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NARAE HWANG, Je Moon Woo, Su Wol Chung; Autophagy confers resistance against endoplasmic reticulum stress-induced cell death in human retinal pigment epithelial cells. Invest. Ophthalmol. Vis. Sci. 2018;59(9):2478.
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© ARVO (1962-2015); The Authors (2016-present)
To investigate the protective role of autophagy on retinal pigment epithelial (RPE) cells during endoplasmic reticulum (ER) stress-induced cell death and its functional mechanisms in vitro.
In human RPE primary cells and ARPE-19 cells, we analyzed autophagy induction in the response to ER stress condition using real-time reverse transcription-polymerase chain reaction (qRT-PCR) and western blotting analysis for 48 and 72 hr. 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) assay was used to examine for the cell viability of ER stress-induced cell death in the presence or absence with chloroquine (CQ) as autophagy inhibitor. ER stress specific inhibitor such as 4-phenylbutyrate (4-PBA) and Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitor as diphenyleneiodonium (DPI) and scavenger of reactive oxygen species (ROS) as N-acetyl-L-cysteine (NAC) and mito-TEMPO were evaluated for determining the transcriptional and translational autophagy level of ER stress in human RPE cells.
Human RPE cells after treatment with tunicamycin as ER stress inducers were markedly elevated with the transcriptional and translational level of autophagy genes as Atg5 and LC3B, but not beclin-1. The inhibition of ER stress by 4-PBA were significantly attenuated by the increased level of autophagy induction such as Atg5, LC3B and p62 by tunicamycin. The inhibitor of ROS production as DPI, NAC, and mito-TEMPO markedly decreased ER stress-induced cell death in primary RPE cells. A significant decrease in the levels of autophagy by chloroquine was evaluated for the increased cell mortality and the pro-survival role of ER stress-induced autophagy.
These data indicate that the induction of autophagy may attenuate acquisition of ER stress-induced cell death through the ROS production by NADPH oxidase, suggesting that autophagy activation may be used as a potential therapeutic strategy for age-related macular degeneration.
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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