A Novel Human Epithelial Enteroid Model of Necrotizing Enterocolitis
April 10th, 2019
7.4K views
Enteroids are emerging as a novel model in the study of human disease. The protocol describes how to simulate an enteroid model of human necrotizing enterocolitis using lipopolysaccharide (LPS) treatment of enteroids generated from neonatal tissue. Collected enteroids demonstrate inflammatory changes akin to those seen in human necrotizing enterocolitis.
Overview
Enteroids are three-dimensional organoids derived from intestinal epithelial cells, providing a novel model for studying human necrotizing enterocolitis. This protocol outlines the culture of human enteroids from neonatal tissue and the effects of lipopolysaccharide treatment.
Key Study Components
Area of Science
- Neuroscience
- Biology
- Gastroenterology
Background
- Enteroids mimic the complex physiology of human intestinal epithelium.
- They are derived from intestinal stem cells, making them biologically relevant.
- Studying necrotizing enterocolitis in humans poses ethical challenges, especially in children.
- Animal models and traditional cell-based systems have limitations that enteroids can overcome.
Purpose of Study
- To develop a model for studying necrotizing enterocolitis using human neonatal tissue.
- To investigate inflammatory responses in enteroids similar to those in human disease.
- To provide a platform for testing potential therapeutic agents.
Methods Used
- Isolation of intestinal stem cells from patients undergoing bowel resection.
- Culturing human enteroids in vitro.
- Co-administration of lipopolysaccharide to induce inflammation.
- Assessment of histologic, genetic, and protein expression changes.
Main Results
- Co-treatment with lipopolysaccharide resulted in inflammatory changes in enteroids.
- Histological and genetic alterations were observed, mirroring human necrotizing enterocolitis.
- Enteroids successfully recapitulated multiple cell types of the intestinal epithelium.
- The model demonstrates potential for future therapeutic research.
Conclusions
- Enteroids are a promising model for studying human intestinal diseases.
- This approach allows for ethical research alternatives in pediatric populations.
- Future studies can leverage this model to explore treatment options for necrotizing enterocolitis.
Frequently Asked Questions
What are enteroids?
Enteroids are three-dimensional organoids derived from intestinal epithelial cells, used to study human intestinal diseases.
Why are enteroids important for studying necrotizing enterocolitis?
They provide a biologically relevant model that overcomes the limitations of traditional animal models and allows for ethical research.
How are enteroids cultured?
Enteroids are cultured from intestinal stem cells isolated from patients, allowing for the study of complex physiological interactions.
What role does lipopolysaccharide play in this study?
Lipopolysaccharide is used to induce an inflammatory response in the enteroids, mimicking conditions seen in human necrotizing enterocolitis.
What are the main findings of the study?
The study found that enteroids exhibit inflammatory changes and alterations similar to those in human necrotizing enterocolitis when treated with lipopolysaccharide.
What are the potential applications of this research?
This research can lead to the development of therapeutic agents for treating necrotizing enterocolitis and improve understanding of intestinal diseases.
Executive Industry Relevance
Necrotizing enterocolitis (NEC) presents a significant unmet need in neonatal care, with limited therapeutic options and high morbidity. A human-derived enteroid model offers a biologically relevant platform for target validation and mechanistic de-risking in early discovery. This system enables predictive confidence in screening and preclinical evaluation by recapitulating human intestinal epithelium pathophysiology.
Strategic Applications in Biopharma R&D
Early Discovery & Target Validation
- Scientific Value: Enables interrogation of therapeutic hypotheses in a human-relevant epithelial system.
- Operational Value: Supports functional target validation through LPS-induced inflammatory response modeling.
- Predictive Value: Facilitates biological de-risking by mirroring histologic, genetic, and protein expression alterations seen in human NEC.
Screening & Assay Development
- Scientific Value: Provides a standardized, reproducible 3D culture system for compound screening.
- Operational Value: Yields quantitative readouts via histologic, genetic, and protein expression analysis.
- Scalability: Supports platform reuse across multiple wells and timepoints for dose-response assessment.
Translational & Preclinical Research
- Scientific Value: Bridges discovery to preclinical work by maintaining human tissue relevance.
- Operational Value: Enables risk-adjusted advancement decisions based on epithelial integrity and inflammatory markers.
- Translational Continuity: Supports biomarker alignment through TLR4 and apoptosis readouts.
Pipeline & Workflow Integration
The enteroid model integrates into the discovery continuum from target validation through lead identification and preclinical assessment, offering a human-relevant alternative to animal models.
- Discovery Biology: Supports hypothesis testing via LPS-induced inflammation and epithelial restitution impairment.
- Screening: Delivers assay readiness through polarized enteroid formation and lumen development.
- Analytics: Enables comparative analysis via qRT-PCR, western blotting, and histologic scoring.
- Translational Research: Connects to preclinical validation through NEC-like phenotypic outcomes.
- Enterprise Reuse: Establishes a scalable, bankable platform for iterative therapeutic testing.
Operational & Enterprise Impact
- Scientific Value: Predictive confidence in target validation through human epithelial pathophysiology modeling.
- Operational Value: Standardization and reproducibility via defined crypt isolation and matrix embedding.
- Strategic Value: Improved go/no-go decisions by reducing reliance on non-predictive animal models.
- Portfolio Impact: Risk-adjusted prioritization based on human-relevant inflammatory and apoptotic responses.
Implementation Considerations
- Requires expertise in intestinal stem cell isolation and 3D organoid culture.
- Dependent on sterile tissue handling, basement membrane matrix, and CO2 incubator infrastructure.
- Necessitates cross-team standardization for consistent enteroid formation and LPS treatment timing.
- Adaptation considerations include donor variability and gestational age-specific epithelial responses.
- Practical limitations include culture duration (10+ days for budding) and yield variability post-LPS treatment.
Biopharma FAQs
Why does LPS-induced inflammation matter for target validation in NEC?
LPS treatment triggers histologic, genetic, and protein expression changes in enteroids that mirror human NEC, enabling mechanistic de-risking of targets involved in epithelial injury and inflammatory pathways.
How does isolating intestinal stem cells support discovery pipeline integration?
Isolating stem cells from resected human tissue ensures biologically relevant enteroid models that recapitulate native epithelial complexity, supporting hypothesis testing in early discovery.
What quantitative measurements enable assessment of enteroid integrity and injury?
Apoptosis rates, actin cytoskeleton integrity, lumen formation, and TLR4 expression provide quantifiable outputs to compare control and LPS-treated conditions for screening decisions.
Why do replication requirements matter for cross-functional collaboration in enteroid models?
Consistent enteroid formation across replicates ensures reliable inflammatory response data, enabling alignment between discovery, screening, and preclinical teams on target validation outcomes.
What statistical analysis capabilities are required before implementing this model in screening?
The model requires comparative statistical analysis of histologic scoring, gene expression (qRT-PCR), and protein levels (western blot) to determine significant differences between control and LPS-treated groups.
