Idebenone inhibiting growth and metastasis of hepatocellular carcinoma via regulating mitochondrial reactive oxygen species
Objective To investigate the function and mechanism of Idebenone(IDE)in inhibiting hepatocellular carcinoma metastasis by regulating mitochondrial reactive oxygen species(ROS).Methods Cell counting kit 8(CCK8)method was used to detect the cell viability of HepG2 liver cancer cells at different concentrations(incubation for 24 hours at 1,2,4,6,8 and 16 μmol/L IDE)and at different time(incubation with 10 μmol/L IDE for 6,12,24,48 and 72 h).HepG2 cells trea-ted with 10 μmol/L IDE were divided into IDE-R group and IDE-T2 group.HepG2 cells treated with dimethyl sulfoxide(DMSO)were divided into DMSO-R group and DMSO-T2 group.Transwell experiment was used to analyze cell invasion and migration in IDE-T2 and DMSO-T2 groups.2,7-Dichlorodihydrofluorescein diacetate(DCFH-DA)and MitoSOX probes were used to detect overall ROS and mitochondrial ROS levels in IDE-R and DMSO-R groups.HepG2 hepatocellu-lar carcinoma orthotopic transplantation mouse models were divided into IDE-L group and DMSO-L group(8 mice in each group),and the lung metastasis ability of the tumors was detected by histopathology.HepG2 cells were used to construct subcutaneous tumor model of hepatocellular carcinoma,which was divided into IDE-M group and DMSO-M group,8 cells in each group,and the tumor growth size was detected.Results IDE could effectively inhibit the HepG2 cell viability.The cell inhibition rates at concentrations of 1,2,4,6,8 and 16 μmol/L IDE treatment for 24 hours were(3.33±0.88)％,(9.67±1.20)％,(16.31±1.15)％,(37.36±2.41)％,(61.42±1.76)％and(86.71±2.02)％,respectively.The 50％inhibitory concentration(IC50)at 24 hours was 8.09±0.28 μmol/L.The cell inhibition rates of HepG2 cells treated with 10 μmol/L IDE for 6,12,24,48 and 72 hours were(11.04±1.52)％,(31.35±1.76)％,(51.37±1.53)％,(75.38± 2.37)％and(90.56±1.93)％,respectively.Intracellular ROS detection revealed that the mitochondrial ROS levels in the IDE-R group(12 017.34±1 647.23)were lower than those in the DMSO-R group(57 495.67±2 512.15),with a statis-tically significant difference,t=-26.30,P＜0.001.The total ROS level in the IDE-R group(11 411.57±1 119.49)was lower than that in the DMSO-R group(51 132.26±2 018.33),with a statistically significant difference,t=-29.81,P=0.009.The Transwell experiment results showed that compared to the DMSO-T2 group[(100.00±2.08)％],the IDE-T2 group[(27.35±3.14)％]had a decrease in cell invasion ability,and the IDE-T2 group[(22.19±2.54)％]had a lower migration ability of liver cancer cells compared to the DMSO-T2 group[(100.00±3.13)％].In vivo tumor metastasis a-nalysis showed that compared with the DMSO-L group(73.12±3.65),the IDE-L group(23.67±2.39)had a signifi-cantly reduced number of lung metastases in nude mice,with a statistically significant difference,t=18.56,P＜0.001.Subcutaneous tumor experiments showed that the tumor volume in the IDE-M group was smaller than that in the DMSO-M group.Conclusion The antioxidant idebenone can inhibit hepatocellular carcinoma metastasis by scavenging mitochon-drial ROS and thereby.
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