HLA-G5介导的hAMSCs免疫炎症调节对ALS运动神经元的保护作用及机制研究

Immune inflammation and Regulatory T cells (Tregs) play important roles in the disease progression of Amyotrophic Lateral Sclerosis (ALS). Human leucocyte antigen-G5 (HLA-G5) is critical for human amniotic mesenchymal stem cells (hAMSCs) to modulate host immune-related inflammatory responses. Our previous study demonstrated that multiple hAMSC transplantations retarded disease progression and improved motor function in an ALS transgenic mouse model. However, the precise mechanisms remain unclear. Thus, we hypothesize that hAMSC transplantation increases HLA-G5 level in vivo, activates downstream signaling pathways, thus promoting the trend of T lymphocytes towards regulatory T lymphocytes, which could exert significant effects in mitigating neuroinflammation. To verify this hypothesis, we will apply lentiviral vector and RNA interference technology to establish the HLA-G over-expression and silence model of hAMSCs, and analyze the biological relationship between hAMSCs and T lymphocytes. Furthermore, we will also transplant the genetically modified hAMSCs by intravenous administration into hSOD1-G93A transgenic mouse and pig models, and analyze the trend of T-lymphocyte subsets, the expression of HLA-G receptors ILT-2/4, anti-inflammatory cytokines and pro-inflammatory cytokines. It is likely that the HLA-G over-expressing hAMSCs have an enhanced immunomodulatory ability that can slow down disease progression via a significant effect they exert on neuroinflammation. Thus, we plan to develop a new strategy for enhancing the therapeutic effects of hAMSCs transplantation therapy for ALS by enhancing their disease modulating properties.

免疫炎症反应和Tregs在肌萎缩侧索硬化症（ALS）疾病进程中起重要作用。HLA-G5是hAMSCs 发挥免疫炎症调节的关键分子。本课题组前期工作结果显示，经静脉移植hAMSCs能延缓转基因ALS模型小鼠的疾病进展，改善运动功能，但具体机制不清楚。我们提出科学假设：hAMSCs移植治疗ALS上调了体内HLA-G5的水平，激活其下游信号通路，诱导T 细胞分化为Tregs，减轻ALS免疫炎症反应，发挥神经保护作用。本项目拟以hAMSCs 表达HLA-G5为切入点，上下调hAMSCs 中HLA-G5 的表达，分别通过体外与外周血单核细胞共培养和体内细胞移植治疗实验，探讨上下调HLA-G5的hAMSCs对T细胞转化、增殖的影响，分析ILT-2/4、各种抗炎和促炎因子的表达水平，从分子、细胞和整体水平探讨HLA-G5在hAMSCs移植保护ALS运动神经元中的作用及机制，为临床治疗ALS提供理论依据。

肌萎缩侧索硬化症（ALS）是一种引起进行性瘫痪和肌肉萎缩的神经系统退行性疾病，目前尚缺乏有效预防和治疗措施。胶质细胞介导的神经炎症在ALS发生和发展过程中起重要作用。人羊膜间充质干细胞（hAMSCs）不但具有干细胞特性，还表现出低免疫源性和免疫抑制作用，可通过多种机制调控机体免疫炎症反应，为各种疾病的细胞治疗提供了新的选择。本项目前期研究已证实hAMSCs静脉移植治疗ALS转基因小鼠能够改善其运动功能，延缓疾病进展并延长生存期，但hAMSCs的治疗作用机制尚不清楚。本项目研究结果初步证实，hAMSCs的治疗机制可能在于通过抑制ALS转基因小鼠CNS病变组织神经炎症反应从而改善微环境起到保护运动神经元的从而发挥治疗作用。本研究结果初步阐明了hAMSCs 移植保护ALS 运动神经元的作用及机制，为临床治疗ALS提供理论依据。