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A critical role for hemolysin in Vibrio fluvialis-induced IL-1β secretion mediated by the NLRP3 inflammasome in macrophages.

Song L, Huang Y, Zhao M, Wang Z, Wang S, Sun H, Kan B, Meng G, Liang W, Ren Z - Front Microbiol (2015)

Bottom Line: The mice treated with 10(8) CFU wild-type V. fluvialis or cell-free supernatant containing VFH induced significantly higher IL-1β production in peritoneal lavage fluid or in colon compared with those infected with the mutant strain, while no effect on TNF and IL-6 production was observed at day 5 or 24 h post-infection.VFH has no effect on the synthesis of pro-IL-1β, but rather it triggers the processing of pro-IL-1β into IL-1β.Summary Sentence: Vibrio fluvialis-secreted hemolysin induces IL-1β secretion through the activation of the NLRP3 inflammasome and contributes to the pathogenicity of V. fluvialis.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory for Infectious Disease Prevention and Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, National Institute for Communicable Disease Control and Prevention - Chinese Center for Disease Control and Prevention Beijing, China.

ABSTRACT
Vibrio fluvialis causes human diarrhea, but the pathogenesis is not well-studied. We hypothesized that V. fluvialis-secreted hemolysin (VFH) may induce IL-1β secretion through the activation of the NLRP3 inflammasome and contribute to the pathogenicity of V. fluvialis. To examine this possibility, we constructed VFH mutant and complement strains and demonstrated that V. fluvialis-induced IL-1β production and cytotoxicity in human monocytic THP-1 cells and mouse macrophages is attributed to VFH. To evaluate the role of VFH in vivo, we infected adult C57BL/6 mice intraperitoneally and suckling C57/B6 mice orally with various strains. The mice treated with 10(8) CFU wild-type V. fluvialis or cell-free supernatant containing VFH induced significantly higher IL-1β production in peritoneal lavage fluid or in colon compared with those infected with the mutant strain, while no effect on TNF and IL-6 production was observed at day 5 or 24 h post-infection. VFH contributed to pathological changes and IL-1β release independent of colonization of V. fluvialis in the colon. VFH has no effect on the synthesis of pro-IL-1β, but rather it triggers the processing of pro-IL-1β into IL-1β. Furthermore, using deficient mouse strains, we verified that V. fluvialis-induced IL-1β is mediated through activation of Caspase-1 and the NLRP3 inflammasome ex vivo. Confocal microscopy suggests that VFH contributes to cathepsin B release. Furthermore, V. fluvialis-induced IL-1β secretion requires potassium (K(+)) efflux and reactive oxygen species production. Our results provide new evidence for the role of VFH in the activation of the NLRP3 inflammasome and pathogenesis in response to V. fluvialis infection. Summary Sentence: Vibrio fluvialis-secreted hemolysin induces IL-1β secretion through the activation of the NLRP3 inflammasome and contributes to the pathogenicity of V. fluvialis.

No MeSH data available.


Related in: MedlinePlus

Cathepsin B is involved in IL-1β secretion in BMMs upon V. fluvialis infection. (A) BMMs were left untreated or were infected with WT, Δvfh, pUC-vfh, or pUC18 V. fluvialis strains for 2 h. Cathepsin B was visualized by confocal laser scanning microscopy using a specific Ab. Actin was stained with phalloidin AF488. (B,C) BMMs were treated with cathepsin B inhibitor CA-074Me at the indicated concentrations for 1 h before infection with WT or Δvfh for 3 h. IL-1β (B) and TNF-α (C) in the supernatants were quantified by ELISA. Results represent mean ± SD of three independent experiments. ∗P < 0.05; ∗∗P < 0.01.
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Figure 6: Cathepsin B is involved in IL-1β secretion in BMMs upon V. fluvialis infection. (A) BMMs were left untreated or were infected with WT, Δvfh, pUC-vfh, or pUC18 V. fluvialis strains for 2 h. Cathepsin B was visualized by confocal laser scanning microscopy using a specific Ab. Actin was stained with phalloidin AF488. (B,C) BMMs were treated with cathepsin B inhibitor CA-074Me at the indicated concentrations for 1 h before infection with WT or Δvfh for 3 h. IL-1β (B) and TNF-α (C) in the supernatants were quantified by ELISA. Results represent mean ± SD of three independent experiments. ∗P < 0.05; ∗∗P < 0.01.

Mentions: To determine whether cathepsin B is activated and released from cells upon V. fluvialis-induced IL-1β release, we performed immunofluorescence staining and confocal microscopy of BMMs. Uninfected cells showed punctate staining, which was obviously diminished in WT-infected cells and pUC-Δvfh-infected cells, but was apparent in Δvfh-infected cells. These results suggest that VFH contributes to cathepsin B release from phagoendosomes to the cytosol (Figure 6A).


A critical role for hemolysin in Vibrio fluvialis-induced IL-1β secretion mediated by the NLRP3 inflammasome in macrophages.

Song L, Huang Y, Zhao M, Wang Z, Wang S, Sun H, Kan B, Meng G, Liang W, Ren Z - Front Microbiol (2015)

Cathepsin B is involved in IL-1β secretion in BMMs upon V. fluvialis infection. (A) BMMs were left untreated or were infected with WT, Δvfh, pUC-vfh, or pUC18 V. fluvialis strains for 2 h. Cathepsin B was visualized by confocal laser scanning microscopy using a specific Ab. Actin was stained with phalloidin AF488. (B,C) BMMs were treated with cathepsin B inhibitor CA-074Me at the indicated concentrations for 1 h before infection with WT or Δvfh for 3 h. IL-1β (B) and TNF-α (C) in the supernatants were quantified by ELISA. Results represent mean ± SD of three independent experiments. ∗P < 0.05; ∗∗P < 0.01.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4440915&req=5

Figure 6: Cathepsin B is involved in IL-1β secretion in BMMs upon V. fluvialis infection. (A) BMMs were left untreated or were infected with WT, Δvfh, pUC-vfh, or pUC18 V. fluvialis strains for 2 h. Cathepsin B was visualized by confocal laser scanning microscopy using a specific Ab. Actin was stained with phalloidin AF488. (B,C) BMMs were treated with cathepsin B inhibitor CA-074Me at the indicated concentrations for 1 h before infection with WT or Δvfh for 3 h. IL-1β (B) and TNF-α (C) in the supernatants were quantified by ELISA. Results represent mean ± SD of three independent experiments. ∗P < 0.05; ∗∗P < 0.01.
Mentions: To determine whether cathepsin B is activated and released from cells upon V. fluvialis-induced IL-1β release, we performed immunofluorescence staining and confocal microscopy of BMMs. Uninfected cells showed punctate staining, which was obviously diminished in WT-infected cells and pUC-Δvfh-infected cells, but was apparent in Δvfh-infected cells. These results suggest that VFH contributes to cathepsin B release from phagoendosomes to the cytosol (Figure 6A).

Bottom Line: The mice treated with 10(8) CFU wild-type V. fluvialis or cell-free supernatant containing VFH induced significantly higher IL-1β production in peritoneal lavage fluid or in colon compared with those infected with the mutant strain, while no effect on TNF and IL-6 production was observed at day 5 or 24 h post-infection.VFH has no effect on the synthesis of pro-IL-1β, but rather it triggers the processing of pro-IL-1β into IL-1β.Summary Sentence: Vibrio fluvialis-secreted hemolysin induces IL-1β secretion through the activation of the NLRP3 inflammasome and contributes to the pathogenicity of V. fluvialis.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory for Infectious Disease Prevention and Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, National Institute for Communicable Disease Control and Prevention - Chinese Center for Disease Control and Prevention Beijing, China.

ABSTRACT
Vibrio fluvialis causes human diarrhea, but the pathogenesis is not well-studied. We hypothesized that V. fluvialis-secreted hemolysin (VFH) may induce IL-1β secretion through the activation of the NLRP3 inflammasome and contribute to the pathogenicity of V. fluvialis. To examine this possibility, we constructed VFH mutant and complement strains and demonstrated that V. fluvialis-induced IL-1β production and cytotoxicity in human monocytic THP-1 cells and mouse macrophages is attributed to VFH. To evaluate the role of VFH in vivo, we infected adult C57BL/6 mice intraperitoneally and suckling C57/B6 mice orally with various strains. The mice treated with 10(8) CFU wild-type V. fluvialis or cell-free supernatant containing VFH induced significantly higher IL-1β production in peritoneal lavage fluid or in colon compared with those infected with the mutant strain, while no effect on TNF and IL-6 production was observed at day 5 or 24 h post-infection. VFH contributed to pathological changes and IL-1β release independent of colonization of V. fluvialis in the colon. VFH has no effect on the synthesis of pro-IL-1β, but rather it triggers the processing of pro-IL-1β into IL-1β. Furthermore, using deficient mouse strains, we verified that V. fluvialis-induced IL-1β is mediated through activation of Caspase-1 and the NLRP3 inflammasome ex vivo. Confocal microscopy suggests that VFH contributes to cathepsin B release. Furthermore, V. fluvialis-induced IL-1β secretion requires potassium (K(+)) efflux and reactive oxygen species production. Our results provide new evidence for the role of VFH in the activation of the NLRP3 inflammasome and pathogenesis in response to V. fluvialis infection. Summary Sentence: Vibrio fluvialis-secreted hemolysin induces IL-1β secretion through the activation of the NLRP3 inflammasome and contributes to the pathogenicity of V. fluvialis.

No MeSH data available.


Related in: MedlinePlus