A comparative study of organoid-derived and cell line-derived intestinal epithelial models
Melis Asal, Nanne J. Paauw, Hetty J. Bontkes, Sandra J. van Vliet, Reina E. Mebius, Susan Gibbs
Abstract
Background
In vitro intestinal epithelial models are being developed for studying gut physiology, barrier function, and drug transport. There are two major in vitro models which are currently used: cell line-derived monolayers (such as Caco-2/HT29 cocultures) and primary cell monolayers derived from human intestinal organoids.
Introduction
The native small intestinal epithelium is a highly specialized, dynamic tissue whose hallmark villus-crypt architecture and heterogeneous cellular composition together amplify the absorptive surface area, strengthen barrier function, and provide structural and functional cues. The epithelial layer consists of multiple differentiated cell types, primarily absorptive enterocytes, which form dense apical microvilli to facilitate nutrient uptake.
Materials and methods
Tissue specimens
Human duodenal tissues were obtained as residual material from Whipple procedures through the Amsterdam University Medical Center biobank. All tissue collection and use were conducted in accordance with the ‘Code of Conduct for Health Research’ established by COREON (Commissie Regelgeving Onderzoek), with written informed consent from patients and approval from the local medical ethics committee of the Amsterdam University Medical Center.
Results
Comparison of structural characteristics of cell line and organoid-derived epithelial models
Epithelial monolayers derived from cell lines or organoids grown on Transwell inserts were compared for morphological and cell type specific (enterocyte and goblet cell) similarities and differences (Fig 1). Both epithelial models formed a confluent layer over the Transwell membrane.
Discussion
In this study, we present a direct comparison of two organotypic intestinal monolayer models composed of a cell line-derived or organoid-derived monolayer epithelium. While previous studies have assessed some features of Caco-2/HT29 versus organoid-derived models [14,18], our study directly compares structural and biochemical properties between the models using TEER and permeability, immunofluorescence for VIL, MUC2, ZO-1, SEM and TEM as well as secretome analysis thus providing a comprehensive side-by-side comparison of morphology and barrier-related physiology of the two models.
Acknowledgments
We acknowledge the Microscopy and Cytometry Core Facility at the Amsterdam UMC (Location VUmc) for providing assistance in microscopy experiments.
Citation: Asal M, Paauw NJ, Bontkes HJ, van Vliet SJ, Mebius RE, Gibbs S (2026) A comparative study of organoid-derived and cell line-derived intestinal epithelial models. PLoS One 21(4): e0346560. https://doi.org/10.1371/journal.pone.0346560
Editor: Paul Kwong-Hang Tam, Macau University of Science and Technology, HONG KONG
Received: November 27, 2025; Accepted: March 20, 2026; Published: April 16, 2026
Copyright: © 2026 Asal et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Data Availability: All raw data files are publicly available from the Mendeley Data repository, doi: 10.17632/44rmzbdmgp.1.
Funding: This project has received funding from the European Union’s Horizon 2020 research and innovation program under grant agreement No 847551. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Competing interests: The authors have declared that no competing interests exist.
