Regulation of vascular leakage in age related macular degeneration

年龄相关性黄斑变性中血管渗漏的调节

• 批准号: 8385057
• 负责人: MEREDITH GREGORY-KSANDER
• 金额: $ 30.02万
• 依托单位: SCHEPENS EYE RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-01 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8385057
• 关键词: Address; Age related macular degeneration; Age-Years; Agonist; American; Angiogenic Factor; Animal Model; Apoptosis; Apoptotic; Atrophic; Binding; Binding Proteins; Blindness; Blood; Blood Vessels; Cell surface; Cells; Choroidal Neovascularization; Cleaved cell; Data; Development; Doxycycline; Endothelial Cells; Extravasation; Eye; Failure; Fluorescein Angiography; Gene Targeting; Goals; Growth; Immune; Induction of Apoptosis; Inflammatory; Inflammatory Infiltrate; Integral Membrane Protein; Interleukin-1; Isolectin; Knock-in Mouse; Laboratories; Lasers; Lead; Lesion; Liquid substance; Maintenance; Mediating; Membrane; Metalloproteases; Modeling; Mouse Strains; Mus; Mutate; Optical Coherence Tomography; Pathogenesis; Pathway interactions; Patients; Phenotype; Play; Population; Protein Isoforms; Proteins; Recombinants; Regulation; Reporting; Retina; Retinal; Role; Site; Staining method; Stains; Structure of retinal pigment epithelium; System; TNF gene; Therapeutic; Time; Tissues; Transgenic Mice; Tumor Necrosis Factor Ligand Superfamily Member 6; Vascular Endothelial Cell; Vascular Endothelial Growth Factors; Vision; Wild Type Mouse; Work; bevacizumab; cell type; cytokine; cytotoxic; extracellular; in vivo; intravitreal injection; macrophage; mouse model; neovascular; new growth; new therapeutic target; novel; overexpression; photoreceptor degeneration; prevent; protein function; success

DESCRIPTION (provided by applicant): Age-related macular degeneration (AMD) is a leading cause of blindness in Americans 60 years of age and older. There are two forms of AMD: Dry or atrophic and Wet or exudative. However, the Wet form of AMD, which is characterized by choroidal neovascularization (CNV), is associated with more severe vision loss caused primarily by the leakage of fluid and blood from the abnormal vessels. Anti-VEGF therapy has emerged as the treatment of choice for patients with wet AMD. However, some patients never respond to anti-VEGF therapy, while still others stop responding after initial success, highlighting the fact that the pathobiology of wet AMD development and progression is still unknown. FasL is an important protein in maintaining immune privilege in the eye, where it is thought to induce apoptosis of infiltrating inflammatory cells. In fact, several reports indicate that FasL expressed on retinal pigment epithelial cells inhibits choroidal neovascularization by triggering apoptosis o vascular endothelial cells. However, FasL exists as both a membrane bound and soluble protein with potentially opposing functions and it is unclear how the different forms of FasL contribute to the development of CNV. We recently developed a unique knock-in mouse strain in which the FasL metalloproteinase cleavage sites were mutated to prevent cleavage of the membrane-bound protein. In these ¿CS mice, FasL is expressed by the physiologically relevant cell types, but these cells are unable to cleave FasL and therefore, can only express mFasL (termed ¿CS mice). For the first time, these mice have allowed us to study the in vivo function of membrane FasL in the absence of soluble FasL. To better understand the function of the different FasL isoforms in the development of choroidal neovascularization, we used a laser-induced murine model of CNV. Our preliminary data indicate that, in ¿CS mice, either the increased expression of mFasL (or the loss of sFasL) prevents vascular leakage following laser induced-CNV. Spectral domain optical coherence tomography (SD-OCT) and Isolectin-B staining demonstrated no difference in the size of the neovascular lesion between ¿CS mice and wild-type mice. However, fluorescein angiography revealed a significant decrease in vascular leakage in ¿CS mice as compared to wild-type mice. These results indicate that membrane FasL does not prevent the ingrowth of choroidal vessels, as previously documented, but rather plays a critical function in preventing vascular leakage, the primary cause of vision loss in patients with wet AMD. We hypothesize that membrane FasL inhibits vascular leakage in laser-induced CNV through Fas mediated pathways triggered in infiltrating macrophages and/or RPE cells. The goals of this proposal are as follows: (Aim 1) Determine whether the decreased vascular leakage in ¿CS mice is due to increased mFasL and/or a lack of sFasL and whether the decreased leakage coincides with a shift in the M1/M2 phenotype of infiltrating macrophages (Aim 2) Establish whether infiltrating macrophages and/or RPE are the critical Fas+ targets through which mFasL mediates inhibition of vascular leakage. PUBLIC HEALTH RELEVANCE: Age Related Macular Degeneration (AMD) is the primary cause of blindness in patients 60 years of age and older. The growth of abnormal blood vessels into the retina is the main cause of severe vision loss, in part due to vessel leakage of blood and fluid that causes damage to the surrounding retinal tissue. We have identified a novel protein that prevents vascular leakage and the goal of this study is to (i) determine whether the administration of this protein can be used to prevent vascular leakage in a mouse model, and (ii) elucidate the mechanism by which this protein functions.

描述（由申请人提供）：视网膜相关性黄斑变性（AMD）是导致60岁及以上美国人失明的主要原因。有两种形式的AMD：干性或萎缩性和湿性或渗出性。然而，以脉络膜新生血管形成（CNV）为特征的湿性AMD与主要由流体和血液从异常血管渗漏引起的更严重的视力丧失相关。抗VEGF疗法已成为湿性AMD患者的治疗选择。然而，一些患者从未对抗VEGF治疗作出反应，而另一些患者在初步成功后停止反应，突出了湿性AMD发展和进展的病理生物学仍然未知的事实。FasL是维持眼睛中免疫赦免的重要蛋白质，其中它被认为诱导浸润性炎性细胞的凋亡。事实上，一些报告表明FasL表达了 通过触发血管内皮细胞凋亡抑制脉络膜新生血管形成。然而，FasL作为具有潜在相反功能的膜结合蛋白和可溶性蛋白两者存在，并且不清楚不同形式的FasL如何促进细胞凋亡。 CNV的发展。我们最近开发了一种独特的敲入小鼠品系，其中FasL金属蛋白酶切割位点发生突变，以防止膜结合蛋白的切割。在这些在CS小鼠中，FasL由生理相关的细胞类型表达，但这些细胞不能切割FasL，因此只能表达mFasL（称为CS小鼠）。这是第一次，这些小鼠使我们能够研究在体内功能的膜FasL的可溶性FasL的情况下。为了更好地了解不同的FasL亚型在脉络膜新生血管形成中的作用，我们使用了激光诱导的CNV小鼠模型。我们的初步数据表明，在<$CS小鼠中，mFasL表达的增加（或sFasL的丢失）防止了激光诱导CNV后的血管渗漏。光谱域光学相干断层扫描（SD-OCT）和Isolectin-B染色显示，CS小鼠和野生型小鼠之间的新生血管病变大小没有差异。然而，荧光素血管造影显示与野生型小鼠相比，CS小鼠的血管渗漏显著减少。这些结果表明，膜FasL不阻止脉络膜血管的向内生长，如先前所记载的，而是在防止血管渗漏中起关键作用，血管渗漏是湿性AMD患者视力丧失的主要原因。我们推测，膜FasL抑制激光诱导的CNV的血管渗漏，通过Fas介导的途径在浸润的巨噬细胞和/或RPE细胞触发。本提案的目标如下：（目的1）确定CS小鼠中血管渗漏减少是否是由于mFasL增加和/或sFasL缺乏所致，以及渗漏减少是否与浸润性巨噬细胞的M1/M2表型转变一致（目的2）确定浸润性巨噬细胞和/或RPE是否是mFasL介导血管渗漏抑制的关键Fas+靶点。 公共卫生相关性：年龄相关性黄斑变性（AMD）是60岁及以上患者失明的主要原因。异常血管生长到视网膜中是严重视力丧失的主要原因，部分原因是血管渗漏血液， 导致周围视网膜组织损伤的液体。我们已经确定了一种新的蛋白质，防止血管渗漏，本研究的目的是（i）确定是否可以使用这种蛋白质的管理，以防止血管渗漏的小鼠模型，和（ii）阐明这种蛋白质的功能机制。