hypoxia, resulting in contraction. On the other hand, bovine coronary arteries maintain a lower level of NADPH and the NADPH generating pentose phosphate pathway (PPP) enzyme glucose-6-phosphate dehydrogenase. We hypothesize that differences in the function of the PPP in coronary arteries results in a hypoxia-elicited oxidation of cytosolic NADPH, which activates relaxation through a mechanism we have shown to be controlled by the PPP. Studies in Aim 1 will investigate how PO2 regulates the expression of vascular responses involving Nox-derived hydrogen peroxide. Aim 2 focuses on understanding how PO2 regulates cytosolic NADH and NADPH redox-linked signaling mechanisms that contribute to the control of force generation. Studies in Aim 3 are to define the origins of differences in the control of Nox oxidase activity and cytosolic NAD(P)H redox that contribute to hypoxia-elicited contractile and relaxing responses observed in pulmonary and coronary arteries. Isolated endothelium-removed coronary and pulmonary conduit and resistance arteries will be studied with perturbations that alter Nox activity and expression, and cytosolic NADH and NADPH redox to define how signaling systems regulated by these processes may function in PO2-elicited responses. The control of cytosolic NAD(P)H redox and oxidant production by PO2 will be examined with a combination of metabolic measurements, tissue fluorescence and chemiluminescent techniques, and cellular fluorescence imaging approaches which are designed to characterize the ROS and redox aspects of the signaling mechanisms that are involved. Rat arteries will also be examined to identify similarities and differences in mechanisms of PO2-elicited responses controlled by Nox and cytosolic NAD(P)H redox. These studies should help define the origins of differences in physiological responses of pulmonary and coronary arteries to changes in O2 tension.