ANALYSIS OF MICRODISSECTED CATARACTOUS HUMAN LENSES

本研究项目的长期目标是了解 导致人类白内障形成的原因和机制。 具体目标是：（1）开展长期的临床生化工作 研究与衰老和药物相关的人类白内障。 在这些 研究，将对患者进行全面的术前评估， 执行。 将随访患者白内障的进展情况 临床上，并将使用标准化裂隙灯程序记录， 偏振反光照明摄影和沙伊姆弗勒摄影。 白内障手术后，将对透镜进行显微解剖和评估 如目标2和目标3所述。 (2)显微解剖单个 人类白内障晶状体，并借助显微镜分离 不透明区域和相邻的正常区域。 高效液相 将使用高效液相色谱法（HPLC）来研究水的性质 可溶性蛋白、尿素可溶性蛋白以及两者中的膜蛋白 透明和不透明的部分。 其他技术包括 聚丙烯酰胺凝胶电泳，等电聚焦，氨基酸 放射性碘标记蛋白质组分的胰蛋白酶肽图谱分析 利用电泳和色谱的二维系统 然后进行放射自显影。 (3)分析主要非蛋白质低 人体透明和不透明切片中的分子量成分 眼镜. (4)开发回收透镜材料的新方法， 囊外手术 采取的方法将是轻敲抽吸管线 手术中使用的设备。 (5)识别和描述 23，000道尔顿晶体蛋白，其在许多情况下被选择性降解和修饰， 人白内障晶状体，并确定和研究蛋白水解酶 在某种程度上，参与了这个过程。 (6)完成 透镜质膜主要结构cDNA克隆的核苷酸序列测定 内源性蛋白（MIP-26）。 (7)为了完成 牛β 23/26晶体蛋白。 (8)为了研究饮食和衰老对 在小鼠中，γ-晶状体蛋白的差异性损失被降低， 卡路里饮食。

The long term objectives of this research project are to understand the causes and mechanisms which lead to the formation of cataract in humans. The specific aims are: (1) To start a long term clinicobiochemical investigation of aging-related and drug-related human cataracts. In these studies, a comprehensive presurgical evaluation of the patient will be performed. The progression of cataract in the patient will be followed clinically, and will be documented using standardized slit lamp procedures, polarized retroillumination photography, and Scheimpflug photography. After cataract surgery, the lens will be microdissected and evaluated biochemically as described under aims 2 and 3. (2) To microdissect single human cataractous lenses and to separate with the aid of a microscope opague areas and adjacent normal areas. High performance liquid chromatography (HPLC) will be used to study the properties of the water soluble proteins, urea soluble proteins, and the membrane proteins in both the clear and opaque sections. Other techniques will include SDS-polyacrylamide gel electrophoresis, isoelectric focusing, amino acid analysis, tryptic peptide mapping of radioiodinated protein fractions employing the two-dimensional system of electrophoresis and chromatography followed by autoradiography. (3) To analyze the major nonprotein low molecular weight constituents in opaque and clear sections of human lenses. (4) To develop new methods for retrieving lens material during extracapsular surgery. To approach taken will be tap the aspiration lines of the equipment used during surgery. (5) To identify and characterize a 23,000 dalton crystallin which is selectively degraded and modified in many human cataractous lenses, and to identify and study the proteolytic enzymes which are involved, in part, in this process. (6) To complete the nucleotide sequencing of cDNA clones of the lens plasma membrane main intrinsic protein (MIP-26). (7) To complete the nucleotide analysis of the cow beta 23/26 crystallin. (8) To study the effects of diet and aging on the differential loss of gamma-crystallin in mice which are given a reduced calorie diet.