Atrial natriuretic peptide reduces expression of TNF-alpha mRNA during reperfusion of the rat liver upon decreased activation of NF-kappa B and AP-1

Background/Aims: The cardiovascular hormone Atrial Natriuretic Peptide (ANP ) attenuates activation of the pro-inflammatory transcription factor NF-kap pa B in macrophages. ANP was also shown to protect from ischemia-reperfusio n injury of the rat liver. This study aimed to investigate the effects of t his immunomodulatory hormone and its second messenger cGMP on the activatio n of the two redox-sensitive transcription factors AP-1 and NF-kappa B and the expression of corresponding pro-inflammatory target genes during ischem ia and reperfusion of the liver, The identification of the mechanisms under lying the protection by ANP should reveal new aspects concerning the pathom echanisms of ischemia/reperfusion injury. Methods: Rat livers were perfused with and without ANP or 8-Br-cGMP precedi ng 24 h of cold storage in University of Wisconsin solution, During reperfu sion NF-kappa B and AP-1 DNA binding activities were determined in freeze-c lamped liver samples by electrophoretic mobility shift assay, Protein level s of p50, p65, and of I kappa B were determined by Western blot, mRNA codin g for inducible nitric oxide synthase, cyclooxygenase-2, and TNF-alpha was determined by RT-PCR and Northern blot. Results: After 45 min of reperfusion DNA binding activities of NF-kappa B w ere increased, whereas in ANP pre-treated livers this effect was markedly r educed. AP-1, another important redox-sensitive transcription factor, mas a ctivated and in the course of reperfusion the subunit composition of AP-1 c hanged as assessed by supershift assays. ANP markedly reduced binding activ ities of both forms of AP-1, 8-Br-cGMP mimicked the effects of;ANP on NF-ka ppa B and AP-1, Neither inducible nitric oxide synthase nor cyclooxygenase- 2 mRNA could be detected. In contrast, a profound expression of transcripts coding for TNF-alpha was detected in the course of reperfusion and ANP mar kedly reduced TNF-alpha mRNA expression, Conclusion: ANP seems to mediate its protective effect during ischemia and reperfusion by reducing the activation of NF-kappa B and AP-1 via cGMP. The reduced binding activity of these redox-sensitive transcription factors wa s accompanied by a diminished mRNA expression of TNF-alpha, a cytokine know n to be involved in cellular damage in ischemia reperfusion injury.