OXIDATIVE INJURY OF CORONARY VENULAR ENDOTHELIAL-CELLS DEPLETES INTRACELLULAR GLUTATHIONE AND INDUCES HSP-70 MESSENGER-RNA

Vascular endothelium is one of the first tissues exposed to reactive o xygen species produced during myocardial ischemia-reperfusion. Bovine coronary venular endothelial cells (CVEC) were evaluated for intracell ular glutathione (GSH) levels and heat shock protein 70 (HSP 70) mRNA and protein during in vitro oxidative stress. CVEC were incubated with 0.01875 U/ml xanthine oxidase (XO) and 0.5 mM hypoxanthine (HX) for 3 0 min and then allowed to recover for 0, 1, 2, or 3 h. Relative GSH le vels were determined by evaluation of monochlorobimane fluorescence. G SH fluorescence was significantly lower in CVEC treated with XO + HX f or 30 min than in controls. GSH fluorescence was also decreased in hea t-shocked CVEC. After oxidative stress, GSH levels were higher than in controls at 1 h, but by 2 or 3 h after treatment, GSH fluorescence fe ll below control values. HSP 70 mRNA was induced in CVEC by a 30-min t reatment with XO + HX followed by a 2- or 3-h recovery; however, HSP 7 0 protein was not significantly increased after XO + HX exposure. Thes e data suggest that CVEC respond to oxidative stress by reducing intra cellular GSH levels and inducing HSP 70 mRNA, although significant inc reases in HSP 70 protein were not detected at the time points tested.