BIOLOGY OF TRABECULAR MESHWORK IN HEALTH AND DISEASE

小梁网在健康和疾病中的生物学

• 批准号: 2159498
• 负责人: BEATRICE Y.J.T. YUE
• 金额: $ 22.03万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-12-01 至 2000-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2159498
• 关键词: biological signal transduction; cell adhesion; collagen; enzyme linked immunosorbent assay; extracellular matrix proteins; fibronectins; flow cytometry; glaucoma; glucocorticoids; growth factor; human tissue; immunocytochemistry; immunoelectron microscopy; immunofluorescence technique; immunoprecipitation; integrins; intraocular pressure; laminin; messenger RNA; northern blottings; organ culture; protein structure function; tissue /cell culture; trabecular meshwork; western blottings

The trabecular meshwork (TM) is believed to be a major site for the regulation of the aqueous outflow. Residing in this tissue are TM cells that are essential for the maintenance of normal outflow. Aberrations of cell integrity may be a key step toward obstruction of the aqueous outflow, intraocular pressure elevations, and glaucomatous conditions. Along with the cells, extracellular matrix (ECM) materials such as fibronectin, laminin, and collagens are also thought to be important for normal functioning of the TM. We propose, in this application, to examine a family of receptors for ECM proteins, the integrins, and the cell-matrix interactions in the human TM. The integrins are known to interact with cytoskeletal proteins, mediate cell attachment, and influence a variety of cellular activities. They are also crucial in the determination of growth, morphology, metabolism, and differentiation of cells. In the TM system, cells line the trabecular beams that are made up of connective tissues. The TM cells in vivo are continually subjected to flows of the aqueous humor and changes in the intraocular pressure. The lining integrity against stress is critically dependent on the adhesion of TM cells to the matrices through integrin receptors. Disruption in the adhesion of TM cells to the ECM would conceivably lead to cell loss, denudation of the beams, and pathologic consequences. Characterization of the integrins and elucidation of the cell-matrix interactions in the TM are therefore of fundamental significance. We have initiated investigations on integrins and have found that the integrin repertoire in cultured TM cells is similar to that in tissues. We will further our efforts to evaluate the roles of integrins in adhesion of cultured TM cells to the ECM and to determine the integrin-mediated signaling pathways by immunoprecipitation, immunofluorescence staining and other methods. Activators and inhibitors will be used to define further the potential signaling mechanisms involved. We propose to dissect systematically the information transduced by cell-ECM interactions using the in vitro human TM cultures coupled with defined matrices. The effects of individual ECM substrata on the integrin distribution and biologic characteristics of TM cells will be assessed by cell biology and molecular biology techniques. To probe the functional roles of integrins, human TM cells will be transfected to result in either enhanced or suppressed expression of specific integrins. The effects of gene alterations will be analyzed. In addition, the integrin distribution and expression in conditions that are known to induce ECM changes in TM cells, such as after phagocytic challenges and after glucocorticoid treatments, will be examined. Modulation in the integrins will be correlated with the ECM changes. The effects of growth factors on the production and expression of fibronectin, laminin, collagen, and integrins by TM cells in both cell culture and organ culture systems will also be investigated. Protocols of immunohistochemical staining, and dot blot, Western blot and Northern blot analyses will be followed. Our goal is to increase the understanding of the biologic characteristics of healthy as well as experimentally-altered TM cells, particularly with regard to the dynamics and control mechanisms involved in the cell-matrix interactions in the TM. This information will assist in uncovering the pathogenic processes of glaucomas.

据信小梁网（TM）是 水流出的调节。居住在该组织中的是TM细胞 这对于维持正常流出至关重要。畸变 细胞完整性可能是障碍物阻塞的关键步骤 流出，眼内压升高和青光眼条件。 与细胞一起，细胞外基质（ECM）材料（例如 纤连蛋白，层粘连蛋白和胶原蛋白也被认为对 TM的正常功能。我们建议在此应用中检查 ECM蛋白质，整合素和细胞矩阵的受体家族 人类TM的相互作用。已知整联蛋白与 细胞骨架蛋白，介导细胞附着并影响多种 细胞活动。它们对于确定增长至关重要， 细胞的形态，代谢和分化。在TM系统中， 细胞为由结缔组织组成的小梁梁排列。 体内的TM细胞不断受到水的流量 眼内压的幽默和变化。衬里完整性 反对压力取决于TM细胞对 通过整联蛋白受体进行矩阵。 TM粘附的破坏 到ECM的细胞可能会导致细胞丢失，剥夺 梁和病理后果。整合素和的表征 因此，阐明TM中的细胞基质相互作用是 基本意义。 我们已经开始对整合素的调查，发现 培养的TM细胞中整合素库与组织中的曲目相似。我们 将进一步努力评估整合素在粘附中的作用 培养的TM细胞到ECM并确定整联蛋白介导的 免疫沉淀，免疫荧光染色和信号通路 其他方法。激活剂和抑制剂将用于进一步定义 潜在的信号传导机制。我们建议剖析 系统地，使用细胞ECM相互作用转导的信息 体外人类TM培养物与定义的矩阵结合。效果 在整联蛋白分布和生物学上的单个ECM基质 TM细胞的特征将通过细胞生物学和分子评估 生物学技术。为了探测整联蛋白的功能作用，人类TM 细胞将被转染以导致增强或抑制 特定整合素的表达。基因改变的影响将是 分析。另外，整联蛋白的分布和表达 已知会在TM细胞中诱导ECM变化的疾病，例如 吞噬挑战和糖皮质激素治疗后，将是 检查。整联蛋白中的调节将与ECM相关 更改。生长因子对生产和表达的影响 两个细胞中TM细胞的纤连蛋白，层粘连蛋白，胶原蛋白和整联蛋白 文化和器官培养系统也将进行研究。协议的 免疫组织化学染色，以及点印迹，蛋白质印迹和北印迹 将遵循分析。 我们的目标是提高对生物学特征的理解 健康和实验改变的TM细胞，特别是 关于细胞矩阵涉及的动力和控制机制 TM中的相互作用。此信息将有助于发现 青光眼的致病过程。