转录因子Eomesodermin表达上调致ALS患者外周免疫失调的机制研究

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterized by selective death of upper and lower motor neurons. Patients usually develop rapid progression and fatal outcome without effective therapies. It’s of great importance to discover potential therapeutic targets. Multiple evidences indicated that T lymphocytes involved deeply in the disease progression and neuron degeneration of ALS. We previously discovered that the Eomes expression in CD4+T lymphocytes of ALS PBMC was obviously elevated, which was negatively correlated with the portion of Treg. One possible explanation is that the cellular distribution of disease-causing protein, also acting as RNA binding protein, was changed. The post-transcriptional regulation was interrupted resulting in an elevation of Eomes expression. In this project, we will further complete the peripheral T cell lineage analysis of ALS patients, as well as the phenotype identification and functional study of Eomes+CD4+T cells. Moreover, we will elucidate the function of Eomes+CD4+T cells in Tardbp mutant mice and clarify the mechanism of Eomes+ CD4+T upregulation and CD4+T lymphocytes abnormality using single-cell sequencing in human samples. Additionally，we will perform functional study in cell line model and conditional deficiency mice, which could provide theoretical basis of immune therapy in ALS patients.

ALS是一种侵犯脊髓前角、脑干运动核团和锥体束的神经系统变性病，进展迅速，现有药物疗效有限，因此寻找潜在治疗靶点至关重要。由于T细胞明确参与ALS的疾病进展及运动神经元变性，申请人前期发现ALS患者外周CD4细胞Eomes表达明显上调，与Treg细胞呈负相关，推测其机制可能是ALS关键毒性蛋白作为RNA结合蛋白在患者CD4细胞中发生定位变化，影响了EOMES基因的转录后调控。本项目将完善ALS患者外周免疫细胞谱系分析及Eomes+CD4+T细胞表型和功能探索，进行Tardbp突变小鼠的表型和Eomes+CD4+T细胞功能探索，并通过分选高、低表达Eomes的人CD4+T细胞进行单细胞测序、细胞模型及Tardbp突变小鼠条件敲除CD4细胞Eomes基因等多个层次阐明ALS患者外周Eomes+CD4+T细胞比例增高及引起外周免疫失调的机制，为ALS的免疫治疗提供理论基础。

ALS是一种侵犯脊髓前角、脑干运动核团和锥体束的神经系统变性病，进展迅速，现有药物疗效有限，因此寻找潜在治疗靶点至关重要。本项目申请人前期发现ALS患者外周CD4细胞Eomes表达明显上调，推测其机制可能是ALS关键毒性蛋白作为RNA结合蛋白在患者CD4细胞中发生定位变化，影响了EOMES基因的转录后调控。本项目通过收集临床样本，对2个队列共148名ALS患者与101名健康正常人的Eomes+CD4+T细胞表达分布和细胞亚群特征进行了鉴定，明确了ALS患者Eomes+CD4+T细胞确实发生了变化，并且这种变化可能影响到了CD4+T细胞谱系，进一步通过RNA免疫共沉淀等功能研究，发现ALS关键致病蛋白TDP-43可与EOMES mRNA结合，且突变型TDP-43蛋白结合的EOMES mRNA水平更高。本研究初步揭示了ALS关键致病蛋白的异常毒性聚集影响了患者外周免疫细胞的调控机制，进一步完善了“ALS的异常蛋白聚集学说”，并提供了一个新的ALS外周免疫学标志物。

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