Establishment of rat ischemia-reperfusion injury model

A total of 40 male SD rats (180–220 g, 6 weeks, SPF grade) were purchased from the Laboratory Animal Center of Peking University Medical Department. All rats were randomly divided into Con group (n=10), I/R group (n=10), miR group (n=10), and IR-C group (n=10). Rats were generally anesthetized by sodium pentobarbital (50 mg/kg), and made every effort to reduce pain during operation. Based on a rapid thoracotomy to expose the heart, the left coronary artery was ligated and compressed with a water-filled balloon (Medtronic, Inc, Santa Rosa, CA, USA) for 20 minutes to cause anterior myocardial ischemia. Then, the water in the balloon was sucked out to relieve the pressure on the blood vessel, and the elevated ST-segment (by electrocardiogram) returned to normal or decreased more than 50% was considered as the successful establishment of I/R injury model. Finally, 2 hours reperfusion was performed on rats. Rats in Con group were placed suture without ligation. Rats in miR group were injected with miR-484 agomir (1×10³mol/L; GenePharma, Shanghai, China) in myocardium at 24 hours before the establishment of MI/R injury model (5 points were randomly selected on the surface of the myocardium, and 10 μL miR-484 agomir were given at each point). Meanwhile, the negative control (NC) sequence was injected into the myocardium at 24 hours before the establishment of I/R model in the IR-C group (5 points were randomly selected on the surface of the myocardium, and 10 μL miR-484 agomir were given at each point). All the experimental schemes were approved by the Ethics Committee of China Medical University (Shenyang, China; permit No. SCXK 2019-0023), and all experiments were in accordance with the guide for the care and use of laboratory animals established by United States National Institutes of Health (Bethesda, MD, USA).