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Figure 3 | BMC Cancer

Figure 3

From: Chemoresistance is associated with increased cytoprotective autophagy and diminished apoptosis in bladder cancer cells treated with the BH3 mimetic (−)-Gossypol (AT-101)

Figure 3

Pharmacological inhibition of autophagy potentiates (−)-gossypol-induced cell death in chemoresistant bladder cancer lines. Bladder cancer cells 5637, 5637rCDDP1000 and 5637rGEMCI20 were treated with 15 μM of pan Bcl-2 inhibitor (−)-gossypol with or without the autophagy inhibitor 3-MA (2 mM) for 48 h and total cell death and apoptotic cell death were quantified by flow cytometry of Annexin V/PI double staining (A and B). *, P < 0.05, compared with the control. #, P < 0.05, compared with cultures not cotreated with 3-MA. n.s., not significant. The inhibitor of autophagic flux Bafilomycin A1 potentiates cell death induced by (−)-gossypol in 5637rCDDP1000 cells (C). Bladder cancer cells 5637 and 5637rCDDP1000 were treated with 15 μM (−)-gossypol with or without Bafilomycin A1 (10 nM) for 48 h and total cell death and apoptotic cell death was quantified by flow cytometry of Annexin V/PI double staining. *, P < 0.05, compared with the control. #, P < 0.05, compared with cultures not co-treated with Bafilomycin A1. n.s., not significant. Stable knockdown of ATG5 in 5637, 5637rGEMCI20 and 5637rCDDP1000 cells as shown by Western blot analysis with an ATG5 antibody (D). Whole-cell lysates of 5637, 5637rGEMCI20 and 5637rCDDP1000 control cells transfected with empty vector (Ø-vec) and ATG5-KD cells were probed and analyzed with an antibody against ATG5. GAPDH served as a loading control.

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