Publications
Browse peer-reviewed literature, posters, webinars, blog articles, and more showing how we and others are using RepliGut Systems to support discovery.
2022
Burclaff, Joseph; Bliton, R. Jarrett; Breau, Keith A.; Ok, Meryem T.; Gomez-Martinez, Ismael; Ranek, Jolene S.; Bhatt, Aadra P.; Purvis, Jeremy E.; Woosley, John T.; Magness, Scott T.
A Proximal-to-Distal Survey of Healthy Adult Human Small Intestine and Colon Epithelium by Single-Cell Transcriptomics Journal Article
In: Cell Mol Gastroenterol Hepatol, vol. 13, no. 5, pp. 1554–1589, 2022, ISSN: 2352-345X.
Abstract | Links | BibTeX | Tags: Differentiate Cell Lineage, Enterochromaffin Cells, Enterocytes, Enteroendocrine Cells, Gut barrier function, Intestinal Epithelial Cells
@article{burclaff_proximal–distal_2022,
title = {A Proximal-to-Distal Survey of Healthy Adult Human Small Intestine and Colon Epithelium by Single-Cell Transcriptomics},
author = {Joseph Burclaff and R. Jarrett Bliton and Keith A. Breau and Meryem T. Ok and Ismael Gomez-Martinez and Jolene S. Ranek and Aadra P. Bhatt and Jeremy E. Purvis and John T. Woosley and Scott T. Magness},
url = {https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9043569/},
doi = {10.1016/j.jcmgh.2022.02.007},
issn = {2352-345X},
year = {2022},
date = {2022-02-15},
urldate = {2022-02-15},
journal = {Cell Mol Gastroenterol Hepatol},
volume = {13},
number = {5},
pages = {1554–1589},
abstract = {Background & Aims
Single-cell transcriptomics offer unprecedented resolution of tissue function at the cellular level, yet studies analyzing healthy adult human small intestine and colon are sparse. Here, we present single-cell transcriptomics covering the duodenum, jejunum, ileum, and ascending, transverse, and descending colon from 3 human beings.
Methods
A total of 12,590 single epithelial cells from 3 independently processed organ donors were evaluated for organ-specific lineage biomarkers, differentially regulated genes, receptors, and drug targets. Analyses focused on intrinsic cell properties and their capacity for response to extrinsic signals along the gut axis across different human beings.
Results
Cells were assigned to 25 epithelial lineage clusters. Multiple accepted intestinal stem cell markers do not specifically mark all human intestinal stem cells. Lysozyme expression is not unique to human Paneth cells, and Paneth cells lack expression of expected niche factors. Bestrophin 4 (BEST4)+ cells express Neuropeptide Y (NPY) and show maturational differences between the small intestine and colon. Tuft cells possess a broad ability to interact with the innate and adaptive immune systems through previously unreported receptors. Some classes of mucins, hormones, cell junctions, and nutrient absorption genes show unappreciated regional expression differences across lineages. The differential expression of receptors and drug targets across lineages show biological variation and the potential for variegated responses.
Conclusions
Our study identifies novel lineage marker genes, covers regional differences, shows important differences between mouse and human gut epithelium, and reveals insight into how the epithelium responds to the environment and drugs. This comprehensive cell atlas of the healthy adult human intestinal epithelium resolves likely functional differences across anatomic regions along the gastrointestinal tract and advances our understanding of human intestinal physiology.},
keywords = {Differentiate Cell Lineage, Enterochromaffin Cells, Enterocytes, Enteroendocrine Cells, Gut barrier function, Intestinal Epithelial Cells},
pubstate = {published},
tppubtype = {article}
}
Background & Aims
Single-cell transcriptomics offer unprecedented resolution of tissue function at the cellular level, yet studies analyzing healthy adult human small intestine and colon are sparse. Here, we present single-cell transcriptomics covering the duodenum, jejunum, ileum, and ascending, transverse, and descending colon from 3 human beings.
Methods
A total of 12,590 single epithelial cells from 3 independently processed organ donors were evaluated for organ-specific lineage biomarkers, differentially regulated genes, receptors, and drug targets. Analyses focused on intrinsic cell properties and their capacity for response to extrinsic signals along the gut axis across different human beings.
Results
Cells were assigned to 25 epithelial lineage clusters. Multiple accepted intestinal stem cell markers do not specifically mark all human intestinal stem cells. Lysozyme expression is not unique to human Paneth cells, and Paneth cells lack expression of expected niche factors. Bestrophin 4 (BEST4)+ cells express Neuropeptide Y (NPY) and show maturational differences between the small intestine and colon. Tuft cells possess a broad ability to interact with the innate and adaptive immune systems through previously unreported receptors. Some classes of mucins, hormones, cell junctions, and nutrient absorption genes show unappreciated regional expression differences across lineages. The differential expression of receptors and drug targets across lineages show biological variation and the potential for variegated responses.
Conclusions
Our study identifies novel lineage marker genes, covers regional differences, shows important differences between mouse and human gut epithelium, and reveals insight into how the epithelium responds to the environment and drugs. This comprehensive cell atlas of the healthy adult human intestinal epithelium resolves likely functional differences across anatomic regions along the gastrointestinal tract and advances our understanding of human intestinal physiology.
Single-cell transcriptomics offer unprecedented resolution of tissue function at the cellular level, yet studies analyzing healthy adult human small intestine and colon are sparse. Here, we present single-cell transcriptomics covering the duodenum, jejunum, ileum, and ascending, transverse, and descending colon from 3 human beings.
Methods
A total of 12,590 single epithelial cells from 3 independently processed organ donors were evaluated for organ-specific lineage biomarkers, differentially regulated genes, receptors, and drug targets. Analyses focused on intrinsic cell properties and their capacity for response to extrinsic signals along the gut axis across different human beings.
Results
Cells were assigned to 25 epithelial lineage clusters. Multiple accepted intestinal stem cell markers do not specifically mark all human intestinal stem cells. Lysozyme expression is not unique to human Paneth cells, and Paneth cells lack expression of expected niche factors. Bestrophin 4 (BEST4)+ cells express Neuropeptide Y (NPY) and show maturational differences between the small intestine and colon. Tuft cells possess a broad ability to interact with the innate and adaptive immune systems through previously unreported receptors. Some classes of mucins, hormones, cell junctions, and nutrient absorption genes show unappreciated regional expression differences across lineages. The differential expression of receptors and drug targets across lineages show biological variation and the potential for variegated responses.
Conclusions
Our study identifies novel lineage marker genes, covers regional differences, shows important differences between mouse and human gut epithelium, and reveals insight into how the epithelium responds to the environment and drugs. This comprehensive cell atlas of the healthy adult human intestinal epithelium resolves likely functional differences across anatomic regions along the gastrointestinal tract and advances our understanding of human intestinal physiology.
Gomez-Martinez, Ismael; Bliton, R. Jarrett; Breau, Keith A.; Czerwinski, Michael J.; Williamson, Ian A.; Wen, Jia; Rawls, John F.; Magness, Scott T.
A Planar Culture Model of Human Absorptive Enterocytes Reveals Metformin Increases Fatty Acid Oxidation and Export Journal Article
In: Cellular and Molecular Gastroenterology and Hepatology, vol. 14, no. 2, pp. 409–434, 2022, ISSN: 2352-345X.
Abstract | Links | BibTeX | Tags: Absorptive Enterocyte Monolayers, Caco-2 Cells, Diabetes Mellitus, Drug Screening, Enterocytes, Fatty Acid Oxidation, Fatty Acids, Humans, Hypoglycemic Agents, intestinal stem cells, Metformin, Type 2
@article{gomez-martinez_planar_2022,
title = {A Planar Culture Model of Human Absorptive Enterocytes Reveals Metformin Increases Fatty Acid Oxidation and Export},
author = {Ismael Gomez-Martinez and R. Jarrett Bliton and Keith A. Breau and Michael J. Czerwinski and Ian A. Williamson and Jia Wen and John F. Rawls and Scott T. Magness},
doi = {10.1016/j.jcmgh.2022.04.009},
issn = {2352-345X},
year = {2022},
date = {2022-01-01},
urldate = {2022-01-01},
journal = {Cellular and Molecular Gastroenterology and Hepatology},
volume = {14},
number = {2},
pages = {409–434},
abstract = {BACKGROUND & AIMS: Fatty acid oxidation by absorptive enterocytes has been linked to the pathophysiology of type 2 diabetes, obesity, and dyslipidemia. Caco-2 and organoids have been used to study dietary lipid-handling processes including fatty acid oxidation, but are limited in physiological relevance or preclude simultaneous apical and basal access. Here, we developed a high-throughput planar human absorptive enterocyte monolayer system for investigating lipid handling, and then evaluated the role of fatty acid oxidation in fatty acid export, using etomoxir, C75, and the antidiabetic drug metformin.
METHODS: Single-cell RNA-sequencing, transcriptomics, and lineage trajectory was performed on primary human jejunum. In vivo absorptive enterocyte maturational states informed conditions used to differentiate human intestinal stem cells (ISCs) that mimic in vivo absorptive enterocyte maturation. The system was scaled for high-throughput drug screening. Fatty acid oxidation was modulated pharmacologically and BODIPY (Thermo Fisher Scientific, Waltham, MA) (B)-labeled fatty acids were used to evaluate fatty acid handling via fluorescence and thin-layer chromatography.
RESULTS: Single-cell RNA-sequencing shows increasing expression of lipid-handling genes as absorptive enterocytes mature. Culture conditions promote ISC differentiation into confluent absorptive enterocyte monolayers. Fatty acid-handling gene expression mimics in vivo maturational states. The fatty acid oxidation inhibitor etomoxir decreased apical-to-basolateral export of medium-chain B-C12 and long-chain B-C16 fatty acids, whereas the CPT1 agonist C75 and the antidiabetic drug metformin increased apical-to-basolateral export. Short-chain B-C5 was unaffected by fatty acid oxidation inhibition and diffused through absorptive enterocytes.
CONCLUSIONS: Primary human ISCs in culture undergo programmed maturation. Absorptive enterocyte monolayers show in vivo maturational states and lipid-handling gene expression profiles. Absorptive enterocytes create strong epithelial barriers in 96-Transwell format. Fatty acid export is proportional to fatty acid oxidation. Metformin enhances fatty acid oxidation and increases basolateral fatty acid export, supporting an intestine-specific role.},
keywords = {Absorptive Enterocyte Monolayers, Caco-2 Cells, Diabetes Mellitus, Drug Screening, Enterocytes, Fatty Acid Oxidation, Fatty Acids, Humans, Hypoglycemic Agents, intestinal stem cells, Metformin, Type 2},
pubstate = {published},
tppubtype = {article}
}
BACKGROUND & AIMS: Fatty acid oxidation by absorptive enterocytes has been linked to the pathophysiology of type 2 diabetes, obesity, and dyslipidemia. Caco-2 and organoids have been used to study dietary lipid-handling processes including fatty acid oxidation, but are limited in physiological relevance or preclude simultaneous apical and basal access. Here, we developed a high-throughput planar human absorptive enterocyte monolayer system for investigating lipid handling, and then evaluated the role of fatty acid oxidation in fatty acid export, using etomoxir, C75, and the antidiabetic drug metformin.
METHODS: Single-cell RNA-sequencing, transcriptomics, and lineage trajectory was performed on primary human jejunum. In vivo absorptive enterocyte maturational states informed conditions used to differentiate human intestinal stem cells (ISCs) that mimic in vivo absorptive enterocyte maturation. The system was scaled for high-throughput drug screening. Fatty acid oxidation was modulated pharmacologically and BODIPY (Thermo Fisher Scientific, Waltham, MA) (B)-labeled fatty acids were used to evaluate fatty acid handling via fluorescence and thin-layer chromatography.
RESULTS: Single-cell RNA-sequencing shows increasing expression of lipid-handling genes as absorptive enterocytes mature. Culture conditions promote ISC differentiation into confluent absorptive enterocyte monolayers. Fatty acid-handling gene expression mimics in vivo maturational states. The fatty acid oxidation inhibitor etomoxir decreased apical-to-basolateral export of medium-chain B-C12 and long-chain B-C16 fatty acids, whereas the CPT1 agonist C75 and the antidiabetic drug metformin increased apical-to-basolateral export. Short-chain B-C5 was unaffected by fatty acid oxidation inhibition and diffused through absorptive enterocytes.
CONCLUSIONS: Primary human ISCs in culture undergo programmed maturation. Absorptive enterocyte monolayers show in vivo maturational states and lipid-handling gene expression profiles. Absorptive enterocytes create strong epithelial barriers in 96-Transwell format. Fatty acid export is proportional to fatty acid oxidation. Metformin enhances fatty acid oxidation and increases basolateral fatty acid export, supporting an intestine-specific role.
METHODS: Single-cell RNA-sequencing, transcriptomics, and lineage trajectory was performed on primary human jejunum. In vivo absorptive enterocyte maturational states informed conditions used to differentiate human intestinal stem cells (ISCs) that mimic in vivo absorptive enterocyte maturation. The system was scaled for high-throughput drug screening. Fatty acid oxidation was modulated pharmacologically and BODIPY (Thermo Fisher Scientific, Waltham, MA) (B)-labeled fatty acids were used to evaluate fatty acid handling via fluorescence and thin-layer chromatography.
RESULTS: Single-cell RNA-sequencing shows increasing expression of lipid-handling genes as absorptive enterocytes mature. Culture conditions promote ISC differentiation into confluent absorptive enterocyte monolayers. Fatty acid-handling gene expression mimics in vivo maturational states. The fatty acid oxidation inhibitor etomoxir decreased apical-to-basolateral export of medium-chain B-C12 and long-chain B-C16 fatty acids, whereas the CPT1 agonist C75 and the antidiabetic drug metformin increased apical-to-basolateral export. Short-chain B-C5 was unaffected by fatty acid oxidation inhibition and diffused through absorptive enterocytes.
CONCLUSIONS: Primary human ISCs in culture undergo programmed maturation. Absorptive enterocyte monolayers show in vivo maturational states and lipid-handling gene expression profiles. Absorptive enterocytes create strong epithelial barriers in 96-Transwell format. Fatty acid export is proportional to fatty acid oxidation. Metformin enhances fatty acid oxidation and increases basolateral fatty acid export, supporting an intestine-specific role.