DIFFERENTIATION OF TRANSPORT MECHANISMS IN LENS CELLS

晶状体细胞中传输机制的差异

• 批准号: 2711067
• 负责人: Nicholas A Delamere
• 金额: $ 24.25万
• 依托单位: UNIVERSITY OF LOUISVILLE
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-01-01 至 2002-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2711067
• 关键词: RNase protection assay; SDS polyacrylamide gel electrophoresis; cell differentiation; electrolyte balance; enzyme activity; enzyme biosynthesis; epithelium; fiber cell; fibroblast growth factor; fluorescent dye /probe; ion transport; isozymes; laboratory rabbit; laboratory rat; lens; membrane permeability; microelectrodes; oxidative phosphorylation; prostaglandin E; protein biosynthesis; sodium potassium exchanging ATPase; western blottings

The project emphasizes the importance of the transport function of the antiporter Na, K-ATPase in maintaining ion homeostasis (Na+ and K+) in the whole lens. The fibers have Na,K-ATPase but the enzyme appears to be inactive save for those newly developed cortical fibers. AIM 1: determine if the lens epithelial cells synthesize new Na,K-ATPase by following labeled methionine incorporation into the transporter, determine if inhibition of protein synthesis or actinomycin D blocks the purported turnover. AIM 2 Determine if the epithelium D blocks the purported turnover. AIM 2 Determine if the epithelium selectively upregulates the Na,K-ATPase alpha 2 isoform thus increasing the pump activity. The up-regulation is though to be a physiological response to compromised permeability found in older lenses. Aim 3: Determine if depolarization triggers increased Na-K- ATPase a2 expression. Determine if depolarization triggers increased Na,K-ATPase a2 expression. Determine if the a2 isoform is insensitive to pump inhibition by increased [Ca]. The PI feels that this may be explained by the lack of a tyrosine phosphorylation site on the a2 isoform. Determine the extent to which the a2 isoform increases with age in human lenses - based on the view tht ionic changes accompany aging in the lens. Determine if isoform change in fibers occurs whenrat lens epithelial cells are induced to differentiate following exposure to fibroblast growth factor. Since fibers have low Na,K- ATPase activity, investigate what inactivtes Na,K-ATPase in the fibers with specific emphasis on possible tyrosine phosphorylation of Na,K- ATPase a1 protein. Determine if slow turnover of Na,K-ATPase in the fibers makes the enzyme susceptible tooxidative modification.

该项目强调了运输功能的重要性 维持离子动态平衡的逆向转运蛋白Na，K-ATPase 和K+)。这些纤维含有Na，K-ATPase，但 酶似乎是不活跃的，除了那些新形成的皮质 纤维。目的1：确定晶状体上皮细胞是否合成新的 Na，K-ATPase通过标记蛋氨酸掺入 转运蛋白，确定是抑制蛋白质合成还是放线菌素 D阻止了所谓的营业额。目的2确定上皮细胞是否 D阻止了所谓的营业额。目的2确定上皮细胞是否 选择性地上调Na，K-ATPaseα2亚型，从而 增加泵的活跃度。上调监管被认为是一种 老年人对通透性受损的生理反应 隐形眼镜。目的3：确定去极化是否会引发钠钾升高 ATPase a2的表达。确定去极化触发是否增加 Na，K-ATPase a2的表达。确定a2亚型是否不敏感 通过增加[Ca]来泵抑制。公安局认为这可能是 由a2上缺乏酪氨酸磷酸化位点解释 异构体。测定a2异构体增加的程度 人类晶状体的年龄--基于伴随离子变化的观点 在晶状体中老化。确定纤维中是否发生异构体变化 大鼠晶状体上皮细胞诱导分化的时间 暴露于成纤维细胞生长因子。由于纤维的钠、钾含量较低， ATPase活性，研究是什么使纤维中的Na，K-ATPase失活 特别强调Na，K-可能的酪氨酸磷酸化。 ATPase A1蛋白。确定Na，K-ATPase在体内的缓慢周转 纤维使这种酶对氧化修饰很敏感。