Superoxide limits cyclosporine-A-induced formation of peroxynitrite in endothelial cells

Peroxynitrite (ONOO−) is a potent oxidant formed by the nonenzymatic reaction between superoxide anion (O2•−) and nitric oxide (NO•) in a one-to-one stoichiometry. Accumulated evidence suggests that endothelial dysfunction coincides with an enhanced NO• synthase expression and O2•− production, facilitating ONOO− formation. In vivo, formation of ONOO− has been associated with atherosclerosis and vascular aging. The immunosuppressor Cyclosporine A (CsA) has been associated to human endothelial dysfunction and accelerated atherosclerosis. We have previously shown that CsA induced a transcriptionally mediated increase of the eNOS gene expression and that CsA induced the formation of nitric oxide, O2•−, and ONOO− in vascular endothelial cells. In this work, we evaluate the CsA-induced relative amounts of formation of O2•− and NO•, providing data consistent with a role of O2•−, and not NO•, as the limiting factor in the CsA-dependent intracellular formation of ONOO− in vascular endothelial cells. Furthermore, when endothelial cells were treated with CsA in a situation of increased generation of superoxide such as that provided by high glucose levels, a further increase in the formation of peroxynitrite was detected. The temporal availability of O2•− for peroxynitrite formation may thus become critical in the pathophysiological scenarios where reactive nitrogen intermediates are operative.