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2015, Nature
Stem cells self-renew and generate specialized progeny through differentiation, but vary in the range of cells and tissues they generate, a property called developmental potency. Pluripotent stem cells produce all cells of an organism, while multipotent or unipotent stem cells regenerate only specific lineages or tissues. Defining stem-cell potency relies upon functional assays and diagnostic transcriptional, epigenetic and metabolic states. Here we describe functional and molecular hallmarks of pluripotent stem cells, propose a checklist for their evaluation, and illustrate how forensic genomics can validate their provenance.
Stem Cell Research, 2013
Much of the excitement generated by induced pluripotent stem cell technology is concerned with the possibility of disease modeling as well as the potential for personalized cell therapy. However, to pursue this it is important to understand the 'normal' pluripotent state including its inherent variability. We have performed various molecular profiling assays for 21 hESC lines and 8 hiPSC lines to generate a comprehensive snapshot of the undifferentiated state of pluripotent stem cells. Analysis of the gene expression data revealed no iPSC-specific gene expression pattern in accordance with previous reports. We further compared cells, differentiated as embryoid bodies in 2 media proposed to initiate differentiation towards separate cell fates, as well as 20 adult tissues. From this analysis we have generated a gene list which defines pluripotency and establishes a baseline for the pluripotent state. Finally, we provide lists of genes enriched under both differentiation conditions which show the proposed bias toward independent cell fates.
Nature reviews. Molecular cell biology, 2016
Basic experimental stem cell research has opened up the possibility of many diverse clinical applications; however, translation to clinical trials has been restricted to only a few diseases. To broaden this clinical scope, pluripotent stem cell derivatives provide a uniquely scalable source of functional differentiated cells that can potentially repair damaged or diseased tissues to treat a wide spectrum of diseases and injuries. However, gathering sound data on their distribution, longevity, function and mechanisms of action in host tissues is imperative to realizing their clinical benefit. The large-scale availability of treatments involving pluripotent stem cells remains some years away, because of the long and demanding regulatory pathway that is needed to ensure their safety.
Nature Communications, 2018
Assessment of established techniques to determine developmental and malignant potential of human pluripotent stem cells The International Stem Cell Initiative # The International Stem Cell Initiative compared several commonly used approaches to assess human pluripotent stem cells (PSC). PluriTest predicts pluripotency through bioinformatic analysis of the transcriptomes of undifferentiated cells, whereas, embryoid body (EB) formation in vitro and teratoma formation in vivo provide direct tests of differentiation. Here we report that EB assays, analyzed after differentiation under neutral conditions and under conditions promoting differentiation to ectoderm, mesoderm, or endoderm lineages, are sufficient to assess the differentiation potential of PSCs. However, teratoma analysis by histologic examination and by TeratoScore, which estimates differential gene expression in each tumor, not only measures differentiation but also allows insight into a PSC's malignant potential. Each of the assays can be used to predict pluripotent differentiation potential but, at this stage of assay development, only the teratoma assay provides an assessment of pluripotency and malignant potential, which are both relevant to the pre-clinical safety assessment of PSCs.
Stem cell reports, 2016
The rigorous characterization of distinct induced pluripotent stem cells (iPSC) derived from multiple reprogramming technologies, somatic sources, and donors is required to understand potential sources of variability and downstream potential. To achieve this goal, the Progenitor Cell Biology Consortium performed comprehensive experimental and genomic analyses of 58 iPSC from ten laboratories generated using a variety of reprogramming genes, vectors, and cells. Associated global molecular characterization studies identified functionally informative correlations in gene expression, DNA methylation, and/or copy-number variation among key developmental and oncogenic regulators as a result of donor, sex, line stability, reprogramming technology, and cell of origin. Furthermore, X-chromosome inactivation in PSC produced highly correlated differences in teratoma-lineage staining and regulator expression upon differentiation. All experimental results, and raw, processed, and metadata from t...
Stem cells (Dayton, Ohio), 2011
Pluripotent stem cells (PSC) are functionally characterized by their capacity to differentiate into all the cell types from the three germ layers. A wide range of markers, the expression of which is associated with pluripotency, has been used as surrogate evidence of PSC pluripotency, but their respective relevance is poorly documented. Here, we compared by polychromatic flow cytometry the kinetics of loss of expression of eight widely used pluripotency markers (SSEA3, SSEA4, TRA-1-60, TRA-1-81, CD24, OCT4, NANOG, and alkaline phosphatase [AP]) at days 0, 5, 7, and 9 after induction of PSC differentiation into cells representative of the three germ layers. Strikingly, each marker showed a different and specific kinetics of disappearance that was similar in all the PSC lines used and for all the induced differentiation pathways. OCT4, SSEA3, and TRA-1-60 were repeatedly the first markers to be downregulated, and their expression was completely lost at day 9. By contrast, AP activity,...
Cell reports, 2017
Human pluripotent stem cells (hPSCs) have been reported in naive and primed states. However, the ability to generate mature cell types remains the imperative property for utility of hPSCs. Here, we reveal that the naive state enhances self-renewal while restricting lineage differentiation in vitro to neural default fate. Molecular analyses indicate expression of multiple lineage-associated transcripts in naive hPSCs that failed to predict biased functional differentiation capacity. Naive hPSCs can be converted to primed state over long-term serial passage that permits recovery of multi-germ layer differentiation. Suppression of OCT4 but not NANOG allows immediate recovery directly from naive state. To this end, we identified chemical inhibitors of OCT4 that restore naive hPSC differentiation. Our study reveals unique cell-fate restrictions in human pluripotent states and provides an approach to overcome these barriers that harness both efficient naive hPSC growth while maintaining i...
Insight into the key genes of pluripotency in human and their interrelationships is necessary for understanding the underlying mechanism of pluripotency and hence their successful application in regenerative medicine. The recent advances in transcriptomics technologies have created new opportunities to decipher the genes involved in pluripotency, genetic network that governs the unique properties of embryonic stem cells and lineage differentiation mechanisms in a deeper scale. There are a large number of experimental studies on human embryonic stem cells (hESCs) being routinely conducted for unfolding the underlying biology of embryogenesis and their clinical prospects. However, the outcome of these studies often lacks consensus due to differences in samples, experimental techniques and/or analysis protocols. A universal stemness gene list is still lacking. In this quest, we compared transcriptomic profiles of pluripotent and non-pluripotent samples from diverse cell lines/types gen...
2021
Human induced pluripotent stem cells (iPSCs) are a powerful tool for studying development and disease. However, different iPSC lines show considerable phenotypic variation. The lack of common well-characterised cell lines that are used widely frustrates efforts to integrate data across research groups or replicate key findings. Inspired by model organism communities who addressed this issue by establishing a limited number of widely accepted strains, we characterised candidate iPSC lines in unprecedented detail to select a well-performing line to underpin collaborative studies. Specifically, we characterised the morphology, growth rates, and single-cell transcriptomes of iPSC lines in the pluripotent state and assessed their genomic integrity using karyotyping, DNA microarrays, whole genome sequencing, and functional assays for p53 activity. We further tested their ability to be edited by CRISPR/Cas9 and used single-cell RNA sequencing to compare the efficiency with which they could...
BioEssays, 2009
The phenomenal proliferation of scientific studies into the nature o nduced pluripotent stem (iPS) cells following publication of the findings of Takahashi and Yamanaka little more than 2 years ago, have significantly expanded our understanding of cellular mechanisms relating to cell lineage, di erentiation, and proliferation. While the full potential o PS cell lineages for both scientific tool and therapeutic applications is as yet unclear, findings from several lines o nvestigation suggests that multipotential and terminally di erentiated cells from an array of cell types are competent to undergo epigenetic reprogramming to a pluripotential state. The nature of this pluripotential state appears to be similar to, but not identical with that previously described for embryonic stem (ES) cells. Understanding the nature of this induced reprogrammed state will be critical to determining the full potential o PS cells. Recently, this issue has been examined through an integrated analysis of the genome in fully and partially reprogrammed iPS cell lineages. These results provide a window onto the temporal components of reprogramming and suggest mechanisms by which the efficacy of reprogramming can be enhanced.
Nature Methods, 2011
Annual review of genomics and human genetics, 2011
Embryonic stem cells (ESCs) first derived from the inner cell mass of blastocyst-stage embryos have the unique capacity of indefinite self-renewal and potential to differentiate into all somatic cell types. Similar developmental potency can be achieved by reprogramming differentiated somatic cells into induced pluripotent stem cells (iPSCs). Both types of pluripotent stem cells provide great potential for fundamental studies of tissue differentiation, and hold promise for disease modeling, drug development, and regenerative medicine. Although much has been learned about the molecular mechanisms that underlie pluripotency in such cells, our understanding remains incomplete. A comprehensive understanding of ESCs and iPSCs requires the deconstruction of complex transcription regulatory networks, epigenetic mechanisms, and biochemical interactions critical for the maintenance of self-renewal and pluripotency. In this review, we will discuss recent advances gleaned from application of gl...
2010
Human pluripotent stem cells (PSCs), encompassing embryonic stem cells and induced pluripotent stem cells, proliferate extensively and differentiate into virtually any desired cell type. PSCs endow regenerative medicine with an unlimited source of replacement cells suitable for human therapy. Several hurdles must be carefully addressed in PSC research before these theoretical possi-bilities are translated into clinical applications. These obstacles are: (1) cell proliferation; (2) cell differentiation;
Cell Stem Cell, 2009
Halo 194, 2020
Induced Pluripotent Stem Cells (iPSCs) are a type of pluripotent stem cells generated by reprogramming an adult somatic cell genome to the stage of a pluripotent stem cell in vitro by inducing a forced expression of specific transcription factors that are important for the maintenance of pluripotency. The iPSCs seem to be very similar to Embryonic Stem Cells (ESCs) in terms of morphology, cell surface markers and gene expression levels, but recent studies have demonstrated some differences between the two cell types. However, iPSCs might have potential application in regenerative medicine, transplantation, drug testing, disease modelling, and avoidance of tissue rejection and with less ethical concern than ESCs. This paper aims to present the most important characteristics of iPSCs which have therapeutic significance.
Journal of Population Therapeutics & Clinical Pharmacology, 2024
Human embryology holds immense promise for medical science, offering insights into the initial establishment of human growth and the potential for regenerative medicine. This paper delves into the captivating realm of self-renewing pluripotent stem cells, which lie at the heart of unlocking the mysteries of human embryology. Pluripotent stem cells hold the outstanding capacity to form into any cell type inside the body, presenting unprecedented opportunities for understanding development, modeling diseases, and facilitating therapeutic interventions. With recent advancements in cellular reprogramming techniques and genome editing technologies, researchers have made significant strides in connecting the potential of pluripotent stem cells for various bids. This paper aims to explore the journey of pluripotent stem cells, from their discovery to current state-of-the-art methodologies, while highlighting the challenges and ethical considerations inherent in their utilization. By elucidating the mechanisms governing pluripotency and differentiation, this research seeks to provide valuable insights into human embryology and pave the way for innovative approaches in regenerative medicine and disease modeling.
PLoS ONE, 2009
Background: Commitment in embryonic stem cells is often depicted as a binary choice between alternate cell states, pluripotency and specification to a particular germ layer or extraembryonic lineage. However, close examination of human ES cell cultures has revealed significant heterogeneity in the stem cell compartment.
Stem Cells, 2010
Pluripotent stem cells (PSCs) have been derived from the embryos of mice and humans, representing the two major sources of PSCs. These cells are universally defined by their developmental properties, specifically their self-renewal capacity and differentiation potential which are regulated in mice and humans by complex transcriptional networks orchestrated by conserved transcription factors. However, significant differences exist in the transcriptional networks and signaling pathways that control mouse and human PSC self-renewal and lineage development. To distinguish between universally applicable and species-specific features, we collated and compared the molecular and cellular descriptions of mouse and human PSCs. Here we compare and contrast the response to signals dictated by the transcriptome and epigenome of mouse and human PSCs that will hopefully act as a critical resource to the field. These analyses underscore the importance of accounting for species differences when desi...
Stem Cell Research, 2021
Pluripotent stem cells (PSCs) have been observed to occur in two distinct statesnaive and primed. Both naive and primed state PSCs can give rise to tissues of all the three germ layers in vitro but differ in their potential to generate germline chimera in vivo. Understanding the molecular mechanisms that govern these two states of pluripotency in human can open a plethora of opportunities for studying early embryonic development and in biomedical applications. In this work, we use weighted gene co-expression network analysis (WGCNA) to identify the key molecular makers and their interactions that define the two distinct pluripotency states. Signed hybrid network was reconstructed from transcriptomic data (RNA-seq) of naive and primed state pluripotent samples. Our analysis revealed two sets of genes that are involved in the establishment and maintenance of naive and primed states. The naive state genes were found to be enriched for biological processes and pathways related to metabolic processes while primed state genes were associated with system development. We further filtered these lists to identify the intra-modular hubs and the hub transcription factors (TFs) for each group. Validation of the identified TFs was carried out using independent microarray datasets and we finally present a list of 52 and 33 TFs as the set of core TFs that are responsible for the induction and maintenance of naive and primed states of pluripotency in human, respectively. Among these, the TFs ZNF275, ZNF232, SP4, and MSANTD3 could be of interest as they were not reported in previous studies.
World Journal of Stem Cells, 2019
Ten years after the initial generation of induced pluripotent stem cells (hiPSCs) from human tissues, their potential is no longer questioned, with over 15000 publications listed on PubMed, covering various fields of research; including disease modeling, cell therapy strategies, pharmacology/toxicology screening and 3D organoid systems. However, despite evidences that the presence of mutations in hiPSCs should be a concern, publications addressing genomic integrity of these cells represent less than 1% of the literature. After a first overview of the mutation types currently reported in hiPSCs, including karyotype abnormalities, copy number variations, single point mutation as well as uniparental disomy, this review will discuss the impact of reprogramming parameters such as starting cell type and reprogramming method on the maintenance of the cellular genomic integrity. Then, a specific focus will be placed on culture conditions and subsequent differentiation protocols and how
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