Magnolol suppresses NF-κB activation and NF-κB regulated gene expression through inhibition of IkappaB kinase activation

Abstract

The mis-regulation of nuclear factor-kappa B (NF-κB) signal pathway is involved in a variety of inflammatory diseases that leds to the production of inflammatory mediators. Our studies using human U937 promonocytes cells suggested that magnolol, a low molecular weight lignan isolated from the medicinal plant Magnolia officinalis, differentially down-regulated the pharmacologically induced expression of NF-κB-regulated inflammatory gene products MMP-9, IL-8, MCP-1, MIP-1α, TNF-α. Pre-treatment of magnolol blocked TNF-α-induced NF-κB activation in different cell types as evidenced by EMSA. Magnolol did not directly affect the binding of p65/p50 heterodimer to DNA. Immunoblot analysis demonstrated that magnolol inhibited the TNF-α-stimulated phosphorylation and degradation of the cytosolic NF-κB inhibitor IκBα and the effects were dose-dependent. Mechanistically, a non-radioactive IκB kinases (IKK) assay using immunoprecipitated IKKs protein demonstrated that magnolol inhibited both intrinsic and TNF-α-stimulated IKK activity, thus suggesting a critical role of magnolol in abrogating the phosphorylation and degradation of IκBα. The involvement of IKK was further verified in a HeLa cell NF-κB-dependent luciferase reporter system. In this system magnolol suppressed luciferase expression stimulated by TNF-α and by the transient transfection and expression of NIK (NF-κB-inducing kinase), wild type IKKβ, constitutively active IKKα and IKKβ, or the p65 subunit. Magnolol was also found to inhibit the nuclear translocation and phosphorylation of p65 subunit of NF-κB. In line with the observation that NF-κB activation may up-regulate anti-apoptotic genes, it was shown in U937 cells that magnolol enhanced TNF-α-induced apoptotic cell death. Our results suggest that magnolol or its derivatives may have potential anti-inflammatory actions through IKK inactivation.