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Proteomic analysis of retinal pigment epithelium cells after exposure to UVA radiation

Authors :
Jiunn-Liang Chen
Chun-Tzu Hung
Joseph Jordan Keller
Hsien-Chung Lin
Yu-Jen Wu
Source :
BMC Ophthalmology, Vol 19, Iss 1, Pp 1-13 (2019)
Publication Year :
2019
Publisher :
BMC, 2019.

Abstract

Abstract Background Age-related macular degeneration (AMD) is the primary cause of blindness and severe vision loss in developed countries and is responsible for 8.7% of blindness globally. Ultraviolet radiation can induce DNA breakdown, produce reactive oxygen species, and has been implicated as a risk factor for AMD. This study investigated the effects of UVA radiation on Human retinal pigment epithelial cell (ARPE-19) growth and protein expression. Methods ARPE-19 cells were irradiated with a UVA lamp at different doses (5, 10, 20, 30 and 40 J/cm2) from 10 cm. Cell viability was determined by MTT assay. Visual inspection was first achieved with inverted light microscopy and then the DeadEnd™ Fluorometric TUNEL System was used to observe nuclear DNA fragmentation. Flow cytometry based-Annexin V-FITC/PI double-staining was used to further quantify cellular viability. Mitochondrial membrane potential was assessed with JC-1 staining. 2D electrophoresis maps of exposed cells were compared to nonexposed cells and gel images analyzed with PDQuest 2-D Analysis Software. Spots with greater than a 1.5-fold difference were selected for LC-MS/MS analysis and some confirmed by western blot. We further investigated whether caspase activation, apoptotic-related mitochondrial proteins, and regulators of ER stress sensors were involved in UVA-induced apoptosis. Results We detected 29 differentially expressed proteins (9 up-regulated and 20 down-regulated) in the exposed cells. Some of these proteins such as CALR, GRP78, NPM, Hsp27, PDI, ATP synthase subunit alpha, PRDX1, and GAPDH are associated with anti-proliferation, induction of apoptosis, and oxidative-stress protection. We also detected altered protein expression levels among caspases (caspase 3 and 9) and in the mitochondrial (cytosolic cytochrome C, AIF, Mcl-1, Bcl-2, Bcl-xl, Bax, Bad, and p-Bad) and ER stress-related (p-PERK, p-eIF2α, ATF4 and CHOP) apoptotic pathways. Conclusions UVA irradiation suppressed the proliferation of ARPE-19 cells in a dose-dependent manner, caused quantitative loses in transmembrane potential (ΔΨm), and induced both early and late apoptosis.

Details

Language :
English
ISSN :
14712415
Volume :
19
Issue :
1
Database :
Directory of Open Access Journals
Journal :
BMC Ophthalmology
Publication Type :
Academic Journal
Accession number :
edsdoj.349183aaa8f34bc8b1df3236edcb6b97
Document Type :
article
Full Text :
https://doi.org/10.1186/s12886-019-1151-9