Fig. 3 in Nerolidol inhibits proliferation of leiomyoma cells via reactive oxygen species-induced DNA damage and downregulation of the ATM/ Akt pathway

Fig. 3. Nerolidol treatment led to the generation of intracellular ROS and impaired the mitochondrial membrane potential. (A) ELT3 cells were harvested after trypsin/EDTA treatment and then treated with 20 mM NAC, followed by DCFDA staining for 30 min. The cells were then treated with nerolidol for another additional 30 min, and DCF fluorescence was detected using flow cytometry. H2O2 was used as the positive control. The data were analyzed using the FlowJo software. (B, C) ELT3 cells were harvested and treated with nerolidol for 30 min. The cells were then stained with JC-1 dye for 15 min, followed by flow cytometry analysis. FCCP was used as the positive control. The data were analyzed using the FlowJo software. (B) The gated cells in the graphs indicate the cells with low mitochondrial membrane potential, and the quantitative results are plotted in (C). (D) ELT3 cells were treated with the indicated concentration of nerolidol for 48 h with or without co-treatment with 5 mM NAC. Cell proliferation was determined using the MTT assay. Ethanol (1 %) was used as the vehicle control. The data are presented as relative percentage compared to that of the vehicle control. (E) ELT3 cells were treated with the indicated concentration of nerolidol for 48 h with or without co-treatment with 20 μM ferrostatin-1. Cell proliferation was determined using the MTT assay. Ethanol (1 %) was used as the vehicle control. The data are presented as relative percentage compared to that of the vehicle control. The p-values were calculated using one-way ANOVA with Tukey's multiple post hoc test for each group. *, p <0.05; ***, p <0.001 compared to the vehicle control group (C). ***, p <0.001 compared to the nerolidol single treatment group (D, E).