VEGF-C/VEGFR3 Regulation of VE-Cadherin in Sinusoidal Angiogenesis and Lymphangiogenesis

VEGF-C/VEGFR3 对 VE-钙粘蛋白在正弦血管生成和淋巴管生成中的调节

• 批准号: 10563110
• 负责人: Derek C Sung
• 金额: $ 3.41万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10563110
• 关键词: Ablation; Binding; Blood; Blood Vessels; Bone Marrow; Cell Culture Techniques; Cells; Cre lox recombination system; Data; Defect; Development; Discontinuous Capillary; Edema; Electrical Resistance; Embryo; Embryonic Development; Endocytosis; Endothelial Cells; Endothelium; Erythroid; Fetal Liver; Fluid Balance; Genetic; Goals; Growth; Growth and Development function; Hematopoiesis; Histologic; Human; Immunofluorescence Immunologic; In Vitro; KDR gene; Knockout Mice; Ligands; Liver; Liver Regeneration; Lymphangiogenesis; Lymphatic; Lymphatic Diseases; Lymphatic Endothelial Cells; Measures; Mediating; Membrane; Modeling; Molecular; Mus; Organ; Pathway interactions; Permeability; Pharmacology; Phenocopy; Phenotype; Phosphorylation; Process; Regulation; Research; Role; Signal Transduction; Signaling Molecule; Skin; Small Interfering RNA; Stains; Testing; Tissues; VEGFA gene; Vascular Endothelial Growth Factor C; Vascular Endothelial Growth Factor Receptor-3; Vascular Endothelial Growth Factors; Western Blotting; Work; angiogenesis; beta-arrestin; blood vessel development; cadherin 5; cell growth; gain of function; improved; in vivo; inhibitor; insight; lymphatic malformations; lymphatic vasculature; novel; reconstitution; regenerative therapy; therapeutic target; vascular bed

Research Summary The blood, lymphatic, and sinusoidal vasculatures are molecularly and functionally distinct vascular networks whose normal growth and development are essential to support embryogenesis. While there has been notable effort to understand mechanisms of blood vessel development, relatively little is known about the role of vascular endothelial growth factor (VEGF) receptor 3 (VEGFR3) and its main ligand VEGF-C in sinusoidal development. Despite decades of work on VEGFR2 and VEGFR3 signaling, insights into the field have been complicated by use of various genetic and pharmacologic models with divergent phenotypes. Furthermore, VEGF-C can weakly bind and activate VEGFR2 in addition to inducing VEGFR2-VEGFR3 heterodimers. As a result, the role of VEGF-C in vascular development remains incompletely understood. Therefore, our goal is to determine the role of VEGF-C/VEGFR3 in vascular development and identify novel signaling mechanisms through which it functions through a combination of in vivo mouse and in vitro cell culture studies. We and others have previously identified VEGF-C as an important regulator of fetal liver hematopoiesis. This was primarily attributed to defects in erythroid maturation, however the sinusoids are the only VEGFR3-positive cells in the fetal liver. Strikingly, endothelial deletion of VEGFR3 and vascular endothelial (VE-)cadherin gain- of-function mice both phenocopy VEGF-C knockout mice, suggesting that diminished fetal liver hematopoiesis is in fact due to sinusoidal defects. These mice also have decreased bone marrow (BM) sinusoids. BM defects due to endothelial loss of VEGFR3 are rescued by VE-cadherin haploinsufficiency, demonstrating that VEGFR3 negatively regulates VE-cadherin to promote sinusoidal growth. Whether this process depends on VEGF-C and the signaling mechanisms through which VEGFR3 regulates VE-cadherin remain unclear. Therefore, I hypothesize that VEGF-C activates VEGFR3 to directly and negatively regulate VE-cadherin during sinusoidal angiogenesis and lymphangiogenesis. Aim 1 will test whether haploinsufficiency of VE- cadherin can rescue fetal liver and lymphatic growth defects, as I have demonstrated to be true in the bone marrow sinusoids. Aim 2 will test whether VEGFR3 functions in a VEGF-C-dependent or independent manner in the skin, liver, and bone marrow during sinusoidal angiogenesis and lymphangiogenesis. Finally, Aim 3 will determine the mechanism by which VEGFR3 negatively regulates VE-cadherin through in vitro studies using human lymphatic endothelial cells. Together, these studies will bring novel insights into the mechanism by which VEGF-C/VEGFR3 may regulate VE-cadherin across various vascular beds in multiple organs, which could have important implications for liver regeneration, BM reconstitution, and lymphatic disorders.

研究综述 血液、淋巴管和窦状隙血管在分子和功能上是不同的血管网 其正常生长和发育对于支持胚胎发生是必需的。虽然有值得注意的 尽管人们努力了解血管发育的机制，但对血管发育的作用知之甚少。 血管内皮生长因子（VEGF）受体3（VEGFR 3）及其主要配体VEGF-C 发展尽管对VEGFR 2和VEGFR 3信号转导进行了数十年的研究，但对该领域的见解仍然存在。 由于使用具有不同表型的各种遗传和药理学模型而变得复杂。此外，委员会认为， 除了诱导VEGFR 2-VEGFR 3异二聚体之外，VEGF-C还可以弱结合并激活VEGFR 2。作为 因此，VEGF-C在血管发育中的作用仍不完全清楚。因此，我们的目标是 确定VEGF-C/VEGFR 3在血管发育中的作用，并确定新的信号传导机制 其通过体内小鼠和体外细胞培养研究的组合发挥作用。我们和 其他人先前已经鉴定VEGF-C是胎儿肝脏造血的重要调节剂。这是 主要归因于红细胞成熟缺陷，然而血窦是唯一的VEGFR 3阳性 胎儿肝脏中的细胞引人注目的是，内皮细胞VEGFR 3缺失和血管内皮细胞（VE-）钙粘蛋白获得- 功能缺陷小鼠均表现为VEGF-C基因敲除小鼠，表明胎肝造血功能减弱 实际上是由于正弦缺陷。这些小鼠还具有减少的骨髓（BM）窦状隙。BM缺陷 由于内皮细胞损失的VEGFR 3被VE-钙粘蛋白单倍不足所挽救，表明 VEGFR 3负调节VE-钙粘蛋白以促进窦状隙生长。这个过程是否取决于 VEGF-C和VEGFR 3调节VE-钙粘蛋白的信号传导机制尚不清楚。 因此，我推测VEGF-C激活VEGFR 3直接负调控VE-钙粘蛋白 在窦状血管生成和淋巴管生成期间。目的1将测试VE-1基因的单倍不足是否 钙粘蛋白可以挽救胎儿肝脏和淋巴生长缺陷，正如我在骨骼中所证明的那样 骨髓窦目标2将测试VEGFR 3是否以VEGF-C依赖或独立的方式发挥功能 在窦状血管生成和淋巴管生成过程中，在皮肤、肝脏和骨髓中。最后，Aim 3将 通过体外研究确定VEGFR 3负调节VE-钙粘蛋白的机制， 人淋巴管内皮细胞总之，这些研究将带来新的见解的机制， 其中VEGF-C/VEGFR 3可以调节VE-钙粘蛋白跨越多个器官中的各种血管床， 可能对肝再生、骨髓重建和淋巴系统疾病有重要意义。