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. 2016 Jan 22;7(5):5157–5175. doi: 10.18632/oncotarget.6986

Figure 8. PI-103 induces cell death independent of necroptosis.

Figure 8

A. RT4 cells were treated with a series of concentrations of PI-103 (PI) alone, or with necroptosis inhibitor necrostatin-1 (Nec) (20uM) for 24 hours. Cell viability was measured via ATP assay kit (Promega) (n = 6 per condition). Data are shown as mean±sd (note that the error bars are not visible due to minor variations). B. HeLa cells were transfected with control and RIP1 siRNA for 24 hours then treated with vehicle or PI (5uM) for 20 hours. Cell death was measured with propidium iodide staining (n = 6 per condition). Data are shown as mean ±sd. *: P < 0.05. Immunoblot was used to verify knockdown efficiency. C. RT4 cells were treated with vehicle, PI (1uM), or artemisinin analogue, artesunate (ART) (25uM) with or without Nec (20uM) for 24 hours. Cell death was measured with propidium iodide staining (n = 6 per condition). ART (25uM) or/and Nec were used as additional controls for necroptosis inhibition. Data are shown as mean ±sd. **: P < 0.01. D. HeLa cells were treated with PI (5uM), zVAD (20uM) and Nec (20uM) for 24 hours. Cell death was measured with propidium iodide staining (n = 3 per condition). Data are shown as mean ±sd. E. RT4 cells were treated with vehicle, PI-103 (PI) (0.5uM), z-VAD (20uM) and nec (20uM) as indicated for 24 hours. Cell viability was measured with MTT assay (n = 6 per condition). Data are shown as mean ±sd. ***: P < 0.001. F. RT4 cells were treated with concentrations of PI as indicated (24 hours). Cell viability was assessed with ATP assay kit (Promega) (n = 6 per condition)). Autophagosome (Apg) numbers were assessed in matching RT4 cells with GFP-LC3 by automated Cellomics microcopy (n = 12 per condition). Data are shown as mean ±sd.