Oxidants and Nitric Oxide in Coronary Vascular Function

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The central hypothesis of this project is that specific mechanisms regulating changes in NAD(P)H oxidase (Nox) activity and subunit expression have important roles, in controlling aspects of mediator release from endothelium and signaling mechanisms observed to control vascular smooth muscle contractile function through both changes in ROS and redox control mechanisms. Studies in Aim 1 will define the influence of pathways of Nox oxidase regulation by interactions between Ang II, stretch, a stimuli of protein kinase C (PKC), and altered availability of NAD(P)H on ROS and redox regulation of vascular contractile function in the absence of vascular disease or endothelial factors. The focus of the systems studied include the control of cytosolic NAD(P)H redox and glutathione redox, mitochondria! function (associated with energy metabolism, oxidant production and redox control), and the impact of these systems on specific ROS and redox-controlled signaling mechanisms known to regulate vascular force generation. Studies in Aim 2 examine how changes in Nox activity control endothelial release of NO, ROS and reactive NO-derived species (RNS), and the influence of endothelium and NO-donor drug derived RNS on the systems studied in Aim 1. Emphasis will be placed on building on our previous signaling studies in bovine coronary arteries and extending these studies into mouse aortas due to the availability of mice deficient in the p47phox and Nox-2 (gp91phox) subunits of Nox oxidases. Freshly isolated vascular smooth muscle cells will be studied by fluorescence imaging in the absence and presence of NO-donors to examine relationships between alterations in Nox activation with the detection of changes in ROS and cytosolic and mitochondria! NAD(P)H redox, and indicators of mitochondrial membrane potential and intra-mitochondrial superoxide. A focus of these studies is to develop an understanding processes that control interactions between cytosolic and mitochondrial NAD(P)H redox and ROS generation that are related to the control of vascular force generation. The studies in Aim 3 examine how many of the mechanisms studied in Aim 1 and Aim 2 are altered in vascular tissue derived from animals studied in the other projects exposed to acute and chronic increases in angiotensin II without increased blood pressure, in vivo to alterations in pressure, diabetics and aging.