BAG5/Hsp70复合体调控外泌体及隧道纳米管介导的TDP-43阳性包涵体在神经元与星形胶质细胞间转运

Amyotrophic lateral sclerosis(ALS) and frontotemporal dementia (FTD) attribute to the formation and transmission of TDP-43-positive inclusions(TDP-43 CIs) in the central nervous system. Current researches have demonstrated that misfolded protein aggregates transfered between cultured cells through exosomes and tunneling nanotubes(TNTs). BAG5/Hsp70 complex participates in protein folding, protein degradation and cytoskeleton rearrangement, and plays an important role in the neurodegenerative disease. Our previous studies have shown that TDP-43-positive exosomes were detected in the cerebrospinal fluid of ALS and ALS-FTD patients, with high-level expression of BAG5 and Hsp70 proteins in the exosomes.Furthermore, TNTs and TDP-43 CIs were detected in the SH-SY5Y and U251 following BAG5/Hsp70-positive exosomes ALS-FTD-CSF treatment, and BAG5/Hsp70 complex was co-localized with TNTs and TDP-43 CIs. Our findings suggests that BAG5/Hsp70 complex takes part in transfering of TDP-43 CIs through exosomes and tunneling nanotubes. This study will investigate the mechanism of BAG5/Hsp70 complex regulating exosomes and tunneling nanotubes-mediated transfer of TDP-43 CIs between neurons and astrocytes,try to find the new targets for preventing progression of ALS.

TDP-43阳性包涵体（TDP-43CIs）形成及在中枢神经系统中播散是肌萎缩侧索硬化症（ALS）和额颞叶痴呆（FTD）的病理机制。研究发现外泌体及隧道纳米管共同介导异常蛋白聚集体在细胞间转运。BAG5/Hsp70复合体参与蛋白折叠及细胞骨架重排，在神经退行性病中扮演重要角色。本项目前期研究发现：ALS及ALS-FTD脑脊液中存在TDP-43阳性外泌体，BAG5/Hsp70在外泌体表达上调；含BAG5/Hsp70阳性外泌体脑脊液孵育神经元及胶质细胞诱导了隧道纳米管及TDP-43CIs形成，二者与BAG5/Hsp70存在亚细胞共定位。此结果提示：BAG5/Hsp70参与了外泌体及隧道纳米管对TDP-43CIs的转运。本研究拟在细胞及TDP-43prp转基因鼠水平研究BAG5/Hsp70调控外泌体及隧道纳米管双渠道介导TDP-43CIs在神经元与胶质细胞间传递机制，寻找阻断ALS进展的新靶点。

TDP-43阳性包涵体（TDP-43 immunoreactive [ir] cytoplasmic inclusions [CIs], TDP-43-ir CIs）的形成及在中枢神经系统中播散是肌萎缩侧索硬化症（Amyotrophic Lateral Sclerosis, ALS）和额颞叶痴呆（frontotemporal dementia, FTD）的主要病理机制。研究发现TDP-43-ir CIs可能的播散方式有两种：其一，沿突触顺行及逆行方式在神经元及胶质细胞间近距离转运；其二，分泌至脑脊液中随脑脊液流动远距离转运。隧道纳米管 (tunneling nanotubes, TNTs)主要介导了TDP-43-ir CIs 在神经元及胶质细胞间的转运；外泌体 (exosomes)主要介导了TDP-43-ir CIs 通过脑脊液在中枢神经系统的播散。BAG5/Hsp70复合体参与蛋白折叠及细胞骨架重排，在神经退行性疾病中扮演重要角色。本研究运用皮层原代神经元和星形胶质细胞培养、质粒转染、细胞免疫荧光、免疫共沉淀、免疫印迹以及TDP-43prp转基因小鼠模型等研究方法发现：TDP-43阳性包涵体可通过exosome及TNTs双渠道在神经元及星型胶质细胞间转运；BAG5/Hsp70复合体促进了神经元和星型胶质细胞TDP-43阳性exosome的释放以及TNTs的形成；进一步使用TDP-43prp转基因小鼠模型证实BAG5/Hsp70复合体促进了TDP-43阳性包涵体在转基因小鼠脑内的播散。通过BAG5/Hsp70调控exosome及TNTs双渠道介导的TDP-43-ir Cis在神经元和胶质细胞之间的传递，可能成为阻断ALS进展的手段。