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J. T. Handa, Y. Yamada, J. Tian, Y. Yang, R. Cutler, T. Wu, R. Telljohann, M. Mattson; Pathologic Gene Expression Mediated by Oxidized LDL in Retinal Pigmented Epithelial Cells. Invest. Ophthalmol. Vis. Sci. 2007;48(13):5637.
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The accumulation of low density lipoproteins (LDL) in Bruch membrane is an early event that is hypothesized to promote age-related macular degeneration (AMD), the most common cause of blindness in western societies.
RPE cells were exposed to LDL or oxidatively modified LDLs (ox-LDL). Total RNA was extracted and subjected to the Affymetrix U133 Plus 2.0 gene chip and/or RT-qPCR. Cell extracts were analyzed by electrospray mass spectrometry or western analysis. To determine a functional impact of ox-LDL treatment, RPE cells were analyzed by TUNEL labeling. Human maculas with AMD were analyzed by immunohistochemistry.
Microarray analysis showed a physiological and pathological transcriptional response after LDL and ox-LDL treatment, respectively. LDL induced a downregulation of cholesterol biosynthesis genes while ox-LDL induced transcriptional alterations in genes related to inflammation including pentraxin-3, matrix expansion, lipid metabolism, and apoptosis. Electrospray mass spectrometry showed that ox-LDL, but not LDL induced large increases in sphingomyelin, ceramides, and cholesterol in RPE cells. With TUNEL labeling, ox-LDL induced 38% apoptosis compared to <1% after LDL or control (p<0.0001). Ox-LDL, but not LDL stimulated RPE cells to secrete pentraxin-3 into the culture medium. Immunohistochemistry of 12 AMD samples showed strong staining for pentraxin-3 in normal Bruch membrane, but light staining within basal deposits.
The transcriptional, biochemical, and functional data support the hypothesis that modified LDLs are one trigger for initiating early events in the pathogenesis of AMD.
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