scholarly journals Context-dependent requirement of H3K9 methyltransferase activity during cellular reprogramming to iPSCs

2019 ◽  
Author(s):  
Simon Vidal ◽  
Alexander Polyzos ◽  
Jorge Morales Valencia ◽  
Hongsu Wang ◽  
Emily Swanzey ◽  
...  
Keyword(s):  
Pluripotent Stem Cells ◽  
Cellular Reprogramming ◽  
Methyltransferase Activity ◽  
Molecular Defects ◽  
Functional Interactions ◽  
Histone 3 ◽  
Epigenetic Remodeling ◽  
Genetic Interference ◽  
Induced Pluripotent ◽  
Epithelial State

SUMMARYMethylation of histone 3 at lysine 9 (H3K9) is widely regarded as a major roadblock for cellular reprogramming and interference with associated methyltransferases such as EHMT1 and EHMT2 (also known as GLP and G9A, respectively) increases the efficiencies at which induced pluripotent stem cells (iPSCs) can be derived. Activation of histone and DNA demethylases by ascorbic acid (AA) has become a common approach to facilitate the extensive epigenetic remodeling required for iPSC formation, but possible functional interactions between the H3K9 methylation machinery and AA-stimulated enzymes remain insufficiently explored. Here we show that reduction of EHMT1/2 activity counteracts iPSC formation in an optimized reprogramming system in the presence of AA. Mechanistically, EHMT1/2 activity under these conditions is required for efficient downregulation of somatic genes and transition into an epithelial state. Of note, transient inhibition of EHMT1/2 during reprogramming yields iPSCs that fail to efficiently give rise to viable mice, suggesting persistent molecular defects in these cells. Genetic interference with the H3K9 demethylase KDM3B ameliorated the adverse effect of EHMT1/2 inhibition on iPSC formation. Together, our observations document novel functions of H3K9 methyltransferases during iPSC formation and suggest that the balancing of AA-stimulated enzymes by EHMT1/2 supports efficient and error-free iPSC reprogramming to pluripotency.

scholarly journals Culture Environment-Induced Pluripotency of SACK-Expanded Tissue Stem Cells

2011 ◽  
Vol 2011 ◽  
pp. 1-12 ◽  
Author(s):  
Jean-François Paré ◽  
James L. Sherley
Keyword(s):  
Stem Cells ◽  
Transcription Factors ◽  
Pluripotent Stem Cells ◽  
Adult Mouse ◽  
Pancreatic Tissue ◽  
Cellular Reprogramming ◽  
Induced Pluripotency ◽  
Pancreatic Stem Cells ◽  
Induced Pluripotent ◽  
Slow Growing

Previous efforts to improve the efficiency of cellular reprogramming for the generation of induced pluripotent stem cells (iPSCs) have focused mainly on transcription factors and small molecule combinations. Here, we report the results of our focus instead on the phenotype of the cells targeted for reprogramming. We find that adult mouse pancreatic tissue stem cells derived by the method of suppression of asymmetric cell kinetics (SACK) acquire increased potency simply by culture under conditions for the production and maintenance of pluripotent stem cells. Moreover, supplementation with the SACK agent xanthine, which promotes symmetric self-renewal, significantly increases the efficiency and degree of acquisition of pluripotency properties. In transplantation analyses, clonal reprogrammed pancreatic stem cells produce slow-growing tumors with tissue derivative of all three embryonic germ layers. This acquisition of pluripotency, without transduction with exogenous transcription factors, supports the concept that tissue stem cells are predisposed to cellular reprogramming, particularly when symmetrically self-renewing.

Regenerative Medicine/Cardiac Cell Therapy: Pluripotent Stem Cells

2017 ◽  
Vol 66 (01) ◽  
pp. 053-062 ◽  
Author(s):  
Ana Duran ◽  
Olivia Reidell ◽  
Harald Stachelscheid ◽  
Kristin Klose ◽  
Manfred Gossen ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Therapy ◽  
Pluripotent Stem Cells ◽  
Cardiac Tissue ◽  
Cellular Reprogramming ◽  
Patient Specific ◽  
Cellular Cardiomyoplasty ◽  
Somatic Cell Therapy ◽  
Cardiac Cell Therapy ◽  
Induced Pluripotent

AbstractFor more than 20 years, tremendous efforts have been made to develop cell-based therapies for treatment of heart failure. However, the results of clinical trials using somatic, nonpluripotent stem or progenitor cells have been largely disappointing in both cardiology and cardiac surgery scenarios. Surgical groups were among the pioneers of experimental and clinical myocyte transplantation (“cellular cardiomyoplasty”), but little translational progress was made prior to the development of cellular reprogramming for creation of induced pluripotent stem cells (iPSC). Ever since, protocols have been developed which allow for the derivation of large numbers of autologous cardiomyocytes (CMs) from patient-specific iPSC, moving translational research closer toward clinical pilot trials. However, compared with somatic cell therapy, the technology required for safe and efficacious pluripotent stem cell (PSC)-based therapies is extremely complex and requires tremendous resources and close interactions between basic scientists and clinicians. This review summarizes PSC sources, strategies to derive CMs, current cardiac tissue engineering approaches, concerns regarding immunogenicity and cellular maturity, and highlights the contributions made by surgical groups.

scholarly journals Pluripotent Cell Models for Gonadal Research

2019 ◽  
Vol 20 (21) ◽  
pp. 5495 ◽  
Author(s):  
Daniel Rodríguez Gutiérrez ◽  
Anna Biason-Lauber
Keyword(s):  
Pluripotent Stem Cells ◽  
Cellular Reprogramming ◽  
Patient Specific ◽  
Cell Models ◽  
In Vitro Differentiation ◽  
Sex Development ◽  
Complex Process ◽  
Differences Of Sex Development ◽  
Induced Pluripotent

Sex development is a complex process involving many genes and hormones. Defects in this process lead to Differences of Sex Development (DSD), a group of heterogeneous conditions not as rare as previously thought. Part of the obstacles in proper management of these patients is due to an incomplete understanding of the genetics programs and molecular pathways involved in sex development and DSD. Several challenges delay progress and the lack of a proper model system for the single patient severely hinders advances in understanding these diseases. The revolutionary techniques of cellular reprogramming and guided in vitro differentiation allow us now to exploit the versatility of induced pluripotent stem cells to create alternatives models for DSD, ideally on a patient-specific personalized basis.

scholarly journals Overcoming barriers to reprogramming and differentiation in nonhuman primate induced pluripotent stem cells

2017 ◽  
Vol 4 (2) ◽  
pp. 153-162 ◽  
Author(s):  
Jacob J. Hemmi ◽  
Anuja Mishra ◽  
Peter J. Hornsby
Keyword(s):  
Stem Cells ◽  
Induced Pluripotent Stem Cells ◽  
Dimethyl Sulfoxide ◽  
Pluripotent Stem Cells ◽  
Ips Cells ◽  
Cellular Reprogramming ◽  
Biological Research ◽  
Differentiation Potential ◽  
Wide Range ◽  
Induced Pluripotent

Abstract. Induced pluripotent stem cells (iPS cells) generated by cellular reprogramming from nonhuman primates (NHPs) are of great significance for regenerative medicine and for comparative biology. Autologously derived stem cells would theoretically avoid any risk of rejection due to host–donor mismatch and may bypass the need for immune suppression post-transplant. In order for these possibilities to be realized, reprogramming methodologies that were initially developed mainly for human cells must be translated to NHPs. NHP studies have typically used pluripotent cells generated from young animals and thus risk overlooking complications that may arise from generating iPS cells from donors of other ages. When reprogramming is extended to a wide range of NHP species, available donors may be middle- or old-aged. Here we have pursued these questions by generating iPS cells from donors across the life span of the common marmoset (Callithrix jacchus) and then subjecting them to a directed neural differentiation protocol. The differentiation potential of different clonal cell lines was assessed using the quantitative polymerase chain reaction. The results show that cells derived from older donors often showed less neural marker induction. These deficits were rescued by a 24 h pretreatment of the cells with 0.5 % dimethyl sulfoxide. Another NHP that plays a key role in biological research is the chimpanzee (Pan troglodytes). iPS cells generated from the chimpanzee can be of great interest in comparative in vitro studies. We investigated if similar deficits in differentiation potential might arise in chimpanzee iPS cells reprogrammed using various technologies. The results show that, while some deficits were observed in iPS cell clones generated using three different technologies, there was no clear association with the vector used. These deficits in differentiation were also prevented by a 24 h pretreatment with 0.5 % dimethyl sulfoxide.

222 INCOMPLETE REPROGRAMMING OF INDUCED PLURIPOTENT STEM CELLS DERIVED FROM PORCINE FETAL FIBROBLASTS

2016 ◽  
Vol 28 (2) ◽  
pp. 242
Author(s):  
K.-H. Choi ◽  
D. Son ◽  
D.-K. Lee ◽  
J.-N. Oh ◽  
S.-H. Kim ◽  
...  
Keyword(s):  
Stem Cells ◽  
Transcription Factors ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Ectopic Expression ◽  
Cellular Reprogramming ◽  
Pluripotent State ◽  
Fetal Fibroblasts ◽  
Endogenous Genes ◽  
Induced Pluripotent

Cellular reprogramming of committed cells into a pluripotent state can be accomplished by ectopic expression of genes such as OCT4, SOX2, KLF4, and MYC. However, during reprogramming, it has been verified that failures of reactivating endogenous genes and epigenetic remodelling lead to partially reprogrammed cells exhibiting features similar to those of fully reprogrammed cells. In this study, partially reprogrammed induced pluripotent stem cells (pre-iPSC) were derived from porcine fetal fibroblasts via drug-inducible vector carrying human transcription factors (OCT4, SOX2, KLF4, and MYC). Therefore, this study aimed to investigate characteristics of pre-iPSC and reprogramming mechanisms. The pre-iPSC were stably maintained over an extended period having in vitro differentiation ability into 3 germ layers. The pluripotent state of pre-iPSC was regulated by modulation of culture condition. They showed naive- or primed-like pluripotent state in leukemia inhibitory factor (LIF) or basic fibroblast growth factor (bFGF) supplemented culture conditions respectively. However, pre-iPSC could not be maintained without ectopic expression of transgenes. The cultured pre-iPSC expressed endogenous transcription factors (OCT4 and SOX2) except for NANOG known as gateway into complete reprogramming. In addition, endogenous genes related to mesenchymal-to-epithelial transition (DPPA2, CDH1, EPCAM, and OCLN) were not sufficiently reactivated as measured by qPCR. DNA methylation analysis for promoters of OCT4, NANOG, and XIST showed that epigenetic reprogramming did not occurred in female pre-iPSC. Given the results, we found that expression of exogenous genes could not sufficiently activate the essential endogenous genes and remodel the epigenetic milieu for achieving faithful pluripotency in pig. Accordingly, investigating pre-iPSC could help us to improve and develop reprogramming methods via understanding reprogramming mechanisms in pig. This work was supported by the Next-generation BioGreen 21 Program (PJ0113002015), Rural Development Administration, Republic of Korea.

scholarly journals Metabolic–Epigenetic Axis in Pluripotent State Transitions

Epigenomes ◽  
2019 ◽  
Vol 3 (3) ◽  
pp. 13 ◽  
Author(s):  
Cristina D’Aniello ◽  
Federica Cermola ◽  
Eduardo J. Patriarca ◽  
Gabriella Minchiotti
Keyword(s):  
Pluripotent Stem Cells ◽  
Pluripotent Stem Cell ◽  
State Transition ◽  
Adult Life ◽  
State Transitions ◽  
Chromatin Modifications ◽  
Epigenetic Changes ◽  
Epigenetic Remodeling ◽  
Induced Pluripotent

Cell state transition (CST) occurs during embryo development and in adult life in response to different stimuli and is associated with extensive epigenetic remodeling. Beyond growth factors and signaling pathways, increasing evidence point to a crucial role of metabolic signals in this process. Indeed, since several epigenetic enzymes are sensitive to availability of specific metabolites, fluctuations in their levels may induce the epigenetic changes associated with CST. Here we analyze how fluctuations in metabolites availability influence DNA/chromatin modifications associated with pluripotent stem cell (PSC) transitions. We discuss current studies and focus on the effects of metabolites in the context of naïve to primed transition, PSC differentiation and reprogramming of somatic cells to induced pluripotent stem cells (iPSCs), analyzing their mechanism of action and the causal correlation between metabolites availability and epigenetic alteration.

scholarly journals Dynamic Changes in Occupancy of Histone Variant H2A.Z during Induced Somatic Cell Reprogramming

2016 ◽  
Vol 2016 ◽  
pp. 1-10 ◽  
Author(s):  
Fulu Dong ◽  
Zhenwei Song ◽  
Jiali Yu ◽  
Baole Zhang ◽  
BaoChun Jiang ◽  
...  
Keyword(s):  
Chromatin Remodeling ◽  
Pluripotent Stem Cells ◽  
Chromatin Immunoprecipitation ◽  
Chromatin Modification ◽  
Cellular Reprogramming ◽  
Transcription Start ◽  
Dynamic Changes ◽  
Go Analysis ◽  
Somatic Cell Reprogramming ◽  
Induced Pluripotent

The development of induced pluripotent stem cells (iPSCs) has enabled study of the mechanisms underlying cellular reprogramming. Here, we have studied the dynamic distribution of H2A.Z during induced reprogramming with chromatin immunoprecipitation deep sequencing (ChIP-Seq). We found that H2A.Z tended to accumulate around transcription start site (TSS) and incorporate in genes with a high transcriptional activity. GO analysis with H2A.Z incorporated genes indicated that most genes are involved in chromatin assembly or disassembly and chromatin modification both in MEF and Day 7 samples, not in iPSCs. Furthermore, we detected the highest level of incorporation of H2A.Z around TSS in Day 7 samples compared to MEFs and iPSCs. GO analysis with only incorporated genes in Day 7 also displayed the function of chromatin remodeling. So, we speculate H2A.Z may be responsible for chromatin remodeling to enhance the access of transcription factors to genes important for pluripotency. This study therefore provides a deeper understanding of the mechanisms underlying induced reprogramming.

scholarly journals Chromosomal Instability and Molecular Defects in Induced Pluripotent Stem Cells from Nijmegen Breakage Syndrome Patients

Cell Reports ◽  
2016 ◽  
Vol 16 (9) ◽  
pp. 2499-2511 ◽  
Author(s):  
Tomer Halevy ◽  
Shira Akov ◽  
Martina Bohndorf ◽  
Barbara Mlody ◽  
James Adjaye ◽  
...  
Keyword(s):  
Stem Cells ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Chromosomal Instability ◽  
Nijmegen Breakage Syndrome ◽  
Molecular Defects ◽  
Induced Pluripotent
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