CTLA4Ig诱导对氧化修饰低密度脂蛋白免疫耐受的机制研究

目的： 动脉粥样硬化（AS）是一种炎症过程，获得性免疫应答参与AS发生和发展。氧化修饰的低密度脂蛋白（ox-LDL）是目前认为最重要的AS相关自身抗原。本研究拟应用融合蛋白CTLA4Ig，在体外建立对ox-LDL的免疫耐受模型，从而有可能预防免疫应答导致的炎症损伤在AS发病中的作用，为防治AS提供新的策略。方法: 分离人外周血单个核细胞诱导树突状细胞（DC）。分别加入LPS、LDL、 ox-LDL刺激48 h，与同种异体淋巴细胞行混合淋巴细胞反应（MLR）。ox-LDL组的MLR中，分别加入不同浓度的CTLA4Ig。以MTT法检测T细胞的增殖。流式细胞仪检测MLR中T细胞活化和T细胞凋亡。ELISpot检测MLR中T细胞分泌IL-2、IFN-γ和IL-4的情况。结果: ox-LDL组MTT中的刺激指数（SI）明显高于LDL组（P<0.05）；应用CTLA4Ig后，SI较未应用时明显降低（P<0.05，P<0.01）；CTLA4Ig可明显减少T细胞CD25的表达（P<0.05，P<0.01），增加T细胞的凋亡（P<0.05，P<0.01）。CTLA4Ig可减少T细胞分泌IL-2和IFN-γ的ELISpot计数(P<0.01），增加IL-4的ELISpot计数（P<0.05）。结论: CTLA4Ig可在体外诱导对ox-LDL的免疫耐受；CTLA4Ig通过抑制T细胞活化、诱导T细胞凋亡和促进Th1/Th2免疫偏移等机制，诱导免疫耐受。

CTLA4Ig induces immune tolerance of T cells to oxidized-low density lipoprotein in vitro

AIM: Recently,it is widely accepted that atherosclerosis (AS) is an auto-immune related disease and the oxidized-low density lipoprotein (ox-LDL) is the most important AS-related antigen.In order to prevent immune injuries in AS and find new strategies to prevent AS,the immune tolerance of T cells to ox-LDL in vitro was induced in this study.METHODS: Human monocytes were separated from peripheral blood to induce dendritic cells (DCs).DCs were treated with LPS (30 μg/L),ox-LDL (10 mg/L) and LDL (10 mg/L) for 48 h.Then DCs were mixed with allogenic T lymphocytes to carry out mixed lymphocytes reaction (MLR).CTLA4Ig in different concentrations was added in the MLR of ox-LDL group.MTT method was used to assay the proliferation of T cells and expressed in stimulation index (IS).The CD25 expression and apoptosis of T cells in MLR were tested by flow cytometry.The excretion of IL-2,IFN-γ and IL-4 was assayed by ELISpot method.RESULTS: SI in ox-LDL group was higher than that in LDL group significantly (P<0.05) and CTLA4Ig inhibited the SI in ox-LDL group with dose-dependent effect (P<0.05,P<0.01).CTLA4Ig decreased the CD25 expression (P<0.05,P<0.01) and induced apoptosis of T cells in MLR (P<0.05,P<0.01).CTLA4Ig decreased the ELISpot counts of IL-2 and IFN-γ (P<0.01),while increased that of IL-4 (P<0.05).CONCLUSION: CTLA4Ig induces T cells tolerance to ox-LDL in vitro.CTLA4Ig inhibits T cells activation,promotes T cells apoptosis and Th1/Th2 immune deviation,which is the important mechanism in it′s induction of tolerance.