XANTHINE OXIDASE-DERIVED SUPEROXIDE CAUSES REOXYGENATION INJURY OF ISCHEMIC CEREBRAL ENDOTHELIAL-CELLS

Oxygen free radicals, generated by cerebral ischemia, have been widely implicated in the damage of vascular endothelium. Endothelial cells h ave been proposed as a significant source of oxygen free radicals. In the present study, we developed an anoxia-reoxygenation (AX/RO) model using pure cultures of cerebral endothelial cells (CECs) isolated from piglet cortex to measure CEC oxygen free radical production and deter mine its role in AX/RO-induced CEC injury. CEC injury, as measured by lactate dehydrogenase efflux into the culture medium, increased progre ssively with the duration of anoxic exposure, becoming significant aft er 10 h. Reoxygenation significantly increased CEC anoxic injury in a time-dependent manner. A 55% increase in oxygen free radical productio n, determined by fluorescence detection of dihydroethidium oxidation, was measured at the end of 4-h reoxygenation in CECs subjected to AX/R O conditions that killed 40% of the cells. Blockade of oxygen free rad ical production with superoxide dismutase (SOD; 250 and 1000 U/ml) or oxypurinol (50 and 200 mu M), a potent xanthine oxidase inhibitor, red uced this injury by 32-36% and 30-39%, respectively. Results from our in vitro model indicate that CECs produce significant amounts of oxyge n free radicals following ischemia, primarily from the xanthine oxidas e pathway. These radicals ultimately have a cytotoxic effect on the ve ry cells that produced them. Thus, reductions in oxygen free radical-m ediated vascular injury may contribute to improvements in neurophysiol ogic outcome following treatment with oxygen free radical inhibitors a nd scavengers. (C) 1998 Elsevier Science B.V.