Generation of hepato-biliary-pancreatic organoid from human pluripotent stem cells

Abstract Organogenesis is a complex and inter-connected process, orchestrated by multiple boundary tissue interactions. Here, we established the protocol of the continuous patterning of hepatic, biliary and pancreatic structures from a three-dimensional culture of human pluripotent stem cell (PSC). The boundary interactions between anterior and posterior gut spheroids differentiated from human PSCs enables autonomous emergence of hepato-biliary-pancreatic (HBP) organ domains in the absence of extrinsic factor supply. This anterior-poterior gut interaction protocol can be used to model the early human HBP organogenesis process in vitro.

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Convergence of human pluripotent stem cell, organoid, and genome editing technologies

Experimental Biology and Medicine ◽

10.1177/1535370220985808 ◽

2021 ◽

pp. 153537022098580

Author(s):

Lin Wang ◽

Zhaohui Ye ◽

Yoon-Young Jang

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Genome Editing ◽

Pluripotent Stem Cells ◽

Pluripotent Stem Cell ◽

Three Dimensional ◽

Experimental Models ◽

Human Pluripotent Stem Cell ◽

Research Areas ◽

Technological Advances

The last decade has seen many exciting technological breakthroughs that greatly expanded the toolboxes for biological and biomedical research, yet few have had more impact than induced pluripotent stem cells and modern-day genome editing. These technologies are providing unprecedented opportunities to improve physiological relevance of experimental models, further our understanding of developmental processes, and develop novel therapies. One of the research areas that benefit greatly from these technological advances is the three-dimensional human organoid culture systems that resemble human tissues morphologically and physiologically. Here we summarize the development of human pluripotent stem cells and their differentiation through organoid formation. We further discuss how genetic modifications, genome editing in particular, were applied to answer basic biological and biomedical questions using organoid cultures of both somatic and pluripotent stem cell origins. Finally, we discuss the potential challenges of applying human pluripotent stem cell and organoid technologies for safety and efficiency evaluation of emerging genome editing tools.

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Induction of functional hypothalamus and pituitary tissues from pluripotent stem cells for regenerative medicine

Journal of the Endocrine Society ◽

10.1210/jendso/bvaa188 ◽

2020 ◽

Author(s):

Mayuko Kano ◽

Hidetaka Suga ◽

Hiroshi Arima

Keyword(s):

Stem Cells ◽

Regenerative Medicine ◽

Pluripotent Stem Cells ◽

Hormone Replacement ◽

Three Dimensional ◽

Embryonic Stem ◽

Hormone Deficiency ◽

Adrenal Crisis ◽

Mouse Escs

Abstract The hypothalamus and pituitary have been identified to play essential roles in maintaining homeostasis. Various diseases can disrupt the functions of these systems, which can often result in serious lifelong symptoms. The current treatment for hypopituitarism involves hormone replacement therapy. However, exogenous drug administration cannot mimic the physiological changes that are a result of hormone requirements. Therefore, patients are at a high risk of severe hormone deficiency, including adrenal crisis. Pluripotent stem cells (PSCs) self-proliferate and differentiate into all types of cells. The generation of endocrine tissues from PSCs has been considered as another new treatment for hypopituitarism. Our colleagues established a three-dimensional culture method for embryonic stem cells (ESCs). In this culture, the ESC-derived aggregates exhibit self-organization and spontaneous formation of highly ordered patterning. Recent results have shown that strict removal of exogenous patterning factors during early differentiation efficiently induces rostral hypothalamic progenitors from mouse ESCs. These hypothalamic progenitors generate vasopressinergic neurons, which release neuropeptides upon exogenous stimulation. Subsequently, we reported adenohypophysis tissue self-formation in three-dimensional cultures of mouse ESCs. The ESCs were found to differentiate into both non-neural oral ectoderm and hypothalamic neuroectoderm in adjacent layers. Interactions between the two tissues appear to be critically important for in vitro induction of a Rathke's pouch-like developing embryo. Various endocrine cells were differentiated from non-neural ectoderm. The induced corticotrophs efficiently secreted adrenocorticotropic hormone when engrafted in vivo, which rescued hypopituitary hosts. For future regenerative medicine, generation of hypothalamic and pituitary tissues from human PSCs is necessary. We and other groups succeeded in establishing a differentiation method with the use of human PSCs. Researchers could use these methods for models of human diseases to elucidate disease pathology or screen potential therapeutics.

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GENERATION OF RABBIT PLURIPOTENT STEM CELL LINES

Reproduction Fertility and Development ◽

10.1071/rdv24n1ab246 ◽

2012 ◽

Vol 24 (1) ◽

pp. 286

Author(s):

A. Dinnyes ◽

M. K. Pirity ◽

E. Gocza ◽

P. Osteil ◽

N. Daniel ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Pluripotent Stem Cells ◽

Pluripotent Stem Cell ◽

Pluripotent Cells ◽

Domestic Species ◽

Isolation And Characterization ◽

Induced Pluripotent ◽

Pluripotency Genes

Pluripotent stem cells have the capacity to divide indefinitely and to differentiate to all the somatic tissues. They can be genetically manipulated in vitro by knocking in and out genes, therefore they serve as an excellent tool for gene-function studies and for the generation of models for human diseases. Since 1981, when the first mouse embryonic stem cell (ESC) line was generated, several attempts have been made to generate pluripotent stem cells from other species as it would help us to understand the differences and similarities of signaling pathways involved in pluripotency and differentiation, and would reveal whether the fundamental mechanism controlling self-renewal of pluripotent cells is conserved among different species. This review gives an overlook of embryonic and induced pluripotent stem cell (iPSCs) research in the rabbit which is one of the most relevant non-rodent species for animal models. To date, several lines of putative ESCs and iPSCs have been described in the rabbit. All expressed stem cell-associated markers and exhibited longevity and pluripotency in vitro, but none have been proven to exhibit full pluripotency in vivo. Moreover, similarly to several domestic species, markers used to characterize the putative ESCs are not fully adequate because studies in domestic species have revealed that they are not specific to the pluripotent inner cell mass. Future validation of rabbit pluripotent stem cells would benefit greatly from a reliable panel of molecular markers specific to pluripotent cells of the developing rabbit embryo. The status of isolation and characterization of the putative pluripotency genes in rabbit will be discussed. Using rabbit specific pluripotency genes we might be able to reprogram somatic cells and generate induced pluripotent stem cells more efficiently thus overcome some of the challenges towards harnessing the potential of this technology. This study was financed by EU FP7 (PartnErS, PIAP-GA-2008-218205; InduHeart, PEOPLE-IRG-2008-234390; InduVir, PEOPLE-IRG-2009-245808; RabPstem, PERG07-GA-2010-268422; PluriSys, HEALTH-2007-B-223485; AniStem, PIAP-GA-2011-286264), NKTH-OTKA-EU-7KP HUMAN-MB08-C-80-205; Plurabbit, OMFB-00130-00131/2010 ANR-NKTH/09-GENM-010-01.

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Differentiating human pluripotent stem cells into vascular smooth muscle cells in three dimensional thermoreversible hydrogels

Biomaterials Science ◽

10.1039/c8bm01128a ◽

2019 ◽

Vol 7 (1) ◽

pp. 347-361 ◽

Cited By ~ 2

Author(s):

Haishuang Lin ◽

Qian Du ◽

Qiang Li ◽

Ou Wang ◽

Zhanqi Wang ◽

...

Keyword(s):

Smooth Muscle ◽

Vascular Smooth Muscle ◽

Smooth Muscle Cells ◽

Vascular Smooth Muscle Cells ◽

Pluripotent Stem Cells ◽

Pluripotent Stem Cell ◽

Three Dimensional ◽

Muscle Cells ◽

Human Pluripotent Stem Cell ◽

Scalable Production

3D thermoreversible PNIPAAm-PEG hydrogels are used for scalable production of human pluripotent stem cell-derived vascular smooth muscle cells.

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Modeling endodermal organ development and diseases using human pluripotent stem cell-derived organoids

Journal of Molecular Cell Biology ◽

10.1093/jmcb/mjaa031 ◽

2020 ◽

Vol 12 (8) ◽

pp. 580-592

Author(s):

Fong Cheng Pan ◽

Todd Evans ◽

Shuibing Chen

Keyword(s):

Pluripotent Stem Cells ◽

Pluripotent Stem Cell ◽

Disease Modeling ◽

State Of The Art ◽

Directed Differentiation ◽

Organ Development ◽

Human Pluripotent Stem Cell ◽

Human Organ ◽

Current State

Abstract Recent advances in development of protocols for directed differentiation from human pluripotent stem cells (hPSCs) to defined lineages, in combination with 3D organoid technology, have facilitated the generation of various endoderm-derived organoids for in vitro modeling of human gastrointestinal development and associated diseases. In this review, we discuss current state-of-the-art strategies for generating hPSC-derived endodermal organoids including stomach, liver, pancreatic, small intestine, and colonic organoids. We also review the advantages of using this system to model various human diseases and evaluate the shortcomings of this technology. Finally, we emphasize how other technologies, such as genome editing and bioengineering, can be incorporated into the 3D hPSC-organoid models to generate even more robust and powerful platforms for understanding human organ development and disease modeling.

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Towards consistent generation of pancreatic lineage progenitors from human pluripotent stem cells

Philosophical Transactions of the Royal Society B Biological Sciences ◽

10.1098/rstb.2014.0365 ◽

2015 ◽

Vol 370 (1680) ◽

pp. 20140365 ◽

Cited By ~ 19

Author(s):

Maria Rostovskaya ◽

Nicholas Bredenkamp ◽

Austin Smith

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Cell Lines ◽

Pluripotent Stem Cells ◽

Pluripotent Stem Cell ◽

Human Pluripotent Stem Cells ◽

Type I ◽

Pancreatic Progenitors ◽

Stem Cell Lines

Human pluripotent stem cells can in principle be used as a source of any differentiated cell type for disease modelling, drug screening, toxicology testing or cell replacement therapy. Type I diabetes is considered a major target for stem cell applications due to the shortage of primary human beta cells. Several protocols have been reported for generating pancreatic progenitors by in vitro differentiation of human pluripotent stem cells. Here we first assessed one of these protocols on a panel of pluripotent stem cell lines for capacity to engender glucose sensitive insulin-producing cells after engraftment in immunocompromised mice. We observed variable outcomes with only one cell line showing a low level of glucose response. We, therefore, undertook a systematic comparison of different methods for inducing definitive endoderm and subsequently pancreatic differentiation. Of several protocols tested, we identified a combined approach that robustly generated pancreatic progenitors in vitro from both embryo-derived and induced pluripotent stem cells. These findings suggest that, although there are intrinsic differences in lineage specification propensity between pluripotent stem cell lines, optimal differentiation procedures may consistently direct a substantial fraction of cells into pancreatic specification.

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Intestinal Organoids—Current and Future Applications

10.20944/preprints201608.0090.v1 ◽

2016 ◽

Author(s):

Andre M. C. Meneses ◽

Kerstin Schneeberger ◽

Hedwig S. Kruitwagen ◽

Louis C. Penning ◽

Frank G. van Steenbeek ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Pluripotent Stem Cells ◽

Disease Modeling ◽

Three Dimensional ◽

Intestinal Stem Cells ◽

Complex Structures ◽

Technical Advances ◽

Induced Pluripotent

Recent technical advances in the stem cell field have enabled the in vitro generation of complex structures resembling whole organs termed organoids. Most of these approaches employ culture systems that allow stem cell-derived or tissue progenitor cells to self-organize into three-dimensional (3D)-structures. Since organoids can be grown from various species, organs and from patient-derived induced pluripotent stem cells, they create significant prospects for modelling development and diseases, for toxicology and drug discovery studies, and in the field of regenerative medicine. Here, we report on intestinal stem cells, organoid culture, organoid disease modeling, transplantation, current and future uses of this exciting new insight model to veterinary medicine field.

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Reconstructing human early embryogenesis in vitro with pluripotent stem cells

10.1101/2021.03.12.435175 ◽

2021 ◽

Author(s):

Berna Sozen ◽

Victoria Jorgensen ◽

Meng Zhu ◽

Tongtong Cui ◽

Magdalena Zernicka-Goetz

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Human Development ◽

Pluripotent Stem Cells ◽

Molecular Mechanisms ◽

Early Embryo ◽

Human Stem Cells ◽

Amniotic Cavity ◽

Early Human

ABSTRACTUnderstanding human development is of fundamental biological and clinical importance. Yet despite its significance, insights into early developmental events in humans still remain largely unknown. While recent advances show that stem cells can mimic embryogenesis1–9 to unravel hidden developmental mechanisms, a stem cell-based model of early human embryogenesis is lacking. Here, we use human extended pluripotent stem cells10to reconstitute early human development in 3-dimensions and recapitulate early embryo-like events. We first perform a systematic characterisation to reveal unique signalling requirements for building the human pre-implantation blastocyst. Further, we show that these in vitro stem cell-derived blastocyst-like structures are able to undertake spatiotemporal self-organisation to mimic peri-implantation remodelling in which a polarised rosette opens up the amniotic cavity within a developing disc. The hallmarks of human early development displayed by this stem cell-based in vitro model mimics features of embryonic day 3 to day 9/10 of natural development. Thus, this platform represents a tractable model system to contribute to the basic understanding of cellular and molecular mechanisms governing early embryonic events in humans and to provide valuable insights into the design of differentiation protocols for human stem cells in clinical applications.

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C-STEM: ENGINEERING NICHE-LIKE MICRO-COMPARTMENTS FOR OPTIMAL AND SCALE-INDEPENDENT EXPANSION OF HUMAN PLURIPOTENT STEM CELLS IN BIOREACTORS

10.1101/2021.07.05.451086 ◽

2021 ◽

Author(s):

Philippe J.R. Cohen ◽

Elisa Luquet ◽

Justine Pletenka ◽

Andrea Leonard ◽

Elise Warter ◽

...

Keyword(s):

Stem Cells ◽

Pluripotent Stem Cells ◽

Large Scale ◽

Scale Up ◽

Three Dimensional ◽

Culture Conditions ◽

Human Pluripotent Stem Cells ◽

Cell Therapies ◽

Cellular Source

Human pluripotent stem cells (hPSCs) have emerged as the most promising cellular source for cell therapies. To overcome scale up limitations of classical 2D culture systems, suspension cultures have been developed to meet the need of large-scale culture in regenerative medicine. Despite constant improvements, current protocols relying on the generation of micro-carriers or cell aggregates only achieve moderate amplification performance. Here, guided by reports showing that hPSCs can self-organize in vitro into cysts reminiscent of the epiblast stage in embryo development, we developed a physio-mimetic approach for hPSC culture. We engineered stem cell niche microenvironments inside microfluidics-assisted core-shell microcapsules. We demonstrate that lumenized three-dimensional colonies maximize viability and expansion rates while maintaining pluripotency. By optimizing capsule size and culture conditions, we scale-up this method to industrial scale stirred tank bioreactors and achieve an unprecedented hPSC amplification rate of 282-fold in 6.5 days.

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Gastrointestinal organoids: a next-generation tool for modeling human development

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00199.2020 ◽

2020 ◽

Vol 319 (3) ◽

pp. G375-G381 ◽

Cited By ~ 1

Author(s):

Akaljot Singh ◽

Holly M. Poling ◽

Jason R. Spence ◽

James M. Wells ◽

Michael A. Helmrath

Keyword(s):

Stem Cells ◽

Human Development ◽

Pluripotent Stem Cells ◽

Three Dimensional ◽

Immunocompromised Host ◽

Dimensional Structure ◽

Gi Tract ◽

Microscopic Anatomy ◽

Complex Signaling

Gastrointestinal organoids are an exciting new tool for modeling human development, physiology, and disease in human tissue. Derived from pluripotent stem cells, gastrointestinal organoids consist of epithelial and mesenchymal cells organized in an intricate, three-dimensional structure that recapitulates the physiology and microscopic anatomy of the human gastrointestinal (GI) tract. In vitro derivation of gastrointestinal organoids from definitive endoderm has permitted an exploration of the complex signaling pathways required for the initial maturation of each individual gastrointestinal organ. Further maturation beyond an early fetal state currently requires transplantation into an immunocompromised host. Transplantation-induced maturation provides an opportunity to functionally interrogate the key mechanisms underlying development of the human GI tract. Gastrointestinal organoids can also be used to model human diseases and ultimately may serve as the basis for developing novel, personalized therapies for human intestinal diseases.

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