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)：老年性黄斑变性(AMD)是60岁及以上美国人失明的主要原因。AMD有两种形式：干性或萎缩性和湿性或渗出性。然而，以脉络膜新生血管(CNV)为特征的湿性AMD与更严重的视力损失有关，主要原因是异常血管的液体和血液泄漏。抗血管内皮生长因子治疗已成为湿性AMD患者的首选治疗方法。然而，一些患者对抗血管内皮生长因子治疗没有反应，还有一些患者在最初成功后停止反应，这突显了湿性AMD的发生和发展的病理生物学仍不清楚的事实。FasL是维持眼睛免疫赦免的重要蛋白质，被认为可以诱导浸润性炎症细胞的凋亡。事实上，几份报告表明，FasL表达了 视网膜色素上皮细胞通过诱导血管内皮细胞凋亡抑制脉络膜新生血管形成。然而，FasL既以膜结合蛋白的形式存在，又以具有潜在相反功能的可溶性蛋白的形式存在，目前尚不清楚不同形式的FasL如何参与 新能源汽车的发展。我们最近开发了一种独特的敲入小鼠品系，其中FasL金属蛋白酶裂解位点发生突变，以防止膜结合蛋白的裂解。在这些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)。这些小鼠第一次允许我们在没有可溶性FasL的情况下研究膜FasL的体内功能。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小鼠CNV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)可以防止激光诱导的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小鼠和野生型小鼠的新生血管病变大小没有差异。然而，荧光血管造影术显示，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并不像先前报道的那样阻止脉络膜血管的内生，而是在防止血管渗漏方面发挥关键作用，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通过Fas介导的途径抑制激光诱导的CNV血管渗漏，Fas介导的通路由浸润的巨噬细胞和/或RPE细胞触发。本研究的目的如下：(1)确定CS小鼠血管渗漏的减少是否由于mFasL的增加和/或sFasL的缺乏，以及渗漏的减少是否与浸润性巨噬细胞M1/M2表型的改变相一致(目的2)确定浸润性巨噬细胞和/或RPE是否是mFasL介导抑制血管渗漏的关键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。进入视网膜的异常血管的生长是严重视力丧失的主要原因，部分原因是血管渗漏血液和 对周围视网膜组织造成损害的液体。我们已经确定了一种防止血管渗漏的新蛋白，本研究的目标是(I)确定该蛋白的给药是否可以用于在小鼠模型中防止血管渗漏，以及(Ii)阐明该蛋白的作用机制。