Molecular pathogenesis of Salmonella enterica serotype Typhimurium-induced inflammatory responses

Abstract

We demonstrated that infection of HeLa cells, which are non-responsive to flagellin, with wild type Salmonella enterica serotype Typhimurium (S. typhimurium) activated chemokine expression at higher level than S. typhimurium lacking sipAsopABDE2, indicating that the corresponding effector proteins (SipA, SopA, SopB, SopD and SopE2) are required to induce chemokines independent of flagellin. The S. typhimurium sipAsopABDE2 mutant complemented with sipA activated IL-8 expression at significantly higher level than a S. typhimurium sipAsopABDE2 mutant. However, extracellular addition of recombinant SipA failed to induce IL-8. Phosphorylation analyses demonstrated that S. typhimurium carrying a chromosomal copy of sipA (sopABDE2 mutant) induced phosphorylation of CREB1, JUN and p38MAPK, which are proteins involved in IL-8 expression. The contribution of effector proteins to S. typhimurium-induced intracellular Ca2+ mobilization and its role in IL-8 expression and bacterial internalization were also investigated. Our results demonstrated that wild type S. typhimurium significantly increased the amplitude of intracellular Ca2+ beginning 30 sec after infection. However, further analyses of intracellular Ca2+ changes in HeLa cells infected with S. typhimurium mutants indicated no correlation between increased intracellular Ca2+ and IL-8 expression or bacterial internalization. To analyze specific cell populations targeted by wild type S. typhimurium or S. typhimurium carrying a chromosomal copy of sipA (sopABDE2 mutant), laser capture microdissection was performed. Our data indicated that in wild type S. typhimurium-infected bovine Peyer’s patches, high levels of IL-8 were expressed in enterocytes of crypts, whereas Gro-α was expressed in enterocytes of both crypts and absorptive villi. A strain carrying a chromosomal copy of sipA colonized the same cell population as wild type, but induced IL8 and Gro-α in enterocytes of both crypts and absorptive villi. In conclusion, we demonstrated that in vitro S. typhimurium effector proteins induce chemokine expression independent of Ca2+ changes through phosphorylation of proteins related to IL-8 pathway. In vivo, we found higher levels of IL-8 expression in enterocytes of crypts than enterocytes of absorptive villi, although both cell populations contributed to Gro-α expression. These data extend the knowledge of the molecular mechanism by which S. typhimurium induces inflammatory genes by identifying pathogen and host molecules involved in inflammation.