Gfi1双重调节高脂血症小鼠单核巨噬细胞分化与泡沫细胞形成的机制

Atherosclerosis and its resultant obstructive diseases of blood vessel is a leading cause of death in China. The development and progression of atherosclerosis is tightly linked to lipid laden monocytes/macrophages that are named foam cells. Previous studies proved the critical role of Gfi1 in constraining expression of PU.1 which drives multipotential progenitor cells differentiating into monocytes and macrophages. Even though such an important role of Gfi1 in balance of monocytes differentiation is well appreciated, very little is known whether this transcription factors is involved in regulation of monocytes differentiation in hyperlipidemia condition, and more importantly regulation of foam cell formation. In pilot experiments, the applicant discovered that the absence of Gfi1 resulted in 3-fold increase of CD11b+CD115+macrophages in the blood. However, strikingly such Gfi1 deficient macrophages were not able to uptake Ox-LDL and develop into foam cells in in vitro experiments, which was also validated by almost absence of MOMA2 positive cells in the intima of aorta in mice with Gfi1-/-ApoE-/- genotype. In the proposed project, we seek to understand the dual role of Gfi1 in monocyte differentiation, and foam cell formation, in hyperlipidemia condition. We will first perform trancriptomic analyses of the Gfi1 deficient and sufficient macrophages in peripheral blood, and identify the enrichment of genes in pathways regulating cell proliferation, apoptosis, and migration. We will perform immunophenotyping of the Gfi1 deficient macrophages in vitro and in vivo to determine their “pro-inflammation” and/or “anti-inflammation” phenotype. then, we will decipher how the signaling pathways implicated in Ox-LDL lipid uptake is affected in the Gfi1 deficient macrophages. Finally，we will explicit wherther SNAG or ZF domain deficency could affect atherosclerosis.

动脉粥样硬化形成与单核巨噬细胞分化、泡沫细胞形成密切相关。研究表明Gfi1在多能祖细胞中抑制PU.1表达，后者为单核巨噬细胞分化驱动因素。Gfi1对维持单核巨噬细胞分化平衡十分重要，但其在高脂血症条件下对单核巨噬细胞分化、泡沫细胞形成尚未研究。申请人发现Gfi1功能缺失高脂血症小鼠单核巨噬细胞增加3倍。令人吃惊的是，Gfi1突变单核巨噬细胞无法形成泡沫细胞，且无法在动脉内膜下沉积。本课题将从4个方面研究Gfi1在高脂血症条件下对“单核巨噬细胞分化”、“泡沫细胞形成”的双向调节机制：RNA测序分析 Gfi1在高脂血症条件下调控单核巨噬细胞分化的转录组机制；FACS分析Gfi1功能缺失对单核巨噬细胞亚群分化的影响；明确Gfi1突变与正常小鼠脂质代谢信号通路的差异；Gfi1结构域对泡沫细胞形成的影响。明确上述机制对深入认识泡沫细胞形成具有重要意义，将为动脉粥样硬化疾病病因探索、治疗提供新视野。

动脉粥样硬化是一种慢性炎症性疾病，其部分原因是炎症细胞渗入血管壁，导致炎症增强，并在主动脉内皮中积累脂质。然而，炎症细胞，特别是巨噬细胞如何影响动脉粥样硬化的发病机制尚未完全明确。Schlafen4 (Slfn4) mRNA在ox-LDL刺激巨噬细胞后显著上调。然而，Slfn4在泡沫细胞形成中的作用尚不清楚。为了明确Slfn4在动脉粥样硬化过程中是否以及如何调节病变巨噬细胞的功能。我们在ApoE–/–背景上构建了ApoE–/–Slfn4–/–双基因敲除小鼠，并通过体内模型研究了Slfn4缺失对动脉粥样硬化病变形成的影响。我们的研究结果表明，在ApoE–/–小鼠中，SLFN4的完全缺失和SLFN4的骨髓限制缺失均显著减少了动脉斑块内的炎症细胞数量，并限制了高胆固醇血症条件下动脉粥样硬化的发展。这与促炎细胞因子表达的显著降低和活性氧(ROS)的产生相关。此外，在Slfn4缺失显著影响Jnk、Erk和p38的激活。终上所述，本研究发现并证明了Slfn4在调节血管炎症和动脉粥样硬化中的作用，并为心血管相关疾病的治疗提 供了潜在的新靶点。