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Long-term expansion of epithelial organoids from human colon, adenoma, adenocarcinoma, and Barrett's epithelium.
Literature Information
| DOI | 10.1053/j.gastro.2011.07.050 |
|---|---|
| PMID | 21889923 |
| Journal | Gastroenterology |
| Impact Factor | 25.1 |
| JCR Quartile | Q1 |
| Publication Year | 2011 |
| Times Cited | 2007 |
| Keywords | epithelial organoids, human colon, adenoma, adenocarcinoma, Barrett's esophagus |
| Literature Type | Journal Article, Research Support, Non-U.S. Gov't |
| ISSN | 0016-5085 |
| Pages | 1762-72 |
| Issue | 141(5) |
| Authors | Toshiro Sato, Daniel E Stange, Marc Ferrante, Robert G J Vries, Johan H Van Es, Stieneke Van den Brink, Winan J Van Houdt, Apollo Pronk, Joost Van Gorp, Peter D Siersema, Hans Clevers |
TL;DR
This study developed optimized culture conditions for expanding epithelial organoids from mouse colon and human intestinal tissues, revealing that the addition of specific growth factors enables indefinite expansion of these cells. The findings suggest potential applications for studying gastrointestinal diseases and regenerative biology, with implications indicating that adult stem cells may not have inherent replicative limits in ex vivo conditions.
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epithelial organoids · human colon · adenoma · adenocarcinoma · Barrett's esophagus
Abstract
BACKGROUND & AIMS We previously established long-term culture conditions under which single crypts or stem cells derived from mouse small intestine expand over long periods. The expanding crypts undergo multiple crypt fission events, simultaneously generating villus-like epithelial domains that contain all differentiated types of cells. We have adapted the culture conditions to grow similar epithelial organoids from mouse colon and human small intestine and colon.
METHODS Based on the mouse small intestinal culture system, we optimized the mouse and human colon culture systems.
RESULTS Addition of Wnt3A to the combination of growth factors applied to mouse colon crypts allowed them to expand indefinitely. Addition of nicotinamide, along with a small molecule inhibitor of Alk and an inhibitor of p38, were required for long-term culture of human small intestine and colon tissues. The culture system also allowed growth of mouse Apc-deficient adenomas, human colorectal cancer cells, and human metaplastic epithelia from regions of Barrett's esophagus.
CONCLUSIONS We developed a technology that can be used to study infected, inflammatory, or neoplastic tissues from the human gastrointestinal tract. These tools might have applications in regenerative biology through ex vivo expansion of the intestinal epithelia. Studies of these cultures indicate that there is no inherent restriction in the replicative potential of adult stem cells (or a Hayflick limit) ex vivo.
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Primary Questions Addressed
- What specific factors contribute to the successful long-term culture of human colon organoids compared to mouse models?
- How do the characteristics of adenoma and adenocarcinoma organoids differ in their growth and differentiation in culture?
- What potential applications do these cultured organoids have in understanding gastrointestinal diseases beyond regenerative biology?
- How does the addition of Wnt3A and other growth factors influence the signaling pathways involved in organoid expansion?
- What challenges might arise when translating findings from these organoid cultures to in vivo models of colorectal cancer?
Key Findings
Key Insights
Research Background and Purpose: The study is built upon previous work that established long-term culture conditions for mouse small intestine crypts and stem cells. The primary aim was to adapt these conditions for the expansion of epithelial organoids from various human gastrointestinal tissues, including the colon, adenomas, adenocarcinomas, and Barrett's esophagus. This research seeks to provide a robust platform for studying the biology of these tissues, particularly in the context of diseases such as cancer and inflammation.
Main Methods and Findings: The researchers optimized culture systems for both mouse and human colon tissues by building on the established mouse small intestinal model. A key modification involved the addition of Wnt3A to the growth factor mix for mouse colon crypts, enabling indefinite expansion. For human tissues, the inclusion of nicotinamide, along with a small molecule inhibitor targeting Alk and an inhibitor of p38, was necessary for long-term culture. These optimized conditions facilitated the growth of mouse Apc-deficient adenomas, human colorectal cancer cells, and human metaplastic epithelia derived from Barrett's esophagus, demonstrating the versatility of the culture system across different types of gastrointestinal epithelium.
Core Conclusions: The study successfully developed a novel technology for the long-term culture of various human gastrointestinal tissues, indicating the absence of a replicative limit (Hayflick limit) in adult stem cells under these specific ex vivo conditions. This finding challenges existing notions about the replicative potential of adult stem cells and opens new avenues for research into the regenerative capabilities of intestinal tissues.
Research Significance and Impact: The implications of this research are significant for both regenerative medicine and cancer biology. By providing a reliable method to grow and study human gastrointestinal epithelial tissues, the findings enable detailed investigations of infection, inflammation, and neoplastic transformations in a controlled environment. Furthermore, the ability to expand organoids indefinitely without a replicative limit can facilitate advancements in regenerative therapies and improve our understanding of intestinal diseases. Overall, this work lays the groundwork for future studies aimed at harnessing the potential of epithelial organoids for therapeutic applications and deeper insights into gastrointestinal pathophysiology.
Literatures Citing This Work
- Controlled gene expression in primary Lgr5 organoid cultures. - Bon-Kyoung Koo;Daniel E Stange;Toshiro Sato;Wouter Karthaus;Henner F Farin;Meritxell Huch;Johan H van Es;Hans Clevers - Nature methods (2011)
- Translational approaches for pharmacotherapy development for acute diarrhea. - Mark Donowitz;David H Alpers;Henry J Binder;Thomas Brewer;Jill Carrington;Michael J Grey - Gastroenterology (2012)
- Identification of a cKit(+) colonic crypt base secretory cell that supports Lgr5(+) stem cells in mice. - Michael E Rothenberg;Ysbrand Nusse;Tomer Kalisky;John J Lee;Piero Dalerba;Ferenc Scheeren;Neethan Lobo;Subhash Kulkarni;Sopheak Sim;Dalong Qian;Philip A Beachy;Pankaj J Pasricha;Stephen R Quake;Michael F Clarke - Gastroenterology (2012)
- Functional engraftment of colon epithelium expanded in vitro from a single adult Lgr5⁺ stem cell. - Shiro Yui;Tetsuya Nakamura;Toshiro Sato;Yasuhiro Nemoto;Tomohiro Mizutani;Xiu Zheng;Shizuko Ichinose;Takashi Nagaishi;Ryuichi Okamoto;Kiichiro Tsuchiya;Hans Clevers;Mamoru Watanabe - Nature medicine (2012)
- Review: Experimental models for Barrett's esophagus and esophageal adenocarcinoma. - Katherine S Garman;Roy C Orlando;Xiaoxin Chen - American journal of physiology. Gastrointestinal and liver physiology (2012)
- The R-spondin protein family. - Wim B M de Lau;Berend Snel;Hans C Clevers - Genome biology (2012)
- A nomenclature for intestinal in vitro cultures. - Matthias Stelzner;Michael Helmrath;James C Y Dunn;Susan J Henning;Courtney W Houchen;Calvin Kuo;John Lynch;Linheng Li;Scott T Magness;Martin G Martin;Melissa H Wong;Jian Yu; - American journal of physiology. Gastrointestinal and liver physiology (2012)
- Cellular origin of Barrett's esophagus: controversy and therapeutic implications. - Wa Xian;Khek Yu Ho;Christopher P Crum;Frank McKeon - Gastroenterology (2012)
- Intestinal crypts reproducibly expand in culture. - Megan K Fuller;Denver M Faulk;Nambirajan Sundaram;Noah F Shroyer;Susan J Henning;Michael A Helmrath - The Journal of surgical research (2012)
- Stem cells: One step closer to gut repair. - Anisa Shaker;Deborah C Rubin - Nature (2012)
... (1997 more literatures)
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