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V. Grami, V.N. Reddy, F.J. Giblin, V.R. Leverenz, D. Tang, D. Borchman; Fluorescence Profiles of Hyperbaric Oxygen/UVA Treated Guinea Pig Lenses, GPX–1 Knock Out Rat Lenses and Clear Human Lenses . Invest. Ophthalmol. Vis. Sci. 2005;46(13):2903.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Fluorophores may contribute to or be markers of cataract. Lenses from guinea pigs (GP) treated with hyperbaric oxygen and treated with chronic UVA light, as well as lenses from glutothione peroxidase–1 knockout rats (GPX1–ko) have shown signs of oxidative damage. By using fluorescent spectroscopy, lens flourophores from human donors were compared to lens flourophores from rat and GP models. Methods: GP lenses from control and treated (66 treatments of hyperbaric oxygen and chronic UVA light over 22 weeks) groups, were obtained at age about 23.5 months. Each GP lens was dissected into nuclear, equatorial and cortical regions. Fifteen month old rat lenses were obtained from GPX1–ko and controls. Human lenses (31–74 y) were separated into cortical and nuclear regions. After homogenization in methanol, fluorescence excitation/emission/intensity spectra were measured with an emission range of 200–650 nm and an excitation range of 200–800 nm. Results:Three peaks were prominent in the 3D excitation/emission fluorescence spectra of rat, GP, and human lens protein suspensions. Peak I near 295/340 (nm excitation/emission) was the most consistent and prominent peak. Peak II 360/445, a diagonal ridge Peak III 265/575, and Peak IV 470/520 were present in all species. Fluorescence spectra from rats and GP were more similar to each other than to human lens spectra. For human data, in relations to Peak I, Peak II was about 75 %. Peak III was about 17 times larger, and peak IV was 10% of Peak I. For the rat and GP data, in relations to Peak I, Peak II was about 0.5%, and Peak IV was about 2%. Peak III varied among the GP and rat lenses. The intensity of Peak II relative to Peak I was smaller in paired controls, consisting of untreated GP and native rats, relative to treated GP and GPX1–ko rats. This was true for all 3 regions of the GP lens. The ratio Peak II/Peak I increased with age in the human lenses. The relative intensities of Peaks III and IV were higher in all 3 regions of treated GP lenses and were higher in GPX1–ko samples compared to controls. Peaks III and IV decreased with increasing age in human lenses. Conclusions: The advantage of 3D excitation/emission spectra over conventional 2D fluorescence spectra is that all changes in the fluorescence characteristics can be quantified that could be potentially missed in 2D spectra. By comparing 3D fluorescence spectra from human and model systems, we may begin to determine the fluorescent components that are markers of oxidation and cataract.
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