Figure 3. IL-6 signals in colon epithelia and enhances epithelial barrier function via induction of claudin-1 and mucin-2.

(a) IL-6Rα protein in T84 and primary murine epithelial cells was determined by Western blot. (b) T84 colonic epithelial cells were cultured to confluence in the absence or presence of 50 ng/ml recombinant human IL-6. Protein was harvested after 0, 10, and 30 minutes of IL-6 exposure. Western blot confirmed phosphorylation of STAT3 after IL-6 exposure. (c) After 24 hours of IL-6 exposure, RNA from T84 cells was harvested and evaluated by qPCR for SOCS3 expression and normalized to actin. Data are the mean ± SEM fold induction of SOCS3 in IL-6 treated cells compared to untreated cells. An unpaired two-tailed Student’s t-test was used to determine statistical significance. *, P<0.05 (d) T84 cells were cultured on membrane permeable supports in the absence or presence of IL-6. Transepithelial resistance (TER) was recorded daily and shown as the mean ± SEM. An unpaired two-tailed Student’s t-test was performed at each time point to determine statistical significance. **, P<0.01 (e) T84 transwells were evaluated for paracellular flux of FITC-dextran. The rate of flux is shown as the mean ± SEM. An unpaired two-tailed Student’s t-test demonstrated significance. ***, P<0.0001 (f) After 24 hours of IL-6 exposure, RNA from T84 cells was extracted evaluated for CLDN1 expression by qPCR. Data are the mean expression of CLDN1 ± SEM in IL-6 treated cells relative to untreated cells. Statistical analysis using an unpaired two-tailed Student’s t-test revealed significance. *P<0.05 (g) Cellular lysates from unexposed and IL-6 exposed T84 cells at 24 hours were evaluated for claudin-1 protein by Western blot. (h) RNA from T84 cells with and without IL-6 treatment was evaluated for MUC2 expression by qPCR. Data are the mean fold induction of MUC2 ± SEM in treated cells compared to untreated cells. Statistical analysis using an unpaired two-tailed Student’s t-test revealed significance. *P<0.05