IL-1β is involved in activating the BMP signaling pathway in HuH7 cells.
A, HuH7 cells were treated with the supernatant of B. longum-infected THP-1 cells for the indicated times. Cell lysates were analyzed by SDS-PAGE and immunoblotting with antibodies to either phosphorylated SMAD1/5/8 (pSMAD1/5/8) or total SMAD1. Con, control. B, HuH7 cells were treated for 4 h with the supernatant of B. longum-infected THP-1 cells (−) or with the supernatant in the presence of 5 μg/ml of either a neutralizing anti-IL-1β antibody or an irrelevant isotype-matched control antibody. Cell lysates were analyzed by SDS-PAGE and immunoblotting with antibodies to either phosphorylated SMAD1/5/8 (pSMAD1/5/8) or total SMAD1. C, quantitation of Western blots corresponding to A. The intensities of the phosphorylated SMAD1/5/8 and total SMAD1 bands in blots from multiple experiments were determined by Li-Cor imaging, and the ratio was calculated. *, p = 0.004; n = 3–5 samples/group from multiple experiments. D, quantitation of Western blots corresponding to B. The intensities of the phosphorylated SMAD1/5/8 and total SMAD1 bands in blots from multiple experiments were determined by Li-Cor imaging, and the ratio was calculated. *, p = 0.006; n = 3–4 samples/group from multiple experiments.