Control of Trabecular Meshwork Cytoskeleton

这项资助的长期目标是确定如何调制αvβ3整合素信号 为了开发治疗靶点以控制青光眼的眼压(IOP)。这个 青光眼会导致视网膜神经节细胞不可逆转的丧失，影响大约6700万人 全世界。它们通常与由于降低眼压而引起的眼压升高有关。 房水从小梁网流出(TM)。已经出现的主要风险因素之一 作为一种重要的流出调节机制，细胞骨架是肌动蛋白。它控制多个按键 维持正常流出功能所涉及的生物过程，包括收缩、吞噬和 细胞外基质沉积。整合素在调节这些细胞骨架介导的过程中起着核心作用 活性和我们的研究表明，αvβ3整合素的失调导致了主要的表型变化。 与青光眼相关的包括吞噬功能减少，细胞外基质沉积增加，家族 形成和眼压升高。我们认为，在青光眼中，这种整合素是通过升高的 转化生长因子β2或在糖皮质激素治疗后，如地塞米松。 在这笔赠款中，我们计划使用RNASEQ研究来确定地塞米松或转化生长因子β2上调的因子 激活αvβ3整合素和邻近连接试验，以确定这些因素是否与 整合素很好用。我们还计划证明NRON/NFAT复合体控制DEX诱导的 αvβ3整合素激活和继发性糖皮质激素反应参与激素性青光眼 最后，我们计划展示对DEX(ECM形成、眼压升高和流出设施)的响应是 在体内受αvβ3整合素激活状态的影响，而不仅仅是其表达水平。为了检验这最后一个假设 我们计划使用他莫昔芬可诱导的Creercag-β3整合素/FLOX小鼠来击倒αvβ3整合素 在小鼠TM中的表达。表达αvβ3整合素3种不同激活状态的腺病毒(Ad5)载体 (野生型、非活性和结构性活性)将被用来改变体内αvβ3整合素的活性水平。 拟议中的研究首次证明了特定整合素信号的变化 体内途径可影响眼压、流出通道和细胞外基质的形成。他们将增进我们的理解 整合素信号事件在TM中是如何控制的，以及这如何影响细胞骨架事件(ECM 沉积、细胞收缩和吞噬)调节流出设施。了解整合素是如何 有助于调节这些过程是重要的，因为它将使我们能够提供新的治疗方法 靶向调节细胞骨架，恢复内环境平衡，降低眼压。

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.