Trabecular Meshwork Cytoskeletal Signaling-Regulation of Aqueous Humor Outflow

小梁网细胞骨架信号传导-房水流出的调节

• 批准号: 7752794
• 负责人: P VASANTHA Rao
• 金额: $ 38.61万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-01-01 至 2011-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7752794
• 关键词: Actomyosin; Agonist; Anterior eyeball segment structure; Aqueous Humor; Cells; Characteristics; Etiology; Event; Extracellular Matrix; Extracellular Matrix Proteins; Eye; Family suidae; Gene Expression; Glaucoma; Homeostasis; Human; Life; Lysophospholipids; Modeling; Molecular; Myosin Type II; Names; Ocular Hypertension; Organ Culture Techniques; Pathway interactions; Phosphorylation; Physiologic Intraocular Pressure; Physiological; Play; Primary Open Angle Glaucoma; Principal Investigator; Public Health; Regulation; Research; Resistance; Resource Sharing; Rho-associated kinase; Rodent Model; Role; Signal Pathway; Signal Transduction; Stream; Thrombin; Trabecular meshwork structure; Transforming Growth Factor beta; aqueous; cytokine; design; effective therapy; in vivo; insight; novel; programs; rho; rho GTP-Binding Proteins

DESCRIPTION (provided by applicant): Identifying potential regulatory mechanisms controlling aqueous humor outflow resistance has important implications for understanding glaucoma and developing effective therapies. We hypothesize that Rho/Rho kinase signaling pathway regulated cytoskeletal tension and cell extracellular matrix interactions of trabecular meshwork (TM) cells play a critical role in the synthesis and turnover of extracellular matrix, and contribute to the homeostasis of aqueous humor outflow resistance. Abnormal activation of Rho/Rho kinase signaling within the outflow pathway is therefore predicted to contribute to the increased aqueous outflow resistance in glaucomatous eyes. Compelling evidence for this hypothesis derives from our recent studies of agonist and RhoA induced changes in TM cell contractile tone and aqueous humor outflow facility in perfused models. Contractile agonists (lysophospholipids and thrombin), TGF beta, extracellular matrix (ECM) proteins and increased intraocular pressure, all stimulated myosin II phosphorylation and actomyosin assembly in TM cells, which are recognized characteristics of increased cellular cytoskeletal tension. These effects were further associated with increased Rho GTPase activation in TM cells, and with significant decreases in aqueous outflow facility in perfused porcine and human eyes. Moreover, expression of constitutively active RhoA significantly decreased aqueous outflow facility in organ cultured porcine eye anterior segments and increased expression of genes encoding various ECM proteins and cytokines in human TM cells. Taken together, these observations indicate the existence of a potential interaction between Rho GTPase signaling, cytoskeletal tension, ECM synthesis and aqueous outflow resistance. Given the convincing evidence in support of a role for Rho GTPase induced cytoskeletal tension and cell ECM interactions in regulation of aqueous humor outflow resistance we propose in this application to investigate: 1. the association between Rho GTPase induced cytoskeletal tension and ECM synthesis and the expression of cytokines in human TM cells; 2. in vivo effects of sustained activation of Rho GTPase on intraocular pressure and ECM synthesis in the outflow pathway in a live rodent model, and 3. the expression and activation status of the Rho/Rho kinase signaling components in the outflow pathway of eyes subjected to elevated intraocular pressure, and in hypertensive open angle glaucomatous human donor eyes. Completion of these studies should unravel the role of TM cytoskeletal tension and cell ECM interactions in the homeostasis of aqueous humor outflow resistance, define the regulation of ECM synthesis by the Rho/Rho kinase signaling pathway, and clarify the significance of these events in increased intraocular pressure in glaucomatous eyes. Relevance to public health: Better understanding of the molecular mechanisms regulating homeostasis of aqueous humor outflow resistance could provide novel avenues for designing targeted therapies for treatment of glaucoma. Understanding the etiology of primary open angle glaucoma and developing effective therapies requires identification of regulatory mechanisms which control aqueous humor outflow. In the proposed study we explore the novel paradigm that there is a strong functional connection between the cytoskeletal signaling and the homeostasis of aqueous humor outflow resistance. Exploring this connection will provide important insights into our understanding of physiological and pathological regulation of aqueous outflow resistance in normal and ocular hypertensive glaucoma eyes and have an important bearing on translational application to novel therapies for glaucoma.

描述（由申请人提供）：识别控制房水流出阻力的潜在调节机制对于了解青光眼和开发有效疗法具有重要意义。我们假设Rho/Rho激酶信号通路调节小梁网（TM）细胞的细胞骨架张力和细胞外基质相互作用在细胞外基质的合成和周转中发挥关键作用，并有助于房水流出阻力的稳态。因此，流出途径中 Rho/Rho 激酶信号传导的异常激活预计会导致青光眼眼中房水流出阻力的增加。这一假设的令人信服的证据来自我们最近对灌注模型中激动剂和 RhoA 诱导 TM 细胞收缩张力和房水流出设施变化的研究。收缩激动剂（溶血磷脂和凝血酶）、TGF β、细胞外基质（ECM）蛋白和眼内压升高，均刺激 TM 细胞中肌球蛋白 II 磷酸化和肌动球蛋白组装，这是公认的细胞骨架张力增加的特征。这些效应进一步与 TM 细胞中 Rho GTP 酶激活的增加有关，并且与灌注猪和人眼中房水流出能力的显着减少有关。此外，组成型活性RhoA的表达显着降低了器官培养的猪眼前节的房水流出能力，并增加了人TM细胞中编码各种ECM蛋白和细胞因子的基因的表达。总而言之，这些观察结果表明 Rho GTPase 信号传导、细胞骨架张力、ECM 合成和房水流出阻力之间存在潜在的相互作用。鉴于支持 Rho GTPase 诱导的细胞骨架张力和细胞 ECM 相互作用在房水流出阻力调节中的作用的令人信服的证据，我们建议在本申请中研究： 1. Rho GTPase 诱导的细胞骨架张力与 ECM 合成和表达之间的关联人类TM细胞中的细胞因子； 2. Rho GTPase 持续激活对活体啮齿动物模型中眼压和流出途径 ECM 合成的体内影响，以及 3. 受试眼睛流出途径中 Rho/Rho 激酶信号传导成分的表达和激活状态眼内压升高，以及高血压开角型青光眼人类供体眼睛。这些研究的完成应阐明 TM 细胞骨架张力和细胞 ECM 相互作用在房水流出阻力稳态中的作用，定义 Rho/Rho 激酶信号通路对 ECM 合成的调节，并阐明这些事件在眼内房水增加中的重要性。青光眼眼内的压力。与公共卫生的相关性：更好地了解调节房水流出阻力稳态的分子机制可以为设计治疗青光眼的靶向疗法提供新途径。 了解原发性开角型青光眼的病因并开发有效的治疗方法需要确定控制房水流出的调节机制。在拟议的研究中，我们探索了一种新的范式，即细胞骨架信号传导与房水流出阻力的稳态之间存在很强的功能联系。探索这种联系将为我们理解正常和高眼压青光眼眼房水流出阻力的生理和病理调节提供重要的见解，并对青光眼新疗法的转化应用产生重要影响。