Inhibition of IκB Kinase and IκB Phosphorylation by 15-Deoxy-Δ(12,14)-Prostaglandin J(2) in Activated Murine Macrophages

Activation of the macrophage cell line RAW 264.7 with lipopolysaccharide (LPS) and gamma interferon (IFN-γ) induces the expression of gene products involved in host defense, among them type 2 nitric oxide synthase. Treatment of cells with 15-deoxy-Δ(12,14)-prostaglandin J(2) (15dPGJ(2)) inhibited the LPS- and IFN-γ-dependent synthesis of NO, a process that was not antagonized by similar concentrations of prostaglandin J(2), prostaglandin E(2), or rosiglitazone, a peroxisomal proliferator-activated receptor γ ligand. Incubation of activated macrophages with 15dPGJ(2) inhibited the degradation of IκBα and IκBβ and increased their levels in the nuclei. NF-κB activity, as well as the transcription of NF-κB-dependent genes, such as those encoding type 2 nitric oxide synthase and cyclooxygenase 2, was impaired under these conditions. Analysis of the steps leading to IκB phosphorylation showed an inhibition of IκB kinase by 15dPGJ(2) in cells treated with LPS and IFN-γ, resulting in an impaired phosphorylation of IκBα, at least in the serine 32 residue required for targeting and degradation of this protein. Incubation of partially purified activated IκB kinase with 2 μM 15dPGJ(2) reduced by 83% the phosphorylation in serine 32 of IκBα, suggesting that this prostaglandin exerts direct inhibitory effects on the activity of the IκB kinase complex. These results show rapid actions of 15dPGJ(2), independent of peroxisomal proliferator receptor γ activation, in macrophages challenged with low doses of LPS and IFN-γ.