MicroRNA-1298 is regulated by DNA methylation and affects vascular smooth muscle cell function by targeting connexin 43
MicroRNA-1298 is regulated by DNA methylation and affects vascular smooth muscle cell function by targeting connexin 43
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MicroRNA-1298 受 DNA 甲基化调节，通过靶向连接蛋白 43 影响血管平滑肌细胞功能
DOI：
10.1093/cvr/cvv160
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发表时间：
2015-09-01
影响因子：
10.8
通讯作者：
Wang, Shenming
中科院分区：
文献类型：
--
作者：
Hu, Wei;Wang, Mian;Wang, Shenming
越来越多的证据将microRNA与外周血管疾病的过程联系起来。最近，我们发现microRNA-1298（miR-1298）是下肢动脉硬化闭塞症（阿索）患者动脉中表达下调最显著的microRNA之一。然而，关于它在阿索过程中的作用知之甚少。本研究旨在探讨miR-1298在阿索中的表达、调控机制及功能。方法与结果采用定量逆转录PCR和原位杂交技术，发现miR-1298在阿索中主要表达于血管平滑肌细胞（VSMCs），与正常动脉相比表达下调。焦磷酸测序分析显示，与正常动脉相比，阿索中CpG位点上游的miR-1298 DNA高甲基化。接下来，荧光素酶报告基因分析揭示miR-1298的下调与上游DNA CpG位点的高甲基化有关。向培养的VSMC中引入miR-1298模拟物显著减弱了细胞增殖和迁移。连接蛋白43（Cx43）被证实是miR-1298的功能性靶点，其参与miR-1298介导的细胞效应。最后，慢病毒介导的miR-1298及其靶点Cx43在大鼠颈动脉球囊损伤模型中的表达表明，miR-1298的再过表达通过靶向连接蛋白43而显著减少新生内膜的形成。结论我们的数据表明上游DNA甲基化/miR-1298/Cx43通路在调节VSMC功能中的特异性作用，并且提示miR-1298的调节可能是通过调节VSMC的功能来实现的。1298水平可能为阿索提供一种新的治疗方法。
Aims Growing evidence links microRNA to the process of peripheral vascular disease. Recently, we have found that microRNA-1298 (miR-1298) is one of the most significantly down-regulated microRNAs in human arteries with arteriosclerosis obliterans (ASO) of the lower extremities. However, little is known regarding its role in the process of ASO. The present study aimed to investigate the expression, regulatory mechanisms, and functions of miR-1298 in the process of ASO.Methods and results Using quantitative reverse-transcription PCR and in situ hybridization assays, miR-1298 was observed predominantly expressed in the vascular smooth muscle cells (VSMCs) and was significantly down-regulated in ASO compared with normal arteries. Pyrosequencing analysis revealed that the miR-1298 DNA upstream of CpG sites were hypermethylated in ASO compared with normal arteries. Next, the luciferase reporter assay revealed that miR-1298 down-regulation is related with upstream DNA CpG site hypermethylation. Introducing a miR-1298 mimic into cultured VSMCs significantly attenuated cell proliferation and migration. Connexin 43 (Cx43) was validated to be a functional target of miR-1298 that was involved in the miR-1298-mediated cellular effects. Finally, lentivirus-mediated delivery of miR-1298 and its target Cx43 into a rat carotid balloon injury model indicated that re-overexpression of miR-1298 significantly decreased neointimal formation by targeting connexin 43.Conclusion Our data demonstrate a specific role of the upstream DNA methylation/miR-1298/Cx43 pathway in regulating VSMC function and suggest that modulation of miR-1298 levels may offer a novel therapeutic approach for ASO.