α-突触核蛋白磷酸化参与小鼠多巴胺能神经元的保护作用  

α-Synuclein phosphorylation takes part in mouse dopaminergic neuron protection

作　　者：刘琦[1] 段春礼[1] 吴波[1] 范春香[1] 张韬[1] 赵焕英[1] 赵春礼[1] 杨慧[1] 

机构地区：[1]首都医科大学北京神经科学研究所北京市神经再生修复研究重点实验室神经变性病教育部重点实验室,北京100069

出　　处：《基础医学与临床》2009年第6期561-565,共5页Basic and Clinical Medicine

基　　金：国家重点基础研究发展计划(2006CB500706);中国高技术研究发展计划(2006AA02A408);国家自然科学基金(30670655;30700199);北京市教育委员会科技发展计划(KM200610025002);教育部高等学校博士学科点专项科研基金(20060025004);北京市自然科学基金(7082011);北京市属高等学校人才强教计划资助项目PHR(IHLB)

摘　　要：目的研究α-突触核蛋白(α-SYN)129位点磷酸化修饰是否具有神经元保护作用。方法应用反转录病毒感染小鼠多巴胺能神经元细胞MN9D,通过实时定量PCR检测各组细胞内α-SYN过表达情况,并通过免疫细胞化学方法检测野生型α-SYN(WT/α-SYN)及129位点磷酸化α-SYN(S129D/α-SYN)过表达后α-SYN在细胞内聚集情况,应用CCK-8检测细胞活力,观察WT/α-SYN及S129D/α-SYN对MN9D的细胞毒性,从而明确S129D/α-SYN对多巴胺能神经元的影响。结果实时定量PCR结果显示,WT/α-SYN及S129D/α-SYN在MN9D细胞内均过表达(P<0.01)。WT/α-SYN过表达组细胞活力明显下降(P<0.01),共聚焦显微镜观察未见明显的α-SYN聚集;S129D/α-SYN过表达组细胞内可观察到明显的α-SYN聚集体,同WT/α-SYN组相比细胞活力有明显提高(P<0.01)。结论129位点丝氨酸的磷酸化修饰在一定程度上减轻了WT/α-SYN过表达所引起的细胞毒性。Objective To study the role of phosphorylation at Serine 129 in regulating the neurotoxicity of α-synuclein. Methods Wild type and phosphomimic mutant α-synuclein genes were over-expressed in mouse dopaminergic cells MN9D using retrovirus. The cell viability was examined using CCK-8 assay and cell morphology was observed by immunofluorescenee microscopy. Results The result of real time PCR showed that WT/α-SYN and S129D/α-SYN genes were overexpressed in MN9D as compared to uninfected MN9D and vector control group （P 〈 0. 01 ）. The cell viability was obviously reduced as a result of over-expressing both WT and phosphorylated α-synuclein genes. The viability of cells over-expressing wild type α-synuelein was much lower than that of the cells over-expressing phosphorylated α-synuclein（ P 〈 0. 05 ,P 〈 0. 01 ,P 〈 0. 01 ）. Insoluble α-synuclein aggregates were observed with confocal microscope in the MN 9 D cells over - expressing phosphorylated α - synuclein . Conclusion Over-expressing phosphorylated α-synuclein may lead to α-synuclein aggregated in MN9D cells. Phosphorylation of 129 Ser may protect the cells from toxicity induced by over expressing α-synuclein.

关 键 词：Α-突触核蛋白 磷酸化α-突触核蛋白 神经元保护 多巴胺神经元 

分 类 号：R742.5[医药卫生—神经病学与精神病学]