Effect Of Rutin On The Antioxidation And Antiapoptosis Of Human Melanocytes Induced By Hydrogen Peroxide

PurposeVitiligo is an acquired skin and mucosal depigmentation disease with high morbidity.The apoptosis of melanocytes is the basis of the occurrence and development of vitiligo,and the oxidative stress of melanocytes is the main cause of its apoptosis.Therefore,it is of great significance to inhibit the damage and apoptosis of melanocytes after oxidative stress for vitiligo.Rutin as a natural antioxidant has a strong antioxidant capacity.In this study,we will establish a model of oxidative damage to melanocytes and explore the effect of rutin on the antioxidant and antiapoptosis of melanocytes induced by hydrogen peroxide,so as to provide a new direction for the treatment of vitiligo.Methods1.The effects of H₂O₂and rutin alone and combined on the viability of PIG1 cells and primary human melanocytes were detected by MTS assay.2.Fluorescence microscopy to observe the effect of rutin on ROS in PIG1 cells under H₂O₂induction.3.The effect of rutin on SOD of PIG1 cells induced by H₂O₂was detected by WST-8.4.The effect of rutin on MDA of PIG1 cells induced by H₂O₂was detected by TBA.5.Western Blot was used to detect the expression of Bcl-2,Bax,pro Caspase-3 and Cleaved Caspase-3 in PIG1 cells pretreated with rutin against H₂O₂induction.Results1.The proliferation activity of PIG1 cells was decreased after being treated with 0.6mmol/L H₂O₂for 24h（P<0.05）.After treated with 0.4 mmol/L H₂O₂for 24h,the proliferation activity of primary human melanocytes was decreased（P<0.05）.2.The proliferation activity of PIG1 cells and primary human melanocytes was increased after treatment with 5,10,20,40 and 80μmol/L rutin for 24h（P<0.05）.3.PIG1 cells and primary human melanocytes were pretreated with rutin at concentrations of 10,20,40,80μmol/L for 24 h,and then treated with H₂O₂for 24 h.The proliferation activity of cells in the experimental group was lower than that in the control group（P<0.05）,but higher than that in the hydrogen peroxide group（P<0.05）.It showed that the pretreatment of rutin could inhibit the proliferation activity of the two kinds of melanocytes damaged by H₂O₂.4.PIG1 cells were pretreated with rutin at concentrations of 10,20 and 40μmol/L for24 h,and then treated with H₂O₂for 24 h.The fluorescence intensity of experimental group was lower than that of the hydrogen peroxide group,and gradually decreased with the increase of the concentration of rutin.It showed that ROS level in PIG1 cells decreased gradually with the increase of rutin concentration.5.PIG1 cells were pretreated with rutin at concentrations of 10,20 and 40μmol/L for24 h,and then treated with H₂O₂for 24 h.SOD activity in the experimental group was higher than that in the hydrogen peroxide group,and increased with the increase of rutin concentration（P<0.05）.MDA content in the experimental group was lower than that in the hydrogen peroxide group,and decreased with the increase of the rutin concentration（P<0.05）.6.PIG1 cells were pretreated with rutin at concentrations of 40μmol/L for 24 h,and then treated with H₂O₂for 24 h.The results showed that the expression of Bcl-2 and Pro Caspase-3 in the experimental group was higher than that in the hydrogen peroxide group（P<0.05）,while the expression of Bax and Cleaved caspase-3 was lower than that of hydrogen peroxide group（P<0.05）.Conclusion1.The proliferative activity of PIG1 cells and primary human melanocytes decreased with the increase of H₂O₂concentration.2.In a certain concentration range,rutin has no obvious toxicity to PIG1 cells and primary human melanocytes,and can promote the proliferative activity of these two kinds of melanocytes.3.Rutin can inhibit oxidative damage of PIG1 cells induced by H₂O₂,reduce ROS production,reduce MDA content and activate SOD activity.4.Rupin can inhibit the apoptosis of PIG1 cells induced by H₂O₂,which is related to the reduction of Bax/Bcl-2 ratio and the reduction of Caspase-3 protein cleavage activation.