Hindlimb ischemia-reperfusion increases complement deposition and glycolysis

Background. Hindlimb ischemia-reperfusion (HIR) impairs cellular energy metabolism and causes local muscle injury possibly through free radical or complement-mediated mechanisms. Materials and methods. To determine the relationship among myocellular energetics, histopathological injury, and mediator activity, male Wistar rats underwent 4 h of Sham (n = 8), Unilateral (n = 8), or Bilateral (n = 8) hindlimb ischemia followed by 4 h of reperfusion. All rats underwent ³¹P magnetic resonance spectroscopy of their right gastrocnemius muscle to determine various high-energy phosphate ratios including ATP to P(i) (ATP/P(i), a measure of energy status) and phosphocreatine to P(i) (PCr/P(i), a measure of thermodynamic capacity). Gastrocnemius muscles were then harvested to determine muscle damage and complement membrane attack complex (MAC) deposition by immunohistochemical staining [grade 0 (none) to 3 (very severe)] and to measure glutathione (GSH), DNA, and enzyme activities: β-hydroxyacyl-CoA dehydrogenase, phosphofructokinase, and citrate synthetase. Results. HIR was associated with significant declines in ATP/P(i) and PCr/P(i) (P < 0.001). Progressively more severe HIR (Sham, Unilateral, Bilateral) was associated with greater MAC deposition (0.0 ± 0.0, 1.0 ± 0.3, 1.5 ± 0.4, P = 0.06, mean ± SEM) and histological damage (0.0 ± 0.0, 0.9 ± 0.3, 1.3 ± 0.4, P < 0.05). GSH levels, β-hydroxyacyl-CoA dehydrogenase, and citrate synthetase activities were not affected by HIR, but phosphofructokinase activity increased (24.09 ± 2.42, 35.16 ± 5.26, 59.29 ± 9.82 mmol/mg of DNA/min, P < 0.05). Although GSH levels were not significantly altered, complement deposition was closely associated with skeletal muscle injury and compensatory changes in glycolysis. Alterations in myocellular bioenergetics after HIR closely paralleled complement deposition rather than GSH depletion. Conclusions. Therapeutic strategies aimed at controlling complement activity and assessment techniques based on bioenergetics may allow more precise determinations of the effects of HIR injury.