CCL21 Facilitates Chemoresistance and Cancer Stem Cell-Like Properties of Colorectal Cancer Cells through AKT/GSK-3β/Snail Signals

Figure 6

AKT/GSK-3β signaling regulates upregulated P-gp and stability of Snail in HCT116 cells. (a) HCT116 cells were treated with CCL21 for 5 min, 15 min, 30 min, and 1 h, and then several signaling pathway key proteins were detected via western blotting. (b) HCT116 cells were pretreated with or without AG490 (20μM), SB431542 (20 μM), SB203580 (20 μM), PD98059 (20 μM), BAY (10 μM), or LY294002 (20 μM) for 2 h, respectively, and treated with CCL21 (100 ng/mL) for 72 h, and then the cell viability was analyzed by MTT. (c) HCT116 cells were pretreated with or without LY294002 (20 μM) for 2 h, respectively, and treated with CCL21 (100 ng/mL), detecting the mammosphere forming ability. (d) HCT116 cells were pretreated with or without AG490 (20 μM), SB431542 (20 μM), SB203580 (20 μM), PD98059 (20 μM), or LY294002 (20 μM) for 2 h, respectively, and treated with CCL21 (100 ng/mL) for 72 h, and then the expressions of P-gp, Bmi-1, Nanog, and OCT-4 were analyzed via western blotting. (e) HCT116 cells were pretreated with or without LY294002 (20 μM) for 2 h and then treated with or without CCL21 (100 ng/mL) for 72 h, and P-gp, Bmi-1, Nanog, and OCT-4 key protein were analyzed by western blotting. (f) HCT116 cells were pretreated with or without AG490 (20 μM), SB431542 (20 μM), SB203580 (20 μM), PD98059 (20 μM), or LY294002 (20 μM) for 2 h, respectively, and treated with CCL21 (100 ng/mL) for 24 h, and then the expression of Snail was analyzed via western blotting. (g) HCT116 cells were pretreated with or without LY294002 (20 μM) for 2 h and then treated with or without CCL21 (100 ng/mL) for 5 min, and Snail, AKT, and GSK-3β pathway key protein were analyzed by western blotting.

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