Interleukin 6 (IL-6) plays a central role in host defense and acute inflammatory responses. In this study, we found that there is a complex cross-regulation between NF-κB p65 and β-catenin pathways for IL-6 production. The β-catenin/NF-κB p65 complex that is translocated to the nucleus was combined with primary TCF-4, and then the NF-κB p65 was sequentially bound to the domain of NF-κB p65 on IL-6 promoter. Importantly, the β-catenin is an essential factor in the production of IL-6 via NF-κB pathways. In contrast, impaired β-catenin signaling decreases IL-6 cytokine production for early immune response. Furthermore, the Ctnnb1loxp/loxp/LysMcre+/+ conditional KO mice did not increase the level of IL-6 cytokine in plasma and CD169(+) macrophage by LPS. Additionally, TCF-4 acted as a co-activator of NF-κB p65 in potentiating the production of cytokine IL-6 in macrophage. Thus β-catenin could play an important role in IL-6 cytokine production via the priming effect for NF-κB p65.