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

Background/Aims: The cardiovascular hormone Atrial Natriuretic Peptide (ANP) attenuates activation of the pro-inflammatory transcription factor NF-kappa B in macrophages. ANP was also shown to protect from ischemia-reperfusion injury of the rat liver. This study aimed to investigate the effects of this immunomodulatory hormone and its second messenger cGMP on the activation of the two redox-sensitive transcription factors AP-1 and NF-kappa B and the expression of corresponding pro-inflammatory target genes during ischemia and reperfusion of the liver, The identification of the mechanisms underlying the protection by ANP should reveal new aspects concerning the pathomechanisms of ischemia/reperfusion injury. Methods: Rat livers were perfused with and without ANP or 8-Br-cGMP preceding 24 h of cold storage in University of Wisconsin solution, During reperfusion NF-kappa B and AP-1 DNA binding activities were determined in freeze-clamped liver samples by electrophoretic mobility shift assay, Protein levels of p50, p65, and of I kappa B were determined by Western blot, mRNA coding 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 were increased, whereas in ANP pre-treated livers this effect was markedly reduced. AP-1, another important redox-sensitive transcription factor, mas activated and in the course of reperfusion the subunit composition of AP-1 changed as assessed by supershift assays. ANP markedly reduced binding activities of both forms of AP-1, 8-Br-cGMP mimicked the effects of;ANP on NF-kappa 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 markedly 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 was accompanied by a diminished mRNA expression of TNF-alpha, a cytokine known to be involved in cellular damage in ischemia reperfusion injury.