Yersinia lipopolysaccharide induces increased pro-inflammatory response in TNFRp55-/-ƒnmacrophages

ELIÇABE JAVIER1, MACCIONI MARINA2 CIUFFO GLADYS1, DI GENARO SILVIA1.

Villa Carlos Paz, Córdoba

Congreso; XLIV Reunion Anual-Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2008

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular

Yersinia enterocolitica causes reactive arthritis and macrophages mediate synovitis. We studied the impact of TNFRp55 deficiency in macrophages stimulated with Yersinia antigens. Peritoneal macrophages from wild-type and TNFRp55-/- C57BL/6 mice were stimulated with Y. enterocolitica O:3 outer membrane (OM) or lipopolysaccharide (LPS). Supernatant nitric oxide (NO) by Griess reaction; and TNF-a and IL-6 by ELISA were measured. Toll-like receptor (TLR)-2 and TLR4 mRNA expression was evaluated by RT-PCR; and iNOS protein expression by western blot. Higher NO levels were secreted by TNFRp55-/- macrophages stimulated with 5 or 10 mg OM (p<0.03 or p<0.006, respectively compared with wilt-type).This was in line with iNOS protein and TLR-4 mRNA expression. TLR-4-/- macrophages did not respond to OM stimulation. In addition, higher NO levels were secreted by of TNFRp55-/- macrophages stimulated with 100 or 1000 ng LPS (p<0.011 or p<0.005, respectively). Similarly, increased IL-6 levels were found after 10, 100 or 1000 ng LPS stimulation (p<0.0004, p<0.05 and p<0.007, respectively).  Polymixine B treatment decreased significantly NO and IL-6 secretions (p<0.05). These results suggested that TNFRp55-/- deficiency increases macrophage pro-inflammatory activity, and that Yersinia LPS is the main OM component that induces this activation.