Objective To evaluate the role of high-mobility group box 1 protein ( HMGB1)∕Toll-like receptor 4 ( TLR4) signaling pathway in myocardial ischemia-reperfusion ( I∕R) injury in rats. Meth-ods Fifty-four clean-grade healthy male Sprague-Dawley rats, aged 7-8 weeks, weighing 200-250 g, were divided into 3 groups ( n=18 each ) using a random number table method: sham operation group ( group Sham) , myocardial I∕R group ( group I∕R) and specific HMGB1 antibody group ( group H) . Myo-cardial I∕R was produced by 30-min occlusion of left anterior descending branch of coronary artery followed by 180-min reperfusion in anesthetized rats. Specific HMGB1 antibody 2 mg∕kg was injected through the femoral vein at 30 min of reperfusion in group H. Twelve rats in each group were randomly selected at 180 min of reperfusion, and blood samples were collected from the femoral vein for determination of plasma in-terleukin-6 ( IL-6) , IL-8, IL-12, tumor necrosis factor-alpha ( TNF-α) and cardiac troponin I ( cTnI ) concentrations. The rats were then sacrificed, hearts were removed and myocardial tissues were obtained forexamination of the pathological changes and for determination of the expression of intercellular adhesion mol-ecule 1 ( ICAM-1) and E-selectin ( by immunohistochemistry) , expression of TLR4 and NF-κB mRNA and protein (by quantitative real-time polymerase chain reaction or by Western blot), activities of glutathione peroxidase ( GSH-PX) , superoxide dismutase ( SOD) and myeloperoxidase ( MPO) and MDA content. Six rats were selected for measurement of the myocardial infarct volume, and the percentage of myocardial in-farct volume was calculated. Results Compared with group Sham, the serum concentrations of IL-6, IL-8, IL-12, TNF-α and cTnI, MPO activity and MDA content were significantly increased, the expression of ICAM-1, E-selectin, TLR4 and NF-κB protein and mRNA was up-regulated, activities of GSH-PX and SOD were decreased, and the percentage of myocardial infarct volume was increased in group I∕R ( P<0. 01) . Compared with group I∕R, the serum concentrations of IL-6, IL-8, IL-12, TNF-α and cTnI, MPO activity and MDA content were significantly decreased, the expression of ICAM-1, E-selectin, TLR4 and NF-κB protein and mRNA was down-regulated, activities of GSH-PX and SOD were increased, and the percentage of myocardial infarct volume was decreased in group H ( P<0. 01) . Conclusion HMGB1∕TLR4 signaling pathway is involved in myocardial I∕R injury in rats.