發(fā)布時間：2019-01-15 21:24

【摘要】：目的觀察miR-363、脫中胚蛋白(Eomes)在人臍帶間充質(zhì)干細(xì)胞(HUMSCs)誘導(dǎo)的神經(jīng)樣細(xì)胞中的表達(dá)變化,并探討其意義。方法培養(yǎng)HUMSCs并應(yīng)用3-異丁基-1-甲基黃嘌呤、視黃酸、丁基羥基茴香醚聯(lián)合誘導(dǎo)分化為神經(jīng)樣細(xì)胞,采用免疫熒光化學(xué)法檢測誘導(dǎo)前后細(xì)胞β-Ⅲ-tubulin和NSE的表達(dá);real-time PCR檢測誘導(dǎo)前后細(xì)胞中miR-363和Eomes mRNA的表達(dá),Western blot檢測誘導(dǎo)前后細(xì)胞中Eomes蛋白的表達(dá);運(yùn)用生物信息學(xué)方法(Target Scan和miRanda)對miR-363和Eomes基因的靶向配對關(guān)系進(jìn)行預(yù)測,采用熒光素酶報告系統(tǒng)進(jìn)行鑒定。結(jié)果光鏡下可見HUMSCs呈梭形均勻分布,誘導(dǎo)分化后細(xì)胞呈圓形、有長的突起;免疫熒光檢測發(fā)現(xiàn)誘導(dǎo)14 d的細(xì)胞β-Ⅲ-tubulin和NSE表達(dá)呈陽性,神經(jīng)樣細(xì)胞誘導(dǎo)成功。HUMSCs誘導(dǎo)分化前后miR-363相對表達(dá)量分別為1.00±0.31、0.32±0.02,Eomes mRNA相對表達(dá)量分別為1.00±0.22、15.73±0.85,Eomes蛋白相對表達(dá)量分別為0.21±0.02、0.78±0.06,兩者比較,P均0.05。生物信息學(xué)軟件結(jié)果顯示miR-363和Eomes基因靶向配對良好,熒光素酶報告系統(tǒng)鑒定發(fā)現(xiàn)miR-363能夠抑制Eomes mRNA表達(dá)。結(jié)論 miR-363能夠靶向作用Eomes 3'UTR負(fù)性調(diào)控HUMSCs神經(jīng)樣細(xì)胞分化后Eomes的表達(dá)。
[Abstract]:Objective to observe the expression of miR-363, demineralized (Eomes) in neural like cells induced by human umbilical cord mesenchymal stem cells (HUMSCs) and its significance. Methods HUMSCs was cultured and differentiated into neuron-like cells with 3-isobutyl 1-methylxanthine, retinoic acid and Ding Ji hydroxyl anisole. The expression of 尾-鈪，

本文編號：2409135