MicroRNAs have been implicated as critical regulatory molecules in many cerebrovascular diseases. Recent studies demonstrated miR-22 might provide a potential neuroprotective effect. However, the neuroprotective effect of miR-22 in ischemia/reperfusion (I/R) injury has not been thoroughly elucidated. In this study, the PC12 cells were subjected to 4 h oxygen and glucose deprivation (I) and 24 h reoxygenation (R). The PC12 cells were pre-transfected with miR-22 or anti-miR-22 or siRNA-mediated downregulation of p53-upregulated-modulator-of-apoptosis (PUMA)(PUMA siRNA) or their controls at 24 h prior to exposure to I/R. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and Western blot were employed to analyze mRNA and protein expression. PI and Annexin V assays and terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling (TUNEL) assay were used to quantify the rate of apoptosis. We found that miR-22 expression was significantly downregulated in the PC12 cells subjected to I/R. Loss of function of miR-22 increased PC12 apoptosis after I/R, and overexpression of miR-22 decreases PC12 apoptosis after I/R. PUMA protein was upregulated in the I/R group as compared with the sham group. The increased PUMA protein expression and apoptosis induced by I/R was reversed by transfection with PUMA siRNA. We concluded that I/R enhanced apoptosis and PUMA expression in PC12 cells via downregulation of miR-22. Enhanced miR-22 expression reversed both PUMA expression and apoptosis induced by I/R in PC12 cells. miR-22/PUMA axis has important implications for their clinical applications.

中文翻译：

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miR-22通过靶向p53上调的凋亡调节剂（PUMA）保护PC12免受缺血/再灌注诱导的损伤。

在许多脑血管疾病中，MicroRNA被认为是关键的调节分子。最近的研究表明，miR-22可能提供潜在的神经保护作用。但是，尚未完全阐明miR-22在缺血/再灌注（I / R）损伤中的神经保护作用。在这项研究中，对PC12细胞进行了4小时的氧和葡萄糖剥夺（I）和24小时的再氧化（R）。在暴露于I的24小时前，将PC12细胞用miR-22或抗miR-22或siRNA介导的p53上调的细胞凋亡调节剂（PUMA）（PUMA siRNA）或它们的对照预转染。 / R。采用逆转录定量聚合酶链反应（RT-qPCR）和蛋白质印迹法分析mRNA和蛋白质表达。PI和膜联蛋白V测定法和末端脱氧核苷酸转移酶介导的dUTP-生物素缺口末端标记（TUNEL）测定法用于定量凋亡率。我们发现miR-22表达在受到I / R作用的PC12细胞中显着下调。miR-22的功能丧失增加I / R后PC12的凋亡，miR-22的过表达降低I / R后的PC12凋亡。与假手术组相比，I / R组的PUMA蛋白上调。通过PUMA siRNA转染可逆转I / R诱导的增加的PUMA蛋白表达和凋亡。我们得出的结论是，I / R通过下调miR-22增强PC12细胞的凋亡和PUMA表达。增强的miR-22表达逆转了PC12细胞中I / R诱导的PUMA表达和细胞凋亡。