Basic Research
Copyright ©2007 Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Oct 21, 2007; 13(39): 5217-5225
Published online Oct 21, 2007. doi: 10.3748/wjg.v13.i39.5217
Exogenous sphingomyelinase causes impaired intestinal epithelial barrier function
Jürgen Bock, Gerhard Liebisch, Joachim Schweimer, Gerd Schmitz, Gerhard Rogler
Jürgen Bock, Joachim Schweimer, Gerhard Rogler, Department of Internal MedicineI, University of Regensburg, Germany
Gerhard Liebisch, Gerd Schmitz, Institute for Clinical Chemistry, University of Regensburg, Germany
Author contributions: All authors contributed equally to the work.
Supported by grants from the University of Regensburg, as part of the ReForM-program, and from the German Research Foundation DFG (BO 2529/2-1) to JB
Correspondence to: Jürgen Bock, MD, Department of Internal MedicineI, University of Regensburg, Regensburg 93042, Germany.
Telephone: +49-941-9447010 Fax: +49-941-9447073
Received: March 29, 2007
Revised: July 12, 2007
Accepted: August 18, 2007
Published online: October 21, 2007

AIM: To test the hypothesis that hydrolysis of sphingomyelin to ceramide changes the composition of tight junctions (TJs) with increasing permeability of the intestinal epithelium.

METHODS: Monolayers of Caco-2 cells were used as an in vitro model for the intestinal barrier. Permeability was determined by quantification of transepithelial flux and transepithelial resistance. Sphingolipid-rich membrane microdomains were isolated by a discontinuous sucrose gradient and characterized by Western-blot. Lipid content of microdomains was analysed by tandem mass spectrometry. Ceramide was subcellularly localized by immunofluorescent staining.

RESULTS: Exogenous sphingomyelinase increased transepithelial permeability and decreased transepithelial resistance at concentrations as low as 0.01 U/mL. Lipid analysis showed rapid accumulation of ceramide in the membrane fractions containing occludin and claudin-4, representing TJs. In these fractions we observed a concomitant decrease of sphingomyelin and cholesterol with increasing concentrations of ceramide. Immunofluorescent staining confirmed clustering of ceramide at the sites of cell-cell contacts. Neutralization of surface ceramide prevented the permeability-increase induced by platelet activating factor.

CONCLUSION: Our findings indicate that changes in lipid composition of TJs impair epithelial barrier functions. Generation of ceramide by sphingomyelinases might contribute to disturbed barrier function seen in diseases such as inflammatory, infectious, toxic or radiogenic bowel disease.

Keywords: Ceramide, Cholesterol, Tight-junction, Caco-2 cells, Permeability, Inflammatory bowel disease