Nurr-1基因转染促进脂肪干细胞的神经分化

Enhanced Neural Differentiation of Adipose Tissue-Derived Stem Cells by Nurr-1 Gene Transfection

目的:研究孤儿核受体相关基因1(Nurr-1)对脂肪干细胞(adipose tissue-derived stem cells,ADSC)向神经元方向分化的潜在作用。方法:流式细胞术与成骨、成脂诱导技术鉴定脂肪干细胞;Nurrr-1基因转染脂肪干细胞后,应用神经特异性标志物MAP-2,β-tubulin的免疫荧光染色评估其向神经方向分化的能力。结果:流式细胞术结果表明培养的细胞CD29,CD44表达90%以上,CD45,CD90表达均低于1.5%,经过诱导后,油红O、茜素红S染色均呈阳性,表明所培养的细胞为脂肪干细胞;慢病毒转染Nurr-1基因后,免疫荧光染色检测MAP-2,β-tubulin的免疫荧光强度显著增加;RT-PCR结果显示Nurr-1转染的脂肪干细胞的MAP-2、β-tubulin、NF200的表达量显著提高。结论:Nurr-1基因转染能促进脂肪干细胞向神经方向分化,为神经损伤和神经退行性病变的治疗提供了新途径。

Objective： To explore the role of Nurr-1 gene over-expression in neuronal differentiation of adipose tissue-derived stem cells （ADSCs）. Methods： The characterization of ADSCs was performed with flow cytometric technology, adipogenic induction and osteogenic induction. The differentiation of ADSCs into neuron-like cells was examined by immunofluorescence staining and RT-PCR. The gene expression of microtubule-associated protein 2 （MAP-2）, β-tubulin and neurofilaments 200 （NF-200）, was examined by RT-PCR. The proliferation of cells after Nurr-1 gene transfection was characterized using a CCK-8 assay. Results： Flow cytometric anal- ysis demonstrated that cells were positive for CD90 and CD29, but only 0.3% of the cells were positive for CD31 and 1.2% of the cells were positive for CD45. After differentiation into adipocytes, the cells were stained with Oil Red O, and red colored lipid vacuoles were visualized. After osteogenic differentiation, the cells were stained with Alizarin-Red-S, and an immense production of mineral deposits was displayed. The MAP-2 and β-tubulin were positive in Nurr-1 group after immunofluorescence staining. The gene and protein ex- pression of MAP-2 and β-tubulin was significantly enhanced compared with control group. Conclusion： Nurr-1 gene transfection is able to enhance the neuronal differentiation of ADSCs, which highlights the potential application in the treatment of neurodegenerative dis- eases and neurotrauma in nervous system.