Pathological flow-induced endothelial damage and plaque erosion

项目摘要/摘要 动脉粥样硬化和高危斑块，包括破裂和侵蚀，主要局限于血管。 非层流扰动血流(d-flow)和层流高血流(h-flow)的壁区。然而， 这种病理性血流诱导的高危斑块的分子机制主要尚不清楚。 由于缺乏这样的高危斑块动物模型，因此迫切需要纠正这一差距。 我们的长期目标是确定高危斑块是如何通过病理性血流形成的。我们发现 在他莫昔芬诱导的LATS(大肿瘤抑制同源物)1/2缺失后14天内 内皮细胞(EC)特异性LATS1Homo/LATS2Homokokout小鼠(LATS1Homo/LATS2Homo-EKO)，ALL小鼠 (28/28)死于严重的全身性水肿，并伴有大量的EC细胞凋亡。接下来，我们生成EC- 特异性LATS1het/LATS2 Homo-EKO高胆固醇血症(H-CHOL)小鼠；我们检测到斑块侵蚀样病变 在主动脉弓和颈动脉的h-和d-血流区域，显示强纤维蛋白/纤维蛋白原阳性组织。 血栓形成，无大的坏死核。我们观察到1)EC增殖显著增加，2) EC凋亡伴衰老表型，3)组织因子(TF)表达，4)炎症，它们是 统称为“血栓前表型”。我们假设病理性流动引起的 LATS1/2降解促进内皮细胞损伤介导的血栓形成和管腔再内皮化 演唱会。这种EC损伤-血栓-再内皮化的循环导致大血栓的形成和 斑块侵蚀。病理性血流诱导的PKC对MAGI1S741的磷酸化 LATS1/2-TERF2IP-MKRN1复合体的形成是LATS1/2降解所必需的。为了检验这一假设， 我们提出了以下三个目标。在目标1中，我们将描述EC过度增殖的作用， 斑块侵蚀样形成中的衰老/凋亡、组织因子(TF)表达和炎症 EC特异性Lats1het/Lats2homo基因敲除(Lats1het/Lats2homo-EKO)小鼠的病变和斑块内出血 在H-chol之下。在目标2中，我们将确定依赖于磷酸化的TERF2IP和MKRN1的关键作用 Ub E3连接酶结合在病理性血流诱导的LATS1/2失稳和随后的血栓形成中的作用 在目标3中，我们将研究PKC诱导的MAGI1S741磷酸化在细胞中的作用。 LATS1/2介导TERF2IP S205的磷酸化和LATS1/2的降解。拟议的工作是可望完成的 建立病理性血流和血栓前表型之间的联系，通过这种联系高危斑块是 形成了。这项拟议的研究是创新的，因为它将提出一个新的概念，即病理性流动如何影响 血管内皮细胞和诱导形成易受侵蚀的斑块，并将提供新的见解 导致这种病理的信号级联反应和分子。拟议的研究还可以提供 预测和预防高危斑块形成的手段。

Project Summary/Abstract Atherosclerotic and high-risk plaques including both rupture and erosion are predominantly localized to vessel wall regions with non-laminar disturbed blood flow (d-flow) and laminar high flow (h-flow). However, the molecular mechanisms of this pathological flow-induced high-risk plaque remain largely unknown mainly because of the lack of such high-risk plaque animal models, hence there is an urgent need to correct this gap. Our long-term goal is to determine how the high-risk plaque is formed by pathological flow. We found that within 14 days after inducing LATS (large tumor suppressor homolog) 1/2 deletion in tamoxifen-inducible endothelial cell (EC) specific LATS1homo/LATS2homoknock-out mice (LATS1homo/LATS2homo-EKO), all mice (28/28) died of severe systemic edema, accompanied by massive EC apoptosis. Next we generated EC- specific LATS1het/LATS2homo-EKO hypercholesteremic (H-chol) mice; we detected a plaque erosion-like lesions at h- and d-flow areas in the aortic arch and carotids, which revealed strong fibrin/fibrinogen positive organized thrombus formation without a large necrotic core. We observed significant increases in 1) EC proliferation, 2) EC apoptosis with senescent phenotype, 3) tissue factor (TF) expression, and 4) inflammation, which are collectively referred to as the “pro-thrombotic phenotype”. We hypothesize that pathological flow-induced LATS1/2 degradation promotes EC damage-mediated thrombus formation and luminal re-endothelialization in concert. This cycle of EC damage-thrombus-re-endothelialization results in largethrombus formation and plaque erosion. Pathological flow-induced MAGI1 S741 phosphorylation by PKC, and the subsequent LATS1/2-TERF2IP-MKRN1 complex formation are essential for LATS1/2 degradation. To test ths hypothesis, we propose the following 3 aims. In aim 1, we will characterize the role of excess EC proliferation, senescence/apoptosis, tissue factor (TF) expression, and inflammation in the formation of plaque erosion-like lesions and intraplaque hemorrhage in EC specific Lats1het/Lats2homo knock-out (Lats1het/Lats2homo-EKO) mice under H-chol. In aim 2, we will determine the crucial role of phosphorylation dependent TERF2IP and MKRN1 Ub E3 ligase binding in pathological flow-induced LATS1/2 destabilization and the consequent pro-thrombotic phenotype, and in aim 3 we will Investigate the role of PKC-induced MAGI1 S741 phosphorylation in LATS1/2-mediated TERF2IP S205 phosphorylation and LATS1/2 degradation. The proposed work is expected to establish the link between pathological flow and pro-thrombotic phenotype, by which high-risk plaques are formed. The proposed study is innovative because it will propose a new concept how pathological flow affects the endothelium and induce the formation of vulnerable plaques of erosion and will provide insights into new signaling cascades and molecules responsible for this pathology. The proposed research may also provide means to predict and prevent high-risk plaque formation.