Reactive oxygen metabolite‐induced toxicity to cultured bovine endothelial cells: Status of cellular iron in mediating injury

Abstract

We aimed to determine the status of iron in mediating oxidant-induced damage to cultured bovine aortic endothelial cells. Chromium-51-labeled cells were exposed to reaction mixtures of xanthine oxidase/hypoxanthine and glucose oxidase/glucose; these produce superoxide and hydrogen peroxide, or hydrogen peroxide, respectively. Xanthine oxidase caused a dose dependent increase of 51Cr release. Damage was prevented by allopurinol, oxypurinol, and extracellular catalase, but not by superoxide dismutase. Prevention of xanthine oxidase-induced damage by catalase was blocked by an inhibitor of catalase, aminotriazole. Glucose oxidase also caused a dose-dependent increase of 51Ci release. Glucose oxidase-induced injury, which was catalase-inhibitable, was not prevented by extracellular superoxide dismutase. Both addition of and pretreatment with deferoxamine (a chelator of Fe3+) prevented glucose oxidase-induced injury. The presence of phenanthroline (a chelator of divalent Fe2+) prevented glucose oxidase-induced 51Cr release, whereas pretreatment with the agent did not. Apotransferrin (a membrane impermeable iron binding protein) failed to influence damage. Neither deferoxamine nor phenanthroline influenced cellular antioxidant defenses, or inhibited lysis by non-oxidant toxic agents. Treatment with allopurinol and oxypurinol, which inhibited cellular xanthine oxidase, failed to prevent glucose oxidase injury. We conclude that (1) among the oxygen species extracellularly generated by xanthine oxidase/hypoxanthine, hydrogen peroxide induces damage via a reaction on cellular iron; (2) deferoxamine and phenanthroline protect cells by chelating Fe3+ and Fe2+, respectively; and (3) reduction of cellular stored iron (Fe3+) to Fe2+ may be prerequisite for mediation of oxidant-induced injury, but this occurs independently of extracellular superoxide or cellular xanthine oxidase-derived superoxide.

Ischemia-Reperfusion Injury of Retinal Endothelium by Cyclooxygenase- and Xanthine Oxidase-Derived Superoxide

“…The resultant duration-dependent injury of MRECs following exposure to this oxidant stress is similar to that reported for endothelial cells from other tissues (Andreoli, 1989;Hiraishi et al, 1994). The speci®city of the response was con®rmed by the ability of oxypurinol to completely block the injury.…”

Mechanisms of shock wave induced endothelial cell injury

“…Although production of ROS generally would not be expected in samples exposed to SWs solely we also elucidated whether scavengers could attenuate effects of SWs per se. The concentration of the scavengers chosen, had previously been shown to abolish effects of ROS at concentrations 100fold higher than expected in our SWC wells [35][36][37]. In this study, no protective effects of the scavengers were seen.…”

Novel biological function of sialic acid (N‐acetylneuraminic acid) as a hydrogen peroxide scavenger

“…Next, we tested the detoxification effect of NANA on cytotoxic proteins producing H 2 O 2 . Glucose oxidase (EC 1.1.3.4), previously known as the toxic antimicrobial protein notatin [11], produces D ‐gluconolactone and H 2 O 2 from D ‐glucose and an oxygen molecule in the culture medium (RPMI‐1640 medium containing approximately 11.1 mM of D ‐glucose), and injures cells by releasing H 2 O 2 . Again, pretreatment with 25 mM of NANA inhibited the cytotoxicity (Fig.…”