Figure 6
The Akt and GSK3α/β signaling pathways are involved in thrombin-enhanced EGFR activation in NP cells. (A) NP cells were treated with thrombin for indicated periods, cell lysates were separated by SDS-PAGE and immunoblotted with anti-phosphorylated-Akt and anti-phosphorylated-GSK3α/β. (B) NP cells were treated with AG1478 (3 μM) for 30 min followed by treatment with thrombin for 30 min, and the levels of phosphorylated-Akt, Akt, phosphorylated GSK3α/β, and GSK3α/β were determined by western blotting. Similar results were obtained from four independent experiments. (C) NP cells were treated with LY294002 (10 μM), or SB216763 (12.5 μM) for 30 min followed by treatment with thrombin (10 U/mL) for 24 h. The production of CXCL8 was determined by ELISA. Each bar represents the mean ± S.E.M. from at least three independent experiments performed in duplicate. *p < 0.05 compared with the control group. # p < 0.05 compared with the thrombin-treated group. (D) NP cells were treated with U0126 (1 μM), PF573228 (1 μM), PP2 (1 μM), LY294002 (10 μM), or SB216763 (12.5 μM) for 30 min followed by treatment with thrombin for 60 min, and phosphorylated-ERK and ERK2 levels were determined by western blot. Similar results were obtained from three independent experiments.
