Enhanced chemiluminescence as a measure of oxygen-derived free radical generation during ischemia and reperfusion

It has been suggested that oxygen-derived free radicals may contribute to the myocardial injury associated with ischemia and reperfusion. As the presence of enhanced free radical generation is a prerequisite for such damage, several techniques have been used to provide evidence of increased oxygen free radical production during reperfusion; however, all such techniques have substantial limitations. In this study, we used enhanced chemiluminescence to evaluate oxygen free radical generation during ischemia and reperfusion in the isolated Langendorff-perfused rat heart. The chemiluminescent technique, which has high sensitivity and can monitor radical generation continuously, avoids some of the limitations of earlier methods. Chemiluminescence (expressed as counts per second) decreased from 219 ± 11 at baseline to 142 ± 9 during ischemia and markedly increased to a peak of 476 ± 36 during the first 3-5 minutes of reperfusion. This was followed by a slow decline over 11-16 minutes to a steady-state level of 253 ± 14 (each sequential change in chemiluminescence was highly significant; p < 0.001). Superoxide dismutase (2,000 units/min) significantly decreased peak reperfusion chemiluminescence to 316 ± 17 (p < 0.01). Hearts subjected to a second period of ischemia and reperfusion had a higher peak chemiluminescence (626 ± 62), which also was significantly attenuated by 1,000 units/min superoxide dismutase (398 ± 16; p < 0.01). These observations indicate 1) that it is feasible to continuously monitor oxygen free radical generation in the perfused rat heart via enhanced chemiluminescence, 2) that there is a pronounced and prolonged increase in free radical generation during the early reperfusion period, and 3) that superoxide dismutase partially attenuates the elevation of free radical content during reperfusion.