Mehri-Ghahfarrokhi Ameneh, Pourteymourfard-Tabrizi Zahra, Farrokhi Effat, Chaleshtori Morteza Hashemzadeh, Jami Mohammad-Saeid
Department of Molecular Medicine, School of Advanced Technologies, Shahrekord University of Medical Sciences, Shahrekord, Iran.
Cellular and Molecular Research Center, Basic Health Sciences Institute, Shahrekord University of Medical Sciences, Shahrekord, Iran.
J Otol. 2019 Dec;14(4):121-127. doi: 10.1016/j.joto.2019.04.001. Epub 2019 Apr 16.
Auditory neuropathy is the particular form of deafness in humans which cannot be treated by replacement therapy. Human dental pulp stem cells (hDPSCs) are derived from an ectomesenchymal neural crest cell population. Therefore, they possess a promising capacity for neuronal differentiation and repair. miR-124, a key regulator of neuronal development in the inner ear, is expressed at high levels in auditory and vestibular neurons. Here, we evaluated the possible effect of miR-124 in alteration of neural protein markers expression. Using quantitative reverse transcription-PCR (qRT-PCR) analyses and immunofluorescence staining, we studied the expression patterns of neural progenitor markers (Nestin, NOTCH1, and SOX2) and neural markers (β-tubulin III, GATA-3, and peripherin) upon transfection of hDPSCs with miR-124. The qRT-PCR results showed that Nestin was upregulated 6 h post-transfection. In contrast, Nestin expression exhibited a decreasing trend 24 h and 48 h post-transfection. Higher levels of β-tubulin III, 6 h and 16 h post transfection in RNA level as compared with control cells, were determined in transfected DPSCs. However, β-tubulin-III expression decreased 48 h post-transfection. The immunoflourescence results indicated that transfection of hDPSCs with miR-124, only affected Nestin among the studied neural progenitor and neural marker expression in protein level.
听觉神经病是人类耳聋的一种特殊形式,无法通过替代疗法进行治疗。人牙髓干细胞(hDPSCs)源自外胚间充质神经嵴细胞群体。因此,它们具有神经元分化和修复的巨大潜力。miR-124是内耳神经元发育的关键调节因子,在听觉和前庭神经元中高水平表达。在此,我们评估了miR-124对神经蛋白标志物表达改变的可能影响。通过定量逆转录PCR(qRT-PCR)分析和免疫荧光染色,我们研究了用miR-124转染hDPSCs后神经祖细胞标志物(巢蛋白、NOTCH1和SOX2)和神经标志物(β-微管蛋白III、GATA-3和外周蛋白)的表达模式。qRT-PCR结果显示,转染后6小时巢蛋白上调。相反,转染后24小时和48小时巢蛋白表达呈下降趋势。与对照细胞相比,转染的DPSCs在RNA水平上转染后6小时和16小时β-微管蛋白III水平更高。然而,转染后48小时β-微管蛋白III表达下降。免疫荧光结果表明,用miR-124转染hDPSCs,在蛋白质水平上,在所研究的神经祖细胞和神经标志物表达中仅影响巢蛋白。