ANALYSIS OF MICRODISSECTED CATARACTOUS HUMAN LENSES

长期目标是了解原因， 导致人类白内障形成的机制。 的 具体目标是：1.为了继续我们长期的临床生物化学 研究与衰老和药物相关的人类白内障。 将对患者进行全面的术前评估， 白内障患者的白内障进展情况将 临床随访，并将使用裂隙灯记录 程序、偏振后向照明和Scheimpflug 摄影. 在白内障手术后，透镜将 显微解剖和生化评估。 2.研究 新鲜离体人透镜上皮细胞的生化特性 白内障手术后 将特别注意 谷胱甘肽还原酶和其它黄素酶。 由于黄素酶 取决于膳食核黄素，膳食核黄素的状况将 通过监测谷胱甘肽对白内障患者进行检查 还原酶的活性在这些患者的红细胞。 其他 还将研究关键酶。 3.研究五味子甲 甲状腺激素（T3和甲状腺素），β受体阻滞剂（如 普萘洛尔）和类固醇（如泼尼松）对酶活性的影响 在人的透镜上皮中。 牛、兔的透镜器官培养， 老鼠也将被用来研究激素的作用 还有毒品 4.根据已知的肽合成短肽 各种晶体蛋白和其他晶状体特异性 proteins. 这些肽将用于生产抗体 特异于这些肽，然后将用于探测 白内障的人类透镜。 5.研究透镜的稳定性 晶体蛋白。 人透镜晶体蛋白的热稳定性 将使用圆二色性（CD）研究年龄的函数， 吸收和荧光光谱。 6.继续并 研究透镜质膜的结构和功能 固有多肽（HIP-26），并尝试将其结晶， X射线衍射研究。 MIP-26有可能 将探索cGMP的结合位点。 7.阐明本 物理化学性质和免疫定位新的 发现了透镜纤维特异性内膜蛋白MP-17。 这将包括纯化的蛋白质表征， 它的二级和三级结构，使用CD和原位 本地化 8.检查白内障和透明晶状体 遗传性视网膜变性（RCS）大鼠。 白内障 形成是一个次要的现象，这些大鼠是一个很好的 对透镜的氧化损伤的动物模型， 白内障

The long term objectives are to understand the causes and mechanisms which lead to the formation of cataract in humans. The specific aims are: 1. To continue our long term clinicobiochemical investigation of aging-related and drug-related human cataracts. A comprehensive presurgical evaluation of the patient will be performed, and the progression of cataract in the patient will be followed clinically, and will be documented using slit-lamp procedures, polarized retroillumination, and Scheimpflug photography. Following cataract surgery, the lens will be microdissected and evaluated biochemically. 2. To study the biochemical properties of freshly excised human lens epithelium following cataract surgery. Special attention will be given to glutathione reductase and other flavoenzymes. Since flavoenzymes depend on dietary riboflavin, the status of dietary riboflavin will be examined in patients with cataract by monitoring glutathione reductase activity in the red blood cells of these patients. Other key enzymes will also be studied. 3. To study the effects of thyroid hormones (T3 and thyroxine), beta-blockers (such as propranolol) and steroids (such as prednisone) on enzyme activity in human lens epithelium. Lens organ culture of bovine, rabbits, and rats will also be used to study the effects of the hormones and drugs. 4. To synthesize short peptides based on the known sequence of the various crystallins and other lens-specific proteins. These peptides will be used to produce antibodies specific to these peptides, and then will be used to probe cataractous human lens. 5. To study the stability of lens crystallins. The thermal stability of human lens crystallin as a function of age will be studied using circular dichroism (CD), absorption, and fluorescence spectroscopy. 6. To continue and study the structure and function of the lens plasma membrane major intrinsic polypeptide (HIP-26), and to try to crystallize it for X-ray diffraction studies. The possibility that MIP-26 has a binding site for cGMP will be explored. 7. To elucidate the physico-chemic properties and to immunolocalize the newly discovered lens fiber-specific intrinsic membrane protein MP-17. This will include purification of the protein characterization of its secondary and tertiary structure using CD and in situ localization. 8. To examine the cataractous and clear lenses of rats with inherited retinal degeneration (RCS). The cataract formation is a secondary phenomenon, these rats are an excellent animal model for oxidative insults to the lens which may cause cataract.