MSCs基于M1/M2-Th17/Treg免疫稳态调控TBI炎症反应的机制研究

The inflammatory response is regarded as a key factor in the secondary injury cascade following traumatic brain injury (TBI), which suggests that anti-inflammatory or immunoregulatory strategies could provide effective treatments for the management of TBI-induced pathology. Although many studies have demonstrated that mesenchymal stem cell (MSCs) possess immunomodulatory properties, the mechanism on immune cell homeostasis remains largely unknown. In this study, we therefore investigated immunomodulatory effects and mechanisms of MSCs from macrophage M1/M2 polarization and Th17/Treg immune balance perspective in TBI-induced neuroinflammation. The study mainly includes three aspects. Firstly, it is important to analyze the correlation between M1/M2 polarization and Th17/Treg immune balance and clinical significance from TBI clinical cases. On this basis, we investigate the effect of umbilical cord mesenchymal stem cell transplantation on M1/M2 polarization and Th17/Treg immune balance in TBI animal model. Finally, we analyze the functions and mechanisms of MSCs on M1/M2 polarization and whether MSC affect the Th17/Treg differentiation by regulating M1/M2 polarized and antigen-presenting function. Overall, this study will clarify the immunomodulatory effects of MSCs-M1/M2 polarization-Th17/Treg homeostasis axis on TBI inflammatory response and may provide a theoretical basis and new therapeutic targets.

炎症反应是颅脑创伤（TBI）后引起继发性损伤的关键因素, 因此抗炎和免疫调节是治疗TBI的有效途径。研究发现间充质干细胞（MSCs）具有抑制TBI后炎症反应的免疫调节作用，但其调控免疫细胞稳态的具体机制尚不清楚。本项目拟从巨噬细胞M1/M2极化以及后续Th17/Treg分化的免疫稳态角度探讨干细胞对TBI后炎症反应的免疫调节作用及其机制。在临床病例分析的基础上，利用TBI动物模型研究MSCs对小胶质细胞/巨噬细胞M1/M2极化和Th17/Treg免疫平衡的影响。通过体外实验进一步阐明MSCs对小胶质细胞M1/M2极化以及后续Th17/Treg分化的影响及可能的分子机制。以上研究将深入探讨MSCs-M1/M2极化-Th17/Treg稳态这一免疫调节轴对TBI后炎症反应的调节作用，从而为更有效地控制TBI后炎症反应提供新的思路及治疗的靶点。

TBI现已成为世界范围内亟待解决的公共卫生问题。有研究显示，炎症反应是TBI后引起继发性脑损害的关键因素。因此，抗炎和免疫调节是治疗TBI诱导损伤的有效治疗措施。研究发现间充质干细胞（MSCs）具有重要的免疫调节功能，然而其调控免疫细胞稳态的具体机制尚不清楚。本课题深入探讨MSCs-M1/M2极化-Th17/Treg稳态这一免疫调节轴对TBI后炎症反应的调节作用。临床实验结果显示，TBI后1周内患者外周血T淋巴细胞亚群向促炎型Th17细胞偏移，Th17/Treg比值明显升高。体外实验结果显示，MSCs处理可导致Th17细胞明显减少，Treg细胞明显增多，Smad3蛋白表达增加，NF-κB表达减少。而Smad3抑制剂SIS3可逆转这种变化。MSCs处理可导致BV2细胞Iba-1和Arg-1表达显著增高，NF-κB p-p65/p65的比值降低。在体实验结果显示，MSCs可减轻TBI大鼠脑组织损伤局部细胞及间质水肿，减少炎性细胞数量。MSCs下调促炎细胞因子（IL-1β、IL-6和TNF-α）而上调抗炎因子（TGF-β、TSG-6和Cox-2）的表达。TBI后，MSCs可使损伤侧海马DG区BrdU/DCX以及BrdU/NeuN双标记阳性细胞均增多。MSCs处理使TBI大鼠外周血或脑组织Th17/Treg比值、p-p65/p65的比值和NF-κB表达有所下降，NF-κB和Smad3表达显著增加。行为学结果显示，MSCs处理后大鼠的逃避潜伏期缩短，平台穿越次数及目标象限停留时间均增加，mNSS评分下降。本研究提示MSCs可通过Smad3信号通路诱导促炎型Th17细胞向抗炎型Treg细胞转分化，调节TBI后炎症反应及中枢及外周Th17/Treg免疫平衡，MSCs还能够调节TBI后大鼠脑组织中小胶质细胞M1/M2极化方向，改善损伤周围免疫微环境，减少促炎因子（IL-1β、IL-6和TNF-α等）的表达，增加抗炎因子（PGE2，TSG-6和TGF-β等）表达，改善TBI后海马神经发生，从而促进TBI大鼠神经功能缺损修复，改善TBI后远期神经功能预后。本课题取得成果如下：发表SCI期刊论文4篇、国内期刊论文4篇；主审学术专著2部；主持召开全国学术会议5次；培养博士1名，硕士3名。