ACTIVATED PROTEIN C ATTENUATES SKELETAL MUSCLE ISCHAEMIA REPERFUSION INJURY THROUGH A DIRECT INHIBITORY EFFECT ON NEUTROPHIL ACTIVATION

Abstract

Aims: Pharmocological modulation of skeletal muscle reperfusion injury after an ischaemic insult may improve limb salvage rates and prevent the associated systemic sequelae. Activated Protein c (APC) is an endogenous anti-coagulant with anti-inflammatory properties. The purpose of our study was to evaluate the effects of APC on skeletal muscle ischaemia reperfusion injury and to examine the direct effects of APC on neutrophil activation.

Methods: Adult male Sprague Dawley rats (n=30) were randomised into three groups: control group, I/R group treated with normal saline and I/R treated with APC. Bilateral hind-limb ischaemia was induced by rubber ban application proximal to the level of the greater trochanters for two hours. Treatment groups received either normal saline or APC prior to tourniquet release. Following twelve hours reperfusion, the tibialis anterior was dissected and muscle function assessed electrophysiologically by electrical field stimulation. The animals were then sacrificed and skeletal muscle harvested for evaluation. Skeletal muscle injury was assessed based on myeloperoxidase content, wet-to-dry ratio and histological analysis. The effect of APC on TNF-α stimulated human peripheral blood neutrophils was also examined by measuring CD 18 expression and reactive oxygen species (ROS) generation.

Results: APC significantly attenuated skeletal muscle reperfusion injury as shown by reduced myeloperoxidase content, wet-to-dry ratio and electrical properties of skeletal muscle. These findings were supported by our histological findings. Our in-vitro work demonstrated a reduction in CD 18 expression and ROS generation.

Conclusion: Activated Protein C may have a protective role in the setting of skeletal muscle ischaemia reperfusion injury and this is in part mediated by a direct inhibitory effect on neutrophil activation.