淀粉样前体蛋白C端片段对细胞骨架状态的影响

The Effects of C-Terminal Fragments of Amyloid Precursor Protein on Cytoskeletal State in Rat Primary Cortical Neurons

摘要: 【目的】探讨大鼠胚胎神经元内转染淀粉样前体蛋白C端片段对细胞骨架状态的影响。【方法】大鼠胚胎大脑皮层获得原代神经元进行培养，利用APP-C端片段、APLP2-C端片段基因重组质粒转染到神经元当中，用免疫染色的实验方法，观察肌动蛋白(actin)、微管蛋白(tubulin)、微管相关蛋白(MAP2)，磷酸化丝切蛋白(Ph-cofilin)的形态，并用Western Blotting方法测定了Ph-cofilin水平，每组实验重复测量十次。【结果】APP/APLP2-CTFs转染组与对照组相比有明显的向细胞外伸展的F-actin的重新聚合，细胞四周伸出的突触有所收缩；tubulin的结构发生紊乱；轴突当中的MAP2蛋白表达显著减弱，而且APP-C99组的MAP2蛋白的表达水平最低。蛋白印迹和免疫组化结果均显示APP/APLP2-CTFs转染组与对照组相比Ph-cofilin水平有所增高(P < 0.05)。【结论】转染淀粉样前体蛋白C端片段可以诱导细胞骨架蛋白和状态的改变。

Abstract: Objective: To study the effects of C-Terminal Fragments of amyloid precursor protein on cytoskeletal state in rat primary cortical neurons. Methods: Transfection of APP/APLP2-CTFs-pEGFP to rat primary cortical neurons investigates the morphology of actin, ph-cofilin, tubulin and MAP2 using immunecytochemistry test method in vitro. We take Western Blotting method for determining the Ph-cofilin transfected with APP-C/APLP2 C-Terminal fragment. Results: In the APP/APLP2- CTFs transfected group, the extracellular direction F-actin polymerization was increased; the extended synapses of Cells around were shrinked. And the structure of tubulin was disordered compared with control group. And the axons of the expression of MAP2 protein significantly reduced, and the APP-C99 group’s MAP2 protein expression level was lower. Immunocytochemistry and Western Blotting test showed that Ph-cofilin expression was increased in the APP/APLP2- CTFs transfection group compared with control group. Conclusion: Transfection of APP/LP-CTFs can induce the change of cytoskeletal protein and state.