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. 2017 Jun 30;7:4411. doi: 10.1038/s41598-017-03619-7

Figure 4.

Figure 4

The effect of β6 integrin-deficiency on inflammatory cytokine expression. (A) SiRNA knockdown of β6 integrin in the GECs after a 48-h transfection relative to β-actin detected by Western blotting. SiRNA that is not homologous to any human gene (C) was used as a control. (B) The effect of β6 integrin knockdown on IL1B expression in biofilm-treated (60 µg protein/ml; 32 h) and non-treated GECs determined by RT-qPCR. Mean ± SEM is presented, n = 3. (C) The effect of TGF-β1 signaling inhibitor SB431542 (5 µM) on ITGB6 and IL1B expression in biofilm-treated (60 µg protein/ml; 32 h) and non-treated GECs. Mean ± SEM, n = 3. (D) GECs were treated with TGF-β1 (2 ng/ml), heated biofilm extract (60 µg protein/ml) or a combination of both for 32 h and analyzed for ITGB6 expression by RT-qPCR. Mean ± SEM, n = 3. (E) GECs were treated with TGF-β1 (2 ng/ml) for 0–120 min and analyzed for Smad2 phosphorylation (activation) relative to total Smad2 by Westerm blotting. (F and G) GECs were treated with TGF-β1 (2 ng/ml), heated biofilm extract (60 µg protein/ml) or a combination of both for 30 min and analyzed for Smad2 phosphorylation relative to total Smad2 by Westerm blotting. The ratio of phosphorylated Smad2 to total Smad2 was determined from triplicated experiments. *p < 0.05; **p < 0.01; ***p < 0.001.