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I. Kozak, L. Cheng, R. Sasik, G. Hardiman, S. Rought, L. J. Pearson, C. Woelk, W. R. Freeman; Non-Infectious HIV Retinopathy: Pathway and Gene Ontology Based Analysis of Gene Expression in Lesions From Human Autopsy Eyes. Invest. Ophthalmol. Vis. Sci. 2009;50(13):3570.
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We and others have shown that HIV retinopathy including cotton wool spots and related non infectious lesions are evidence of focal retinal damage and may be associated with accumulating vision dysfunction and structural damage to the retina. We therefore applied microarray analysis techniques to study expression of various genes in retinal cotton wool spots in HIV retinopathy from human autopsy eyes.
Autopsy eyes from patients who had died of acquired immunodeficiency syndrome (AIDS) were placed in RNA-later less than 24 hours after enucleation. After opening they were flat mounted and visible cotton wool spots were punched using a PCR-clean 2 mm trephine. Control punches from the posterior pole and peripheral retina were procured from eyes with and without lesions and from eyes of age-matched HIV-negative individuals. Each punch was placed into separate vial containing 1.0 ml Trizol and stored at -80C until processing. After mRNA extraction, the genetic expression profile of the mRNA was analyzed using Illumina Microarray Sentrix Expression Bead Chips (Illumina, San Diego). The arrays were scanned on BeadArray reader imaging system with 532 (cye3) nm laser illumination. Data analysis was carried out using the BeadStudio software. Statistical analysis proceeded in three steps: 1. normalization of microarray data, 2. sorting of genes according to interest and 3. statistical analysis of pathways and gene ontology terms represented by the sorted list of genes.
Bonferroni-adjusted comparison of genes from areas with cotton wool spots has shown significantly increased expression of genes involved in immune response (p=1.3 E-27) compared to non-lesion areas of HIV+ eyes. The same comparison of non-lesion areas from HIV+ eyes has shown significantly increased expression of genes involved in immune system process (p=4.1 E-27) compared to HIV-negative retinae.
Microarray analysis is a powerful tool in the genomic research of ocular HIV disease. It has shown difference in gene expression between HIV+ and HIV-negative retinas and involvement of several pathophysiologic pathways in HIV retinopathy lesions.
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