Effect of remote ischemic preconditioning on reperfusion arrhythmia in rats and the role of Shh/Gli1 signaling pathway

Abstract

Objective To evaluate the effect of remote ischemic preconditioning (RIPC) on reperfusion arrhythmia and the role of Sonic hedgehog (Shh)/glioma associated oncogene homolog 1 (Gli-1) signaling pathway in rats. Methods Forty clean-grade healthy male SD rats, aged 6-8 weeks, weighing 220-280 g, were divided into 4 groups (n=10 each) using a random number table method: sham operation group (Sham group), myocardial I/R group (I/R group), remote ischemic preconditioning group (RIPC group) and cyclopamine + remote ischemic preconditioning group (CYC+ RIPC group). Myocardial I/R was produced by occlusion of left anterior descending branch of coronary artery for 30 min followed by 2 h reperfusion in anesthetized rats.In RIPC group, an elastic rubber tourniquet was used to block the lower limb blood flow for 10 min in the right groin and then released for 30-min reperfusion, and then the myocardial I/R model was established.In CYC+ RIPC group, a Shh specific inhibitor cyclopamine 10 mg/kg was intraperitoneally injected at 30 min before ischemia, and then RIPC was performed.The development of ventricular arrhythmia was recorded.Blood samples were taken at 120 min of reperfusion for determination of the concentrations of serum creatine kinase-MB (CK-MB) and cardiac troponin I (cTnI) using a 7600 automatic biochemical analyzer.The myocardial tissues in the ischemic area were taken after collecting blood for examination of pathological changes and for determination of tumor necrosis factor α (TNF-α) content (by enzyme-linked immunosorbent assay), cell apoptosis (using TUNEL), expression sites of Shh and Gli1 (by immunofluorescence) and expression of Shh and Gli1 (by Western blot). AI was calculated. Results Compared with Sham group, the incidence and times of arrhythmia were significantly increased, the total duration of arrhythmia was prolonged, the arrhythmia score was increased, the serum cTnI and CK-MB concentrations, myocardial TNF-α content and AI were increased, the expression of Shh and Gli1 was up-regulated (P 0.05). Compared with group RIPC, the incidence and times of arrhythmia were significantly increased, the total duration of arrhythmia was prolonged, the serum cTnI and CK-MB concentrations, myocardial TNF-α content and AI were increased, and the expression of Shh and Gli1 was down-regulated in group CYC+ RIPC (P<0.05). Conclusion RIPC can reduce reperfusion arrhythmia, and the mechanism is related to activating Shh/Gli1 signaling pathway in rats. Key words: Ischemic preconditioning; Extremities; Hedgehog proteins; Myocardial reperfusion injury; Arrhythmias, cardiac