LENS EPITHELIUM AND HELIUM ION CATARACTOGENESIS

晶状体上皮和氦离子白内障发生

• 批准号: 2164817
• 负责人: ELEANOR A BLAKELY
• 金额: $ 28.45万
• 依托单位: UNIVERSITY OF CALIF-LAWRENC BERKELEY LAB
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-09-01 至 1999-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2164817
• 关键词: DNA damage; HeLa cells; SDS polyacrylamide gel electrophoresis; cataract; cell differentiation; cellular pathology; chromosome aberrations; cytogenetics; enzyme linked immunosorbent assay; eye neoplasms; fibroblast growth factor; helium; human tissue; iatrogenic disease; in situ hybridization; lens proteins; melanoma; messenger RNA; neoplasm /cancer radiation therapy; northern blottings; radiation genetics; radiation therapy dosage; tissue /cell culture; tissue mosaicism; western blottings

Cataracts arise in uveal melanoma patients as a complication following their successful treatment with helium-ion radiotherapy. The objective of the proposed research is to determine the helium-ion-induced alterations in chromosomes and in protein expression that are important to cataractogenesis, and to develop strategies to diminish the incidence or severity of these changes. The current paradigm for cataractogenesis is focussed on genomic damage of lens epithelial cells leading to altered crystallin proteins. This proposal will examine two alternative mechanisms of cataractogenesis involving radiation-induction of basic Fibroblast Growth Factor (bFGF) in human lens epithelial (HLE) cells functioning either to alter the normal program of crystallin expression and thereby disrupting normal fiber formation, or the radiation-induced bFGF acting to hinder cell loss processes which leads to the formation of aberrant lens fiber formation. To this end experiments have been designed with three specific aims that will: Firstly, characterize acute and residual responses to helium-ion irradiation of cultured HLE cells grown on extracellular matrix. Quantitative dose-response measurements between helium-ion induced survival, repair capacity and yields of micronuclei, apoptotic nuclei, and persistent chromosomal rearrangements will be determined. Secondly, since radiation is known to induce bFGF in endothelial cells leading to inhibition of apoptosis, experiments are proposed that will determine whether helium-ion radiation changes levels of bFGF mRNA or protein in HLE cells, and whether there is a correlation with apoptotic events. Finally, an investigation of the possible modification of intracellular lens proteins by helium-ion radiation is proposed. The program will elucidate relationships between helium-ion- induced damage to the chromatin of HLE cells in vitro, and biological consequences to the cells surviving the resulting damage. In the revised application significant changes include: 1) an increased source of HLE cells, 2) new preliminary data from mRNA and protein analyses (SDS-PAGE) of exponential and confluent cell cultures, 3) new preliminary data with immunological evidence confirming feasibility to characterize lens cell differentiation, 4) some revised experimental protocols, and 5) and an updated review of literature published in this rapidly increasing area. This knowledge will allow correlative comparisons with available experimental work in vivo, and may provide a basis for potential changes to the treatment protocol that could reduce the risk of cataract. The biological mechanisms contributing to cataractogenesis may also be elucidated.

白内障是葡萄膜黑色素瘤患者的并发症， 氦离子放射治疗的成功。客观 建议的研究是确定氦离子诱导的 染色体和蛋白质表达的改变， 白内障的发生，并制定战略，以减少发病率 这些变化的严重性。白内障发生的当前模式 重点关注导致改变的透镜上皮细胞基因组损伤 晶状体蛋白质。本提案将审查两种备选方案， 白内障发生的机制涉及辐射诱导的基础 人透镜上皮细胞中的成纤维细胞生长因子 其功能是改变晶体蛋白表达的正常程序 从而破坏正常的纤维形成，或辐射诱导的 碱性成纤维细胞生长因子的作用，以阻止细胞损失的过程，这导致形成 异常的透镜纤维形成。为此， 设计有三个具体目标，将：首先， 培养的HLE细胞对氦离子照射的残留反应 生长在细胞外基质上定量剂量反应测量 氦离子诱导的存活、修复能力和 微核、凋亡核和持续性染色体重排 将被确定。其次，由于已知辐射可诱导bFGF， 导致细胞凋亡抑制的内皮细胞，实验是 这将决定氦离子辐射是否会改变 bFGF mRNA或蛋白在HLE细胞中的表达，以及是否存在相关性 凋亡事件。最后，对可能的 通过氦离子辐射修饰细胞内透镜蛋白， 提出了该计划将阐明氦离子之间的关系， 在体外诱导HLE细胞染色质损伤， 对细胞造成的伤害。订正 应用显著变化包括：1）HLE来源增加 细胞，2）来自mRNA和蛋白质分析（SDS-PAGE）的新的初步数据 指数和汇合细胞培养物，3）新的初步数据， 免疫学证据证实表征透镜细胞可行性 分化，4）一些修订的实验方案，和5）和一个 对这一快速增长领域发表的文献进行更新综述。 这些知识将允许与现有的 在体内的实验工作，并可能提供潜在的变化的基础 降低白内障风险的治疗方案。的 导致白内障发生的生物学机制也可能 阐明。