SPECTROSCOPIC AND RELATED STUDIES ON LENS MEMBRANE LIPID

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The overall objective of our proposed investigation is to use state-of-the-art spectroscopic techniques to examine regional conformational; compositional and structural characteristics of lens membrane lipids and proteins. Spectroscopic findings could be related to membrane composition and performance. We will investigate normal bovine and human lenses, cataractous human lenses and lenses subjected to a variety of physicochemical stresses. Such studies well permit us to develop an in depth appreciation of relatively subtle changes in membrane properties which can lead to considerable changes in membrane performance contributing to cataractogenesis. The results of this study will strengthen the foundation of knowledge necessary to assure future development of medical therapy of human cataract. The specific aims of this study are as follows: 1. Measure primary infrared and Raman spectroscopic characteristics of lipid vesicle dispersions obtained from normal bovine and human lenses. Spectroscopic characteristics will include: a) fluidity and phase transition temperatures; b) orthorhombic or monoclinic packing order; c) lipid head group interactions; d) lipid acyl-interface region order and packing; e) quantitate lipid saturation; f) confirm lipid composition. The membrane lipid composition of the vesicle dispersions will be determine by thin layer chromatography and high performance liquid chromatography. 2. Determine the influence of intrinsic and extrinsic membrane proteins upon spectroscopic characteristics of lens cell membranes. 3. Explore changes in spectroscopic characteristics accompanying aging of the lens and in human cataract. 4. Determine the impact of oxidative stress, UV light exposure, calcium and the cholesterol binding antibiotic filipin upon the spectroscopic characteristics of lens cell membranes and membrane lipids. 5. Explore the correlation of membrane lipid spectroscopic characteristics with lens membrane performance i.e. ATPase activity and membrane vesicle solute flux.