Identification for serum biomarker for ALS associated with the function of TDP-43

肌萎缩侧索硬化症(ALS)是一种成人起病的运动神经元选择性丧失引起的神经退行性疾病。在ALS运动神经元中，43 kDa的TAR DNA结合蛋白(TDP-43)从胞核移位到胞浆并形成包涵体，提示TDP-43核功能的丧失可能是ALS发病的基础。TDP-43在RNA代谢中的功能包括转录调节、mRNA稳定性和Pre-mRNA的选择性剪接。然而，TDP-43在肌萎缩侧索硬化症患者组织中的功能尚未阐明。我们试图确定反映TDP-43功能的分子指示剂。利用外显子阵列分析，我们观察到在两种类型的培养细胞中，由于TDP-43表达缺失，几个基因的剪接发生了显著的变化。在TDP-43siRNA处理的细胞中，野生型剪接变异体减少，缺乏某些外显子的转录本增加。TDP-43的RNA结合能力是包含和排除这些外显子所必需的。此外，我们还发现，激光显微切割ALS后，在运动皮质、脊髓和脊髓运动神经元中拼接的内翻细胞数量增加。我们的结果提示在ALS影响的组织中TDP-43功能丧失，支持TDP-43功能丧失是ALS发病机制的假说。

Amyotrophic lateral sclerosis(ALS) is an adult-onset neurodegenerative disease caused by selective loss of motor neurons. In the ALS motor neurons, TAR DNA-binding protein of 43 kDa(TDP-43) is dislocated from the nucleus to cytoplasm and forms inclusions, suggesting that loss of a nuclear function of TDP-43 may underlie the pathogenesis of ALS. TDP-43 functions in RNA metabolism include regulation of transcription, mRNA stability, and alternative splicing of pre-mRNA. However, a function of TDP-43 in tissue affected with ALS has not been elucidated. We sought to identify the molecular indicators reflecting on a TDP-43 function. Using exon array analysis, we observed a remarkable alteration of splicing in several genes as a result of the depletion of TDP-43 expression in two types of cultured cells. In the cells treated with TDP-43 siRNA, wild-type splicing variants decreased and transcripts lacking some exons increased. The RNA binding ability of TDP-43 was necessary for inclusion and exclusion of these exons. Moreover, we found an increment of splicing varinats in motor cortex, spinal cord and spinal motor neurons collected by laser microdissection with ALS. Our results suggest a loss of TDP-43 function in tissues affected with ALS, supporting the hypothesis that a loss of function of TDP-43 underlies the pathogenesis of ALS.