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G. Malek, J. Ebright, B. Mace, P. Sullivan, C. Bowes Rickman; Oxidative Stress and Immune–Related Genes Are Upregulated in the Retina/RPE/Choroid of Mice With Age–Related Macular Degeneration Pathology . Invest. Ophthalmol. Vis. Sci. 2006;47(13):4161.
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© ARVO (1962-2015); The Authors (2016-present)
Age–related macular degeneration (AMD) is a degenerative retinal eye disease that causes progressive loss of central vision. Aged APOE4 targeted replacement mice fed a high fat, cholesterol–rich diet develop characteristic lesions of AMD including basal deposits and neovascularization, and serve as an animal model for AMD. These changes were not seen in APOE3 mice. A comprehensive transcript profile in mice with ‘disease’ vs. ‘normal’ morphology was obtained by microarray analysis and used to elucidate differential gene expression contributing to the disease phenotype.
Right eyes from aged, APOE4 and APOE3 mice, fed normal or high fat, cholesterol–rich diets (n=3) were enucleated and fixed in RNAlater. The left eyes were used to confirm morphology, previously determined to be bilateral, through histochemistry. Total RNA was isolated from the retina/RPE/choroid samples using TriZol–glycogen, amplified and biotin–labeled for hybridization. Expression analysis was performed using Operon Mouse Oligo Set GeneChips, representing 30,000 genes. Samples were hybridized onto microarray slides, visualized with the GeneChip scanner. Data were normalized and clustered using GeneSpring GX software. The criteria used for gene selection was 1.5 fold or greater change and p–value of less than 0.05.
GeneSpring array analysis revealed significant changes in the expression of 2961 genes in samples from ‘diseased’ versus ‘normal’ mouse eyes. Candidate genes were prioritized based on pathways and processes implicated in AMD pathogenesis. Using gene ontology, genes were clustered based on their biological function into five major categories: inflammation–, oxidative stress–, lipid and cholesterol–, cell death– and extracellular matrix–related. We are currently confirming differential gene expression changes using quantitative real–time PCR, immunohistochemistry and in situ hybridization on mice with and without AMD pathology.
Comparative analysis of the gene expression profiles of aged APOE4 mice with AMD pathology versus controls supports the activation of inflammatory and oxidative stress related genes in the pathology of the disease. These results provide us with insight into the processes that underlie, age–, APOE4 isoform– and diet–induced changes/damage in the eye and their possible involvement in AMD.
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