HDL通过SR-BI调节造血干细胞抑制动脉粥样硬化的作用和机制研究

Inflammatory cells in atherosclerotic plaque are exclusively derived from hematopoietic stem/progenitor cells (HSPCs). Recently, we and Yves-Charvet demonstrated that infusion of reconstituted HDL (rHDL) or lipid poor human apolipoprotein A-I (apoA-I) inhibits hematopoietic stem/progenitor cells (HSPCs) proliferation in hypercholesterolemic Abca1-/-Abcg1-/- mice and C57BL/6 mice, therefore, limiting white blood cell expansion in the peripheral blood (PB). In patients with coronary heart disease, we identified that HDL levels were negatively correlated with white blood cell count and HSPC frequency in PB (unpublished data). In addition, by flow cytometry, we demonstrated that the HDL receptor, scavenger receptor type BI (SR-BI), is expressed on murine and human HSPC. SR-BI is expressed on hepatocytes and facilitates selective cholesterol ester uptake from HDL. Previously, others and we have illustrated the anti-atherosclerotic function of SR-BI in different mouse models. Moreover, SR-BI deficiency is associated with impaired HDL function, intracellular cholesterol accumulation and increased oxidative stress. This led us to hypothesize that SR-BI deficiency impairs the regulation of HDL on HSPC, modulates Akt and MAPK phosphorylation and leads to increased reactive oxygen species production in HSPC, resulting in enhanced HSPC proliferation and differentiation and accelerated atherosclerosis. This study aims to investigate how HDL and SR-BI modulate HSPC proliferation and differentiation in the treatment of atherosclerosis.Ultimately，we hope to provide a new therapeutic target in the treatment of atheroscleross.

目前大多数研究发现高密度脂蛋白(HDL)通过抑制炎症细胞侵入动脉粥样硬化(As)病变血管而起到抗As作用,但对HDL是否、如何抑制炎症细胞的祖/前体细胞,即造血干/前体细胞(HSPC)从而减少炎症细胞数量、抗As尚不明确。我们新近发现①注射重组HDL明显抑制小鼠HSPC的增殖进而减少炎症细胞生成；②HSPC上表达HDL的受体SR-BI,高脂血症的SR-BI缺陷小鼠其HSPC增生、炎症细胞增多、As加重。据此，我们提出"HDL通过SR-BI抑制HSPC增殖、减少炎症细胞生成从而减轻As"科学假说。在本课题中，我们将用高血脂的SR-BI缺陷小鼠模型探索①HDL和SR-BI调节HSPC增殖的主要分子机制，着重于SR-BI对HSPC磷酸化通路、活性氧生成和转录因子的影响；②抑制HSPC增殖对As进程的影响。该课题旨在揭示HDL通过调控HSPC来抑制炎症和As,可望为冠心病治疗提供新的思路。

背景：冠心病的病理基础是动脉粥样硬化（As），斑块上的炎症细胞均来自造血祖、前体细胞(HSPC)，我们报道了注射重组HDL(rHDL) 或apoA-I抑制了C57BL/6小鼠HSPC的增生和高血脂引起的白细胞增多症、人和小鼠HSPC都表达HDL受体,即清道夫BI(SR-BI)。在该课题中，我们验证假说：HDL通过SR-BI调节HSPCs增殖，进而抑制白细胞增多症、炎症和As的进展。.主要研究内容：将野生、SR-BI-/-骨髓细胞移植到致命X线照射的LDLr-/-小鼠移植8周后，开始喂饲高脂，移植16周，取心脏制成组织切片，H&E 染色分析As大小，组化染色CD45和CD45.2,分析移植的HSPC对As炎症细胞的贡献;给喂饲高脂饮食的SR-BI-/-、LDLr-/-apoA-I-/-小鼠皮下注射HDL的主要功能成分apoA-I，与之并行，给喂饲高脂的野生和SR-BI-/-小鼠腹腔注射生理盐水或N-acetylcysteine (NAC),12周后评价对HSPCs增殖、As的抑制作用；从正常和高脂喂饲的野生和SR-BI-/-小鼠分离出骨髓HSPCs，开展RNASeq测序，在转录组水平系统研究HDL/SR-BI调控HSPCs增殖的机制；收集到心脏中心做冠脉造影的病人，流式细胞仪定量HSPCs，多因素回归分析HSPCs频率与冠状动脉狭窄度的相关性。.重要结果和关键数据：(1)骨髓移植实验表明，与野生HSPC相比，SR-BI-/-HSPC增殖分化成更多的炎症细胞；与移植野生骨髓细胞的受体LDLr-/-小鼠相比，移植SR-BI-/-骨髓细胞的LDLr-/-小鼠，高脂喂饲后As比明显加重，且As斑块炎症细胞明显增多。(2)注射apoA-I有效地抑制了LDLr-/-apoA-I-/-小鼠HSPCs的增殖并降低As进展；但对SR-BI-/-小鼠HSPCs的增殖、As没有抑制作用。(3)与注射生理盐水组相比，注射NAC有效的降低了高脂野生和SR-BI-/-小鼠外周血白细胞数量和骨髓HSPCs频率。（4）小鼠HSPCs的RNAseq结果正在分析中。(5)经多因素logistic回归分析和Bonferroni矫正后，HSPCs是所有检测因子中唯一与轻度狭窄到重度狭窄危险性成正相关的因子。.科学意义：HDL通过SR-BI抑制HSPCs增殖进而抑制As进展，HSPCs对As进展中起关键作用。