Haemolysin produced by Vibrio mimicus activates two Cl- secretory pathways in cultured ntestinal-like Caco-2 cells

Akira Takahashi, Shin-ichi Miyoshi, Noriko Takata, Masayuki Nakano, Akiko Hamamoto, Kazuaki Mawatari, Nagakatsu Harada, Sumio Shinoda, Yutaka Nakaya

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Abstract

Haemolysin (VMH) is a virulent factor produced by Vibrio mimicus, a human pathogen that causes diarrhoea. As intestinal epithelial cells are the primary targets of haemolysin, we investigated its effects on ion transport in human colonic epithelial Caco-2 cells. VMH increased the cellular short circuit current (Isc), used to estimated ion fluxes, and 125I efflux of the cells. The VMH-induced increases in Isc and 125I efflux were suppressed by depleting Ca from the medium or by pretreating the cells with BAPTA-AM or by Rp-adenosin 3′,5′-cyclic monophosphorothioate triethylammonium salt (Rp-cAMPS). The Cl - channel inhibitors 4,4′-disothiocyanatostibene-2,2′-disulfonic acid (DIDS), glybenclamide, and 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB) suppressed the VMH-induced increases in Isc and 125I efflux. Moreover, VMH increased the intracellular concentrations of Ca and cAMP. Thus, VMH stimulates Caco-2 cells to secrete Cl- by activating both Ca -dependent and cAMP-dependent Cl- secretion mechanisms. VMH forms ion-permeable pores in the lipid bilayer that are non-selectively permeable to small ions. However, the ion permeability of these pores was not inhibited by glybenclamide and DIDS, and VMH did not change the cell membrane potential. These observations indicate that the pores formed on the cell membrane by VMH are unlikely to be involved in VMH-induced Cl- secretion. Notably, VMH stimulated fluid accumulation in the iliac loop test that was fully suppressed by a combination of DIDS and glybenclamide. Thus, Ca -dependent and cAMP-dependent Cl- secretion may be important therapeutic targets with regard to the diarrhoea that is induced by Vibrio mimicus.

Original languageEnglish
Pages (from-to)583-595
Number of pages13
JournalCellular Microbiology
Volume9
Issue number3
DOIs
Publication statusPublished - Mar 2007

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Caco-2 Cells
Secretory Pathway
Vibrio mimicus
Glyburide
Ions
Hemolysin Proteins
Acids
Diarrhea
Cell membranes
Epithelial Cells
Cell Membrane
Ion Transport
Lipid Bilayers
Lipid bilayers
Membrane Potentials
Pathogens
Permeability
Salts
Short circuit currents
Vibrio hemolysin

ASJC Scopus subject areas

  • Clinical Biochemistry
  • Microbiology

Cite this

Haemolysin produced by Vibrio mimicus activates two Cl- secretory pathways in cultured ntestinal-like Caco-2 cells. / Takahashi, Akira; Miyoshi, Shin-ichi; Takata, Noriko; Nakano, Masayuki; Hamamoto, Akiko; Mawatari, Kazuaki; Harada, Nagakatsu; Shinoda, Sumio; Nakaya, Yutaka.

In: Cellular Microbiology, Vol. 9, No. 3, 03.2007, p. 583-595.

Research output: Contribution to journalArticle

Takahashi, A, Miyoshi, S, Takata, N, Nakano, M, Hamamoto, A, Mawatari, K, Harada, N, Shinoda, S & Nakaya, Y 2007, 'Haemolysin produced by Vibrio mimicus activates two Cl- secretory pathways in cultured ntestinal-like Caco-2 cells', Cellular Microbiology, vol. 9, no. 3, pp. 583-595. https://doi.org/10.1111/j.1462-5822.2006.00809.x
Takahashi, Akira ; Miyoshi, Shin-ichi ; Takata, Noriko ; Nakano, Masayuki ; Hamamoto, Akiko ; Mawatari, Kazuaki ; Harada, Nagakatsu ; Shinoda, Sumio ; Nakaya, Yutaka. / Haemolysin produced by Vibrio mimicus activates two Cl- secretory pathways in cultured ntestinal-like Caco-2 cells. In: Cellular Microbiology. 2007 ; Vol. 9, No. 3. pp. 583-595.
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abstract = "Haemolysin (VMH) is a virulent factor produced by Vibrio mimicus, a human pathogen that causes diarrhoea. As intestinal epithelial cells are the primary targets of haemolysin, we investigated its effects on ion transport in human colonic epithelial Caco-2 cells. VMH increased the cellular short circuit current (Isc), used to estimated ion fluxes, and 125I efflux of the cells. The VMH-induced increases in Isc and 125I efflux were suppressed by depleting Ca2± from the medium or by pretreating the cells with BAPTA-AM or by Rp-adenosin 3′,5′-cyclic monophosphorothioate triethylammonium salt (Rp-cAMPS). The Cl - channel inhibitors 4,4′-disothiocyanatostibene-2,2′-disulfonic acid (DIDS), glybenclamide, and 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB) suppressed the VMH-induced increases in Isc and 125I efflux. Moreover, VMH increased the intracellular concentrations of Ca2± and cAMP. Thus, VMH stimulates Caco-2 cells to secrete Cl- by activating both Ca2± -dependent and cAMP-dependent Cl- secretion mechanisms. VMH forms ion-permeable pores in the lipid bilayer that are non-selectively permeable to small ions. However, the ion permeability of these pores was not inhibited by glybenclamide and DIDS, and VMH did not change the cell membrane potential. These observations indicate that the pores formed on the cell membrane by VMH are unlikely to be involved in VMH-induced Cl- secretion. Notably, VMH stimulated fluid accumulation in the iliac loop test that was fully suppressed by a combination of DIDS and glybenclamide. Thus, Ca2± -dependent and cAMP-dependent Cl- secretion may be important therapeutic targets with regard to the diarrhoea that is induced by Vibrio mimicus.",
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AU - Harada, Nagakatsu

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AU - Nakaya, Yutaka

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