Fig. 3

Effects of GKN2 on NF-κB and JNK signaling. a-b Western blotting detection of pro-apoptotic protein in GC cell lines after H₂O₂ (300 uM) treatment for 6 h. c The Phospho Explorer Antibody Array was used to screen total cell lysates from untreated and H₂O₂-treated cell cultures. Data show fold change of indicated phosphoproteins upon H₂O₂ (300 uM) treatment for 6 h after normalization to total protein expression. d & f Western blotting detection of the protein expression and phosphorylation in GC cell lines after H₂O₂ (300 uM) treatment for 6 h. (e) Comparison of the proliferation of GC cell lines after H₂O₂ (300 uM) treatment for 6 h with or without inhibition of NF-κB by 10 uM bay11–7082. Cells were pretreated with the inhibitors for 2 h and maintained in culture. GKN2 silencing MGC cells (IC50 676.7 ± 27.9 μM) and SGC cells (IC50 644.8 ± 30.0 μM) with inhibition of NF-κB were significantly more sensitive to H₂O₂ compared to GKN2 silencing MGC cells (IC50 807.4 ± 45.4 μM, P < 0.05) and SGC cells (IC50 836.8 ± 30.5 μM, P < 0.01), respectively. (g) Comparison of the proliferation of GC cell lines after H₂O₂ (300 uM) treatment for 6 h with or without inhibition of JNK by 10 uM SP600125. Cells were pretreated with the inhibitors for 2 h and maintained in culture. GKN2 overexpressing MGC cells (IC50 603.4 ± 23.2 μM) and SGC cells (IC50 611.3 ± 25.5 μM) with inhibition of JNK were significantly less sensitive to H₂O₂ compared to GKN2 overexpressing MGC cells (IC50 528.4 ± 29.5 μM, P < 0.05) and SGC cells (IC50 539.2 ± 31.6 μM, P < 0.05), respectively. Data are presented as mean ± SD from three independent experiments with each running in triplicate. (n = 3, **p < 0.01, ***p < 0.001)