Biochanin A impedes STAT3 activation by upregulating p38δ MAPK phosphorylation in IL-6-stimulated macrophages.
Inflamm Res. 2020 Nov ;69(11):1143-1156. Epub 2020 Aug 27. PMID: 32852592
OBJECTIVE: IL-6-induced STAT3 activation is associated with various chronic inflammatory diseases. In this study, we investigated the anti-STAT3 mechanism of the dietary polyphenol, biochanin A (BCA), in IL-6-treated macrophages.
METHODS: The effect of BCA on STAT3 and p38 MAPK was analyzed by immunoblot. The localization of both these transcription factors was determined by immunofluorescence and fractionation studies. The impact on DNA-binding activity of STAT3 was studied by luciferase assay. To understand which of the isoforms of p38 MAPK was responsible for BCA-mediated regulation of STAT3, overexpression of the proteins, site-directed mutagenesis, pull-down assays and computational analysis were performed. Finally, adhesion-migration assays and semi-quantitative PCR were employed to understand the biological effects of BCA-mediated regulation of STAT3.
RESULTS: BCA prevented STAT3 phosphorylation (Tyr) and increased p38 MAPK phosphorylation (Thr/Tyr) in IL-6-stimulated differentiated macrophages. This opposing modulatory effect of BCA was not observed in cells treated with other stress-inducing stimuli that activate p38 MAPK. BCA abrogated IL-6-induced nuclear translocation of phospho-STAT3 and its transcriptional activity, while increasing the cellular abundance of phospho-p38 MAPK. BCA-induced phosphorylation of p38δ, but not α, β, or γ was responsible for impeding IL-6-induced STAT3 phosphorylation. Interestingly, interaction with phospho-p38δ masked the Tyrresidue of STAT3, preventing its phosphorylation. BCA significantly reduced STAT3-dependent expression of icam-1 and mcp-1 diminishing IL-6-mediated monocyte adhesion and migration.
CONCLUSION: This differential regulation of STAT3 and p38 MAPK in macrophages establishes a novel anti-inflammatory mechanism of BCA which could be important for the prevention of IL-6-associated chronic inflammatory diseases.