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.

研究总结 血液、淋巴管和血窦血管在分子和功能上是不同的血管网络。 它们的正常生长和发育对支持胚胎发育是必不可少的。虽然有值得注意的是 为了了解血管发育的机制，人们对血管生长因子的作用知之甚少 血管内皮细胞生长因子受体3及其主要配体血管内皮生长因子C在肝窦中的表达 发展。尽管在VEGFR2和VEGFR3信号方面做了几十年的工作，但对该领域的见解 通过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， VEGF-C除了能诱导VEGFR2-VEGFR3异源二聚体外，还能弱结合和激活VEGFR2。作为一名 结果，血管内皮生长因子-C在血管发育中的作用尚不完全清楚。因此，我们的目标是 确定VEGF-C/VEGFR3在血管发育中的作用并确定新的信号机制 通过体内小鼠和体外细胞培养研究相结合的方式发挥作用。我们和 此前已有研究表明，血管内皮生长因子-C是胎肝造血的重要调节因子。这是 主要归因于红系成熟缺陷，然而，血窦是唯一的VEGFR3阳性 胎儿肝脏中的细胞。值得注意的是，血管内皮细胞VEGFR3的缺失和血管内皮细胞(VE-)钙粘附素的增加- 功能不全的小鼠两种表型均为血管内皮生长因子-C基因敲除小鼠，提示胎肝造血功能减弱 实际上是由于正弦缺陷造成的。这些小鼠的骨髓(BM)正弦波也减少了。黑石缺陷 由于内皮细胞的丢失，VEGFR3被VE-钙粘蛋白单倍体功能不全所拯救，表明 VEGFR3负性调节VE-钙粘蛋白促进肝窦生长。这一过程是否取决于 VEGF-C和VEGFR3调节VE-钙粘蛋白的信号机制尚不清楚。 因此，我推测，VEGF-C激活VEGFR3直接负向调节VE-钙粘附素 在正弦血管生成和淋巴管生成过程中。目标1将测试VE是否单倍性不足- 钙粘附素可以挽救胎儿肝脏和淋巴生长缺陷，我已经在骨骼上证明了这一点。 骨髓血窦。AIM 2将测试VEGFR3是否以依赖或独立的方式发挥作用 在皮肤、肝脏和骨髓中，在正弦血管生成和淋巴管生成过程中。最后，《目标3》将 通过体外研究确定VEGFR3负调控VE-钙粘附素的机制 人淋巴管内皮细胞。总之，这些研究将通过以下方式为这一机制带来新的见解 哪个VEGF-C/VEGFR3可以调节VE-钙粘附素跨多个器官的不同血管床， 可能对肝脏再生、骨髓重建和淋巴疾病有重要意义。