Figure 3.

IL-6 and MCP-1 production with HBHA stimulation might be associated with CHOP expression in RAW 264.7 cells. (a and b) RAW264.7 cells were preincubated for 30 min with the IKK2 inhibitor IKK2 IV and then stimulated with HBHA (10 μg/ml) for 24 h. (a) Effects of IKK2 inhibition on IL-6 and MCP-1 in supernatants 24 h after HBHA stimulation. Measurement of IL-6 and MCP-1 levels from the supernatant was performed using a CBA flow cytometric assay. LPS (100 ng/ml, 24 h) was used as a positive control. (b) The cells were harvested 24 h after HBHA stimulation, and each total cell lysate was analysed by western blotting for CHOP. (c) RAW264.7 cells were preincubated for 1 h with the JNK inhibitor SP600125 and then stimulated with HBHA (10 μg/ml) for 24 h. The cell lysates were analysed by western blotting with antibodies for CHOP, GRP78, p-eIF2α, and β-actin. Cell lysate treated with 2 μg/ml TM for 6 h was used as a positive control to induce ER stress. Data are representative of at least three independent experiments with similar results. A significant difference was observed in HBHA-stimulated cells compared with unstimulated control cells (***P<0.001)