The Integrin Ligand MFG-E8 and Photoreceptor Outer Segment Renewal.

整合素配体 MFG-E8 和光感受器外段更新。

• 批准号: 7229911
• 负责人: SILVIA C FINNEMANN
• 金额: $ 17.57万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-01-01 至 2008-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7229911
• 关键词: Affinity; Age; Age related macular degeneration; Apoptotic; Appearance; Binding; Binding Sites; Biochemistry; Biological Assay; Cells; Characteristics; Circadian Rhythms; Daily; Data; Detection; Development; Failure; Focal Adhesion Kinase 1; Functional disorder; Genetic Transcription; Homeostasis; Human; In Vitro; Inclusion Bodies; Integrins; Knockout Mice; Ligands; Lipofuscin; MFG; Mediating; Microscopy; Molecular; Molecular Biology; Morphology; Mus; Mutant Strains Mice; Phagocytosis; Phosphatidylserines; Photoreceptors; Process; Protein Tyrosine Kinase; Proteins; Reagent; Recombinants; Recruitment Activity; Retina; Retinal; Retinal Dystrophy; Rod Outer Segments; Rodent; Signal Pathway; Signal Transduction; Structure of retinal pigment epithelium; Surface; Testing; Time; Tissues; Vision; adhesion receptor; age related; aged; extracellular; in vivo; integrin beta5; intracellular protein transport; mouse model; novel; protein expression; protein localization location; receptor; research study; retinal rods

DESCRIPTION (provided by applicant): Shedding of aged photoreceptor outer segment tips (OS) and their phagocytosis by the retinal pigment epithelium (RPE) take place daily in the retina. Failure of RPE cells to phagocytose OS causes retinal dystrophy in rodents and humans. Furthermore, delayed OS degradation may contribute to development or progression of age-related macular degeneration. It is thus important to understand the molecular mechanisms of outer segment renewal. We have identified a signaling pathway in intact retina that is required to clear shed OS promptly. Lack of synchronized phagocytosis in mice deficient in alpha-v-beta5 integrin receptors causes age-related photoreceptor dysfunction and lipofuscin accumulation. Integrin signaling requires ligand engagement of integrin receptors. We thus hypothesize that timely appearance or recruitment of integrin ligand/s serves to stimulate rhythmic signaling in the retina. No such integrin ligand has been identified to date. Our preliminary data suggest the secreted integrin ligand protein MFG-E8 as candidate for this function. MFG-E8 mediates integrin-dependent phagocytosis of apoptotic cells, a process similar to OS phagocytosis. MFG-E8 is expressed by the RPE and participates in RPE phagocytosis in vitro. This exploratory project will achieve two specific objectives: Aim 1 will explore visual function, retinal morphology, in vivo and in vitro RPE phagocytosis and phagocytic signaling in a new mouse model that lacks MFG-E8. These experiments will determine if MFG-E8 functions as a necessary ligand for av|35 integrin in the retina. Aim 2 will investigate if MFG-E8 activity in the sub-retinal space correlates with avB5 ihtegrin's activation, daily OS shedding and phagocytosis. It will further explore if photoreceptor tips destined for shedding expose phosphatidylserine to increase MFG-E8 binding sites. We will combine molecular biology, microscopy, and biochemistry to detect MFG-E8 transcription, protein expression, and protein localization in mouse retina in relation to the time course of outer segment renewal. We will develop novel affinity assays to quantify phosphatidylserine exposure and MFG-E8 binding at the time of OS shedding in intact retina. The results of this exploratory project are an important prerequisite for comprehensive studies of changes in photoreceptor tip and secreted proteins that promote the timely outer segment renewal that is essential for long-term retinal function.

描述（由申请人提供）：视网膜中每天都会发生老化的光感受器外节尖端（OS）的脱落及其被视网膜色素上皮（RPE）的吞噬作用。 RPE 细胞无法吞噬 OS 会导致啮齿动物和人类视网膜营养不良。此外，延迟的操作系统退化可能会导致年龄相关性黄斑变性的发生或进展。因此，了解外节更新的分子机制非常重要。我们已经在完整的视网膜中发现了及时清除脱落的操作系统所需的信号通路。缺乏α-V-β5整合素受体的小鼠缺乏同步吞噬作用会导致与年龄相关的光感受器功能障碍和脂褐素积累。整合素信号传导需要整合素受体的配体参与。因此，我们假设整合素配体的及时出现或募集有助于刺激视网膜中的节律信号传导。迄今为止尚未鉴定出这样的整联蛋白配体。我们的初步数据表明分泌性整联蛋白配体蛋白 MFG-E8 是该功能的候选蛋白。 MFG-E8 介导凋亡细胞的整合素依赖性吞噬作用，该过程类似于 OS 吞噬作用。 MFG-E8由RPE表达并参与体外RPE吞噬作用。该探索性项目将实现两个具体目标：目标 1 将在缺乏 MFG-E8 的新小鼠模型中探索视觉功能、视网膜形态、体内和体外 RPE 吞噬作用以及吞噬信号传导。这些实验将确定 MFG-E8 是否作为视网膜中 av|35 整合素的必要配体发挥作用。目标 2 将研究视网膜下间隙中的 MFG-E8 活性是否与 avB5 ihtegrin 的激活、每日 OS 脱落和吞噬作用相关。它将进一步探索注定要脱落的光感受器尖端是否会暴露磷脂酰丝氨酸以增加 MFG-E8 结合位点。我们将结合分子生物学、显微镜学和生物化学来检测小鼠视网膜中 MFG-E8 转录、蛋白质表达和蛋白质定位与外节更新时间过程的关系。我们将开发新的亲和力测定法，以量化完整视网膜中 OS 脱落时的磷脂酰丝氨酸暴露和 MFG-E8 结合。这个探索性项目的结果是全面研究光感受器尖端和分泌蛋白变化的重要先决条件，这些变化促进了视网膜外节的及时更新，这对于长期视网膜功能至关重要。