【摘要】：背景:微小RNA參與調(diào)控干細(xì)胞增殖、分化、衰老等過程,通過了解Let-7家族成員Let-7c對骨髓間充質(zhì)干細(xì)胞向神經(jīng)細(xì)胞分化的影響可以為干細(xì)胞移植治療提供新思路。目的:探討Let-7c慢病毒載體在體外誘導(dǎo)大鼠骨髓間充質(zhì)干細(xì)胞向神經(jīng)細(xì)胞分化中的作用。方法:構(gòu)建Let-7c上調(diào)及下調(diào)慢病毒載體并體外轉(zhuǎn)染大鼠骨髓間充質(zhì)干細(xì)胞,篩選最適轉(zhuǎn)染復(fù)數(shù);實(shí)驗分為未轉(zhuǎn)染組、陰性轉(zhuǎn)染組(轉(zhuǎn)染空白慢病毒)、轉(zhuǎn)染上調(diào)組(轉(zhuǎn)染LV-rno-Let-7c-up)、轉(zhuǎn)染下調(diào)組(轉(zhuǎn)染LV-rno-Let-7c-5p-inhibition);采用法舒地爾誘導(dǎo)轉(zhuǎn)染后的大鼠骨髓間充質(zhì)干細(xì)胞分化為神經(jīng)細(xì)胞,倒置熒光顯微鏡下觀察骨髓間充質(zhì)干細(xì)胞轉(zhuǎn)染后熒光表達(dá),免疫細(xì)胞化學(xué)染色檢測神經(jīng)元標(biāo)記物NSE和MAP-2的表達(dá),RT-PCR檢測MAP-2 m RNA的表達(dá),MTT法檢測細(xì)胞存活率。結(jié)果與結(jié)論:倒置熒光顯微鏡下觀察大鼠LV-rno-Let-7c-up和LV-rno-Let-7c-5p-inhibition慢病毒載體轉(zhuǎn)染成功。法舒地爾可以誘導(dǎo)骨髓間充質(zhì)干細(xì)胞分化為神經(jīng)細(xì)胞,與陰性轉(zhuǎn)染組相比,轉(zhuǎn)染上調(diào)組的誘導(dǎo)效率升高(P0.05),NSE和MAP-2表達(dá)率上升(P0.05),轉(zhuǎn)染下調(diào)組的誘導(dǎo)效率下降,NSE和MAP-2表達(dá)率下降(P0.05)。轉(zhuǎn)染LV-rno-Let-7c-up后骨髓間充質(zhì)干細(xì)胞經(jīng)法舒地爾誘導(dǎo)向神經(jīng)細(xì)胞分化比率增加,Let-7c可能具有促進(jìn)骨髓間充質(zhì)干細(xì)胞向神經(jīng)細(xì)胞分化的作用。
[Abstract]:Background: micro RNA is involved in the regulation of stem cell proliferation, differentiation and senescence. Understanding the effect of Let-7c, a member of Let-7 family, on the differentiation of bone marrow mesenchymal stem cells into neural cells can provide new ideas for stem cell transplantation. Aim: to investigate the role of Let-7c lentivirus vector in inducing the differentiation of rat bone marrow mesenchymal stem cells into neural cells in vitro. Methods: Let-7c up-regulated and down-regulated lentivirus vectors were constructed and transfected into rat bone marrow mesenchymal stem cells in vitro. Negative transfection group (transfection of blank lentivirus), transfection upregulation group (LV-rno-Let-7c-up), transfection down-regulation group (LV-rno-Let-7c-5p-inhibition), induced differentiation of rat bone marrow mesenchymal stem cells into neural cells by fasudil. Fluorescence expression of bone marrow mesenchymal stem cells after transfection was observed under inverted fluorescence microscope. The expression of NSE and MAP-2 were detected by immunocytochemistry. The expression of MAP-2 m RNA was detected by RT-PCR. Results and conclusion: rat LV-rno-Let-7c-up and LV-rno-Let-7c-5p-inhibition lentivirus vectors were successfully transfected with inverted fluorescence microscope. Mesenchymal stem cells were induced to differentiate into neural cells. Compared with negative transfection group, the induction efficiency of transfection up-regulated group was higher (P0.05) and the expression rate of NSE and MAP-2 was higher (P0.05), but the induction efficiency of down-transfected group was lower than that of negative transfection group (P0.05). After transfection of LV-rno-Let-7c-up, the ratio of differentiation of bone marrow mesenchymal stem cells induced by fasudil was increased. Let-7c may promote the differentiation of bone marrow mesenchymal stem cells into neural cells.
【作者單位】： 鄭州大學(xué)第一附屬醫(yī)院神經(jīng)內(nèi)科;
【基金】：國家自然科學(xué)基金項目(骨髓間充質(zhì)干細(xì)胞神經(jīng)命運(yùn)決定中的自噬現(xiàn)象及其m TOR信號通路的作用/81071114;Lin28/let-7調(diào)控的骨髓間充質(zhì)干細(xì)胞移植治療阿爾茲海默病的實(shí)驗研究/81371385)~~
【分類號】：R457.7

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