Differentiation of trophoblast cells from human embryonic stem cells: to be or not to be?

It is imperative to unveil the full range of differentiated cell types into which human pluripotent stem cells (hPSCs) can develop. The need is twofold: it will delimit the therapeutic utility of these stem cells and is necessary to place their position accurately in the developmental hierarchy of lineage potential. Accumulated evidence suggested that hPSC could develop in vitro into an extraembryonic lineage (trophoblast (TB)) that is typically inaccessible to pluripotent embryonic cells during embryogenesis. However, whether these differentiated cells are truly authentic TB has been challenged. In this debate, we present a case for and a case against TB differentiation from hPSCs. By analogy to other differentiation systems, our debate is broadly applicable, as it articulates higher and more challenging standards for judging whether a given cell type has been genuinely produced from hPSC differentiation.

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Defining totipotency using criteria of increasing stringency

10.1101/2020.03.02.972893 ◽

2020 ◽

Cited By ~ 7

Author(s):

Eszter Posfai ◽

John Paul Schell ◽

Adrian Janiszewski ◽

Isidora Rovic ◽

Alexander Murray ◽

...

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

Single Cell ◽

Pluripotent Stem Cells ◽

Embryonic Stem ◽

Cell Types ◽

Differentiated Cells ◽

Totipotent Cell

AbstractTotipotency is the ability of a single cell to give rise to all the differentiated cells that build the conceptus, yet how to capture this property in vitro remains incompletely understood. Defining totipotency relies upon a variety of assays of variable stringency. Here we describe criteria to define totipotency. We illustrate how distinct criteria of increasing stringency can be used to judge totipotency by evaluating candidate totipotent cell types in the mouse, including early blastomeres and expanded or extended pluripotent stem cells. Our data challenge the notion that expanded or extended pluripotent states harbor increased totipotent potential relative to conventional embryonic stem cells under in vivo conditions.

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Establishing a deeper understanding of the osteogenic differentiation of monolayer cultured human pluripotent stem cells using novel and detailed analyses

Stem Cell Research & Therapy ◽

10.1186/s13287-020-02085-9 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Ping Zhou ◽

Jia-Min Shi ◽

Jing-E Song ◽

Yu Han ◽

Hong-Jiao Li ◽

...

Keyword(s):

Stem Cells ◽

Osteogenic Differentiation ◽

Pluripotent Stem Cells ◽

Embryonic Stem ◽

Telomerase Activity ◽

Human Pluripotent Stem Cells ◽

Cell Counting ◽

Cell Type ◽

Alizarin Red

Abstract Background Derivation of osteoblast-like cells from human pluripotent stem cells (hPSCs) is a popular topic in bone tissue engineering. Although many improvements have been achieved, the low induction efficiency because of spontaneous differentiation hampers their applications. To solve this problem, a detailed understanding of the osteogenic differentiation process of hPSCs is urgently needed. Methods Monolayer cultured human embryonic stem cells and human-induced pluripotent stem cells were differentiated in commonly applied serum-containing osteogenic medium for 35 days. In addition to traditional assays such as cell viability detection, reverse transcription-polymerase chain reaction, immunofluorescence, and alizarin red staining, we also applied studies of cell counting, cell telomerase activity, and flow cytometry as essential indicators to analyse the cell type changes in each week. Results The population of differentiated cells was quite heterogeneous throughout the 35 days of induction. Then, cell telomerase activity and cell cycle analyses have value in evaluating the cell type and tumourigenicity of the obtained cells. Finally, a dynamic map was made to integrate the analysis of these results during osteogenic differentiation of hPSCs, and the cell types at defined stages were concluded. Conclusions Our results lay the foundation to improve the in vitro osteogenic differentiation efficiency of hPSCs by supplementing with functional compounds at the desired stage, and then establishing a stepwise induction system in the future.

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Pancreatic Progenitors and Organoids as a Prerequisite to Model Pancreatic Diseases and Cancer

Stem Cells International ◽

10.1155/2019/9301382 ◽

2019 ◽

Vol 2019 ◽

pp. 1-11 ◽

Cited By ~ 3

Author(s):

Meike Hohwieler ◽

Martin Müller ◽

Pierre-Olivier Frappart ◽

Sandra Heller

Keyword(s):

Stem Cells ◽

Pluripotent Stem Cells ◽

Disease Modeling ◽

Embryonic Stem ◽

Cell Types ◽

Genetically Engineered ◽

Hereditary Diseases ◽

Pancreatic Diseases ◽

Pancreatic Progenitors

Embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) are characterized by their unique capacity to stepwise differentiate towards any particular cell type in an adult organism. Pluripotent stem cells provide a beneficial platform to model hereditary diseases and even cancer development. While the incidence of pancreatic diseases such as diabetes and pancreatitis is increasing, the understanding of the underlying pathogenesis of particular diseases remains limited. Only a few recent publications have contributed to the characterization of human pancreatic development in the fetal stage. Hence, most knowledge of pancreatic specification is based on murine embryology. Optimizing and understanding current in vitro protocols for pancreatic differentiation of ESCs and iPSCs constitutes a prerequisite to generate functional pancreatic cells for better disease modeling and drug discovery. Moreover, human pancreatic organoids derived from pluripotent stem cells, organ-restricted stem cells, and tumor samples provide a powerful technology to model carcinogenesis and hereditary diseases independent of genetically engineered mouse models. Herein, we summarize recent advances in directed differentiation of pancreatic organoids comprising endocrine cell types. Beyond that, we illustrate up-and-coming applications for organoid-based platforms.

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Generation of Sheep Induced Pluripotent Stem Cells With Defined DOX-Inducible Transcription Factors via piggyBac Transposition

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.785055 ◽

2021 ◽

Vol 9 ◽

Author(s):

Moning Liu ◽

Lixia Zhao ◽

Zixin Wang ◽

Hong Su ◽

Tong Wang ◽

...

Keyword(s):

Stem Cells ◽

Induced Pluripotent Stem Cells ◽

Pluripotent Stem Cells ◽

Embryonic Stem ◽

Cell Types ◽

T Antigen ◽

Sv40 Large T Antigen ◽

Adult Individual ◽

Induced Pluripotent

Pluripotent stem cells (PSCs) have the potential to differentiate to all cell types of an adult individual and are useful for studying mammalian development. Establishing induced pluripotent stem cells (iPSCs) capable of expressing pluripotent genes and differentiating to three germ layers will not only help to explain the mechanisms underlying somatic reprogramming but also lay the foundation for the establishment of sheep embryonic stem cells (ESCs) in vitro. In this study, sheep somatic cells were reprogrammed in vitro into sheep iPSCs with stable morphology, pluripotent marker expression, and differentiation ability, delivered by piggyBac transposon system with eight doxycycline (DOX)-inducible exogenous reprogramming factors: bovine OCT4, SOX2, KLF4, cMYC, porcine NANOG, human LIN28, SV40 large T antigen, and human TERT. Sheep iPSCs exhibited a chimeric contribution to the early blastocysts of sheep and mice and E6.5 mouse embryos in vitro. A transcriptome analysis revealed the pluripotent characteristics of somatic reprogramming and insights into sheep iPSCs. This study provides an ideal experimental material for further study of the construction of totipotent ESCs in sheep.

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Generation of defined neural populations from pluripotent stem cells

Philosophical Transactions of the Royal Society B Biological Sciences ◽

10.1098/rstb.2017.0214 ◽

2018 ◽

Vol 373 (1750) ◽

pp. 20170214 ◽

Cited By ~ 7

Author(s):

Sarah F. McComish ◽

Maeve A. Caldwell

Keyword(s):

Stem Cells ◽

Pluripotent Stem Cells ◽

Embryonic Stem ◽

Cell Types ◽

Disease Modelling ◽

Human Neural Stem Cells ◽

Neural Populations ◽

Human Disorders ◽

Induced Pluripotent

Effective and efficient generation of human neural stem cells and subsequently functional neural populations from pluripotent stem cells has facilitated advancements in the study of human development and disease modelling. This review will discuss the established protocols for the generation of defined neural populations including regionalized neurons and astrocytes, oligodendrocytes and microglia. Early protocols were established in embryonic stem cells (ESC) but the discovery of induced pluripotent stem cells (iPSC) in 2006 provided a new platform for modelling human disorders of the central nervous system (CNS). The ability to produce patient- and disease-specific iPSC lines has created a new age of disease modelling. Human iPSC may be derived from adult somatic cells and subsequently patterned into numerous distinct cell types. The ability to derive defined and regionalized neural populations from iPSC provides a powerful in vitro model of CNS disorders. This article is part of the theme issue ‘Designer human tissue: coming to a lab near you’.

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Human Pluripotent Stem Cell-Derived Cardiomyocytes as Research and Therapeutic Tools

BioMed Research International ◽

10.1155/2014/512831 ◽

2014 ◽

Vol 2014 ◽

pp. 1-14 ◽

Cited By ~ 32

Author(s):

Ivana Acimovic ◽

Aleksandra Vilotic ◽

Martin Pesl ◽

Alain Lacampagne ◽

Petr Dvorak ◽

...

Keyword(s):

Stem Cells ◽

Pluripotent Stem Cells ◽

Drug Testing ◽

Embryonic Stem ◽

Cell Types ◽

Patient Specific ◽

Disease Modelling ◽

Therapeutic Tools ◽

Induced Pluripotent

Human pluripotent stem cells (hPSCs), namely, embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), with their ability of indefinite self-renewal and capability to differentiate into cell types derivatives of all three germ layers, represent a powerful research tool in developmental biology, for drug screening, disease modelling, and potentially cell replacement therapy. Efficient differentiation protocols that would result in the cell type of our interest are needed for maximal exploitation of these cells. In the present work, we aim at focusing on the protocols for differentiation of hPSCs into functional cardiomyocytesin vitroas well as achievements in the heart disease modelling and drug testing on the patient-specific iPSC-derived cardiomyocytes (iPSC-CMs).

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Establishing a deeper understanding of the osteogenic differentiation of monolayer cultured human pluripotent stem cells using novel and detailed analyses

10.21203/rs.3.rs-45401/v3 ◽

2020 ◽

Author(s):

Ping Zhou ◽

Jia-Min Shi ◽

Jing-E Song ◽

Yu Han ◽

Hong-Jiao Li ◽

...

Keyword(s):

Stem Cells ◽

Osteogenic Differentiation ◽

Pluripotent Stem Cells ◽

Embryonic Stem ◽

Telomerase Activity ◽

Human Pluripotent Stem Cells ◽

Cell Counting ◽

Cell Type ◽

Alizarin Red

Abstract Background: Derivation of osteoblast-like cells from human pluripotent stem cells (hPSCs) is a popular topic in bone tissue engineering. Although many improvements have been achieved, the low induction efficiency because of spontaneous differentiation hampers their applications. To solve this problem, a detailed understanding of the osteogenic differentiation process of hPSCs is urgently needed.Methods: Monolayer cultured human embryonic stem cells and human induced pluripotent stem cells were differentiated in commonly applied serum-containing osteogenic medium for 35 days. In addition to traditional assays such as cell viability detection, reverse transcription-polymerase chain reaction, immunofluorescence, and alizarin red staining, we also applied studies of cell counting, cell telomerase activity, and flow cytometry as essential indicators to analyse the cell type changes in each week.Results: The population of differentiated cells was quite heterogeneous throughout the 35 days of induction. Then, cell telomerase activity and cell cycle analyses have value in evaluating the cell type and tumourigenicity of the obtained cells. Finally, a dynamic map was made to integrated the analysis of these results during osteogenic differentiation of hPSCs, and the cell types at defined stages were concluded.Conclusions: Our results lay the foundation to improve the in vitro osteogenic differentiation efficiency of hPSCs by supplementing with functional compounds at the desired stage, and then establishing a step-wise induction system in the future.

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Prunellae Spica Extract Suppresses Teratoma Formation of Pluripotent Stem Cells through p53-Mediated Apoptosis

Nutrients ◽

10.3390/nu12030721 ◽

2020 ◽

Vol 12 (3) ◽

pp. 721 ◽

Cited By ~ 2

Author(s):

Aeyung Kim ◽

Seo-Young Lee ◽

Chang-Seob Seo ◽

Sun-Ku Chung

Keyword(s):

Stem Cells ◽

Pluripotent Stem Cells ◽

Apoptotic Cell ◽

Embryonic Stem ◽

M Cell ◽

In Ovo ◽

Differentiated Cells ◽

Teratoma Formation

Induced pluripotent stem cells (iPSCs) have similar properties to embryonic stem cells in terms of indefinite self-renewal and differentiation capacity. After in vitro differentiation of iPSCs, undifferentiated iPSCs (USCs) may exist in cell therapy material and can form teratomas after in vivo transplantation. Selective elimination of residual USCs is, therefore, very important. Prunellae Spica (PS) is a traditional medicinal plant that has been shown to exert anti-cancer, antioxidant, and anti-inflammatory activities; however, its effects on iPSCs have not been previously characterized. In this study, we find that ethanol extract of PS (EPS) effectively induces apoptotic cell death of USCs through G2/M cell cycle arrest, generation of intracellular reactive oxygen species, alteration of mitochondrial membrane potentials, and caspase activation of USCs. In addition, EPS increases p53 accumulation and expression of its downstream targets. In p53 knockout (KO) iPSCs, the EPS did not induce apoptosis, indicating that EPS-mediated apoptosis of USCs was p53-dependent. In addition, EPS was not genotoxic towards iPSCs-derived differentiated cells. EPS treatment before injection efficiently prevented in ovo teratoma formation of p53 wild-type (WT) iPSCs but not p53KO iPSCs. Collectively, these results indicate that EPS has potent anti-teratoma activity and no genotoxicity to differentiated cells. It can, therefore, be used in the development of safe and efficient iPSC-based cell therapies.

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A Web-Server of Cell Type Discrimination System

The Scientific World JOURNAL ◽

10.1155/2014/459064 ◽

2014 ◽

Vol 2014 ◽

pp. 1-5 ◽

Cited By ~ 1

Author(s):

Anyou Wang ◽

Yan Zhong ◽

Yanhua Wang ◽

Qianchuan He

Keyword(s):

Stem Cells ◽

Pluripotent Stem Cells ◽

Embryonic Stem ◽

Web Server ◽

Cell Types ◽

Cell Type ◽

User Input ◽

The Common ◽

Induced Pluripotent ◽

User Friendly

Discriminating cell types is a daily request for stem cell biologists. However, there is not a user-friendly system available to date for public users to discriminate the common cell types, embryonic stem cells (ESCs), induced pluripotent stem cells (iPSCs), and somatic cells (SCs). Here, we develop WCTDS, a web-server of cell type discrimination system, to discriminate the three cell types and their subtypes like fetal versus adult SCs. WCTDS is developed as a top layer application of our recent publication regarding cell type discriminations, which employs DNA-methylation as biomarkers and machine learning models to discriminate cell types. Implemented by Django, Python, R, and Linux shell programming, run under Linux-Apache web server, and communicated through MySQL, WCTDS provides a friendly framework to efficiently receive the user input and to run mathematical models for analyzing data and then to present results to users. This framework is flexible and easy to be expended for other applications. Therefore, WCTDS works as a user-friendly framework to discriminate cell types and subtypes and it can also be expended to detect other cell types like cancer cells.

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Cell type determination for cardiac differentiation occurs soon after seeding of human induced pluripotent stem cells

10.1101/2021.08.08.455532 ◽

2021 ◽

Author(s):

Connie L Jiang ◽

Yogesh Goyal ◽

Naveen Jain ◽

Qiaohong Wang ◽

Rachel E Truitt ◽

...

Keyword(s):

Stem Cells ◽

Induced Pluripotent Stem Cells ◽

Pluripotent Stem Cells ◽

Cell Types ◽

Cardiac Differentiation ◽

Marker Genes ◽

Directed Differentiation ◽

Cell Type ◽

Differentiated Cells ◽

Induced Pluripotent

Cardiac directed differentiation of human induced pluripotent stem cells consistently produces a mixed population of cardiomyocytes and non-cardiac cell types even when using very well-characterized protocols. We wondered whether differentiated cell types might result from intrinsic differences in hiPS cells prior to the onset of differentiation. By associating individual differentiated cells that share a common hiPS cell precursor, we were able to test whether expression variability in differentiated cells was pre-determined from the hiPS cell state. Although within a single experiment, differentiated cells that share an hiPS cell progenitor were more transcriptionally similar to each other than to other cells in the differentiated population, when the same hiPS cells were differentiated in parallel, we did not observe high transcriptional similarity across differentiations. Additionally, we found that substantial cell death occurred during differentiation in a manner that suggested that all cells were equally likely to survive or die, suggesting that there was no intrinsic selection bias for cells descended from particular hiPS cell progenitors. These results led us to wonder about how cells grow out spatially during the directed differentiation process. Labeling cells by their expression of a few canonical cell type marker genes, we showed that cells expressing the same marker tended to occur in patches observable by visual inspection, suggesting that cell type determination across multiple cell types, once initiated, is maintained in a cell-autonomous manner for multiple divisions. Altogether, our results show that while there is substantial heterogeneity in the initial hiPS cell population, that heterogeneity is not responsible for heterogeneous outcomes, and that the window during which cell type specification occurs is likely to begin shortly after the seeding of hiPS cells for differentiation.

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