Protective Effects And Its Mechanism Of Sodium Aescinate Against H₂O₂-Induced Oxidative Injury In Human Retinal Pigment Epithelial Cells

OBJECTIVE:The present study is to observe the effect of sodium aescinate（SA）on H₂O₂-induced human retinal pigment epithelium（ARPE-19）cells oxidative injury and to investigate the mechanism of the effect.To explore whether SA has the potential as an antioxidant at the cellular level to provide a new idea for clinical treatment of age-related macular degeneration（AMD）.METHODS:In this study,the oxidative damaged model of ARPE-19 cells was established by H₂O₂,and the effective range of SA was also determined.Then SA and H₂O₂ were added to the cells for co-culture,and the cell viability and apoptosis rate were detected to see if SA can protect H₂O₂ injured retinal cells.Real-time quantitative polymerase chain reaction（RT-qPCR）was used to detect the mRNA expression of antioxidant genes including NQO1 and HO-1 in the downstream of Nrf2/ARE pathway and apoptosis-related genes including bcl-2 and Bax.RESULTS:H₂O₂ could induce oxidative damage and apoptosis in ARPE-19 cells,which was marked by a significant increase in intracellular reactive oxygen species（ROS）,a significant decrease in superoxide dismutase（SOD）,and an increase in early apoptotic rate.When the cells were pre-incubated with SA and then treated with H₂O₂,the intracellular ROS decreased,the SOD increased,and the early apoptosis of the cells decreased.Compared with the purely H₂O₂ treated cells,the mRNA expression of HO-1 significantly increased in SA pre-cultured cells in a concentration-dependent manner;The mRNA expression of NQO1 increased at 25μmol/L and 50μmol/L concentrations of SA,but there was no significant change at 5μmol/L concentrations of SA.In addition,treating ARPE-19cells with H₂O₂ for 24h could reduce the mRNA expression of Bcl-2 by about 5-fold,while pre-culture of SA increased it.On the contrary,the mRNA expression of Bax in the H₂O₂ treatment group increased significantly,but the increase trend could be alleviated by SA pre-culture.Conclusion:This study demonstrates that H₂O₂ can cause oxidative damage in ARPE-19 cells,mimic the RPE environment when AMD occurs,induce the cell vitality,promote cell apoptosis and increase the production of ROS.In ARPE-19 cells,SA can increase the mRNA expression of antioxidant genes including NQO1and HO-1 in the downstream of Nrf2/ARE pathway and inhibit the decrease of SOD activity induced by H₂O₂,indicating a jointly antioxidant effect.This study also demonstrates that SA can inhibit apoptosis in ARPE-19 cells by adjusting the ratio of bcl-2/Bax,and reduce cell damage.In conclusion,sodium aescinate is expected to be an effective antioxidant,providing a new idea for clinical prevention and treatment of AMD.