發(fā)布時間：2018-06-23 08:33

  本文選題：Nurr + 基因修飾　； 參考：《中風(fēng)與神經(jīng)疾病雜志》2017年05期

【摘要】：目的利用慢病毒載體建立GFP-Nurr1基因修飾的原代神經(jīng)干細(xì)胞(NSCs)模型并觀察Nurr1過表達(dá)后NSCs向多巴胺神經(jīng)元的分化影響。方法利用基因重組構(gòu)建pLenO-DCE-Nurr1慢病毒載體,用慢病毒轉(zhuǎn)染第三代NSCs,轉(zhuǎn)染72 h后熒光檢測轉(zhuǎn)染效果;設(shè)置空白對照組、空載體組及DCE-Nurr1組,分別用Western blot及PCR檢測Nurr1的表達(dá)差異;并將轉(zhuǎn)染后的NSCs分化培養(yǎng)7 d后分別用免疫細(xì)胞化學(xué)檢測、Western blot及PCR檢測酪氨酸羥化酶(TH)的表達(dá)。結(jié)果慢病毒轉(zhuǎn)染NSCs 72 h后,轉(zhuǎn)染率可達(dá)90%,與對照組相比DCE-Nurr1組高表達(dá)Nurr1。經(jīng)慢病毒載體感染后的NSCs仍具備分化潛能,分化培養(yǎng)后發(fā)現(xiàn)DCE-Nurr1組分化的神經(jīng)細(xì)胞中TH陽性細(xì)胞分化率90.60%,對照組為21.2%。結(jié)論慢病毒載體可高效轉(zhuǎn)染NSCs過表達(dá)Nurr1;Nurr1基因過表達(dá)可以促進(jìn)中腦腹側(cè)來源NSCs向TH陽性多巴胺能神經(jīng)元方向分化。
[Abstract]:Objective to establish the primary neural stem cell (NSCs) model modified by GFP-Nurr1 gene using lentivirus vector and to observe the effect of Nurr1 overexpression on the differentiation of NSCs into dopamine neurons. Methods the lentivirus vector pLenO-DCE-Nurr1 was constructed by gene recombination, the third generation of NSCs was transfected with lentivirus, the transfection effect was detected by fluorescence after 72 h, the difference of Nurr1 expression in blank control group, empty vector group and DCE-Nurr1 group was detected by Western blot and PCR, respectively. The expression of tyrosine hydroxylase (th) was detected by immunocytochemistry and Western blot. Results after 72 hours of lentivirus transfection, the transfection rate of NSCs reached 90%, and the expression of Nurr1 in DCE-Nurr1 group was higher than that in control group. NSCs infected by lentivirus vector still had differentiation potential. After differentiation and culture, it was found that the differentiation rate of th positive cells in DCE-Nurr1 group was 90.60%, and that in control group was 21.2%. Conclusion the overexpression of Nurr1N Nurr1 gene in NSCs transfected with lentivirus vector can promote the differentiation of NSCs from ventral mesencephalon into th positive dopaminergic neurons.
【作者單位】： 昆明醫(yī)科大學(xué)第一附屬醫(yī)院神經(jīng)外科;
【基金】：國家自然科學(xué)基金地區(qū)科學(xué)基金項(xiàng)目(No.81360197);國家自然科學(xué)基金項(xiàng)目(No.81241126) 云南省教育廳科學(xué)研究基金項(xiàng)目(No.2013C227) 云南省干細(xì)胞與再生醫(yī)學(xué)重點(diǎn)實(shí)驗(yàn)室開放課題(No.201401UH00160)
【分類號】：R742.5
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本文編號：2056517