Effects of genetic and pharmacological inhibition of TNF-α in the regulation of inflammation in macrophages

Tumour necrosis factor-alpha (TNF-alpha) is an inflammatory cytokine produced by circulating monocytes and resident macrophages during acute inflammation and is responsible for a diverse range of signalling events within cells, leading to necrosis or apoptosis. The biologic activities of TNF are mediated by two receptors TNFR1 and TNFR2, although a lot of studies demonstrated that most of the biological activities of TNF-alpha are mediated through TNFR1. In the present study, we want to evaluate the role of TNF-alpha on regulation of in vitro models of inflammation. In particular we used peritoneal macrophages, from TNF-alpha R1 knock out and TNF-alpha R1 wild-type mice, stimulated with LPS 10 mu g/ml and IFN-gamma 100U/ml. Our results showed that the deletion of TNF-alpha R1 gene significantly reduced the degree of (i) MAPK activation, (ii) I kappa B-alpha degradation, (iii) phosphorylation of Ser536 on the NF-kappa B subunit p65 and (iv) iNOS and COX-2 expression. In addition, to confirm the pivotal role of TNF-alpha on regulation of peritoneal macrophages inflammation, we have also investigated the protective effects of infliximab, a TNF-alpha chimeric mouse/human IgG1 antibody against TNF-alpha. As shown in the present study, the cell incubation with infliximab (0.1 mu g/ml, 1 mu g/ml and 10 mu g/ml) significantly leads to a concentration-dependent inhibition of the inflammatory mediators above described. In conclusion, our study demonstrates that pharmacological and genetic inhibition of the TNF/TNFR1 binding reduce the degree of macrophages inflammation caused by LPS/IFN-gamma stimulation. (C) 2009 Published by Elsevier Ltd.