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M. Liu, E. Y. Chew, C. Meyerle, C.-C. Chan, J. Tuo; Copy Number Variations for Eight Genes in Neovascular Age-Related Macular Degeneration. Invest. Ophthalmol. Vis. Sci. 2010;51(13):1273.
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Age-related macular degeneration (AMD) is the leading cause of central vision loss among the elderly in developed countries, and its pathogenesis is strongly influenced by genetic risk factors. Single nucleotide polymorphisms (SNPs) in several genes have been consistently linked to AMD. Copy number variation (CNV), or variation in the number of copies of a DNA segment, may be an additional type of genetic variation that contributes to AMD pathogenesis. This study investigated CNVs in 8 AMD-relevant genes for AMD cases and controls.
Peripheral blood was collected from 88 neovascular AMD cases and 89 age-matched controls at the National Eye Institute. Genomic DNA was extracted and quantified by UV absorption and RT-PCR amplification of RNase P using standard DNA as a calibrator. The RT-PCR based TaqMan Copy Number Assays were performed for the genes CCR3, CFH, CX3CR1, ERCC6, GSTM1, GSTT1, HtrA1, and VEGF. Samples were analyzed in triplicate using RNase P as an internal control. Copy numbers were computed using the ΔΔCt method with the aid of the Applied Biosystems CopyCaller Software. Samples had been previously genotyped for AMD-relevant SNPs, and these results have been reported.
Copy numbers (0, 1, 2, or 3+ copies) were reliably determined for the 8 genes of interest in each sample. Assays were repeated on 7% of samples for each gene, and copy number calls were consistent in >97% of duplicates. CNV frequencies were compared between cases and controls using the Chi-square test. Although no statistically significant CNVs were observed (p<0.05), the data suggest that CNV in CX3CR1 may be relevant to AMD pathogenesis. Independently, a gain in CX3CR1 copy number may be protective (OR=0.58, p=0.12). Loss in CX3CR1 copy number may account for the reported suppressed CX3CR1 expression levels in AMD maculae. Homozygosity for the CX3CR1 T280M SNP and 3 copies of CX3CR1 confers a statistically significant increase in AMD risk (OR=10.2, p=0.009).
This study investigated CNVs in 8 AMD-relevant genes, most of which had previously never been characterized for CNV. Although no statistically significant CNVs were found, the current sample size likely offers insufficient power due to heavy stratification by CNV category. A trend suggests that CNV in CX3CR1 may contribute to neovascular AMD pathogenesis, both independently and through a statistical interaction with CX3CR1 T280M SNP genotype.
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