Juan Wu1, Zhen-Ya Gao2, Dong-Mei Cui1, Hong-Hui Li3, Jun-Wen Zeng1. 1. State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou 510060, Guangdong Province, China. 2. Xuchang University, School of Medicine, Xuchang 461000, Henan Province, China. 3. Chengdu Aier Eye Hospital, Chengdu 610000, Sichuan Province, China.
Abstract
AIM: To explore the apoptosis of ARPE-19 cells after the treatment with different doses of all-trans-retinoic acid (ATRA). METHODS: ARPE-19 cells were used in the in-vitro experiment. Flow cytometry assay was employed to evaluate the level of reactive oxygen species (ROS) and apoptosis. The effects of ATRA (concentrations from 2.5 to 20 µmol/L) on the expression of endoplasmic reticulum stress (ERS) markers in vitro were evaluated by Western blot and real-time quantitative polymerase chain reaction (qRT-PCR) assays. The contribution of ROS and ERS-induced apoptosis in vitro was determined by using N-acetyl-L-cysteine (NAC) and Salubrinal, an antagonist of NAC and ERS, respectively. RESULTS: Flow cytometry showed that ATRA significantly increased ARPE-19 cell apoptosis and ROS levels in each group (F=86.39, P<0.001; F=116.839, P<0.001). Western blot and qRT-PCR revealed that levels of CHOP and BIP were elevated in a concentration-dependent pattern after the cells were incubated with ATRA (2.5-20 µmol/L). The upregulation of VEGF-A and CHOP induced by ATRA could be inhibited by NAC (antioxidant) and Salubrinal (ERS inhibitor) in vitro. CONCLUSION: ATRA induces the apoptosis of ARPE-19 cells via activated ROS and ERS signaling pathways. International Journal of Ophthalmology Press.
AIM: To explore the apoptosis of ARPE-19 cells after the treatment with different doses of all-trans-retinoic acid (ATRA). METHODS:ARPE-19 cells were used in the in-vitro experiment. Flow cytometry assay was employed to evaluate the level of reactive oxygen species (ROS) and apoptosis. The effects of ATRA (concentrations from 2.5 to 20 µmol/L) on the expression of endoplasmic reticulum stress (ERS) markers in vitro were evaluated by Western blot and real-time quantitative polymerase chain reaction (qRT-PCR) assays. The contribution of ROS and ERS-induced apoptosis in vitro was determined by using N-acetyl-L-cysteine (NAC) and Salubrinal, an antagonist of NAC and ERS, respectively. RESULTS: Flow cytometry showed that ATRA significantly increased ARPE-19 cell apoptosis and ROS levels in each group (F=86.39, P<0.001; F=116.839, P<0.001). Western blot and qRT-PCR revealed that levels of CHOP and BIP were elevated in a concentration-dependent pattern after the cells were incubated with ATRA (2.5-20 µmol/L). The upregulation of VEGF-A and CHOP induced by ATRA could be inhibited by NAC (antioxidant) and Salubrinal (ERS inhibitor) in vitro. CONCLUSION:ATRA induces the apoptosis of ARPE-19 cells via activated ROS and ERS signaling pathways. International Journal of Ophthalmology Press.
Authors: Julia Leitman; Boaz Barak; Ron Benyair; Marina Shenkman; Uri Ashery; F Ulrich Hartl; Gerardo Z Lederkremer Journal: PLoS One Date: 2014-03-03 Impact factor: 3.240