Regulation of Lysosomal pH in RPE cells

RPE 细胞中溶酶体 pH 值的调节

• 批准号: 8048058
• 负责人: CLAIRE H MITCHELL
• 金额: $ 33.68万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-02-01 至 2013-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8048058
• 关键词: ATP phosphohydrolase; Acidity; Acids; Age; Agonist; Alkalinization; Biological Assay; Cells; Cyclic AMP; Cystic Fibrosis Transmembrane Conductance Regulator; Data; Defect; Development; Ensure; Environment; Enzymes; Functional disorder; Gases; Genes; Health; In Vitro; Link; Lysosomes; Macular degeneration; Manuscripts; Mus; Mutant Strains Mice; Mutate; Mutation; Organelles; Patients; Photoreceptors; Process; Proteins; Protocols documentation; Proton Pump; Recovery of Function; Regulation; Reporting; Research; Rodent Model; Stargardt' s disease; Structure of retinal pigment epithelium; Therapeutic; Translating; Wild Type Mouse; Work; disease characteristic; early onset; enzyme activity; high throughput screening; improved; in vivo; mouse model; mutant; novel strategies; receptor coupling; repaired; restoration; treatment strategy

DESCRIPTION (provided by applicant): Lysosomal enzymes require an acidic pH to degrade engulfed material. We show here that the lysosomal pH of RPE cells from ABCA4-/- mice is elevated as compared to age matched controls. As the ABCA4 gene is mutated in Stargardt's disease, and as accumulation of partially degraded material in and around RPE cells is a key characteristic of the disease, we hypothesize that restoring an acid pH to the lysosomes will be beneficial. High-throughput screening indicates that lysosomes of perturbed RPE cells in culture are acidified by cytoplasmic cAMP, and by agonists which stimulate receptors coupled to the Gas protein. Elevation of cAMP increases the clearance of outer segments by RPE cells, demonstrating an improvement in function. This proposal investigates the contribution of cAMP, the Cl- channel CFTR, and the vesicular proton pump vH+ATPase to the restoration of lysosomal acidity. It will determine the relationship between age, A2E and lysosomal pH in RPE cells from ABCA4-/- mice and confirm that cAMP restores lysosomal acidity the these RPE cells using both in vitro and in vivo treatment strategies. PUBLIC HEALTH RELEVANCE: Stargardt's disease is an early-onset form of macular degeneration characterized by the accumulation of partially degraded material in and around RPE cells. This proposal will explore a newly identified defect in the lysosomal pH of RPE cells from the ABCA4-/- mouse model of Stargardt's disease that may slow processing of photoreceptor outer segments, develop treatments to restore this defect, and determine how these treatments work.

描述（由申请人提供）：溶酶体酶需要酸性pH值来降解被吞噬的物质。我们在此表明​​，与年龄匹配的对照组相比，ABCA4-/- 小鼠的 RPE 细胞的溶酶体 pH 值升高。由于ABCA4基因在Stargardt病中发生突变，并且RPE细胞内和周围部分降解物质的积累是该疾病的关键特征，因此我们假设恢复溶酶体的酸性pH值将是有益的。高通量筛选表明，培养中受干扰的 RPE 细胞的溶酶体被细胞质 cAMP 和刺激与 Gas 蛋白偶联的受体的激动剂酸化。 cAMP 的升高增加了 RPE 细胞对外部节段的清除，表明功能得到改善。该提案研究了 cAMP、Cl-通道 CFTR 和囊泡质子泵 vH+ATPase 对溶酶体酸度恢复的贡献。它将确定 ABCA4-/- 小鼠 RPE 细胞中年龄、A2E 和溶酶体 pH 之间的关系，并使用体外和体内治疗策略确认 cAMP 恢复这些 RPE 细胞的溶酶体酸度。公众健康相关性：斯塔加特氏病是一种早发性黄斑变性，其特征是部分降解物质在 RPE 细胞内和周围积聚。该提案将探索 Stargardt 病 ABCA4-/- 小鼠模型 RPE 细胞溶酶体 pH 值中新发现的缺陷，该缺陷可能会减慢光感受器外节的处理，开发治疗方法来恢复这种缺陷，并确定这些治疗方法如何发挥作用。