星形胶质细胞影响神经干细胞突触表达的神经营养家族基因表达机制探讨  

Regulation of Synaptophysin and GAP-43 Expression in Neural Stem Cells Affected by 1-methyl-4 phenyl Pyridinium and Astrocyte

作　　者：闫荣[1] 罗晓光[2] 张尧[1] 李宇凤[1] 冯娟[1] 

机构地区：[1]中国医科大学附属盛京医院神经内科,沈阳110004 [2]中国医科大学附属第一医院神经内科,沈阳110001

出　　处：《中国医科大学学报》2013年第11期989-995,共7页Journal of China Medical University

基　　金：辽宁省教育厅高校科研计划(2009A780);辽宁省博士科研启动基金(20091106)

摘　　要：目的探讨1甲基4苯基-吡啶离子(MPP+)和星形胶质细胞对神经干细胞突触素和生长相关蛋白43(GAP 43)表达的影响,以及星形胶质细胞神经营养素家族基因表达变化,包括脑源性神经营养因子(BDNF),神经生长因子(NGF),神经营养素3(NT 3)。方法实验分为两步骤,第一步骤:PC12细胞分别经MPP+诱导不同时间点(0 h、24 h、48 h、72 h、96 h)后分为两部分,一部分应用流式细胞技术检测不同时间点PC12细胞凋亡率;另一部分与星形胶质细胞共育2 d,然后收集各组细胞和各组细胞条件培养液,应用RT PCR检测各组细胞BDNF mRNA、NGF mRNA和NT 3 mRNA表达变化;将各组细胞条件培养液对神经干细胞(NSC)进行诱导分化,免疫荧光检测神经干细胞(NSC)突触素(SYN)和GAP 43表达。结果在MPP+作用48 h时间点,PC12细胞凋亡率达高峰(P<0.05);与各组比较,与MPP+诱导48 h后的PC12细胞共育48 h后的星形胶质细胞(C3组)BDNF mRNA表达(A=93.96±4.89)明显增高(P<0.05),差异有统计学意义(P<0.05),并且C3组星形胶质细胞条件培养液诱导的神经干细胞(NSC)突触素(SYN)(A=34.09±2.69)、GAP 43(A=49.36±5.98)表达明显升高,与其他组比较,差异有统计学意义(P<0.05);NGF mRNA与各组比较差异无统计学意义(P>0.05);NT 3 mRNA在各组中均未见到表达。结论星形胶质细胞与MPP+诱导凋亡的PC12细胞共育后,星形胶质细胞BDNF基因表达明显增高,星形胶质细胞促进神经干细胞(NSC)突触素(SYN)和GAP 43表达,BDNF可能参与了这一过程。Objetedve To observe whether 1-methyl-4 phenyl pyridinium （MPP＋） and Astrocyte （AS） affected the expression of synaptophysin and GAP-43 in neural stem cells （ NSC ）, and discuss whether neuretrophins （ BDNF, NGF, NT-3 ） were involved in the process. Methods Experi- ments were designed with two steps. First, PC12 cells were induced by MPP＋ for different time periods （0 h, 24 h, 48 h, 72 h, 96 h）. Then the PC 12 cells were divided into two pans. The apoptosis rates ofMPP＋ -induced PCI2 cells （13 h,24 h, 48 h, 72 h,96 h） were measured at different time points by Flow cytometry. The rest of the MPP＋ -induced PC12 cells were incubated with astrocytes for 48 hours. Finally ,cells of all groups were col- lected, and the expression of BDNF mRNA, NGF mRNA and NT-3 mRNA were detecled by RT-PCR. The conditioned medium collected from each group was used to induce cell differentiation of NSC. The expression of synaptophysin （ SYN ） and growth-associated protein -43 （ GAP-43 protein） of NSC were analyzed by immunofluorescen assay. Results The Flow cytometry results showed that the MPP induced PC12 cell apoptesis rates reached the peak at 48h （ P 〈 0.05 ）. The expression of BDNF mRNA in the astrocytes （A = 93.96±4.89 ） were significantly up-regulated after 48 h induction with MPP＋-treated PC 12 （ P 〈 0.05 ）. The expression of synaptophysin （ SYN ） （ A = 34.09±2.69 ） and GAP-43 （ A = 49.36±5.98 ） of NSC induced by astrocytes -conditioned medium from 48 h treatment group were significantly higher than other groups （ P 〈 0.05 ）. There was no statisti- cal difference of NGF mRNA among different groups （P 〉 0.05 ）. NT-3 mRNA was not detected in all groups. Conclusion After incubating with MPP-induced PC12 cells（MPP-PC12）, the expression of BDNF mRNA of astrocytes were remarkably increased. Astrocytes promoted the expression of synaptophysin （ SYN ）and GAP-43 in NSC, and BDNF probably war involved in this process.

关 键 词：星形胶质细胞 神经干细胞 突触素 生长相关蛋白-43 脑源性神经营养因子 神经生长因子 神经营养素-3 

分 类 号：R741.02[医药卫生—神经病学与精神病学]