Figure 2. The growth of HCC827 and HCC827ER cells depends on glutamine.
(A) HCC827 (1 × 105 cells per well) and HCC827ER cells (1 × 105 cells per well) were cultured in RPMI 1640 medium with glutamine or without for 6 days. The cell numbers were counted every day. The number of cells cultured in medium with glutamine (300 mg/L L-Glutamine) was calculated as control. Data are shown as means ± S.D. of three experiments. ***P < 0.001. (B) HCC827 cells were transfected with control siRNA or siRNA1 and siRNA2 targeting GAC. Cells were grown for the indicated number days and counted. Untreated cells were used as control. Data represent the average of three independent experiments (mean ± SD). *P < 0.05, **P < 0.01. (C) HCC827ER cells were transfected with control siRNA or siRNA1 and siRNA2 targeting GAC. Cells were grown for the indicated number days and counted. Untreated cells were used as control. Data represent the average of three independent experiments (mean ± SD). **P < 0.01, ***P < 0.001. (D) The efficiencies of the siRNA knocking down of GAC in both HCC827 and HCC827ER cells were determined by Western blot (top panel). The rescue efficiencies of GAC in HCC827 and HCC827ER cells were determined after the transfection of pCDNA 3.1-V5-GAC (bottom panel). (E) HCC827 cells were transfected with control siRNA or siRNA1 and siRNA2 targeting GAC. After 24 hours, V5-GAC plasmid was transfected into HCC827 cells. The cell numbers were counted on the indicated days. Data represent the average of three independent experiments (mean ± SD). *P < 0.05, **P < 0.01. (F) HCC827ER cells were transfected with control siRNA or siRNA1 and siRNA2 targeting GAC. After 24 hours, V5-GAC plasmid was transfected into HCC827ER cells. The cell numbers were counted on the indicated days. Data represent the average of three independent experiments (mean ± SD). ***P < 0.001.