cGMP-mediated inhibition of TNF-α production by the atrial natriuretic peptide in murine macrophages

The atrial natriuretic peptide (ANP) is suggested to regulate inflammatory response by alteration of macrophage functions. The aim of this study was to investigate whether ANP influences production of TNF-α. TNF-α production in murine bone marrow-derived macrophages was induced by LPS, and TNF-α secretion (±ANP) was determined by L929 bioassay. ANP dose dependently (10^-8-10^-6 M) inhibited TNF-α release by up to 95%. The effect was mediated via the guanylate cyclase-coupled A receptor, as was shown by employing dibutyryl-cGMP, the cGMP-inhibitory compound Ly-83583, and the A receptor antagonist HS-142-1. A specific ligand of the natriuretic peptide 'clearance' receptor inhibited TNF-α production only at 10^-7 and 10^-8 M, but not at 10^-6 M. The B receptor ligand C-type natriuretic peptide showed no TNF-α-inhibitory effect. To investigate the underlying mechanism of ANP-mediated TNF-α inhibition, Northern blot was performed. ANP-treated macrophages displayed decreased TNF-α-mRNA levels. Besides the known inhibition of NF-κB activation, in this study we demonstrated that ANP also attenuates the activation of the proinflammatory transcription factor AP-1 (gel shift assay). ANP did not alter subunit composition of AP-1 complexes, as was shown by supershift assays applying anti-c-jun and anti-c- fos Abs. To get information on the ANP effect for human inflammatory processes, we investigated cytokine production in human LPS-activated blood. ANP significantly attenuated production of TNF-α and IL-1β without affecting production of IL.10 and IL-1ra. In summary, ANP was shown to attenuate TNF-α production of LPS-activated macrophages via cGMP. The inhibition is suggested to involve transcriptional processes that are the result of reduced activation of responsible transcription factors.