Inhibition of IκB kinase and IκB phosphorylation by 15-deoxy-Δ12,14-prostaglandin J2 in activated murine macrophages

Activation of the macrophage cell line RAW 264.7 with lipopolysaccharide (LPS) and gamma interferon (IFN-gamma) induces the expression of gene products involved in host defense, among them type 2 nitric oxide synthase. Treatment of cells with 15-deoxy-Delta(12,14)-prostaglandin J(2) (15dPGJ(2)) inhibited the LPS- and IFN-gamma-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 gamma ligand, Incubation of activated macrophages with 15dPGJ(2) inhibited the degradation of I kappa B alpha and I kappa B beta and increased their levels in the nuclei. NF-kappa B activity, as well as the transcription of NF-kappa B 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 I kappa B phosphorylation showed an inhibition of I kappa B kinase by 15dPGJ(2) in cells treated with LPS and IFN-gamma, resulting in an impaired phosphorylation of I kappa B alpha, at least in the serine 32 residue required for targeting and degradation of this protein. Incubation of partially purified activated I kappa B kinase with 2 mu M 15dPGJ(2) reduced by 83% the phosphorylation in serine 32 of I kappa B alpha, suggesting that this prostaglandin exerts direct inhibitory effects on the activity of the I kappa B kinase complex. These results show rapid actions of 15dPGJ(2), independent of peroxisomal proliferator receptor gamma activation, in macrophages challenged,vith low doses of LPS and IFN-gamma.