CALCIUM AND THE NORMAL LENS AND CATARACT

钙在多种细胞信号系统中起着关键作用 越来越多的人认识到在发展中发挥着重要作用 皮质性白内障。这个项目的主要目的将是调查 晶状体中实现钙稳态的机制以及 这些机制对外界侮辱的敏感性。进一步的目标将是 为了阐明钙激活的酶，钙激活酶，在 镜片混浊的产生。我们将对钙稳态进行研究， 主要使用人类晶状体组织，但也用于体外培养的大鼠晶状体， 通过电生理、放射性示踪剂和荧光染料的组合 技巧。Thapsigargin，选择性地抑制血管内皮细胞CaATPase 内质网在这些研究中将是一个无价的工具。 并将使用数学建模技术来解释45Ca通量 来自整个人类晶状体的数据。钙离子通透性的敏感性 (渠道)氧化侮辱的途径也将在 完整的人和大鼠晶状体和组织培养细胞中。阳离子通道 抗体也将与免疫荧光和 金标记法研究阳离子通道的定位 晶状体上皮细胞和纤维细胞。 单个晶状体细胞和整个大鼠晶状体细胞内钙离子浓度升高 外源性钙离子载体(A23187)对人晶状体的诱导作用 膜SH络合作用(PCMPS)与钙信号激动剂 (乙酰胆碱)，并将用上述技术进行定量测定。 这些药剂对晶状体透明度的影响将被整体量化。 透镜通过成像散射光进行实验。对晶状体的影响 细胞骨架(主要是肌动蛋白、微管蛋白、波形蛋白和血影蛋白)将 DSS-PAGE电泳法和免疫荧光法相结合的研究 本地化技术。晶状体共焦的伴随研究 阳离子通道和细胞骨架元素将在 正常和钙超载的晶状体组织。 这项研究将在以下两个方面产生有价值的信息 钙是受控制的，钙的产生机制也是如此- 失控后导致蛋白分解和混浊。在.期间 在项目的过程中，将采取一种综合的方法，以实现这两个目标 应用通道阻滞剂减少钙超载及预防 钙诱导的蛋白水解酶的抗钙蛋白化合物。

Calcium plays a critical role in a variety of cell-signalling systems and is increasingly recognised to play a significant part in athe development of cortical cataract. The main aims of this project will be to investigate the mechanisms for achieving calcium homeostasis in the lens and also the sensitivity of these mechanisms to external insult. A further aim will be to elucidate the role of the calcium-activated enzyme, calpain, in the production of lens opacities. Calcium homeostasis will be studied, principally using human lens tissues, but also on athe rat lens in vitro, by a combination of elecrophysiological, radiotracer and fluorimetric dye techniques. Thapsigargin, which selectively inhibits the CaATPase of the endoplasmic reticulum, will be an invaluable tool in these investigations and mathematical modelling techniques will be used to interpret 45Ca flux data from whole human lenses. The sensitivity of calcium permeability (channel) pathways to oxidative insult will also be studied in detail in whole human and rat lenses and in tissue cultured cells. A cation channel antibody will also be used with a combination of immunofluorescence and gold-labelling technique to study the localization of cation channels in lens epithelia and fibre cells. Increases in cytosolic calcium in single lens cells and in whole rat and human lenses will be induced by calcium ionophore (A23187), external membrane SH complexation (pCMPS) and calcium signalling agonists (acetylcholine) and will be quantitatively assayed by the above technique. The effect of these agents on lens transparency will be quantified in whole lens experiments by imaging scattered light. The effect on the lens cytoskeleton (principally actin, tubulin, vimentin and spectrin) will be studied by a combination of DSS-PAGE electrophoresis and immunofluorescence localization techniques. A concomitant study of the colocalization of lens cation channels and cytoskeletal elements will be carried out both in normal and calcium overloaded lens tissues. This study will yield valuable information both on the mechanisms whereby calcium is controlled and also the mechanism responsible for calcium- induced proteolysis and opacification following loss of control. During the course of the project a combined approach will be made that aims both to reduce calcium overload by channel blocking agents and to prevent calcium-induced proteolysis by anti-calpain compounds.