Figure 4

TLR-5 mediated IL-17C induce pBD-2, claudin-1 and -2 expression in IPEC-J2 cells. IPEC-J2 monolayers were pre-incubated with 0.5 μM TH1020 or the equivalent amount of its solvent DMSO for 2 h. Then, the monolayers were inoculated with HB101 or C83901 at MOI 100 for 1 h, washed and further incubated for 23 h. Non-treated cells were supplemented with PBS or IL-17C (20 or 40 ng/mL). A pBD-2, claudin-1 and -2 mRNA expression in the IPEC-J2 monolayers was assessed by qPCR. The mRNA expression level was normalized to the reference genes and then to the control group. Different letters indicate significant differences (p < 0.05). Data are presented as the mean ± SD (n = 5 per group). B pBD-2, claudin-1 and -2 protein were detected by Western blotting. IPEC-J2 monolayers were pre-incubated with TH1020 (0.5 μM) or the equivalent amount of its solvent DMSO for 2 h. Then, the monolayers were inoculated with HB101 and C83901 at MOI 100 for 1 h, washed and further incubated for 23 h. Non-treated cells were supplemented 20 ng/mL IL-17C. C Cellular localization of claudin-1 and -2 in IPEC-J2 cells 24 h post PBS (left panel) and C83901 (right pane, MOI 100) stimulation. The cells were stained with anti-claudin-1 (Texas-Red, Red), anti-claudin-2 (FITC, green). The nuclei were counterstained with Hoechst and representative confocal images are shown (n = 3). Scale bar = 10 μm.