Circpostn shrna

The AC16 cell lines (ventricular cardiomyocyte of humans) were obtained from the American Type Tissue Culture Collection. The cells were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM; Solarbio, China) containing 10% fetal bovine serum (Gibco, United States), 0.1 mg/ml streptomycin (Solarbio, China), and 100 units/ml penicillin (Solarbio, China) at a condition of 37°C with 5% CO₂. The cell injury model was established by hypoxia and reoxygenation (H/R). The cells without H/R served as control. The cells were cultured in a medium without glucose and fetal bovine serum for 6 h at a hypoxia condition of 5% CO₂ and 0.1% O₂ and then were reoxygenated in DMEM containing normal glucose and 10% fetal bovine serum for 12 h. The circPostn shRNA, control shRNA, miR-96-5p mimic, control mimic, miR-96-5p inhibitor, control inhibitor, and the pcDNA3.1-BNIP3 overexpression vector were obtained (GenePharma, China). The transfection in the cells was performed by Liposome 3000 (Invitrogen, United States) according to the manufacturer’s instructions.

To establish the MI mouse model, the C57BL/6 mice (4–6 months old, male, 25–30 g) were intraperitoneally injected with ketamine (120 mg/kg) and xylazine (5 mg/kg body weight) for anesthetization. The left anterior descending coronary artery occlusion/reperfusion (LAD/reperfusion) was performed. Briefly, we placed the LAD on the heart surface by applying an anatomy microscope and ligated the LAD for 30 min, followed by the restoration of blood flow. Sham mice were conducted the surgery without occlusion of LAD. We injected the lentiviral vectors comprising shRNA of circPostn (Genechem, China) (1 × 10⁷ TU/mice) or the lentivirus comprising control shRNA in the mice ventricular cavity, followed by the construction of the MI mouse model. Then, the heart tissues and blood were collected from mice for further analysis. The cardiac injury was assessed by hematoxylin and eosin (H&E) staining. The cardiac function was analyzed by echocardiography measurement with an ultrasound system (Panoview, China) furnished with a 30-MHz phased-array transducer in 3-day post-MI mice. The circPostn shRNA, control shRNA, miR-96-5p mimic, control mimic, miR-96-5p inhibitor, control inhibitor, and the pcDNA3.1-BNIP3 overexpression vector were obtained (GenePharma, China). Animal care and method procedure were authorized by the Animal Ethics Committee of PLA General Hospital.