Salmonella enterica serovar Enteritidis Dam mutant infection induces attenuated NF-kB and MAPK activation in macrophages.

CERQUETTI, MC; HOVSEPIÁN, E; SARNACKI, SH; GOREN, NB.

Estambul, Turquía.

Congreso; XII International Congress of Bacteriology and Applied Microbiology.; 2008

International Union of Microbiological Societies (IUMS)

Although dam mutants of Salmonella have been proposed as live vaccines, their capacity to trigger cell inflammatory cascades has not been fully elucidated.  In this study we investigated in detail the ability of S. Enteritidis dam mutant to activate the signaling pathways of the inflammatory response in RAW 264.7 cells. The rate of apoptosis, determined by flow cytometry and TUNEL, was higher in macrophages treated with the virulent Salmonella than in dam mutant-infected cells.  Both, the expression of NOS-2 and COX-2 and subsequently the production of NO and PGE2 were significantly reduced in cells infected with the mutant strain.  Interesting, Q-RT-PCR assay showed not only a diminished amount of, but also a delay in the expression of COX-2 mRNA in cells infected with the dam mutant.  Our results also indicate that the dam mutant of S. Enteritidis induces an attenuated activation of the NF-kB pathway.  Degradation of IêBá and IêBâ was diminished in dam mutant-infected cells and the capacity of these bacteria to induce IêBá phosphorylation was also impaired.  Translocation of p65 to the nucleus, determined by Western blot and EMSA, was notably reduced in cells infected with the dam mutant of S. Enteritidis. The amount of phosphorylated p44, p42 and p38 MAPKs was clearly reduced in extracts from dam-infected macrophages, particularly at early time points after stimulation. These results indicate that the lack of ERK and p38 phosphorylation at the proper time in dam-infected cells notably reduces the engagement of subsequent signaling pathways involved in the full activation of NF-kB in response to infection.  Taken together, these results suggest that Salmonella activation of both signaling cascades in the inflammatory response is a mechanism requiring Dam protein participation.