Zfp521在大鼠骨髓间充质干细胞向神经元分化过程中的表达变化及意义

目的: 探讨锌指蛋白521(Zfp521)在大鼠骨髓间充质干细胞分化为神经元过程中的表达变化及意义。方法: 体外培养大鼠MSCs,实验分为未转染组、转染组(转染Rn-Zfp521-siRNA)和阴性对照组(转染negative control siRNA),采用β-巯基乙醇诱导骨髓间充质干细胞分化为神经元。倒置荧光显微镜下观察MSCs转染后荧光表达情况。采用免疫细胞化学法、RT-PCR法及Western blotting法检测诱导后神经元特异性烯醇化酶(NSE)和微管相关蛋白2(MAP-2)的表达情况及诱导前、后Zfp521的表达变化。结果: (1)siRNA转染72 h荧光表达最强,转染率可达84.1%±2.3%,转染组骨髓间充质干细胞的Zfp521 mRNA表达下降(P<0.05);(2)β-巯基乙醇可以诱导骨髓间充质干细胞分化为神经元,以转染组诱导效果最佳,NSE和MAP-2表达显著高于其它各组(P<0.05);(3)Zfp521在各组细胞中均有表达,诱导后Zfp521表达明显低于诱导前(P<0.01)。结论: Zfp521在大鼠骨髓间充质干细胞神经分化中表达下降,抑制Zfp521表达可促进神经元的分化,提示Zfp521在骨髓间充质干细胞的神经分化中可能发挥重要调控作用。

Effect of Zfp521 expression on differentiation of rat bone marrow mesenchymal stem cells into neurons

AIM: To investigate the effect of zinc finger protein 521(Zfp521) on the differentiation of rat bone marrow mesenchymal stem cells(MSCs) into neurons.METHODS: Rat MSCs were cultured by conventional method in vitro and divided into non-transfection group,transfection group(transfected with Rn-Zfp521-siRNA) and negative control group(transfected with negative control siRNA).MSCs were induced by β-mercaptoethanol(β-ME) to differentiate into neurons.The fluorescence expressed by transfected MSCs was observed under inverted fluorescence microscope.The expression of Zfp521 was detected after transfection by RT-PCR.Immunohistochemistry,RT-PCR and Western blotting were used to detect the expression levels of neuron-specific enolase(NSE),microtubule-associated protein 2(MAP-2) and Zfp521 after induction.RESULTS: The fluorescence of MSCs was mostly displayed 72 h after transfection and the efficiency of transfection was up to 84.1%±2.3%.Meanwhile,the mRNA expression of Zfp521 was decreased(P<0.05).MSCs were induced by β-ME to differentiate into neurons.The differentiation efficiency of MSCs transfected with Rn-Zfp521-siRNA was the highest and the expression of NSE and MAP-2 was significantly increased compared with other groups(P<0.05).Zfp521 was detected in all groups,and the expression level of Zfp521 was significantly decreased after induction(P<0.01).CONCLUSION: Zfp521 may be down-regulated during the differentiation.The inhibition of Zfp521 promotes the neural differentiation of MSCs.Zfp521 may play an important role in regulating MSCs differentiation into neurons.