RETINAL ATTACHMENT AND OUTER SEGMENT TURNOVER IN VITRO

体外视网膜附着和外节翻转

• 批准号: 3263879
• 负责人: Dennis Michael Defoe
• 金额: $ 9.5万
• 依托单位: AUGUSTA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-09-30 至 1994-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3263879
• 关键词: Rana; Xenopus; active transport; antiserum; biological fluid transport; cell adhesion; cell cell interaction; cellular polarity; chlorine; cyclic AMP; glutamates; histology; intraocular fluid; ion transport; laboratory rabbit; ligands; organ culture; phagocytosis; receptor binding; retina; retina detachment; retinal pigment epithelium; rod cell; tissue /cell culture; visual photoreceptor; western blottings

The studies outlined in this proposal focus upon mechanisms which may be responsible for maintaining the attachment of visual cells to the retinal pigment epithelium (RPE). Two independent procedures will be utilized to assess the relative strength of adhesive interactions under a wide range of physiological conditions. First, methods for experimentally detaching Xenopus laevis retinas and reconstituting their interaction with monolayer cultures of RPE will be used to measure spontaneous and L-glutamate-induced tissue reassociation in vitro. Second, a peeling technique, which assays the tendency of apical RPE cytoplasts to partition with mechanically detached retinas, will be utilized to distinguish normal retinal adhesion from the enhanced adhesiveness apparent during active turnover of outer segment membranes. Overall, experiments have been designed to test the involvement of two general mechanisms which have been proposed as determinants of retinal attachment: (1) force generated by active ion- coupled H2O transport by the RPE and (2) formation of non-covalent receptor-ligand bonds between interacting cell surfaces. The possibility that transport-induced transretinal fluid flux influences photoreceptor- epithelium interdigitation will be examined by correlating electrical and ion transport properties of RPE monolayers with their ability to support retinal reapposition. In addition, we will attempt to modulate cell interaction using pharmacological manipulations which previously have been established to alter epithelial fluid transport. Alternatively, involvement of receptor-ligand interactions will be evaluated using highly sensitive and selective immunochemical reagents. Heteroantisera directed against epithelial and photoreceptor surfaces will be generated in rabbits and, after appropriate selection procedures, used as probes of retinal reattachment and uptake of shed outer segment discs. These immunoglobulin molecules will in turn be used as tools for possible biochemical isolation of the cell constituents involved. It is hoped that information gained through this research will lead to better strategies for managing retinal detachment and encouraging photoreceptor-RPE attachment.

本提案中概述的研究重点关注可能是 负责维持视觉细胞与视网膜的附着 色素上皮（RPE）。 将利用两个独立的程序来 评估在广泛的范围内粘合剂相互作用的相对强度 生理条件。 一、实验分离方法 非洲爪蟾视网膜及其与单层细胞的相互作用的重建 RPE 培养物将用于测量自发性和 L-谷氨酸诱导的 体外组织重组。 二、剥离技术，测定 顶端 RPE 细胞质通过机械方式分配的趋势 视网膜脱离，将用于区分正常视网膜粘连 从外层主动周转过程中明显增强的粘合性 段膜。 总的来说，实验的目的是测试 涉及两个一般机制，已提议为 视网膜附着的决定因素：（1）活性离子产生的力 RPE 耦合 H2O 传输和 (2) 非共价键的形成 相互作用的细胞表面之间的受体-配体键。 可能性 运输引起的跨视网膜液体通量影响光感受器 将通过关联电和 RPE 单层的离子传输特性及其支持能力 视网膜再附着。 此外，我们将尝试调节细胞 使用以前已被证实的药理学操作进行相互作用 旨在改变上皮液体运输。 或者， 受体-配体相互作用的参与将使用高度评估 敏感和选择性的免疫化学试剂。 异源抗血清定向 兔子体内会产生针对上皮和光感受器表面的 并且，经过适当的选择程序后，用作视网膜的探针 棚外段圆盘的重新附着和吸收。 这些免疫球蛋白 分子将反过来用作可能的生化分离的工具 所涉及的细胞成分。 希望获得的信息 通过这项研究将带来更好的视网膜管理策略 分离并促进光感受器-RPE 附着。