Skip to main content


Effects of Clostridium perfringens enterotoxin via claudin-4 on normal human pancreatic duct epithelial cells and cancer cells

Article metrics

  • 317 Accesses

  • 14 Citations


The tight junction protein claudin-4 is frequently overexpressed in pancreatic cancer, and is also a receptor for Clostridium perfringens enterotoxin (CPE). The cytotoxic effects of CPE are thought to be useful as a novel therapeutic tool for pancreatic cancer. However, the responses to CPE via claudin-4 remain unknown in normal human pancreatic duct epithelial (HPDE) cells. We introduced the human telomerase reverse transcriptase (hTERT) gene into HPDE cells in primary culture as a model of normal HPDE cells in vitro. hTERT-HPDE cells treated with or without 10% FBS and pancreatic cancer cell lines PANC-1, BXPC3, HPAF-II and HPAC were treated with CPE. In Western blotting, the expression of claudin-4 protein in hTERT-HPDE cells treated with 10% FBS was as high as it was in all of the pancreatic cancer cell lines. In hTERT-HPDE cells with or without 10% FBS, cytotoxicity was not observed at any concentration of CPE, whereas in all pancreatic cancer cell lines, CPE had a dose-dependent cytotoxic effect. In hTERT-HPDE cells with 10% FBS, claudin-4 was localized in the apical-most regions, where there are tight junction areas, in which in all pancreatic cancer cell lines claudin-4 was found not only in the apical-most regions but also at basolateral membranes. In hTERT-HPDE cells with 10% FBS after treatment with CPE, downregulation of barrier function and claudin-4 expression at the membranes was observed. In HPAC cells, the sensitivity to CPE was significantly decreased by knockdown of claudin-4 expression using siRNA compared to the control. These findings suggest that, in normal HPDE cells, the lack of toxicity of CPE was probably due to the localization of claudin-4, which is different from that of pancreatic cancer cells. hTERT-HPDE cells in this culture system may be a useful model of normal HPDE cells not only for physiological regulation of claudin-4 expression but also for developing safer and more effective therapeutic methods targeting claudin-4 in pancreatic cancer.



carbonic anhydrase isozyme 2




Clostridium perfringens enterotoxin


4′,6-diamidino-2-phenylindole dihydrochloride


Dulbecco’s modified Eagle’s medium


fetal bovine serum


human pancreatic duct epithelial


human telomerase reverse transcriptase


phosphate-buffered saline


room temperature


small interference RNAs


Tris-buffered saline


transepithelial electrical resistance


  1. 1.

    Morin, P.J. Claudin proteins in human cancer: promising new targets for diagnosis and therapy. Cancer Res. 65 (2005) 9603–9606.

  2. 2.

    Tsukita, S., Yamazaki, Y., Katsuno, T., Tamura, A. and Tsukita, S. Tight junction-based epithelial microenvironment and cell proliferation. Oncogene 27 (2008) 6930–6938.

  3. 3.

    Tsukita, S., Furuse, M. and Itoh, M. Multifunctional strands in tight junctions. Nat. Rev. Mol. Cell Biol. 2 (2001) 285–293.

  4. 4.

    Fujita, K., Katahira, J., Horiguchi, Y., Sonoda, N., Furuse, M. and Tsukita, S. Clostridium perfringens enterotoxin binds to the second extracellular loop of claudin-3, a tight junction integral membrane protein. FEBS Lett. 476 (2000) 258–261.

  5. 5.

    McClane, B.A. and Chakrabarti, G. New insights into the cytotoxic mechanisms of Clostridium perfringens enterotoxin. Anaerobe 10 (2004) 107–114.

  6. 6.

    Michl, P., Buchholz, M., Rolke, M., Kunsch, S., Löhr, M., McClane, B., Tsukita, S., Leder, G., Adler, G. and Gress, T.M. Claudin-4: a new target for pancreatic cancer treatment using Clostridium Perfringens enterotoxin. Gastroenterology 121 (2001) 678–684.

  7. 7.

    Katahira, J., Sugiyama, H,, Inoue, N., Horiguchi, Y., Matsuda, M. and Sugimoto, N. Clostridium perfringens enterotoxin utilizes two structurally related membrane proteins as functional receptors in vivo. J. Biol. Chem. 272 (1997) 26652–26658.

  8. 8.

    Yamaguchi, H., Kojima, T., Ito, T., Kimura, Y., Imamura, M., Son, S., Koizumi, J., Murata, M., Nagayama, M., Nobuoka, T., Tanaka, S., Hirata, K. and Sawada, N. Transcriptional control of tight junction proteins via a PKC signal pathway in hTERT-transfected human pancreatic duct epithelial cells. Am. J. Pathol. 177 (2010) 698–712.

  9. 9.

    Kojima, T., Fuchimoto, J., Takasawa, A., Yamaguchi, H., Ito, T., Ninomiya, T., Kikuchi, S., Ogasawara, N., Ohkuni, T., Masaki, T., Hirata, K., Himi, T. and Sawada, N. c-Jun N-terminal kinase is largely involved in the regulation of tricellular tight junctions via tricellulin in human pancreatic duct epithelial cells. J. Cell. Physiol. 225 (2010) 720–733.

  10. 10.

    Deer, E.L., González-Hernández, J., Coursen, J.D., Shea, J.E., Ngatia, J., Scaife, C.L., Firpo, M.A. and Mulvihill, S.J. Phenotype and genotype of pancreatic cancer cell lines. Pancreas 39 (2010) 425–435.

  11. 11.

    11 Winkler, L., Gehring, C., Wenzel, A., Müller, S.L., Piehl, C., Krause, G., Blasig, I.E. and Piontek, J. Molecular determinants of the interaction between Clostridium perfringens enterotoxin fragments and claudin-3. J. Biol. Chem. 284 (2009) 18863–18872.

  12. 12.

    Sonoda, N., Furuse, M., Sasaki, H., Yonemura, S., Katahira, J., Horiguchi, Y. and Tsukita, S. Clostridium perfringens enterotoxin fragment removes specific claudins from tight junction strands: Evidence for direct involvement of claudins in tight junction barrier. J. Cell Biol. 147 (1999) 195–204.

Download references

Author information

Correspondence to Takashi Kojima.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Yamaguchi, H., Kojima, T., Ito, T. et al. Effects of Clostridium perfringens enterotoxin via claudin-4 on normal human pancreatic duct epithelial cells and cancer cells. Cell Mol Biol Lett 16, 385–397 (2011) doi:10.2478/s11658-011-0014-z

Download citation

Key words

  • Tight junction
  • Claudin-4
  • CPE
  • Human pancreatic duct epithelial cells
  • Human pancreatic cancer cells