Caveolin-1在大鼠骨髓间质干细胞分化为神经细胞中的作用

目的:探讨caveolin-1在骨髓间质干细胞(MSCs)分化为神经细胞中的作用。方法:实验分为未转染组、转染组(转染Rn-caveolin-1-siRNA)、阳性对照组(转染Rn-MAPK-1control siRNA)及阴性对照组(转染negative control siRNA)4组。采用β-巯基乙醇诱导大鼠MSCs分化为神经细胞。倒置荧光显微镜下观察MSCs转染后荧光表达情况;RT-PCR检测caveolin-1和促分裂原活化蛋白激酶1(MAPK1)mRNA的表达变化;免疫细胞化学法检测caveolin-1、神经元烯醇化酶(NSE)、神经微丝亚单位(NF-M)和胶质纤维酸性蛋白(GFAP)的表达变化;MTT方法检测细胞存活率。结果:(1)siRNA转染72h,MSCs荧光表达最强,转染率可达(81.5±2.8)%;而且转染组MSCs的caveolin-1mRNA转录下降(P0.05);MTT提示转染组细胞存活率无显著变化(P0.05)。(2)β-巯基乙醇可以诱导MSCs向神经细胞分化,其中以转染组诱导效果最佳,NSE、NF-M的表达率显著高于其它各组(P0.01)。(3)随诱导时间延长,各组caveolin-1表达持续增加,诱导6d达到峰值,与诱导前、诱导6h相比有显著差异(P0.05)。此外,转染组与其它各组同时点的caveolin-1表达均显著降低(P0.01)。结论:以caveolin-1为标记蛋白的脂筏在MSCs诱导分化为神经细胞中可能起到重要的调控作用。

Role of caveolin-1 in differentiation of rat bone marrow mesenchymal stem cells into neurons

AIM: To investigate the role of caveolin-1 in bone marrow mesenchymal stem cells (MSCs) differentiation into neurons. METHODS: The MSCs used in the experiment were divided into non-transfected group, transfected group (transfected with Rn-caveolin-1 siRNA), positive control group (transfected with Rn-MAPK-1 control siRNA) and negative control group (transfected with negative control siRNA). The MSCs were induced by β-ME to differentiate into neurons. The fluorescence expressed by transfected MSCs was observed under inverted fluorescence microscope. The mRNA expression of caveolin-1 and MAPK-1 was detected by RT-PCR. NSE, NF-M and GFAP, the neural cell specific markers, were detected by immunocytochemistry staining. The survival ratio of MSCs was detected by MTT method. RESULTS: The fluorescence of MSCs was mostly displayed at 72th h after transfection and the efficiency of transfection was up to 81.5%±2.8%. Meanwhile, the mRNA expression of caveolin-1 in transfected MSCs was decreased (P<0.05). No significant difference of the survival MSCs ratio between transfected group and other groups was observed by MTT method. β-ME induced MSCs into neural cells. The differentiation efficiency of MSCs transfected with Rn-caveolin-1 siRNA was the highest and the expression of NSE and NF-M was increased significantly compared to the other groups (P<0.01). The expression of caveolin-1 was increased persistently with time and the highest expression was observed 6 d after induction. Furthermore, there was significant difference between before induction and 6 d after induction. CONCLUSION: Lipid raft labeled with careolin as marker protein has important role in regulating MSCs differentiation into neurons.