Inhibition of nuclear factor-κb activation in mouse macrophages and the RAW 264.7 cell line by a synthetic adenyl carbocyclic nucleoside

Abstract

Adenyl carbocyclic nucleosides have potent anti-inflammatory effects on a number of cell types. Notable in this regard is their ability to inhibit the production of tumor necrosis factor-α (TNF-α) by mouse macrophages that have been activated with bacterial lipopolysaccharide (LPS). Because the transcriptional activation of the mouse TNF-α gene is highly dependent on κB enhancers, the present study determined whether the synthetic carbocyclic nucleoside 9-[(1 S,3 R)- cis-cyclopentan-3-ol]adenine (cPA) inhibited LPS-induced nuclear factor-κB (NF-κB) activation in these cells. Stimulation of either mouse peritoneal macrophages or RAW 264.7 macrophage-like cells with LPS led to the appearance of four distinct κB-binding nucleoprotein complexes detected by gel mobility shift assays. Cells treated with 100 μM cPA showed significantly reduced levels of NF-κB activation as evidenced by measurements of nucleoprotein κB-binding activity and diminished κB-dependent transcriptional activation. However, both the LPS-induced degradation of the cytoplasmic NF-κB inhibitor IκBα and the nuclear translocation of the NF-κB p50, p65, and c-Rel peptides were unaffected by treatment of the cells with the nucleoside. These findings suggest that certain adenyl carbocyclic nucleosides inhibit the activation of NF-κB/Rel complexes by a novel mechanism that results in an inhibition of their DNA-binding activities, without blocking their dissociation from IκBα or their nuclear translocation.