Fig. 1

Expression of HK1 and HK2 in ovarian cancer and effects of HK1 and HK2 deletions. a Immunoblot of HK1 and HK2 in ovarian cancer cell lines. b, c The expression of HK1 and HK2 in primary ovarian cancer cells that were isolated and cultivated from high-grade serous ovarian carcinomas (HGSOC) of patients. The data are shown at the mRNA transcripts (b) and the protein (c). b mRNA: Pearson’s r = 0.747, p < 0.001, n = 26; protein: Fisher’s z transformed Pearson correlations of individual blots, fixed effects r = 0.938, z = 6.20, p < 0.001, random effects r = 0.936, z = 4.48, p < 0.001, n = 22. c Representative blot of hexokinases in primary HGSOC cancer cells. d The expression of HK1 and HK2 as analyzed by immunoblot of fibroblasts isolated from matched normal adjacent ovarian tissue of the examined ovarian cancer patients (Pearson’s r = 0.866, p = 0.005, n = 8). e, f Expression of HK1 and HK2 in HK1⁻ and HK2⁻ TOV-112D cells at the protein (e) and mRNA (f) levels. g The differences in hexokinase activity in the HK1⁻ or HK2⁻ TOV-112D cells. The differences in hexokinase activity were tested by one-way ANOVA (F = 6.0, df = 8, p < 0.001); Student–Newman–Keuls post-test p < 0.05 are indicated by asterisks. h Changes in extracellular acidification rates (ECAR) in the HK1⁻ or HK2⁻ TOV-112D cells. Asterisks indicate p < 0.05 in one-way ANOVA followed by Student–Newman–Keuls post-tests. i Changes in the intracellular ATP levels in the HK1⁻ or HK2⁻ TOV-112D cells. The differences in hexokinase activity were tested by one-way ANOVA (F = 33.1, df = 8, p = 0.04); Student–Newman–Keuls post-test p < 0.05 are indicated by asterisks. j Doubling time of the HK1⁻ or HK2⁻ TOV-112D cells in DMEM with altered glucose (5.5 mM or 0.4 mM), glutamine (2 mM or 0 mM), and pyruvate (1.2 mM or 0 mM) concentrations. Asterisks indicate p < 0.05 in one-way ANOVA followed by Student–Newman–Keuls post-tests. k Mitochondrial membrane potential visualized as the ratio of JC-1 aggregates (red fluorescence) towards the sum of red and green fluorescence in the HK1⁻ or HK2⁻ TOV-112D cells; cells that were incubated with 100 μM trifluoromethoxy carbonylcyanide phenylhydrazone (FCCP) were used as a depolarization control. Asterisks indicate p < 0.05 in one-way ANOVA followed by Student–Newman–Keuls post-tests. l Reactive oxygen species visualized by CM-H₂DCFDA in the HK1⁻ or HK2⁻ TOV-112D cells. Data are shown relative to the control TOV-112D cells; the CM-H₂DCFDA fluorescence following the incubation with 100 μM H₂O₂ is indicated as a positive control. Asterisks indicate p < 0.05 in one-way ANOVA followed by Student–Newman–Keuls post-tests. m Mitochodrial mass, superoxide, and superoxide relative to the mitochondrial mass in the HK1⁻ or HK2⁻ TOV-112D cells. Data are shown relative to the control TOV-112D cells (mitochondrial mass, superoxide). Asterisks indicate p < 0.05 in one-way ANOVA followed by Student–Newman–Keuls post-tests