Fig. 4

CDDP resistance in A2780cis cells is caused by DNA methylation-medicated silencing of GS expression. a RT-PCR analysis of glutamine metabolism enzymes. b Western blotting of glutamine metabolism enzymes. “+“indicates culture medium with normal glutamine concentration, and “−“indicates culture medium without glutamine. Cells were cultured in the absence or presence of glutamine for 48 h. c RT-PCR analysis of GS expression in A2780cis cells. Cells were cultured for 72 h in the absence or presence of 5-Aza-dC (2 μM). d Effects of 5-Aza-dC on CDDP-induced cytotoxicity. A2780cis cells were cultured for 72 h in medium containing CDDP (0 or 10 μM) and 5-Aza-dC (0, 1, 2, or 5 μM). Cell viability was measured using the MTT assay. e Confirmation of GS knockdown in A2780 cells using RT-PCR analysis. f Effects of CDDP knockdown on CDDP-induced cytotoxicity. A2780 cells transfected with control siRNA or GS siRNA were cultured for 72 h in the presence of CDDP (concentrations as indicated). Cell viability was measured using the MTT assay. Data are shown as the mean ± SD of the three independent experiments. The differences were analyzed using the Student’s t-test or one-way ANOVA with Dunnett’s multiple comparison (*p < 0.05, **p < 0.01)