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Cody Ronald Fisher, Mara Supik, Sandra Rocio Montezuma, Deborah A. Ferrington; Effects of oxidative stress and AMD on mitochondrial homeostasis in cultured human primary RPE. Invest. Ophthalmol. Vis. Sci. 2018;59(9):345. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Oxidative stress and age-related macular degeneration (AMD) have been shown to adversely affect mitochondrial function. The purpose of this study was to determine how the cell regulates mitochondria under oxidative stress and how this differs in primary retinal pigment epithelium (RPE) cultures from donors with (AMD) or without AMD (No AMD).
The presence of AMD in donor eyes was determined using the Minnesota Grading System (MGS). Primary RPE cell cultures were developed from donors (ages 50-80 yrs) without AMD (MGS1,n=5) and with AMD (MGS2 and 3, early and intermediate AMD, n=8). Cells were grown to confluence and treated with 300uM hydrogen peroxide for 0 (no peroxide), 6 or 24 hours. Expression of genes associated with mitochondrial biogenesis (AMPK, TFAM), mitochondrial autophagy (PINK1,PARKIN,VDAC), and fusion/fission (DRP1,FIS1,OPA1) were determined using quantitative reverse transcription PCR (qRT-PCR). Gene expression was compared between treatments (One Way ANOVA with repeated measures, and Dunnet’s post hoc) as well as No AMD and AMD donors (Student's t-test).
Expression of AMPK decreased in both AMD (p<0.0001) and No AMD (p<0.0001) donors at both 6 and 24 hours peroxide treatment. Peroxide treatment increased expression of VDAC, FIS1 and TFAM in both AMD (p=0.004,0.007,0.024) and No AMD donors (p<0.0001,<0.0001,p=0.002). AMD donors had higher expression of AMPK (p=0.024) and PARKIN (p=0.056) at 6 hours post peroxide relative to No AMD donors. Additionally, TFAM had significantly higher expression in AMD donors (p=0.042) at basal levels.
Genes that play a role in mitochondrial biogenesis, autophagy, and fusion/fission have altered expression in the presence of increased oxidative stress. Importantly, we see significant differences in how primary RPE from AMD and non AMD donors respond to oxidative stress. These differences in the cell’s ability to regulate mitochondrial homeostasis in the presence of increased oxidative stress may contribute to AMD progression.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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