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Qu N, Daoud A, Kechele DO, Cleary CE, Múnera JO. Differentiation of human pluripotent stem cells into urothelial organoids via transient activation of WNT signaling. iScience 2025; 28:112398. [PMID: 40322079 PMCID: PMC12049843 DOI: 10.1016/j.isci.2025.112398] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/28/2024] [Revised: 10/15/2024] [Accepted: 04/07/2025] [Indexed: 05/08/2025] Open
Abstract
The cloaca is a transient structure that forms in the terminal hindgut giving rise to the rectum dorsally and the urogenital sinus ventrally. Similarly, human hindgut cultures derived from human pluripotent stem cells generate human colonic organoids (HCOs) which also contain co-developing urothelial tissue. In this study, our goal was to identify pathways involved in cloacal patterning and apply this to human hindgut cultures. RNA sequencing (RNA-seq) data comparing dorsal versus ventral cloaca in e10.5 mice revealed that WNT signaling was elevated in the ventral versus dorsal cloaca. Inhibition of WNT signaling in hindgut cultures maintained their differentiation toward colonic organoids. WNT activation promoted differentiation toward human urothelial organoids (HUOs). HUOs contained developmental stage specific cell types present in mammalian urothelial tissue including co-developing mesenchyme. Therefore, HUOs offer a powerful in vitro model for dissecting the regulatory pathways that control the dynamic emergence of stage specific cell types within the human urothelium.
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Affiliation(s)
- Na Qu
- Department of Regenerative Medicine and Cell Biology, Medical University of South Carolina, Charleston, SC 29425, USA
| | - Abdelkader Daoud
- Department of Regenerative Medicine and Cell Biology, Medical University of South Carolina, Charleston, SC 29425, USA
| | - Daniel O. Kechele
- Division of Developmental Biology, Cincinnati Children’s Hospital Medical Center, Cincinnati, OH 45229-3039, USA
| | - Cassie E. Cleary
- Department of Regenerative Medicine and Cell Biology, Medical University of South Carolina, Charleston, SC 29425, USA
| | - Jorge O. Múnera
- Department of Regenerative Medicine and Cell Biology, Medical University of South Carolina, Charleston, SC 29425, USA
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2
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Bandral M, Sussel L, Lorberbaum DS. Retinoid signaling in pancreas development, islet function, and disease. Curr Top Dev Biol 2024; 161:297-318. [PMID: 39870436 DOI: 10.1016/bs.ctdb.2024.10.007] [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] [Indexed: 01/29/2025]
Abstract
All-trans retinoic acid (ATRA) signaling is essential in numerous different biological contexts. This review highlights the diverse roles of ATRA during development, function, and diseases of the pancreas. ATRA is essential to specify pancreatic progenitors from gut tube endoderm, endocrine and exocrine differentiation, and adult islet function. ATRA concentration must be carefully regulated during the derivation of islet-like cells from human pluripotent stem cells (hPSCs) to optimize the expression of key pancreatic transcription factors while mitigating adverse and unwanted cell-types in these cultures. The ATRA pathway is integral to the pancreas and here we will present selected studies from decades of research that has laid the essential groundwork for ongoing projects dedicated to unraveling the complexities of ATRA signaling in the pancreas.
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Affiliation(s)
- Manuj Bandral
- University of Michigan, Department of Pharmacology, Caswell Diabetes Institute, Ann Arbor, MI, United States
| | - Lori Sussel
- University of Colorado Denver Anschutz Medical Campus, Barbara Davis Center for Diabetes, Aurora, CO, United States
| | - David S Lorberbaum
- University of Michigan, Department of Pharmacology, Caswell Diabetes Institute, Ann Arbor, MI, United States.
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3
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Kane E, Mak TC, Latreille M. MicroRNA-7 regulates endocrine progenitor delamination and endocrine cell mass in developing pancreatic islets. iScience 2024; 27:110332. [PMID: 39055950 PMCID: PMC11269303 DOI: 10.1016/j.isci.2024.110332] [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: 02/01/2024] [Revised: 05/28/2024] [Accepted: 06/18/2024] [Indexed: 07/28/2024] Open
Abstract
β-cell replenishment in patients with diabetes through cadaveric islet transplantation has been successful; however, it requires long-term immunosuppression and suitable islet donors are scarce. Stepwise in vitro differentiation of pluripotent stem cells into β-cells represents a viable alternative, but limitations in our current understanding of in vivo islet endocrine differentiation constrains its clinical use. Here, we show that microRNA-7 (miR-7) is highly expressed in embryonic pancreatic endocrine progenitors. Genetic deletion of the miR-7 gene family in endocrine progenitors leads to reduced islet endocrine cell mass, due to endocrine progenitors failing to delaminate from the epithelial plexus. This is associated with a reduction in neurogenin-3 levels and increased expression of Sry-box transcription factor 9. Further, we observe that a significant number of endocrine progenitors lacking miR-7 differentiate into ductal cells. Our study suggests that increasing miR-7 expression could improve efficiency of in vitro differentiation and augment stem cell-derived β-cell terminal maturity.
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Affiliation(s)
- Eva Kane
- MRC Laboratory of Medical Sciences, Du Cane Road, London W12 0NN, UK
| | - Tracy C.S. Mak
- MRC Laboratory of Medical Sciences, Du Cane Road, London W12 0NN, UK
| | - Mathieu Latreille
- MRC Laboratory of Medical Sciences, Du Cane Road, London W12 0NN, UK
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4
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Ashok A, Kalthur G, Kumar A. Degradation meets development: Implications in β-cell development and diabetes. Cell Biol Int 2024; 48:759-776. [PMID: 38499517 DOI: 10.1002/cbin.12155] [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] [Received: 11/03/2023] [Revised: 02/22/2024] [Accepted: 03/04/2024] [Indexed: 03/20/2024]
Abstract
Pancreatic development is orchestrated by timely synthesis and degradation of stage-specific transcription factors (TFs). The transition from one stage to another stage is dependent on the precise expression of the developmentally relevant TFs. Persistent expression of particular TF would impede the exit from the progenitor stage to the matured cell type. Intracellular protein degradation-mediated protein turnover contributes to a major extent to the turnover of these TFs and thereby dictates the development of different tissues. Since even subtle changes in the crucial cellular pathways would dramatically impact pancreatic β-cell performance, it is generally acknowledged that the biological activity of these pathways is tightly regulated by protein synthesis and degradation process. Intracellular protein degradation is executed majorly by the ubiquitin proteasome system (UPS) and Lysosomal degradation pathway. As more than 90% of the TFs are targeted to proteasomal degradation, this review aims to examine the crucial role of UPS in normal pancreatic β-cell development and how dysfunction of these pathways manifests in metabolic syndromes such as diabetes. Such understanding would facilitate designing a faithful approach to obtain a therapeutic quality of β-cells from stem cells.
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Affiliation(s)
- Akshaya Ashok
- Manipal Institute of Regenerative Medicine, Bangalore, Manipal Academy of Higher Education, Manipal, India
| | - Guruprasad Kalthur
- Division of Reproductive and Developmental Biology, Department of Reproductive Science, Kasturba Medical College, Manipal Academy of Higher Education, Manipal, India
| | - Anujith Kumar
- Manipal Institute of Regenerative Medicine, Bangalore, Manipal Academy of Higher Education, Manipal, India
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5
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Qu N, Daoud A, Kechele DO, Múnera JO. Human Pluripotent Stem Cell Derived Organoids Reveal a Role for WNT Signaling in Dorsal-Ventral Patterning of the Hindgut. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.03.04.583343. [PMID: 38496665 PMCID: PMC10942392 DOI: 10.1101/2024.03.04.583343] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 03/19/2024]
Abstract
The cloaca is a transient structure that forms in the terminal hindgut giving rise to the rectum dorsally and the urogenital sinus ventrally. Similarly, human hindgut cultures derived from human pluripotent stem cells generate human colonic organoids (HCOs) which also contain co-developing urothelial tissue. In this study, our goal was to identify pathways involved in cloacal patterning and apply this to human hindgut cultures. RNA-seq data comparing dorsal versus ventral cloaca in e10.5 mice revealed that WNT signaling was elevated in the ventral versus dorsal cloaca. Inhibition of WNT signaling in hindgut cultures biased their differentiation towards a colorectal fate. WNT activation biased differentiation towards a urothelial fate, giving rise to human urothelial organoids (HUOs). HUOs contained cell types present in human urothelial tissue. Based on our results, we propose a mechanism whereby WNT signaling patterns the ventral cloaca, prior to cloacal septation, to give rise to the urogenital sinus.
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6
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Bishop D, Schwarz Q, Wiszniak S. Endothelial-derived angiocrine factors as instructors of embryonic development. Front Cell Dev Biol 2023; 11:1172114. [PMID: 37457293 PMCID: PMC10339107 DOI: 10.3389/fcell.2023.1172114] [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: 02/23/2023] [Accepted: 06/19/2023] [Indexed: 07/18/2023] Open
Abstract
Blood vessels are well-known to play roles in organ development and repair, primarily owing to their fundamental function in delivering oxygen and nutrients to tissues to promote their growth and homeostasis. Endothelial cells however are not merely passive conduits for carrying blood. There is now evidence that endothelial cells of the vasculature actively regulate tissue-specific development, morphogenesis and organ function, as well as playing roles in disease and cancer. Angiocrine factors are growth factors, cytokines, signaling molecules or other regulators produced directly from endothelial cells to instruct a diverse range of signaling outcomes in the cellular microenvironment, and are critical mediators of the vascular control of organ function. The roles of angiocrine signaling are only beginning to be uncovered in diverse fields such as homeostasis, regeneration, organogenesis, stem-cell maintenance, cell differentiation and tumour growth. While in some cases the specific angiocrine factor involved in these processes has been identified, in many cases the molecular identity of the angiocrine factor(s) remain to be discovered, even though the importance of angiocrine signaling has been implicated. In this review, we will specifically focus on roles for endothelial-derived angiocrine signaling in instructing tissue morphogenesis and organogenesis during embryonic and perinatal development.
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Ebrahim N, Shakirova K, Dashinimaev E. PDX1 is the cornerstone of pancreatic β-cell functions and identity. Front Mol Biosci 2022; 9:1091757. [PMID: 36589234 PMCID: PMC9798421 DOI: 10.3389/fmolb.2022.1091757] [Citation(s) in RCA: 28] [Impact Index Per Article: 9.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2022] [Accepted: 12/01/2022] [Indexed: 12/23/2022] Open
Abstract
Diabetes has been a worldwide healthcare problem for many years. Current methods of treating diabetes are still largely directed at symptoms, aiming to control the manifestations of the pathology. This creates an overall need to find alternative measures that can impact on the causes of the disease, reverse diabetes, or make it more manageable. Understanding the role of key players in the pathogenesis of diabetes and the related β-cell functions is of great importance in combating diabetes. PDX1 is a master regulator in pancreas organogenesis, the maturation and identity preservation of β-cells, and of their role in normal insulin function. Mutations in the PDX1 gene are correlated with many pancreatic dysfunctions, including pancreatic agenesis (homozygous mutation) and MODY4 (heterozygous mutation), while in other types of diabetes, PDX1 expression is reduced. Therefore, alternative approaches to treat diabetes largely depend on knowledge of PDX1 regulation, its interaction with other transcription factors, and its role in obtaining β-cells through differentiation and transdifferentiation protocols. In this article, we review the basic functions of PDX1 and its regulation by genetic and epigenetic factors. Lastly, we summarize different variations of the differentiation protocols used to obtain β-cells from alternative cell sources, using PDX1 alone or in combination with various transcription factors and modified culture conditions. This review shows the unique position of PDX1 as a potential target in the genetic and cellular treatment of diabetes.
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Affiliation(s)
- Nour Ebrahim
- Center for Precision Genome Editing and Genetic Technologies for Biomedicine, Pirogov Russian National Research Medical University, Moscow, Russia,Moscow Institute of Physics and Technology (State University), Dolgoprudny, Russia
| | - Ksenia Shakirova
- Center for Precision Genome Editing and Genetic Technologies for Biomedicine, Pirogov Russian National Research Medical University, Moscow, Russia
| | - Erdem Dashinimaev
- Center for Precision Genome Editing and Genetic Technologies for Biomedicine, Pirogov Russian National Research Medical University, Moscow, Russia,Moscow Institute of Physics and Technology (State University), Dolgoprudny, Russia,*Correspondence: Erdem Dashinimaev,
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8
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Nihad M, Shenoy P S, Bose B. Cell therapy research for Diabetes: Pancreatic β cell differentiation from pluripotent stem cells. Diabetes Res Clin Pract 2021; 181:109084. [PMID: 34673084 DOI: 10.1016/j.diabres.2021.109084] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/23/2021] [Revised: 09/24/2021] [Accepted: 09/28/2021] [Indexed: 12/16/2022]
Abstract
Human pluripotent stem cells (PSCs), both embryonic and induced pluripotent stem cells (iPSCs), have been differentiated into pancreatic β isletsin vitrofor more than a decade. The idea is to get enough β cells for cell transplantation for diabetics. Finding a standard cell therapy for diabetes is essential because of the logarithmic increase in the global population of people with diabetes and the insufficient availability of the human cadaveric pancreas. Moreover, with better insights into developmental biology, thein vitroβ cell differentiation protocols have depended on thein vivoβ cell organogenesis. Various protocols for pancreatic β cell differentiation have been developed. Such protocols are based on the modulation of cell signalling pathways with growth factors, small molecules, RNAi approaches, directed differentiation using transcription factors, genome editing. Growth factor free differentiation protocols, epigenetic modulations, 3D differentiation approaches, and encapsulation strategies have also been reported for better glycemic control and endocrine modulations. Here, we have reviewed various aforementionedin vitroβ cell differentiation protocols from human PSCs, their respective comparisons, challenges, past, present, and future. The literature has been reviewed primarily from PubMed from the year 2000 till date using the mentioned keywords.
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Affiliation(s)
- Muhammad Nihad
- Stem Cells and Regenerative Medicine Centre, Yenepoya Research Centre, Yenepoya (Deemed to be University), Mangalore, Pincode-575 018, Karnataka, India
| | - Sudheer Shenoy P
- Stem Cells and Regenerative Medicine Centre, Yenepoya Research Centre, Yenepoya (Deemed to be University), Mangalore, Pincode-575 018, Karnataka, India
| | - Bipasha Bose
- Stem Cells and Regenerative Medicine Centre, Yenepoya Research Centre, Yenepoya (Deemed to be University), Mangalore, Pincode-575 018, Karnataka, India.
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9
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Abstract
Pancreatic islet beta cells (β-cells) synthesize and secrete insulin in response to rising glucose levels and thus are a prime target in both major forms of diabetes. Type 1 diabetes ensues due to autoimmune destruction of β-cells. On the other hand, the prevailing insulin resistance and hyperglycemia in type 2 diabetes (T2D) elicits a compensatory response from β-cells that involves increases in β-cell mass and function. However, the sustained metabolic stress results in β-cell failure, characterized by severe β-cell dysfunction and loss of β-cell mass. Dynamic changes to β-cell mass also occur during pancreatic development that involves extensive growth and morphogenesis. These orchestrated events are triggered by multiple signaling pathways, including those representing the transforming growth factor β (TGF-β) superfamily. TGF-β pathway ligands play important roles during endocrine pancreas development, β-cell proliferation, differentiation, and apoptosis. Furthermore, new findings are suggestive of TGF-β's role in regulation of adult β-cell mass and function. Collectively, these findings support the therapeutic utility of targeting TGF-β in diabetes. Summarizing the role of the various TGF-β pathway ligands in β-cell development, growth and function in normal physiology, and during diabetes pathogenesis is the topic of this mini-review.
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Affiliation(s)
- Ji-Hyun Lee
- Cell Growth and Metabolism Section, Diabetes, Endocrinology & Obesity Branch, National Institutes of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Clinical Research Center, Bethesda, MD, USA
| | - Ji-Hyeon Lee
- Cell Growth and Metabolism Section, Diabetes, Endocrinology & Obesity Branch, National Institutes of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Clinical Research Center, Bethesda, MD, USA
| | - Sushil G Rane
- Cell Growth and Metabolism Section, Diabetes, Endocrinology & Obesity Branch, National Institutes of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Clinical Research Center, Bethesda, MD, USA
- Correspondence: Sushil G. Rane, PhD, Cell Growth and Metabolism Section, Diabetes, Endocrinology and Obesity Branch, National Institutes of Diabetes, Digestive and Kidney Diseases, National Institutes of Health, Clinical Research Center, Building 10, CRC-West 5-5940, 10 Center Drive, Bethesda, MD 20892, USA.
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10
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Dumasia NP, Pethe PS. Pancreas development and the Polycomb group protein complexes. Mech Dev 2020; 164:103647. [PMID: 32991980 DOI: 10.1016/j.mod.2020.103647] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/07/2020] [Revised: 09/02/2020] [Accepted: 09/17/2020] [Indexed: 02/07/2023]
Abstract
The dual nature of pancreatic tissue permits both endocrine and exocrine functions. Enzymatic secretions by the exocrine pancreas help digestive processes while the pancreatic hormones regulate glucose homeostasis and energy metabolism. Pancreas organogenesis is defined by a conserved array of signaling pathways that act on common gut progenitors to bring about the generation of diverse cell types. Multiple cellular processes characterize development of the mature organ. These processes are mediated by signaling pathways that regulate lineage-specific transcription factors and chromatin modifications guiding long-term gene expression programs. The chromatin landscape is altered chiefly by DNA or histone modifications, chromatin remodelers, and non-coding RNAs. Amongst histone modifiers, several studies have identified Polycomb group (PcG) proteins as crucial determinants mediating transcriptional repression of genes involved in developmental processes. Although PcG-mediated chromatin modifications define cellular transitions and influence cell identity of multipotent progenitors, much remains to be understood regarding coordination between extracellular signals and their impact on Polycomb functions during the pancreas lineage progression. In this review, we discuss interactions between sequence-specific DNA binding proteins and chromatin regulators underlying pancreas development and insulin producing β-cells, with particular focus on Polycomb group proteins. Understanding such basic molecular mechanisms would improve current strategies for stem cell-based differentiation while also help elucidate the pathogenesis of several pancreas-related maladies, including diabetes and pancreatic cancer.
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Affiliation(s)
- Niloufer P Dumasia
- Department of Biological Sciences, Sunandan Divatia School of Science, SVKM's NMIMS (deemed to-be) University, Mumbai 400 056, India
| | - Prasad S Pethe
- Symbiosis Centre for Stem Cell Research (SCSCR), Symbiosis International University, Lavale, Pune 412 115, India.
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11
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Single-cell patterning and axis characterization in the murine and human definitive endoderm. Cell Res 2020; 31:326-344. [PMID: 33106598 DOI: 10.1038/s41422-020-00426-0] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/07/2020] [Accepted: 09/28/2020] [Indexed: 12/20/2022] Open
Abstract
Defining the precise regionalization of specified definitive endoderm progenitors is critical for understanding the mechanisms underlying the generation and regeneration of respiratory and digestive organs, yet the patterning of endoderm progenitors remains unresolved, particularly in humans. We performed single-cell RNA sequencing on endoderm cells during the early somitogenesis stages in mice and humans. We developed molecular criteria to define four major endoderm regions (foregut, lip of anterior intestinal portal, midgut, and hindgut) and their developmental pathways. We identified the cell subpopulations in each region and their spatial distributions and characterized key molecular features along the body axes. Dorsal and ventral pancreatic progenitors appear to originate from the midgut population and follow distinct pathways to develop into an identical cell type. Finally, we described the generally conserved endoderm patterning in humans and clear differences in dorsal cell distribution between species. Our study comprehensively defines single-cell endoderm patterning and provides novel insights into the spatiotemporal process that drives establishment of early endoderm domains.
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12
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Huang H, Bader TN, Jin S. Signaling Molecules Regulating Pancreatic Endocrine Development from Pluripotent Stem Cell Differentiation. Int J Mol Sci 2020; 21:E5867. [PMID: 32824212 PMCID: PMC7461594 DOI: 10.3390/ijms21165867] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/09/2020] [Revised: 08/08/2020] [Accepted: 08/09/2020] [Indexed: 12/24/2022] Open
Abstract
Diabetes is one of the leading causes of death globally. Currently, the donor pancreas is the only source of human islets, placing extreme constraints on supply. Hence, it is imperative to develop renewable islets for diabetes research and treatment. To date, extensive efforts have been made to derive insulin-secreting cells from human pluripotent stem cells with substantial success. However, the in vitro generation of functional islet organoids remains a challenge due in part to our poor understanding of the signaling molecules indispensable for controlling differentiation pathways towards the self-assembly of functional islets from stem cells. Since this process relies on a variety of signaling molecules to guide the differentiation pathways, as well as the culture microenvironments that mimic in vivo physiological conditions, this review highlights extracellular matrix proteins, growth factors, signaling molecules, and microenvironments facilitating the generation of biologically functional pancreatic endocrine cells from human pluripotent stem cells. Signaling pathways involved in stepwise differentiation that guide the progression of stem cells into the endocrine lineage are also discussed. The development of protocols enabling the generation of islet organoids with hormone release capacities equivalent to native adult islets for clinical applications, disease modeling, and diabetes research are anticipated.
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Affiliation(s)
- Hui Huang
- Department of Biomedical Engineering, Thomas J. Watson School of Engineering and Applied Sciences, State University of New York at Binghamton, Binghamton, NY 13902, USA; (H.H.); (T.N.B.)
| | - Taylor N. Bader
- Department of Biomedical Engineering, Thomas J. Watson School of Engineering and Applied Sciences, State University of New York at Binghamton, Binghamton, NY 13902, USA; (H.H.); (T.N.B.)
| | - Sha Jin
- Department of Biomedical Engineering, Thomas J. Watson School of Engineering and Applied Sciences, State University of New York at Binghamton, Binghamton, NY 13902, USA; (H.H.); (T.N.B.)
- Center of Biomanufacturing for Regenerative Medicine, State University of New York at Binghamton, Binghamton, NY 13902, USA
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13
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Bukys MA, Mihas A, Finney K, Sears K, Trivedi D, Wang Y, Oberholzer J, Jensen J. High-Dimensional Design-Of-Experiments Extracts Small-Molecule-Only Induction Conditions for Dorsal Pancreatic Endoderm from Pluripotency. iScience 2020; 23:101346. [PMID: 32745983 PMCID: PMC7398937 DOI: 10.1016/j.isci.2020.101346] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/28/2019] [Revised: 04/15/2020] [Accepted: 07/02/2020] [Indexed: 01/27/2023] Open
Abstract
The derivation of endoderm and descendant organs, such as pancreas, liver, and intestine, impacts disease modeling and regenerative medicine. Use of TGF-β signaling agonism is a common method for induction of definitive endoderm from pluripotency. By using a data-driven, High-Dimensional Design of Experiments (HD-DoE)-based methodology to address multifactorial problems in directed differentiation, we found instead that optimal conditions demanded BMP antagonism and retinoid input leading to induction of dorsal foregut endoderm (DFE). We demonstrate that pancreatic identity can be rapidly, and robustly, induced from DFE and that such cells are of dorsal pancreatic identity. The DFE population was highly competent to differentiate into both stomach organoids and pancreatic tissue types and able to generate fetal-type β cells through two subsequent differentiation steps using only small molecules. This alternative, rapid, and low-cost basis for generating pancreatic insulin-producing cells may have impact for the development of cell-based therapies for diabetes.
Method development for addressing multifactorial problems in directed differentiation Generation of endodermal populations without the use of TGF-β agonism Small-molecule-based pancreatic differentiation protocol
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Affiliation(s)
- Michael A Bukys
- Department of Biomedical Engineering, Lerner Research Institute, Cleveland Clinic Foundation, 9500 Euclid Avenue, Cleveland, OH 44195, USA
| | - Alexander Mihas
- Department of Biomedical Engineering, Lerner Research Institute, Cleveland Clinic Foundation, 9500 Euclid Avenue, Cleveland, OH 44195, USA
| | - Krystal Finney
- Trailhead Biosystems Inc, 10000 Cedar Avenue, Cleveland, OH, USA; Cleveland Clinic, Cleveland, OH 44195, USA
| | - Katie Sears
- Department of Biomedical Engineering, Lerner Research Institute, Cleveland Clinic Foundation, 9500 Euclid Avenue, Cleveland, OH 44195, USA
| | - Divya Trivedi
- Trailhead Biosystems Inc, 10000 Cedar Avenue, Cleveland, OH, USA; Cleveland Clinic, Cleveland, OH 44195, USA
| | - Yong Wang
- Division of Transplantation, University of Virginia, Charlottesville, VA 22903, USA
| | - Jose Oberholzer
- Division of Transplantation, University of Virginia, Charlottesville, VA 22903, USA
| | - Jan Jensen
- Department of Biomedical Engineering, Lerner Research Institute, Cleveland Clinic Foundation, 9500 Euclid Avenue, Cleveland, OH 44195, USA; Trailhead Biosystems Inc, 10000 Cedar Avenue, Cleveland, OH, USA; Cleveland Clinic, Cleveland, OH 44195, USA.
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14
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Durel JF, Nerurkar NL. Mechanobiology of vertebrate gut morphogenesis. Curr Opin Genet Dev 2020; 63:45-52. [PMID: 32413823 DOI: 10.1016/j.gde.2020.04.002] [Citation(s) in RCA: 13] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/19/2019] [Accepted: 04/09/2020] [Indexed: 01/15/2023]
Abstract
Approximately a century after D'Arcy Thompson's On Growth and Form, there continues to be widespread interest in the biophysical and mathematical basis of morphogenesis. Particularly over the past 20 years, this interest has led to great advances in our understanding of a broad range of processes in embryonic development through a quantitative, mechanically driven framework. Nowhere in vertebrate development is this more apparent than the development of endodermally derived organs. Here, we discuss recent advances in the study of gut development that have emerged primarily from mechanobiology-motivated approaches that span from gut tube morphogenesis and later organogenesis of the respiratory and gastrointestinal systems.
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Affiliation(s)
- John F Durel
- Department of Biomedical Engineering, Columbia University, New York, NY 10027, United States
| | - Nandan L Nerurkar
- Department of Biomedical Engineering, Columbia University, New York, NY 10027, United States; Department of Genetics and Development, Columbia University Medical Center, New York, NY 10032, United States.
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15
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Lv YQ, Wu J, Li XK, Zhang JS, Bellusci S. Role of FGF10/FGFR2b Signaling in Mouse Digestive Tract Development, Repair and Regeneration Following Injury. Front Cell Dev Biol 2019; 7:326. [PMID: 31921841 PMCID: PMC6914673 DOI: 10.3389/fcell.2019.00326] [Citation(s) in RCA: 13] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/14/2019] [Accepted: 11/25/2019] [Indexed: 12/18/2022] Open
Abstract
During embryonic development, the rudimentary digestive tract is initially a tube-like structure. It is composed of epithelial cells surrounded by mesenchymal cells. Reciprocal epithelial–mesenchymal interactions progressively subdivide this primitive tube into distinct functional regions: the tongue, the pharynx, the esophagus, the stomach, the duodenum, the small intestine, the cecum, the large intestine, the colon, and the anus as well as the pancreas and the liver. Fibroblast growth factors (Fgfs) constitute a family of conserved small proteins playing crucial roles during organogenesis, homeostasis, and repair after injury. Among them, fibroblast growth factor 10 (Fgf10) has been reported to orchestrate epithelial–mesenchymal interactions during digestive tract development. In mice, loss of function of Fgf10 as well as its receptor fibroblast growth factor receptor 2b (Fgfr2b) lead to defective taste papillae in the tongue, underdeveloped and defective differentiation of the stomach, duodenal, cecal, and colonic atresias, anorectal malformation, as well as underdeveloped pancreas and liver. Fgf signaling through Fgfr2b receptor is also critical for the repair process after gut injury. In the adult mice, a malabsorption disorder called small bowel syndrome is triggered after massive small bowel resection (SBR). In wild-type mice, SBR leads to a regenerative process called gut adaptation characterized by an increase in the diameter of the remaining small intestine as well as by the presence of deeper crypts and longer villi, altogether leading to increased intestinal surface. Intestinal stem cells are key for this regeneration process. Induction of Fgf10 expression in the Paneth cells located in the crypt following SBR suggests a critical role for this growth factor in the process of gut adaptation.
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Affiliation(s)
- Yu-Qing Lv
- Key Laboratory of Interventional Pulmonology of Zhejiang Province, Department of Pulmonary and Critical Care Medicine, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China.,Institute of Life Sciences, Wenzhou University, Wenzhou, China
| | - Jin Wu
- Institute of Life Sciences, Wenzhou University, Wenzhou, China
| | - Xiao-Kun Li
- Key Laboratory of Interventional Pulmonology of Zhejiang Province, Department of Pulmonary and Critical Care Medicine, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China.,Institute of Life Sciences, Wenzhou University, Wenzhou, China
| | - Jin-San Zhang
- Key Laboratory of Interventional Pulmonology of Zhejiang Province, Department of Pulmonary and Critical Care Medicine, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China.,Institute of Life Sciences, Wenzhou University, Wenzhou, China
| | - Saverio Bellusci
- Key Laboratory of Interventional Pulmonology of Zhejiang Province, Department of Pulmonary and Critical Care Medicine, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China.,Institute of Life Sciences, Wenzhou University, Wenzhou, China.,Department of Internal Medicine II, Cardio-Pulmonary Institute, University of Giessen and Marburg Lung Center, Giessen, Germany
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16
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Villamayor L, Cano DA, Rojas A. GATA factors in pancreas development and disease. IUBMB Life 2019; 72:80-88. [PMID: 31580534 DOI: 10.1002/iub.2170] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/06/2019] [Accepted: 09/05/2019] [Indexed: 12/29/2022]
Abstract
There is an urgent need for the development of novel therapeutics options for diabetic patients given the high prevalence of diabetes worldwide and that, currently, there is no cure for this disease. The transplantation of pancreatic islets that contain insulin-producing cells is a promising therapeutic alternative, particularly for type 1 diabetes. However, the shortage of organ donors constitutes a major limitation for this approach; thus, developing alternative sources of insulin-producing cells is of critical importance. In the last decade, our knowledge of the molecular mechanisms controlling embryonic pancreas development has significantly advanced. More importantly, this knowledge has provided the basis for the in vitro generation of insulin-producing cells from stem cells. Recent studies have revealed that GATA transcription factors are involved in various stages of pancreas formation and in the adult ß cell function. Here, we review the fundamental role of GATA transcription factors in pancreas morphogenesis and their association with congenital diseases associated with pancreas.
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Affiliation(s)
- Laura Villamayor
- Centro Andaluz de Biología Molecular y Medicina Regenerativa-CABIMER, Universidad Pablo de Olavide, Universidad de Sevilla, Consejo Superior de Investigaciones Científicas (CSIC), Seville, Spain
| | - David A Cano
- Instituto de Biomedicina de Sevilla (IBIS), Hospital Universitario Virgen del Rocío/CSIC/Universidad de Sevilla, Seville, Spain
| | - Anabel Rojas
- Centro Andaluz de Biología Molecular y Medicina Regenerativa-CABIMER, Universidad Pablo de Olavide, Universidad de Sevilla, Consejo Superior de Investigaciones Científicas (CSIC), Seville, Spain.,Centro de Investigación Biomédica en Red de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM), Madrid, Spain
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17
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Guerriero I, De Angelis MT, D'Angelo F, Leveque R, Savignano E, Roberto L, Lucci V, Mazzone P, Laurino S, Storto G, Nardelli A, Sgambato A, Ceccarelli M, De Felice M, Amendola E, Falco G. Exploring the Molecular Crosstalk between Pancreatic Bud and Mesenchyme in Embryogenesis: Novel Signals Involved. Int J Mol Sci 2019; 20:ijms20194900. [PMID: 31623299 PMCID: PMC6811752 DOI: 10.3390/ijms20194900] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/21/2019] [Revised: 09/23/2019] [Accepted: 09/27/2019] [Indexed: 01/07/2023] Open
Abstract
Pancreatic organogenesis is a multistep process that requires the cooperation of several signaling pathways. In this context, the role of pancreatic mesenchyme is important to define the epithelium development; nevertheless, the precise space–temporal signaling activation still needs to be clarified. This study reports a dissection of the pancreatic embryogenesis, highlighting the molecular network surrounding the epithelium–mesenchyme interaction. To investigate this crosstalk, pancreatic epithelium and surrounding mesenchyme, at embryonic day 10.5, were collected through laser capture microdissection (LCM) and characterized based on their global gene expression. We performed a bioinformatic analysis to hypothesize crosstalk interactions, validating the most promising genes and verifying the precise localization of their expression in the compartments, by RNA in situ hybridization (ISH). Our analyses pointed out also the c-Met gene, a very well-known factor involved in stimulating motility, morphogenesis, and organ regeneration. We also highlighted the potential crosstalk between Versican (Vcan) and Syndecan4 (Sdc4) since these genes are involved in pancreatic tissue repair, strengthening the concept that the same signaling pathways required during pancreatic embryogenesis are also involved in tissue repair. This finding leads to novel strategies for obtaining functional pancreatic stem cells for cell replacement therapies.
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Affiliation(s)
- Ilaria Guerriero
- Istituto di RicercheGenetiche G. Salvatore, Biogems.c.ar.l, ArianoIrpino, 83031 Avellino, Italy.
| | - Maria Teresa De Angelis
- Istituto di RicercheGenetiche G. Salvatore, Biogems.c.ar.l, ArianoIrpino, 83031 Avellino, Italy.
| | - Fulvio D'Angelo
- Istituto di RicercheGenetiche G. Salvatore, Biogems.c.ar.l, ArianoIrpino, 83031 Avellino, Italy.
| | - Rita Leveque
- Dipartimento di Biologia, Universita' degliStudi di Napoli, Federico II, 80126 Napoli, Italy.
| | - Eleonora Savignano
- Istituto di RicercheGenetiche G. Salvatore, Biogems.c.ar.l, ArianoIrpino, 83031 Avellino, Italy.
| | - Luca Roberto
- Istituto di RicercheGenetiche G. Salvatore, Biogems.c.ar.l, ArianoIrpino, 83031 Avellino, Italy.
| | - Valeria Lucci
- Dipartimento di Biologia, Universita' degliStudi di Napoli, Federico II, 80126 Napoli, Italy.
| | - Pellegrino Mazzone
- Dipartimento di Biologia, Universita' degliStudi di Napoli, Federico II, 80126 Napoli, Italy.
| | - Simona Laurino
- IRCCS-Referral Cancer Center of Basilicata (CROB), 85028 Rionero in Vulture (PZ), Italy.
| | - Giovanni Storto
- IRCCS-Referral Cancer Center of Basilicata (CROB), 85028 Rionero in Vulture (PZ), Italy.
| | - Anna Nardelli
- Istituto di Biostrutture e Bioimmagini-CNR, Via De Amicis No. 95, 80145 Napoli, Italy.
| | - Alessandro Sgambato
- IRCCS-Referral Cancer Center of Basilicata (CROB), 85028 Rionero in Vulture (PZ), Italy.
| | - Michele Ceccarelli
- Department of Science and Technology, University of Sannio, 82100 Benevento, Italy.
| | - Mario De Felice
- Istituto per l'Endocrinologia e l'OncologiaSperimentale "G. Salvatore", CNR, 80131 Napoli, Italy.
| | - Elena Amendola
- Dipartimento di Biologia, Universita' degliStudi di Napoli, Federico II, 80126 Napoli, Italy.
- Istituto per l'Endocrinologia e l'OncologiaSperimentale "G. Salvatore", CNR, 80131 Napoli, Italy.
| | - Geppino Falco
- Istituto di RicercheGenetiche G. Salvatore, Biogems.c.ar.l, ArianoIrpino, 83031 Avellino, Italy.
- Dipartimento di Biologia, Universita' degliStudi di Napoli, Federico II, 80126 Napoli, Italy.
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18
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Asymmetric paralog evolution between the "cryptic" gene Bmp16 and its well-studied sister genes Bmp2 and Bmp4. Sci Rep 2019; 9:3136. [PMID: 30816280 PMCID: PMC6395752 DOI: 10.1038/s41598-019-40055-1] [Citation(s) in RCA: 236] [Impact Index Per Article: 39.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/25/2018] [Accepted: 02/07/2019] [Indexed: 12/05/2022] Open
Abstract
The vertebrate gene repertoire is characterized by “cryptic” genes whose identification has been hampered by their absence from the genomes of well-studied species. One example is the Bmp16 gene, a paralog of the developmental key genes Bmp2 and -4. We focus on the Bmp2/4/16 group of genes to study the evolutionary dynamics following gen(om)e duplications with special emphasis on the poorly studied Bmp16 gene. We reveal the presence of Bmp16 in chondrichthyans in addition to previously reported teleost fishes and reptiles. Using comprehensive, vertebrate-wide gene sampling, our phylogenetic analysis complemented with synteny analyses suggests that Bmp2, -4 and -16 are remnants of a gene quartet that originated during the two rounds of whole-genome duplication (2R-WGD) early in vertebrate evolution. We confirm that Bmp16 genes were lost independently in at least three lineages (mammals, archelosaurs and amphibians) and report that they have elevated rates of sequence evolution. This finding agrees with their more “flexible” deployment during development; while Bmp16 has limited embryonic expression domains in the cloudy catshark, it is broadly expressed in the green anole lizard. Our study illustrates the dynamics of gene family evolution by integrating insights from sequence diversification, gene repertoire changes, and shuffling of expression domains.
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19
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Bloise E, Ciarmela P, Dela Cruz C, Luisi S, Petraglia F, Reis FM. Activin A in Mammalian Physiology. Physiol Rev 2019; 99:739-780. [DOI: 10.1152/physrev.00002.2018] [Citation(s) in RCA: 78] [Impact Index Per Article: 13.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022] Open
Abstract
Activins are dimeric glycoproteins belonging to the transforming growth factor beta superfamily and resulting from the assembly of two beta subunits, which may also be combined with alpha subunits to form inhibins. Activins were discovered in 1986 following the isolation of inhibins from porcine follicular fluid, and were characterized as ovarian hormones that stimulate follicle stimulating hormone (FSH) release by the pituitary gland. In particular, activin A was shown to be the isoform of greater physiological importance in humans. The current understanding of activin A surpasses the reproductive system and allows its classification as a hormone, a growth factor, and a cytokine. In more than 30 yr of intense research, activin A was localized in female and male reproductive organs but also in other organs and systems as diverse as the brain, liver, lung, bone, and gut. Moreover, its roles include embryonic differentiation, trophoblast invasion of the uterine wall in early pregnancy, and fetal/neonate brain protection in hypoxic conditions. It is now recognized that activin A overexpression may be either cytostatic or mitogenic, depending on the cell type, with important implications for tumor biology. Activin A also regulates bone formation and regeneration, enhances joint inflammation in rheumatoid arthritis, and triggers pathogenic mechanisms in the respiratory system. In this 30-yr review, we analyze the evidence for physiological roles of activin A and the potential use of activin agonists and antagonists as therapeutic agents.
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Affiliation(s)
- Enrrico Bloise
- Department of Morphology, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil; Department of Experimental and Clinical Medicine, Università Politecnica delle Marche, Ancona, Italy; Department of Obstetrics and Gynecology, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil; Department of Molecular and Developmental Medicine, Obstetrics and Gynecological Clinic, University of Siena, Siena, Italy; and Department of Biomedical, Experimental and Clinical Sciences, Division of Obstetrics and
| | - Pasquapina Ciarmela
- Department of Morphology, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil; Department of Experimental and Clinical Medicine, Università Politecnica delle Marche, Ancona, Italy; Department of Obstetrics and Gynecology, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil; Department of Molecular and Developmental Medicine, Obstetrics and Gynecological Clinic, University of Siena, Siena, Italy; and Department of Biomedical, Experimental and Clinical Sciences, Division of Obstetrics and
| | - Cynthia Dela Cruz
- Department of Morphology, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil; Department of Experimental and Clinical Medicine, Università Politecnica delle Marche, Ancona, Italy; Department of Obstetrics and Gynecology, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil; Department of Molecular and Developmental Medicine, Obstetrics and Gynecological Clinic, University of Siena, Siena, Italy; and Department of Biomedical, Experimental and Clinical Sciences, Division of Obstetrics and
| | - Stefano Luisi
- Department of Morphology, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil; Department of Experimental and Clinical Medicine, Università Politecnica delle Marche, Ancona, Italy; Department of Obstetrics and Gynecology, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil; Department of Molecular and Developmental Medicine, Obstetrics and Gynecological Clinic, University of Siena, Siena, Italy; and Department of Biomedical, Experimental and Clinical Sciences, Division of Obstetrics and
| | - Felice Petraglia
- Department of Morphology, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil; Department of Experimental and Clinical Medicine, Università Politecnica delle Marche, Ancona, Italy; Department of Obstetrics and Gynecology, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil; Department of Molecular and Developmental Medicine, Obstetrics and Gynecological Clinic, University of Siena, Siena, Italy; and Department of Biomedical, Experimental and Clinical Sciences, Division of Obstetrics and
| | - Fernando M. Reis
- Department of Morphology, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil; Department of Experimental and Clinical Medicine, Università Politecnica delle Marche, Ancona, Italy; Department of Obstetrics and Gynecology, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil; Department of Molecular and Developmental Medicine, Obstetrics and Gynecological Clinic, University of Siena, Siena, Italy; and Department of Biomedical, Experimental and Clinical Sciences, Division of Obstetrics and
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20
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Pancreas organogenesis: The interplay between surrounding microenvironment(s) and epithelium-intrinsic factors. Curr Top Dev Biol 2019; 132:221-256. [DOI: 10.1016/bs.ctdb.2018.12.005] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
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21
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Mesodermal induction of pancreatic fate commitment. Semin Cell Dev Biol 2018; 92:77-88. [PMID: 30142440 DOI: 10.1016/j.semcdb.2018.08.008] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/05/2018] [Revised: 07/29/2018] [Accepted: 08/20/2018] [Indexed: 12/27/2022]
Abstract
The pancreas is a compound gland comprised of both exocrine acinar and duct cells as well as endocrine islet cells. Most notable amongst the latter are the insulin-synthesizing β-cells, loss or dysfunction of which manifests in diabetes mellitus. All exocrine and endocrine cells derive from multipotent pancreatic progenitor cells arising from the primitive gut epithelium via inductive interactions with adjacent mesodermal tissues. Research in the last two decades has revealed the identity of many of these extrinsic cues and they include signaling molecules used in many other developmental contexts such as retinoic acid, fibroblast growth factors, and members of the TGF-β superfamily. As important as these inductive cues is the absence of other signaling molecules such as hedgehog family members. Much has been learned about the interactions of extrinsic factors with fate regulators intrinsic to the pancreatic endoderm. This new knowledge has had tremendous impact on the development of directed differentiation protocols for converting pluripotent stem cells to β-cells in vitro.
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22
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Li LC, Qiu WL, Zhang YW, Xu ZR, Xiao YN, Hou C, Lamaoqiezhong, Yu P, Cheng X, Xu CR. Single-cell transcriptomic analyses reveal distinct dorsal/ventral pancreatic programs. EMBO Rep 2018; 19:embr.201846148. [PMID: 30065074 DOI: 10.15252/embr.201846148] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/19/2018] [Revised: 06/30/2018] [Accepted: 07/09/2018] [Indexed: 11/09/2022] Open
Abstract
The pancreas of vertebrates is separately derived from both the dorsal and ventral endodermal domains. However, the difference between these two programs has been unclear. Here, using a pancreatic determination gene, Pdx1, driven GFP transgenic mouse strain, we identified Pdx1-GFP highly expressing cells (Pdx1high) and Pdx1-GFP lowly expressing cells (Pdx1low) in both embryonic dorsal Pdx1-expressing region (DPR) and ventral Pdx1-expressing region (VPR). We analyzed the transcriptomes of single Pdx1low and Pdx1high cells from the DPR and VPR. In the VPR, Pdx1low cells have an intermediate progenitor identity and can generate hepatoblasts, extrahepatobiliary cells, and Pdx1high pancreatic progenitor cells. In the DPR, Pdx1high cells are directly specified as pancreatic progenitors, whereas Pdx1low cells are precocious endocrine cells. Therefore, our study defines distinct road maps for dorsal and ventral pancreatic progenitor specification. The findings provide guidance for optimization of current β-cell induction protocols by following the in vivo dorsal pancreatic specification program.
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Affiliation(s)
- Lin-Chen Li
- Ministry of Education Key Laboratory of Cell Proliferation and Differentiation, College of Life Sciences, Peking-Tsinghua Center for Life Sciences, Beijing, China
| | - Wei-Lin Qiu
- Ministry of Education Key Laboratory of Cell Proliferation and Differentiation, College of Life Sciences, Peking-Tsinghua Center for Life Sciences, Beijing, China.,PKU-Tsinghua-NIBS Graduate Program, Peking University, Beijing, China
| | - Yu-Wei Zhang
- Ministry of Education Key Laboratory of Cell Proliferation and Differentiation, College of Life Sciences, Peking-Tsinghua Center for Life Sciences, Beijing, China
| | - Zi-Ran Xu
- Ministry of Education Key Laboratory of Cell Proliferation and Differentiation, College of Life Sciences, Peking-Tsinghua Center for Life Sciences, Beijing, China.,PKU-Tsinghua-NIBS Graduate Program, Peking University, Beijing, China
| | - Yi-Ni Xiao
- State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Caiying Hou
- General Hospital of PLA Rocket Force, Beijing, China
| | - Lamaoqiezhong
- Ministry of Education Key Laboratory of Cell Proliferation and Differentiation, College of Life Sciences, Peking-Tsinghua Center for Life Sciences, Beijing, China
| | - Peng Yu
- Ministry of Education Key Laboratory of Cell Proliferation and Differentiation, College of Life Sciences, Peking-Tsinghua Center for Life Sciences, Beijing, China
| | - Xin Cheng
- State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Cheng-Ran Xu
- Ministry of Education Key Laboratory of Cell Proliferation and Differentiation, College of Life Sciences, Peking-Tsinghua Center for Life Sciences, Beijing, China
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23
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Angelo JR, Tremblay KD. Identification and fate mapping of the pancreatic mesenchyme. Dev Biol 2018; 435:15-25. [PMID: 29329912 DOI: 10.1016/j.ydbio.2018.01.003] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/14/2017] [Revised: 01/06/2018] [Accepted: 01/06/2018] [Indexed: 12/25/2022]
Abstract
The murine pancreas buds from the ventral embryonic endoderm at approximately 8.75 dpc and a second pancreas bud emerges from the dorsal endoderm by 9.0 dpc. Although it is clear that secreted signals from adjacent mesoderm-derived sources are required for both the appropriate emergence and further refinement of the pancreatic endoderm, neither the exact signals nor the requisite tissue sources have been defined in mammalian systems. Herein we use DiI fate mapping of cultured murine embryos to identify the embryonic sources of both the early inductive and later condensed pancreatic mesenchyme. Despite being capable of supporting pancreas induction from dorsal endoderm in co-culture experiments, we find that in the context of the developing embryo, the dorsal aortae as well as the paraxial, intermediate, and lateral mesoderm derivatives only transiently associate with the dorsal pancreas bud, producing descendants that are decidedly anterior to the pancreas bud. Unlike these other mesoderm derivatives, the axial (notochord) descendants maintain association with the dorsal pre-pancreatic endoderm and early pancreas bud. This fate mapping data points to the notochord as the likely inductive source in vivo while also revealing dynamic morphogenetic movements displayed by individual mesodermal subtypes. Because none of the mesoderm examined above produced the pancreatic mesenchyme that condenses around the induced bud to support exocrine and endocrine differentiation, we also sought to identify the mesodermal origins of this mesenchyme. We identify a portion of the coelomic mesoderm that contributes to the condensed pancreatic mesenchyme. In conclusion, we identify a portion of the notochord as a likely source of the signals required to induce and maintain the early dorsal pancreas bud, demonstrate that the coelomic mesothelium contributes to the dorsal and ventral pancreatic mesenchyme, and provide insight into the dynamic morphological rearrangements of mesoderm-derived tissues during early organogenesis stages of mammalian development.
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Affiliation(s)
- Jesse R Angelo
- Department of Veterinary&Animal Sciences, University of Massachusetts, Amherst, MA, USA
| | - Kimberly D Tremblay
- Department of Veterinary&Animal Sciences, University of Massachusetts, Amherst, MA, USA.
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24
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Bastidas-Ponce A, Scheibner K, Lickert H, Bakhti M. Cellular and molecular mechanisms coordinating pancreas development. Development 2017; 144:2873-2888. [PMID: 28811309 DOI: 10.1242/dev.140756] [Citation(s) in RCA: 104] [Impact Index Per Article: 13.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022]
Abstract
The pancreas is an endoderm-derived glandular organ that participates in the regulation of systemic glucose metabolism and food digestion through the function of its endocrine and exocrine compartments, respectively. While intensive research has explored the signaling pathways and transcriptional programs that govern pancreas development, much remains to be discovered regarding the cellular processes that orchestrate pancreas morphogenesis. Here, we discuss the developmental mechanisms and principles that are known to underlie pancreas development, from induction and lineage formation to morphogenesis and organogenesis. Elucidating such principles will help to identify novel candidate disease genes and unravel the pathogenesis of pancreas-related diseases, such as diabetes, pancreatitis and cancer.
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Affiliation(s)
- Aimée Bastidas-Ponce
- Institute of Diabetes and Regeneration Research, Helmholtz Zentrum München, D-85764 Neuherberg, Germany.,Institute of Stem Cell Research, Helmholtz Zentrum München, D-85764 Neuherberg, Germany.,German Center for Diabetes Research (DZD), D-85764 Neuherberg, Germany.,Technical University of Munich, Medical Faculty, 81675 Munich, Germany
| | - Katharina Scheibner
- Institute of Diabetes and Regeneration Research, Helmholtz Zentrum München, D-85764 Neuherberg, Germany.,Institute of Stem Cell Research, Helmholtz Zentrum München, D-85764 Neuherberg, Germany.,German Center for Diabetes Research (DZD), D-85764 Neuherberg, Germany.,Technical University of Munich, Medical Faculty, 81675 Munich, Germany
| | - Heiko Lickert
- Institute of Diabetes and Regeneration Research, Helmholtz Zentrum München, D-85764 Neuherberg, Germany.,Institute of Stem Cell Research, Helmholtz Zentrum München, D-85764 Neuherberg, Germany.,German Center for Diabetes Research (DZD), D-85764 Neuherberg, Germany.,Technical University of Munich, Medical Faculty, 81675 Munich, Germany
| | - Mostafa Bakhti
- Institute of Diabetes and Regeneration Research, Helmholtz Zentrum München, D-85764 Neuherberg, Germany .,Institute of Stem Cell Research, Helmholtz Zentrum München, D-85764 Neuherberg, Germany.,German Center for Diabetes Research (DZD), D-85764 Neuherberg, Germany
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25
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Gnatenko DA, Kopantzev EP, Sverdlov ED. [Fibroblast growth factors and their effects in pancreas organogenesis]. BIOMEDIT︠S︡INSKAI︠A︡ KHIMII︠A︡ 2017; 63:211-218. [PMID: 28781254 DOI: 10.18097/pbmc20176303211] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/23/2022]
Abstract
Fibroblast growth factors (FGF) - growth factors that regulate many important biological processes, including proliferation and differentiation of embryonic cells during organogenesis. In this review, we will summarize current information about the involvement of FGFs in the pancreas organogenesis. Pancreas organogenesis is a complex process, which involves constant signaling from mesenchymal tissue. This orchestrates the activation of various regulator genes at specific stages, determining the specification of progenitor cells. Alterations in FGF/FGFR signaling pathway during this process lead to incorrect activation of the master genes, which leads to different pathologies during pancreas development. Understanding the full picture about role of FGF factors in pancreas development will make it possible to more accurately understand their role in other pathologies of this organ, including carcinogenesis.
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Affiliation(s)
- D A Gnatenko
- Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences
| | - E P Kopantzev
- Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences
| | - E D Sverdlov
- Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences
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26
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Múnera JO, Sundaram N, Rankin SA, Hill D, Watson C, Mahe M, Vallance JE, Shroyer NF, Sinagoga KL, Zarzoso-Lacoste A, Hudson JR, Howell JC, Chatuvedi P, Spence JR, Shannon JM, Zorn AM, Helmrath MA, Wells JM. Differentiation of Human Pluripotent Stem Cells into Colonic Organoids via Transient Activation of BMP Signaling. Cell Stem Cell 2017; 21:51-64.e6. [PMID: 28648364 PMCID: PMC5531599 DOI: 10.1016/j.stem.2017.05.020] [Citation(s) in RCA: 188] [Impact Index Per Article: 23.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/19/2016] [Revised: 02/28/2017] [Accepted: 05/25/2017] [Indexed: 02/07/2023]
Abstract
Gastric and small intestinal organoids differentiated from human pluripotent stem cells (hPSCs) have revolutionized the study of gastrointestinal development and disease. Distal gut tissues such as cecum and colon, however, have proved considerably more challenging to derive in vitro. Here we report the differentiation of human colonic organoids (HCOs) from hPSCs. We found that BMP signaling is required to establish a posterior SATB2+ domain in developing and postnatal intestinal epithelium. Brief activation of BMP signaling is sufficient to activate a posterior HOX code and direct hPSC-derived gut tube cultures into HCOs. In vitro, HCOs express colonic markers and contained colon-specific cell populations. Following transplantation into mice, HCOs undergo morphogenesis and maturation to form tissue that exhibits molecular, cellular, and morphologic properties of human colon. Together these data show BMP-dependent patterning of human hindgut into HCOs, which will be valuable for studying diseases including colitis and colon cancer.
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Affiliation(s)
- Jorge O Múnera
- Division of Developmental Biology, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA
| | - Nambirajan Sundaram
- Division of Pediatric General and Thoracic Surgery, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA
| | - Scott A Rankin
- Division of Developmental Biology, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA
| | - David Hill
- University of Michigan, Ann Arbor, MI 48109, USA
| | - Carey Watson
- Division of Pediatric General and Thoracic Surgery, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA
| | - Maxime Mahe
- Division of Pediatric General and Thoracic Surgery, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA
| | - Jefferson E Vallance
- Division of Gastroenterology, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA
| | - Noah F Shroyer
- Division of Gastroenterology, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA
| | - Katie L Sinagoga
- Division of Developmental Biology, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA
| | - Adrian Zarzoso-Lacoste
- Division of Developmental Biology, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA
| | - Jonathan R Hudson
- Division of Developmental Biology, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA
| | - Jonathan C Howell
- Division of Endocrinology, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA
| | - Praneet Chatuvedi
- Division of Developmental Biology, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA
| | | | - John M Shannon
- Division of Pulmonary Biology, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA
| | - Aaron M Zorn
- Division of Developmental Biology, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA; Center for Stem Cell and Organoid Medicine, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA
| | - Michael A Helmrath
- Division of Pediatric General and Thoracic Surgery, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA; Center for Stem Cell and Organoid Medicine, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA
| | - James M Wells
- Division of Developmental Biology, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA; Division of Endocrinology, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA; Center for Stem Cell and Organoid Medicine, Cincinnati Children's Hospital Research Foundation, Cincinnati, OH 45229, USA.
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27
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McCracken KW, Wells JM. Mechanisms of embryonic stomach development. Semin Cell Dev Biol 2017; 66:36-42. [PMID: 28238948 DOI: 10.1016/j.semcdb.2017.02.004] [Citation(s) in RCA: 32] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/24/2017] [Accepted: 02/20/2017] [Indexed: 12/18/2022]
Abstract
The stomach is a digestive organ that has important roles in human physiology and pathophysiology. The developmental origin of the stomach is the embryonic foregut, which also gives rise a number of other structures. There are several signaling pathways and transcription factors that are known to regulate stomach development at different stages, including foregut patterning, stomach specification, and gastric regionalization. These developmental events have important implications in later homeostasis and disease in the adult stomach. Here we will review the literature that has shaped our current understanding of the molecular mechanisms that coordinate gastric organogenesis. Further we will discuss how developmental paradigms have guided recent efforts to differentiate stomach tissue from pluripotent stem cells.
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Affiliation(s)
- Kyle W McCracken
- Division of Developmental Biology, 3333 Burnet Avenue, Cincinnati, OH 45229-3039, USA; Boston Children's Hospital, 300 Longwood Avenue, Boston, MA 02115, USA.
| | - James M Wells
- Division of Developmental Biology, 3333 Burnet Avenue, Cincinnati, OH 45229-3039, USA; Division of Endocrinology Cincinnati Children's Hospital Medical Center, 3333 Burnet Avenue, Cincinnati, OH 45229-3039, USA.
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28
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Zhang X, Chen Y, Ye Y, Wang J, Wang H, Yuan G, Lin Z, Wu Y, Zhang Y, Lin X. Wnt signaling promotes hindgut fate commitment through regulating multi-lineage genes during hESC differentiation. Cell Signal 2016; 29:12-22. [PMID: 27693749 DOI: 10.1016/j.cellsig.2016.09.009] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/07/2016] [Revised: 09/22/2016] [Accepted: 09/27/2016] [Indexed: 12/22/2022]
Abstract
Wnt signaling plays essential roles in both embryonic pattern formation and postembryonic tissue homoestasis. High levels of Wnt activity repress foregut identity and facilitate hindgut fate through forming a gradient of Wnt signaling activity along the anterior-posterior axis. Here, we examined the mechanisms of Wnt signaling in hindgut development by differentiating human embryonic stem cells (hESCs) into the hindgut progenitors. We observed severe morphological changes when Wnt signaling was blocked by using Wnt antagonist Dkk1. We performed deep-transcriptome sequencing (RNA-seq) and identified 240 Wnt-activated genes and 2023 Wnt-repressed genes, respectively. Clusters of Wnt targets showed enrichment in specific biological functions, such as "gastrointestinal or skeletal development" in the Wnt-activated targets and "neural or immune system development" in the Wnt-repressed targets. Moreover, we adopted a high-throughput chromatin immunoprecipitation and deep sequencing (ChIP-seq) approach to identify the genomic regions through which Wnt-activated transcription factor TCF7L2 regulated transcription. We identified 83 Wnt direct target candidates, including the hindgut marker CDX2 and the genes relevant to morphogenesis (MSX1, MSX2, LEF1, T, PDGFRB etc.) through combinatorial analysis of the RNA-seq and ChIP-seq data. Together, our study identified a series of direct and indirect Wnt targets in hindgut differentiation, and uncovered the diverse mechanisms of Wnt signaling in regulating multi-lineage differentiation.
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Affiliation(s)
- Xiujuan Zhang
- State Key Laboratory of Membrane Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China; University of Chinese Academy of Sciences, Beijing, China
| | - Ying Chen
- State Key Laboratory of Membrane Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China
| | - Ying Ye
- State Key Laboratory of Membrane Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China
| | - Jianfeng Wang
- Core Genomic Facility, CAS Key Laboratory of Genome Sciences & Information, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, China
| | - Hong Wang
- State Key Laboratory of Membrane Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China
| | - Guohong Yuan
- State Key Laboratory of Membrane Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China
| | - Zhe Lin
- State Key Laboratory of Integrated Management of Pest Insects and Rodents, Institute of Zoology, Chinese Academy of Sciences, Beijing, China
| | - Yihui Wu
- State Key Laboratory of Membrane Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China
| | - Yan Zhang
- State Key Laboratory of Membrane Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China
| | - Xinhua Lin
- State Key Laboratory of Membrane Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China; Division of Developmental Biology, Cincinnati Childrens Hospital Medical Center, Cincinnati, OH, United States; State Key Laboratory of Genetic Engineering, Institute of Genetics, Collaborative Innovation Center of Genetics and Development, School of Life Sciences, Fudan University, Shanghai, China.
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29
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Huang W, Beer RL, Delaspre F, Wang G, Edelman HE, Park H, Azuma M, Parsons MJ. Sox9b is a mediator of retinoic acid signaling restricting endocrine progenitor differentiation. Dev Biol 2016; 418:28-39. [PMID: 27565026 DOI: 10.1016/j.ydbio.2016.08.019] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/16/2015] [Revised: 08/16/2016] [Accepted: 08/17/2016] [Indexed: 10/21/2022]
Abstract
Centroacinar cells (CACs) are ductal Notch-responsive progenitors that in the larval zebrafish pancreas differentiate to form new islets and ultimately contribute to the majority of the adult endocrine mass. Uncovering the mechanisms regulating CAC differentiation will facilitate understanding how insulin-producing β cells are formed. Previously we reported retinoic acid (RA) signaling and Notch signaling both regulate larval CAC differentiation, suggesting a shared downstream intermediate. Sox9b is a transcription factor important for islet formation whose expression is upregulated by Notch signaling in larval CACs. Here we report that sox9b expression in larval CACs is also regulated by RA signaling. Therefore, we hypothesized that Sox9b is an intermediate between both RA- and Notch-signaling pathways. In order to study the role of Sox9b in larval CACs, we generated two cre/lox based transgenic tools, which allowed us to express full-length or truncated Sox9b in larval CACs. In this way we were able to perform spatiotemporal-controlled Sox9b gain- and loss-of-function studies and observe the subsequent effect on progenitor differentiation. Our results are consistent with Sox9b regulating CAC differentiation by being a downstream intermediate of both RA- and Notch-signaling pathways. We also demonstrate that adult zebrafish with only one functional allele of sox9b undergo accelerated β-cell regeneration, an observation consistent with sox9b regulating CAC differentiation in adults.
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Affiliation(s)
- Wei Huang
- McKusick-Nathans Institute for Genetic Medicine, Johns Hopkins University School of Medicine, 733 N. Broadway, 470 Miller Research Building, Baltimore, MD 21205, USA
| | - Rebecca L Beer
- McKusick-Nathans Institute for Genetic Medicine, Johns Hopkins University School of Medicine, 733 N. Broadway, 470 Miller Research Building, Baltimore, MD 21205, USA
| | - Fabien Delaspre
- McKusick-Nathans Institute for Genetic Medicine, Johns Hopkins University School of Medicine, 733 N. Broadway, 470 Miller Research Building, Baltimore, MD 21205, USA
| | - Guangliang Wang
- McKusick-Nathans Institute for Genetic Medicine, Johns Hopkins University School of Medicine, 733 N. Broadway, 470 Miller Research Building, Baltimore, MD 21205, USA
| | - Hannah E Edelman
- McKusick-Nathans Institute for Genetic Medicine, Johns Hopkins University School of Medicine, 733 N. Broadway, 470 Miller Research Building, Baltimore, MD 21205, USA
| | - Hyewon Park
- Department of Molecular Biosciences, University of Kansas, 1200 Sunnyside Avenue, Lawrence, KS 66045, USA
| | - Mizuki Azuma
- Department of Molecular Biosciences, University of Kansas, 1200 Sunnyside Avenue, Lawrence, KS 66045, USA
| | - Michael J Parsons
- McKusick-Nathans Institute for Genetic Medicine, Johns Hopkins University School of Medicine, 733 N. Broadway, 470 Miller Research Building, Baltimore, MD 21205, USA; Department of Surgery, Johns Hopkins University School of Medicine, 733 N. Broadway, 470 Miller Research Building, Baltimore, MD 21205, USA
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30
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Xuan S, Sussel L. GATA4 and GATA6 regulate pancreatic endoderm identity through inhibition of hedgehog signaling. Development 2016; 143:780-6. [PMID: 26932670 DOI: 10.1242/dev.127217] [Citation(s) in RCA: 37] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022]
Abstract
GATA4 and GATA6 are zinc finger transcription factors that have important functions in several mesodermal and endodermal organs, including heart, liver and pancreas. In humans, heterozygous mutations of either factor are associated with pancreatic agenesis; however, homozygous deletion of both Gata4 and Gata6 is necessary to disrupt pancreas development in mice. In this study, we demonstrate that arrested pancreatic development in Gata4(fl/fl); Gata6(fl/fl); Pdx1:Cre (pDKO) embryos is accompanied by the transition of ventral and dorsal pancreatic fates into intestinal or stomach lineages, respectively. These results indicate that GATA4 and GATA6 play essential roles in maintaining pancreas identity by regulating foregut endodermal fates. Remarkably, pancreatic anlagen derived from pDKO embryos also display a dramatic upregulation of hedgehog pathway components, which are normally absent from the presumptive pancreatic endoderm. Consistent with the erroneous activation of hedgehog signaling, we demonstrate that GATA4 and GATA6 are able to repress transcription through the sonic hedgehog (Shh) endoderm-specific enhancer MACS1 and that GATA-binding sites within this enhancer are necessary for this repressive activity. These studies establish the importance of GATA4/6-mediated inhibition of hedgehog signaling as a major mechanism regulating pancreatic endoderm specification during patterning of the gut tube.
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Affiliation(s)
- Shouhong Xuan
- Department of Genetics and Development, Columbia University, New York, NY 10032, USA
| | - Lori Sussel
- Department of Genetics and Development, Columbia University, New York, NY 10032, USA
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31
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Singh V, Saluja N. Phylogenetic and promoter analysis of islet amyloid polypeptide gene causing type 2 diabetes in mammalian species. Int J Diabetes Dev Ctries 2016. [DOI: 10.1007/s13410-016-0508-3] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
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32
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Dedhia PH, Bertaux-Skeirik N, Zavros Y, Spence JR. Organoid Models of Human Gastrointestinal Development and Disease. Gastroenterology 2016; 150:1098-1112. [PMID: 26774180 PMCID: PMC4842135 DOI: 10.1053/j.gastro.2015.12.042] [Citation(s) in RCA: 171] [Impact Index Per Article: 19.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/13/2015] [Revised: 12/23/2015] [Accepted: 12/23/2015] [Indexed: 12/21/2022]
Abstract
We have greatly advanced our ability to grow a diverse range of tissue-derived and pluripotent stem cell-derived gastrointestinal (GI) tissues in vitro. These systems, broadly referred to as organoids, have allowed the field to move away from the often nonphysiological, transformed cell lines that have been used for decades in GI research. Organoids are derived from primary tissues and have the capacity for long-term growth. They contain varying levels of cellular complexity and physiological similarity to native organ systems. We review the latest discoveries from studies of tissue-derived and pluripotent stem cell-derived intestinal, gastric, esophageal, liver, and pancreatic organoids. These studies have provided important insights into GI development, tissue homeostasis, and disease and might be used to develop personalized medicines.
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Affiliation(s)
- Priya H. Dedhia
- Department of Surgery, University of Michigan Medical School, Ann Arbor, MI 48109, USA,Center for Organogenesis, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Nina Bertaux-Skeirik
- Department of Molecular and Cellular Physiology, University of Cincinnati, Cincinnati, OH 45229, USA
| | - Yana Zavros
- Department of Molecular and Cellular Physiology, University of Cincinnati, Cincinnati, Ohio.
| | - Jason R. Spence
- Division of Gastroenterology, Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA,Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI 48109, USA,Center for Organogenesis, University of Michigan Medical School, Ann Arbor, MI 48109, USA,Authors for Correspondence: Jason R. Spence – , Twitter: @TheSpenceLab, Yana Zavros –
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Abstract
OBJECTIVES In injury conditions, myofibroblasts are induced to lay down matrix proteins and support the repair process. In this study, we investigated the role of myofibroblasts, particularly stellate cells, in the growth and regeneration of pancreatic β cells. METHODS We used both in vitro and in vivo approaches to address whether stellate cells may promote the growth of β cells. RESULTS Our experiments demonstrated that activated stellate cells support the proliferation of β cells in vitro. In vivo, mesenchymals surrounding the pancreatic islets are activated (induced to proliferate) in the islet regeneration model of Pten null mice. These mesenchymals display markers of pancreatic stellate cells, such as desmin and to a lesser extent, smooth muscle actin α. We have shown previously that targeted β-cell deletion of Pten lead to a significant increase in total islet mass. This phenotype was accompanied by an increase in peri-islet mitotic activity, particularly in islets injured by streptozotocin, a β cell-specific toxin. CONCLUSIONS Together with the in vitro observations, our data, here, suggest that that these mesenchymal cells may support the regeneration of the islets. Identifying how the communication occurs may provide clinically relevant mechanism for inducing β-cell regeneration.
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34
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Yang SF, Xue WJ, Duan YF, Xie LY, Lu WH, Zheng J, Yin AP. Nicotinamide Facilitates Mesenchymal Stem Cell Differentiation Into Insulin-Producing Cells and Homing to Pancreas in Diabetic Mice. Transplant Proc 2015; 47:2041-9. [DOI: 10.1016/j.transproceed.2015.05.019] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/13/2014] [Revised: 04/13/2015] [Accepted: 05/15/2015] [Indexed: 01/09/2023]
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35
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Christensen GL, Jacobsen MLB, Wendt A, Mollet IG, Friberg J, Frederiksen KS, Meyer M, Bruun C, Eliasson L, Billestrup N. Bone morphogenetic protein 4 inhibits insulin secretion from rodent beta cells through regulation of calbindin1 expression and reduced voltage-dependent calcium currents. Diabetologia 2015; 58:1282-90. [PMID: 25828920 DOI: 10.1007/s00125-015-3568-x] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/20/2014] [Accepted: 03/04/2015] [Indexed: 01/09/2023]
Abstract
AIMS/HYPOTHESIS Type 2 diabetes is characterised by progressive loss of pancreatic beta cell mass and function. Therefore, it is of therapeutic interest to identify factors with the potential to improve beta cell proliferation and insulin secretion. Bone morphogenetic protein 4 (BMP4) expression is increased in diabetic animals and BMP4 reduces glucose-stimulated insulin secretion (GSIS). Here, we investigate the molecular mechanism behind this inhibition. METHODS BMP4-mediated inhibition of GSIS was investigated in detail using single cell electrophysiological measurements and live cell Ca(2+) imaging. BMP4-mediated gene expression changes were investigated by microarray profiling, quantitative PCR and western blotting. RESULTS Prolonged exposure to BMP4 reduced GSIS from rodent pancreatic islets. This inhibition was associated with decreased exocytosis due to a reduced Ca(2+) current through voltage-dependent Ca(2+) channels. To identify proteins involved in the inhibition of GSIS, we investigated global gene expression changes induced by BMP4 in neonatal rat pancreatic islets. Expression of the Ca(2+)-binding protein calbindin1 was significantly induced by BMP4. Overexpression of calbindin1 in primary islet cells reduced GSIS, and the effect of BMP4 on GSIS was lost in islets from calbindin1 (Calb1) knockout mice. CONCLUSIONS/INTERPRETATION We found BMP4 treatment to markedly inhibit GSIS from rodent pancreatic islets in a calbindin1-dependent manner. Calbindin1 is suggested to mediate the effect of BMP4 by buffering Ca(2+) and decreasing Ca(2+) channel activity, resulting in diminished insulin exocytosis. Both BMP4 and calbindin1 are potential pharmacological targets for the treatment of beta cell dysfunction.
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Affiliation(s)
- Gitte L Christensen
- Department of Biomedical Sciences, University of Copenhagen, Nørre Alle 20, 2100, Copenhagen, Denmark
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36
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Talavera-Adame D, Dafoe DC. Endothelium-derived essential signals involved in pancreas organogenesis. World J Exp Med 2015; 5:40-49. [PMID: 25992319 PMCID: PMC4436939 DOI: 10.5493/wjem.v5.i2.40] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/06/2014] [Revised: 03/18/2015] [Accepted: 04/14/2015] [Indexed: 02/06/2023] Open
Abstract
Endothelial cells (ECs) are essential for pancreas differentiation, endocrine specification, and endocrine function. They are also involved in the physiopathology of type 1 and type 2 diabetes. During embryogenesis, aortic ECs provide specific factors that maintain the expression of key genes for pancreas development such as pancreatic and duodenal homeobox-1. Other unknown factors are also important for pancreatic endocrine specification and formation of insulin-producing beta cells. Endocrine precursors proliferate interspersed with ductal cells and exocrine precursors and, at some point of development, these endocrine precursors migrate to pancreatic mesenchyme and start forming the islets of Langerhans. By the end of the gestation and close to birth, these islets contain immature beta cells with the capacity to express vascular endothelial growth factor and therefore to recruit ECs from the surrounding microenvironment. ECs in turn produce factors that are essential to maintain insulin secretion in pancreatic beta cells. Once assembled, a cross talk between endocrine cells and ECs maintain the integrity of islets toward an adequate function during the whole life of the adult individual. This review will focus in the EC role in the differentiation and maturation of pancreatic beta cells during embryogenesis as well as the current knowledge about the involvement of endothelium to derive pancreatic beta cells in vitro from mouse or human pluripotent stem cells.
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37
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Duan FF, Liu JH, March JC. Engineered commensal bacteria reprogram intestinal cells into glucose-responsive insulin-secreting cells for the treatment of diabetes. Diabetes 2015; 64:1794-803. [PMID: 25626737 PMCID: PMC4407861 DOI: 10.2337/db14-0635] [Citation(s) in RCA: 149] [Impact Index Per Article: 14.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/22/2014] [Accepted: 12/10/2014] [Indexed: 12/25/2022]
Abstract
The inactive full-length form of GLP-1(1-37) stimulates conversion of both rat and human intestinal epithelial cells into insulin-secreting cells. We investigated whether oral administration of human commensal bacteria engineered to secrete GLP-1(1-37) could ameliorate hyperglycemia in a rat model of diabetes by reprogramming intestinal cells into glucose-responsive insulin-secreting cells. Diabetic rats were fed daily with human lactobacilli engineered to secrete GLP-1(1-37). Diabetic rats fed GLP-1-secreting bacteria showed significant increases in insulin levels and, additionally, were significantly more glucose tolerant than those fed the parent bacterial strain. These rats developed insulin-producing cells within the upper intestine in numbers sufficient to replace ∼25-33% of the insulin capacity of nondiabetic healthy rats. Intestinal tissues in rats with reprogrammed cells expressed MafA, PDX-1, and FoxA2. HNF-6 expression was observed only in crypt epithelia expressing insulin and not in epithelia located higher on the villous axis. Staining for other cell markers in rats treated with GLP-1(1-37)-secreting bacteria suggested that normal function was not inhibited by the close physical proximity of reprogrammed cells. These results provide evidence of the potential for a safe and effective nonabsorbed oral treatment for diabetes and support the concept of engineered commensal bacterial signaling to mediate enteric cell function in vivo.
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Affiliation(s)
- Franklin F Duan
- Department of Biological and Environmental Engineering, Cornell University, Ithaca, NY
| | - Joy H Liu
- Department of Biological and Environmental Engineering, Cornell University, Ithaca, NY
| | - John C March
- Department of Biological and Environmental Engineering, Cornell University, Ithaca, NY
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38
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39
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Sinagoga KL, Wells JM. Generating human intestinal tissues from pluripotent stem cells to study development and disease. EMBO J 2015; 34:1149-63. [PMID: 25792515 DOI: 10.15252/embj.201490686] [Citation(s) in RCA: 70] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/27/2014] [Accepted: 03/09/2015] [Indexed: 01/05/2023] Open
Abstract
As one of the largest and most functionally complex organs of the human body, the intestines are primarily responsible for the breakdown and uptake of macromolecules from the lumen and the subsequent excretion of waste from the body. However, the intestine is also an endocrine organ, regulating digestion, metabolism, and feeding behavior. Intricate neuronal, lymphatic, immune, and vascular systems are integrated into the intestine and are required for its digestive and endocrine functions. In addition, the gut houses an extensive population of microbes that play roles in digestion, global metabolism, barrier function, and host-parasite interactions. With such an extensive array of cell types working and performing in one essential organ, derivation of functional intestinal tissues from human pluripotent stem cells (PSCs) represents a significant challenge. Here we will discuss the intricate developmental processes and cell types that are required for assembly of this highly complex organ and how embryonic processes, particularly morphogenesis, have been harnessed to direct differentiation of PSCs into 3-dimensional human intestinal organoids (HIOs) in vitro. We will further describe current uses of HIOs in development and disease research and how additional tissue complexity might be engineered into HIOs for better functionality and disease modeling.
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Affiliation(s)
- Katie L Sinagoga
- Division of Developmental Biology, Cincinnati Children's Hospital Medical Center, Cincinnati, OH, USA
| | - James M Wells
- Division of Developmental Biology, Cincinnati Children's Hospital Medical Center, Cincinnati, OH, USA Division of Endocrinology, Cincinnati Children's Hospital Medical Center, Cincinnati, OH, USA
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40
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Ikonomou L, Kotton DN. Derivation of Endodermal Progenitors From Pluripotent Stem Cells. J Cell Physiol 2015; 230:246-58. [PMID: 25160562 PMCID: PMC4344429 DOI: 10.1002/jcp.24771] [Citation(s) in RCA: 21] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/19/2014] [Accepted: 08/22/2014] [Indexed: 01/18/2023]
Abstract
Stem and progenitor cells play important roles in organogenesis during development and in tissue homeostasis and response to injury postnatally. As the regenerative capacity of many human tissues is limited, cell replacement therapies hold great promise for human disease management. Pluripotent stem cells such as embryonic stem (ES) cells and induced pluripotent stem (iPS) cells are prime candidates for the derivation of unlimited quantities of clinically relevant cell types through development of directed differentiation protocols, that is, the recapitulation of developmental milestones in in vitro cell culture. Tissue-specific progenitors, including progenitors of endodermal origin, are important intermediates in such protocols since they give rise to all mature parenchymal cells. In this review, we focus on the in vivo biology of embryonic endodermal progenitors in terms of key transcription factors and signaling pathways. We critically review the emerging literature aiming to apply this basic knowledge to achieve the efficient and reproducible in vitro derivation of endodermal progenitors such as pancreas, liver and lung precursor cells.
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Affiliation(s)
- Laertis Ikonomou
- Center for Regenerative Medicine, Boston University and Boston
Medical Center, Boston, MA, USA
- Boston University Pulmonary Center, Boston University School of
Medicine, Boston, MA, USA
| | - Darrell N. Kotton
- Center for Regenerative Medicine, Boston University and Boston
Medical Center, Boston, MA, USA
- Boston University Pulmonary Center, Boston University School of
Medicine, Boston, MA, USA
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41
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Ramasamy SK, Kusumbe AP, Adams RH. Regulation of tissue morphogenesis by endothelial cell-derived signals. Trends Cell Biol 2014; 25:148-57. [PMID: 25529933 DOI: 10.1016/j.tcb.2014.11.007] [Citation(s) in RCA: 139] [Impact Index Per Article: 12.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/12/2014] [Revised: 11/21/2014] [Accepted: 11/24/2014] [Indexed: 02/08/2023]
Abstract
Endothelial cells (ECs) form an extensive network of blood vessels that has numerous essential functions in the vertebrate body. In addition to their well-established role as a versatile transport network, blood vessels can induce organ formation or direct growth and differentiation processes by providing signals in a paracrine (angiocrine) fashion. Tissue repair also requires the local restoration of vasculature. ECs are emerging as important signaling centers that coordinate regeneration and help to prevent deregulated, disease-promoting processes. Vascular cells are also part of stem cell niches and have key roles in hematopoiesis, bone formation, and neurogenesis. Here, we review these newly identified roles of ECs in the regulation of organ morphogenesis, maintenance, and regeneration.
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Affiliation(s)
- Saravana K Ramasamy
- Max-Planck-Institute for Molecular Biomedicine, Department of Tissue Morphogenesis, University of Münster, Faculty of Medicine, D-48149 Münster, Germany
| | - Anjali P Kusumbe
- Max-Planck-Institute for Molecular Biomedicine, Department of Tissue Morphogenesis, University of Münster, Faculty of Medicine, D-48149 Münster, Germany
| | - Ralf H Adams
- Max-Planck-Institute for Molecular Biomedicine, Department of Tissue Morphogenesis, University of Münster, Faculty of Medicine, D-48149 Münster, Germany.
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42
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Kumar SS, Alarfaj AA, Munusamy MA, Singh AJAR, Peng IC, Priya SP, Hamat RA, Higuchi A. Recent developments in β-cell differentiation of pluripotent stem cells induced by small and large molecules. Int J Mol Sci 2014; 15:23418-47. [PMID: 25526563 PMCID: PMC4284775 DOI: 10.3390/ijms151223418] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/29/2014] [Revised: 12/03/2014] [Accepted: 12/08/2014] [Indexed: 12/21/2022] Open
Abstract
Human pluripotent stem cells, including human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs), hold promise as novel therapeutic tools for diabetes treatment because of their self-renewal capacity and ability to differentiate into beta (β)-cells. Small and large molecules play important roles in each stage of β-cell differentiation from both hESCs and hiPSCs. The small and large molecules that are described in this review have significantly advanced efforts to cure diabetic disease. Lately, effective protocols have been implemented to induce hESCs and human mesenchymal stem cells (hMSCs) to differentiate into functional β-cells. Several small molecules, proteins, and growth factors promote pancreatic differentiation from hESCs and hMSCs. These small molecules (e.g., cyclopamine, wortmannin, retinoic acid, and sodium butyrate) and large molecules (e.g. activin A, betacellulin, bone morphogentic protein (BMP4), epidermal growth factor (EGF), fibroblast growth factor (FGF), keratinocyte growth factor (KGF), hepatocyte growth factor (HGF), noggin, transforming growth factor (TGF-α), and WNT3A) are thought to contribute from the initial stages of definitive endoderm formation to the final stages of maturation of functional endocrine cells. We discuss the importance of such small and large molecules in uniquely optimized protocols of β-cell differentiation from stem cells. A global understanding of various small and large molecules and their functions will help to establish an efficient protocol for β-cell differentiation.
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Affiliation(s)
- S Suresh Kumar
- Department of Medical Microbiology and Parasitology, Universities Putra Malaysia, Serdang 43400, Selangor, Malaysia.
| | - Abdullah A Alarfaj
- Department of Botany and Microbiology, College of Science, King Saud University, Riyadh 11451, Saudi Arabia.
| | - Murugan A Munusamy
- Department of Botany and Microbiology, College of Science, King Saud University, Riyadh 11451, Saudi Arabia.
| | - A J A Ranjith Singh
- Department of Bioscience, Jacintha Peter College of Arts and Sciences, Ayakudi, Tenkasi, Tamilnadu 627852, India.
| | - I-Chia Peng
- Department of Chemical and Materials Engineering, National Central University, No. 300, Jhongda RD., Jhongli, Taoyuan 32001, Taiwan.
| | - Sivan Padma Priya
- Department of Basic Science and Department of Surgical Sciences, Ajman University of Science and Technology-Fujairah Campus, P.O. Box 9520, Al Fujairah, United Arab Emirates.
| | - Rukman Awang Hamat
- Department of Medical Microbiology and Parasitology, Universities Putra Malaysia, Serdang 43400, Selangor, Malaysia.
| | - Akon Higuchi
- Department of Botany and Microbiology, College of Science, King Saud University, Riyadh 11451, Saudi Arabia.
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Tan G, Elefanty AG, Stanley EG. β-cell regeneration and differentiation: how close are we to the 'holy grail'? J Mol Endocrinol 2014; 53:R119-29. [PMID: 25385843 DOI: 10.1530/jme-14-0188] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Diabetes can be managed by careful monitoring of blood glucose and timely delivery of exogenous insulin. However, even with fastidious compliance, people with diabetes can suffer from numerous complications including atherosclerosis, retinopathy, neuropathy, and kidney disease. This is because delivery of exogenous insulin coupled with glucose monitoring cannot provide the fine level of glucose control normally provided by endogenous β-cells in the context of intact islets. Moreover, a subset of people with diabetes lack awareness of hypoglycemic events; a status that can have grave consequences. Therefore, much effort has been focused on replacing lost or dysfunctional β-cells with cells derived from other sources. The advent of stem cell biology and cellular reprogramming strategies have provided impetus to this work and raised hopes that a β-cell replacement therapy is on the horizon. In this review, we look at two components that will be required for successful β-cell replacement therapy: a reliable and safe source of β-cells and a mechanism by which such cells can be delivered and protected from host immune destruction. Particular attention is paid to insulin-producing cells derived from pluripotent stem cells because this platform addresses the issue of scale, one of the more significant hurdles associated with potential cell-based therapies. We also review methods for encapsulating transplanted cells, a technique that allows grafts to evade immune attack and survive for a long term in the absence of ongoing immunosuppression. In surveying the literature, we conclude that there are still several substantial hurdles that need to be cleared before a stem cell-based β-cell replacement therapy for diabetes becomes a reality.
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Affiliation(s)
- Gemma Tan
- Department of Anatomy and Developmental BiologyMonash University, Building 73, Clayton, Victoria 3800, AustraliaMurdoch Childrens Research InstituteThe Royal Children's Hospital, Flemington Road, Parkville, Victoria 3052, AustraliaDepartment of PaediatricsThe Royal Children's Hospital, University of Melbourne, Flemington Road, Parkville, Victoria 3052, Australia Department of Anatomy and Developmental BiologyMonash University, Building 73, Clayton, Victoria 3800, AustraliaMurdoch Childrens Research InstituteThe Royal Children's Hospital, Flemington Road, Parkville, Victoria 3052, AustraliaDepartment of PaediatricsThe Royal Children's Hospital, University of Melbourne, Flemington Road, Parkville, Victoria 3052, Australia
| | - Andrew G Elefanty
- Department of Anatomy and Developmental BiologyMonash University, Building 73, Clayton, Victoria 3800, AustraliaMurdoch Childrens Research InstituteThe Royal Children's Hospital, Flemington Road, Parkville, Victoria 3052, AustraliaDepartment of PaediatricsThe Royal Children's Hospital, University of Melbourne, Flemington Road, Parkville, Victoria 3052, Australia Department of Anatomy and Developmental BiologyMonash University, Building 73, Clayton, Victoria 3800, AustraliaMurdoch Childrens Research InstituteThe Royal Children's Hospital, Flemington Road, Parkville, Victoria 3052, AustraliaDepartment of PaediatricsThe Royal Children's Hospital, University of Melbourne, Flemington Road, Parkville, Victoria 3052, Australia Department of Anatomy and Developmental BiologyMonash University, Building 73, Clayton, Victoria 3800, AustraliaMurdoch Childrens Research InstituteThe Royal Children's Hospital, Flemington Road, Parkville, Victoria 3052, AustraliaDepartment of PaediatricsThe Royal Children's Hospital, University of Melbourne, Flemington Road, Parkville, Victoria 3052, Australia
| | - Edouard G Stanley
- Department of Anatomy and Developmental BiologyMonash University, Building 73, Clayton, Victoria 3800, AustraliaMurdoch Childrens Research InstituteThe Royal Children's Hospital, Flemington Road, Parkville, Victoria 3052, AustraliaDepartment of PaediatricsThe Royal Children's Hospital, University of Melbourne, Flemington Road, Parkville, Victoria 3052, Australia Department of Anatomy and Developmental BiologyMonash University, Building 73, Clayton, Victoria 3800, AustraliaMurdoch Childrens Research InstituteThe Royal Children's Hospital, Flemington Road, Parkville, Victoria 3052, AustraliaDepartment of PaediatricsThe Royal Children's Hospital, University of Melbourne, Flemington Road, Parkville, Victoria 3052, Australia Department of Anatomy and Developmental BiologyMonash University, Building 73, Clayton, Victoria 3800, AustraliaMurdoch Childrens Research InstituteThe Royal Children's Hospital, Flemington Road, Parkville, Victoria 3052, AustraliaDepartment of PaediatricsThe Royal Children's Hospital, University of Melbourne, Flemington Road, Parkville, Victoria 3052, Australia
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Modelling human development and disease in pluripotent stem-cell-derived gastric organoids. Nature 2014; 516:400-4. [PMID: 25363776 PMCID: PMC4270898 DOI: 10.1038/nature13863] [Citation(s) in RCA: 705] [Impact Index Per Article: 64.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/12/2014] [Accepted: 09/12/2014] [Indexed: 12/16/2022]
Abstract
Gastric diseases, including peptic ulcer disease and gastric cancer, affect 10% of the world’s population and are largely due to chronic H. pylori infection1–3. Species differences in embryonic development and architecture of the adult stomach make animal models suboptimal for studying human stomach organogenesis and pathogenesis4, and there is no experimental model of normal human gastric mucosa. Here we report the de novo generation of three-dimensional human gastric tissue in vitro through the directed differentiation of human pluripotent stem cells (hPSCs). We identified that temporal manipulation of the FGF, WNT, BMP, retinoic acid and EGF signaling pathways and three-dimensional growth are sufficient to generate human gastric organoids (hGOs). Developing hGOs progressed through molecular and morphogenetic stages that were nearly identical to the developing antrum of the mouse stomach. Organoids formed primitive gastric gland- and pit-like domains, proliferative zones containing LGR5-expressing cells, surface and antral mucous cells, and a diversity of gastric endocrine cells. We used hGO cultures to identify novel signaling mechanisms that regulate early endoderm patterning and gastric endocrine cell differentiation upstream of the transcription factor NEUROG3. Using hGOs to model pathogenesis of human disease, we found that H. pylori infection resulted in rapid association of the virulence factor CagA with the c-Met receptor, activation of signaling and induction of epithelial proliferation. Together, these studies describe a novel and robust in vitro system for elucidating the mechanisms underlying human stomach development and disease.
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Abstract
With the high prevalence of gastrointestinal disorders, there is great interest in establishing in vitro models of human intestinal disease and in developing drug-screening platforms that more accurately represent the complex physiology of the intestine. We will review how recent advances in developmental and stem cell biology have made it possible to generate complex, three-dimensional, human intestinal tissues in vitro through directed differentiation of human pluripotent stem cells. These are currently being used to study human development, genetic forms of disease, intestinal pathogens, metabolic disease and cancer.
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Affiliation(s)
- James M Wells
- Division of Developmental Biology, Cincinnati Children's Hospital Medical Center, 3333 Burnet Avenue, Cincinnati, OH 45229-3039, USA
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46
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Bose B, Katikireddy KR, Shenoy PS. Regenerative medicine for diabetes: differentiation of human pluripotent stem cells into functional β-cells in vitro and their proposed journey to clinical translation. VITAMINS AND HORMONES 2014; 95:223-48. [PMID: 24559920 DOI: 10.1016/b978-0-12-800174-5.00009-0] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/04/2022]
Abstract
Diabetes is a group of metabolic diseases, rising globally at an alarming rate. Type 1 (juvenile diabetes) is the autoimmune version of diabetes where the pancreas is unable to produce insulin, whereas type 2 (adult onset diabetes) is caused due to insulin resistance of the cells. In either of the cases, elevated blood glucose levels are observed which leads to progressive comorbidity like renal failure, cardiovascular disease, retinopathy, etc. Metformin, sulphonyl urea group of drugs, as well as insulin injections are the available therapies. In advanced cases of diabetes, the drug alone or drug in combination with insulin injections are not able to maintain a steady level of blood glucose. Moreover, frequent insulin injections are rather cumbersome for the patient. So, regenerative medicine could be a permanent solution for fighting diabetes. Islet transplantation has been tried with a limited amount of success on a large population of diabetics because of the shortage of cadaveric pancreas. Therefore, the best proposed alternative is regenerative medicine involving human pluripotent stem cell (hPSC)-derived beta islet transplantation which can be obtained in large quantities. Efficient protocols for in vitro differentiation of hPSC into a large number of sustained insulin-producing beta cells for transplantation will be considered to be a giant leap to address global rise in diabetic cases. Although most of the protocols mimic in vivo pancreatic development in humans, considerable amount of lacuna persists for near-perfect differentiation strategies. Moreover, beta islets differentiated from hPSC have not yet been successfully translated under clinical scenario.
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Affiliation(s)
- Bipasha Bose
- Nanyang Technological University, School of Biological Sciences, NTU Lab Location @ Level 2 Singapore Institute for Clinical Sciences, Brenner Centre for Molecular Medicine, Singapore, Singapore.
| | | | - P Sudheer Shenoy
- Nanyang Technological University, School of Biological Sciences, NTU Lab Location @ Level 2 Singapore Institute for Clinical Sciences, Brenner Centre for Molecular Medicine, Singapore, Singapore
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Lancman JJ, Zvenigorodsky N, Gates KP, Zhang D, Solomon K, Humphrey RK, Kuo T, Setiawan L, Verkade H, Chi YI, Jhala US, Wright CVE, Stainier DYR, Dong PDS. Specification of hepatopancreas progenitors in zebrafish by hnf1ba and wnt2bb. Development 2013; 140:2669-79. [PMID: 23720049 PMCID: PMC3678338 DOI: 10.1242/dev.090993] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 04/25/2013] [Indexed: 12/16/2022]
Abstract
Although the liver and ventral pancreas are thought to arise from a common multipotent progenitor pool, it is unclear whether these progenitors of the hepatopancreas system are specified by a common genetic mechanism. Efforts to determine the role of Hnf1b and Wnt signaling in this crucial process have been confounded by a combination of factors, including a narrow time frame for hepatopancreas specification, functional redundancy among Wnt ligands, and pleiotropic defects caused by either severe loss of Wnt signaling or Hnf1b function. Using a novel hypomorphic hnf1ba zebrafish mutant that exhibits pancreas hypoplasia, as observed in HNF1B monogenic diabetes, we show that hnf1ba plays essential roles in regulating β-cell number and pancreas specification, distinct from its function in regulating pancreas size and liver specification, respectively. By combining Hnf1ba partial loss of function with conditional loss of Wnt signaling, we uncover a crucial developmental window when these pathways synergize to specify the entire ventrally derived hepatopancreas progenitor population. Furthermore, our in vivo genetic studies demonstrate that hnf1ba generates a permissive domain for Wnt signaling activity in the foregut endoderm. Collectively, our findings provide a new model for HNF1B function, yield insight into pancreas and β-cell development, and suggest a new mechanism for hepatopancreatic specification.
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Affiliation(s)
- Joseph J. Lancman
- Sanford Children’s Health Research Center, Programs in Genetic Disease, Development and Aging, and Stem Cell and Regenerative Biology, Graduate School of Biomedical Sciences, Sanford-Burnham Medical Research Institute, 10901 North Torrey Pines Road, La Jolla, CA 92037, USA
| | - Natasha Zvenigorodsky
- Department of Biochemistry and Biophysics, Programs in Developmental Biology, Genetics and Human Genetics, and the Diabetes Center and Liver Center, University of California, San Francisco, 1550 Fourth Street, San Francisco, CA 94158, USA
| | - Keith P. Gates
- Sanford Children’s Health Research Center, Programs in Genetic Disease, Development and Aging, and Stem Cell and Regenerative Biology, Graduate School of Biomedical Sciences, Sanford-Burnham Medical Research Institute, 10901 North Torrey Pines Road, La Jolla, CA 92037, USA
| | - Danhua Zhang
- Sanford Children’s Health Research Center, Programs in Genetic Disease, Development and Aging, and Stem Cell and Regenerative Biology, Graduate School of Biomedical Sciences, Sanford-Burnham Medical Research Institute, 10901 North Torrey Pines Road, La Jolla, CA 92037, USA
| | - Keely Solomon
- Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA
| | - Rohan K. Humphrey
- Pediatric Diabetes Research Center, UCSD School of Medicine, La Jolla CA 92037, USA
| | - Taiyi Kuo
- Department of Biochemistry and Biophysics, Programs in Developmental Biology, Genetics and Human Genetics, and the Diabetes Center and Liver Center, University of California, San Francisco, 1550 Fourth Street, San Francisco, CA 94158, USA
| | - Linda Setiawan
- Department of Biochemistry and Biophysics, Programs in Developmental Biology, Genetics and Human Genetics, and the Diabetes Center and Liver Center, University of California, San Francisco, 1550 Fourth Street, San Francisco, CA 94158, USA
| | - Heather Verkade
- Department of Biochemistry and Biophysics, Programs in Developmental Biology, Genetics and Human Genetics, and the Diabetes Center and Liver Center, University of California, San Francisco, 1550 Fourth Street, San Francisco, CA 94158, USA
- School of Biological Sciences, Monash University, Clayton, VIC 3800, Australia
| | - Young-In Chi
- Hormel Institute, University of Minnesota, Austin, MN 55912, USA
| | - Ulupi S. Jhala
- Pediatric Diabetes Research Center, UCSD School of Medicine, La Jolla CA 92037, USA
| | - Christopher V. E. Wright
- Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA
| | - Didier Y. R. Stainier
- Department of Biochemistry and Biophysics, Programs in Developmental Biology, Genetics and Human Genetics, and the Diabetes Center and Liver Center, University of California, San Francisco, 1550 Fourth Street, San Francisco, CA 94158, USA
| | - P. Duc Si Dong
- Sanford Children’s Health Research Center, Programs in Genetic Disease, Development and Aging, and Stem Cell and Regenerative Biology, Graduate School of Biomedical Sciences, Sanford-Burnham Medical Research Institute, 10901 North Torrey Pines Road, La Jolla, CA 92037, USA
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48
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Stanger BZ, Hebrok M. Control of cell identity in pancreas development and regeneration. Gastroenterology 2013; 144:1170-9. [PMID: 23622126 PMCID: PMC3639438 DOI: 10.1053/j.gastro.2013.01.074] [Citation(s) in RCA: 107] [Impact Index Per Article: 8.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/19/2012] [Revised: 01/05/2013] [Accepted: 01/14/2013] [Indexed: 02/07/2023]
Abstract
The endocrine and exocrine cells in the adult pancreas are not static, but can change their differentiation state in response to injury or stress. This concept of cells in flux means that there may be ways to generate certain types of cells (such as insulin-producing β-cells) and prevent formation of others (such as transformed neoplastic cells). We review different aspects of cell identity in the pancreas, discussing how cells achieve their identity during embryonic development and maturation, and how this identity remains plastic, even in the adult pancreas.
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Affiliation(s)
- Ben Z. Stanger
- Division of Gastroenterology, Department of Medicine, Department of Cell and Developmental Biology and Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104
| | - Matthias Hebrok
- Diabetes Center, Department of Medicine, University of California, San Francisco, CA 94143, USA
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Abstract
Liver is a prime organ responsible for synthesis, metabolism, and detoxification. The organ is endodermal in origin and its development is regulated by temporal, complex, and finely balanced cellular and molecular interactions that dictate its origin, growth, and maturation. We discuss the relevance of endoderm patterning, which truly is the first step toward mapping of domains that will give rise to specific organs. Once foregut patterning is completed, certain cells within the foregut endoderm gain competence in the form of expression of certain transcription factors that allow them to respond to certain inductive signals. Hepatic specification is then a result of such inductive signals, which often emanate from the surrounding mesenchyme. During hepatic specification bipotential hepatic stem cells or hepatoblasts become apparent and undergo expansion, which results in a visible liver primordium during the stage of hepatic morphogenesis. Hepatoblasts next differentiate into either hepatocytes or cholangiocytes. The expansion and differentiation is regulated by cellular and molecular interactions between hepatoblasts and mesenchymal cells including sinusoidal endothelial cells, stellate cells, and also innate hematopoietic elements. Further maturation of hepatocytes and cholangiocytes continues during late hepatic development as a function of various growth factors. At this time, liver gains architectural novelty in the form of zonality and at cellular level acquires polarity. A comprehensive elucidation of such finely tuned developmental cues have been the basis of transdifferentiation of various types of stem cells to hepatocyte-like cells for purposes of understanding health and disease and for therapeutic applications.
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Affiliation(s)
- Donghun Shin
- Department of Developmental Biology, University of Pittsburgh, School of Medicine, Pittsburgh, Pennsylvania, USA.
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50
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Li J, Zhu L, Qu X, Li J, Lin R, Liao L, Wang J, Wang S, Xu Q, Zhao RC. Stepwise differentiation of human adipose-derived mesenchymal stem cells toward definitive endoderm and pancreatic progenitor cells by mimicking pancreatic development in vivo. Stem Cells Dev 2013; 22:1576-87. [PMID: 23259909 DOI: 10.1089/scd.2012.0148] [Citation(s) in RCA: 32] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
Pancreatic progenitor (PP) cells are tissue-committed cells, which can differentiate into all kinds of pancreatic cells. They are potential candidates for regeneration of pancreatic tissue. However, it is unfeasible to acquire PP cells from pancreatic tissues and expand them in vitro. Generation of PP cells from adipose tissue-derived mesenchymal stem cells (AD-MSCs) would provide an unlimited source of PP cells. Here we developed a 2-step stepwise protocol, which induced AD-MSCs to generate FOXA2- or SOX17-positive definitive endoderm (DE) (5 days) and pancreatic and duodenal homeobox gene 1 (PDX1)-positive PP cells (4-6 days). By mimicking the developmental progress in embryonic development, we optimized the timing and combination of cytokines to activate the key signaling pathways during pancreatic development. We found that activating the Nodal/Activin signal with Activin A could induce differentiation of AD-MSCs toward DE, which could be further promoted by the Wnt signaling pathway activator Wnt3a. Besides, transient T (BRACHYURY)(+) mesendodermal cells were observed during formation of DE from AD-MSCs. Subsequently, the Wnt signaling pathway inhibitor Dkk1 along with retinoic acid/FGF2 (60 ng/mL) further induced AD-MSC-derived DE cells to differentiate into PDX1-positive PP cells. The derived PP cells were capable to form pancreatic endocrine or exocrine cells. In conclusion, we established a stepwise protocol that could derive DE and PP cells from AD-MSCs. It might provide an unlimited source of autologous PP cells for pancreatic diseases.
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Affiliation(s)
- Jing Li
- Center of Excellence in Tissue Engineering, Department of Cell Biology, Institute of Basic Medical Sciences and School of Basic Medicine, Peking Union Medical College Hospital, Beijing, People's Republic of China
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