BIOCHEMICAL RESPONSES OF HUMAN CILIARY EPITHELIAL CELLS

人睫状上皮细胞的生化反应

• 批准号: 3259439
• 负责人: MIGUEL COCA-PRADOS
• 金额: $ 13.8万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-07-01 至 1992-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3259439
• 关键词: active transport; adenosinetriphosphatase; adenylate cyclase; beta adrenergic receptor; biochemistry; calcium; cell sorting; chromatography; cyclic AMP; density gradient ultracentrifugation; epithelium; fluorescence; fluorescence microscopy; gel electrophoresis; glaucoma; human fetus tissue; human tissue; hybridomas; immunofluorescence technique; immunologic techniques; inositol phosphates; ion transport; laboratory mouse; laboratory rabbit; membrane proteins; monoclonal antibody; phosphorylation; protein kinase; protein kinase C; radioassay; secretion; sodium potassium exchanging ATPase; sympathetic nervous system; tissue /cell culture; transport proteins; uvea ciliary body

Knowledge of aqueous humor secretion and regulation of aqueous production in the human eye is relevant to the understanding and treatment of glaucoma. Aqueous humor is secreted by the ciliary epithelium and its composition is largely dependent on the mechanism of active transport of plasma proteins an electrolites in these cells. The rate of formation of aqueous humor is modulated by the sympathetic (adrenergic) system (Neufeld, A.H., 1984). The development of a tissue culture system for the ciliary epithelial (CE) cells, namely the nonpigmented (NPE) and pigmented (PE) epithelia is important because it permits the in vitro study of the cellular and molecular functions of these cells which are very difficult to carry out in vivo. The aim of this proposal is to develop antibodies to both membrane-bound surface proteins and internal proteins which are specific markers for NPE and PE cells using the hybridoma technology. These antibodies will serve as probes to investigate the differentiated properties of these cells in culture. In addition, it will allow for the isolation (i.e. fluorescence activated cell sorter) of NPE and PE cells based on their potential immunological differences. These antibodies will be used to facilitate the understanding of the secretory function and transport of NPE and PE cells. Secretory function will be studied by developing antibodies to a 76KDa protein secreted by CE in culture. Ion transport will be studied by immunobloting analysis of (Na+, K+)-ATPase activity by antibodies to the alpha subunit of this enzyme. Sodium pump transport activity will assessed by ouabain-sensitive 86Rb+ uptake. In addition, the mechanism of transduction by external signals into intracellular events (i.e., protein phosphorylation) will be studied.

房水分泌和房水调节的知识 人眼的产生与理解和 青光眼的治疗。 房水由睫状体分泌 上皮细胞及其组成很大程度上取决于 血浆蛋白和电解质的主动运输机制 在这些细胞中。 房水形成速率为 由交感神经（肾上腺素能）系统调节（Neufeld，A.H.， 1984）。 睫状体组织培养系统的开发 上皮细胞（CE），即非色素细胞（NPE）和 色素 (PE) 上皮细胞很重要，因为它允许在 这些细胞的细胞和分子功能的体外研究 这在体内很难进行。 该提案的目的是开发针对这两种疾病的抗体 膜结合表面蛋白和内部蛋白 使用杂交瘤对 NPE 和 PE 细胞进行特异性标记 技术。 这些抗体将作为探针进行研究 这些细胞在培养物中的分化特性。 此外， 它将允许分离（即荧光激活细胞 根据 NPE 和 PE 细胞的潜力进行分选 免疫学差异。 这些抗体将用于促进理解 NPE和PE细胞的分泌功能和运输。 将通过开发抗体来研究分泌功能 培养物中 CE 分泌的 76KDa 蛋白质。 离子传输将 通过 (Na+, K+)-ATPase 活性的免疫印迹分析进行研究 通过该酶的α亚基的抗体。 钠泵 转运活性将通过哇巴因敏感的 86Rb+ 进行评估 吸收。 此外，外部信号的转导机制 进入细胞内事件（即蛋白质磷酸化）将是 研究过。