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. 2017 Jun 2;8(9):1530–1541. doi: 10.7150/jca.18628

Figure 5.

Figure 5

STAT3 mediates Pim-3-induced stemness in PC cells.A, Expression level of pSTAT3 and total STAT3 were detected by immunoblotting in PANC-1-Pim-3-sh, MIAPaCa-2-Pim-3 and their parent cells as well as M-110 treated PANC-1 and MIAPaCa-2-Pim-3. β-actin was used as internal control. B, Transcriptional activity of STAT3 in PANC-1-Pim-3-sh, MIAPaCa-2-Pim-3 and their parent cells. Cells were co-transfected with a mixture of the STAT3 luciferase reporter and Renilla plasmid. After 48 h, cells were harvested in passive lysis buffer and luciferase activity was assessed using a dual-luciferase assay system. Data are presented as normalized relative luciferase activity (mean ± SD; n = 3, **p < 0.01). C, Expression levels of stemness-associated markers in MIAPaCa-2-Pim-3-STAT3-si and PANC-1-Pim-3-sh-STAT3 cells. MIAPaCa-2-Pim-3 was transiently transfected with STAT3 siRNA and PANC-1-Pim-3-sh was transiently transfected with STAT3 plasmid. Expression level of pSTAT3 and total STAT3 were detected by immunoblotting in MIAPaCa-2-Pim-3-STAT3-si and PANC-1-Pim-3-STAT3 cells. β-actin was used as internal control. D, Expression level of CD24, CD44 and ESA in MIAPaCa-2-Pim-3-STAT3-si and PANC-1-Pim-3-sh-STAT3 were analyzed by flow cytometry. Single positive cells (CD24+, CD44+ and ESA+) and their percentage in total cells were drawn in the rectangle gate. CD24+ESA+ PANC-1-Pim-3-sh cells were shown in the first quadrant as well as their percentage in total cells.