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G Malek, C Guidry, NE Medeiros, CA Curcio; Localization of Apolipoproteins and Cholesterol in Macular and Peripheral Bruch's Membrane, Basal Deposits (BD) and Drusen of Normal and Age-related Maculopathy (ARM) eyes . Invest. Ophthalmol. Vis. Sci. 2002;43(13):2789.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: To localize apolipoproteins E, B (apo E, apo B) and cholesterol in macular and peripheral Bruch's membrane, BD and drusen of normal and ARM eyes; to examine the presence of apo E and B in human retinal pigment epithelium (RPE). Methods: Normal (n=5) and ARM (n=5) donor eyes were preserved <4 hrs after death in 4% paraformaldehyde. Serial cryosections 10µm thick were cut. ARM eyes contained RPE changes, BD and/or drusen ≷63µm. Apo E and B were identified through indirect immunofluorescence using polyclonal antibodies. Antibody specificity was confirmed through western blot analysis of human plasma. The histochemical stain filipin was used to demonstrate unesterified cholesterol (UC), or esterified cholesterol (EC) following extraction and hydrolysis. Western blot analysis was also used to screen human RPE extracts (n=3) for apo E and B. Results: 1) Histopathologic examination of eyes revealed: many peripheral and few macular deposits in normal eyes; peripheral and macular deposits in ARM eyes. 2) Apo E and B immunoreactivity was present in peripheral BD and drusen of normal and ARM eyes, but varied in pattern and extent in macular BD and drusen. 3) EC/UC and apo B co-localized in peripheral BD and drusen of normal and ARM eyes. EC/UC was present in all macular apo B-positive BD and drusen of ARM eyes. However the converse was not the case: apo B was not present in all EC/UC-rich BD and drusen. 4) Apo E immunoreactivity was present in normal and ARM Bruch's membrane. Apo B immunoreactivity was not present in Bruch's membrane except in association with BD and drusen. 5) Human RPE extracts contained apo E but not apo B. Conclusion: Since apo B is the principal protein in low density lipoproteins (LDL) and LDL transports cholesterol in plasma, it is plausible that apo B deposition in BD and drusen indicates LDL retention. The partial co-localization of apo B and EC/UC suggests LDL may be one means by which cholesterol accumulates in BD and drusen. While peripheral BD and drusen in our eyes consistently contained apo E and B, apolipoprotein content of macular BD and drusen was much more variable. To our knowledge this is the first demonstration of differences in biochemical constituents of BD and drusen between the macula and periphery. Results of the western blot analysis suggest RPE is a potential local source for apo E but not apo B, consistent with previous reports of apo E mRNA in RPE.
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