The analysis of PAR-3 in flow-mediated endothelial planar cell polarity

在发育过程中，血管系统随着血液流动而扩张。现有血管系统的扩张，称为血管生成，必须紧密协调一系列复杂的步骤。以前的工作，包括我的发现，已经揭示了这个过程的分子基础。血管功能的关键调节因子是血管内皮生长因子及其酪氨酸激酶受体（vegfr）。我已经证明，ephrin-B2与分区缺陷3 （PAR-3）和非典型PKC （aPKC）形成蛋白复合物，控制VEGFR信号传导。尽管内皮细胞的协调运动和增殖行为对血管生成至关重要，但内皮细胞的基本功能是在血液和身体其他部分之间建立屏障。内皮细胞对血流动力学剪切应力表现出深刻的形态适应性。内皮细胞有三种不同的细胞极性；发芽的内皮细胞具有前后极性，内皮管具有顶基极性和平行于血流的平面细胞极性（PCP）。因此，极性转换和维护是该过程的关键特征。越来越多的证据表明，PAR-3和aPKC对于控制许多细胞类型和动物物种的细胞极性至关重要。然而，PAR-3和aPKC在血管生成和血管稳态过程中内皮极性的作用尚不清楚。根据这些观察结果，我们研究了PAR-3在生理和病理条件下内皮极性中的作用。我们已经研究了EC特异性基因PAR-3失活的影响。我们的初步数据表明，PAR-3控制EC高尔基体取向，从而控制内皮细胞PCP在体外和体内的流动。此外，我们发现gsk3 β活性受血流控制，而血流在体外被PAR-3敲除而受损。此外，通过VCAM1表达检测的炎症反应在PAR-3 KD培养的ECs和EC特异性KO小鼠中上调。主动脉血流中断的区域通常表现为无组织的高尔基定向。临床观察表明，血管壁剪切应力对动脉粥样硬化病变等血管发病的部位特异性有一定影响。然而，内皮PCP与血管疾病进展之间的因果关系尚未得到证实。目前尚不清楚ECs中PCP紊乱是疾病进展的结果还是原因。提出研究的目的是澄清这一重要问题。总的计划工作将提供实质性的新见解内皮极性的形成和体内稳态。鉴于血管系统的重要功能，了解EC PCP在发病机制中的作用将对基本生物学机制产生新的见解，并可能为未来的药物开发开辟新的令人兴奋的机会。这可能会导致新的治疗策略的发展，以治疗病理。

The vasculature expands substantially during development with blood flow. Expansion of existing vasculature, termed angiogenesis, must tightly coordinate a complex series of steps. Previous works, including my findings, have revealed the molecular basis of this process. Key regulators of blood vessel function are Vascular Endothelial Growth Factor and its tyrosine kinase receptors, VEGFRs. I have shown that ephrin-B2 forms a protein complex with Partitioning defective 3 (PAR-3) and atypical PKC (aPKC), controlling VEGFR signaling. While coordinated motile and proliferative behavior of ECs are crucial for angiogenesis, the fundamental function of ECs is to establish a barrier between the blood and the rest of the body. ECs show profound morphological adaptation to hemodynamic shear stress. ECs have three different cellular polarities; sprouting ECs have front-rear polarity, and established endothelial tubes have apical-basal polarity and planar cell polarity (PCP) parallel to blood flow. Thus, polarity switching and maintenance are a key feature of this process. A growing body of evidence is accumulating to show that PAR-3 and aPKC are crucial for controlling cell polarity across many cell types and animal species. However, the role of PAR-3 and aPKC in endothelial polarity during angiogenesis and blood vessel homeostasis remains elusive. In light of these observations, we have addressed the role of PAR-3 in endothelial polarity in physiological and pathological conditions. We have already examined the effect of EC specific gene inactivation of PAR-3. Our preliminary data have shown that PAR-3 controls EC Golgi orientation and thereby endothelial PCP under flow in vitro and in vivo. Furthermore, we found that GSK3beta activity was controlled by flow, which was impaired by PAR-3 knockdown in vitro. Moreover, inflammatory response examined by VCAM1 expression was upregulated in PAR-3 KD cultured ECs and EC specific KO mice.It is well appreciated that regions of disrupted flow in the aorta often show disorganized Golgi orientation. Clinical observations indicate that shear stress on the blood vessel wall contributes to the site specificity of vessel pathogenesis such as atherosclerosis lesion. However, a causative association between endothelial PCP and vascular disease progression has not yet been shown. It is unclear whether disrupted PCP in ECs is a consequence or causation of disease progression. The goal of proposed study is to clarify this important issue. The sum of the planned work will provide substantial new insights into endothelial polarity formation and homeostasis in vivo. Given the important function of the vasculature, understanding the contribution of EC PCP to pathogenesis will yield novel insights into fundamental biological mechanisms and is likely to open up new exciting opportunities for future drug development. This may well lead to the development of new therapeutic strategies for the treatment of pathologies.