The effect of ischemia-reperfusion derived oxygen free radicals on skeletal muscle calcium metabolism.

Abstract

This study investigated the contribution of cytotoxic oxygen-derived free radicals to the skeletal muscle injury seen in a rat hindlimb tourniquet model after ischemia and reperfusion. The free radical scavengers superoxide dismutase (SOD) and catalase (CAT) were used as biologic probes to detect free radical activity, while Ca2+ uptake by sarcoplasmic reticulum (SR) was used to measure subcellular muscle function. Anesthetized rats received SOD (2 mg/kg IV) plus CAT (3.5 mg/kg IV, n = 6 treated group) or saline alone (4 ml/kg, n = 6 control group) 5 min before unilateral hindlimb tourniquet ischemia of 3 hr duration. SOD and CAT were conjugated to polyethylene glycol to increase their plasma half-life. After 19 hr reperfusion, muscle from ischemic and non-ischemic lower legs of each rat was excised and homogenized. Skeletal muscle SR was isolated by differential centrifugation and ATP-dependent Ca2+ uptake by SR was then measured with dual wavelength spectrophotometry and a calcium-sensitive dye. In control rats, Ca2+ uptake velocity by SR from ischemic muscle was reduced by 48% compared with contralateral non-ischemic muscle (p less than .001). Rats pretreated with SOD + CAT showed a less severe (27%) reduction in Ca2+ uptake velocity by SR from ischemic muscle. Thus, SOD + CAT significantly (p less than .01) reduced the dysfunction of SR Ca2+ transport seen in this tourniquet ischemia model. These results strongly implicate the involvement of oxygen-derived free radicals in abnormal Ca2+ transport observed in skeletal muscle after ischemia and reperfusion.