Immune Netw.  2013 Oct;13(5):213-217. 10.4110/in.2013.13.5.213.

Bacteroides fragilis Toxin Induces IL-8 Secretion in HT29/C1 Cells through Disruption of E-cadherin Junctions

• Hwang S
• Gwon SY
• Kim MS
• Lee S
• Rhee KJ

• KMID: 1493616
• DOI: http://doi.org/10.4110/in.2013.13.5.213

Abstract

Enterotoxigenic Bacteroides fragilis (ETBF) is a human gut commensal bacteria that causes inflammatory diarrhea and colitis. ETBF also promotes colorectal tumorigenesis in the Min mouse model. The key virulence factor is a secreted metalloprotease called B. fragilis toxin (BFT). BFT induces E-cadherin cleavage, cell rounding, activation of the beta-catenin pathway and secretion of IL-8 in colonic epithelial cells. However, the precise mechanism by which these processes occur and how these processes are interrelated is still unclear. E-cadherin form homophilic interactions which tethers adjacent cells. Loss of E-cadherin results in detachment of adjacent cells. Prior studies have suggested that BFT induces IL-8 expression by inducing E-cadherin cleavage; cells that do not express E-cadherin do not secrete IL-8 in response to BFT. In the current study, we found that HT29/C1cells treated with dilute trypsin solution induced E-cadherin degradation and IL-8 secretion, consistent with the hypothesis that E-cadherin cleavage causes IL-8 secretion. However, physical damage to the cell monolayer did not induce IL-8 secretion. We also show that EDTA-mediated disruption of E-cadherin interactions without E-cadherin degradation was sufficient to induce IL-8 secretion. Finally, we determined that HT29/C1 cells treated with LiCl (beta-catenin activator) induced IL-8 secretion in a dose-dependent and time-dependent manner. Taken together, our results suggest that BFT induced IL-8 secretion may occur by the following process: E-cadherin cleavage, disruption of cellular interactions, activation of the beta-catenin pathway and IL-8 expression. However, we further propose that E-cadherin cleavage per se may not be required for BFT induced IL-8 secretion.

Keyword

Bacteroides fragilis toxin; Interleukin-8; E-cadherin; beta-catenin; EDTA; LiCl

MeSH Terms

Animals
Bacteria
Bacterial Toxins
Bacteroides fragilis*
Bacteroides*
beta Catenin
Cadherins*
Cell Transformation, Neoplastic
Colitis
Colon
Diarrhea
Edetic Acid
Epithelial Cells
Fibrinogen
Humans
Interleukin-8*
Metalloendopeptidases
Mice
Trypsin
Bacterial Toxins
Cadherins
Edetic Acid
Fibrinogen
Interleukin-8
Metalloendopeptidases
Trypsin
beta Catenin

Figure

• 

  Figure 1 Induction of IL-8 secretion by various stimuli. (A) HT29/C1 cells or HEK293/17 cells were cultured with BFT (100 ng/ml) or IL-1β (50 ng/ml) for 24 hr and IL-8 secretion assessed by ELISA. (B) HT29/C1 cells were cultured with diluted trypsin solution (1/100, 1/1,000) or dextran sulfate sodium (DSS) for 24 hr and IL-8 secretion assessed by ELISA. To induce physical damage, the subconfluent cell monolayer was scratched in a line with a pipette tip either 5 or 20 times. Asterisks indicate p<0.05. Data are shown as mean and SEM.

• 

  Figure 2 Disruption of E-cadherin interaction but not physical damage induces IL-8 secretion. Subconfluent HT29/C1 cells were cultured with EDTA, scratched (20 times) or treated with purified BFT (100 ng/ml) for 24 hr. (A) IL-8 secretion. (B) Western blot analysis of E-cadherin and β-actin. Cells were treated with Asterisks indicate p<0.05. Data are shown as mean and SEM.

• 

  Figure 3 LiCl treatment induces IL-8 secretion in HT29/C1 cells. Cells were cultured with (A) different concentrations of LiCl for 24 hr or (B) different lengths of time with 50 mM of LiCl. IL-8 secretion was assessed by ELISA. Asterisks indicate p<0.05. Data are shown as mean and SEM.

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