LENS PROTEINS & FLUORESCENCE: IN VIVO & IN VITRO STUDY

晶状体蛋白

• 批准号: 3256016
• 负责人: SIDNEY LERMAN
• 金额: $ 19万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1978
• 资助国家: 美国
• 起止时间: 1978-06-01 至 1986-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3256016
• 关键词: adult human (21+); aging; biophysics; cataract; chromatography; eye bank /preservation; fluorescence; fluorescence spectrometry; human tissue; lens proteins; mature animal; photochemistry; radiobiology; spectrometry; ultraviolet radiation; visual pigments

The specific aims for this project are: 1) To isolate and identify some of the molecular species responsible for the photochemically induced age-related fluorescence (particularly in the 360 nm excitation/440 nm emission, and 435 nm excitation/520 nm emission) in the human lens. This will be performed by the isolation and purification of specific fluorescent peptides (enzymatic hydrolysis and chromatography); amino acid composition and sequencing, and further hydrolysis of the peptides to isolate the purified chromophores. Spectroscopic analyses (UV, fluorescence, phosphorescence, GC mass spectrometry and NMR spectroscopy will be performed at certain stages of this procedure to monitor and identify the compounds isolated. 2) To elucidate the molecular mechanisms of photochemical lens damage by studying the photochemistry of purified lens proteins and appropriate model compounds. 3) To probe the distribution of lens protein fractions (alpha, beta, gamma-crystallin and insoluble) using laser excited tryptophan fluorescence decay measurements on intact human lenses ("optical dissection"). Knowing the photochemical reactivities of the various lens protein fractions and their distribution should permit us to predict which region(s) of the intact lens are most susceptible to photochemical damage. 4) In addition to our conventional irradiation source which enables us to bathe whole lenses with photons, we also propose to utilize our laser photolysis apparatus to study localized photodamage in whole lenses and to detect such damage by laser excited fluorescence and ESR spectroscopy. 5) To correlate our in vitro fluorescence studies (on human lenses) with in vivo UV-slit lamp densitography. We have recently demonstrated the feasibility of this approach and we have instituted a study on normal individuals as well as patients on phototherapy. This may enable us to detect changes in lens fluorescence several years prior to their becoming manifest by usual slit lamp examinations. 6) To isolate and identify the photochemical product(s) produced when animals are exposed to UV radiation following a dose of the sensitizer 8-methoxypsoralen. Spectroscopic analyses (as outlined in 1) will be employed. 7) To relate the molecular changes caused by direct and photosensitized UV radiation to morphological events.

该项目的具体目的是：1）隔离并确定一些 负责光化学引起的年龄相关的分子物种 荧光（特别是在360 nm激发/440 nm发射中，435 nm 兴奋/520 nm的发射）。 这将由 特异性荧光肽的分离和纯化（酶促的 水解和色谱）；氨基酸组成和测序，以及 肽进一步水解以分离纯化的发色团。 光谱分析（UV，荧光，磷光，GC质谱法 NMR光谱将在此过程的某些阶段进行 监测并识别分离的化合物。 2）阐明分子 通过研究光化学损伤的机制通过研究光化学 纯化的透镜蛋白和适当的模型化合物。 3）探测 透镜蛋白分数的分布（alpha，beta，伽马晶蛋白和 使用激光激发色氨酸荧光衰减测量的不溶性） 完整的人透镜（“光学解剖”）。 知道光化学 各种晶状体蛋白分数及其分布的反应性应 允许我们预测完整镜头的哪个区域最容易受到影响 光化学损坏。 4）除了我们的常规辐照源外 这使我们能够用光子洗澡，我们还建议使用 我们的激光光解仪研究整个镜片中的局部光损伤 并通过激光激发荧光和ESR光谱检测这种损害。 5）将我们的体外荧光研究（在人镜上）与体内相关联 紫外线灯光密摄影。 我们最近证明了 这种方法，我们已经就普通人以及 接受光疗的患者。 这可能使我们能够检测到镜头的变化 荧光前几年通过通常的缝隙灯体现出来 考试。 6）隔离并识别产生的光化学产品 剂量后，动物暴露于紫外线辐射时 8-甲氧基索拉伦。 光谱分析（如1中所述）将被采用。 7）关联直接和光敏紫外线引起的分子变化 形态事件的辐射。