microsurgical model of ischemia reperfusion in rat muscle: evidence of free radical formation by spin trapping

The purpose of developing the experimental model described in this study was to verify the hypothesis that free radicals are formed during ischemia- reperfusion of skeletal muscle. Spin trapping technique, along with electron spin resonance spectroscopy (ESR), directly indicates the presence of reactive radicals, which are widely considered to be important in tissue injury. The experimental model was a rat pedicled rectus femoris muscle flap. The femoral artery and vein were cannulated to inject the 'spin trap' and collect the effluent flow. The spin trap agent was phenyl-t-butyl nitrone (PBN) and Hank's balanced salt solution. Three injections and collections were made: a) before ischemia; b) after ischemia of 15, 30, 60, 120, and 180 minutes, but before blood flow had been restored; and c) after blood flow had been restored. No ESR signal was detected either before the ischemic period or after only 15 minutes of ischemia. PBN radical adducts were detected after 30, 60, 120, and 180 minutes of ischemia. A similar signal was detected when PBN was injected during reperfusion 10 minutes after the ischemic periods. The study demonstrated the presence of free radicals in an in vivo intact skeletal muscle ischemia-reperfusion model.