Figure 3.

Sirt3 amplifies E2-induced metabolic activity and mitochondrial fitness of MCF-7 cells. (A) Metabolic activity measured with MTT assay. Two-way ANOVA revealed significant interaction effect between Sirt3 and treatment on metabolic activity F (2,34) = 7.051, p = 0.003, partial η² = 0.293; *** p < 0.001 MCF-7S3 vs. MCF-7C. In MCF-7C, higher in E2 vs. other groups (^a p < 0.001). In MCF-7S3, higher in E2-treated vs. other groups (^b p < 0.001). Results are shown as mean ± SD (n ≥ 3); (B) Immunoblots of mitochondrial respiration complexes. Two-way ANOVA revealed significant interaction effect between Sirt3 and treatment on SDH-A expression F (2,6) = 10,846, p = 0.010, partial η² = 0.783; *** p < 0.001 MCF-7C vs. MCF-7S3 cells. In MCF-7C, higher SDH-A expression in E2-treated vs. other groups (^a p < 0.001). In MCF-7S3, higher SDH-A expression in E2-treated vs. other groups (^b p < 0.001). Higher UQCRC2 and NDUFA9 protein expression in MCF-7S3 cells as compared with MCF-7C (*** p < 0.001). Results are shown as mean ± SD normalized to the control MCF-7C (n ≥ 3). Amidoblack was used as a loading control and representative immunoblots are shown; (C) Mitochondrial membrane potential (ΔΨm) measured with MitoTracker Deep Red dye using flow cytometry. Two-way ANOVA revealed significant interaction effect between Sirt3 and treatment F (2,12) = 11,019, p = 0,010, partial η² = 0.786; higher ΔΨm in the control (** p = 0.003), E2, and E2 + ICI-treated (*** p < 0.001) MCF-7S3 vs. MCF-7C. In MCF-7S3, higher ΔΨm in E2 vs. the control and E2 + ICI-treated (^a p < 0.001). Results are shown as mean ± S.D (n ≥ 3).