Escherichia coli strain Nissle 1917 ameliorates experimental colitis via toll-like receptor 2- and toll-like receptor 4-dependent pathways - PubMed (original) (raw)

. 2006 Jul;74(7):4075-82.

doi: 10.1128/IAI.01449-05.

D Paclik, C Guzy, A Dankof, D C Baumgart, J Erckenbrecht, B Raupach, U Sonnenborn, J Eckert, R R Schumann, B Wiedenmann, A U Dignass, A Sturm

Affiliations

Escherichia coli strain Nissle 1917 ameliorates experimental colitis via toll-like receptor 2- and toll-like receptor 4-dependent pathways

A Grabig et al. Infect Immun. 2006 Jul.

Abstract

Toll-like receptors (TLRs) are key components of the innate immune system that trigger antimicrobial host defense responses. The aim of the present study was to analyze the effects of probiotic Escherichia coli Nissle strain 1917 in experimental colitis induced in TLR-2 and TLR-4 knockout mice. Colitis was induced in wild-type (wt), TLR-2 knockout, and TLR-4 knockout mice via administration of 5% dextran sodium sulfate (DSS). Mice were treated with either 0.9% NaCl or 10(7) E. coli Nissle 1917 twice daily, followed by the determination of disease activity, mucosal damage, and cytokine secretion. wt and TLR-2 knockout mice exposed to DSS developed acute colitis, whereas TLR-4 knockout mice developed significantly less inflammation. In wt mice, but not TLR-2 or TLR-4 knockout mice, E. coli Nissle 1917 ameliorated colitis and decreased proinflammatory cytokine secretion. In TLR-2 knockout mice a selective reduction of gamma interferon secretion was observed after E. coli Nissle 1917 treatment. In TLR-4 knockout mice, cytokine secretion was almost undetectable and not modulated by E. coli Nissle 1917, indicating that TLR-4 knockout mice do not develop colitis similar to the wt mice. Coculture of E. coli Nissle 1917 and human T cells increased TLR-2 and TLR-4 protein expression in T cells and increased NF-kappaB activity via TLR-2 and TLR-4. In conclusion, our data provide evidence that E. coli Nissle 1917 ameliorates experimental induced colitis in mice via TLR-2- and TLR-4-dependent pathways.

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Figures

FIG. 1.

FIG. 1.

E. coli Nissle 1917-conditioned medium increases NF-κB activity via TLR-2- and TLR-4-dependent pathways. After TLR-2 and TLR-4 transfection, E. coli Nissle 1917-conditioned medium increases NF-κB activity. HEK 293 cells were transiently transfected as described in Materials and Methods. At 24 h after transfection cells were stimulated with LP3 or LPS (concentration in ng/ml) or E. coli Nissle 1917 supernatants (EcN) (concentration in μl/ml) or controls in Dulbecco modified Eagle medium. At 20 h after stimulation, cell extracts were prepared for determination of luciferase activity by using a Luciferase Reporter Gene Assay. LP3 was used at 20 ng/ml only. Each bar represents mean ± the SEM value of three separate experiments.

FIG. 2.

FIG. 2.

Distinct effect of DSS application and E. coli Nissle 1917 treatment in wt mice and in TLR-2 and TLR-4 knockout mice. In TLR-4 knockout mice, DSS application induced significantly fewer symptoms of colitis compared to wt or TLR-2 knockout mice. wt mice, but not TLR-2 knockout or TLR-4 knockout mice, treated with E. coli Nissle 1917 showed a significant clinical improvement in disease activity, as evaluated with the DAI, compared to mice treated with 0.9% NaCl. Colitis was induced by DSS application in wt mice, TLR-2 knockout mice, and TLR-4 knockout mice. Mice were subsequently treated with an oral or rectal application of 107 E. coli Nissle 1917 (100 μl) or 0.9% NaCl (100 μl) twice daily for 8 days. After sacrifice, the DAI was determined as described in Materials and Methods. Each bar represents mean ± the SEM value of 10 animals. *, P < 0.05 for differences in E. coli Nissle 1917-treated groups versus saline-treated animals.

FIG. 3.

FIG. 3.

Differential effect of E. coli Nissle 1917 treatment on inflammation and regeneration in wt mice but not in TLR-2 or TLR-4 knockout mice. In TLR-4 knockout mice DSS application induced significantly less inflammation compared to wt or TLR-2 knockout mice. wt mice, but not TLR-2 knockout or TLR-4 knockout mice, treated with E. coli Nissle 1917 showed significantly less inflammation or crypt damage and more regeneration, as evaluated with the Dieleman index, compared to mice treated with 0.9% NaCl. Colitis was induced by DSS application in wt mice, TLR-2 knockout mice, and TLR-4 knockout mice. Mice were subsequently treated with an oral or rectal application of 107 E. coli Nissle 1917 (100 μl) or 0.9% NaCl (100 μl) twice daily for 8 days. After sacrifice, tissue samples were obtained from the distal 5 cm of the colon and prepared for histological evaluation as described in Materials and Methods. Each bar represents mean ± the SEM value of 10 animals. *, P < 0.05 for differences in E. coli Nissle 1917-treated groups versus saline-treated animals.

FIG. 4.

FIG. 4.

Differential effect of E. coli Nissle 1917 treatment on mucosal damage in wt mice but not in TLR-2 or TLR-4 knockout mice. wt (A) and TLR-2 knockout (C) mice exposed to 5% DSS developed symptoms of acute colitis to a comparable degree, with a patchy pattern of severe mucosal damage characterized by a loss of crypts, necrosis, and the local influx of inflammatory cells into the lamina propria. (E) In contrast, in TLR-4 knockout mice, DSS application induced significantly fewer symptoms of colitis compared to wt and TLR-2 knockout mice. E. coli Nissle 1917 treatment ameliorated the severity of DSS-induced colitis in wt mice (B) but not in TLR-2 knockout (D) or TLR-4 knockout (F) mice. DSS colitis was induced as described in Materials and Methods, and mice were treated orally with 100 μl of 0.9% NaCl (A, C, and E) or 100 μl of 107 E. coli Nissle 1917 (B, D, and F). Control, non-DSS-treated wt C57BL/6 mice received, by gastric gavage, either 100 μl of isotonic sterile saline (G) or 100 μl of 107 E. coli Nissle 1917 (H). After sacrifice at day 8, tissue samples were obtained from the distal 5 cm of the colon and prepared for histological evaluation as described in Materials and Methods (hematoxylin and eosin stain; original magnifications, ×100).

FIG. 5.

FIG. 5.

Distinct effect of DSS application and E. coli Nissle 1917 treatment on MPO activity in wt, TLR-2 knockout, and TLR-4 knockout mice. In TLR-4 knockout mice, DSS application induced significantly less MPO activity in colonic tissue compared to wt or TLR-2 knockout mice. wt mice, but not TLR-2 knockout or TLR-4 knockout mice, treated rectally with 107 E. coli Nissle 1917 showed a significant decrease in MPO activity compared to mice treated with 0.9% NaCl. Colitis was induced by DSS application in wt, TLR-2 knockout, and TLR-4 knockout mice. Mice were subsequently treated with an oral or rectal application of 107 E. coli Nissle 1917 (100 μl) or 0.9% NaCl (100 μl) for 8 days. After sacrifice, the MPO activity was determined as described in Materials and Methods. Each bar represents mean ± the SEM value of 10 animals. *, P < 0.05 for differences in E. coli Nissle 1917-treated groups versus saline-treated animals.

FIG. 6.

FIG. 6.

Differential cytokine secretion in wt, TLR-2 knockout, and TLR-4 knockout mice. In wt animals, E. coli Nissle 1917 treatment decreased the secretion of TNF-α, IFN-γ, MCP-1, and IL-10. In TLR-2 knockout mice, only IFN-γ secretion was still high in control animals and reduced by E. coli Nissle 1917, whereas the secretion of TNF-α, MCP-1, and IL-10 was reduced compared to wt controls and not altered by E. coli Nissle 1917 treatment. In TLR-4 knockout mice, TNF-α, IFN-γ, IL-6, and IL-10 secretion was nearly abolished, and MCP-1 secretion was dramatically reduced compared to wt animals. A total of 105 freshly isolated PBT from wt, TLR-2 knockout, and TLR-4 knockout mice, treated orally or rectally with 0.9% NaCl or 107 E. coli Nissle 1917, were cultured for 3 days in the presence of anti-CD3 and CD28 MAbs. Supernatants were then collected, and cytokine secretion was analyzed by using a cytometric bead array assay. Each bar represents the mean ± the SEM value of 10 animals. *, P < 0.05 for differences in E. coli Nissle 1917-treated groups versus saline-treated animals.

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