Fiber Cell Junctions in Normal and Cataractous Lenses

描述（由申请人提供）：这是一项长期资助的修订竞争性更新，旨在了解人类年龄相关的核性白内障形成的细胞基础，核性白内障是人类白内障最常见的形式，也是全球失明的主要原因。在上一个授权期内，在表征纤维细胞膜的损伤、纤维细胞质中晶体蛋白的分布以及包含被多个脂质双层覆盖的细胞质核心的独特球形颗粒（称为多片层体）方面取得了重大进展。通过对每种类型的细胞损伤的预测散射进行理论分析，对该领域做出了值得注意的贡献。这些研究导致了一个重要的修改，通常持有的建议，核白内障是由蛋白质聚集成高分子量的聚集体引起的。我们的研究结果表明，早期核性白内障的纤维细胞超微结构与老化的透明晶状体几乎相同，这表明最初的蛋白质修饰和关联是自然衰老过程的一部分。只有在印度晚期白内障中进行了广泛的蛋白质修饰后，才有可能使用电子断层扫描识别潜在的HMW聚集体。我们的研究结果表明，多层体对干扰中央凹正常成像的前向散射起主要作用，并且随着年龄的增长和主要由氧化应激引起的细胞损伤的积累，这种影响会增加视力障碍。我们建议扩大我们的超微结构研究，使用扫描和透射电子显微镜，电子断层扫描，冷冻电子显微镜和共聚焦/多光子光学显微镜来表征各种核性白内障的细胞损伤，可以与年龄相关的核性白内障进行比较。改进的保存和分辨率将产生有关晶体蛋白的年龄相关转化及其与膜的关联的有价值的新信息。定量结构数据将进行理论分析，以评估它们对透镜散射的预测贡献。我们最近的证据首次表明，自噬和有丝自噬发生在晶状体中，并且自噬很可能是多层小体的来源。我们建议在不同年龄的人和鸡胚晶状体中探索多层体的形成，作为一种动物模型，这将允许识别和调节参与自噬的基因。在晶状体早期发育过程中，营养压力可能会增强自噬。这些超微结构研究和理论分析有望更好地理解核性白内障形成过程中产生过度光散射的细胞损伤机制。

DESCRIPTION (provided by applicant): This is a revised competitive renewal of a long-standing grant to understand the cellular basis for the formation of human age-related nuclear cataracts, the most common form of human cataract and a leading cause of worldwide blindness. In the last granting period significant progress was made in characterizing damage to fiber cell membranes, the distribution of crystallins in fiber cell cytoplasm and unique spherical particles containing a core of cytoplasm covered by multiple lipid bilayers, termed multi-lamellar bodies. Noteworthy contributions were made to the field by providing theoretical analyses of the predicted scattering from each type of cellular damage. These studies have led to an important modification to commonly held proposal that nuclear cataracts are caused by protein clumping into high molecular weight aggregates. Our results suggest that the fiber cell ultrastructure of early stage nuclear cataracts is nearly identical to aged transparent lenses, suggesting that initial protein modifications and associations are part of the natural aging process. Only after extensive protein modification in advanced cataracts from India, has it been possible to identify potential HMW aggregates using electron tomography. Our results suggest that multi-lamellar bodies make a major contribution to forward scatter that interferes with normal image formation at the fovea and this contribution to visual impairment increases with age and accumulation of cell damage caused mainly by oxidative stress. We propose to expand our ultrastructural studies employing scanning and transmission electron microscopy, electron tomography, cryo-electron microscopy and confocal/multi-photon light microscopy to characterize cellular damage in a variety of nuclear cataracts that can be compared to age-related nuclear cataracts. Improved preservation and resolution will yield valuable new information about the age-related transformations of crystallins and their associations with membranes. Quantitative structural data will be analyzed theoretically to evaluate their predicted contribution to lens scattering. Ou recent evidence shows, for the first time, that autophagy and mitophagy occur in the lens and that autophagy is most likely the source of multi-lamellar bodies. We propose to explore multi-lamellar body formation in humans of varying ages and in chick embryonic lenses as an animal model that will allow identification and regulation of genes involved in autophagy. Autophagy is enhanced by nutritional stress that could be important during early lens development. These ultrastructural studies and theoretical analyses are expected to lead to better understanding of the mechanisms of cell damage that produce excessive light scattering in nuclear cataract formation.