Figure 4 | Cell Death & Disease

Figure 4

From: Glucose restriction induces cell death in parental but not in homeodomain-interacting protein kinase 2-depleted RKO colon cancer cells: molecular mechanisms and implications for tumor therapy

Figure 4

Targeting glycolysis by 2-DG or siGlut-1 does not re-establish glucose-starving siHIPK2 cell death. (a) HIPK2+/+ and siHIPK2 subconfluent cells were seeded and the day after untreated (−) or treated with 2-DG at 4, 8 and 12 mM. Forty-eight hours later, trypan blue staining was performed. The result is the mean of three independent experiments performed in triplicate ±S.D. (b) HIPK2+/+ and siHIPK2 were treated with 12 mM 2-DG for 48 h, after which live cell images were taken, or fresh medium was replaced for 48 h (+rec) before imaging. A representative result of three independent experiments is shown. (c) HIPK2+/+ and siHIPK2 subconfluent cells were seeded and the day after untreated (−) or treated with 12 mM 2-DG for 48 h before fresh medium was replaced for 48 h (+rec). Trypan blue staining was performed to assess live proliferating cells (upper panel). The result is the mean of three independent experiments performed in triplicate ±S.D. Below is shown the western immunoblotting of cyclin B1. L.c, loading control. (d) HIPK2+/+ and siHIPK2 cells were assayed for RT-PCR analysis of HIPK2 and Glut-1 mRNA expression or Glut-1 protein levels. β-actin was used as loading control for both RT-PCR and western immunoblotting analyses. (e) siHIPK2 cells were transfected with siRNA for Glut-1 interference (Glut-1 mRNA is shown in the inset) and 36 h after transfection cultured in glucose-free mediun (glu stv). Forty-eight hours after starvation, trypan blue staining was performed. The result is the mean of two independent experiments performed in duplicate ±S.D

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