Oxidative inhibition of the vascular Na+-K+ pump via NADPH oxidase-dependent pi-subunit glutathionylation: Implications for angiotensin β1-induced vascular dysfunction

Glutathionylation of the Na+-K+ pump's beta(1)-subunit is a key molecular mechanism of physiological and pathophysiological pump inhibition in cardiac myocytes. Its contribution to Na+-K+ pump regulation in other tissues is unknown, and cannot be assumed given the dependence on specific beta-subunit isoform expression and receptor-coupled pathways. As Na+-K+ pump activity is an important determinant of vascular tone through effects on [Ca2+](i), we have examined the role of oxidative regulation of the Na+-K+ pump in mediating angiotensin II (Ang II)-induced increases in vascular reactivity. beta(1)-subunit glutathione adducts were present at baseline and increased by exposure to Ang II in rabbit aortic rings, primary rabbit aortic vascular smooth muscle cells (VSMCs), and human arterial segments. In VSMCs, Ang II-induced glutathionylation was associated with marked reduction in Na+-K+ ATPase activity, an effect that was abolished by the NADPH oxidase inhibitory peptide, tat-gp91 ds. In aortic segments, Ang II-induced glutathionylation was associated with decreased K+-induced vasorelaxation, a validated index of pump activity. Ang II-induced oxidative inhibition of Na+-K+ ATPase and decrease in K+-induced relaxation were reversed by preincubation of VSMCs and rings with recombinant FXYD3 protein that is known to facilitate deglutathionylation of beta(1)-subunit. Knock-out of FXYD1 dramatically decreased K+-induced relaxation in a mouse model. Attenuation of Ang II signaling in vivo by captopril (8 mg/kg/day for 7 days) decreased superoxide-sensitive DHE levels in the media of rabbit aorta, decreased (beta(1)-subunit glutathionylation, and enhanced K+-induced vasorelaxation. Ang II inhibits the Na+-K+ pump in VSMCs via NADPH oxidase-dependent glutathionylation of the pump's beta(1)-subunit, and this newly identified signaling pathway may contribute to altered vascular tone. FXYD proteins reduce oxidative inhibition of the Na+-K+ pump and may have an important protective role in the vasculature under conditions of oxidative stress. (C) 2013 Elsevier Inc. All rights reserved.