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目的：观察非对称性二甲基精氨酸（ADMA）对内皮细胞中microRNA-21(miR-21)表达的影响，探讨microRNA-21 在ADMA 诱导的内皮细胞衰老中的作用。方法: 人脐静脉内皮细胞（HUVEC）与10 uM 的ADMA 孵育48 小时后收集细胞提取总RNA 及蛋白，荧光定量实时RT-PCR 检测miR-21 表达，Western blot 检测超氧化物歧化酶2 （SOD2）表达，衰老相关半乳糖苷酶（SA-β-gal）染色鉴定衰老的内皮细胞；然后HUVEC 与miR-21 抑制剂转染6 小时后继续与10 uM 的ADMA 孵育48 小时留取细胞按上述方法检测相关指标。结果：HUVEC 与ADMA 孵育后miR-21 表达量明显增加（P＜0.01），同时衰老的内皮细胞数量增多（P＜0.05），而SOD2 表达减少（P＜0.01）；MiR-21 抑制剂转染HUVEC 后ADMA 诱导的miR-21 表达明显减少，同时衰老的内皮细胞减少，而SOD2 表达明显增加（所有P＜0.01）。结论：ADMA 诱导了HUVEC 中miR-21 表达及细胞衰老，miR-21 介导了 ADMA 诱导的内皮细胞衰老作用，其机制可能与其抑制SOD2 表达有关。

英文摘要:

Objective: To investigate the effects of asymmetric dimethylarginine (ADMA) on the expression of microRNA-21 (miR-21) in endothelial cells in order to explore the role of miR-21 in ADMA-induced endothelial cell senescence. Methods: After human umbilical vein endothelial cells (HUVECs) were incubated with 10 uM ADMA for 48 hours, the cells were collected for extraction of total RNA and protein. Fluorescence quantitative real-time PCR was applied to determine the expression of miR-21 in HUVECs. Western Blot was used to detect the expression of superoxide dismutase 2 (SOD2) in HUVECs, and senescence associated galactosidase (SA-β-gal) staining was employed to identify the senescent endothelial cells. Then after the transfection of miR-21 inhibitors for 6 hours, HUVEC were incubated with 10 uM ADMA for 48 hours, and then the cells were collected for the determination of miR-21 and SOD2 expression and senescent endothelial cells according to the above-mentioned methods. Results: After HUVECs were incubated with ADMA, the expression of miR-21 was increased significantly (P＜0.01) and meanwhile the number of SA-β-gal positive endothelial cells were elevated (P＜0.05) , while the SOD2 expression decreased (P＜0.01); After the transfection of miR-21 inhibitors the ADMA-induced miR-21 expression in HUVECs was significantly reduced and meanwhile the number of SA-β-gal positive endothelial cells decreased while the SOD2 expression was significantly increased (all P＜0.01). Conclusions: ADMA induced the expression of miR-21 in HUVECs and endothelial cell senescence, further miR-21 mediated the endothelial cell senescence induced by ADMA. This mechanism might be related to the inhibition of miR-21 on SOD2 expression.

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