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Real-time Measurement of Epithelial Barrier Permeability in Human Intestinal Organoids
Published on: December 18, 2017
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Development of Physiologically Responsive Human iPSC-Derived Intestinal Epithelium to Study Barrier Dysfunction in
John P Gleeson1, Hannah Q Estrada1, Michifumi Yamashita2
1Board of Governors Regenerative Medicine Institute, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA.
International Journal of Molecular Sciences
|February 26, 2020
Summary
Induced pluripotent stem cell-derived organoids model intestinal barrier dysfunction in inflammatory bowel disease (IBD). These models show increased permeability and altered junction proteins when exposed to inflammatory cytokines, aiding IBD research.
Area of Science:
- Gastroenterology and Regenerative Medicine
- Stem Cell Biology
- Epithelial Biology
Background:
- Inflammatory bowel disease (IBD) is associated with increased intestinal permeability, but its genetic basis is poorly understood due to limited modeling systems.
- Studying epithelial barrier dysfunction in IBD requires advanced in vitro models that recapitulate human intestinal physiology.
Purpose of the Study:
- To develop and validate induced pluripotent stem cell (iPSC)-derived human intestinal organoids (HIOs) and human colonic organoids (HCOs) as a model for studying intestinal barrier dysfunction in IBD.
- To investigate the impact of pro-inflammatory cytokines on epithelial barrier integrity and tight/adherens junction proteins in iPSC-derived organoids.
Main Methods:
- Generation of iPSCs from healthy controls and IBD patients (adult-onset and very early-onset IBD [VEO-IBD]).
- Differentiation of iPSCs into HIOs and HCOs, followed by seeding onto Transwell inserts for barrier integrity studies.
- Assessment of barrier function using transepithelial electrical resistance (TEER) measurements, quantitative real-time PCR (qRT-PCR), transmission electron microscopy (TEM), and immunofluorescence.
Main Results:
- HCOs exhibited higher gene expression of CDX2, CD147, and CA2, and increased basal TEER compared to HIOs.
- Exposure to TNFα and IFNγ significantly increased FD4 permeability in both HIO- and HCO-derived epithelia.
- TEM and immunofluorescence revealed mislocalization of E-cadherin and ZO-1, with decreased mRNA expression, upon cytokine stimulation.
Conclusions:
- iPSC-derived HIOs and HCOs provide a physiologically relevant and responsive in vitro model for investigating epithelial barrier dysfunction in IBD and VEO-IBD.
- These organoid models effectively mimic cytokine-induced barrier defects, offering a platform for studying the molecular mechanisms underlying IBD pathogenesis.
