Control of Trabecular Meshwork Cytoskeleton

小梁网细胞骨架的控制

• 批准号: 10557068
• 负责人: Donna M Peters
• 金额: $ 41.37万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-09-15 至 2024-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10557068
• 关键词: Actins; Adenovirus Vector; Affect; African American population; Aqueous Humor; Binding Proteins; Biological Assay; Biological Process; Blindness; Cannulations; Cell Line; Cells; Cilengitide; Complex; Cyclosporine; Cytoskeleton; Deposition; Dexamethasone; Event; Extracellular Matrix; Family; Glaucoma; Glucocorticoids; Goals; Grant; Guanosine Triphosphate Phosphohydrolases; Homeostasis; Human; ITGB3 gene; Immunofluorescence Microscopy; Immunoprecipitation; Injections; Integrin Binding; Integrin Signaling Pathway; Integrin alphaVbeta3; Integrins; Ligation; LoxP-flanked allele; Measures; Mediating; Microscopy; Monitor; Mus; NFAT Pathway; Ocular Hypertension; Optic Nerve; Persons; Phagocytosis; Phagocytosis Inhibition; Phenotype; Physiologic Intraocular Pressure; Play; Process; Production; Recombinant Proteins; Regulation; Retinal Ganglion Cells; Risk Factors; Role; Signal Pathway; Signal Transduction; Sirolimus; Small Interfering RNA; Steroids; Talin; Tamoxifen; Testing; Tissues; Trabecular meshwork structure; Transforming Growth Factor beta; Transgenes; Up-Regulation; Western Blotting; anterior chamber; caveolin 1; connective tissue growth factor; crosslink; in vivo; inhibitor; intravitreal injection; knock-down; nerve damage; new therapeutic target; nuclear factors of activated T-cells; optic nerve disorder; prevent; protein expression; response; therapeutic target; transcription factor; transcriptome; transcriptome sequencing

The long-term objective of this grant is to identify how to modulate αvβ3 integrin signaling pathways in order to develop therapeutic targets to control intraocular pressure (IOP) in glaucoma. The glaucomas, which lead to irreversible loss of retinal ganglion cells, affect approximately 67 million people worldwide. They are commonly associated with elevated levels of intraocular pressure (IOP) due to a reduction in aqueous humor outflow from the trabecular meshwork (TM). One of the major risk factors that has emerged as an important regulatory mechanism for outflow facility is the actin cytoskeleton. It controls a number of key biological processes involved in maintaining normal outflow facility including contractility, phagocytosis, and deposition of the extracellular matrix. Integrins play a central role in regulating these cytoskeleton-mediated activities and our studies suggest that dysregulation of the αvβ3 integrin causes the major phenotypic changes associated with glaucoma including decreased phagocytosis, increased extracellular matrix deposition, CLAN formation and an elevation in IOP. We propose that this integrin is activated in glaucoma by elevated levels of TGFβ2 or following treatments with glucocorticoids, like dexamethasone (DEX). In this grant, we plan to use RNAseq studies to identify the factors upregulated by DEX or TGFβ2 that activate αvβ3 integrin and proximity ligation assays to determine if these factors are associated with the integrin adhesome. We also plan to demonstrate that an NRON/NFAT complex controls DEX-induced activation of αvβ3 integrin and the secondary glucocorticoid response involved in steroid-induced glaucoma. Finally, we plan to show that the responses to DEX (ECM formation, IOP elevation, and outflow facility) are affected by the activated state of αvβ3 integrin in vivo, not just its expression level. To test this last hypothesis we plan to use a tamoxifen inducible CreERcag-β3 integrinflox/flox mouse to knock down αvβ3 integrin expression in the mouse TM. Adenoviral (Ad5) vectors expressing 3 different activated states of αvβ3 integrin (wildtype, inactive and constitutively active) will be used to alter the activity levels of αvβ3 integrins in vivo. The proposed studies are the first to demonstrate that changes in a specific integrin signaling pathway can affect IOP, outflow facility, and ECM formation in vivo. They will enhance our understanding of how integrin signaling events are controlled in the TM and how this affects the cytoskeletal events (ECM deposition, cell contractility and phagocytosis) that regulate outflow facility. Understanding how integrins contribute to the regulation of these processes is important because it will enable us to provide new therapeutic targets to regulate the cytoskeleton in order to restore homeostasis and decrease IOP.

该赠款的长期目标是确定如何调节αVβ3整合素信号传导 途径以开发治疗靶标，以控制青光眼中的眼内压（IOP）。这 绿木瘤导致视网膜神经节细胞的不可逆转损失，影响约6700万人 全世界。由于减少，它们通常与眼内压（IOP）升高有关 在小梁网（TM）的水性幽默出口中。出现的主要风险因素之一 作为出口设施的重要调节机制是肌动蛋白细胞骨架。它控制了许多密钥 维持正常出口设施涉及的生物过程，包括收缩性，吞噬作用和 细胞外基质的沉积。整合素在调查这些细胞骨架介导的核心作用中起着核心作用 活动和我们的研究表明，αVβ3整合素的失调会导致主要的表型变化 与青光眼有关，包括减少吞噬作用，细胞外基质沉积增加，氏族 IOP中的形成和高程。我们建议该整联蛋白在青光眼中被升高的水平激活 TGFβ2或使用糖皮质激素的处理，例如地塞米松（DEX）。 在这笔赠款中，我们计划使用RNASEQ研究来确定DEX或TGFβ2更新的因素 激活αVβ3整联蛋白和接近连接测定法，以确定这些因素是否与 整合素a ad夫。我们还计划证明NRON/NFAT复合物控制DEX诱导的 αVβ3整联蛋白的激活和涉及类固醇诱导的青光眼的二级糖皮质激素反应。 最后，我们计划表明对DEX的反应（ECM组，IOP高程和出口设施）是 受体内αVβ3整联蛋白的活化状态的影响，而不仅仅是其表达水平。检验最后一个假设 我们计划使用他莫昔芬诱导的creercag-β3整合素/flox小鼠击倒αVβ3整合素 在小鼠TM中的表达。表达3种不同激活状态的αVβ3整合素的腺病毒（AD5）载体 （WildType，非活性和组成性活性）将用于改变体内αVβ3整合素的活性水平。 拟议的研究是第一个证明特定整联蛋白信号传导的变化的研究 途径可以影响体内的IOP，出口设施和ECM形成。他们将增强我们的理解 关于在TM中如何控制整联蛋白信号事件以及这如何影响细胞骨架事件（ECM） 调节出口设施的沉积，细胞收缩性和吞噬作用）。了解整联蛋白 为这些过程的调节做出贡献很重要，因为它将使我们能够提供新的疗法 靶标调节细胞骨架以恢复体内稳态和减少IOP。