ROLE OF NF-KB IN TNFRP55-/- MACROPHAGE HYPER-ACTIVATION BY YERSINIA LPS

ARIAS JL; ELIÇABE RJ; CARGNELUTTI E; DI GENARO MS

Mendoza

Congreso; XXVI Reunión Científica Anual de la Sociedad de Biología de Cuyo; 2008

Sociedad de Biología de Cuyo

The inducible transcription factor nuclear factor-kappaB (NF-kB) regulates gene expression during inflammatory and immune responses. Previously, we have demonstrated that TNFRp55 deficiency increases macrophage pro-inflammatory activity induced by Yersinia lipopolysaccharide (LPS) stimulation. The objective of the present work was to investigate whether NF-kB mediates this LPS-induced activation. Peritoneal macrophages from wild-type and TNFRp55-/- C57BL/6 mice were stimulated with Yersinia LPS (10, 100 or 1000 ng). Since the most abundant NF-kB form found in stimulated cells is the p65/p50 heterodimer, we analysed the effect of NF-kB inhibition by incubation of 100 ng LPS-stimulated macrophages with an oligonucleotide sequence of p65 antisense (1mM). Nitric oxide (NO) concentrations were measured in the supernatants by Griess reaction.  Higher NO levels were secreted by TNFRp55-/- macrophages stimulated with 100 or 1000 ng LPS (p<0.01 or p<0.005, respectively) compared with wild-type macrophages. In addition, polymixine B treatment, a LPS inhibitor, decreased significantly NO secretion (p<0.003). Interesting, antisense p65 significantly reduced NO secretion of LPS-stimulated TNFRp55-/- macrophages (p<0.05). Taken together, the above evidence implies that TNFRp55 deficiency increases susceptibility of macrophages to LPS stimulation, and NF-kB p65 is a transcriptional regulator of this LPS-induced hyper-activation.