Effects of sinomenine on hind limb ischemia-reperfusion injury and expression of Bcl-2 and Bax in skeletal muscle cells of rats

Abstract

Objective To investigate the effects of sinomenine on hind limb ischemia-reperfusion (I/R) injury and expression of Bcl-2 and Bax in skeletal muscle cells of rats. Methods Fifty-four healthy adult male Wistar rats, aged 6-8 weeks, weighing 180-220 g, were divided into 3 groups (n=18 each) using a random number table: sham operation group (group S), group I/R and sinomenine group (group SIN). The rats were subjected to 4 h of ischemia on the proximal part of the right hind limb using elastic rubber bands followed by reperfusion in I/R and SIN groups.Sinomenine 60 mg/kg was injected intraperitoneally at 30 min before reperfusion in group SIN, and the equal volume of normal saline was given instead of sinomenine at 30 min before reperfusion in S and I/R groups.Immediately after onset of reperfusion and at 4 and 24 h of reperfusion, blood samples were collected from the cardiac apex to measure the concentrations of serum lactate dehydrogenase (LDH) and creatine kinase (CK). The animals were sacrificed immediately after blood sampling, and the gastrocnemius specimens of the hind limb were immediately removed for determination of the wet to dry weight ratio (W/D ratio) and expression of Bcl-2 and Bax in gastrocnemius cells (by immunohistochemistry) and for examination of the pathological changes after haematoxylin and eosin staining.The Bcl-2/Bax ratio was calculated. Results Compared with group S, the gastrocnemius W/D ratio and concentrations of serum LDH and CK were significantly increased, the expression of Bcl-2 was significantly down-regulated, the expression of Bax was significantly up-regulated, and the Bcl-2/Bax ratio was significantly decreased in I/R and SIN groups (P<0.05). Compared with group I/R, the gastrocnemius W/D ratio and concentrations of serum LDH and CK were significantly decreased, the expression of Bcl-2 was significantly up-regulated, the expression of Bax was significantly down-regulated, and the Bcl-2/Bax ratio was significantly increased in group SIN (P<0.05). The pathological changes of the gastrocnemius were significantly attenuated in group SIN as compared with group I/R. Conclusion Sinomenine can attenuate hind limb I/R injury, and the mechanism may be related to maintenance of the balance between Bcl-2 and Bax and to inhibition of apoptosis in skeletal muscle cells of rats. Key words: Sinomenium; Reperfusion injury; Extremities; Proto-oncogene proteins c-bcl-2; bcl-2-associated X protein