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R. Farjo, T.S. Kern, U.P. Andley, A. Swaroop; Microarray Analysis of Mouse Models of Diabetic Retinopathy Reveals Dramatic Modulation of Crystallin Genes in the Retina . Invest. Ophthalmol. Vis. Sci. 2003;44(13):3297.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: To identify changes in retinal gene expression in mouse models of diabetic retinopathy (DR) and in response to pharmacological treatments. The primary goals of our studies are: To elucidate molecular pathways underlying the pathogenesis of diabetic complications; Develop surrogate markers for early detection of diabetic complications and evaluation of treatment parameters. Methods: Diabetes was induced in male C57BL/6 mice by daily injections of streptozotocin for 5 successive days, each injection following an 8-hour fast. Insulin was given to diabetics as needed to achieve slow weight gain without preventing hyperglycemia. Diabetic mice were randomized to receive diet supplemented with aminoguanidine, aspirin, or α-tocopherol. Total RNA samples isolated from retinas of control, diabetic, and treated mice were labeled using the Submicro kit (Genisphere) and hybridized to microarray slides containing 6500 eye-expressed genes printed in duplicate. Microarray image analysis was performed with DigitalGenome software (Molecularware). Raw intensity and background data were normalized using a print-tip based lowess smoothing algorithm. Gene expression changes were extracted after statistical analysis and validated by real-time RT-PCR. Anti-Crystallin antibodies are being used to validate if RNA changes are reflected in corresponding protein levels. Results: Using custom I-Gene microarrays, we have identified over 30 genes that are differentially expressed in the retina of DR mice. Of these, many genes belonging to the crystallin family are drastically downregulated in the retinas of DR mice. Real-time RT-PCR confirmed these changes and revealed that drug-supplementation of DR mice had a positive effect and the expression levels of many crystallin genes approached wild-type retinal expression. We will present baseline gene expression profile of DR retina with a focus on crystallin genes and their modulation in response to various treatments. Conclusions: Expression profiles of retinal disease states can be readily accomplished with microarray technology. We propose that crystallins play a major, as yet unidentified, protective role in the retina. These studies should provide better understanding of molecular and cellular networks by which DR manifests itself and assist in evaluating the role of drug supplements and designing more rational drug design.
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