胆固醇转运子ABCA1调控CD4+T细胞免疫应答抑制动脉粥样硬化的新机制研究

Atherosclerosis is a complex disease with the involvement of many cell types and circulating mediators that result in an inflammatory state. Despite the extensive application of lipid lowering drugs, the high prevalence of myocardial infarction and stoke indicates the urgent need for the development/discovery of new therapeutic strategies/targets. ABCA1, a cholesterol transporter, protects against atherosclerosis. However, the effector cells and mechanisms underlying the atheroprotective effects of ABCA1 are still not clear. This greatly hampers the novel ABCA1-based therapeutic strategy for atherosclerosis. Our previous studies indicate that CD4+ T cells are effector cells for bone marrow-derived ABCA1 in prevention of atherosclerosis. In the current study, we will generate T cell-specific ABCA1 knockout (KO)(ABCA1-CD4/-CD4) mice to determine the regulatory function of ABCA1 on the development, phenotype and function of CD4+ T cells. The important molecular events underlying this regulatory function will be identified, e.g. signaling pathways and the phosphorylation of ABCA1. The atherosclerotic lesion development will be induced in both LDLr-/- and LDLr-/-/ABCA1-CD4/-CD4 mice to determine the anti-atherogenic effects of ABCA1 on CD4+ T cells. Finally, we will test the effect of ABCA1 knockdown on the function of human T cells, and compare the ABCA1 expression and function of CD4+T cells between healthy controls and patients of coronary heart diseases. The current study will unravel novel mechanisms for the atheroprotection of ABCA1: through the regulation of CD4+ T cell-mediated immune responses. Identification of underlying molecular events will likely discover novel promising therapeutic targets for atherosclerosis.

动脉粥样硬化的发生发展机制复杂，缺乏理想的治疗药，亟需寻找可调控的靶点。尽管胆固醇转运子ABCA1 具有抗动脉粥样硬化作用，但其作用的靶细胞和机制尚不清楚，这极大地阻碍了其相关治疗靶点的发现。我们前期的研究提示CD4+T细胞可能是骨髓源性ABCA1发挥抗动脉粥样硬化作用的靶细胞。 本研究拟构建T细胞上ABCA1的条件性敲除小鼠, 比较条件敲除鼠与野生型小鼠的CD4+T细胞在发育、表型及功能上的差异, 发现ABCA1调控CD4+T细胞免疫应答的重要功能分子事件, 并在动脉粥样硬化小鼠模型中明确CD4+T细胞上ABCA1的抗动脉粥样硬化作用。最后我们将在人CD4+T细胞上进一步验证ABCA1对CD4+T细胞免疫应答的调控作用，并探讨ABCA1的表达与CD4+T细胞功能和冠心病发病间的相关性。本研究旨在通过对ABCA1抗动脉粥样硬化新机制的探讨，寻找治疗动脉粥样硬化的靶点,为临床提供新的治疗策略

动脉粥样硬化是脂质沉积引发的慢性血管炎症。胆固醇转运子ABCA1可主动将巨噬细胞内过多的胆固醇排出减少泡沫细胞的生成。因此，过去一直认为巨噬细胞是ABCA1发挥抗动脉粥样硬化的靶细胞。但是巨噬细胞特异性敲除ABCA1并不影响动脉粥样硬化的发展。我们前期发现骨髓源性ABCA1的缺失可促进外周血T淋巴细胞的增多及其在病灶管周的聚集，且与病灶的发展呈正相关性。因此，我们大胆的推测T细胞是ABCA1抗动脉粥样硬化的靶细胞。在国家自然科学基金委的支持下，课题组对胆固醇转运子ABCA1调节T淋巴细胞功能及其对动脉粥样硬化发展的影响进行了细致深入的研究。我们首先利用cre-loxp系统构建了低密度脂蛋白基因敲除背景下的T细胞特异性ABCA1基因敲除小鼠，并成功验证了基因敲除的细胞特异性。ABCA1通路的缺失可致T细胞胆固醇流出障碍及细胞表面富含胆固醇的脂筏区增多。尽管特异性ABCA1的缺失并不影响T细胞在体内的发育、生成和表型，但是在-CD3/CD28抗体刺激下，CD4+和CD8+ T细胞都因ABCA1的缺失出现了活化与增殖水平的降低。更重要地是，T细胞特异性缺失ABCA1可显著抑制小鼠体内动脉粥样硬化的发展，且与病灶及管周T细胞的浸润减少相关。这可能是T细胞活化增殖能力减弱，记忆性细胞凋亡增加的结果。有趣地是，临床急性心梗患者外周血中ABCA1的表达水平也显著地高于无冠心病的高血压患者。因此，我们明确了T细胞不是ABCA1抗动脉粥样硬化作用的靶细胞。考虑到B淋巴细胞也可通过T细胞影响炎症的发展，我们构建了B细胞特异性ABCA1敲除小鼠。ABCA1缺失可增强B2细胞的活化、增殖及抗体生成能力，这些结果都提示B细胞可能是ABCA1抗动脉粥样硬化的靶细胞。此外，我们还研究了脂肪ABCA1在动脉粥样硬化发展中的作用，发现了动脉粥样硬化、脂肪炎症、脂肪肝和组织泡沫细胞沉积间的有趣关系。总之，本研究不仅阐明了ABCA1介导的胆固醇流出通路在调控CD4+ T细胞功能的作用，还为动脉粥样硬化的治疗开辟了新的思路。本课题执行期间发表论文5篇，均标注本项目自然科学基金资助。