RETINAL ATTACHMENT AND OUTER SEGMENT TURNOVER IN VITRO

体外视网膜附着和外节翻转

• 批准号: 3263877
• 负责人: Dennis Michael Defoe
• 金额: $ 11.73万
• 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/3263877
• 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.

该提案中概述的研究重点是可能是 负责维持视觉细胞附着在视网膜上 色素上皮（RPE）。 将使用两个独立程序 评估在广泛范围内的粘合剂相互作用的相对强度 生理条件。 首先，实验分离的方法 Xenopus laevis视网膜并重建与单层的互动 RPE的培养物将用于测量自发和L-谷氨酸诱导的 组织在体外重新关联。 其次，一种剥离技术，该技术测定 顶端RPE细胞体的趋势与机械分割 分离的视网膜将用于区分正常的视网膜粘附 从外部主动周转期间明显的粘附性增强 细分膜。 总体而言，已经设计了实验来测试 提出的两种一般机制的参与 视网膜附着的决定因素：（1）活性离子产生的力 通过RPE耦合H2O传输和（2）非共价的形成 相互作用的细胞表面之间的受体配体键。 可能性 转运诱导的转视网膜流体会影响光感受器 - 将通过将电气相关的和 RPE单层的离子运输特性具有支持的能力 视网膜重置。 此外，我们将尝试调节单元格 使用以前曾经是的药理操作的相互作用 建立以改变上皮流体的运输。 或者， 将使用高度评估受体配体相互作用的参与 敏感和选择性免疫化学试剂。 异性恋指导 在兔子中将产生针对上皮和光感受器的表面 并且，经过适当的选择程序，用作视网膜的探针 棚外部圆盘的重新计算和吸收。 这些免疫球蛋白 分子又将用作可能生化隔离的工具 涉及的细胞成分。 希望获得信息 通过这项研究将为管理视网膜提供更好的策略 分离并鼓励光感受器-RPE附着。