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Ralph Michael, Cees Otto, A. Lenferink, Gustavo Montenegro, Rafael Barraquer, Gijs Vrensen; Conformational characteristics of proteins and lipids in transparent human lenses and lenses with cortical opacities - A Raman spectroscopic study. Invest. Ophthalmol. Vis. Sci. 2013;54(15):5732. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Comparison of the conformational characteristics of the molecular bonds of proteins and lipids in cortical cataracts in comparison with transparent lenses by Raman spectroscopy.
Eleven human donor lenses (aged 63-93 years) with pronounced cortical opacities (n = 5), minor cortical opacities (n =2) or without cortical opacities (n = 4) were fixed in paraformaldehyde and cut in axial slices (± 500 µm). Raman spectra were taken for areas of 30 x 30 µm. All lenses were measured at three sites below the capsular surface at: the superficial cortex (200 µm), the deep cortex (800 µm) and the lens nucleus (about 1000 µm off lens center). Protein and lipid conformations were analysed in the 500-1800 cm-1 fingerprint region. Protein concentration was calculated from the ratio of the Raman peaks at 3390 cm-1 (water) and 2935 cm-1 (CH-stretch of proteins).
Cortical cataracts were restricted to a depth of 400-1000 µm below the lens capsule, mostly leaving the superficial cortex transparent. Lens nuclei showed some minor decrease in transparency and had different degrees of coloration. The Raman spectra exhibited a varying overall intensity, increasing from the superficial cortex towards the nucleus with a pronounced increase in lenses with more nuclear coloration. This variation was due to variation in local protein content and fluorescence. After correction for these variations, unrelated to lipid and protein conformations, all Raman peaks and peak ratios were very similar in all lenses, irrespective of the presence of opacities. Specifically the cholesterol peak at 1586 cm-1, the phenylalanine peak at 1003 cm-1 and the amide (beta-sheet) peak at 1666 cm-1 were very much alike. Mean protein concentration in the superficial cortex was 13% in lenses with cortical cataract and 35% in transparent lenses, in the deeper cortex and the nucleus it was 35%, independent of the presence of cortical cataract.
Raman spectroscopy showed that cortical cataracts are not accompanied by conformational changes of lens proteins and lipids. A low protein concentration in the superficial cortex is correlated with the presence of pronounced cortical opacities in the deep lens cortex.
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