Specification And Patterning of the Lymphatic System

淋巴系统的规格和模式

基本信息

批准号：
7968722
负责人：
Brant Weinstein
金额：
$ 29.65万
依托单位：
EUNICE KENNEDY SHRIVER NATIONAL INSTITUTE OF CHILD HEALTH & HUMAN DEVELOPMENT
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/7968722
关键词：
Anatomy Animal Model Animals Antisense Oligonucleotides Blood Blood Vessels Cardiovascular system Cells Characteristics Clinical Computer Systems Development Development Drainage procedure Dyes Embryo Experimental Genetics Fatty acid glycerol esters Fishes Genes Genetic Screening Goals Growth Histology Homeostasis Human Image Imaging Device Immune response Individual Life Liquid substance Lymph Lymphangiogenesis Lymphangiography Lymphatic Lymphatic Endothelial Cells Lymphatic Endothelium Lymphatic System Lymphatic vessel Lymphedema Malignant Neoplasms Maps Methods Microscopic Modeling Molecular Neoplasm Metastasis Pathologic Processes Pattern Phenotype Play Regulation Resolution Role Route Scanning Transmission Electron Microscopy Procedures Specific qualifier value Structure Technology Time Transgenic Animals Transgenic Organisms Trees Ursidae Family Vascular System Venous Vertebrates Zebrafish absorption in vivo insight interest loss of function migration mutant neuronal guidance novel positional cloning stem time use tool tumor two-photon

项目摘要

Our recent identification and preliminary characterization of a lymphatic vascular system in the zebrafish has made it possible to bring the powerful genetic, experimental embryologic,and imaging tools available in this model organism to bear on the question of how lymphatic vessels form and what regulates their growth and assembly. Understanding how to regulate and control lymphatic vessel formation is a topic of considerable clinical interest given recent evidence suggesting that lymphatics are the major route for tumor metastasis in many if not most cancers. The long-term goal of this project is to expand on our preliminary findings in order to gain a more detailed understanding of how lymphatic vessels assemble in the fish, and then use this powerful model to study how lymphangiogenesis is regulated in vivo. The lymphatic system has become the subject of great interest in recent years because of its important role in normal and pathological processes, but progress in understanding the origins and early development of this system has been hampered by difficulties in observing lymphatic cells in vivo and performing defined genetic and experimental manipulation of the lymphatic system in currently available model organisms. We recently showed for the first time that the zebrafish possesses a lymphatic system that shares many of the morphological, molecular, and functional characteristics of the lymphatic vessels found in other vertebrates (including humans), providing a superb new model for imaging and studying lymphatic development. Using two-photon time-lapse imaging of transgenic zebrafish, we also traced the migration and lineage of individual cells incorporating into the lymphatic endothelium, providing the first conclusive in vivo evidence establishing that early lymphatic endothelial cells are derived from primitive venous blood vessels. We are continuing to examine the assembly and origins of the lymphatic system of the zebrafish. In ongoing studies we are using time-lapse two-photon imaging of transgenic animals, lymphangiography (imaging of dye injected into lymphatic vessels), histology, and scanning and transmission electron microscopy to further characterize how the lymphatic vascular system grows and assembles during development. We are also examining the spatial and temporal expression of lymphatic genes to gain a better understanding of where and how cells become specified as lymphatic endothelial cells. In addition, we are characterizing a number of novel as well as previously characterized genes. Our goal is to determine what functional role they play in lymphatic endothelial specification and differentiation, and in lymphatic vessel formation and growth. We have identified several lymphatic specific mutants in genetic screens and are now carrying out positional cloning and characterization of these mutants. We are also using antisense oligonucleotide knockdown technologies to examine the loss-of-function phenotypes of a number of genes. As one example of the results of these studies we have recently used these knockdown methods to show that netrin, a secreted molecule previously shown to have an important role in neuronal guidance, is also required for lymphatic assembly and patterning. The phenotypes of mutants and antisense oligonucleotide-treated embryos are being assessed by imaging lymphatic vessels in transgenic animals and examining expression of lymphatic marker genes. The results of our studies, combining the genetic and experimental tools available in the zebrafish with the ability to perform high-resolution microscopic imaging of developing vascular structures in living animals, are leading to important new insights into the origins and assembly of the lymphatic system.

我们最近对斑马鱼中淋巴管系统的鉴定和初步表征使该模型生物体中可用的强大遗传，实验性胚胎学和成像工具成为可能，以解决淋巴管如何形成的问题以及是什么调节其生长和组装的问题。鉴于最近的证据表明，在许多癌症中，了解淋巴管是肿瘤转移的主要途径，因此了解如何调节和控制淋巴管的形成是一个相当大的临床兴趣的话题。该项目的长期目标是扩大我们的初步发现，以便对淋巴管如何在鱼中组装，然后使用这种强大的模型来研究如何在体内调节淋巴管生成。由于其在正常和病理过程中的重要作用，淋巴系统已成为引起人们兴趣的主题，但是在了解该系统的起源和早期发展方面的进展受到了在体内观察淋巴细胞的困难，并在当前可用模型生物体中对淋巴细胞的淋巴细胞进行了困难。我们最近首次表明斑马鱼具有一种淋巴系统，该系统具有在其他脊椎动物（包括人类）中发现的淋巴管的许多形态，分子和功能特征，为成像和研究淋巴发育提供了出色的新模型。使用转基因斑马鱼的两光子延时成像，我们还追踪了掺入淋巴内皮中的单个细胞的迁移和谱系，从而在体内证据证明了早期淋巴内皮细胞的第一个结论性证据是从原始静脉血管中得出的。我们正在继续检查斑马鱼的淋巴系统的组装和起源。在正在进行的研究中，我们使用转基因动物的延时两光子成像，淋巴管造影（注射到淋巴管中的染料成像），组织学和扫描和透射电子显微镜进一步表征淋巴管血管系统在开发过程中的生长和组件。我们还研究了淋巴基因的空间和时间表达，以更好地了解细胞在哪里和如何将其指定为淋巴内皮细胞。此外，我们正在表征许多新颖的以及先前表征的基因。我们的目标是确定它们在淋巴内皮规范和分化以及淋巴管的形成和生长中的功能作用。我们已经确定了遗传筛选中的几个淋巴特异性突变体，现在正在进行这些突变体的位置克隆和表征。我们还使用反义寡核苷酸敲低技术来检查许多基因的功能丧失表型。作为这些研究结果的一个例子，我们最近使用了这些敲低方法来表明，淋巴组装和图案化也需要Netrin（先前证明的一个分泌的分子）在神经元引导中具有重要作用。通过成像转基因动物中的淋巴管和检查淋巴标记基因的表达来评估突变体和反义寡核苷酸处理的胚胎的表型。我们的研究结果将斑马鱼中可用的遗传和实验工具与对活动物中发展血管结构进行高分辨率显微成像的能力相结合，这使人们对淋巴系统的起源和组装有了重要的新见解。