The pluripotency factor OCT4 plays an athero-protective role in endothelial cells

多能因子 OCT4 在内皮细胞中发挥动脉粥样硬化保护作用

• 批准号: 10266228
• 负责人: Olga Cherepanova
• 金额: $ 40.25万
• 依托单位: CLEVELAND CLINIC LERNER COM-CWRU
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-30 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10266228
• 关键词: ABCG2 gene; ATP-Binding Cassette Transporters; Adhesions; Aorta; ApoE knockout mouse; Apolipoprotein E; Arterial Fatty Streak; Arteries; Atherosclerosis; Attenuated; Binding; Blood Vessels; Brain; Cell Adhesion Molecules; Cell Lineage; Cells; Cessation of life; ChIP-seq; Cholesterol; Chronic Disease; Clinical; Cluster Analysis; Coronary artery; Data; Development; Disease; Endothelial Cells; Endothelium; Functional disorder; Gatekeeping; Gene Expression Profiling; Genes; Goals; Growth; Heme; High Fat Diet; Human; Impairment; In Vitro; Inflammation; Inflammatory; Inflammatory Response; Knock-out; Lead; Lesion; Lipids; Lung; Mediating; Mesenchymal; Metabolic; Metabolism; Modification; Molecular; Mus; Myocardial; Myocardial Infarction; NF-kappa B; Oxidative Stress; Pathogenesis; Phenotype; Play; Population Heterogeneity; Porphyrins; Production; Proteins; Pulmonary artery structure; Role; Signal Transduction; Site-Directed Mutagenesis; Smooth Muscle Myocytes; Somatic Cell; Specimen; Testing; Therapeutic; Time; Tissue-Specific Gene Expression; Up-Regulation; Vascular Cell Adhesion Molecule-1; Vascular Smooth Muscle; Xenobiotics; atherogenesis; atheroprotective; base; cardiac repair; cell injury; chromatin immunoprecipitation; cytokine; embryonic stem cell; gene repression; in vivo; indexing; induced pluripotent stem cell; intercellular cell adhesion molecule; macrophage; monocyte; mouse model; neuroinflammation; novel; novel therapeutic intervention; oxidized lipid; pluripotency; prevent; promoter; recruit; response; single-cell RNA sequencing; stem cell division; transcriptome; western diet

ABSTRACT Atherosclerosis is a chronic disease of the arterial wall which contributes to >40% of all deaths in the USA. Although previous studies provide clear evidence for the importance of endothelial cell (EC) activation and phenotypic modulation during atherosclerosis, little is known about key regulatory mechanisms for EC plasticity that influence lesion growth and/or stability. Recently, we found that the embryonic stem cell/iPS factor OCT4 that was believed to be silenced in somatic cells plays an athero-protective role in smooth muscle cells (SMC) by regulating SMC phenotypic transitions. We then asked if OCT4 might also play a key role in regulating plasticity of EC during atherosclerosis development. Our preliminary results using a novel EC-specific Oct4 knockout Apoe-/- mouse model demonstrated that loss of Oct4 in EC increases the total lipid burden within aortas and occurrence of plaques within pulmonary arteries of Apoe-/- mice fed a long-term high-fat diet, indicating that OCT4 plays a key functional role within EC. To further understand molecular mechanisms whereby EC-derived OCT4 inhibits the accumulation of lipids we performed a single cell (sc)RNA-seq analysis on the aortic and lung EC sorted from EC-Oct4 knockout and wild-type Apoe-/- mice fed a Western diet for five weeks. Differential gene expression analyses identified 405 (aorta), and 250 (lung) significantly dysregulated genes in Oct4 knockout EC as compared to wild type EC, including upregulation of genes involved into Endothelial-to-Mesenchymal transition (Endo-MT), pro-inflammatory and adhesion molecules, and down-regulation of genes involved into xenobiotic and heme metabolism. Also, using results from the in vivo OCT4 ChIP-seq, we found that one of the genes significantly down-regulated in the Oct4 knockout lung and aortic EC, Abcg2, is a putative target of OCT4. ABCG2 is an ATP-binding cassette transporter responsible for extruding toxic xenobiotics, including heme from cells. Previous studies demonstrated that EC-derived ABCG2 plays a pivotal role in cardiac repair after myocardial infarction and protective role against oxidative stress and neuroinflammatory response in mouse brains by inhibiting NF-kB-signaling. However, there is no data about the potential protective role of ABCG2 in EC during atherosclerosis development. Our overall hypothesis is that the pluripotency factor OCT4 plays a critical athero-protective role in EC during the development of atherosclerosis through activation of the gatekeeper protein ABCG2, an adaptive mechanism to decrease oxidative stress and protect EC from ROS- induced activation and inflammatory response. Aim 1 will test the hypothesis that OCT4 plays an athero- protective role by preventing EC from activation and phenotypic modification at the early and late stages of atherosclerosis. Aim 2 will test the hypothesis that OCT4 attenuates EC-activation and EC inflammatory response at least in part through up-regulating Abcg2 that protects EC from oxidative stress by decreasing intracellular heme levels and ROS-mediated NF-kB-dependent EC inflammatory response. These studies will lead to an identification of novel cell-specific therapeutic approaches for treating or preventing atherosclerosis.

摘要 动脉粥样硬化是一种动脉壁的慢性疾病，在美国占所有死亡人数的40%以上。 虽然先前的研究提供了明确的证据表明内皮细胞（EC）活化的重要性， 动脉粥样硬化过程中的表型调节，对EC可塑性的关键调节机制知之甚少 其影响损伤生长和/或稳定性。最近，我们发现胚胎干细胞/iPS因子OCT 4 据信在体细胞中沉默，在平滑肌细胞（SMC）中发挥动脉粥样硬化保护作用 通过调节SMC表型转变。然后，我们问OCT 4是否也可能在调节 动脉粥样硬化发展过程中EC的可塑性。我们使用一种新的EC特异性Oct 4 ApoE基因敲除小鼠模型证实EC中Oct 4缺失增加了动脉内的总脂质负荷 和Apoe-/-小鼠肺动脉内斑块的发生，表明 OCT 4在EC中起着关键的功能作用。为了进一步了解EC衍生的 我们对主动脉和肺进行了单细胞（sc）RNA-seq分析， EC分选自EC-Oct 4敲除和野生型Apoe-/-小鼠，喂食西方饮食五周。差异基因 在Oct 4基因敲除的EC中，表达分析鉴定出405个（主动脉）和250个（肺）显著失调的基因。 与野生型EC相比，包括参与内皮细胞到间充质细胞的基因的上调， 过渡（Endo-MT），促炎和粘附分子，以及下调基因参与 异生素和血红素代谢。此外，使用来自体内OCT 4 ChIP-seq的结果，我们发现其中一个 在Oct 4敲除的肺和主动脉EC中显著下调的基因Abcg 2是OCT 4的推定靶标。 ABCG 2是一种ATP结合盒转运蛋白，负责从细胞中排出有毒的外源性物质，包括血红素。 细胞先前的研究表明，EC衍生的ABCG 2在心脏损伤后的心脏修复中起关键作用。 小鼠心肌梗死及其对氧化应激和神经炎症反应的保护作用 通过抑制NF-kB信号传导。然而，没有关于ABCG 2在细胞凋亡中的潜在保护作用的数据。 动脉粥样硬化发展过程中的EC。我们的总体假设是，多能性因子OCT 4发挥作用， 在动脉粥样硬化的发展过程中，EC通过激活 门控蛋白ABCG 2是一种降低氧化应激和保护EC免受ROS侵害的适应性机制。 诱导活化和炎症反应。目的1将检验OCT 4在动脉粥样硬化中起作用的假设。 通过阻止EC在早期和晚期的活化和表型修饰来保护作用。 动脉粥样硬化目的2将检验OCT 4减弱EC活化和EC炎性的假设。 至少部分地通过上调Abcg 2来保护EC免受氧化应激， 细胞内血红素水平和ROS介导的NF-κ B依赖性EC炎症反应。这些研究将 导致鉴定用于治疗或预防动脉粥样硬化的新的细胞特异性治疗方法。