Toll-like receptor-4 is essential for Arcobacter butzleri-induced colonic and systemic immune responses in gnotobiotic IL-10−/− mice

Abstract

Arcobacter butzleri causes sporadic cases of gastroenteritis, but the underlying immunopathological mechanisms of infection are unknown. We have recently demonstrated that A. butzleri-infected gnotobiotic IL-10–/– mice were clinically unaffected but exhibited intestinal and systemic inflammatory immune responses. For the first time, we here investigated the role of Toll-like receptor (TLR)-4, the main receptor for lipopolysaccharide and lipooligosaccharide of Gram-negative bacteria, in murine arcobacteriosis. Gnotobiotic TLR-4/IL-10-double deficient (TLR-4–/– IL-10–/–) and IL-10–/– control mice generated by broad-spectrum antibiotics were perorally infected with A. butzleri. Until day 16 postinfection, mice of either genotype were stably colonized with the pathogen, but fecal bacterial loads were approximately 0.5–2.0 log lower in TLR-4–/– IL-10–/– as compared to IL-10–/– mice. A. butzleri-infected TLR-4–/– IL-10–/– mice displayed less pronounced colonic apoptosis accompanied by lower numbers of macrophages and monocytes, T lymphocytes, regulatory T-cells, and B lymphocytes within the colonic mucosa and lamina propria as compared to IL-10–/– mice. Furthermore, colonic concentrations of nitric oxide, TNF, IL-6, MCP-1, and, remarkably, IFN-γ and IL-12p70 serum levels were lower in A. butzleri-infected TLR-4–/– IL-10–/– versus IL-10–/– mice. In conclusion, TLR-4 is involved in mediating murine A. butzleri infection. Further studies are needed to investigate the molecular mechanisms underlying Arcobacter–host interactions in more detail.

Intestinal expression of genes encoding inflammatory mediators and gelatinases during Arcobacter butzleri infection of gnotobiotic IL-10 deficient mice

“…5 ). Hence, large intestinal immune responses upon A. butzleri infection shown previously [ 8 , 13 ] are associated with a strain-dependent expression pattern of mucus constituents, pro-inflammatory mediators including Th17 cytokines and their regulatory molecules as well as of matrix-degrading gelatinases.…”

“…Following stable intestinal colonization, A. butzleri- infected mice did not exhibit macroscopic disease but distinct pro-inflammatory sequelae that were not restricted to the small and large intestines, given that pronounced immune responses could be observed in extra-intestinal including systemic compartments [ 8 , 9 , 13 , 14 ]. Both innate and adaptive immune responses upon C. jejuni infection were characterized by elevated expression of genes encoding Th17 cytokines in human intestinal ex vivo biopsies [ 29 ].…”

“…Four-month old female gnotobiotic IL-10 –/– mice were perorally infected with approximately 10 9 viable colony forming units (CFU) of two different A. butzleri strains either (CCUG 30485 or C1 strain, respectively) by gavage in a total volume of 0.3 ml phosphate buffered saline (PBS) on two consecutive days (day 0 and day 1) as described previously [ 13 , 14 ] . Naive age-matched gnotobiotic IL-10 –/– mice served as uninfected controls.…”

Toll-like receptor-4 dependent intestinal gene expression during Arcobacter butzleri infection of gnotobiotic IL-10 deficient mice

“…Age-matched female gnotobiotic TLR-4 –/– IL-10 –/– and IL-10 –/– mice were perorally infected with approximately 10 9 colony forming units (CFU) of two different A. butzleri strains (either CCUG 30485 or C1 strain, respectively) by gavage in a total volume of 0.3 ml phosphate buffered saline (PBS) on two consecutive days (day 0 and day 1) as described previously [ 35 , 36 ]. Naive age-matched female gnotobiotic IL-10 –/– and TLR-4 –/– IL-10 –/– mice served as uninfected controls.…”

“…Whereas mice did not display any overt clinial signs of infection-induced disease, the bacteria did not only induce small and large intestinal but also extra-intestinal including systemic immune responses depending on the respective strain and the time course of infection [ 33 , 34 ]. These immune responses were dependent on Toll-like receptor (TLR) -4, the innate immune receptor for bacterial lipooligosaccharide (LOS), and lipopolysaccharide (LPS) derived from cell walls of gram-negative bacteria, given that gnotobiotic IL-10 –/– mice lacking TLR-4 presented with less pronounced intestinal and systemic immune responses [ 35 , 36 ]. The exact mechanisms underlying arcobacteriosis, however, are virtually unknown.…”