The role of ABCG1 in Atherosclerosis Progression and TH17 Differentiation

描述（由申请人提供）：动脉粥样硬化是导致心血管疾病的主要原因，而心血管疾病是西方世界导致死亡的主要原因。动脉粥样硬化斑块见于动脉壁，由脂质和免疫细胞组成。T淋巴细胞已被证明在动脉粥样硬化中起积极作用。调节性T细胞（Tregs）是表达FoxP3并抑制自身免疫的T细胞的一个亚群。据报道，Tregs具有抗动脉粥样硬化的作用。TH17细胞是表达ROR3t和分泌IL-17的T细胞的一个亚群，IL-17参与促进自身免疫，但其在动脉粥样硬化中的作用存在争议。Tregs和Th17细胞有着密切的发育关系，研究表明Tregs在适当的刺激下能够转化为Th17细胞。ABCG1是一种atp结合盒转运蛋白，通过将多余的胆固醇从外周细胞转运到HDL参与逆向胆固醇转运。造血细胞中缺乏ABCG1的小鼠可以防止发生严重的动脉粥样硬化。提供这种动脉粥样硬化保护的机制目前尚不清楚。我们有初步数据表明，ApoE-/-小鼠喂养致动脉粥样硬化饮食后，treg中ROR3t表达增加；然而，饲喂相同饮食的ApoE-/- ABCG1-/-小鼠的ROR3T表达并未增加。ROR3t是TH17细胞的主要调节因子，其表达的增加可能表明在动脉粥样硬化早期从抗动脉粥样硬化的Tregs细胞向TH17细胞转变。我们也有数据表明，在喂食西方饮食的ApoE-/- abcg1 -/-小鼠中，T细胞产生的IL-17显著减少。我们假设由于TH17分化受损，CD4 T细胞中ABCG1的缺失具有动脉粥样硬化保护作用。本提案将研究ABCG1在动脉粥样硬化进展过程中对CD4+ T细胞功能的作用，具体目的如下：特异性Aim 1将验证CD4+ T细胞中ABCG1缺失抑制TH17分化的假设。我们有初步的数据显示，ABCG1缺失大大降低了TH17细胞的分化。我们将研究这一现象所涉及的机制。特异性目的2将验证CD4+ T细胞中缺乏ABCG1对动脉粥样硬化具有保护作用的假设。我们将通过ABCG1fl/flApoE-/-小鼠与CD4- cre小鼠杂交，在动脉粥样硬化易感小鼠中产生CD4+ T细胞特异性ABCG1缺失。这些小鼠将分别进行8周或20周的致动脉粥样硬化饮食，以研究早期和晚期动脉粥样硬化。然后测量动脉粥样硬化的严重程度。流式细胞术检测动脉粥样硬化进展过程中小鼠主动脉壁和脾脏中TH1、TH2、Tregs和TH17细胞的数量。ABCG1在CD4+ T细胞中的作用，特别是在Tregs和TH17细胞中的作用，可能在促进动脉粥样硬化进展中起重要作用。这项研究可能会澄清目前存在争议的ABCG1和TH17细胞在动脉粥样硬化中的作用，并有助于确定它们作为该疾病治疗靶点的潜力。

DESCRIPTION (provided by applicant): Atherosclerosis is the major cause of cardiovascular disease, which is the leading cause of death in the western world. Atherosclerotic plaques are found in the arterial wall and consist of lipids and immune cells. T lymphocytes have been shown to play an active role in atherosclerosis. Regulatory T cells (Tregs) are a subset of T cells that express FoxP3 and suppress autoimmunity. Tregs have been reported to be antiatherogenic. TH17 cells are a subset of T cells that express ROR3t and secrete IL-17 that are implicated in promoting autoimmunity but their role in atherosclerosis is controversial. Tregs and Th17 cells have an intimate developmental relationship and Tregs have been shown to be able to convert to TH17 cells with the appropriate stimuli. ABCG1 is an ATP-binding cassette transporter that participates in reverse cholesterol transport by transferring excess cholesterol from peripheral cells to HDL. Mice that lack ABCG1 in hematopoietic cells are protected from developing severe atherosclerosis. The mechanism that confers this atheroprotection is currently unknown. We have preliminary data that there is increased ROR3t expression in Tregs from ApoE-/- mice fed an atherogenic diet; however ROR3T expression is not increased in ApoE-/- ABCG1-/- mice fed the same diet. ROR3t is the master regulator of TH17 cells and an increase in its expression may indicate a shift from antiatherogenic Tregs to TH17 cells during early atherogenesis. We also have data that there is a marked reduction in IL-17 production by T cells in ApoE-/-ABCG1-/- mice fed western diet. We hypothesize that absence of ABCG1 in CD4 T cells is atheroprotective due to impaired TH17 differentiation. This proposal will study the role of ABCG1 on CD4+ T cell function during atherosclerosis progression in the following specific aims. Specific Aim 1 will test the hypothesis that absence of ABCG1 in CD4+ T cells inhibits TH17 differentiation. We have preliminary data showing that ABCG1 deficiency greatly reduces the differentiation of TH17 cells. We will investigate the mechanisms involved in this phenomenon. Specific Aim 2 will test the hypothesis that absence of ABCG1 in CD4+ T cells is atheroprotective. We will produce a CD4+ T cell specific deletion of ABCG1 in atherosclerosis prone mice by crossing ABCG1fl/flApoE-/- mice with CD4-Cre mice. These mice will be put on an atherogenic diet for 8 or 20 weeks to investigate early and advanced atherosclerosis respectively. Atherosclerosis severity will then be measured. The numbers of TH1, TH2, Tregs and TH17 cells present in the aortic walls and spleens of the mice during atherosclerosis progression using flow cytometry. The role of ABCG1 in CD4+ T cells, particularly it's role in Tregs and TH17 cells, may be important in promoting atherosclerosis progression. This study may clarify the currently controversial roles of ABCG1 and TH17 cells in atherosclerosis and help determine their potential as therapeutic targets of this disease.