Figure 4
Hepatoma-derived HMGB1 contributes to induce M2 macrophage polarization. (a) Cell lysates and conditioned medium from shLuc or shHMGB1 ML-14a cells were collected and the expression of HMGB1 was determined by Western blotting. (b) BMDMs were treated with control or HMGB1-low MCM for 24 h and the ROS production was determined by flow cytometry. (c) BMDMs were treated with control or HMGB1-low MCM for 24 h and the CD206 expression was determined by flow cytometry. (d) BMDMs were treated with control or HMGB1-low MCM for the indicated time and their cell lysates were collected to determine the expression of NF-κB p65, Arg-1, and LC3-I/II by Western blotting. (e) BMDMs were treated with MCM in the presence of isotype or anti-HMGB1 antibodies for 24 h. ROS production was analyzed by flow cytometry. (f) BMDMs were treated with MCM in the presence of isotype or anti-HMGB1 antibodies for 24 h to determine the production of IL-10 by ELISA and CD204 or CD206 by flow cytometry. (g) BMDMs were treated with MCM in the presence of isotype or anti-HMGB1 antibodies for 24 h and their cell lysates were collected to determine the expression of NF-κB p65, Arg-1 and LC3-I/II by Western blotting. *p < 0.05; ** < 0.001; ***p < 0.0001.
