BLOOD-LENS TRANSFER OF GLUTATHIONE & SULFUR AMINO ACIDS

谷胱甘肽的血晶状体转移

• 批准号: 3266828
• 负责人: Berislav V Zlokovic
• 金额: $ 14.61万
• 依托单位: UNIVERSITY OF SOUTHERN CALIFORNIA
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-08-01 至 1996-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3266828
• 关键词: aminoacid transport; biological transport; blood aqueous barrier; eye circulation; glutathione; guinea pigs; high performance liquid chromatography; lens; perfusion; radiotracer; sulfur aminoacid

Lens transparency is essential for proper visual function. Glutathione (GSH) plays an important role in preventing lens opacification (cataract) by maintaining SH-groups of the lens proteins (membrane components, enzymes and crystallins) in reduced state. The lens is rich in GSH, and the current concept states that low levels of GSH are not consistent with lens clarity, but themselves are cataract inducing. Uptake of GSH by the lens and GSH lens synthesis de novo from sulfur amino acids (SAA) have been demonstrated in vitro, but direct evidence is lacking to conclude that this approximates the situation invivo. A new invivo vascular eye perfusion (VEP) model in the guinea-pig has been developed in our laboratory. The use of this species as an animal model for proposed studies has been justified during the course of our preliminary work. Biochemical characterization of guinea-pig lens suggested that metabolic scheme for GSH homeostasis is similar to that in other mammalian lenses. Using present VEP model, we obtained strong preliminary evidence indicating significant insitu rapid cellular uptake of newly secreted plasma-derived [35S]-GSH by the lens. A rapid in situ GSH lens synthesis from newly secreted blood-borne (35S)-cysteine was shown by radio HPLC analysis. The focus of this proposal is directed at plasma-derived GSH and SAA in the lens. The proposal is designed to test in a comprehensive fashion the hypothesis that blood-to-lens transport of circulating GSH and SAA is essential for the regulation of GSH levels in normal lenses. Towards this end, several experiments are proposed to test the following two hypotheses: I. Plasma-derived GSH is rapidly taken up at the lenticular epithelium by a specific transport system. II. GSH lens synthesis de novo is dependent on blood-to-lens transport of SAA. All transport and metabolic studies in normal guinea-pigs will use the VEP model, and consider for kinetic analysis four eye compartments including plasma, aqueous humor, lens capsule/epithelium and lens/cortex. Molecular forms of uptake during compartmental blood-to-lens transfer will be determined by radio-HPLC. The specificity and kinetic properties of GSH transport system in the lens will be characterized in situ, and GSH de novo synthesis from circulating SAA precursors will be estimated. These studies will help us understand the significance of blood-to-lens transport of GSH and SAA for normal lens function. Defining the role of plasma-derived GSH and SAA in normal lenses may be important in designing therapeutic strategies to decelerate cataract-inducing processes and/or to prevent formation of cataract.

透镜的透明度对正常的视觉功能至关重要。谷胱甘肽 (GSH)在预防透镜混浊（白内障）中起重要作用 通过保持透镜蛋白质（膜组分， 酶和晶体蛋白）。透镜富含GSH， 目前的概念指出，低水平的GSH不符合 透镜清晰，但本身是诱发白内障。谷胱甘肽的摄取 从含硫氨基酸（SAA）从头合成透镜和GSH透镜， 已经在体外证明，但缺乏直接证据来得出结论， 这和活体情况很接近一种新的体内血管眼 在我们的实验室里，我们建立了豚鼠视觉诱发电位（VEP）模型， 实验室使用该物种作为动物模型， 在我们的初步工作过程中，研究是有道理的。 豚鼠透镜的生化特征表明， GSH体内平衡的机制与其他哺乳动物晶状体相似。 利用现有的视觉诱发电位模型，我们获得了强有力的初步证据， 这表明新分泌的蛋白质的显著原位快速细胞摄取 血浆来源的[35 S]-GSH通过透镜。谷胱甘肽透镜的快速原位合成 放射性高效液相色谱法显示新分泌的血源性（35 S）-半胱氨酸 分析.该提案的重点是针对血浆来源的GSH， 透镜中的SAA。 该提案旨在全面测试 形成了这样的假设，即血液向透镜转运循环GSH和 SAA对于调节正常晶状体中的GSH水平是必不可少的。 为此，提出了几个实验来测试以下内容 两个假设：I.血浆来源的GSH在 晶状体上皮细胞通过特定的运输系统。二. GSH透镜 从头合成依赖于SAA的血液至透镜转运。所有 正常豚鼠的转运和代谢研究将使用VEP 模型，并考虑动力学分析的四个眼睛隔间，包括 血浆、房水、透镜囊/上皮和透镜/皮质。分子 隔室血液转移至透镜期间的摄取形式将是 通过放射性HPLC测定。还原型谷胱甘肽的特异性和动力学性质 运输系统中的透镜将在原位进行表征，并GSH de 将估计来自循环SAA前体的从头合成。这些 研究将帮助我们理解血液对透镜的重要性 正常透镜功能的GSH和SAA的转运。定义的作用 正常晶状体中的血浆源性GSH和SAA可能在设计晶状体时是重要的。 减缓白内障诱导过程和/或 防止白内障的形成。