IJPN  >> Vol. 5 No. 3 (August 2016)

    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

  • 全文下载: PDF(739KB) HTML   XML   PP.40-47   DOI: 10.12677/IJPN.2016.53007  
  • 下载量: 1,068  浏览量: 2,986   国家自然科学基金支持

作者:  

许敢峰:延边第二人民医院内分泌科,吉林 延吉;
刘 芳,陈 慧,许妍姬:延边大学医学院,预防医学教研室,吉林 延吉

关键词:
APP-C-Terminals Fragment Actin Tubulin MAP2APP-C端片段肌动蛋白微管蛋白微管相关蛋白

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。

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.

文章引用:
许敢峰, 刘芳, 陈慧, 许妍姬. 淀粉样前体蛋白C端片段对细胞骨架状态的影响[J]. 国际神经精神科学杂志, 2016, 5(3): 40-47. http://dx.doi.org/10.12677/IJPN.2016.53007

参考文献

[1] 贾艳滨, 何红波, 肖计划, 等. 阿尔茨海默病与血管性痴呆患者血浆微管蛋白和APP的变化[J]. 暨南大学学报(医学版), 2008, 29(2): 180-183.
[2] Kim, H.S., Kim, E.M., Lee, J.P., Park, C.H., Kim, S.H., Seo, J.H., Chang, K.A., Yu, E.A., Jeong, S.J., Chong, Y.H. and Suh, Y.H. (2003) C-Terminal Fragments of Amyloid Precursor Protein Exert Neurotoxicity by Inducing Glycogen Synthase Kinase-3Beta Expression. The FASEB Journal, 17, 1951-1953.
[3] Xu, Y., Kim, H.S., Joo, Y., Choi, Y., Chang, K.A., Park, C.H., Shin, K.Y., Kim, S., Cheon, Y.H., Baik, T.K., Kim, J.H. and Suh, Y.H. (2007) Intracellular Domains of Amyloid Precursor-Like Protein2 Interact with CP2 Transcription Factor in the Nucleus and Induce Glycogen Synthase Kinase-3β Expression. Cell Death and Differentiation, 14, 79-91.
http://dx.doi.org/10.1038/sj.cdd.4401928
[4] Munsie, L.N., Desmond, C.R. and Truant, R. (2012) Cofilin Nuc-lear-Cytoplasmic Shuttling Affects Cofilin-Actin Rod Formation During Stress. Journal of Cell Science, 125, 3977-3988.
http://dx.doi.org/10.1242/jcs.097667
[5] Heredia, L., Helguera, P., de Olmos, S., et al. (2006) Phosphorylation of Ac-tin-Depolymerizing Factor/Cofilin by LIM-Kinase Mediates Amyloid_β-Induced Degeneration: A Potential Mechanism of Neuronal Dystrophy in Alzheimer’s Disease. The Journal of Neuroscience, 26, 6533-6542.
http://dx.doi.org/10.1523/JNEUROSCI.5567-05.2006
[6] 田磊, 廖明芳, 李冀洲, 等. 细胞骨架蛋白中ADF/cofilin蛋白家系的作用: 如何与肌动蛋白结合及发挥解聚作用[J]. 中国组织工程研究与临床康复, 2009, 13(50): 9941-9945.
[7] 郭林林, 李学宝. 肌动蛋白解聚因子/丝切蛋白: 肌动蛋白重塑蛋白质家系[J]. 生命的化学, 2005, 25(3): 216-218.
[8] 陈显久, 胥显民, 等. 菌体中大量抽提质粒DNA方法的改进[J]. 中国药物与临床, 2003, 3(2): 103-105.
[9] Dopie, J., Skarp, K.P., Rajakyla, E.K., et al. (2012) Active Maintenance of Nuclear Actin by Importin 9 Supports Transcription. Proceedings of the National Academy of Sciences of the United States of America, 109, 544-552.
http://dx.doi.org/10.1073/pnas.1118880109
[10] Nogales, E. (2001) Structural Insights into Microtubule Function. Annual Review of Biophysics and Biomolecular Structure, 30, 397-420.
http://dx.doi.org/10.1146/annurev.biophys.30.1.397
[11] Ferrer, I., Blanco, R., Carmona, M., et al. (2002) Phosphorylated Mitogen-Activated Protein Kinase(MAPK/ERK) and Calcium/Calmodulin-Dependent Kinase II(CaM Kinase II) Are Differentially Expressed in Tau Deposits in Neurons and Galial in Tau Opathies. Journal of Neural Transmission, 108, 1397-1415.
http://dx.doi.org/10.1007/s007020100016
[12] 金善, 曹秉振, 张秀花, 等. 静脉注射同种异体骨髓间充质干细胞对血管性痴呆大鼠海马区脑组织形态及微管相关蛋白2表达的影响[J]. 中国组织工程研究与临床康复, 2007, 11(33): 6637-6640.
[13] Gong, C.-X., Wegiel, J., Lidsky, T., et al. (2000) Regulation of Phosphorylation of Neuronal Microtu-bule-Associated Proteins MAP1b and MAP2 by Protein Phosphatase-2A and -2B in Rat Brain. Brain Research, 853, 299-309.
http://dx.doi.org/10.1016/S0006-8993(99)02294-5
[14] 马志健, 牛海艳, 易西南. 血管性痴呆模型大鼠海马CA3区微管相关蛋白-2的表达及其意义[J]. 现代生物医学进展, 2009, 9(12): 2231-2233.
[15] 安改红, 陈学伟, 王静, 等. 慢性不可预知性心理应激对大鼠认知功能和海马微管相关蛋白-2表达的影响[J]. 解放军预防医学杂志, 2012, 30(4): 239-242.