Mechanisms That Mediate The Absence of Lymphatics in the Retina

描述(申请人提供)：淋巴系统起双重作用：1)从组织中排出间质液体并将其返回血液；2)参与免疫反应。除中枢神经系统(CNS)外，所有有血管的组织都有淋巴管。尽管最近在了解调节淋巴管生成的分子方面取得了重大进展，但对阻止淋巴管生长的因素或条件知之甚少。因此，我们建议研究这个问题，并检验中枢神经系统中淋巴管的缺失是主动抑制过程的结果的假设。我们认为，中枢神经系统淋巴管的抑制是通过Sema3F与NRP2的结合来介导的，NRP2可以阻断血管内皮生长因子C和血管内皮生长因子D的促淋巴效应。我们建议通过以下三个目标来检验这一假说。(1)检测Sema3F、NRP2和VEGF-C在视网膜中的表达，进一步研究视网膜淋巴样结构。将通过原位杂交在出生后的小鼠和成人的视网膜中检测Sema3F和3B。将使用NRP2-LacZ小鼠和原位杂交来评估NRP2的表达。随着时间的推移，将对淋巴样结构进行检查，从P0开始，并将评估其他淋巴标志物的表达，包括NRP2、VEGFR3、PodoPlanin和Prox-1。(2)确定阻断Sema3F是否导致视网膜淋巴管的形成。给予可溶性NRP2或中和Sema3F可中和Sema3F的作用。此外，还将评估血管内皮生长因子-C过度表达的影响。(3)确定Sema3F能否抑制角膜炎症/伤口愈合模型中淋巴管的生成。Sema3F阻断视网膜外淋巴管形成的能力将通过在具有良好特征的角膜缝合模型中注射Sema3F蛋白来检验。

DESCRIPTION (provided by applicant): The lymphatic system plays a dual role: 1) draining interstitial fluid from tissues and returning it to the blood and, 2) participating in the immune response. All vascularized tissues, except the central nervous system (CNS), are invested with lymphatic vessels. Although significant progress has recently been made in understanding the molecules that regulate lymphangiogenesis, very little is known about factors or conditions that deter lymphatic growth. We, thus, propose to investigate this question and to test the hypothesis that the absence of lymphatic vessels in the CNS is the result of an active inhibitory process. We propose that suppression of lymphatic vessels in the CNS is mediated by Sema3F binding to NRP2, which acts to block the pro-lymphatic effects of VEGF-C and VEGF-D. We propose to test this hypothesis with the following aims three aims. (1) To examine the expression of Sema3F, NRP2 and VEGF-C in the retina and to further characterize the lymphatic-like structures in the retina. Sema3F and 3B will be examined by in situ hybridization in postnatal mouse retinas and in the adult. NRP2 expression will be assessed using NRP2-LacZ mice and by in situ hybridization. Lymphatic-like structures will be examined over time, beginning at P0, and will be assessed for expression of other lymphatic markers, including NRP2, VEGFR3, podoplanin and Prox-1. (2) To determine if blocking Sema3F leads to the formation of lymphatic vessels in the retina. The action of Sema3F will be neutralized by administration of soluble NRP2 or neutralizing Sema3F. In addition, the effect of VEGF- C overexpression will be assessed. (3) To determine if the Sema3F administration can suppress lymphangiogenesis in a model of corneal inflammation/wound healing. The ability of Sema3F to block the formation of lymphatic vessels outside of the retina will be examined by injection of Sema3F protein in a well-characterized corneal suture model.