miR-17-5p调控PTEN对大鼠中动脉阻塞的神经保护作用研究

Neuroprotective effects of microRNA-17-5p on MCAO rats by regulating PTEN

目的探究microRNA-17-5p(miR-17-5p)在中动脉阻塞(MCAO)中的神经保护作用及其可能机制。方法将60只清洁级雄性SD大鼠随机分为假手术组(Sham组)、模型组(MCAO组)、MCAO+mimic组、MCAO+inhibitor组和MCAO+BPV组,每组12只。采用线栓法建立MCAO大鼠模型。建模24 h后,检测大鼠神经功能和脑组织含水量;HE染色观察大鼠脑组织病理学损伤;TUNEL染色检测大鼠脑组织神经元细胞凋亡情况;RT-PCR检测miR-17-5p和第10号染色体缺失的磷酸酶及张力蛋白同源蛋白(PTEN)的mRNA表达水平,预测两者是否存在连续的结合位点;Western blot检测脑组织通路蛋白磷酸化水平。结果MCAO+mimic组和MCAO+BPV组大鼠神经功能评分、脑组织含水量、神经细胞凋亡率、PTEN的mRNA和蛋白表达水平均低于MCAO组,差异有统计学意义(P<0.05);MCAO+mimic组和MCAO+BPV组脑组织病理学损伤明显改善,miR-17-5p和PTEN存在连续结合位点,而这2组的miR-17-5p表达水平、AKT和mTOR磷酸化水平高于MCAO组,差异有统计学意义(P<0.05);MCAO+inhibitor组神经功能评分、脑组织含水量、神经细胞凋亡率、PTEN的mRNA和蛋白表达水平高于MCAO组,差异有统计学意义(P<0.05);MCAO+inhibitor组脑组织病理学损伤加重,同时AKT和mTOR磷酸化水平低于MCAO组。结论miR-17-5p表达上调对MCAO大鼠神经具有保护作用,其功能发挥可能与介导调控PTEN信号通路相关。

Objective To study the neuroprotective effect of microRNA-17-5p(miR-17-5p)on the middle cerebral artery occlusion(MCAO)and its possible mechanism.Methods Sixty male SD rats were randomly divided into sham operation group(Sham group),model group(MCAO group),MCAO+mimic group,MCAO+inhibitor group and MCAO+BPV group,with 12 rats in each group.Thread embolism method was used to establish MCAO rat model,brain water content and neurological function were detected after 24 hours;HE staining was used to observe the pathological changes of rat brain;TUNEL staining was used to detect neuronal apoptosis in brain tissue of rats.Predicting the existence of continuous binding sites of miR-17-5p and PTEN;RT-PCR was used to detect the expression level of miR-17-5p and the mRNA level of phosphatase and tensin homolog deleted on chromosome ten(PTEN);Western blot was used to detect the phosphorylation level of pathway protein in brain tissue.Results Compared with the MCAO group,the neurological function score,brain water content,neuronal apoptosis rate,the mRNA and protein levels of PTEN were significantly decreased in MCAO+mimic group and MCAO+BPV group,the differences were significant(P<0.05),and the brain histopathological damage was significantly improved.There were continuous binding sites between miR-17-5p and PTEN,while the expression levels of miR-17-5p,and the phosphorylation levels of pathway protein in MCAO+mimic group and MCAO+BPV group were significantly increased,the differences were significant(P<0.05).The neurological function score,brain water content,neuronal apoptosis rate,the mRNA and protein levels of PTEN in MCAO+inhibitor group were all higher than those in the MCAO group,the differences were significant( P <0.05),while the brain histopathological damage was aggravated,and the phosphorylation levels of pathway protein in MCAO+inhibitor group were lower than those in MCAO group. Conclusion The up-regulation of the expression of miR-17-5p has neuroprotective effect on MCAO rats,and its function may be related to the regulation of PTEN signaling pathway.