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B. Bakall, R.A. Radu, J.B. Stanton, J. Burke, B.S. MacKay, C. Wadelius, C. Appelqvist, G.H. Travis, A.D. Marmorstein; Analysis of Donor Eyes From a Best Vitelliform Macular Dystrophy Patient, Homozygous for the Bestrophin W93C Mutation . Invest. Ophthalmol. Vis. Sci. 2006;47(13):5817.
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To determine the biochemical and histopathologic consequences of Best Vitelliform Macular Dystrophy, in eyes from a donor homozygous for the W93C mutation in bestrophin.
Eyes were enucleated post mortem. One eye was fixed in 4% Paraformaldehyde, embedded in paraffin, and sectioned. Sections were stained for histological analysis and immunocytochemistry was performed to localize bestrophin. The remaining eye was frozen directly. Retinal pigment epithelium (RPE) was isolated for biochemical analysis. The expression of bestrophin was confirmed by western blot. Intracellular granules were subject to sucrose gradient sedimentation. Granules were counted using a hemocytometer, examined by electron microscopy and analyzed for A2E and its chemical precursors.
Histologic analysis confirmed complete degeneration and scarring of the macula. Throughout the eye, photoreceptor degeneration was observed and RPE cells appeared hypertrophic with substantial accumulation of intracellular granules. Fluorescence microscopy indicated substantial autofluorescence of granules which were confirmed to contain A2E and its precursors. While A2E levels per granule were lower than in age matched control eyes, the increase in total granules resulted in a 5–7 fold increase in total A2E/eye in the Best donor eye. Analysis of the granules by electron microscopy indicated a complex multilobular structure that differed substantially from granules isolated from age matched control eyes. Immunohistochemistry for bestrophin showed localization exclusive to the RPE.
These donor eyes provide, for the first time, the histopathology of eyes homozygous for a bestrophin mutation. The study confirms prior findings, where RPE appear increased in size with lipofuscin accumulation. This is the first report indicating that levels of A2E are increased in Best disease.
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