Intracellular galectin-9 activates inflammatory cytokines in monocytes

Whether galectin-9 plays a role in inflammatory responses remains elusive. The present study was designed to determine the role of intracellular galectin-9 in activation of inflammatory cytokine genes in human monocytes. Galectin-9 expression vector pBKCMV3-G9 was transiently co-transfected into THP-1 monocytic cells along with luciferase reporters carrying gene promoters of IL-1alpha (IL1A), IL-1beta (IL1B) and IFNgamma. Transient transfection studies showed that galectin-9 over-expression activated all three gene promoters, suggesting that intracellular galectin-9 induces inflammatory cytokine genes in monocytes. Galectin-9 over-expression also activated NF-IL6 (C/EBP beta) and AP-1, but not NF-kappaB. In contrast, extracellular galectin-9 is not involved in regulation of inflammatory cytokines. Immunoprecipitation/Western blotting, using anti-galectin-9 Ab and anti-NF-IL6 Ab, showed physical association of intracellular galectin-9 with NF-IL6. RT-PCR confirmed that galectin-9 over-expression increased IL-1alpha and IL-1beta mRNA levels in THP-1 cells. The interaction of galectin-9 with NF-IL6 was enhanced following LPS treatment in THP-1 cells. Intracellular galectin-9 synergized with LPS to activate NF-IL6. Nuclear translocation of galectin-9 was also observed in THP-1 cells treated with LPS. Our results indicate that galectin-9 is a LPS-responsive factor, and further demonstrate that intracellular galectin-9 transactivates inflammatory cytokine genes in monocytes through direct physical interaction with NF-IL6.