Angiotensin II Stimulates NADH andNADPH Oxidase Activity inCultured Vascular SmoothMuscleCells

Thesignaling pathways involved inthelong-term metabolic effects ofangiotensin II (AngII)invascular smooth musclecells are incompletely understood butinclude the generation ofmolecules likely toaffect oxidase activity. We examined theability ofAngII tostimulate superoxide anion formation andinvestigated theidentity oftheoxidases respon- sible foritsproduction. Treatment ofvascular smoothmuscle cells withAngIIfor4 to6 hourscaused a 2.7+0.4-fold increase inintracellular superoxide anionformation as de- tected bylucigenin assay.Thissuperoxide appeared toresult fromactivation ofboththeNADPH andNADH oxidases. NADPH oxidaseactivity increased from3.23±0.61to 11.80±1.72 nmol 02-/min permilligram protein after 4hours ofAngII,whereas NADH oxidase activity increased from 16.76+2.13 to45.00+4.57 nmol02-/min permilligram pro- tein. TheNADPH oxidase activity was stimulated byexoge- nous phosphatidic andarachidonic acids andwas partially inhibited bythespecific inhibitor diphenylene iodinium. NADH oxidase activity was increased byarachidonic and linoleic acids, was insensitive toexogenous phosphatidic acid, andwas inhibited byhighconcentrations ofquinacrine. Both ofthese oxidases appeartoreside intheplasma membrane, on thebasis ofmigration oftheactivity after cellular fractionation andtheir apparentinsensitivity tothemitochondrial poison KCN.Theseobservations suggestthatAngIIspecifically activates enzyme systems that promotesuperoxide generation andraise thepossibility thatthesepathways function as second messengersforlong-term responses,such ashypertro- phyorhyperplasia. (Circ Res.1994;74:1141-1148.)