Key Laboratory for Biotechnology on Medicinal Plants of Jiangsu Province, School of Life Science, Jiangsu Normal University, Xuzhou, P. R. China.
Department of Neurology, The Fourth Affiliated Hospital of China Medical University, Shenyang, P. R. China.
J Cell Physiol. 2018 Sep;233(9):6632-6643. doi: 10.1002/jcp.26297. Epub 2018 Apr 11.
This study aims to investigate the effect of microRNA-129 (miR-129) on proliferation and apoptosis of hippocampal neurons in epilepsy rats by targeting c-Fos via the MAPK signaling pathway. Thirty rats were equally classified into a model group (successfully established as chronic epilepsy models) and a normal group. Expression of miR-129, c-Fos, bax, and MAPK was detected by RT-qPCR and Western blotting. Hippocampal neurons were assigned into normal, blank, negative control (NC), miR-129 mimic, miR-129 inhibitor, siRNA-c-Fos, miR-129 inhibitor+siRNA-c-Fos groups. The targeting relationship between miR-129 and c-Fos was predicted and verified by bioinformatics websites and dual-luciferase reporter gene assay. Cell proliferation after transfection was measured by MTT assay, and cell cycle and apoptosis by flow cytometry. c-Fos is a potential target gene of miR-129. Compared with the normal group, the other six groups showed a decreased miR-129 expression; increased expression of expression of c-Fos, Bax, and MAPK; decreased proliferation; accelerated apoptosis; more cells arrested in the G1 phase; and fewer cells arrested in the S phase. Compared with the blank and NC groups, the miR-129 mimic group and the siRNA-c-Fos group showed decreased expression of c-Fos, Bax, and MAPK, increased cells proliferation, and decreased cell apoptosis, fewer cells arrested in the G1 phase and more cells arrested in the S phase. However, the miR-129 inhibitor groups showed reverse consequences. This study suggests that miR-129 could inhibit the occurrence and development of epilepsy by repressing c-Fos expression through inhibiting the MAPK signaling pathway.
本研究旨在通过靶向 MAPK 信号通路探讨 microRNA-129(miR-129)对癫痫大鼠海马神经元增殖和凋亡的影响。30 只大鼠等分为模型组(成功建立慢性癫痫模型)和正常组。采用 RT-qPCR 和 Western blot 检测 miR-129、c-Fos、bax 和 MAPK 的表达。将海马神经元分为正常组、空白组、阴性对照组(NC)、miR-129 模拟物组、miR-129 抑制剂组、siRNA-c-Fos 组、miR-129 抑制剂+siRNA-c-Fos 组。通过生物信息学网站和双荧光素酶报告基因检测预测和验证 miR-129 与 c-Fos 的靶向关系。MTT 法检测转染后细胞增殖情况,流式细胞术检测细胞周期和凋亡情况。c-Fos 是 miR-129 的潜在靶基因。与正常组相比,其余 6 组 miR-129 表达下调;c-Fos、Bax 和 MAPK 表达上调;细胞增殖减少;细胞凋亡加速;更多细胞停滞在 G1 期;更少细胞停滞在 S 期。与空白组和 NC 组相比,miR-129 模拟物组和 siRNA-c-Fos 组 c-Fos、Bax 和 MAPK 表达下调,细胞增殖增加,细胞凋亡减少,G1 期细胞减少,S 期细胞增加。但是,miR-129 抑制剂组则呈现相反的结果。本研究表明,miR-129 可能通过抑制 MAPK 信号通路抑制 c-Fos 表达从而抑制癫痫的发生和发展。