MOLECULAR BIOLOGY OF DIABETIC CATARACT FORMATION

糖尿病性白内障形成的分子生物学

• 批准号: 6518340
• 负责人: Patrick Ross Cammarata
• 金额: $ 25.41万
• 依托单位: UNIVERSITY OF NORTH TEXAS HLTH SCI CTR
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-07-01 至 2005-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6518340
• 关键词: aldehyde reductase; chloride channels; diabetic cataract; diacylglycerols; disease /disorder model; gene expression; genetically modified animals; immunoelectron microscopy; laboratory mouse; lens; membrane transport proteins; model design /development; northern blottings; osmotic pressure; pathologic process; polymerase chain reaction; sodium; taurine; tissue /cell culture

DESCRIPTION (Adapted from applicant's abstract): Diabetic cataract is a significant and costly worldwide health problem. Intracellular osmotic stress has been implicated in the etiology of diabetic cataract. The definition of the sequence of basic molecular and cellular processes leading to the complications of diabetic cataract have yet to be established. The accumulation of organic osmolytes (sorbitol, myo-inositol, taurine) normally protects the lens against osmotic imbalance by maintaining intracellular osmotic homoeostasis. In the course of maintaining lens homeostasis, the lens epithelial layer preserves itself by utilizing several osmotic compensatory mechanisms, whereas the subjacent fiber cells, likely because of a diminished capacity to osmoregulate, swell and bleb. To obtain a realistic view of the pathophysiological impact of osmotic stress on diabetic cataract formation, a novel transgenic animal model has been developed useful to exploring the pathogenesis and therapy of osmotic cataractogenesis. We have successfully introduced the bovine sodium/myo-inositol cotransporter gene (bSMIT) in several mouse lines and have shown the transgene is functionally expressed in developing lens fibers. Lens fiber swelling and consequent cataractous formation provide a physiological surrogate that simulates the progression of human diabetic cataract. Careful scrutiny of mouse lens regional development and early-onset swelling allows for verification of the hypothesis that the lens fibers are incapable of osmoregulation. The molecular biology and the pathophysiology of the transgenic mouse model exhibiting diabetic cataract will be linked by correlating the level of bSMIT gene expression via in situ hybridization and coupled reverse transcription/polymerase chain reaction with the intralenticular content of free myo-inositol. Lens morphology will be followed by light and electron microscopic evaluation of transgenic and nontransgenic littermates using embryonic lenses up through lenses from six month-old mice. Low transgene-expressing mice, which do not form lens opacities with normal rearing and diet, will be made to do so with a myo-inositol supplemented diet. The determination of the sequence of events in the pathophysiology of diabetic cataract formation will identify sites of intervention and allow for the development of innovative pharmacological agents to prevent or halt the progression of diabetic cataract. One such intervention is to activate chloride channels, which enhances both chloride exit and myo-inositol efflux. This study proposes to test the potential usefulness of enhancing myo-inositol efflux through chloride channels as a means for drug therapy to relieve intrafiber osmotic stress.

描述（改编自申请人的摘要）：糖尿病性白内障是一种 重大且代价高昂的全球健康问题。细胞内渗透压 与糖尿病性白内障的病因有关。的定义 导致并发症的基本分子和细胞过程的顺序 糖尿病性白内障的发病率尚未确定。有机积累 渗透调节剂（山梨醇、肌醇、牛磺酸）通常可以保护镜片免受 通过维持细胞内渗透压稳态来调节渗透压失衡。在 在维持晶状体稳态的过程中，晶状体上皮层得以保留 本身通过利用几种渗透补偿机制，而 下面的纤维细胞，可能是因为渗透调节能力减弱， 膨胀并起泡。为了获得对病理生理学影响的现实看法 渗透压对糖尿病白内障形成的影响，一种新型转基因动物模型 已开发出有助于探索渗透压的发病机制和治疗 白内障发生。我们已成功引进牛 钠/肌醇协同转运蛋白基因（bSMIT）存在于多个小鼠品系中，并具有 显示转基因在发育中的晶状体纤维中功能性表达。镜片 纤维肿胀和随后的白内障形成提供了生理学 模拟人类糖尿病性白内障进展的替代物。小心 对小鼠晶状体区域发育和早发性肿胀的检查允许 验证晶状体纤维不能的假设 渗透压调节。转基因的分子生物学和病理生理学 表现出糖尿病白内障的小鼠模型将通过关联 通过原位杂交和反向耦合检测 bSMIT 基因表达水平 转录/聚合酶链反应与晶状体内的内容物 游离肌醇。透镜形态将跟随光和电子 使用显微镜评估转基因和非转基因同窝仔猪 胚胎晶状体通过六个月大小鼠的晶状体进行观察。低的 表达转基因的小鼠，正常饲养时不会形成晶状体混浊 和饮食，将通过补充肌醇的饮食来做到这一点。这 确定糖尿病病理生理学中的事件顺序 白内障的形成将确定干预部位并允许 开发创新药物以预防或阻止 糖尿病性白内障的进展。其中一种干预措施是激活氯化物 通道，增强氯离子排出和肌醇流出。这项研究 提议测试增强肌醇流出的潜在用途 通过氯离子通道作为药物治疗的手段来缓解纤维内 渗透压。