Publications
Browse peer-reviewed literature, posters, webinars, blog articles, and more showing how we and others are using RepliGut Systems to support discovery.
2025
Pike, Colleen M.; Levi, James A.; Boone, Lauren A.; Peddibhotla, Swetha; Johnson, Jacob; Zwarycz, Bailey; Bunger, Maureen K.; Thelin, William; Boazak, Elizabeth M.
High-throughput assay for predicting diarrhea risk using a 2D human intestinal stem cell-derived model Journal Article
In: Toxicology In Vitro, vol. 106, pp. 106040, 2025, ISSN: 0887-2333.
Abstract | Links | BibTeX | Tags: Adverse events, Diarrhea, Epithelium, High throughput, In vitro model, Intestine
@article{pike_high-throughput_2025,
title = {High-throughput assay for predicting diarrhea risk using a 2D human intestinal stem cell-derived model},
author = {Colleen M. Pike and James A. Levi and Lauren A. Boone and Swetha Peddibhotla and Jacob Johnson and Bailey Zwarycz and Maureen K. Bunger and William Thelin and Elizabeth M. Boazak},
url = {https://www.sciencedirect.com/science/article/pii/S0887233325000347},
doi = {10.1016/j.tiv.2025.106040},
issn = {0887-2333},
year = {2025},
date = {2025-06-01},
urldate = {2025-06-01},
journal = {Toxicology In Vitro},
volume = {106},
pages = {106040},
abstract = {Gastrointestinal toxicities (GITs) in clinical trials often lead to dose-limitations that reduce drug efficacy and delay treatment optimization. Preclinical animal models do not accurately replicate human physiology, leaving few options for early detection of GITs, such as diarrhea, before human studies. Chemotherapeutic agents, known to cause clinical diarrhea, frequently target mitotic cells. Therefore, we hypothesized a model utilizing proliferative cell populations derived from human intestinal crypts would predict clinical diarrhea occurrence with high accuracy. Here, we describe the development of a diarrhea prediction assay utilizing RepliGut® Planar, a primary intestinal stem cell-derived platform. To evaluate the ability of this model to predict clinical diarrhea risk, we assessed toxicity of 30 marketed drugs by measuring cell proliferation (EdU incorporation), cell abundance (nuclei quantification), and barrier formation (TEER) in cells derived from three human donors. Dose response curves were generated for each drug, and the IC15 to Cmax ratio was used to identify a threshold for assay positivity. This model accurately predicted diarrhea potential, achieving an accuracy of 91 % for proliferation, 90 % for abundance, and 88 % for barrier formation. In vitro toxicity screening using primary proliferative cells may reduce clinical diarrhea and ultimately lead to safer and more effective treatments for patients.},
keywords = {Adverse events, Diarrhea, Epithelium, High throughput, In vitro model, Intestine},
pubstate = {published},
tppubtype = {article}
}
Gastrointestinal toxicities (GITs) in clinical trials often lead to dose-limitations that reduce drug efficacy and delay treatment optimization. Preclinical animal models do not accurately replicate human physiology, leaving few options for early detection of GITs, such as diarrhea, before human studies. Chemotherapeutic agents, known to cause clinical diarrhea, frequently target mitotic cells. Therefore, we hypothesized a model utilizing proliferative cell populations derived from human intestinal crypts would predict clinical diarrhea occurrence with high accuracy. Here, we describe the development of a diarrhea prediction assay utilizing RepliGut® Planar, a primary intestinal stem cell-derived platform. To evaluate the ability of this model to predict clinical diarrhea risk, we assessed toxicity of 30 marketed drugs by measuring cell proliferation (EdU incorporation), cell abundance (nuclei quantification), and barrier formation (TEER) in cells derived from three human donors. Dose response curves were generated for each drug, and the IC15 to Cmax ratio was used to identify a threshold for assay positivity. This model accurately predicted diarrhea potential, achieving an accuracy of 91 % for proliferation, 90 % for abundance, and 88 % for barrier formation. In vitro toxicity screening using primary proliferative cells may reduce clinical diarrhea and ultimately lead to safer and more effective treatments for patients.
2022
Breau, Keith A.; Ok, Meryem T.; Gomez-Martinez, Ismael; Burclaff, Joseph; Kohn, Nathan P.; Magness, Scott T.
Efficient transgenesis and homology-directed gene targeting in monolayers of primary human small intestinal and colonic epithelial stem cells Journal Article
In: vol. 17, no. 6, pp. 1493–1506, 2022, ISSN: 2213-6711.
Abstract | Links | BibTeX | Tags: 2D monolayer cultures, electroporation, Gene Editing, Gene Targeting, human {ISC} marker, Humans, Intestine, microphysiological device, Organoids, planar crypt-microarray, Small, stem cells, transfection, transgenic
@article{breau_efficient_2022,
title = {Efficient transgenesis and homology-directed gene targeting in monolayers of primary human small intestinal and colonic epithelial stem cells},
author = {Keith A. Breau and Meryem T. Ok and Ismael Gomez-Martinez and Joseph Burclaff and Nathan P. Kohn and Scott T. Magness},
doi = {10.1016/j.stemcr.2022.04.005},
issn = {2213-6711},
year = {2022},
date = {2022-06-14},
urldate = {2022-06-14},
volume = {17},
number = {6},
pages = {1493–1506},
abstract = {Two-dimensional (2D) cultures of intestinal and colonic epithelium can be generated using human intestinal stem cells (hISCs) derived from primary tissue sources. These 2D cultures are emerging as attractive and versatile alternatives to three-dimensional organoid cultures; however, transgenesis and gene-editing approaches have not been developed for hISCs grown as 2D monolayers. Using 2D cultured hISCs we show that electroporation achieves up to 80% transfection in hISCs from six anatomical regions with around 64% survival and produces 0.15% transgenesis by PiggyBac transposase and 35% gene edited indels by electroporation of Cas9-ribonucleoprotein complexes at the OLFM4 locus. We create OLFM4-emGFP knock-in hISCs, validate the reporter on engineered 2D crypt devices, and develop complete workflows for high-throughput cloning and expansion of transgenic lines in 3-4 weeks. New findings demonstrate small hISCs expressing the highest OLFM4 levels exhibit the most organoid forming potential and show utility of the 2D crypt device to evaluate hISC function.},
keywords = {2D monolayer cultures, electroporation, Gene Editing, Gene Targeting, human {ISC} marker, Humans, Intestine, microphysiological device, Organoids, planar crypt-microarray, Small, stem cells, transfection, transgenic},
pubstate = {published},
tppubtype = {article}
}
Two-dimensional (2D) cultures of intestinal and colonic epithelium can be generated using human intestinal stem cells (hISCs) derived from primary tissue sources. These 2D cultures are emerging as attractive and versatile alternatives to three-dimensional organoid cultures; however, transgenesis and gene-editing approaches have not been developed for hISCs grown as 2D monolayers. Using 2D cultured hISCs we show that electroporation achieves up to 80% transfection in hISCs from six anatomical regions with around 64% survival and produces 0.15% transgenesis by PiggyBac transposase and 35% gene edited indels by electroporation of Cas9-ribonucleoprotein complexes at the OLFM4 locus. We create OLFM4-emGFP knock-in hISCs, validate the reporter on engineered 2D crypt devices, and develop complete workflows for high-throughput cloning and expansion of transgenic lines in 3-4 weeks. New findings demonstrate small hISCs expressing the highest OLFM4 levels exhibit the most organoid forming potential and show utility of the 2D crypt device to evaluate hISC function.
2019
Dutton, Johanna S.; Hinman, Samuel S.; Kim, Raehyun; Wang, Yuli; Allbritton, Nancy L.
Primary Cell-Derived Intestinal Models: Recapitulating Physiology Journal Article
In: Trends in Biotechnology, vol. 37, no. 7, pp. 744, 2019, (Publisher: NIH Public Access).
Abstract | Links | BibTeX | Tags: Biological, Cell Culture Techniques, Cells, Cultured, Humans, in vitro models, Intestine, Intestines, Models, monolayers, organ-on-chips, Organoids, stem cells, Tissue Engineering
@article{dutton_primary_2019,
title = {Primary Cell-Derived Intestinal Models: Recapitulating Physiology},
author = {Johanna S. Dutton and Samuel S. Hinman and Raehyun Kim and Yuli Wang and Nancy L. Allbritton},
url = {https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6571163/},
doi = {10.1016/j.tibtech.2018.12.001},
year = {2019},
date = {2019-07-01},
journal = {Trends in Biotechnology},
volume = {37},
number = {7},
pages = {744},
abstract = {The development of physiologically relevant intestinal models fueled by breakthroughs in primary cell-culture methods, has enabled successful recapitulation of key features of intestinal physiology. These advances when paired with engineering methods, ...},
note = {Publisher: NIH Public Access},
keywords = {Biological, Cell Culture Techniques, Cells, Cultured, Humans, in vitro models, Intestine, Intestines, Models, monolayers, organ-on-chips, Organoids, stem cells, Tissue Engineering},
pubstate = {published},
tppubtype = {article}
}
The development of physiologically relevant intestinal models fueled by breakthroughs in primary cell-culture methods, has enabled successful recapitulation of key features of intestinal physiology. These advances when paired with engineering methods, ...
2017
Wang, Yuli; Gunasekara, Dulan B.; Reed, Mark I.; DiSalvo, Matthew; Bultman, Scott J.; Sims, Christopher E.; Magness, Scott T.; Allbritton, Nancy L.
A microengineered collagen scaffold for generating a polarized crypt-villus architecture of human small intestinal epithelium Journal Article
In: vol. 128, pp. 44–55, 2017, ISSN: 0142-9612.
Abstract | Links | BibTeX | Tags: Crypt, Intestine, Microfabrication, Scaffold, Stem cell, Villus
@article{wang_microengineered_2017,
title = {A microengineered collagen scaffold for generating a polarized crypt-villus architecture of human small intestinal epithelium},
author = {Yuli Wang and Dulan B. Gunasekara and Mark I. Reed and Matthew DiSalvo and Scott J. Bultman and Christopher E. Sims and Scott T. Magness and Nancy L. Allbritton},
url = {https://www.sciencedirect.com/science/article/pii/S0142961217301412},
doi = {10.1016/j.biomaterials.2017.03.005},
issn = {0142-9612},
year = {2017},
date = {2017-06-01},
urldate = {2023-02-20},
volume = {128},
pages = {44–55},
abstract = {The human small intestinal epithelium possesses a distinct crypt-villus architecture and tissue polarity in which proliferative cells reside inside crypts while differentiated cells are localized to the villi. Indirect evidence has shown that the processes of differentiation and migration are driven in part by biochemical gradients of factors that specify the polarity of these cellular compartments; however, direct evidence for gradient-driven patterning of this in vivo architecture has been hampered by limitations of the in vitro systems available. Enteroid cultures are a powerful in vitro system; nevertheless, these spheroidal structures fail to replicate the architecture and lineage compartmentalization found in vivo, and are not easily subjected to gradients of growth factors. In the current work, we report the development of a micropatterned collagen scaffold with suitable extracellular matrix and stiffness to generate an in vitro self-renewing human small intestinal epithelium that replicates key features of the in vivo small intestine: a crypt-villus architecture with appropriate cell-lineage compartmentalization and an open and accessible luminal surface. Chemical gradients applied to the crypt-villus axis promoted the creation of a stem/progenitor-cell zone and supported cell migration along the crypt-villus axis. This new approach combining microengineered scaffolds, biophysical cues and chemical gradients to control the intestinal epithelium ex vivo can serve as a physiologically relevant mimic of the human small intestinal epithelium, and is broadly applicable to model other tissues that rely on gradients for physiological function.},
keywords = {Crypt, Intestine, Microfabrication, Scaffold, Stem cell, Villus},
pubstate = {published},
tppubtype = {article}
}
The human small intestinal epithelium possesses a distinct crypt-villus architecture and tissue polarity in which proliferative cells reside inside crypts while differentiated cells are localized to the villi. Indirect evidence has shown that the processes of differentiation and migration are driven in part by biochemical gradients of factors that specify the polarity of these cellular compartments; however, direct evidence for gradient-driven patterning of this in vivo architecture has been hampered by limitations of the in vitro systems available. Enteroid cultures are a powerful in vitro system; nevertheless, these spheroidal structures fail to replicate the architecture and lineage compartmentalization found in vivo, and are not easily subjected to gradients of growth factors. In the current work, we report the development of a micropatterned collagen scaffold with suitable extracellular matrix and stiffness to generate an in vitro self-renewing human small intestinal epithelium that replicates key features of the in vivo small intestine: a crypt-villus architecture with appropriate cell-lineage compartmentalization and an open and accessible luminal surface. Chemical gradients applied to the crypt-villus axis promoted the creation of a stem/progenitor-cell zone and supported cell migration along the crypt-villus axis. This new approach combining microengineered scaffolds, biophysical cues and chemical gradients to control the intestinal epithelium ex vivo can serve as a physiologically relevant mimic of the human small intestinal epithelium, and is broadly applicable to model other tissues that rely on gradients for physiological function.