title: Increased superoxide production in nitrate tolerance is associated with NAD(P)H oxidase and aldehyde dehydrogenase 2 downregulation.
abstract: Chronic administration of nitroglycerin (NTG) induces nitrate tolerance. Among possible underlying mechanisms, increased vascular production of reactive oxygen species (ROS) has emerged as a principal mechanism. Using cell culture and animal models of nitrate tolerance, we aimed to assess the impact of nitrates on NAD(P)H oxidases and aldehyde dehydrogenase 2 (ALDH2) expression. Rats and vascular smooth muscle cells were treated with NTG. Vascular reactivity was assessed by isometric tension studies. Superoxide was detected by dihydroethidium staining. Gene expression was measured by real-time polymerase chain reaction. NAD(P)H oxidase activity was measured using lucigenin-enhanced chemiluminescence. ALDH activity was measured biochemically, and NO consumption electrochemically. Nitrate tolerance was induced in rats by treatment with NTG for 3 days, and detected as impaired endothelium-dependent and -independent relaxation of aortic segments. Although superoxide production was increased in all aortic layers, expression of nox1, nox2 and nox4 was significantly decreased. Similarly, in vascular smooth muscle cells exposed to NTG for 6-24 h, NAD(P)H oxidase activity was increased, in spite of nox1 downregulation. In addition, expression and activity of ALDH-2 was decreased in nitrate-tolerant rings. Furthermore, exogenous addition of ALDH decreased superoxide generation in vitro and attenuated NO consumption in vascular smooth muscle cell homogenates. Our data suggest that in nitrate tolerance, activation of nox enzymes more than compensates for their downregulation, resulting in a net increase in superoxide and NO consumption. Furthermore, reduced ALDH-2 activity and expression leads to decreased NTG bioconversion. Therefore, both mechanisms reduce NO availability and impair vasorelaxation.
authors.affiliation: Department of Cardiology, University Hospital Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany.
authors.forename: Katalin
authors.initials: K
authors.lastname: Szöcs
authors.forename: Bernard
authors.initials: B
authors.lastname: Lassègue
authors.forename: Philip
authors.initials: P
authors.lastname: Wenzel
authors.forename: Maria
authors.initials: M
authors.lastname: Wendt
authors.forename: Andreas
authors.initials: A
authors.lastname: Daiber
authors.forename: Matthias
authors.initials: M
authors.lastname: Oelze
authors.forename: Thomas
authors.initials: T
authors.lastname: Meinertz
authors.forename: Thomas
authors.initials: T
authors.lastname: Münzel
authors.forename: Stephan
authors.initials: S
authors.lastname: Baldus
mesh_terms: D000444:Aldehyde Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D004361:Drug Tolerance; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D008297:Male; D019255:NADPH Oxidases / Q000235:genetics / Q000378:metabolism*; D005996:Nitroglycerin / Q000494:pharmacology*; D051381:Rats; D017208:Rats, Wistar; D013481:Superoxides / Q000378:metabolism*; D014665:Vasodilator Agents / Q000494:pharmacology*
chemical_list: D014665:Vasodilator Agents; D013481:Superoxides; D000444:Aldehyde Dehydrogenase; D019255:NADPH Oxidases; D005996:Nitroglycerin