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Cordeiro S, Oliveira BB, Valente R, Ferreira D, Luz A, Baptista PV, Fernandes AR. Breaking the mold: 3D cell cultures reshaping the future of cancer research. Front Cell Dev Biol 2024; 12:1507388. [PMID: 39659521 PMCID: PMC11628512 DOI: 10.3389/fcell.2024.1507388] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/07/2024] [Accepted: 11/13/2024] [Indexed: 12/12/2024] Open
Abstract
Despite extensive efforts to unravel tumor behavior and develop anticancer therapies, most treatments fail when advanced to clinical trials. The main challenge in cancer research has been the absence of predictive cancer models, accurately mimicking the tumoral processes and response to treatments. The tumor microenvironment (TME) shows several human-specific physical and chemical properties, which cannot be fully recapitulated by the conventional 2D cell cultures or the in vivo animal models. These limitations have driven the development of novel in vitro cancer models, that get one step closer to the typical features of in vivo systems while showing better species relevance. This review introduces the main considerations required for developing and exploiting tumor spheroids and organoids as cancer models. We also detailed their applications in drug screening and personalized medicine. Further, we show the transition of these models into novel microfluidic platforms, for improved control over physiological parameters and high-throughput screening. 3D culture models have provided key insights into tumor biology, more closely resembling the in vivo TME and tumor characteristics, while enabling the development of more reliable and precise anticancer therapies.
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Affiliation(s)
- Sandra Cordeiro
- UCIBIO, Departamento de Ciências da Vida, Faculdade de Ciências e Tecnologia, Universidade NOVA de Lisboa, Caparica, Portugal
- i4HB, Associate Laboratory – Institute for Health and Bioeconomy, Faculdade de Ciências e Tecnologia, Universidade NOVA de Lisboa, Caparica, Portugal
| | - Beatriz B. Oliveira
- UCIBIO, Departamento de Ciências da Vida, Faculdade de Ciências e Tecnologia, Universidade NOVA de Lisboa, Caparica, Portugal
- i4HB, Associate Laboratory – Institute for Health and Bioeconomy, Faculdade de Ciências e Tecnologia, Universidade NOVA de Lisboa, Caparica, Portugal
| | - Ruben Valente
- UCIBIO, Departamento de Ciências da Vida, Faculdade de Ciências e Tecnologia, Universidade NOVA de Lisboa, Caparica, Portugal
- i4HB, Associate Laboratory – Institute for Health and Bioeconomy, Faculdade de Ciências e Tecnologia, Universidade NOVA de Lisboa, Caparica, Portugal
| | - Daniela Ferreira
- UCIBIO, Departamento de Ciências da Vida, Faculdade de Ciências e Tecnologia, Universidade NOVA de Lisboa, Caparica, Portugal
- i4HB, Associate Laboratory – Institute for Health and Bioeconomy, Faculdade de Ciências e Tecnologia, Universidade NOVA de Lisboa, Caparica, Portugal
| | - André Luz
- UCIBIO, Departamento de Ciências da Vida, Faculdade de Ciências e Tecnologia, Universidade NOVA de Lisboa, Caparica, Portugal
- i4HB, Associate Laboratory – Institute for Health and Bioeconomy, Faculdade de Ciências e Tecnologia, Universidade NOVA de Lisboa, Caparica, Portugal
| | - Pedro V. Baptista
- UCIBIO, Departamento de Ciências da Vida, Faculdade de Ciências e Tecnologia, Universidade NOVA de Lisboa, Caparica, Portugal
- i4HB, Associate Laboratory – Institute for Health and Bioeconomy, Faculdade de Ciências e Tecnologia, Universidade NOVA de Lisboa, Caparica, Portugal
| | - Alexandra R. Fernandes
- UCIBIO, Departamento de Ciências da Vida, Faculdade de Ciências e Tecnologia, Universidade NOVA de Lisboa, Caparica, Portugal
- i4HB, Associate Laboratory – Institute for Health and Bioeconomy, Faculdade de Ciências e Tecnologia, Universidade NOVA de Lisboa, Caparica, Portugal
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Choi H, Kwak MJ, Kang AN, Mun D, Lee S, Park MR, Oh S, Kim Y. Limosilactobacillus fermentum SLAM 216-Derived Extracellular Vesicles Promote Intestinal Maturation in Mouse Organoid Models. J Microbiol Biotechnol 2024; 34:2091-2099. [PMID: 39252638 PMCID: PMC11540603 DOI: 10.4014/jmb.2405.05028] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/25/2024] [Revised: 07/23/2024] [Accepted: 07/31/2024] [Indexed: 09/11/2024]
Abstract
Probiotics, when consumed in adequate amounts, can promote the health of the host and beneficially modulate the host's immunity. Particularly during the host's early life, the gut intestine undergoes a period of epithelial maturation in which epithelial cells organize into specific crypt and villus structures. This process can be mediated by the gut microbiota. Recent studies have reported that the administration of probiotics can further promote intestinal maturation in the neonatal intestine. Therefore, in this study, we investigated the effects of extracellular vesicles derived from the Limosilactobacillus fermentum SLAM 216 strain, which is an established probiotic with known immune and anti-aging effects on intestinal epithelial maturation and homeostasis, using mouse small intestinal organoids. As per our findings, treatment with L. fermentum SLAM 216-derived LF216EV (LF216EV) has significantly increased the bud number and size of organoid buds. Furthermore, extracellular vesicle (EV) treatment upregulated the expression of maturation-related genes, including Ascl2, Ephb2, Lgr5, and Sox9. Tight junctions are known to have an important role in the intestinal immune barrier, and EV treatment has significantly increased the expression of genes associated with tight junctions, such as Claudin, Muc2, Occludin, and Zo-1, indicating that it can promote intestinal development. This was supported by RNA sequencing, which revealed the upregulation of genes associated with cAMP-mediated signaling, which is known to regulate cellular processes including cell differentiation. Additionally, organoids exposed to LF216EV exhibited upregulation of genes associated with maintaining brain memory and neurotransmission, suggesting possible future functional implications.
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Affiliation(s)
- Hyejin Choi
- Department of Agricultural Biotechnology and Research Institute of Agriculture and Life Science, Seoul National University, Seoul 08826, Republic of Korea
| | - Min-Jin Kwak
- Department of Agricultural Biotechnology and Research Institute of Agriculture and Life Science, Seoul National University, Seoul 08826, Republic of Korea
| | - An Na Kang
- Department of Agricultural Biotechnology and Research Institute of Agriculture and Life Science, Seoul National University, Seoul 08826, Republic of Korea
| | - Daye Mun
- Department of Agricultural Biotechnology and Research Institute of Agriculture and Life Science, Seoul National University, Seoul 08826, Republic of Korea
| | - Suengwon Lee
- Department of Pharmacology and Systems Physiology, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA
| | - Mi Ri Park
- Korea Food Research Institute, Wanju 55365, Republic of Korea
| | - Sangnam Oh
- Department of Functional Food and Biotechnology, Jeonju University, Jeonju 55069, Republic of Korea
| | - Younghoon Kim
- Department of Agricultural Biotechnology and Research Institute of Agriculture and Life Science, Seoul National University, Seoul 08826, Republic of Korea
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Wang X, Jing Y, Zheng C, Huang C, Yao H, Guo Z, Wu Y, Wang Z, Wu Z, Ge R, Cheng W, Yan Y, Jiang S, Sun J, Li J, Xie Q, Li X, Wang H. Using integrated transcriptomics and metabolomics to explore the effects of infant formula on the growth and development of small intestinal organoids. Food Funct 2024; 15:9191-9209. [PMID: 39158038 DOI: 10.1039/d4fo01723d] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 08/20/2024]
Abstract
Infant formulas are designed to provide sufficient energy and the necessary nutrients to support the growth and development of newborns. Currently, research on the functions of formula milk powder focuses on clinical research and cell experiments, and there were many cell experiments that investigated the effect of infant formulas on cellular growth. However, most of the cells used are tumor cell lines, which are unable to simulate the real digestion process of an infant. In this study, we innovatively proposed a method that integrates human small intestinal organoids (SIOs) with transcriptomics and metabolomics analysis. We induced directed differentiation of human embryonic stem cells into SIOs and simulated the intestinal environment of newborns with them. Then, three kinds of 1-stage infant formulas from the same brand were introduced to simulate the digestion, absorption, and metabolism of the infant intestine. The nutritional value of each formula milk powder was examined by multi-omics sequencing methods, including transcriptomics and metabolomics analysis. Results showed that there were significant alterations in gene expression and metabolites in the three groups of SIOs after absorbing different infant formulas. By analyzing transcriptome and metabolome data, combined with GO, KEGG, and GSEA analysis, we demonstrated the ability of SIOs to model the different aspects of the developing process of the intestine and discovered the correlation between formula components and their effects, including Lactobacillus lactis and lactoferrin. The study reveals the effect and mechanisms of formula milk powder on the growth and development of infant intestines and the formation of immune function. Furthermore, our method can help to construct a multi-level assessment model, detect the effects of nutrients, and evaluate the interactions between nutrients, which is helpful for future research and development of infant powders.
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Affiliation(s)
- Xianli Wang
- School of Public Health, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Yuxin Jing
- Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Chengdong Zheng
- Heilongjiang Feihe Dairy Co., Ltd, C-16, 10A Jiuxianqiao Rd, Chaoyang, Beijing 100015, China
| | - Chenxuan Huang
- Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Haiyang Yao
- Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Zimo Guo
- Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Yilun Wu
- Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Zening Wang
- Institutes of Biomedical Sciences, Fudan University, 131 Dongan Road, Shanghai, 200032, China
| | - Zhengyang Wu
- Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Ruihong Ge
- School of Public Health, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Wei Cheng
- School of Public Health, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Yuanyuan Yan
- School of Public Health, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Shilong Jiang
- Heilongjiang Feihe Dairy Co., Ltd, C-16, 10A Jiuxianqiao Rd, Chaoyang, Beijing 100015, China
| | - Jianguo Sun
- Heilongjiang Feihe Dairy Co., Ltd, C-16, 10A Jiuxianqiao Rd, Chaoyang, Beijing 100015, China
| | - Jingquan Li
- School of Public Health, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Qinggang Xie
- Heilongjiang Feihe Dairy Co., Ltd, C-16, 10A Jiuxianqiao Rd, Chaoyang, Beijing 100015, China
| | - Xiaoguang Li
- State Key Laboratory of Systems Medicine for Cancer, Center for Single-Cell Omics, School of Public Health, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China.
| | - Hui Wang
- State Key Laboratory of Systems Medicine for Cancer, Center for Single-Cell Omics, School of Public Health, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China.
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Piraino F, Costa M, Meyer M, Cornish G, Ceroni C, Garnier V, Hoehnel-Ka S, Brandenberg N. Organoid models: the future companions of personalized drug development. Biofabrication 2024; 16:032009. [PMID: 38608454 DOI: 10.1088/1758-5090/ad3e30] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/30/2023] [Accepted: 04/12/2024] [Indexed: 04/14/2024]
Abstract
High failure rates of the current drug development process are driving exemplary changes toward methodologies centered on human diseasein-vitromodeling. Organoids are self-organized tissue sub-units resembling their organ of origin and are widely acknowledged for their unique potential in recapitulating human physio-pathological mechanisms. They are transformative for human health by becoming the platform of choice to probe disease mechanisms and advance new therapies. Furthermore, the compounds' validation as therapeutics represents another point of the drug development pipeline where organoids may provide key understandings and help pharma organizations replace or reduce animal research. In this review, we focus on gastrointestinal organoid models, which are currently the most advanced organoid models in drug development. We focus on experimental validations of their value, and we propose avenues to enhance their use in drug discovery and development, as well as precision medicine and diagnostics.
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Affiliation(s)
| | - Mariana Costa
- Doppl SA, EPFL Innovation Park, Lausanne, Switzerland
| | - Marine Meyer
- Doppl SA, EPFL Innovation Park, Lausanne, Switzerland
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Greiner TU, Koh A, Peris E, Bergentall M, Johansson MEV, Hansson GC, Drucker DJ, Bäckhed F. GLP-1R signaling modulates colonic energy metabolism, goblet cell number and survival in the absence of gut microbiota. Mol Metab 2024; 83:101924. [PMID: 38521185 PMCID: PMC11002751 DOI: 10.1016/j.molmet.2024.101924] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/19/2023] [Revised: 03/11/2024] [Accepted: 03/18/2024] [Indexed: 03/25/2024] Open
Abstract
OBJECTIVES Gut microbiota increases energy availability through fermentation of dietary fibers to short-chain fatty acids in conventionally raised mice. Energy deficiency in germ-free (GF) mice increases glucagon-like peptide-1 (GLP-1) levels, which slows intestinal transit. To further analyze the role of GLP-1-mediated signaling in this model of energy deficiency, we re-derived mice lacking GLP-1 receptor (GLP-1R KO) as GF. METHODS GLP-1R KO mice were rederived as GF through hysterectomy and monitored for 30 weeks. Mice were subjected to rescue experiments either through feeding an energy-rich diet or colonization with a normal cecal microbiota. Histology and intestinal function were assessed at different ages. Intestinal organoids were assessed to investigate stemness. RESULTS Unexpectedly, 25% of GF GLP-1R KO mice died before 20 weeks of age, associated with enlarged ceca, increased cecal water content, increased colonic expression of apical ion transporters, reduced number of goblet cells and loss of colonic epithelial integrity. Colonocytes from GLP-1R KO mice were energy-deprived and exhibited increased ER-stress; mitochondrial fragmentation, increased oxygen levels and loss of stemness. Restoring colonic energy levels either by feeding a Western-style diet or colonization with a normal gut microbiota normalized gut phenotypes and prevented lethality. CONCLUSIONS Our findings reveal a heretofore unrecognized role for GLP-1R signaling in the maintenance of colonic physiology and survival during energy deprivation.
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Affiliation(s)
- Thomas U Greiner
- Wallenberg Laboratory, Department of Molecular and Clinical Medicine, Institute of Medicine, University of Gothenburg, Sweden
| | - Ara Koh
- Wallenberg Laboratory, Department of Molecular and Clinical Medicine, Institute of Medicine, University of Gothenburg, Sweden; Department of Life Sciences, Pohang University of Science and Technology, Pohang 37673 South Korea
| | - Eduard Peris
- Wallenberg Laboratory, Department of Molecular and Clinical Medicine, Institute of Medicine, University of Gothenburg, Sweden
| | - Mattias Bergentall
- Wallenberg Laboratory, Department of Molecular and Clinical Medicine, Institute of Medicine, University of Gothenburg, Sweden
| | - Malin E V Johansson
- Department of Medical Biochemistry and Cell Biology, Institute of Biomedicine, University of Gothenburg, Sweden
| | - Gunnar C Hansson
- Department of Medical Biochemistry and Cell Biology, Institute of Biomedicine, University of Gothenburg, Sweden
| | - Daniel J Drucker
- Department of Medicine, Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, University of Toronto, Toronto ON, Canada
| | - Fredrik Bäckhed
- Wallenberg Laboratory, Department of Molecular and Clinical Medicine, Institute of Medicine, University of Gothenburg, Sweden; Novo Nordisk Foundation Center for Basic Metabolic Research, Faculty of Health Sciences, University of Copenhagen, Denmark; Region Västra Götaland, Sahlgrenska University Hospital, Department of Clinical Physiology, Gothenburg, Sweden.
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6
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Wang Y, Tan R, Chen YG. Organoid Culture of Different Intestinal Segments from Human and Mouse. Methods Mol Biol 2024. [PMID: 38647862 DOI: 10.1007/7651_2024_542] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 04/25/2024]
Abstract
The intestine comprises distinct segments, each characterized by unique cell populations and functions. Intestinal organoids faithfully replicate the cellular composition and functions of the intestine. Over the past decade, the organoid model has garnered considerable attention for its application in investigation of organ development, renewal and functional performance. While the organoid culture systems for mouse small intestine and human large intestine have widely adopted, a comparison summary for different segments of the human or mouse intestine is lacking. In this study, we present a systematically detailed culture methodology for intestinal organoids, encompassing both the small intestine and the large intestine from humans or mice. This method provides a robust in vitro tool for intestinal research, and expands the possible clinical application of organoids.
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Affiliation(s)
- Yalong Wang
- Guangzhou National Laboratory, Guangzhou, China
| | - Ronghui Tan
- Guangzhou National Laboratory, Guangzhou, China
| | - Ye-Guang Chen
- Guangzhou National Laboratory, Guangzhou, China.
- The State Key Laboratory of Membrane Biology Tsinghua-Peking Center for Life Sciences School of Life Sciences, Tsinghua University, Beijing, China.
- Jiangxi Medical College, Nanchang University, Nanchang, China.
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Jiang KL, Jia YB, Liu XJ, Jia QL, Guo LK, Wang XX, Yang KM, Wu CH, Liang BB, Ling JH. Bibliometrics analysis based on the Web of Science: Current trends and perspective of gastric organoid during 2010-2023. World J Gastroenterol 2024; 30:969-983. [PMID: 38516239 PMCID: PMC10950634 DOI: 10.3748/wjg.v30.i8.969] [Citation(s) in RCA: 4] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/25/2023] [Revised: 01/02/2024] [Accepted: 02/01/2024] [Indexed: 02/26/2024] Open
Abstract
BACKGROUND Three-dimensional organoid culture systems have been established as a robust tool for elucidating mechanisms and performing drug efficacy testing. The use of gastric organoid models holds significant promise for advancing personalized medicine research. However, a comprehensive bibliometric review of this bur-geoning field has not yet been published. AIM To analyze and understand the development, impact, and direction of gastric organoid research using bibliometric methods using data from the Web of Science Core Collection (WoSCC) database. METHODS This analysis encompassed literature pertaining to gastric organoids published between 2010 and 2023, as indexed in the WoSCC. CiteSpace and VOSviewer were used to depict network maps illustrating collaborations among authors, institutions and keywords related to gastric organoid. Citation, co-citation, and burst analysis methodologies were applied to assess the impact and progress of research. RESULTS A total of 656 relevant studies were evaluated. The majority of research was published in gastroenterology-focused journals. Globally, Yana Zavros, Hans Clevers, James M Wells, Sina Bartfeld, and Chen Zheng were the 5 most productive authors, while Hans Clevers, Huch Meritxell, Johan H van Es, Marc Van de Wetering, and Sato Toshiro were the foremost influential scientists in this area. Institutions from the University Medical Center Utrecht, Netherlands Institute for Developmental Biology (Utrecht), and University of Cincinnati (Cincinnati, OH, United States) made the most significant contributions. Currently, gastric organoids are used mainly in studies investigating gastric cancer (GC), Helicobacter pylori-infective gastritis, with a focus on the mechanisms of GC, and drug screening tests. CONCLUSION Key focus areas of research using gastric organoids include unraveling disease mechanisms and enhancing drug screening techniques. Major contributions from renowned academic institutions highlight this field's dynamic growth.
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Affiliation(s)
- Kai-Lin Jiang
- Department of Gastroenterology, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 200021, China
- Laboratory of Cancer Biology, University of Oxford, Oxford OX37DQ, United Kingdom
| | - Yue-Bo Jia
- Department of Gastroenterology, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 200021, China
| | - Xue-Jiao Liu
- Department of Gastroenterology, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 200021, China
| | - Qing-Ling Jia
- Department of Gastroenterology, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 200021, China
| | - Li-Kun Guo
- Department of Gastroenterology, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 200021, China
| | - Xiang-Xiang Wang
- School of Traditional Chinese Medicine, Shanghai University of Chinese Medicine, Shanghai 200021, China
| | - Ke-Ming Yang
- Department of Gastroenterology, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 200021, China
| | - Chen-Heng Wu
- Department of Gastroenterology, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 200021, China
| | - Bei-Bei Liang
- Shanghai Key Laboratory of Molecular Imaging, Shanghai University of Medicine and Health Sciences, Shanghai 201318, China
| | - Jiang-Hong Ling
- Department of Gastroenterology, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 200021, China
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Mai S, Inkielewicz-Stepniak I. Graphene Oxide Nanoparticles and Organoids: A Prospective Advanced Model for Pancreatic Cancer Research. Int J Mol Sci 2024; 25:1066. [PMID: 38256139 PMCID: PMC10817028 DOI: 10.3390/ijms25021066] [Citation(s) in RCA: 2] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/03/2023] [Revised: 01/02/2024] [Accepted: 01/04/2024] [Indexed: 01/24/2024] Open
Abstract
Pancreatic cancer, notorious for its grim 10% five-year survival rate, poses significant clinical challenges, largely due to late-stage diagnosis and limited therapeutic options. This review delves into the generation of organoids, including those derived from resected tissues, biopsies, pluripotent stem cells, and adult stem cells, as well as the advancements in 3D printing. It explores the complexities of the tumor microenvironment, emphasizing culture media, the integration of non-neoplastic cells, and angiogenesis. Additionally, the review examines the multifaceted properties of graphene oxide (GO), such as its mechanical, thermal, electrical, chemical, and optical attributes, and their implications in cancer diagnostics and therapeutics. GO's unique properties facilitate its interaction with tumors, allowing targeted drug delivery and enhanced imaging for early detection and treatment. The integration of GO with 3D cultured organoid systems, particularly in pancreatic cancer research, is critically analyzed, highlighting current limitations and future potential. This innovative approach has the promise to transform personalized medicine, improve drug screening efficiency, and aid biomarker discovery in this aggressive disease. Through this review, we offer a balanced perspective on the advancements and future prospects in pancreatic cancer research, harnessing the potential of organoids and GO.
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Affiliation(s)
| | - Iwona Inkielewicz-Stepniak
- Department of Pharmaceutical Pathophysiology, Faculty of Pharmacy, Medical University of Gdańsk, 80-210 Gdańsk, Poland;
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Liu W, Wang Q, Bai Y, Xiao H, Li Z, Wang Y, Wang Q, Yang J, Sun H. Potential Application of Intestinal Organoids in Intestinal Diseases. Stem Cell Rev Rep 2024; 20:124-137. [PMID: 37938407 DOI: 10.1007/s12015-023-10651-w] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 10/30/2023] [Indexed: 11/09/2023]
Abstract
To accurately reveal the scenario and mecahnism of gastrointestinal diseases, the establishment of in vitro models of intestinal diseases and drug screening platforms have become the focus of attention. Over the past few decades, animal models and immortalized cell lines have provided valuable but limited insights into gastrointestinal research. In recent years, the development of intestinal organoid culture system has revolutionized in vitro studies of intestinal diseases. Intestinal organoids are derived from self-renewal and self-organization intestinal stem cells (ISCs), which can replicate the genetic characteristics, functions, and structures of the original tissues. Consequently, they provide new stragety for studying various intestinal diseases in vitro. In the review, we will discuss the culture techniques of intestinal organoids and describe the use of intestinal organoids as research tools for intestinal diseases. The role of intestinal epithelial cells (IECs) played in the pathogenesis of inflammatory bowel diseases (IBD) and the treatment of intestinal epithelial dysfunction will be highlighted. Besides, we review the current knowledge on using intestinal organoids as models to study the pathogenesis of IBD caused by epithelial dysfunction and to develop new therapeutic approaches. Finally, we shed light on the current challenges of using intestinal organoids as in vitro models.
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Affiliation(s)
- Wenxiu Liu
- Cuiying Biomedical Research Center, Lanzhou University Second Hospital, Lanzhou, 730030, Gansu, China
- Lanzhou Huazhitiancheng Biotechnologies Co., Ltd, Lanzhou, 730000, Gansu, China
| | - Qian Wang
- Cuiying Biomedical Research Center, Lanzhou University Second Hospital, Lanzhou, 730030, Gansu, China
| | - Yanrui Bai
- Cuiying Biomedical Research Center, Lanzhou University Second Hospital, Lanzhou, 730030, Gansu, China
| | - Han Xiao
- Cuiying Biomedical Research Center, Lanzhou University Second Hospital, Lanzhou, 730030, Gansu, China
| | - Zhunduo Li
- Cuiying Biomedical Research Center, Lanzhou University Second Hospital, Lanzhou, 730030, Gansu, China
| | - Yan Wang
- Cuiying Biomedical Research Center, Lanzhou University Second Hospital, Lanzhou, 730030, Gansu, China
| | - Qi Wang
- Lanzhou Huazhitiancheng Biotechnologies Co., Ltd, Lanzhou, 730000, Gansu, China.
| | - Jing Yang
- Cuiying Biomedical Research Center, Lanzhou University Second Hospital, Lanzhou, 730030, Gansu, China.
| | - Hui Sun
- Cuiying Biomedical Research Center, Lanzhou University Second Hospital, Lanzhou, 730030, Gansu, China.
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10
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Saorin G, Caligiuri I, Rizzolio F. Microfluidic organoids-on-a-chip: The future of human models. Semin Cell Dev Biol 2023; 144:41-54. [PMID: 36241560 DOI: 10.1016/j.semcdb.2022.10.001] [Citation(s) in RCA: 62] [Impact Index Per Article: 31.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/31/2022] [Revised: 10/06/2022] [Accepted: 10/06/2022] [Indexed: 11/06/2022]
Abstract
Microfluidics opened the possibility to model the physiological environment by controlling fluids flows, and therefore nutrients supply. It allows to integrate external stimuli such as electricals or mechanicals and in situ monitoring important parameters such as pH, oxygen and metabolite concentrations. Organoids are self-organized 3D organ-like clusters, which allow to closely model original organ functionalities. Applying microfluidics to organoids allows to generate powerful human models for studying organ development, diseases, and drug testing. In this review, after a brief introduction on microfluidics, organoids and organoids-on-a-chip are described by organs (brain, heart, gastrointestinal tract, liver, pancreas) highlighting the microfluidic approaches since this point of view was overlooked in previously published reviews. Indeed, the review aims to discuss from a different point of view, primary microfluidics, the available literature on organoids-on-a-chip, standing out from the published literature by focusing on each specific organ.
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Affiliation(s)
- Gloria Saorin
- Department of Molecular Sciences and Nanosystems, Ca' Foscari University of Venice, 30123 Venezia, Italy
| | - Isabella Caligiuri
- Pathology Unit, Centro di Riferimento Oncologico di Aviano (CRO) IRCCS, 33081 Aviano, Italy
| | - Flavio Rizzolio
- Department of Molecular Sciences and Nanosystems, Ca' Foscari University of Venice, 30123 Venezia, Italy; Pathology Unit, Centro di Riferimento Oncologico di Aviano (CRO) IRCCS, 33081 Aviano, Italy.
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Kakni P, Jutten B, Teixeira Oliveira Carvalho D, Penders J, Truckenmüller R, Habibovic P, Giselbrecht S. Hypoxia-tolerant apical-out intestinal organoids to model host-microbiome interactions. J Tissue Eng 2023; 14:20417314221149208. [PMID: 36699634 PMCID: PMC9869231 DOI: 10.1177/20417314221149208] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/14/2022] [Accepted: 12/16/2022] [Indexed: 01/20/2023] Open
Abstract
Microbiome is an integral part of the gut and is essential for its proper function. Imbalances of the microbiota can be devastating and have been linked with several gastrointestinal conditions. Current gastrointestinal models do not fully reflect the in vivo situation. Thus, it is important to establish more advanced in vitro models to study host-microbiome/pathogen interactions. Here, we developed for the first time an apical-out human small intestinal organoid model in hypoxia, where the apical surface is directly accessible and exposed to a hypoxic environment. These organoids mimic the intestinal cell composition, structure and functions and provide easy access to the apical surface. Co-cultures with the anaerobic strains Lactobacillus casei and Bifidobacterium longum showed successful colonization and probiotic benefits on the organoids. These novel hypoxia-tolerant apical-out small intestinal organoids will pave the way for unraveling unknown mechanisms related to host-microbiome interactions and serve as a tool to develop microbiome-related probiotics and therapeutics.
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Affiliation(s)
- Panagiota Kakni
- Department of Instructive Biomaterials Engineering, MERLN Institute for Technology-Inspired Regenerative Medicine, Maastricht University, Maastricht, The Netherlands
| | - Barry Jutten
- Department of Instructive Biomaterials Engineering, MERLN Institute for Technology-Inspired Regenerative Medicine, Maastricht University, Maastricht, The Netherlands
| | - Daniel Teixeira Oliveira Carvalho
- Department of Instructive Biomaterials Engineering, MERLN Institute for Technology-Inspired Regenerative Medicine, Maastricht University, Maastricht, The Netherlands
| | - John Penders
- Department of Medical Microbiology, Infectious Diseases and Infection Prevention, NUTRIM School of Nutrition and Translational Research in Metabolism, CAPHRI Care and Public Health Research Institute, Maastricht University Medical Centre+, Maastricht, The Netherlands
| | - Roman Truckenmüller
- Department of Instructive Biomaterials Engineering, MERLN Institute for Technology-Inspired Regenerative Medicine, Maastricht University, Maastricht, The Netherlands
| | - Pamela Habibovic
- Department of Instructive Biomaterials Engineering, MERLN Institute for Technology-Inspired Regenerative Medicine, Maastricht University, Maastricht, The Netherlands
| | - Stefan Giselbrecht
- Department of Instructive Biomaterials Engineering, MERLN Institute for Technology-Inspired Regenerative Medicine, Maastricht University, Maastricht, The Netherlands,Stefan Giselbrecht, Department of Instructive Biomaterials Engineering, MERLN Institute for Technology-Inspired Regenerative Medicine, Maastricht University, Universiteitssingel 40, Maastricht 6229 ER, The Netherlands.
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12
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Li G, Liu L, Yin Y, Wang M, Wang L, Dou J, Wu H, Yang Y, He B. Network pharmacology and experimental verification-based strategy to explore the underlying mechanism of Liu Jun An Wei formula in the treatment of gastrointestinal reactions caused by chemotherapy for colorectal cancer. Front Pharmacol 2022; 13:999115. [PMID: 36204230 PMCID: PMC9530632 DOI: 10.3389/fphar.2022.999115] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/20/2022] [Accepted: 09/05/2022] [Indexed: 11/13/2022] Open
Abstract
Background: Liu Jun An Wei formula (LJAW), derived from “Liu Jun Zi Decoction”, is a classical prescription of Tradition Chinese Medicine and has been used for the treatment of gastrointestinal reactions caused by chemotherapy for colorectal cancer (CRC) for many years. Its molecular mechanism remains to be further explored.Objective: To clarify the mechanism of LJAW in attenuating gastrointestinal reactions caused by chemotherapy for CRC.Methods: The 5-fluorouracil (5-FU) induced mouse and intestine organoid models were established to observe the effect of LJAW. The ingredients of LJAW were analyzed and identified by UPLC-Q-TOF-MS technology. Targets of LJAW and chemotherapy-induced gastrointestinal reactions were collected from several databases. “Ingredient-target” network and protein-protein interaction network were constructed based on network pharmacology. Then, gene ontology (GO) functional analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were performed. Subsequently, molecular docking method was used to verify the interaction between the core ingredients and key targets. The results were validated by both in vivo experiments and organoid experiments. Western Blot was used to analyze the influence of LJAW on key targets including PI3K, AKT1, MAPK1, MAPK14 proteins and their phosphorylated proteins. RT-qPCR and Western Blot were used to detect the mRNA and protein levels of apoptosis-related gene PUMA.Results: Compared with the 5-FU group, the LJAW group had better morphology in mouse small intestine and intestine organoids. In total, 18 core ingredients and 19 key targets were obtained from 97 ingredients and 169 common targets. KEGG analysis showed that the common targets were involved in PI3K/Akt, MAPK, apoptosis and other signal pathways, which are closely related to gastrointestinal injury. Experiments confirmed that LJAW lowered the expressions of phosphorylated proteins including p-PI3K, p-AKT1, p-MAPK1, and p-MAPK14 and reduced the mRNA and protein levels of PUMA.Conclusion: LJAW shows protective effect on 5-FU induced small intestine and intestinal organoids injury. LJAW attenuates gastrointestinal reactions caused by chemotherapy for CRC probably by regulating apoptosis-related genes through PI3K/AKT and MAPK signaling pathways.
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Affiliation(s)
- Gaobiao Li
- Xiyuan Hospital, China Academy of Chinese Medical Sciences, Beijing, China
- Graduate School, Beijing University of Chinese Medicine, Beijing, China
| | - Liying Liu
- Dongzhimen Hospital, Beijing University of Chinese Medicine, Beijing, China
| | - Yiran Yin
- Xiyuan Hospital, China Academy of Chinese Medical Sciences, Beijing, China
| | - Mengmeng Wang
- Xiyuan Hospital, China Academy of Chinese Medical Sciences, Beijing, China
| | - Lei Wang
- Xiyuan Hospital, China Academy of Chinese Medical Sciences, Beijing, China
| | - Jianwei Dou
- School of Pharmacy, Xi’an Jiaotong University, Xi’an, Shaanxi, China
| | - Hongwei Wu
- Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing, China
| | - Yufei Yang
- Xiyuan Hospital, China Academy of Chinese Medical Sciences, Beijing, China
- *Correspondence: Yufei Yang, ; Bin He,
| | - Bin He
- Xiyuan Hospital, China Academy of Chinese Medical Sciences, Beijing, China
- *Correspondence: Yufei Yang, ; Bin He,
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13
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Huang S, Zhang S, Chen L, Pan X, Wen Z, Chen Y, Zhang L, Liu J, Chen D. Lipopolysaccharide induced intestinal epithelial injury: a novel organoids-based model for sepsis in vitro. Chin Med J (Engl) 2022; 135:2232-2239. [PMID: 36355867 PMCID: PMC9771316 DOI: 10.1097/cm9.0000000000002348] [Citation(s) in RCA: 12] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/30/2021] [Indexed: 11/12/2022] Open
Abstract
BACKGROUND Advances in organoid culture technology have provided a greater understanding of disease pathogenesis, which has been rarely studied in sepsis before. We aim to establish a suitable organoids-based intestinal injury model for sepsis. METHODS Stable passaged organoids were constructed and pre-treated with lipopolysaccharide (LPS) to mimic sepsis-induced intestinal injury. The LPS-induced sepsis model was used as a reference. We used quantitative real-time polymerase chain reaction to evaluate the RNA levels of inflammatory factors and antimicrobial peptides. Enzyme-linked immunosorbent assay was used to evaluate the protein levels, hematoxylin and eosin staining was used to evaluate the pathology of the small intestine of mice, and immunohistochemistry and immunofluorescence were used to evaluate the intestinal epithelial barrier function. Perkin Elmer Operetta™ was used to obtain high-resolution images of three-dimensional organoids. RESULTS An LPS concentration >150 μg/mL after 24 h was identified to cause organoid growth restriction. The fluorescence intensity of zonula occludens-1 and occludins at LPS concentrations >100 μg/mL decreased significantly after 24 h. After LPS stimulation for 8 h, the RNA expression levels of interleukin (IL)-1α, tumor necrosis factor alpha, granulocyte-macrophage colony-stimulating factor, IL-6, and regenerating islet-derived protein 3 alpha, beta, and gamma increased. These results resembled those of intestinal epithelial layer alterations in a mouse sepsis model. For IL-10, the RNA expression level increased only when the LPS level >200 μg/mL for 24 h. CONCLUSIONS This study provides the primary intestinal in vitro model to study the effects of LPS-induced intestinal injury resembling sepsis. This model provides a platform for immune associated mechanism exploration and effective drug screening.
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Affiliation(s)
- Sisi Huang
- Department of Critical Care Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200020, China
| | - Sheng Zhang
- Department of Critical Care Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200020, China
| | - Limin Chen
- Department of Critical Care Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200020, China
| | - Xiaojun Pan
- Department of Critical Care Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200020, China
| | - Zhenliang Wen
- Department of Critical Care Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200020, China
| | - Yizhu Chen
- Department of Critical Care Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200020, China
| | - Lidi Zhang
- Department of Critical Care Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200020, China
| | - Jiao Liu
- Department of Critical Care Medicine, Ruijin Hospital North, Shanghai Jiao Tong University School of Medicine, Shanghai 201801, China
| | - Dechang Chen
- Department of Critical Care Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200020, China
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14
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Elbadawi M, Ammar RM, Rabini S, Klauck SM, Efferth T. Modulation of Intestinal Corticotropin-Releasing Hormone Signaling by the Herbal Preparation STW 5-II: Possible Mechanisms for Irritable Bowel Syndrome Management. Pharmaceuticals (Basel) 2022; 15:ph15091121. [PMID: 36145342 PMCID: PMC9504045 DOI: 10.3390/ph15091121] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/02/2022] [Revised: 08/31/2022] [Accepted: 09/04/2022] [Indexed: 11/25/2022] Open
Abstract
Corticotropin-releasing factor (CRF) mediates stress responses and alters the gut-brain axis, contributing to the pathogenesis of irritable bowel syndrome (IBS), which is recognized by abdominal pain accompanied by bowel habit disturbance. STW 5-II, a mixture of six herbal extracts, is clinically effective in functional dyspepsia and IBS. Here we aimed to establish an organoid-based stress-induced IBS-like model to investigate the mechanisms of action of STW 5-II. STW 5-II (10, 20, and 30 g/mL) was applied to intestinal organoids for 24 h before being treated with CRF (100 nM) for 48 h. The effects of STW 5-II on CRF signaling were investigated using several in vitro and in silico approaches. STW 5-II activities were further explored by in silico PyRx screening followed by molecular docking of the main 52 identified compounds in STW 5-II with both CRF receptors CRFR1 and CRFR2. CRF exposure stimulated inflammation and increased proinflammatory mediators, while STW 5-II dose-dependently counteracted these effects. STW 5-II inhibited CRF-induced claudin-2 overexpression and serotonin release. Docking of the STW 5-II constituents oleanolic acid and licorice saponin G2 to CRFR1 and CRFR2, respectively, showed a good affinity. These multi-target activities support and elucidate the clinically proven efficacy of STW 5-II in disorders of gut-brain interaction.
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Affiliation(s)
- Mohamed Elbadawi
- Department of Pharmaceutical Biology, Institute of Pharmaceutical and Biomedical Sciences, Johannes Gutenberg University-Mainz, 55128 Mainz, Germany
| | - Ramy M. Ammar
- Medical Affairs, Bayer Consumer Health, 64295 Darmstadt, Germany
| | - Sabine Rabini
- Medical Affairs, Bayer Consumer Health, 64295 Darmstadt, Germany
| | - Sabine M. Klauck
- Division of Cancer Genome Research, German Cancer Research Center (DKFZ), German Cancer Consortium (DKTK), National Center for Tumor Diseases (NCT), 69120 Heidelberg, Germany
| | - Thomas Efferth
- Department of Pharmaceutical Biology, Institute of Pharmaceutical and Biomedical Sciences, Johannes Gutenberg University-Mainz, 55128 Mainz, Germany
- Correspondence:
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15
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Mussard E, Lencina C, Gallo L, Barilly C, Poli M, Feve K, Albin M, Cauquil L, Knudsen C, Achard C, Devailly G, Soler L, Combes S, Beaumont M. The phenotype of the gut region is more stably retained than developmental stage in piglet intestinal organoids. Front Cell Dev Biol 2022; 10:983031. [PMID: 36105361 PMCID: PMC9465596 DOI: 10.3389/fcell.2022.983031] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/30/2022] [Accepted: 07/29/2022] [Indexed: 11/13/2022] Open
Abstract
Intestinal organoids are innovative in vitro tools to study the digestive epithelium. The objective of this study was to generate jejunum and colon organoids from suckling and weaned piglets in order to determine the extent to which organoids retain a location-specific and a developmental stage-specific phenotype. Organoids were studied at three time points by gene expression profiling for comparison with the transcriptomic patterns observed in crypts in vivo. In addition, the gut microbiota and the metabolome were analyzed to characterize the luminal environment of epithelial cells at the origin of organoids. The location-specific expression of 60 genes differentially expressed between jejunum and colon crypts from suckling piglets was partially retained (48%) in the derived organoids at all time point. The regional expression of these genes was independent of luminal signals since the major differences in microbiota and metabolome observed in vivo between the jejunum and the colon were not reproduced in vitro. In contrast, the regional expression of other genes was erased in organoids. Moreover, the developmental stage-specific expression of 30 genes differentially expressed between the jejunum crypts of suckling and weaned piglets was not stably retained in the derived organoids. Differentiation of organoids was necessary to observe the regional expression of certain genes while it was not sufficient to reproduce developmental stage-specific expression patterns. In conclusion, piglet intestinal organoids retained a location-specific phenotype while the characteristics of developmental stage were erased in vitro. Reproducing more closely the luminal environment might help to increase the physiological relevance of intestinal organoids.
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Affiliation(s)
- Eloïse Mussard
- GenPhySE, Université de Toulouse, INRAE, ENVT, Castanet-Tolosan, France
- Lallemand Animal Nutrition, Blagnac Cedex, France
| | - Corinne Lencina
- GenPhySE, Université de Toulouse, INRAE, ENVT, Castanet-Tolosan, France
| | - Lise Gallo
- GenPhySE, Université de Toulouse, INRAE, ENVT, Castanet-Tolosan, France
| | - Céline Barilly
- GenPhySE, Université de Toulouse, INRAE, ENVT, Castanet-Tolosan, France
| | - Maryse Poli
- GenPhySE, Université de Toulouse, INRAE, ENVT, Castanet-Tolosan, France
| | - Katia Feve
- GenPhySE, Université de Toulouse, INRAE, ENVT, Castanet-Tolosan, France
| | - Mikael Albin
- Toxalim (Research Centre in Food Toxicology), Université de Toulouse, INRAE, ENVT, INP-Purpan, UPS, Toulouse, France
| | - Laurent Cauquil
- GenPhySE, Université de Toulouse, INRAE, ENVT, Castanet-Tolosan, France
| | | | | | | | - Laura Soler
- Toxalim (Research Centre in Food Toxicology), Université de Toulouse, INRAE, ENVT, INP-Purpan, UPS, Toulouse, France
| | - Sylvie Combes
- GenPhySE, Université de Toulouse, INRAE, ENVT, Castanet-Tolosan, France
| | - Martin Beaumont
- GenPhySE, Université de Toulouse, INRAE, ENVT, Castanet-Tolosan, France
- *Correspondence: Martin Beaumont,
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16
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Weiß F, Holthaus D, Kraft M, Klotz C, Schneemann M, Schulzke JD, Krug SM. Human duodenal organoid-derived monolayers serve as a suitable barrier model for duodenal tissue. Ann N Y Acad Sci 2022; 1515:155-167. [PMID: 35666953 DOI: 10.1111/nyas.14804] [Citation(s) in RCA: 15] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Usually, duodenal barriers are investigated using intestinal cell lines like Caco-2, which in contrast to native tissue are limited in cell-type representation. Organoids can consist of all intestinal cell types and are supposed to better reflect the in vivo situation. Growing three-dimensionally, with the apical side facing the lumen, application of typical physiological techniques to analyze the barrier is difficult. Organoid-derived monolayers (ODMs) were developed to overcome this. After optimizing culturing conditions, ODMs were characterized and compared to Caco-2 and duodenal tissue. Tight junction composition and appearance were analyzed, and electrophysiological barrier properties, like paracellular and transcellular barrier function and macromolecule permeability, were evaluated. Furthermore, transcriptomic data were analyzed. ODMs had tight junction protein expression and paracellular barrier properties much more resembling the originating tissue than Caco-2. Transcellular barrier was similar between ODMs and native tissue but was increased in Caco-2. Transcriptomic data showed that Caco-2 expressed fewer solute carriers than ODMs and native tissue. In conclusion, while Caco-2 cells differ mostly in transcellular properties, ODMs reflect trans- and paracellular properties of the originating tissue. If cultured under optimized conditions, ODMs possess reproducible functionality, and the variety of different cell types makes them a suitable model for human tissue-specific investigations.
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Affiliation(s)
- Franziska Weiß
- Clinical Physiology/Nutritional Medicine, Charité - Universitätsmedizin Berlin, CBF, Berlin, Germany
| | - David Holthaus
- Department of Infectious Diseases, Unit 16 Mycotic and Parasitic Agents and Mycobacteria, Robert Koch-Institute, Berlin, Germany
| | - Martin Kraft
- Department of Infectious Diseases, Unit 16 Mycotic and Parasitic Agents and Mycobacteria, Robert Koch-Institute, Berlin, Germany
| | - Christian Klotz
- Department of Infectious Diseases, Unit 16 Mycotic and Parasitic Agents and Mycobacteria, Robert Koch-Institute, Berlin, Germany
| | - Martina Schneemann
- Clinical Physiology/Nutritional Medicine, Charité - Universitätsmedizin Berlin, CBF, Berlin, Germany
| | - Jörg D Schulzke
- Clinical Physiology/Nutritional Medicine, Charité - Universitätsmedizin Berlin, CBF, Berlin, Germany
| | - Susanne M Krug
- Clinical Physiology/Nutritional Medicine, Charité - Universitätsmedizin Berlin, CBF, Berlin, Germany
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Liabeuf D, Oshima M, Stange DE, Sigal M. Stem Cells, Helicobacter pylori, and Mutational Landscape: Utility of Preclinical Models to Understand Carcinogenesis and to Direct Management of Gastric Cancer. Gastroenterology 2022; 162:1067-1087. [PMID: 34942172 DOI: 10.1053/j.gastro.2021.12.252] [Citation(s) in RCA: 27] [Impact Index Per Article: 9.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/24/2021] [Revised: 12/14/2021] [Accepted: 12/15/2021] [Indexed: 12/20/2022]
Abstract
Several genetic and environmental factors increase gastric cancer (GC) risk, with Helicobacter pylori being the main environmental agent. GC is thought to emerge through a sequence of morphological changes that have been elucidated on the molecular level. New technologies have shed light onto pathways that are altered in GC, involving mutational and epigenetic changes and altered signaling pathways. Using various new model systems and innovative approaches, the relevance of such alterations for the emergence and progression of GC has been validated. Here, we highlight the key strategies and the resulting achievements. A major step is the characterization of epithelial stem cell behavior in the healthy stomach. These data, obtained through new reporter mouse lines and lineage tracing, enabled insights into the processes that control cellular proliferation, self-renewal, and differentiation of gastric stem cells. It has become evident that these cells and pathways are often deregulated in carcinogenesis. Second, insights into how H pylori colonizes gastric glands, directly interacts with stem cells, and alters cellular and genomic integrity, as well as the characterization of tissue responses to infection, provide a comprehensive picture of how this bacterium contributes to gastric carcinogenesis. Third, the development of stem cell- and tissue-specific reporter mice have driven our understanding of the signals and mutations that promote different types of GC and now also enable the study of more advanced, metastasized stages. Finally, organoids from human tissue have allowed insights into gastric carcinogenesis by validating mutational and signaling alterations in human primary cells and opening a route to predicting responses to personalized treatment.
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Affiliation(s)
- Dylan Liabeuf
- Department of Visceral, Thoracic and Vascular Surgery, University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany
| | - Masanobu Oshima
- Division of Genetics, Cancer Research Institute, Kanazawa University, Kanazawa, Japan
| | - Daniel E Stange
- Department of Visceral, Thoracic and Vascular Surgery, University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany; National Center for Tumor Diseases (NCT/UCC), Dresden, Germany, German Cancer Research Center (DKFZ), Heidelberg, Germany, Faculty of Medicine and University Hospital Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany, Helmholtz-Zentrum Dresden - Rossendorf (HZDR), Dresden, Germany
| | - Michael Sigal
- Department of Internal Medicine, Division of Hepatology and Gastroenterology, Charité Universitätsmedizin Berlin, Germany; Berlin Institute for Medical Systems Biology, Max Delbrück Center for Molecular Medicine, Berlin, Germany.
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18
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Xi W, Saleh J, Yamada A, Tomba C, Mercier B, Janel S, Dang T, Soleilhac M, Djemat A, Wu H, Romagnolo B, Lafont F, Mège RM, Chen Y, Delacour D. Modulation of designer biomimetic matrices for optimized differentiated intestinal epithelial cultures. Biomaterials 2022; 282:121380. [DOI: 10.1016/j.biomaterials.2022.121380] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/03/2021] [Revised: 01/07/2022] [Accepted: 01/16/2022] [Indexed: 12/22/2022]
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Challenges to, and prospects for, reverse engineering the gastrointestinal tract using organoids. Trends Biotechnol 2022; 40:932-944. [DOI: 10.1016/j.tibtech.2022.01.006] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/19/2021] [Revised: 01/09/2022] [Accepted: 01/10/2022] [Indexed: 12/29/2022]
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20
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The Fast Track for Intestinal Tumor Cell Differentiation and In Vitro Intestinal Models by Inorganic Topographic Surfaces. Pharmaceutics 2022; 14:pharmaceutics14010218. [PMID: 35057113 PMCID: PMC8781367 DOI: 10.3390/pharmaceutics14010218] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/23/2021] [Revised: 01/11/2022] [Accepted: 01/14/2022] [Indexed: 11/17/2022] Open
Abstract
Three-dimensional (3D) complex in vitro cell systems are well suited to providing meaningful and translatable results in drug screening, toxicity measurements, and biological studies. Reliable complex gastrointestinal in vitro models as a testbed for oral drug administration and toxicity are very valuable in achieving predictive results for clinical trials and reducing animal testing. However, producing these models is time-consuming due to the lengthy differentiation of HT29 or other cells into mucus-producing goblet cells or other intestinal cell lineages. In the present work, HT29 cells were grown on an inorganic topographic surface decorated with a periodic pattern of micrometre-sized amorphous SiO2 structures for up to 35 days. HT29 cells on topographic surfaces were compared to undifferentiated HT29 in glucose-containing medium on glass or culture dish and with HT29 cells differentiated for 30 days in the presence of methotrexate (HT29-MTX). The cells were stained with Alcian blue for mucus, antibodies for mucus 2 (goblet cells), villin (enterocytes), lysozyme (Paneth cells), and FITC-labeled lectins to identify different cells, glycomic profiles, and cell features. We observed that HT29 cells on topographic surfaces showed more similarities with the differentiated HT29-MTX than with undifferentiated HT29. They formed islands of cell clusters, as observed for HT29-MTX. Already after 2 days, the first mucus secretion was shown by Alcian blue stain and FITC-wheat germ agglutinin. After 4–6 days, mucus was observed on the cell surface and in the intercellular space. The cell layer was undulated, and in 3D reconstruction, the cells showed a clear polarisation with a strong actin signal to one membrane. The lectins and the antibody-staining confirmed the heterogeneous composition of differentiated HT29 cells on topographic surfaces after 6–8 days, or after 6–8 days following MTX differentiation (30 days).
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21
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Ma HC, Zhu YJ, Zhou R, Yu YY, Xiao ZZ, Zhang HB. Lung cancer organoids, a promising model still with long way to go. Crit Rev Oncol Hematol 2022; 171:103610. [DOI: 10.1016/j.critrevonc.2022.103610] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/09/2021] [Revised: 01/26/2022] [Accepted: 01/27/2022] [Indexed: 12/13/2022] Open
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22
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Elbadawi M, Ammar RM, Aziz-Kalbhenn H, Rabini S, Klauck SM, Dawood M, Saeed MEM, Kampf CJ, Efferth T. Anti-inflammatory and tight junction protective activity of the herbal preparation STW 5-II on mouse intestinal organoids. PHYTOMEDICINE : INTERNATIONAL JOURNAL OF PHYTOTHERAPY AND PHYTOPHARMACOLOGY 2021; 88:153589. [PMID: 34111617 DOI: 10.1016/j.phymed.2021.153589] [Citation(s) in RCA: 18] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 03/26/2021] [Revised: 04/17/2021] [Accepted: 04/29/2021] [Indexed: 06/12/2023]
Abstract
BACKGROUND Irritable bowel syndrome (IBS) is a functional bowel disorder, in which recurrent abdominal pain is associated with defecation or a change in bowel habits. STW 5-II is a combination of six medicinal herbs with a clinically proven efficacy in managing IBS. AIM This study aims to establish an in vitro IBS model using mouse intestinal organoids and to explore the anti-inflammatory and tight junction protective activities of the multi-herbal preparation STW 5-II. METHODS Intestinal organoids were cultured in 1:1 Matrigel™ and medium domes. Inflammation and tight junction disruption were induced by a cocktail of cytokines (TNFα, IFNγ, IL-1β, IL-6) and bacterial proteins (LPS, flagellin). Organoids were treated with different concentrations of STW 5-II, and its multi-target activity was assessed using microarray analyses, RT-qPCR, immunofluorescence, western blot, immunohistochemistry, and a FITC permeability assay. In addition, we analyzed the expression of pNF-κB, pSTAT1, iNOS and ZO-1. In silico analyses were conducted to predict and identify the active components that may be responsible in mediating the multi-target anti-inflammatory activity of STW 5-II. RESULTS An organoid based IBS model was successfully established. STW 5-II effectively reduced the cytokines-induced overexpression of the pro-inflammatory mediators pNF-κB, pSTAT1 and iNOS. Moreover, STW 5-II attenuated cytokine-mediated downregulation of the tight junction protein, ZO-1. This finding was confirmed by a FITC permeability assay. In silico analyses revealed a promising inhibitory activity of some isolated compounds from STW 5-II against NF-κB, STAT1 and iNOS. CONCLUSION STW 5-II possesses multiple anti-inflammatory as well as tight junction protective activities that could explain its clinically proven efficacy in managing IBS symptoms.
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Affiliation(s)
- Mohamed Elbadawi
- Department of Pharmaceutical Biology, Institute of Pharmaceutical and Biomedical Sciences, Johannes Gutenberg University, Mainz, Germany
| | - Ramy M Ammar
- Bayer Consumer Health, Steigerwald Arzneimittelwerk GmbH, Darmstadt, Germany
| | - Heba Aziz-Kalbhenn
- Bayer Consumer Health, Steigerwald Arzneimittelwerk GmbH, Darmstadt, Germany
| | - Sabine Rabini
- Bayer Consumer Health, Steigerwald Arzneimittelwerk GmbH, Darmstadt, Germany
| | - Sabine M Klauck
- Division of Cancer Genome Research, German Cancer Research Center (DKFZ), German Cancer Consortium (DKTK), National Center for Tumor Diseases (NCT), Heidelberg, Germany
| | - Mona Dawood
- Department of Pharmaceutical Biology, Institute of Pharmaceutical and Biomedical Sciences, Johannes Gutenberg University, Mainz, Germany; Department of Molecular Biology, Faculty of Medical Laboratory Science, Al-Neelain University, Khartoum, Sudan
| | - Mohamed E M Saeed
- Department of Pharmaceutical Biology, Institute of Pharmaceutical and Biomedical Sciences, Johannes Gutenberg University, Mainz, Germany
| | | | - Thomas Efferth
- Department of Pharmaceutical Biology, Institute of Pharmaceutical and Biomedical Sciences, Johannes Gutenberg University, Mainz, Germany.
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New Stable Cell Lines Derived from the Proximal and Distal Intestine of Rainbow Trout ( Oncorhynchus mykiss) Retain Several Properties Observed In Vivo. Cells 2021; 10:cells10061555. [PMID: 34205481 PMCID: PMC8235179 DOI: 10.3390/cells10061555] [Citation(s) in RCA: 16] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/18/2021] [Revised: 06/15/2021] [Accepted: 06/17/2021] [Indexed: 12/22/2022] Open
Abstract
We derived two novel cell lines from rainbow trout (RT) proximal (RTpi-MI) and distal intestine (RTdi-MI) and compared them with the previously established continuous cell line RTgutGC. Intestinal stem cells, differentiating and differentiated epithelial cells, and connective cells were found in all cell lines. The cell lines formed a polarized barrier, which was not permeable to large molecules and absorbed proline and glucose. High seeding density induced their differentiation into more mature phenotypes, as indicated by the downregulation of intestinal stem cell-related genes (i.e., sox9, hopx and lgr5), whereas alkaline phosphatase activity was upregulated. Other enterocyte markers (i.e., sglt1 and pept1), however, were not regulated as expected. In all cell lines, the presence of a mixed population of epithelial and stromal cells was characterized for the first time. The expression by the stromal component of lgr5, a stem cell niche regulatory molecule, may explain why these lines proliferate stably in vitro. Although most parameters were conserved among the three cell lines, some significant differences were observed, suggesting that characteristics typical of each tract are partly conserved in vitro as well.
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24
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Tallapragada NP, Cambra HM, Wald T, Keough Jalbert S, Abraham DM, Klein OD, Klein AM. Inflation-collapse dynamics drive patterning and morphogenesis in intestinal organoids. Cell Stem Cell 2021; 28:1516-1532.e14. [PMID: 33915079 DOI: 10.1016/j.stem.2021.04.002] [Citation(s) in RCA: 51] [Impact Index Per Article: 12.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/26/2020] [Revised: 12/29/2020] [Accepted: 04/01/2021] [Indexed: 02/07/2023]
Abstract
How stem cells self-organize to form structured tissues is an unsolved problem. Intestinal organoids offer a model of self-organization as they generate stem cell zones (SCZs) of typical size even without a spatially structured environment. Here we examine processes governing the size of SCZs. We improve the viability and homogeneity of intestinal organoid cultures to enable long-term time-lapse imaging of multiple organoids in parallel. We find that SCZs are shaped by fission events under strong control of ion channel-mediated inflation and mechanosensitive Piezo-family channels. Fission occurs through stereotyped modes of dynamic behavior that differ in their coordination of budding and differentiation. Imaging and single-cell transcriptomics show that inflation drives acute stem cell differentiation and induces a stretch-responsive cell state characterized by large transcriptional changes, including upregulation of Piezo1. Our results reveal an intrinsic capacity of the intestinal epithelium to self-organize by modulating and then responding to its mechanical state.
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Affiliation(s)
- Naren P Tallapragada
- Department of Systems Biology, Blavatnik Institute, Harvard Medical School, Boston, MA 02115, USA
| | - Hailey M Cambra
- Department of Systems Biology, Blavatnik Institute, Harvard Medical School, Boston, MA 02115, USA
| | - Tomas Wald
- Program in Craniofacial Biology and Department of Orofacial Sciences, University of California, San Francisco, San Francisco, CA, USA; Department of Pediatrics and Institute for Human Genetics, University of California, San Francisco, San Francisco, CA, USA
| | - Samantha Keough Jalbert
- Department of Systems Biology, Blavatnik Institute, Harvard Medical School, Boston, MA 02115, USA
| | - Diana M Abraham
- Department of Systems Biology, Blavatnik Institute, Harvard Medical School, Boston, MA 02115, USA
| | - Ophir D Klein
- Program in Craniofacial Biology and Department of Orofacial Sciences, University of California, San Francisco, San Francisco, CA, USA; Department of Pediatrics and Institute for Human Genetics, University of California, San Francisco, San Francisco, CA, USA
| | - Allon M Klein
- Department of Systems Biology, Blavatnik Institute, Harvard Medical School, Boston, MA 02115, USA.
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25
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Gao M, Harper MM, Lin M, Qasem SA, Patel RA, Mardini SH, Gabr MM, Cavnar MJ, Pandalai PK, Kim J. Development of a Single-Cell Technique to Increase Yield and Use of Gastrointestinal Cancer Organoids for Personalized Medicine Application. J Am Coll Surg 2021; 232:504-514. [PMID: 33253861 PMCID: PMC8005421 DOI: 10.1016/j.jamcollsurg.2020.11.009] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/13/2020] [Accepted: 11/13/2020] [Indexed: 12/15/2022]
Abstract
BACKGROUND Organoids are excellent 3-dimensional in vitro models of gastrointestinal cancers. However, patient-derived organoids (PDOs) remain inconsistent and unreliable for rapid actionable drug sensitivity testing due to size variation and limited material. STUDY DESIGN On day10/passage 2 after standard creation of organoids, half of PDOs were dissociated into single-cells with TrypLE Express Enzyme/DNase I and mechanical dissociation; and half of PDOs were expanded by the standard technique. Hematoxylin and eosin and immunohistochemistry with CK7 and CK20 were performed for characterization. Drug sensitivity testing was completed for single-cells and paired standard PDOs to assess reproducibility. RESULTS After 2 to 3 days, >50% of single-cells reformed uniform miniature PDOs (∼50 μm). We developed 10 PDO single-cell lines (n = 4, gastric cancer, [GC]; and n = 6, pancreatic ductal adenocarcinoma, [PDAC]), which formed epithelialized cystic structures and by IHC, exhibited CK7(high)/CK20(low) expression patterns mirroring parent tissues. Compared with paired standard PDOs, single-cells (n = 2, PDAC; = 2, GC) showed similar architecture, albeit smaller and more uniform. Importantly, single cells demonstrated similar sensitivity to cytotoxic drugs to matched PDOs. CONCLUSIONS PDO single-cells are accurate for rapid clinical drug testing in gastrointestinal cancers. Using early passage PDO single-cells facilitates high-volume drug testing, decreasing time from tumor sampling to actionable clinical decisions, and provides a personalized medicine platform to optimally select drugs for gastrointestinal cancer patients.
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Affiliation(s)
- Mei Gao
- Division of Surgical Oncology, University of Kentucky, Lexington, KY
| | - Megan M Harper
- Division of Surgical Oncology, University of Kentucky, Lexington, KY
| | - Miranda Lin
- Division of Surgical Oncology, University of Kentucky, Lexington, KY
| | - Shadi A Qasem
- Department of Pathology and Laboratory Medicine, University of Kentucky, Lexington, KY
| | - Reema A Patel
- Divisions of Hematology and Medical Oncology, University of Kentucky, Lexington, KY
| | | | - Moamen M Gabr
- Gastroenterology, University of Kentucky, Lexington, KY
| | - Michael J Cavnar
- Division of Surgical Oncology, University of Kentucky, Lexington, KY
| | | | - Joseph Kim
- Division of Surgical Oncology, University of Kentucky, Lexington, KY.
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26
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Culture and differentiation of rabbit intestinal organoids and organoid-derived cell monolayers. Sci Rep 2021; 11:5401. [PMID: 33686141 PMCID: PMC7940483 DOI: 10.1038/s41598-021-84774-w] [Citation(s) in RCA: 16] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/07/2020] [Accepted: 02/22/2021] [Indexed: 12/14/2022] Open
Abstract
Organoids emulate many aspects of their parental tissue and are therefore used to study pathogen-host interactions and other complex biological processes. Here, we report a robust protocol for the isolation, maintenance and differentiation of rabbit small intestinal organoids and organoid-derived cell monolayers. Our rabbit intestinal spheroid and monolayer cultures grew most efficiently in L-WRN-conditioned medium that contained Wnt, R-spondin and Noggin, and that had been supplemented with ROCK and TGF-β inhibitors. Organoid and monolayer differentiation was initiated by reducing the concentration of the L-WRN-conditioned medium and by adding ROCK and Notch signalling inhibitors. Immunofluorescence staining and RT-qPCR demonstrated that our organoids contained enterocytes, enteroendocrine cells, goblet cells and Paneth cells. Finally, we infected rabbit organoids with Rabbit calicivirus Australia-1, an enterotropic lagovirus that—like many other caliciviruses—does not grow in conventional cell culture. Despite testing various conditions for inoculation, we did not detect any evidence of virus replication, suggesting either that our organoids do not contain suitable host cell types or that additional co-factors are required for a productive infection of rabbit organoids with Rabbit calicivirus Australia-1.
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Soto F, Guimarães CF, Reis RL, Franco W, Rizvi I, Demirci U. Emerging biofabrication approaches for gastrointestinal organoids towards patient specific cancer models. Cancer Lett 2021; 504:116-124. [PMID: 33577978 DOI: 10.1016/j.canlet.2021.01.023] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/11/2020] [Revised: 01/14/2021] [Accepted: 01/23/2021] [Indexed: 01/12/2023]
Abstract
Tissue engineered organoids are simple biomodels that can emulate the structural and functional complexity of specific organs. Here, we review developments in three-dimensional (3D) artificial cell constructs to model gastrointestinal dynamics towards cancer diagnosis. We describe bottom-up approaches to fabricate close-packed cell aggregates, from the use of biochemical and physical cues to guide the self-assembly of organoids, to the use of engineering approaches, including 3D printing/additive manufacturing and external field-driven protocols. Finally, we outline the main challenges and possible risks regarding the potential translation of gastrointestinal organoids from laboratory settings to patient-specific models in clinical applications.
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Affiliation(s)
- Fernando Soto
- Canary Center at Stanford for Cancer Early Detection, Bio-Acoustic MEMS in Medicine (BAMM) Laboratory, Department of Radiology, School of Medicine Stanford University, Palo Alto, California, 94304-5427, USA
| | - Carlos F Guimarães
- Canary Center at Stanford for Cancer Early Detection, Bio-Acoustic MEMS in Medicine (BAMM) Laboratory, Department of Radiology, School of Medicine Stanford University, Palo Alto, California, 94304-5427, USA; 3B's Research Group, Research Institute on Biomaterials, Biodegradables and Biomimetics, University of Minho, Headquarters of the European Institute of Excellence on Tissue Engineering and Regenerative Medicine, AvePark, Parque de Ciência e Tecnologia, Zona Industrial da Gandra, 4805-017 Barco, Guimarães, Portugal; ICVS/3B's, PT Government Associate Laboratory, University of Minho, Braga/Guimarães, Portugal
| | - Rui L Reis
- 3B's Research Group, Research Institute on Biomaterials, Biodegradables and Biomimetics, University of Minho, Headquarters of the European Institute of Excellence on Tissue Engineering and Regenerative Medicine, AvePark, Parque de Ciência e Tecnologia, Zona Industrial da Gandra, 4805-017 Barco, Guimarães, Portugal; ICVS/3B's, PT Government Associate Laboratory, University of Minho, Braga/Guimarães, Portugal
| | - Walfre Franco
- Department of Biomedical Engineering, University of Massachusetts, Lowell, 01854, MA, USA; Wellman Center for Photomedicine, Massachusetts General Hospital, Boston, 02114, MA, USA
| | - Imran Rizvi
- Joint Department of Biomedical Engineering, University of North Carolina at Chapel Hill, Chapel Hill, NC and North Carolina State University, Raleigh, NC, 27599, USA; Lineberger Comprehensive Cancer Center, University of North Carolina School of Medicine, Chapel Hill, NC, 27599, USA
| | - Utkan Demirci
- Canary Center at Stanford for Cancer Early Detection, Bio-Acoustic MEMS in Medicine (BAMM) Laboratory, Department of Radiology, School of Medicine Stanford University, Palo Alto, California, 94304-5427, USA.
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28
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O'Neill JD, Pinezich MR, Guenthart BA, Vunjak-Novakovic G. Gut bioengineering strategies for regenerative medicine. Am J Physiol Gastrointest Liver Physiol 2021; 320:G1-G11. [PMID: 33174453 PMCID: PMC8112187 DOI: 10.1152/ajpgi.00206.2020] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/01/2020] [Revised: 10/23/2020] [Accepted: 11/05/2020] [Indexed: 01/31/2023]
Abstract
Gastrointestinal disease burden continues to rise in the United States and worldwide. The development of bioengineering strategies to model gut injury or disease and to reestablish functional gut tissue could expand therapeutic options and improve clinical outcomes. Current approaches leverage a rapidly evolving gut bioengineering toolkit aimed at 1) de novo generation of gutlike tissues at multiple scales for microtissue models or implantable grafts and 2) regeneration of functional gut in vivo. Although significant progress has been made in intestinal organoid cultures and engineered tissues, development of predictive in vitro models and effective regenerative therapies remains challenging. In this review, we survey emerging bioengineering tools and recent methodological advances to identify current challenges and future opportunities in gut bioengineering for disease modeling and regenerative medicine.
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Affiliation(s)
- John D O'Neill
- Department of Biomedical Engineering, Columbia University, New York, New York
- Department of Cell Biology, State University of New York Downstate Medical Center, Brooklyn, New York
| | - Meghan R Pinezich
- Department of Biomedical Engineering, Columbia University, New York, New York
| | - Brandon A Guenthart
- Department of Cardiothoracic Surgery, Stanford University, Stanford, California
| | - Gordana Vunjak-Novakovic
- Department of Biomedical Engineering, Columbia University, New York, New York
- Department of Medicine, Columbia University Medical Center, New York, New York
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29
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Seidlitz T, Koo BK, Stange DE. Gastric organoids-an in vitro model system for the study of gastric development and road to personalized medicine. Cell Death Differ 2021; 28:68-83. [PMID: 33223522 PMCID: PMC7852679 DOI: 10.1038/s41418-020-00662-2] [Citation(s) in RCA: 57] [Impact Index Per Article: 14.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/25/2020] [Revised: 10/24/2020] [Accepted: 10/26/2020] [Indexed: 12/12/2022] Open
Abstract
Gastric cancer ranks as the fifth most common human malignancy and the third leading cause of cancer related deaths. Depending on tumor stage, endoscopic or surgical resection supported by perioperative chemotherapy is the only curative option for patients. Due to late clinical manifestation and missing reliable biomarkers, early detection is challenging and overall survival remains poor. Organoids are cell aggregates cultured in three-dimensions that grow with similar characteristics as their tissue-of-origin. Due to their self-renewal and proliferative capacity, organoids can be maintained long term in culture and expanded in many cases in an unlimited fashion. Patient-derived organoid (PDO) libraries function as living biobanks, allowing the in depth analysis of tissue specific function, development and disease. The recent successful establishment of gastric cancer PDOs opens up new perspectives for multiple translational clinical applications. Here, we review different adult stem cell derived gastric organoid model systems and focus on their establishment, phenotypic and genotypic characterizations as well as their use in predicting therapy response.
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Affiliation(s)
- Therese Seidlitz
- Department of Visceral, Thoracic and Vascular Surgery, Medical Faculty and University Hospital Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany
| | - Bon-Kyoung Koo
- Institute of Molecular Biotechnology of the Austrian Academy of Sciences, Vienna Biocenter (VBC), Vienna, Austria
| | - Daniel E Stange
- Department of Visceral, Thoracic and Vascular Surgery, Medical Faculty and University Hospital Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany.
- National Center for Tumor Diseases (NCT), Dresden, Germany: German Cancer Research Center (DKFZ), Heidelberg, Germany; Faculty of Medicine and University Hospital Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany; Helmholtz-Zentrum Dresden - Rossendorf (HZDR), Dresden, Germany.
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30
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Mussard E, Pouzet C, Helies V, Pascal G, Fourre S, Cherbuy C, Rubio A, Vergnolle N, Combes S, Beaumont M. Culture of rabbit caecum organoids by reconstituting the intestinal stem cell niche in vitro with pharmacological inhibitors or L-WRN conditioned medium. Stem Cell Res 2020; 48:101980. [PMID: 32920507 DOI: 10.1016/j.scr.2020.101980] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/18/2020] [Revised: 08/19/2020] [Accepted: 08/28/2020] [Indexed: 12/16/2022] Open
Abstract
Intestinal organoids are self-organized 3-dimensional (3D) structures formed by a single layer of polarized epithelial cells. This innovative in vitro model is highly relevant to study physiology of the intestinal epithelium and its role in nutrition and barrier function. However, this model has never been developed in rabbits, while it would have potential applications for biomedical and veterinary research. Here, we cultured rabbit caecum organoids with either pharmacological inhibitors (2Ki medium) or L-WRN cells conditioned medium (L-WRN CM) to reconstitute the intestinal stem cell niche in vitro. Large spherical organoids were obtained with the 2Ki medium and this morphology was associated with a high level of proliferation and stem cells markers gene expression. In contrast, organoids cultured with L-WRN CM had a smaller diameter; a greater cell height and part of them were not spherical. When the L-WRN CM was used at low concentration (5%) for two days, the gene expression of stem cells and proliferation markers were very low, while absorptive and secretory cells markers and antimicrobial peptides were elevated. Epithelial cells within organoids were polarized in 3D cultures with 2Ki medium or L-WRN CM (apical side towards the lumen). We cultured dissociated organoid cells in 2D monolayers, which allowed accessibility to the apical compartment. Under these conditions, actin stress fibers were observed with the 2Ki medium, while perijonctionnal localization of actin was observed with the L-WRN CM suggesting, in 2D cultures as well, a higher differentiation level in the presence of L-WRN CM. In conclusion, rabbit caecum organoids cultured with the 2Ki medium were more proliferative and less differentiated than organoids cultured with L-WRN CM. We propose that organoids cultured with the 2Ki medium could be used to rapidly generate in vitro a large number of rabbit intestinal epithelial stem cells while organoids cultured with the L-WRN CM used at low concentration represent a suitable model to study differentiated rabbit epithelium.
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Affiliation(s)
- Eloïse Mussard
- GenPhySE, Université de Toulouse, INRAE, ENVT, F-31326 Castanet Tolosan, France
| | - Cécile Pouzet
- FRAIB-TRI Imaging Platform Facilities, Université de Toulouse, CNRS, 31320 Castanet-Tolosan, France
| | - Virginie Helies
- GenPhySE, Université de Toulouse, INRAE, ENVT, F-31326 Castanet Tolosan, France
| | - Géraldine Pascal
- GenPhySE, Université de Toulouse, INRAE, ENVT, F-31326 Castanet Tolosan, France
| | - Sandra Fourre
- GeT-PlaGe, Genotoul, INRAE, Castanet Tolosan, France
| | - Claire Cherbuy
- Micalis Institute, INRAE, AgroParisTech, Université Paris-Saclay, 78350 Jouy-en-Josas, France
| | - Aude Rubio
- IRSD, Université de Toulouse, INSERM, INRAE, ENVT, UPS, U1220, CHU Purpan, CS60039, 31024 Toulouse, France
| | - Nathalie Vergnolle
- IRSD, Université de Toulouse, INSERM, INRAE, ENVT, UPS, U1220, CHU Purpan, CS60039, 31024 Toulouse, France
| | - Sylvie Combes
- GenPhySE, Université de Toulouse, INRAE, ENVT, F-31326 Castanet Tolosan, France
| | - Martin Beaumont
- GenPhySE, Université de Toulouse, INRAE, ENVT, F-31326 Castanet Tolosan, France.
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31
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Biel C, Bigaeva E, Hesse M, Bomers JJM, van Summeren K, Teunis MAT, Vaessen S, Ten Klooster JP, Olinga P. Survival and cellular heterogeneity of epithelium in cultured mouse and rat precision-cut intestinal slices. Toxicol In Vitro 2020; 69:104974. [PMID: 32828807 DOI: 10.1016/j.tiv.2020.104974] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/25/2020] [Revised: 08/05/2020] [Accepted: 08/19/2020] [Indexed: 11/30/2022]
Abstract
Precision-cut intestinal slices (PCIS) are used to study intestinal (patho)physiology, drug efficacy, toxicity, transport and metabolism ex vivo. One of the factors that limit the use of PCIS is a relatively short life-span. Moreover, culture-induced changes in cellular composition of PCIS remain largely uncharacterized. In this study, we demonstrated the epithelial cell heterogeneity in mouse and rat PCIS and its alterations during culture. In addition, we evaluated whether the presence of niche growth factors impacts the survival of PCIS epithelial cells. We showed that freshly prepared PCIS retained the main epithelial cell types, namely absorptive enterocytes, goblet cells, enteroendocrine cells, stem cells, transit-amplifying cells and Paneth cells. Once placed in culture, PCIS displayed progressive epithelial damage, and loss of these epithelial cell types. Cells comprising the intestinal stem cell niche were especially sensitive to the damage, and the addition of niche growth factors beneficially affected the survival of stem cells and transit-amplifying cells in PCIS during culture. In conclusion, this study provides new insights into the dynamic changes in cellular composition of epithelium in cultured PCIS, paving the way to future toxicological and pharmacological studies in an informed and reliable ex vivo setting.
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Affiliation(s)
- Carin Biel
- Department of Pharmaceutical Technology and Biopharmacy, University of Groningen, the Netherlands
| | - Emilia Bigaeva
- Department of Pharmaceutical Technology and Biopharmacy, University of Groningen, the Netherlands; Department of Gastroenterology and Hepatology, University of Groningen, University Medical Center Groningen, Groningen, the Netherlands
| | - Melanie Hesse
- Department of Pharmaceutical Technology and Biopharmacy, University of Groningen, the Netherlands
| | - Jordy J M Bomers
- Department of Pharmaceutical Technology and Biopharmacy, University of Groningen, the Netherlands; PROdermpath, Labor für Dermatohistopathology, Vreden, Germany
| | - Kitty van Summeren
- Research Centre for Healthy and Sustainable Living, Innovative Testing in Life Sciences and Chemistry, University of Applied Sciences, Utrecht, the Netherlands
| | - Marc A T Teunis
- Research Centre for Healthy and Sustainable Living, Innovative Testing in Life Sciences and Chemistry, University of Applied Sciences, Utrecht, the Netherlands
| | - Stefan Vaessen
- Research Centre for Healthy and Sustainable Living, Innovative Testing in Life Sciences and Chemistry, University of Applied Sciences, Utrecht, the Netherlands
| | - Jean Paul Ten Klooster
- Research Centre for Healthy and Sustainable Living, Innovative Testing in Life Sciences and Chemistry, University of Applied Sciences, Utrecht, the Netherlands
| | - Peter Olinga
- Department of Pharmaceutical Technology and Biopharmacy, University of Groningen, the Netherlands.
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32
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Arachidonic Acid Promotes Intestinal Regeneration by Activating WNT Signaling. Stem Cell Reports 2020; 15:374-388. [PMID: 32649903 PMCID: PMC7419670 DOI: 10.1016/j.stemcr.2020.06.009] [Citation(s) in RCA: 38] [Impact Index Per Article: 7.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/26/2019] [Revised: 06/09/2020] [Accepted: 06/09/2020] [Indexed: 12/17/2022] Open
Abstract
Intestinal regeneration is crucial for functional restoration after injury, and nutritional molecules can play an important role in this process. Here, we found that arachidonic acid (AA) serves as a direct proliferation promoter of intestinal epithelial cells that facilitates small intestinal regeneration in both three-dimensional cultured organoids and mouse models. As shown in the study, during post-irradiation regeneration, AA positively regulates intestinal epithelial cell proliferation by upregulating the expression of Ascl2 and activating WNT signaling, but negatively regulates intestinal epithelial cell differentiation. AA acts as a delicate regulator that efficiently facilitates epithelial tissue repair by activating radiation-resistant Msi1+ cells rather than Lgr5+ cells, which are extensively considered WNT-activated crypt base stem cells. Additionally, short-term AA treatment maintains optimal intestinal epithelial homeostasis under physiological conditions. As a result, AA treatment can be considered a potential therapy for irradiation injury repair and tissue regeneration.
AA promotes regeneration of intestinal epithelium after irradiation injury AA triggers Ascl2 expression and activates WNT signaling in intestinal epithelium AA facilitates intestinal repair by activating Msi1+ populations
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33
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Silva-Almeida C, Ewart MA, Wilde C. 3D gastrointestinal models and organoids to study metabolism in human colon cancer. Semin Cell Dev Biol 2020; 98:98-104. [DOI: 10.1016/j.semcdb.2019.05.019] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/08/2019] [Revised: 05/17/2019] [Accepted: 05/17/2019] [Indexed: 12/22/2022]
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34
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Xie Y, Ding F, Di W, Lv Y, Xia F, Sheng Y, Yu J, Ding G. Impact of a high‑fat diet on intestinal stem cells and epithelial barrier function in middle‑aged female mice. Mol Med Rep 2020; 21:1133-1144. [PMID: 32016468 PMCID: PMC7003032 DOI: 10.3892/mmr.2020.10932] [Citation(s) in RCA: 41] [Impact Index Per Article: 8.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/24/2019] [Accepted: 12/06/2019] [Indexed: 12/25/2022] Open
Abstract
A high-fat diet (HFD) or obesity-promoting diet is closely associated with metabolic diseases and intestinal tumors, particularly in middle-aged individuals (typically 45–64 years old). The intestinal epithelium constitutes a barrier that separates the host from the food and microbiota in the gut, and thus, a dysfunctional epithelium is associated with a number of diseases. However, the changes caused to the function of intestinal epithelium in response to an HFD have not been well-studied to date. In the present study, middle-aged female mice (12 months old) fed an HFD for a period of 14 weeks were used to determine the effects of HFD on the intestine. Characteristics including the body weight, fat deposition, glucose metabolism, inflammatory state and intestinal morphology were assessed, while the intestinal stem cell (ISC) counts and the ability of isolated intestinal crypts to form organoid bodies in 3D culture were examined. Intestinal epithelial barrier function, including secretory defense, tight junctions and cell apoptosis, were also studied. Morphologically, the HFD resulted in a mild reduction in the length of villi of the small intestine, the colon length and the depth of colon crypts. In addition, the ISC counts were increased in the small intestine and colon in HFD-fed mice. The ability of crypts to grow into organoids (mini-guts) was also increased in crypts obtained from mice fed an HFD, while HFD compromised the epithelial barrier function of the colon. These results demonstrated how an HFD affects the intestinal epithelium and highlighted the need to carefully consider dietary patterns.
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Affiliation(s)
- Yu Xie
- Department of Geriatrics, Division of Geriatric Endocrinology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210029, P.R. China
| | - Fei Ding
- Department of Geriatrics, Division of Geriatric Endocrinology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210029, P.R. China
| | - Wenjuan Di
- Department of Geriatrics, Division of Geriatric Endocrinology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210029, P.R. China
| | - Yifan Lv
- Department of Geriatrics, Division of Geriatric Endocrinology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210029, P.R. China
| | - Fan Xia
- Department of Geriatrics, Division of Geriatric Endocrinology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210029, P.R. China
| | - Yunlu Sheng
- Department of Geriatrics, Division of Geriatric Endocrinology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210029, P.R. China
| | - Jing Yu
- Department of Geriatrics, Division of Geriatric Endocrinology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210029, P.R. China
| | - Guoxian Ding
- Department of Geriatrics, Division of Geriatric Endocrinology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210029, P.R. China
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Oremek ME, Cartwright JA, Rossi AG. Epithelium; Repair of. ENCYCLOPEDIA OF GASTROENTEROLOGY 2020:314-320. [DOI: 10.1016/b978-0-12-801238-3.65902-0] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/02/2025]
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Taggart MW, Foo WC, Lee SM. Tumors of the Gastrointestinal System Including the Pancreas. ONCOLOGICAL SURGICAL PATHOLOGY 2020:691-870. [DOI: 10.1007/978-3-319-96681-6_12] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/06/2025]
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Abstract
Recent advances in culturing of intestinal stem cells and pluripotent stem cells have led to the development of intestinal organoids. These are self-organizing 3D structures, which recapitulate the characteristics and physiological features of in vivo intestinal epithelium. Intestinal organoids have allowed the development of novel in vitro models to study various gastrointestinal diseases expanding our understanding of the pathophysiology of diseases and leading to the development of innovative therapies. This article aims to summarize the current usage of intestinal organoids as a model of gastrointestinal diseases and the potential applications of intestinal organoids in infants and children. Intestinal organoids allow the study of intestinal epithelium responses to stress factors. Mimicking intestinal injury such as necrotizing enterocolitis, intestinal organoids increases the expression of pro-inflammatory cytokine genes and shows disruption of tight junctions after they are injured by lipopolysaccharide and hypoxia. In cystic fibrosis, intestinal organoids derived from rectal biopsies have provided benefits in genetic studies and development of novel therapeutic gene modulation. Transplantation of intestinal organoids via enema has been shown to rescue damaged colonic epithelium in mice. In addition, tissue-engineered small intestine derived from intestinal organoids have been successfully established providing a potential novel treatment and a new hope for children with short bowel syndrome.
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Fois CAM, Le TYL, Schindeler A, Naficy S, McClure DD, Read MN, Valtchev P, Khademhosseini A, Dehghani F. Models of the Gut for Analyzing the Impact of Food and Drugs. Adv Healthc Mater 2019; 8:e1900968. [PMID: 31592579 DOI: 10.1002/adhm.201900968] [Citation(s) in RCA: 20] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/22/2019] [Revised: 08/30/2019] [Indexed: 12/16/2022]
Abstract
Models of the human gastrointestinal tract (GIT) can be powerful tools for examining the biological interactions of food products and pharmaceuticals. This can be done under normal healthy conditions or using models of disease-many of which have no curative therapy. This report outlines the field of gastrointestinal modeling, with a particular focus on the intestine. Traditional in vivo animal models are compared to a range of in vitro models. In vitro systems are elaborated over time, recently culminating with microfluidic intestines-on-chips (IsOC) and 3D bioengineered models. Macroscale models are also reviewed for their important contribution in the microbiota studies. Lastly, it is discussed how in silico approaches may have utility in predicting and interpreting experimental data. The various advantages and limitations of the different systems are contrasted. It is posited that only through complementary use of these models will salient research questions be able to be addressed.
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Affiliation(s)
- Chiara Anna Maria Fois
- School of Chemical and Biomolecular Engineering Centre for Advanced Food Enginomics University of Sydney Sydney NSW 2006 Australia
| | - Thi Yen Loan Le
- School of Chemical and Biomolecular Engineering Centre for Advanced Food Enginomics University of Sydney Sydney NSW 2006 Australia
| | - Aaron Schindeler
- School of Chemical and Biomolecular Engineering Centre for Advanced Food Enginomics University of Sydney Sydney NSW 2006 Australia
| | - Sina Naficy
- School of Chemical and Biomolecular Engineering Centre for Advanced Food Enginomics University of Sydney Sydney NSW 2006 Australia
| | - Dale David McClure
- School of Chemical and Biomolecular Engineering Centre for Advanced Food Enginomics University of Sydney Sydney NSW 2006 Australia
| | - Mark Norman Read
- School of Chemical and Biomolecular Engineering Centre for Advanced Food Enginomics University of Sydney Sydney NSW 2006 Australia
| | - Peter Valtchev
- School of Chemical and Biomolecular Engineering Centre for Advanced Food Enginomics University of Sydney Sydney NSW 2006 Australia
| | - Ali Khademhosseini
- Department of Chemical and Biomolecular Engineering Department of Bioengineering Department of Radiology California NanoSystems Institute (CNSI) University of California Los Angeles CA 90095 USA
| | - Fariba Dehghani
- School of Chemical and Biomolecular Engineering Centre for Advanced Food Enginomics University of Sydney Sydney NSW 2006 Australia
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Yi J, Bergstrom K, Fu J, Shan X, McDaniel JM, McGee S, Qu D, Houchen CW, Liu X, Xia L. Dclk1 in tuft cells promotes inflammation-driven epithelial restitution and mitigates chronic colitis. Cell Death Differ 2019; 26:1656-1669. [PMID: 30478383 PMCID: PMC6748088 DOI: 10.1038/s41418-018-0237-x] [Citation(s) in RCA: 61] [Impact Index Per Article: 10.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/01/2018] [Revised: 09/28/2018] [Accepted: 10/22/2018] [Indexed: 12/16/2022] Open
Abstract
Ulcerative colitis (UC) is a chronic inflammatory bowel disease characterized by defective intestinal barrier integrity toward the microbiota and epithelial damage. Double cortin-like kinase 1 (Dclk1), a marker of intestinal tuft cells, can regulate tissue regenerative responses, but its role in epithelial repair during bacterial-dependent chronic colitis is unclear. We addressed this question using our recently developed mouse model of spontaneous microbiota-dependent colitis induced by mucin-type O-glycan deficiency (DKO), which recapitulates most features of human UC. We generated DKO mice lacking intestinal epithelial Dclk1 (DKO;Dclk1ΔIEC) and analyzed colitis onset and severity using clinical and histologic indices, immune responses by qPCR and immunostaining, and epithelial responses using proliferation markers and organoid culture. We found 3-4-week-old DKO;Dclk1ΔIEC mice developed worsened spontaneous colitis characterized by reduced body weight, loose stool, severe colon thickening, epithelial lesions, and inflammatory cell infiltrates compared with DKO mice. The primary defect was an impaired epithelial proliferative response during inflammation. Dclk1 deficiency also reduced inflammation-induced proliferation and growth of colon organoids ex vivo. Mechanistically, Dclk1 expression was important for inflammation-induced Cox2 expression and prostaglandin E2 (PGE2) production in vivo, and PGE2 rescued proliferative defects in Dclk1-deficient colonic organoids. Although tuft cells were expanded in both DKO and DKO;Dclk1ΔIEC relative to WT mice, loss of Dclk1 was associated with reduced tuft cell activation (i.e., proliferation) during inflammation. Similar results were found in DKO vs. DKO;Dclk1ΔIEC mice at 3-6 months of age. Our results support that tuft cells, via Dclk1, are important responders to bacterial-induced colitis by enhancing epithelial repair responses, which in turn limits bacterial infiltration into the mucosa.
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Affiliation(s)
- Jun Yi
- Department of Gastroenterology, Xiangya Hospital of Central South University, Changsha, Hunan, 410008, China
- Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, 73104, USA
| | - Kirk Bergstrom
- Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, 73104, USA
| | - Jianxin Fu
- Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, 73104, USA
| | - Xindi Shan
- Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, 73104, USA
| | - J Michael McDaniel
- Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, 73104, USA
| | - Samuel McGee
- Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, 73104, USA
| | - Dongfeng Qu
- Department of Internal Medicine, University of Oklahoma Health Sciences Center, Oklahoma City, OK, 73104, USA
| | - Courtney W Houchen
- Department of Internal Medicine, University of Oklahoma Health Sciences Center, Oklahoma City, OK, 73104, USA
| | - Xiaowei Liu
- Department of Gastroenterology, Xiangya Hospital of Central South University, Changsha, Hunan, 410008, China.
| | - Lijun Xia
- Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, 73104, USA.
- Department of Biochemistry and Molecular Biology, University of Oklahoma Health Sciences Center, Oklahoma City, OK, 73104, USA.
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Lin M, Gao M, Cavnar MJ, Kim J. Utilizing gastric cancer organoids to assess tumor biology and personalize medicine. World J Gastrointest Oncol 2019; 11:509-517. [PMID: 31367270 PMCID: PMC6657221 DOI: 10.4251/wjgo.v11.i7.509] [Citation(s) in RCA: 19] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/22/2019] [Revised: 04/25/2019] [Accepted: 06/13/2019] [Indexed: 02/05/2023] Open
Abstract
While the incidence and mortality of gastric cancer (GC) have declined due to public health programs, it remains the third deadliest cancer worldwide. For patients with early disease, innovative endoscopic and complex surgical techniques have improved survival. However, for patients with advanced disease, there are limited treatment options and survival remains poor. Therefore, there is an urgent need for more effective therapies. Since novel therapies require extensive preclinical testing prior to human trials, it is important to identify methods to expedite this process. Traditional cancer models are restricted by the inability to accurately recapitulate the primary human tumor, exorbitant costs, and the requirement for extended periods of development time. An emerging in vitro model to study human disease is the patient-derived organoid, which is a three-dimensional system created from fresh surgical or biopsy tissues of a patient’s gastric tumor. Organoids are cultured in plastic wells and suspended in a gelatinous matrix, providing a substrate for extension and growth in all dimensions. They are rapid-growing and highly representative of the molecular landscape, histology, and morphology of the various subtypes of GC. Organoids uniquely model tumor initiation and growth, including steps taken by normal stomach cells to transform into invasive, intestinal-type tumor cells. Additionally, they provide ample material for biobanking and screening novel therapies. Lastly, organoids are a promising model for personalized therapy and warrant further investigation in drug sensitivity studies for GC patients.
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Affiliation(s)
- Miranda Lin
- Department of Surgery, University of Kentucky, Lexington, KY 40536, United States
| | - Mei Gao
- Department of Surgery, University of Kentucky, Lexington, KY 40536, United States
| | - Michael J Cavnar
- Department of Surgery, University of Kentucky, Lexington, KY 40536, United States
| | - Joseph Kim
- Department of Surgery, University of Kentucky, Lexington, KY 40536, United States
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Abstract
Organoid cultures have emerged as powerful model systems accelerating discoveries in cellular and cancer biology. These three-dimensional cultures are amenable to diverse techniques, including high-throughput genome and transcriptome sequencing, as well as genetic and biochemical perturbation, making these models well suited to answer a variety of questions. Recently, organoids have been generated from diverse human cancers, including breast, colon, pancreas, prostate, bladder, and liver cancers, and studies involving these models are expanding our knowledge of the etiology and characteristics of these malignancies. Co-cultures of cancer organoids with non-neoplastic stromal cells enable investigation of the tumor microenvironment. In addition, recent studies have established that organoids have a place in personalized medicine approaches. Here, we describe the application of organoid technology to cancer discovery and treatment.
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Affiliation(s)
- Hans Clevers
- Hubrecht Institute, Royal Netherlands Academy of Arts and Sciences, and Utrecht University Medical Center, 3584 CT Utrecht, The Netherlands
- Cancer Genomics Netherlands, Utrecht University Medical Center, 3584 GC Utrecht, The Netherlands
- Princess Máxima Centre, 3584 CT Utrecht, The Netherlands
| | - David A. Tuveson
- Cold Spring Harbor Laboratory, Cold Spring Harbor, New York 11724, USA
- Lustgarten Foundation Pancreatic Cancer Research Laboratory, Cold Spring Harbor, New York 11724, USA
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Sims KC, Schwendinger KL, Szymkowicz DB, Swetenburg JR, Bain LJ. Embryonic arsenic exposure reduces intestinal cell proliferation and alters hepatic IGF mRNA expression in killifish (Fundulus heteroclitus). JOURNAL OF TOXICOLOGY AND ENVIRONMENTAL HEALTH. PART A 2019; 82:142-156. [PMID: 30729860 PMCID: PMC6397093 DOI: 10.1080/15287394.2019.1571465] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/09/2023]
Abstract
Arsenic (As) is a toxicant found in food and water throughout the world, and studies suggested that exposure early in life reduces growth. Thus, the goal of this study was to examine mechanisms by which As impacted organismal growth. Killifish (Fundulus heteroclitus) were exposed to 0, 10, 50, or 200 ppb As as embryos and, after hatching, were reared in clean water for up to 40 weeks. Metabolism studies revealed that killifish biotransform As such that monomethylated and dimethylated arsenicals account for 15-17% and 45-61%, respectively, of the total metal. Growth, as measured by condition factor (CF), was significantly and dose-dependently reduced at 8 weeks of age but was similar to controls by 40 weeks. To determine mechanisms underlying the observed initial decrease, intestinal proliferation and morphology were examined. Arsenic-exposed fish exhibited significant 1.3- to 1.5-fold reduction in intestinal villus height and 1.4- to 1.6-fold decrease in proliferating cell nuclear antigen (PCNA+) intestinal cells at all weeks examined. In addition, there were significant correlations between CF, PCNA+ cells, and intestinal villus height. Upon examining whether fish might compensate for the intestinal changes, it was found that hepatic mRNA expression of insulin-like growth factor 1 (IGF-1) and its binding protein (IGFBP-1) were dose-dependently increased. These results indicate that embryonic exposure initially diminished growth, and while intestinal cell proliferation remained reduced, fish appear to compensate by enhancing transcript levels of hepatic IGF-1 and IGFBP-1.
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Affiliation(s)
- Kaleigh C. Sims
- Environmental Toxicology Graduate Program, Clemson University, Clemson, SC, USA
| | | | - Dana B. Szymkowicz
- Environmental Toxicology Graduate Program, Clemson University, Clemson, SC, USA
| | | | - Lisa J. Bain
- Environmental Toxicology Graduate Program, Clemson University, Clemson, SC, USA
- Department of Biological Sciences, Clemson University, Clemson, SC, USA
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Seidlitz T, Merker SR, Rothe A, Zakrzewski F, von Neubeck C, Grützmann K, Sommer U, Schweitzer C, Schölch S, Uhlemann H, Gaebler AM, Werner K, Krause M, Baretton GB, Welsch T, Koo BK, Aust DE, Klink B, Weitz J, Stange DE. Human gastric cancer modelling using organoids. Gut 2019; 68:207-217. [PMID: 29703791 PMCID: PMC6352409 DOI: 10.1136/gutjnl-2017-314549] [Citation(s) in RCA: 228] [Impact Index Per Article: 38.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/23/2017] [Revised: 04/07/2018] [Accepted: 04/10/2018] [Indexed: 12/12/2022]
Abstract
OBJECTIVE Gastric cancer is the second leading cause of cancer-related deaths and the fifth most common malignancy worldwide. In this study, human and mouse gastric cancer organoids were generated to model the disease and perform drug testing to delineate treatment strategies. DESIGN Human gastric cancer organoid cultures were established, samples classified according to their molecular profile and their response to conventional chemotherapeutics tested. Targeted treatment was performed according to specific druggable mutations. Mouse gastric cancer organoid cultures were generated carrying molecular subtype-specific alterations. RESULTS Twenty human gastric cancer organoid cultures were established and four selected for a comprehensive in-depth analysis. Organoids demonstrated divergent growth characteristics and morphologies. Immunohistochemistry showed similar characteristics to the corresponding primary tissue. A divergent response to 5-fluoruracil, oxaliplatin, irinotecan, epirubicin and docetaxel treatment was observed. Whole genome sequencing revealed a mutational spectrum that corresponded to the previously identified microsatellite instable, genomic stable and chromosomal instable subtypes of gastric cancer. The mutational landscape allowed targeted therapy with trastuzumab for ERBB2 alterations and palbociclib for CDKN2A loss. Mouse cancer organoids carrying Kras and Tp53 or Apc and Cdh1 mutations were characterised and serve as model system to study the signalling of induced pathways. CONCLUSION We generated human and mouse gastric cancer organoids modelling typical characteristics and altered pathways of human gastric cancer. Successful interference with activated pathways demonstrates their potential usefulness as living biomarkers for therapy response testing.
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Affiliation(s)
- Therese Seidlitz
- Department of Visceral, Thoracic and Vascular Surgery, University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany
| | - Sebastian R Merker
- Department of Visceral, Thoracic and Vascular Surgery, University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany
| | - Alexander Rothe
- Department of Visceral, Thoracic and Vascular Surgery, University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany
| | - Falk Zakrzewski
- Core Unit for Molecular Tumour Diagnostics (CMTD), National Center for Tumour Diseases (NCT) Dresden, Dresden, Germany
| | - Cläre von Neubeck
- Department of Radiotherapy and Radiation Oncology and National Center for Radiation Research in Oncology (OncoRay), University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany,Partner Site Dresden, German Cancer Consortium (DKTK), Heidelberg, Germany
| | - Konrad Grützmann
- Core Unit for Molecular Tumour Diagnostics (CMTD), National Center for Tumour Diseases (NCT) Dresden, Dresden, Germany
| | - Ulrich Sommer
- Institute for Pathology and Tumour and Normal Tissue Bank of the University Cancer Center (UCC), University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany
| | - Christine Schweitzer
- Department of Visceral, Thoracic and Vascular Surgery, University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany
| | - Sebastian Schölch
- Department of Visceral, Thoracic and Vascular Surgery, University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany,Partner Site Dresden, German Cancer Consortium (DKTK), Heidelberg, Germany,Partner Site Dresden, National Center for Tumor Diseases (NCT), Heidelberg, Germany
| | - Heike Uhlemann
- Department of Visceral, Thoracic and Vascular Surgery, University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany
| | - Anne-Marlene Gaebler
- Department of Visceral, Thoracic and Vascular Surgery, University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany
| | - Kristin Werner
- Department of Visceral, Thoracic and Vascular Surgery, University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany
| | - Mechthild Krause
- Department of Radiotherapy and Radiation Oncology and National Center for Radiation Research in Oncology (OncoRay), University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany,Partner Site Dresden, German Cancer Consortium (DKTK), Heidelberg, Germany,Partner Site Dresden, National Center for Tumor Diseases (NCT), Heidelberg, Germany
| | - Gustavo B Baretton
- Core Unit for Molecular Tumour Diagnostics (CMTD), National Center for Tumour Diseases (NCT) Dresden, Dresden, Germany,Institute for Pathology and Tumour and Normal Tissue Bank of the University Cancer Center (UCC), University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany
| | - Thilo Welsch
- Department of Visceral, Thoracic and Vascular Surgery, University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany,Partner Site Dresden, German Cancer Consortium (DKTK), Heidelberg, Germany,Partner Site Dresden, National Center for Tumor Diseases (NCT), Heidelberg, Germany
| | - Bon-Kyoung Koo
- Institute of Molecular Biotechnology of the Austrian Academy of Sciences (IMBA), Vienna, Austria
| | - Daniela E Aust
- Core Unit for Molecular Tumour Diagnostics (CMTD), National Center for Tumour Diseases (NCT) Dresden, Dresden, Germany,Institute for Pathology and Tumour and Normal Tissue Bank of the University Cancer Center (UCC), University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany
| | - Barbara Klink
- Core Unit for Molecular Tumour Diagnostics (CMTD), National Center for Tumour Diseases (NCT) Dresden, Dresden, Germany,Partner Site Dresden, German Cancer Consortium (DKTK), Heidelberg, Germany,Partner Site Dresden, National Center for Tumor Diseases (NCT), Heidelberg, Germany,Institute for Clinical Genetics, University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany
| | - Jürgen Weitz
- Department of Visceral, Thoracic and Vascular Surgery, University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany,Partner Site Dresden, German Cancer Consortium (DKTK), Heidelberg, Germany,Partner Site Dresden, National Center for Tumor Diseases (NCT), Heidelberg, Germany
| | - Daniel E Stange
- Department of Visceral, Thoracic and Vascular Surgery, University Hospital Carl Gustav Carus, Medical Faculty, Technische Universität Dresden, Dresden, Germany,Partner Site Dresden, German Cancer Consortium (DKTK), Heidelberg, Germany,Partner Site Dresden, National Center for Tumor Diseases (NCT), Heidelberg, Germany
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Intestinal organoids: A new paradigm for engineering intestinal epithelium in vitro. Biomaterials 2019; 194:195-214. [DOI: 10.1016/j.biomaterials.2018.12.006] [Citation(s) in RCA: 39] [Impact Index Per Article: 6.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/30/2018] [Revised: 11/22/2018] [Accepted: 12/08/2018] [Indexed: 12/11/2022]
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Hung L, Obernolte H, Sewald K, Eiwegger T. Human ex vivo and in vitro disease models to study food allergy. Asia Pac Allergy 2019; 9:e4. [PMID: 30740352 PMCID: PMC6365658 DOI: 10.5415/apallergy.2019.9.e4] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/05/2018] [Accepted: 01/11/2019] [Indexed: 12/25/2022] Open
Abstract
Food allergy is a growing global public health concern. As treatment strategies are currently limited to allergen avoidance and emergency interventions, there is an increasing demand for appropriate models of food allergy for the development of new therapeutics. Many models of food allergy rely heavily on the use of animals, and while useful, many are unable to accurately reflect the human system. In order to bridge the gap between in vivo animal models and clinical trials with human patients, human models of food allergy are of great importance. This review will summarize the commonly used human ex vivo and in vitro models of food allergy and highlight their advantages and limitations regarding how accurately they represent the human in vivo system. We will cover biopsy-based systems, precision cut organ slices, and coculture systems as well as organoids and organ-on-a-chip. The availability of appropriate experimental models will allow us to move forward in the field of food allergy research, to search for effective treatment options and to further explore the cause and progression of this disorder.
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Affiliation(s)
- Lisa Hung
- Department of Immunology, University of Toronto, Toronto, ON, Canada.,Translational Medicine Program, Research Institute, The Hospital for Sick Children, Toronto, ON, Canada
| | - Helena Obernolte
- Department of Preclinical Pharmacology and In-Vitro Toxicology, Fraunhofer ITEM, Hannover, Germany
| | - Katherina Sewald
- Department of Preclinical Pharmacology and In-Vitro Toxicology, Fraunhofer ITEM, Hannover, Germany
| | - Thomas Eiwegger
- Department of Immunology, University of Toronto, Toronto, ON, Canada.,Translational Medicine Program, Research Institute, The Hospital for Sick Children, Toronto, ON, Canada.,Division of Clinical Immunology and Allergy, Food Allergy and Anaphylaxis Program, The Hospital for Sick Children, Toronto, ON, Canada
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Gagné-Sansfacon J, Langlois A, Langlois MJ, Coulombe G, Tremblay S, Vaillancourt-Lavigueur V, Qu CK, Menendez A, Rivard N. The tyrosine phosphatase Shp-2 confers resistance to colonic inflammation by driving goblet cell function and crypt regeneration. J Pathol 2018; 247:135-146. [PMID: 30376595 PMCID: PMC6519201 DOI: 10.1002/path.5177] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/30/2018] [Revised: 08/30/2018] [Accepted: 12/25/2018] [Indexed: 12/18/2022]
Abstract
The Src homology‐2 domain‐containing tyrosine phosphatase 2 (SHP‐2) regulates many cellular processes, including proliferation, differentiation and survival. Polymorphisms in the gene encoding SHP‐2 are associated with an increased susceptibility to develop ulcerative colitis. We recently reported that intestinal epithelial cell (IEC)‐specific deletion of Shp‐2 in mice (Shp‐2IEC‐KO) leads to chronic colitis and colitis‐associated cancer. This suggests that SHP‐2‐dependent signaling protects the colonic epithelium against inflammation and colitis‐associated cancer development. To verify this hypothesis, we generated mice expressing the Shp‐2 E76K activated form specifically in IEC. Our results showed that sustained Shp‐2 activation in IEC increased intestine and crypt length, correlating with increased cell proliferation and migration. Crypt regeneration capacity was also markedly enhanced, as revealed by ex vivo organoid culture. Shp‐2 activation alters the secretory cell lineage, as evidenced by increased goblet cell numbers and mucus secretion. Notably, these mice also demonstrated elevated ERK signaling in IEC and exhibited resistance against both chemical‐ and Citrobacter rodentium‐induced colitis. In contrast, mice with IEC‐specific Shp‐2 deletion displayed reduced ERK signaling and rapidly developed chronic colitis. Remarkably, expression of an activated form of Braf in Shp‐2‐deficient mice restored ERK activation, goblet cell production and prevented colitis. Altogether, our results indicate that chronic activation of Shp‐2/ERK signaling in the colonic epithelium confers resistance to mucosal erosion and colitis. © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.
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Affiliation(s)
- Jessica Gagné-Sansfacon
- Department of Anatomy and Cell Biology, Cancer Research Pavilion, Faculty of Medicine and Health Sciences, Université de Sherbrooke, Sherbrooke, Canada
| | - Ariane Langlois
- Department of Anatomy and Cell Biology, Cancer Research Pavilion, Faculty of Medicine and Health Sciences, Université de Sherbrooke, Sherbrooke, Canada
| | - Marie-Josée Langlois
- Department of Anatomy and Cell Biology, Cancer Research Pavilion, Faculty of Medicine and Health Sciences, Université de Sherbrooke, Sherbrooke, Canada
| | - Geneviève Coulombe
- Department of Anatomy and Cell Biology, Cancer Research Pavilion, Faculty of Medicine and Health Sciences, Université de Sherbrooke, Sherbrooke, Canada
| | - Sarah Tremblay
- Department of Microbiology and Infectiology, Cancer Research Pavilion, Faculty of Medicine and Health Sciences, Université de Sherbrooke, Sherbrooke, Canada
| | - Vanessa Vaillancourt-Lavigueur
- Department of Anatomy and Cell Biology, Cancer Research Pavilion, Faculty of Medicine and Health Sciences, Université de Sherbrooke, Sherbrooke, Canada
| | - Cheng-Kui Qu
- Department of Pediatrics, Emory University School of Medicine, Atlanta, GA, USA
| | - Alfredo Menendez
- Department of Microbiology and Infectiology, Cancer Research Pavilion, Faculty of Medicine and Health Sciences, Université de Sherbrooke, Sherbrooke, Canada
| | - Nathalie Rivard
- Department of Anatomy and Cell Biology, Cancer Research Pavilion, Faculty of Medicine and Health Sciences, Université de Sherbrooke, Sherbrooke, Canada
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Lin M, Kim J. ASO Author Reflections: We Can Stomach It: Personalizing Medicine with EGD-Derived Gastric Cancer Organoids. Ann Surg Oncol 2018; 25:972-973. [PMID: 30374916 DOI: 10.1245/s10434-018-6962-z] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/13/2018] [Indexed: 11/18/2022]
Affiliation(s)
- Miranda Lin
- Department of Surgery, Division of Surgical Oncology, University of Kentucky and Markey Cancer Center, Lexington, KY, USA
| | - Joseph Kim
- Department of Surgery, Division of Surgical Oncology, University of Kentucky and Markey Cancer Center, Lexington, KY, USA.
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Modeling Host-Pathogen Interactions in the Context of the Microenvironment: Three-Dimensional Cell Culture Comes of Age. Infect Immun 2018; 86:IAI.00282-18. [PMID: 30181350 DOI: 10.1128/iai.00282-18] [Citation(s) in RCA: 104] [Impact Index Per Article: 14.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022] Open
Abstract
Tissues and organs provide the structural and biochemical landscapes upon which microbial pathogens and commensals function to regulate health and disease. While flat two-dimensional (2-D) monolayers composed of a single cell type have provided important insight into understanding host-pathogen interactions and infectious disease mechanisms, these reductionist models lack many essential features present in the native host microenvironment that are known to regulate infection, including three-dimensional (3-D) architecture, multicellular complexity, commensal microbiota, gas exchange and nutrient gradients, and physiologically relevant biomechanical forces (e.g., fluid shear, stretch, compression). A major challenge in tissue engineering for infectious disease research is recreating this dynamic 3-D microenvironment (biological, chemical, and physical/mechanical) to more accurately model the initiation and progression of host-pathogen interactions in the laboratory. Here we review selected 3-D models of human intestinal mucosa, which represent a major portal of entry for infectious pathogens and an important niche for commensal microbiota. We highlight seminal studies that have used these models to interrogate host-pathogen interactions and infectious disease mechanisms, and we present this literature in the appropriate historical context. Models discussed include 3-D organotypic cultures engineered in the rotating wall vessel (RWV) bioreactor, extracellular matrix (ECM)-embedded/organoid models, and organ-on-a-chip (OAC) models. Collectively, these technologies provide a more physiologically relevant and predictive framework for investigating infectious disease mechanisms and antimicrobial therapies at the intersection of the host, microbe, and their local microenvironments.
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Nakamura T. Recent progress in organoid culture to model intestinal epithelial barrier functions. Int Immunol 2018; 31:13-21. [DOI: 10.1093/intimm/dxy065] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/28/2018] [Accepted: 10/02/2018] [Indexed: 12/30/2022] Open
Affiliation(s)
- Tetsuya Nakamura
- Department of Advanced Therapeutics for GI Diseases, Tokyo Medical and Dental University Yushima, Bunkyo-ku, Tokyo, Japan
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Bhasin N, Alleyne D, Gray OA, Kupfer SS. Vitamin D Regulation of the Uridine Phosphorylase 1 Gene and Uridine-Induced DNA Damage in Colon in African Americans and European Americans. Gastroenterology 2018; 155:1192-1204.e9. [PMID: 29964038 PMCID: PMC6866230 DOI: 10.1053/j.gastro.2018.06.049] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/30/2017] [Revised: 06/08/2018] [Accepted: 06/25/2018] [Indexed: 12/24/2022]
Abstract
BACKGROUND & AIMS African Americans have the greatest colorectal cancer (CRC) burden in the United States; interethnic differences in protective effects of vitamin D might contribute to disparities. 1α,25(OH)2D3 vitamin D (the active form of vitamin D) induces transcription of the uridine phosphorylase gene (UPP1) in colon tissues of European Americans but to a lesser extent in colon tissues of African Americans. UPP1-knockout mice have increased intestinal concentrations of uridine and Deoxyuridine triphosphate (dUTP), have increased uridine-induced DNA damage, and develop colon tumors. We studied 1α,25(OH)2D3 regulation of UPP1 and uridine-induced DNA damage in the colon and differences in these processes between African and European Americans. METHODS We quantified expression and activity of UPP1 in response to 1α,25(OH)2D3 in young adult mouse colonic cells, human CRC cells (LS174T), and organoids (derived from rectosigmoid biopsy samples of healthy individuals undergoing colonoscopies) using quantitative polymerase chain reaction, immunoblot, and immunocytochemistry assays. Binding of the vitamin D receptor to UPP1 was tested by chromatin immunoprecipitation. Uridine-induced DNA damage was measured by fragment-length analysis in repair enzyme assays. Allele-specific 1α,25(OH)2D3 responses were tested using luciferase assays. RESULTS Vitamin D increased levels of UPP1 mRNA, protein, and enzymatic activity and increased vitamin D receptor binding to the UPP1 promoter in young adult mouse colonic cells, LS174T cells, and organoids. 1α,25(OH)2D3 significantly reduced levels of uridine and uridine-induced DNA damage in these cells, which required UPP1 expression. Organoids derived from colon tissues of African Americans expressed lower levels of UPP1 after exposure to 1α,25(OH)2D3 and had increased uridine-induced DNA damage compared with organoids derived from tissues of European Americans. Luciferase assays with the T allele of single nucleotide polymorphism rs28605337 near UPP1, which is found more frequently in African Americans than European Americans, expressed lower levels of UPP1 after exposure to 1α,25(OH)2D3 than assays without this variant. CONCLUSIONS We found vitamin D to increase expression of UPP1, leading to reduce uridine-induced DNA damage, in colon cells and organoids. A polymorphism in UPP1 found more frequently in African Americans than European Americans reduced UPP1 expression upon cell exposure to 1α,25(OH)2D3. Differences in expression of UPP1 in response to vitamin D could contribute to the increased risk of CRC in African Americans.
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Affiliation(s)
- Nobel Bhasin
- Section of Gastroenterology, Hepatology and Nutrition, Department of Medicine, University of Chicago, Chicago, Illinois
| | - Dereck Alleyne
- Section of Gastroenterology, Hepatology and Nutrition, Department of Medicine, University of Chicago, Chicago, Illinois
| | - Olivia A. Gray
- Department of Human Genetics, University of Chicago, Chicago, Illinois
| | - Sonia S. Kupfer
- Section of Gastroenterology, Hepatology and Nutrition, Department of Medicine, University of Chicago, Chicago, Illinois
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