目的：探讨依达拉奉通过微小RNA－25（microRNA－25， miR－25）对高糖诱导的人神经母细胞瘤SH－SY5Y细胞凋亡的抑制作用及其机制。方法：将SH－SY5Y细胞用含高浓度葡萄糖的DMEM培养基和依达拉奉的联合培养液共同培养24 h。MTT比色法测定SH－SY5Y细胞存活率；DCFH－DA荧光探针法检测SH－SY5Y细胞中活性氧簇（ROS）的水平；采用流式细胞术检测SH－SY5Y细胞的凋亡率；Western blot法检测凋亡相关蛋白Bax和Bcl－2的表达水平；实时定量PCR检测细胞中miR－25的表达水平。为进一步阐明依达拉奉抑制高糖诱导的神经细胞凋亡的作用靶点，我们将miR－25抑制剂应用于细胞，之后采用caspase－3凋亡试剂盒检测细胞的凋亡率。结果：与对照组相比，高糖诱导后细胞存活率明显降低，细胞中的ROS水平和细胞凋亡率明显升高，Bax的表达明显增加， Bcl－2的表达明显降低， miR－25的表达水平也明显降低。给予依达拉奉治疗之后，细胞存活率明显升高，ROS含量和细胞凋亡率明显降低，Bax的蛋白水平明显降低，Bcl－2蛋白水平明显升高，miR－25的表达水平亦明显升高。进一步给予miR－25抑制剂后，caspase－3的水平明显升高，此时同时给予依达拉奉后并不能抑制高糖引起的神经细胞的凋亡。结论：依达拉奉对高糖诱导的SH－SY5Y细胞凋亡具有抑制作用，其作用靶点可能是miR－25。%AIM:To observe the effects of edaravone on high glucose-induced apoptosis of SH-SY5Y cells and its potential mechanism .METHODS:The SH-SY5Y cells were cultured in the DMEM medium with 100 mmol/L glucose and 100μmol/L edaravone for 24 h.The viability of the SH-SY5Y cells was detected by MTT assay .The levels of ROS in the cells were determined by DCFH-DA fluorescent probing .The apoptotic rates of the cells were analyzed by flow cytome-try.The protein expression of Bax and Bcl-2 in the cells were detected by Western blot .The expression levels of micro-RNA-25 (miR-25) were determined by real-time PCR.To further clarify the target sites of edaravone on inhibiting apopto-sis induced by high glucose , miR-25 inhibitor was applied to the SH-SY5Y cells and the activity of caspase-3 was meas-ured.RESULTS:Compared with control group , the cell viability was decreased significantly in model group , and the ROS level was increased significantly .The protein expression of Bax was up-regulated significantly , while the expression levels of Bcl-2 and miR-25 were significantly down-regulated .Compared with model group , the cell viability was increased signifi-cantly in edaravone group .The ROS level was decreased significantly .Meanwhile, the expression of Bax was down-regula-ted, while the expression of Bcl-2 and miR-25 was up-regulated with statistical significance .The caspase-3 activity of the cells incubated with 100 mmol/L glucose and miR-25 inhibitor was increased .However, no alteration of caspase-3 activity with edaravone added simultaneously was observed .CONCLUSION: Edaravone inhibits the apoptosis of SH-SY5Y cells induced by high glucose with the potential target site of miR-25.
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