Comparative Effects of Antioxidants on Ang II‐induced Superoxide Generation by SHR Preglomerular Vascular Smooth Muscle Cells (PGVSMCs)

Abstract

We studied inhibition of Ang II-induced superoxide (O2.−) generation in primary cultures of SHR PGVSMCs by catalytic antioxidants (SOD, PEG-SOD, Tempol and Cat-1), by uncharacterized antioxidants (Fe-TPPS, -Epicatechin and Ebselen ), by an O2.− spin trap (Nitroblue tetrazolium,NBT) or by vitamins (ascorbic acid, α-tocopherol and Trolox). Ang II (1μM) increased O2.− generation by 94% determined by lucigenin-derived chemiluminescence (P<0.01, N=3). This was prevented by diphenyleneiodonium (DPI) and apocynin. Antioxidant drugs (10−4 - 10−8M ) over 4 hours caused dose-dependent decreases in O2.− over 4 hours with a maximum inhibition of 91±1; 82±0; 78±0; 65±2; 61±5; 57±2 and 54±13% for PEG-SOD, SOD, Tempol, Ebselen, NBT, -Epicatechin and Fe-TPPS, respectively ( P<0.01 vs. control ). The rank order of sensitivity ( ED50 ) was PEG-SOD > SOD > Tempol > Ebselen > Fe-TPPS >-Epicatechin > NBT. Cat-1, ascorbic acid, α-tocopherol and Trolox were minimally effective even over 20 hours (17–31% inhibition). In conclusion, Ang II stimulates NADPH-oxidase-dependent O2.− generation in PGVSMCs. The sensitivity and effectiveness of catalytic antioxidants (SOD, PEG-SOD and tempol ) are high, whereas that of Fe-TPPS, -Epicatechin, Ebselen and NBT are moderate, and Cat-1, ascorbic acid, α-tocopherol and Trolox are almost ineffective. These results may explain the efficacy of Tempol and SOD in reversing oxidative stress and the disappointing results with vitamins C and E.