Role of Cytoglobin in the Regulation of Vascular Tone

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Endothelium-derived nitric oxide (NO), is a key mediator regulating vascular tone and blood pressure. NO mediates vascular relaxation through activation of soluble guanylate cyclase (sGC) in the smooth muscle. While NO is synthesized by NO synthase in the endothelium, the process of vascular NO degradation and metabolism in smooth muscle is poorly understood. NO degradation in the vessel wall is mediated by an O2- dependent NO dioxygenase (NOD) that oxidizes NO to nitrate. Cytoglobin (Cgb) is a recently discovered globin expressed in smooth muscle (but not in the endothelium) with previously unknown function. Cgb is proposed to serve as a critical regulator of the rate of O2-dependent NO metabolism in the vessel wall, in turn regulating vascular tone. Over the prior grant period: 1) we demonstrated that Cgb is the major heme protein that regulates the rate of O2-dependent NO metabolism in the smooth muscle of both conduit and resistance vessels and, in turn, profoundly modulates vascular tone; 2) we identified that cytochrome b5 (B5)/ B5 reductase (B5R) constitutes the major Cgb reducing system that supports this NOD function; 3) we discovered that Cgb has potent superoxide dismutase (SOD) function accounting for its previously unexplained antioxidant effects; 4) most recently, we identified novel selective inhibitors of Cgb NOD function that do not impair its SOD function and were shown in vessels to enhance NO mediated sGC activation. However, major questions remain regarding the overall process of NO decay in the vessel wall, how this varies in disease, and how it can be modulated to ameliorate disease. In the next stage of this grant program, we seek to determine how the NO degrading and SOD/antioxidant properties of Cgb in the vessel wall control the processes of NO and redox metabolism in normal and hypertensive vessels, and how this in turn regulates vessel tone and systemic blood pressure. The critical effects of Cgb expression levels and the modulation of its NOD activity will be determined. Studies will be performed first in isolated vessels and then in the in vivo cardiovascular system, with measurements in our genetically modified mouse lines with Cgb-/-, Cgb-/+ and Cgb overexpression as well as compounds that selectively inhibit Cgb NOD function. Studies will also be performed in our recently developed conditional smooth muscle-selective Cgb-/- mouse to definitively characterize the role of Cgb in smooth muscle. Studies will focus on the physiological regulation that occurs in normal non-hypertensive mice and then on angiotensin-induced hypertension. All the data obtained will be used to support computational modeling that will enable us to predict the effects of modulating Cgb expression and its NO metabolizing function. Accomplishment of this research plan will elucidate how Cgb levels and its NOD and SOD function regulates O2-dependent NO metabolism and the redox state of the vessel, thus providing important insights into the regulation of vascular tone in normal physiology and cardiovascular disease. This knowledge will lead to new therapeutic approaches to treat or reverse hypertension and other cardiovascular disease.