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Updated: Sep 1, 2025
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Determining Intestinal Permeability Using Lucifer Yellow in an Apical-Out Enteroid Model
Published on: July 27, 2022
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Determining Intestinal Permeability using Lucifer Yellow in an Apical-Out Enteroid Model
Heather Liebe1, Camille Schlegel2, Xue Cai2
1Division of Pediatric Surgery, Oklahoma Children's Hospital.
Journal of Visualized Experiments : Jove
|August 15, 2022
Summary
This study introduces a novel apical-out enteroid model to better study necrotizing enterocolitis (NEC) and intestinal permeability. The new model, using lucifer yellow, enhances understanding of host-pathogen interactions in NEC research.
Area of Science:
- Gastroenterology and Hepatology
- Developmental Biology
- Infectious Diseases
Background:
- Enteroids are valuable research models for inflammatory bowel diseases like necrotizing enterocolitis (NEC).
- Traditional basolateral-out (BO) enteroid models present challenges in accessing the luminal surface for studying host-pathogen interactions.
- Necrotizing enterocolitis is characterized by significant changes in intestinal epithelial cell permeability.
Purpose of the Study:
- To develop and validate a neonatal apical-out (AO) enteroid model for studying necrotizing enterocolitis (NEC).
- To assess the utility of lucifer yellow (LY) as a marker for paracellular permeability in the AO NEC model.
- To compare the AO model with the traditional BO model for investigating NEC-related permeability changes and host-pathogen interactions.
Main Methods:
- Human intestinal stem cells from preterm neonates were used to generate enteroids.
- Enteroids were cultured to maturity and then either maintained in basolateral-out (BO) or transformed into apical-out (AO) conformation.
- In vitro necrotizing enterocolitis (NEC) was induced using lipopolysaccharide (LPS) and hypoxia; paracellular permeability was assessed using lucifer yellow (LY) and confirmed via immunofluorescence staining for ZO-1 and β-catenin.
Main Results:
- Immunofluorescence confirmed the successful transformation of enteroids into the apical-out (AO) conformation.
- Both AO and BO enteroids treated with LPS and hypoxia exhibited significantly increased paracellular permeability compared to controls.
- Increased uptake of LY into the lumen was observed in treated AO and BO enteroids, indicating heightened permeability.
Conclusions:
- The apical-out (AO) enteroid model provides improved access for studying luminal events in necrotizing enterocolitis (NEC).
- Lucifer yellow (LY) effectively measures paracellular permeability across all three major pathways in the AO NEC model.
- This AO enteroid model facilitates comprehensive investigation of host-pathogen interactions and their impact on intestinal permeability in NEC.
