Objective: To investigate the precise molecular mechanisms by which baicalein exerts beneficial biochemical activities in RAW264.7 macrophages treated with LPS.
Materials and methods: RAW264.7 cells were cultured in the absence or presence of baicalein together with or without LPS. iNOS and COX-2 expression were measured by western blot and RT-PCR analyses. TNF-α, IL-1β, and IL-6 were determined by using double-antibody sandwich ELISA. Phosphorylations of JAK1 and JAK2, and of STAT1 and STAT3 were detected by western blotting. Nuclear translocation of STAT1 and STAT3 was visualized by confocal microscopy. ROS production was detected by ROS assay.
Results: Baicalein significantly reduced the phosphorylation of STAT1 and STAT3 and the phosphorylation of JAK1 and JAK2, but without affecting MAPKs phosphorylation in LPS-stimulated RAW264.7 cells. Baicalein suppressed the nuclear translocation of STAT1 and STAT3 and inhibited production of iNOS upon LPS-stimulation, resulting in the inhibition of releases of NO and pro-inflammatory cytokines such as IL-1β, IL-6, and TNF-α, in a dose-dependent manner. In addition, we found that baicalein reduced the LPS-induced accumulation of ROS, confirming that baicalein serves as an antioxidant.
Conclusions: Our results suggested that suppressing JAK/STATs activation and interfering with ROS production might contribute to the anti-inflammatory action of baicalein in macrophages.