PrP106-126神经毒性多肽对原代神经元造成病理性损伤  

Neurotoxic PrP106-126 Fragmentinduces Neuropathological Changes in Primary Cortical Neurons

作　　者：宋志琦[1] 王运盛[2] 朱婷[3] 杨威[1] 崔永勇[1] 周向梅[1] 杨利峰[1] 赵德明[1] 

机构地区：[1]中国农业大学动物医学院,国家动物海绵状脑病实验室,北京100193 [2]北京动物园兽医院,北京100044 [3]福建农林大学动物科学学院,福州350002

出　　处：《实验动物科学》2017年第4期4-10,共7页Laboratory Animal Science

基　　金：国家自然科学基金(No.31472166)

摘　　要：目的系统地研究PrP106-126神经毒性多肽对原代神经元造成的损伤。方法使用出生24 h内的SD乳鼠分离培养原代神经元,经过PrP106-126处理后,利用免疫印迹技术检测凋亡相关蛋白的变化;利用透射电镜观察功毒后原代神经元亚细胞结构的变化情况;利用流式细胞仪检测细胞活性;利用免疫荧光技术观察细胞凋亡情况。结果随着PrP106-126刺激原代神经元的时间延长,抑制凋亡蛋白的表达量不断下降,促凋亡蛋白的表达量不断升高;透射电镜显示,经过多肽刺激的原代神经元的细胞器结构有不同程度的损伤,细胞胞浆内出现大小不等的单层或多层空泡结构;TUNEL细胞凋亡检测和Annexin V-FITC细胞活力检测试剂盒的检测结果都表明,经多肽刺激后,原代神经元的细胞活力显著下降,并有大量神经元发生凋亡。结论 PrP106-126神经毒性多肽激活促凋亡信号通路,造成细胞内的平稳紊乱和超微结构损伤,诱导原代神经元发生凋亡,为朊病毒疾病研究模型的建立提供全面可靠的证据。To investigate the neuropathological damage of prion diseases, we observed the apoptosis-associated protein changes by immunoblottingand the morphological changes of primary cortical neurons （PCCN） exposed to PrP106-126 by transmission electron microscopy （TEM）. Furthermore, PrP106-126-stimulated primary neuronal death was verified by TUNEL and Annexin V-FITC/PI apoptosis assay. After exposure to PrP106-126 for 24 h, the PCCN showed ultrastructural pathology similar to typical ＂ spongiform vacuoles＂ in TSEs. A large number of vacuoles of various sizes and swollen and disorganized smooth endoplasmie reticulum （SER） appeared in the cytoplasm. Nuclear fragments and condensed chromatin were released into the cytoplasm. Consistent with previous researches, we confirmed that PrP106-126 peptide induced mitochondrial cytochrome c release and caspase 3 activation, increased Bcl-2-associated X protein （BAX） and reduced B-cell lymphoma protein 2 （Bcl-2） in a time- dependent manner. Our observations proved that PrP106-126 is a valid model for researching PrPs~ neuropathological damage and neurotoxicity, which provide the theoretical basis for the researehin prion diseases and associated neurodegenerative diseases in vivo.

关 键 词：PrP106-126毒性多肽 原代神经元 病理性损伤 

分 类 号：R741.02[医药卫生—神经病学与精神病学]