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Albert R. Wielgus, Scott W. Cousins; The Effect of -3 and -6 Polyunsaturated Fatty Acid Peroxidation Products on RPE Cell Morphology: Implications for Development of Sub-retinal Deposits in Early AMD. Invest. Ophthalmol. Vis. Sci. 2011;52(14):2309.
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© ARVO (1962-2015); The Authors (2016-present)
To investigate RPE cellular changes relevant to the formation of sub-RPE deposits induced by the addition of 4-HNE and 4-HHE, the peroxidation products of ω-6 and ω-3 polyunsaturated fatty acids (PUFA-s), respectively. Epidemiological data implicate a pathological role for ω-6 fatty acids and a potential protective role for ω-3 fatty acids in the development of drusen and early AMD. We hypothesize that the lipid peroxidation by-products of ω-6 can directly trigger biochemical changes in cytoskeleton and injury kinase pathways that would predispose to deposit formation.
Differentiated ARPE19 cells were fed arachidonic (ω-6 PUFA), docosahexaenoic (ω-3 PUFA), 4-HNE or 4-HHE. Morphologic changes of the cells and cytoskeleton alterations during the incubation process were monitored using immunocytochemical staining. Protein expression was determined with Western blot analysis. The effect of PUFA-s and their peroxidation products on cell membrane permeability was detected LDH assay.
We found that ARPE19 cells treated with native PUFA-s at concentration up to 20 µM did not show significant changes in their morphology. However, cells treated with sublethal doses of 4-HNE demonstrated dramatic alterations in their shape and modification of cytoskeleton structure. The morphological changes in the cells were accompanied by modified expression and activation of proteins involved in actin filament dynamics, such as HSP27 phosphorylation and increased permeability of cytoplasm membrane. Interestingly, the changes of the cell shape, modification of filamentous actin and increased cell membrane permeability were less significant in RPE cells treated with the same sub-lethal doses of 4-HHE.
4-HNE as a product of ω-6 PUFA peroxidation shows stronger modifications in RPE cell morphology and cytoskeleton structure than 4-HHE - the product of ω-3 PUFA oxidation. 4-HNE causes modification of the RPE cell membrane, which can induce release of cellular material that contributes to formation of deposits and drusen in Bruch’s membrane.
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