新型内源性警报素IL-33/ST2途径介导的炎症损伤对狼疮性肾炎肾小球内皮细胞功能的影响及其分子机制研究

What are the antigens that induce Systemic Lupus Erythematosus (SLE)? Are the stimulating antigens foreign (cross-reacting or mimicking "self", such as virus), or are they normal or altered self-molecules (e.g., necrotic bodies) that induce responses that overwhelm tolerance? The innate immune system constitutes an important defense system to respond rapidly to both endogenous and exogenous molecules.Effective sensing of these molecules promotes autoreactivity via effects on immune activation and antigen presentation."Alarmins" are prototypic endogenous pro-inflammatory factors as they are released from necotic cells and provoke local damage or systemic inflammation. Evidences are accumulating to support the inclusion of "Alarmins" as targets of autoreactivity as well as inducers in the pathogenesis of SLE. Interleukin (IL)-33 is a novel member of the family of "Alarmins" because of its characteristics and functions in mediating host immune responses.On this background, we sought to determine the role of IL-33/ST2 axis in lupus pathogenesis.The role of IL-33/ST2 axis has not previously been described in lupus nephritis.This project will study the followings: (1)To determine whether IL-33 was present in renal glomerular endothelial cells; (2); To assess the functional and intracellular signal transuction mechanisms regulating the link between IL-33/ST2-mediated innate immunity and inflammation in CD4+ T cells/CD14+ monocytes-endothelial cells co-culture system of lupus patients; (3) To explore whether the inhibitors of IL-33/ST2 axis can be the potential therapeutic targets using MRLlpr lupus mice.Hypothesis:As a result of external stimuli or infection, renal glomerular endothelial cells undergo cellular death and release the "Alarmin", IL-33, to alert the lupus immune system.Released IL-33 interact with their target cells, CD4+ T cells and CD14+ monocytes via their specific receptor ST2 to subsequently induce innate and adaptive responses, activate inflammatory pathways in the pathogenesis of lupus nephritis.Understanding of the immunopathogenic mechanism of inflammation induced by IL-33 binding to ST2 in lupus nephritis, and evaluation of potential therapeutic role of its inhibitors will provide important biochemical basis for targeting ST2 and the downstream intracellular signaling pathways, which may serve as promising tools to control the inflammation of lupus nephritis.

究竟什么是诱发狼疮的抗原？免疫原性相关因子在狼疮发病中的角色非常值得研究。基于本组前期在外源性危险因子及早期内源性警报素（HMGB1）的工作积累，我们将进一步探讨新型内源性警报素IL-33/ST2途径介导的炎症损伤在狼疮性肾炎发病机制中的作用，本项目拟解决三个关键问题：1.是否IL-33/ST2途径以肾脏为靶点介导狼疮发病？2.如果是，该途径又如何被激活、募集效应细胞发挥生物学效应？3.该通路的负向调节剂是否具有潜在治疗作用？初步结果已证实狼疮肾损伤释放的IL-33来源于肾小球基底膜并可活化表面高表达受体ST2的T细胞及单核细胞。进一步研究将首先运用T细胞/单核细胞-内皮细胞共培养体系阐明IL-33/ST2体外调控机制，研究其细胞间粘附效应及胞内信号功能；运用MRLlpr转基因小鼠研究IL-33对狼疮肾靶点损伤的促进作用；最后应用拮抗剂阻断IL-33/ST2途径，研究靶点抑制剂的生物功能。

研究探讨IL-33/ST2通路介导的炎症损伤在狼疮肾炎（LN）发病中作用：1. 是否IL-33/ST2途径以肾脏为靶点介导狼疮（SLE）发病？2. 该途径如何被激活？运用T细胞/单核-内皮细胞共培养体系阐明IL-33/ST2体外调控机制，研究细胞间粘附效应及胞内信号功能；3. 研究IL-33对狼疮肾靶点的促进作用，阻断IL-33/ST2途径研究靶点抑制剂的生物功能。结果：1. SLE血清中IL-33、sST2表达及与疾病活动相关性：a, SLE血清sST2、IL-33浓度及IL-33/sST2比值均高于正常组；SLE血清sST2浓度与SLEDAI、ESR、24h尿蛋白、甘油三酯正相关，与C3负相关，其中SLEDAI、甘油三酯独立相关；b, SLE血清IL-33浓度与SLEDAI、ESR、24h尿蛋白、CKD分期、甘油三酯正相关，与C3、C4、eGFR、白蛋白负相关，其中SLEDAI、C3、eGFR独立相关；c, SLE 血清IL-33/sST2比值与SLEDAI、eGRF正相关，与病程、起病年龄、激素用量、CKD分期、血肌酐、C3、C4、ALB负相关，其中CKD、白蛋白独立相关，CKD3期以上该比值显著下降；2. IL-33在肾脏的储备细胞及与LN相关性：LN肾组织切片见IL-33表达，与CD34阳性细胞内DAPI共表达；LN肾脏高表达IL-33，与肾脏肿瘤患者相比无统计学意义；3.体外探讨IL-33对人肾脏细胞的影响：IL-33刺激可使内皮细胞-肾小球上皮细胞生长受抑，并释放TNF-a、IL-4、IL-5、IL-6、IL-13，以IL-6为主，但对肾小球系膜细胞无明显影响。结论：1. IL-33/ST2参与SLE发病，血清IL-33、sST2均能一定程度地反映病情，sST2较IL-33与疾病活动度关联性更强；2. IL-33、sST2不能直接反映SLE的肾脏受累，但与间接反映SLE肾损伤的24h尿蛋白及其危险因素有关，IL-33/sST2比值的降低更能提示SLE肾脏的慢性、中重度损害；2. 肾小球内皮细胞是IL-33在LN肾脏的储备细胞之一，细胞损伤后可释放IL-33；3.LN肾脏IL-33基因及蛋白水平表达增加；4.损伤释放的IL-33可能过内皮细胞、肾小管上皮细胞加重LN肾脏损害，并产生以IL-6为主的促炎细胞因子。