Specification And Patterning of the Lymphatic System

我们最近对斑马鱼淋巴管系统的鉴定和初步表征使我们能够利用这种模式生物中可用的强大遗传、实验胚胎学和成像工具来解决淋巴管如何形成以及是什么调节淋巴管的问题。它们的生长和组装。 了解如何调节和控制淋巴管是一个相当大的临床兴趣的话题，最近的证据表明，在许多（如果不是大多数）癌症中，淋巴管是肿瘤转移的主要途径。 这个新成立的项目的长期目标是扩展我们的初步发现，以更详细地了解淋巴管如何在鱼中组装，然后使用这个强大的模型来研究淋巴管生成如何在体内调节。 淋巴系统由于其在正常和病理过程中的重要作用，近年来已成为人们非常感兴趣的主题，但在了解该系统的起源和早期发育方面的进展受到阻碍，因为难以在体内观察淋巴细胞并在目前可用的模型生物体中对淋巴系统进行定义的遗传和实验操作。 我们最近首次表明，斑马鱼拥有一个淋巴系统，该系统与其他脊椎动物（包括人类）中发现的淋巴管具有许多形态，分子和功能特征，为成像和研究淋巴发育提供了极好的新模型。 使用转基因斑马鱼的双光子延时成像，我们还追踪了掺入淋巴内皮的单个细胞的迁移和谱系，提供了第一个确凿的体内证据，证明早期淋巴内皮细胞来源于原始静脉血管。 我们正在继续研究斑马鱼淋巴系统的组装和起源。 在正在进行的研究中，我们正在使用转基因动物的延时双光子成像，淋巴管造影术（注射到淋巴管中的染料成像），组织学，扫描和透射电子显微镜，以进一步表征淋巴管系统在发育过程中如何生长和组装。 我们也在研究淋巴管基因的空间和时间表达，以更好地了解细胞在何处以及如何被指定为淋巴管内皮细胞。 此外，我们已经开始表征一些新的和以前表征的基因，以确定它们在淋巴管内皮细胞的特化和分化，以及淋巴管的形成和生长中发挥什么样的功能作用。 我们正在使用遗传突变体和反义寡核苷酸敲除技术来检查这些基因的功能丧失表型。 这些表型正在通过转基因动物的淋巴管成像和淋巴标记基因的表达进行评估。 初步结果表明，我们正在测试的一些基因参与了血管内皮细胞谱系的初始形成，而其他基因对该系统的模式化很重要。

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 is a topic of considerable clinical interest given recent evidence suggesting that lympahtics are the major route for tumor metastasis in many if not most cancers. The long-term goal of this newly formed project is to expand on our preliminary findings 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 have begun characterization of a number of both novel and previously characterized genes to determine what functional role they play in lymphatic endothelial specification and differentiation, and lymphatic vessel formation and growth. We are using both both genetic mutants and antisense oligonucleotide knockdown technologies to examine the loss-of-function phenotypes of these genes. These phenotypes are being assessed by imaging of lymphatic vessels in transgenic animals and expression of lymphatic marker genes. Preliminary results suggest that some of the genes we are testing are involved in the initial formation of the lympathic endothelial cell lineage while others are important for the patterning of this system.