Comparison of inhibitors of superoxide generation in vascular smooth muscle cells

Background and purpose:  We compared the dose‐dependent reductions in cellular superoxide anion (O 2 ‐ ) by catalytic agents: superoxide dismutase (SOD), polyethylene glycol (PEG)‐SOD and the nitroxide 4‐hydroxy‐2,2,6,6,‐tetramethylpiperidine‐1‐oxyl (tempol) with uncharacterized antioxidants: 5,10,15,20‐tetrakis (4‐sulphonatophenyl) porphyrinate iron (III)(Fe‐TTPS), (‐)‐ cis ‐3,3′,4′,5,7‐pentahydroxyflavane (2 R ,3 R )‐2‐(3,4‐dihydroxyphenyl)‐3,4‐dihydro‐1(2 H )‐benzopyran‐3,5,7‐triol (‐epicatechin), 2‐phenyl‐1,2‐benzisoselenazol‐3(2H)‐one (ebselen) and N‐acetyl‐L‐cysteine (NAC) with the spin trap nitroblue tetrazolium (NBT) and with the vitamins or their analogues: ascorbate, α‐tocopherol and 6‐hydroxy‐2,5,7,8‐tetramethylkroman‐2‐carboxy acid (trolox). Experimental approach:  O 2 ‐ was generated in primary cultures of angiotensin II‐stimulated preglomerular vascular smooth muscle cells from spontaneously hypertensive rats and detected by lucigenin‐enhanced chemiluminescence. Key results:  SOD, PEG‐SOD, NAC and tempol produced a similar maximum inhibition of O 2 ‐ of 80–90%. ‐Epicatechin, NBT, ebselen and Fe‐TTPS were significantly ( P < 0.0125) less effective (50–70%), whereas trolox, α‐tocopherol and ascorbate had little action even over 24 h of incubation (<31%). Effectiveness in disrupted and intact cells was similar for the permeable agents, PEG‐SOD and tempol, but was enhanced for SOD. Generation of O 2 ‐ was increased by NAC and NBT at low concentrations but reduced at high concentrations. Conclusions and implications:  Maximum effectiveness against cellular production of O 2 ‐ requires cell membrane permeability and catalytic action as exemplified by PEG‐SOD or tempol. NAC and NBT have biphasic effects on O 2 ‐ production. Vitamins C and E or analogues have low efficacy.