Protective effect of coumarin on hydrogen peroxide induced oxidative damage in human ovarian granulosa cells by regulating the Keap1/Nrf2/ARE signaling pathway
10.13699/j.cnki.1001-6821.2024.04.016
- VernacularTitle:香豆素调节Keap1/Nrf2/ARE信号通路对过氧化氢诱导的人卵巢颗粒细胞氧化损伤的保护作用
- Author:
Huan QIAN
1
;
Xiao-Yang SHEN
;
Fang WANG
Author Information
1. 湖州市第一人民医院妇科,浙江湖州 313000
- Keywords:
coumarin;
Kelch like ECH associated protein 1/nuclear factor E2 related factor 2/antioxidant response element signaling pathway;
hydrogen peroxide;
ovarian granulosa cells;
oxidative damage
- From:
The Chinese Journal of Clinical Pharmacology
2024;40(4):554-558
- CountryChina
- Language:Chinese
-
Abstract:
Objective To investigate the improvement effect of coumarin on hydrogen peroxide(H2 O2)induced oxidative damage in human ovarian granulosa cells by regulating the Kelch like ECH associated protein 1(Keap1)/nuclear factor E2 related factor 2(Nrf2)/antioxidant response element(ARE)signaling pathway.Methods The human ovarian granulosa cell line COV434 was grouped into control group(normal culture),model group(0.5 mmol·L-1 H2 O2),coumarin group(0.5 mmol·L-1 H2 O2+320 μmol·L 1 coumarin),suppressor group(0.5 mmol·L-1 H2 O2+10 μmol·L-1 Keap1/Nrf2/ARE pathway inhibitor compound 20c)and combination group(0.5 mmol·L-1 H2O2+320 μmol·L-1coumarin+10 μmol·L-1 compound 20c).Cell counting kit 8(CCK-8)method was applied to detect the optical density(OD)of COV434 cells;2',7'-dichlorodihydrofluorescein diacetate(DCFH-DA)fluorescence probe method was applied to detect intracellular reactive oxygen species(ROS);the levels of superoxide dismutase(SOD)and interleukin-18(IL-18)were detected by enzyme linked immunosorbent assay(ELISA);flow cytometry was applied to detect the apoptosis rate of COV434 cells;Western blot was applied to detect the expression of Keap1,Nrf2.Results OD values of blank group,model group,coumarin group,suppressor group and combination group were 1.53±0.20,0.91±0.10,1.35±0.14,0.56±0.05 and 1.06±0.12;the ROS levels were 1.00±0.00,2.24±0.35,1.18±0.13,3.97±0.58 and 2.09±0.20;the SOD levels were(73.43±9.76),(43.32±5.88),(71.54±8.76),(27.64±3.12)and(52.46±6.45)U·mL-1;the levels of IL-18 were(205.90±20.43),(334.56±35.68),(233.24±24.58),(456.54±47.83)and(301.13±37.64)pg·mL-1;the apoptosis rates were(5.96±0.69)％,(19.62±1.77)％,(8.89±0.97)％,(30.23±3.12)％and(17.65±1.07)％;the Keap1 protein levels were 0.95±0.10,0.66±0.08,0.93±0.09,0.31±0.04 and 0.69±0.07;the Nrf2 protein levels were 1.31±0.12,0.76±0.07,1.25±0.14,0.21±0.02 and 0.90±0.11,respectively.The differences between model group with blank group,between model group with coumarin group,suppressor group,between combination group with coumarin group,were also statistically significant(all P＜0.05).Conclusion Coumarin may improve H2 O2-induced oxidative damage in human ovarian granulosa cells by up-regulating the Keap1/Nrf2/ARE signaling pathway.
