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Related Experiment Video

Updated: Jul 11, 2026

Live Imaging Assay for Assessing the Roles of Ca2+ and Sphingomyelinase in the Repair of Pore-forming Toxin Wounds
18:25

Live Imaging Assay for Assessing the Roles of Ca2+ and Sphingomyelinase in the Repair of Pore-forming Toxin Wounds

Published on: August 25, 2013

Exogenous sphingomyelinase causes impaired intestinal epithelial barrier function.

Jurgen Bock1, Gerhard Liebisch, Joachim Schweimer

  • 1Department of Internal Medicine I, University of Regensburg, Regensburg, Germany. juergen.bock@klinik.uni-regensburg.de

World Journal of Gastroenterology
|September 19, 2007
PubMed
Summary

This study investigated how sphingomyelinase affects intestinal barrier function. Using Caco-2 cells as a model, researchers found that even low concentrations of the enzyme increased permeability and decreased resistance. They observed that sphingomyelinase activity led to ceramide accumulation in tight junctions, which correlated with decreased sphingomyelin and cholesterol levels. Immunofluorescence confirmed ceramide clustering at cell-cell contacts. Neutralizing ceramide prevented permeability increases. The findings suggest that sphingomyelinase-induced lipid changes disrupt tight junction integrity. This mechanism may explain barrier dysfunction in various bowel diseases. The study highlights the role of sphingolipid metabolism in epithelial barrier regulation.

Keywords:
Intestinal permeabilityTight junction compositionSphingomyelinase activityEpithelial barrier dysfunction

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Related Experiment Videos

Last Updated: Jul 11, 2026

Live Imaging Assay for Assessing the Roles of Ca2+ and Sphingomyelinase in the Repair of Pore-forming Toxin Wounds
18:25

Live Imaging Assay for Assessing the Roles of Ca2+ and Sphingomyelinase in the Repair of Pore-forming Toxin Wounds

Published on: August 25, 2013

Area of Science:

  • Intestinal physiology within gastrointestinal medicine
  • Membrane lipid biochemistry in cell biology
  • Epithelial barrier function research in pharmacology

Background:

It was already known that tight junctions (TJs) regulate intestinal permeability. Prior research has shown that sphingolipids influence membrane structure and function. No prior work had resolved how sphingomyelinase affects TJ composition. This gap motivated investigation into whether sphingomyelin hydrolysis alters TJ integrity. The role of ceramide in TJ remodeling remains unclear. Researchers propose that sphingolipid metabolism may modulate barrier function. Existing studies suggest that lipid composition affects paracellular permeability. That uncertainty drove the need to test the hypothesis that sphingomyelinase activity changes TJ structure.

Purpose Of The Study:

The aim was to determine if sphingomyelinase-induced ceramide production alters intestinal barrier integrity. Researchers wanted to assess whether TJ composition changes with sphingomyelin hydrolysis. The specific problem addressed was the mechanism by which sphingomyelinase affects epithelial permeability. This study sought to clarify the relationship between lipid metabolism and TJ structure. The motivation came from observing TJ dysfunction in various bowel diseases. The hypothesis proposed that ceramide accumulation disrupts TJ organization. The study tested whether sphingomyelinase activity leads to TJ permeability changes. The researchers aimed to identify the role of sphingolipid metabolism in epithelial barrier function.

Main Methods:

Caco-2 cell monolayers were used as an in vitro model for intestinal epithelium. Transepithelial permeability was measured using flux and resistance quantification. Sphingolipid-rich microdomains were isolated via sucrose gradient centrifugation. Western blot analysis identified TJ proteins in membrane fractions. Tandem mass spectrometry quantified lipid content in isolated fractions. Immunofluorescence localized ceramide within cell-cell junctions. Exogenous sphingomyelinase was applied at varying concentrations. The effects on TJ composition and permeability were analyzed over time.

Main Results:

Exogenous sphingomyelinase increased permeability at concentrations as low as 0.01 U/mL. Transepithelial resistance decreased significantly with enzyme treatment. Ceramide levels rose rapidly in membrane fractions containing occludin and claudin-4. Sphingomyelin and cholesterol decreased as ceramide concentrations increased. Immunofluorescence confirmed ceramide clustering at cell-cell contacts. Neutralizing surface ceramide prevented permeability increases. Platelet activating factor-induced permeability was blocked by ceramide neutralization. These findings suggest that TJ lipid composition changes correlate with barrier dysfunction.

Conclusions:

The authors propose that sphingomyelinase activity alters TJ lipid composition. Ceramide accumulation correlates with increased intestinal permeability. The observed decrease in sphingomyelin and cholesterol supports this mechanism. TJ disruption may explain barrier dysfunction in various bowel diseases. The findings suggest that lipid metabolism influences epithelial barrier integrity. The study supports a role for sphingomyelinase in TJ remodeling. Researchers suggest that ceramide generation may contribute to disease-related barrier failure. These results may inform understanding of TJ dynamics in intestinal pathophysiology.

Exogenous sphingomyelinase increases permeability and decreases resistance in Caco-2 monolayers.

Ceramide clusters at cell-cell contacts and correlates with decreased sphingomyelin and cholesterol.

Ceramide clustering at junctions suggests direct impact on tight junction structure and permeability.

Ceramide neutralization prevented permeability increases induced by platelet activating factor.

TJ proteins and lipids were isolated via sucrose gradient and analyzed by Western blot and mass spectrometry.

The authors suggest sphingomyelinase may contribute to barrier dysfunction in inflammatory and toxic bowel diseases.