Wang Ye, Wang Desheng, Guo Dawen
Department of Neurology, First Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang, 150001, China.
Department of Clinical Laboratory, First Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang, 150001, China.
PLoS One. 2016 Jan 8;11(1):e0146646. doi: 10.1371/journal.pone.0146646. eCollection 2016.
Some recent studies suggest that multiple miRNAs might regulate neurogenic transdifferentiation of mesenchymal stromal cells (MSCs). In the present study, we hypothesized that the miR-124 can repress the expression of RhoA upon the neurogenesis of adipose derived MSCs (ADMSCs).
MiRNA expression dynamics during neurogenic transdifferentiation of ADMSCs were measured. The expression of neuron-specific enolase (NSE), Tuj-1 (Neuron-specific class III beta-tubulin) and glial fibrillary acidic protein (GFAP), as well as electrophysiological properties, were detected after neurogenic transdifferentiation. The targeting of miR-124 over RhoA was verified by dual luciferase assay, qRT-PCR and western blot. The functions of miR-124 and the RhoA/ROCK signaling pathway were studied using gain and loss of function experiments in vitro.
MiR-124 is significantly upregulated during neurogenic transdifferentiation of ADMSCs. Knockdown of endogenous miR-124 hampered neurogenic transdifferentiation and the acquired electrophysiological properties. MiR-124 could directly target RHOA mRNA and repress its expression, through which it increased the proportion of transdifferentiated (transdiff.) cells with positive NSE, Tuj-1 and GFAP. RhoA/ROCK1, but not ROCK2 is a downstream signaling pathway of miR-124 in the process of transdifferentiation.
MiR-124 is an important miRNA modulating neurogenic transdifferentiation of ADMSCs at least partly via the miR-124/RhoA/ROCK1 signaling pathway. These findings provided some fundamental information for future use of ADMSCs as an agent for regenerative medicine and cell therapy for neurological diseases.
近期一些研究表明，多种微小RNA（miRNA）可能调控间充质基质细胞（MSC）的神经源性转分化。在本研究中，我们假设miR-124在脂肪来源的间充质干细胞（ADMSC）神经发生过程中可抑制RhoA的表达。
测定ADMSC神经源性转分化过程中miRNA的表达动态。在神经源性转分化后，检测神经元特异性烯醇化酶（NSE）、Tuj-1（神经元特异性III类β-微管蛋白）和胶质纤维酸性蛋白（GFAP）的表达以及电生理特性。通过双荧光素酶测定、qRT-PCR和蛋白质免疫印迹法验证miR-124对RhoA的靶向作用。在体外利用功能获得和功能缺失实验研究miR-124和RhoA/ROCK信号通路的功能。
在ADMSC神经源性转分化过程中，miR-124显著上调。敲低内源性miR-124会阻碍神经源性转分化及所获得的电生理特性。miR-124可直接靶向RHOA mRNA并抑制其表达，由此增加NSE、Tuj-1和GFAP阳性的转分化细胞比例。在转分化过程中，RhoA/ROCK1而非ROCK2是miR-124的下游信号通路。
miR-124是一种重要的miRNA，至少部分通过miR-124/RhoA/ROCK1信号通路调控ADMSC的神经源性转分化。这些发现为未来将ADMSC用作再生医学和神经疾病细胞治疗的药物提供了一些基础信息。
