Department of Human Anatomy, College of Basic Medical Sciences, China Medical University, Shenyang, China.
PLoS One. 2022 Aug 2;17(8):e0272482. doi: 10.1371/journal.pone.0272482. eCollection 2022.
DNA methylation regulates epigenetic gene expression in ischemic stroke. Decitabine attenuates ischemic stroke by inhibiting DNA methylation. However, the underlying mechanism of this effect is not known. A model of ischemic stroke in Sprague-Dawley rats was induced through middle cerebral artery occlusion followed by reperfusion step. The rats were randomly treated with decitabine or vehicle by a one-time intraperitoneal injection. Sham rats received similar treatments. Four days after treatment, the rats were perfused with saline or 4% paraformaldehyde after which the brain was excised. DNA methylation level and brain infarct volume were determined by dot blot and histochemistry, respectively. The cellular co-localization and quantitative analysis of DNA methylation were assessed by immunohistochemistry and expression levels of cdkn1b (p27) mRNA and protein were measured by qRT-PCR and immunohistochemistry, respectively. The proliferation of astrocytes and number of neurons were determined by immunohistochemistry. Rats treated with decitabine showed hypomethylation and reduced infarct volume in the cortex. DNA methylation was decreased in astrocytes. Decitabine upregulated p27 mRNA and protein expression levels and attenuated the proliferation of astrocytes in vivo and vitro. Decitabine promotes p27 gene expression possibly by inhibiting its DNA methylation, thereby decreases the proliferation of astrocytes, neuronal death and infarct volume after ischemic stroke.
DNA 甲基化调控缺血性卒中的表观基因表达。地西他滨通过抑制 DNA 甲基化来减轻缺血性卒中。然而，其作用机制尚不清楚。通过大脑中动脉闭塞和再灌注来诱导 Sprague-Dawley 大鼠的缺血性卒中模型。大鼠通过一次性腹腔注射随机接受地西他滨或载体处理。假手术大鼠接受类似处理。治疗 4 天后，用生理盐水或 4%多聚甲醛灌注大鼠，然后取出大脑。通过斑点印迹和组织化学分别测定 DNA 甲基化水平和脑梗死体积。通过免疫组织化学评估 DNA 甲基化的细胞共定位和定量分析，通过 qRT-PCR 和免疫组织化学分别测量 cdkn1b(p27)mRNA 和蛋白的表达水平。通过免疫组织化学测定星形胶质细胞的增殖和神经元数量。用地西他滨处理的大鼠表现出皮质区低甲基化和梗死体积减小。星形胶质细胞中的 DNA 甲基化减少。地西他滨上调 p27mRNA 和蛋白表达水平，并在体内和体外减轻星形胶质细胞的增殖。地西他滨可能通过抑制其 DNA 甲基化促进 p27 基因表达，从而减少缺血性卒中后的星形胶质细胞增殖、神经元死亡和梗死体积。
