Figure 6.
Other signaling pathways are also involved in active or inactive arazyme activation of GM-CSF-derived bone marrow-derived dendritic cells (BMDCs). (A) BMDCs were obtained from C57BL/6 mice, incubated in V bottom plates (1 × 105 cells/well), pretreated for 1 h with SP600125 (JNK inhibitor, 50 µM), SB203580 (P38 inhibitor, 20 µM) or PD98059 (MAPK/ERK inhibitor, 100 µM) and stimulated with lipopolysaccharide (LPS) (200 ng/mL) or with indicated concentrations of active or heat-inactivated arazyme for 48 h. IL-12 was quantified in culture supernatants by ELISA. (B) BMDCs were treated for 2 h with LPS (200 ng/mL) or heat-inactivated arazyme (10 μg/mL) and the levels of p-STAT1, p-STAT5, p-JNK, p-MAPK/ERK, P38, and p-P38 in cell lysates were analyzed by western blotting. (C) Densitometric quantification of bands in (B). Medium, unstimulated cells; Control, stimulated BMDCs in the absence of inhibitors. **p < 0.01 and ****p < 0.0001, analyzed by one-way ANOVA with Dunnett's multiple comparisons.