免疫相关GTP酶（IGRM/Irgm1）在多发性硬化症发病过程中对巨噬细胞型别转换的影响及相关机制研究

Multiple sclerosis (MS) is a chronic inflammatory disease of central nerve system. Studies mainly focus on the effect of T cells on the occurrence and result of MS, however macrophage (M?), as the necessary condition of the activation and playing effect of T cells, has not been emphasized enoughly. Latest research and our previous work showed that M? could differentiate into different subtypes, and different subtypes could play different role during the pathology of MS and its animal model-EAE. At the same time we found that IRGM/Irgm1 is the important factor for the regulation of M?. The severity of EAE based on Irgm1 knock out mouse is obviously lower than WT EAE mice. Combining our previous work and nowadays knowledge about the switch of M?, our project employ mice EAE model and blood sample of MS patients as study objects, explore from a completely new view the effect of different subtype of M? on the different stage of disease and the relative mechanism. We will resolve through the project: 1, What character does different subtype of M? play in the pathology of MS/EAE? 2, How does the subtype of M? feedback on the effective T cells? 3, What kind of mechanism does IRGM/Irgm1 play in the regulation of the switch of M?? The resolution of the above questions would provide important theoretical basis and entirely new idea for the therapy of MS.

多发性硬化(MS)是中枢神经系统慢性炎性疾病。大量研究集中在T细胞对MS发生和转归的影响，而作为T细胞活化和发挥效应的必要条件-单核/巨噬细胞（M?）在MS中的作用并没给予足够重视。最新的研究及我们的前期工作表明：M?可分为不同型别，不同M?亚型在MS及其动物模型EAE的发病机制中扮演着不同的角色；同时发现IRGM/Irgm1是调节M?功能的重要因子；Irgm1基因敲除小鼠EAE发病明显低于野生型小鼠。结合前期工作和现有M?型别转换的知识，本项目以小鼠EAE模型和MS病人外周血样本为研究对象，从全新的角度探讨不同类型M?对疾病不同阶段的影响及相关机制。通过本项目我们将解决：1、不同型别M?在MS/EAE病程中扮演什么角色？2、不同型别M?如何反馈性调控效应性T细胞？3、IRGM/Irgm1通过何种机制调控M?型别转化？这些问题的解决将为MS临床治疗提供重要的理论依据和全新的思路。

经过四年的科研工作，我们如期完成了课题内容。首先，我们建立了稳定的Irgm1+/-和Irgm1-/-的转基因小鼠模型，在此基础上证实了Irgm1对于EAE发病的促进作用，同时揭示了Irgm1在EAE早期的M1型巨噬细胞中高表达，且调控M1型巨噬细胞极化，而Irgm1在体内可调控Th细胞分化，但不直接影响Th17细胞分化，该结果提示我们Irgm1可能通过调节巨噬细胞功能来影响Th细胞分化。另外，我们成功建立M1型和M2型巨噬细胞体外诱导模型，在此基础上沉默M1细胞中Irgm1并检测TNF-α，IL-1β和IL-6的分泌，结果显示Irgm1显著促进TNF-α和IL-1β分泌；将Irgm1敲减的M1细胞和淋巴细胞共培养，结果显示Irgm1敲减降低了M1细胞促Th1和Th17分化作用。后续我们进一步探究了巨噬细胞通过何种细胞因子调控Th1和Th17分化作用，中和抗体分别中和M1细胞-CD4+T细胞共培养体系中TNF-α，IL-1β，结果显示中和TNF-α，IL-1β均能够抑制Th1和Th17分化。脂质体删除EAE体内巨噬细胞的实验也证实了巨噬细胞在疾病早期对Th1，Th17和Treg的调控作用。在Irgm1调节M1细胞极化的机制研究中，结果显示Irgm1在M1分化过程中抑制Akt活化，促进NF-κB活化和入核，Irgm1可能通过Akt/NF-κB通路促进巨噬细胞向M1型转化。综上所述，Irgm1参与EAE发病过程，通过Akt/NF-κB信号通路调控EAE早期M1型巨噬细胞极化和促炎性因子释放，进而间接实现对Th细胞亚型分化的调节作用。我们所取得的成果以科研论文的形式完成，发表SCI文章3篇，培养博士研究生3名，硕士研究生5名，圆满完成了科研课题。