Publications by authors named "Karen Sermon"

59 Publications

MSH2 knock-down shows CTG repeat stability and concomitant upstream demethylation at the DMPK locus in myotonic dystrophy type 1 human embryonic stem cells.

Hum Mol Genet 2021 Jan;29(21):3566-3577

Department Reproduction and Genetics, Vrije Universiteit Brussel, Brussels 1090, Belgium.

Myotonic dystrophy type 1 (DM1) is caused by expansion of a CTG repeat in the DMPK gene, where expansion size and somatic mosaicism correlates with disease severity and age of onset. While it is known that the mismatch repair protein MSH2 contributes to the unstable nature of the repeat, its role on other disease-related features, such as CpG methylation upstream of the repeat, is unknown. In this study, we investigated the effect of an MSH2 knock-down (MSH2KD) on both CTG repeat dynamics and CpG methylation pattern in human embryonic stem cells (hESC) carrying the DM1 mutation. Repeat size in MSH2 wild-type (MSH2WT) and MSH2KD DM1 hESC was determined by PacBio sequencing and CpG methylation by bisulfite massive parallel sequencing. We found stabilization of the CTG repeat concurrent with a gradual loss of methylation upstream of the repeat in MSH2KD cells, while the repeat continued to expand and upstream methylation remained unchanged in MSH2WT control lines. Repeat instability was re-established and biased towards expansions upon MSH2 transgenic re-expression in MSH2KD lines while upstream methylation was not consistently re-established. We hypothesize that the hypermethylation at the mutant DM1 locus is promoted by the MMR machinery and sustained by a constant DNA repair response, establishing a potential mechanistic link between CTG repeat instability and upstream CpG methylation. Our work represents a first step towards understanding how epigenetic alterations and repair pathways connect and contribute to the DM1 pathology.
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http://dx.doi.org/10.1093/hmg/ddaa250DOI Listing
January 2021

Uncovering low-level mosaicism in human embryonic stem cells using high throughput single cell shallow sequencing.

Sci Rep 2019 10 16;9(1):14844. Epub 2019 Oct 16.

Research Group Reproduction and Genetics, Vrije Universiteit Brussel, Laarbeeklaan 103, 1090, Brussels, Belgium.

Human pluripotent stem cells (hPSCs) have significant levels of low-grade genetic mosaicism, which commonly used techniques fail to detect in bulk DNA. These copy number variations remain a hurdle for the clinical translation of hPSC, as their effect in vivo ranges from unknown to dangerous, and the ability to detect them will be necessary as the field advances. As such there is need for techniques which can efficiently analyse genetic content in single cells with higher throughput and lower costs. We report here on the use of the Fluidigm C1 single cell WGA platform in combination with shallow whole genome sequencing to analyse the genetic content of single hPSCs. From a hPSC line carrying an isochromosome 20, 56 single cells were analysed and found to carry a total of 50 aberrations, across 23% of cells, which could not be detected by bulk analysis. Aberrations were predominantly segmental gains, with a fewer number of segmental losses and aneuploidies. Interestingly, 40% of the breakpoints seen here correspond to known DNA fragile sites. Our results therefore demonstrate the feasibility of single cell shallow sequencing of hPSC and further expand upon the biological importance and frequency of single cell mosaicism in hPSC.
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http://dx.doi.org/10.1038/s41598-019-51314-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6796059PMC
October 2019

Publisher Correction: In silico discovery of a FOXM1 driven embryonal signaling pathway in therapy resistant neuroblastoma tumors.

Sci Rep 2019 Jun 4;9(1):8360. Epub 2019 Jun 4.

Center for Medical Genetics (CMGG), Ghent University, Ghent, Belgium.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.
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http://dx.doi.org/10.1038/s41598-019-44435-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6547841PMC
June 2019

Iodine Dose of Administered Contrast Media Affects the Level of Radiation-Induced DNA Damage During Cardiac CT Scans.

AJR Am J Roentgenol 2019 08 23;213(2):404-409. Epub 2019 Apr 23.

1 Department of Radiology, Universitair Ziekenhuis Brussel, Vrije Universiteit Brussel, Laarbeeklaan 101, Brussels 1090, Belgium.

The purpose of this study is to investigate the contributing effect of contrast media (CM) iodine dose on radiation-induced DNA damage in blood lymphocytes during a cardiac CT scan. The minipigs were exposed 12 times in total to a fixed cardiac CT scan protocol. An unenhanced and two CM injection protocols were considered, the latter with 50% saline diluted (160 mg I/mL) and standard iodixanol. Blood samples were collected before and after CT, and radiation-induced DNA double-strand breaks were assessed using γ-H2AX (H2A histone family member X) immunofluorescent staining of the blood lymphocytes. Significant differences in foci numbers were investigated with an independent sample test. In addition, a numeric dosimetry model was applied that simulates the cardiac CT scan, with the heart represented by a blood volume containing a mixture of six iodine concentrations (0, 10, 20, 30, 40, and 50 mg I/mL). Compared with the unenhanced (0 mg I/mL) protocol, the number of γ-H2AX foci per cell increased significantly ( < 0.038), by 56.1% for the reduced iodine dose (160 mg I/mL) and by 141.1% for the standard iodine dose (320 mg I/mL) protocols. These in vivo results are confirmed by the dosimetry simulation model, in which 78.8% and 133.7% increases in locally absorbed blood dose in the left ventricle were observed for the reduced and standard iodine dose protocols, respectively. Administration of CM during a cardiac CT examination significantly increases radiation-induced DNA damage in blood lymphocytes. Moreover, a lower CM iodine dose results in a reduced level of DNA damage, at constant radiation exposure.
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http://dx.doi.org/10.2214/AJR.18.20950DOI Listing
August 2019

In silico discovery of a FOXM1 driven embryonal signaling pathway in therapy resistant neuroblastoma tumors.

Sci Rep 2018 11 30;8(1):17468. Epub 2018 Nov 30.

Center for Medical Genetics (CMGG), Ghent University, Ghent, Belgium.

Chemotherapy resistance is responsible for high mortality rates in neuroblastoma. MYCN, an oncogenic driver in neuroblastoma, controls pluripotency genes including LIN28B. We hypothesized that enhanced embryonic stem cell (ESC) gene regulatory programs could mark tumors with high pluripotency capacity and subsequently increased risk for therapy failure. An ESC miRNA signature was established based on publicly available data. In addition, an ESC mRNA signature was generated including the 500 protein coding genes with the highest positive expression correlation with the ESC miRNA signature score in 200 neuroblastomas. High ESC m(i)RNA expression signature scores were significantly correlated with poor neuroblastoma patient outcome specifically in the subgroup of MYCN amplified tumors and stage 4 nonamplified tumors. Further data-mining identified FOXM1, as the major predicted driver of this ESC signature, controlling a large set of genes implicated in cell cycle control and DNA damage response. Of further interest, re-analysis of published data showed that MYCN transcriptionally activates FOXM1 in neuroblastoma cells. In conclusion, a novel ESC m(i)RNA signature stratifies neuroblastomas with poor prognosis, enabling the identification of therapy-resistant tumors. The finding that this signature is strongly FOXM1 driven, warrants for drug design targeted at FOXM1 or key components controlling this pathway.
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http://dx.doi.org/10.1038/s41598-018-35868-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6269481PMC
November 2018

Preimplantation genetic testing for aneuploidy by microarray analysis of polar bodies in advanced maternal age: a randomized clinical trial.

Hum Reprod 2018 09;33(9):1767-1776

Research Group Reproduction and Genetics, Vrije Universiteit Brussel, Laarbeeklaan 103, Brussels, Belgium.

Study Question: Does preimplantation genetic testing for aneuploidy (PGT-A) by comprehensive chromosome screening (CCS) of the first and second polar body to select embryos for transfer increase the likelihood of a live birth within 1 year in advanced maternal age women aged 36-40 years planning an ICSI cycle, compared to ICSI without chromosome analysis?

Summary Answer: PGT-A by CCS in the first and second polar body to select euploid embryos for transfer does not substantially increase the live birth rate in women aged 36-40 years.

What Is Known Already: PGT-A has been used widely to select embryos for transfer in ICSI treatment, with the aim of improving treatment effectiveness. Whether PGT-A improves ICSI outcomes and is beneficial to the patients has remained controversial.

Study Design, Size, Duration: This is a multinational, multicentre, pragmatic, randomized clinical trial with intention-to-treat analysis. Of 396 women enroled between June 2012 and December 2016, 205 were allocated to CCS of the first and second polar body (study group) as part of their ICSI treatment cycle and 191 were allocated to ICSI treatment without chromosome screening (control group). Block randomization was performed stratified for centre and age group. Participants and clinicians were blinded at the time of enrolment until the day after intervention.

Participants/materials, Setting, Methods: Infertile couples in which the female partner was 36-40 years old and who were scheduled to undergo ICSI treatment were eligible. In those assigned to PGT-A, array comparative genomic hybridization (aCGH) analysis of the first and second polar bodies of the fertilized oocytes was performed using the 24sure array of Illumina. If in the first treatment cycle all oocytes were aneuploid, a second treatment with PB array CGH was offered. Participants in the control arm were planned for ICSI without PGT-A. Main exclusion criteria were three or more previous unsuccessful IVF or ICSI cycles, three or more clinical miscarriages, poor response or low ovarian reserve. The primary outcome was the cumulative live birth rate after fresh or frozen embryo transfer recorded over 1 year after the start of the intervention.

Main Results And The Role Of Chance: Of the 205 participants in the chromosome screening group, 50 (24%) had a live birth with intervention within 1 year, compared to 45 of the 191 in the group without intervention (24%), a difference of 0.83% (95% CI: -7.60 to 9.18%). There were significantly fewer participants in the chromosome screening group with a transfer (relative risk (RR) = 0.81; 95% CI: 0.74-0.89) and fewer with a miscarriage (RR = 0.48; 95% CI: 0.26-0.90).

Limitations, Reasons For Caution: The targeted sample size was not reached because of suboptimal recruitment; however, the included sample allowed a 90% power to detect the targeted increase. Cumulative outcome data were limited to 1 year. Only 11 patients out of 32 with exclusively aneuploid results underwent a second treatment cycle in the chromosome screening group.

Wider Implications Of The Findings: The observation that the similarity in birth rates was achieved with fewer transfers, less cryopreservation and fewer miscarriages points to a clinical benefit of PGT-A, and this form of embryo selection may, therefore, be considered to minimize the number of interventions while producing comparable outcomes. Whether these benefits outweigh drawbacks such as the cost for the patient, the higher workload for the IVF lab and the potential effect on the children born after prolonged culture and/or cryopreservation remains to be shown.

Study Funding/competing Interest(s): This study was funded by the European Society of Human Reproduction and Embryology. Illumina provided microarrays and other consumables necessary for aCGH testing of polar bodies. M.B.'s institution (UZBrussel) has received educational grants from IBSA, Ferring, Organon, Schering-Plough, Merck and Merck Belgium. M.B. has received consultancy and speakers' fees from Organon, Serono Symposia and Merck. G.G. has received personal fees and non-financial support from MSD, Ferring, Merck-Serono, Finox, TEVA, IBSA, Glycotope, Abbott and Gedeon-Richter as well as personal fees from VitroLife, NMC Healthcare, ReprodWissen, BioSilu and ZIVA. W.V., C.S., P.M.B., V.G., G.A., M.D., T.E.G., L.G., G.Ka., G.Ko., J.L., M.C.M., M.P., A.S., M.T., K.V., J.G. and K.S. declare no conflict of interest.

Trial Registration Number: NCT01532284.

Trial Registration Date: 7 February 2012.

Date Of First Patient’s Enrolment: 25 June 2012.
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http://dx.doi.org/10.1093/humrep/dey262DOI Listing
September 2018

Random Mutagenesis, Clonal Events, and Embryonic or Somatic Origin Determine the mtDNA Variant Type and Load in Human Pluripotent Stem Cells.

Stem Cell Reports 2018 07 14;11(1):102-114. Epub 2018 Jun 14.

Research Group Reproduction and Genetics, Faculty of Medicine and Pharmacy, Vrije Universiteit Brussel (VUB), Laarbeeklaan 103, Brussels 1090, Belgium. Electronic address:

In this study, we deep-sequenced the mtDNA of human embryonic and induced pluripotent stem cells (hESCs and hiPSCs) and their source cells and found that the majority of variants pre-existed in the cells used to establish the lines. Early-passage hESCs carried few and low-load heteroplasmic variants, similar to those identified in oocytes and inner cell masses. The number and heteroplasmic loads of these variants increased with prolonged cell culture. The study of 120 individual cells of early- and late-passage hESCs revealed a significant diversity in mtDNA heteroplasmic variants at the single-cell level and that the variants that increase during time in culture are always passenger to the appearance of chromosomal abnormalities. We found that early-passage hiPSCs carry much higher loads of mtDNA variants than hESCs, which single-fibroblast sequencing proved pre-existed in the source cells. Finally, we show that these variants are stably transmitted during short-term differentiation.
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http://dx.doi.org/10.1016/j.stemcr.2018.05.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6117474PMC
July 2018

Genetic and epigenetic factors which modulate differentiation propensity in human pluripotent stem cells.

Hum Reprod Update 2018 Mar;24(2):162-175

Research group Reproduction and Genetics (REGE), Vrije Universiteit Brussel (VUB), Laarbeeklaan 103, 1090 Jette, Belgium.

Background: Human pluripotent stem cell (hPSC) lines are known to have a bias in their differentiation. This gives individual cell lines a propensity to preferentially differentiate towards one germ layer or cell type over others. Chromosomal aberrations, mitochondrial mutations, genetic diversity and epigenetic variance are the main drivers of this phenomenon, and can lead to a wide range of phenotypes.

Objective And Rationale: Our aim is to provide a comprehensive overview of the different factors which influence differentiation propensity. Specifically, we sought to highlight known genetic variances and their mechanisms, in addition to more general observations from larger abnormalities. Furthermore, we wanted to provide an up-to-date list of a growing number of predictive indicators which are able to identify differentiation propensity before the initiation of differentiation. As differentiation propensity can lead to difficulties in both research as well as clinical translation, our thorough overview could be a useful tool.

Search Methods: Combinations of the following key words were applied as search criteria in the PubMed database: embryonic stem cells, induced pluripotent stem cells, differentiation propensity (also: potential, efficiency, capacity, bias, variability), epigenetics, chromosomal abnormalities, genetic aberrations, X chromosome inactivation, mitochondrial function, mitochondrial metabolism, genetic diversity, reprogramming, predictive marker, residual stem cell, clinic. Only studies in English were included, ranging from 2000 to 2017, with a majority ranging from 2010 to 1017. Further manuscripts were added from cross-references.

Outcomes: Differentiation propensity is affected by a wide variety of (epi)genetic factors. These factors clearly lead to a loss of differentiation capacity, preference towards certain cell types and oftentimes, phenotypes which begin to resemble cancer. Broad changes in (epi)genetics, such as aneuploidies or wide-ranging modifications to the epigenetic landscape tend to lead to extensive, less definite changes in differentiation capacity, whereas more specific abnormalities often have precise ramifications in which certain cell types become more preferential. Furthermore, there appears to be a greater, though often less considered, contribution to differentiation propensity by factors such as mitochondria and inherent genetic diversity. Varied differentiation capacity can also lead to potential consequences in the clinical translation of hPSC, including the occurrence of residual undifferentiated stem cells, and the transplantation of potentially transformed cells.

Wider Implications: As hPSC continue to advance towards the clinic, our understanding of them progresses as well. As a result, the challenges faced become more numerous, but also more clear. If the transition to the clinic is to be achieved with a minimum number of potential setbacks, thorough evaluation of the cells will be an absolute necessity. Altered differentiation propensity represents at least one such hurdle, for which researchers and eventually clinicians will need to find solutions. Already, steps are being taken to tackle the issue, though further research will be required to evaluate any long-term risks it poses.
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http://dx.doi.org/10.1093/humupd/dmx042DOI Listing
March 2018

Accurate and comprehensive analysis of single nucleotide variants and large deletions of the human mitochondrial genome in DNA and single cells.

Eur J Hum Genet 2017 11 23;25(11):1229-1236. Epub 2017 Aug 23.

Research Group Reproduction and Genetics, Faculty of Medicine and Pharmacy, Vrije Universiteit Brussel (VUB), Brussels, Belgium.

Massive parallel sequencing (MPS) can accurately quantify mitochondrial DNA (mtDNA) single nucleotide variants (SNVs), but no MPS methods are currently validated to simultaneously and accurately establish the breakpoints and frequency of large deletions at low heteroplasmic loads. Here we present the thorough validation of an MPS protocol to quantify the load of very low frequency, large mtDNA deletions in bulk DNA and single cells, along with SNV calling by standard methods. We used a set of well-characterized DNA samples, DNA mixes and single cells to thoroughly control the study. We developed a custom script for the detection of mtDNA rearrangements that proved to be more accurate in detecting and quantifying deletions than pre-existing tools. We also show that PCR conditions and primersets must be carefully chosen to avoid biases in the retrieved variants and an increase in background noise, and established a lower detection limit of 0.5% heteroplasmic load for large deletions, and 1.5 and 2% for SNVs, for bulk DNA and single cells, respectively. Finally, the analysis of different single cells provided novel insights into mtDNA cellular mosaicism.
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http://dx.doi.org/10.1038/ejhg.2017.129DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5643964PMC
November 2017

CpG Methylation, a Parent-of-Origin Effect for Maternal-Biased Transmission of Congenital Myotonic Dystrophy.

Am J Hum Genet 2017 Mar;100(3):488-505

Genetics & Genome Biology, The Hospital for Sick Children, Toronto, ON M5G 0A4, Canada; The Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 1A8, Canada. Electronic address:

CTG repeat expansions in DMPK cause myotonic dystrophy (DM1) with a continuum of severity and ages of onset. Congenital DM1 (CDM1), the most severe form, presents distinct clinical features, large expansions, and almost exclusive maternal transmission. The correlation between CDM1 and expansion size is not absolute, suggesting contributions of other factors. We determined CpG methylation flanking the CTG repeat in 79 blood samples from 20 CDM1-affected individuals; 21, 27, and 11 individuals with DM1 but not CDM1 (henceforth non-CDM1) with maternal, paternal, and unknown inheritance; and collections of maternally and paternally derived chorionic villus samples (7 CVSs) and human embryonic stem cells (4 hESCs). All but two CDM1-affected individuals showed high levels of methylation upstream and downstream of the repeat, greater than non-CDM1 individuals (p = 7.04958 × 10). Most non-CDM1 individuals were devoid of methylation, where one in six showed downstream methylation. Only two non-CDM1 individuals showed upstream methylation, and these were maternally derived childhood onset, suggesting a continuum of methylation with age of onset. Only maternally derived hESCs and CVSs showed upstream methylation. In contrast, paternally derived samples (27 blood samples, 3 CVSs, and 2 hESCs) never showed upstream methylation. CTG tract length did not strictly correlate with CDM1 or methylation. Thus, methylation patterns flanking the CTG repeat are stronger indicators of CDM1 than repeat size. Spermatogonia with upstream methylation may not survive due to methylation-induced reduced expression of the adjacent SIX5, thereby protecting DM1-affected fathers from having CDM1-affected children. Thus, DMPK methylation may account for the maternal bias for CDM1 transmission, larger maternal CTG expansions, age of onset, and clinical continuum, and may serve as a diagnostic indicator.
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http://dx.doi.org/10.1016/j.ajhg.2017.01.033DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5339342PMC
March 2017

Adjuncts in the IVF laboratory: where is the evidence for 'add-on' interventions?

Hum Reprod 2017 03;32(3):485-491

Reproductive Medicine, Sahlgrenska University Hospital, Goteborg, Sweden.

Globally, IVF patients are routinely offered and charged for a selection of adjunct treatments and tests or 'add-ons' that they are told may improve their chance of a live birth, despite there being no clinical evidence supporting the efficacy of the add-on. Any new IVF technology claiming to improve live birth rates (LBR) should, in most cases, first be tested in an appropriate animal model, then in clinical trials, to ensure safety, and finally in a randomized controlled trial (RCT) to provide high-quality evidence that the procedure is safe and effective. Only then should the technique be considered as 'routine' and only when applied to the similar patient population as those studied in the RCT. Even then, further pediatric and long-term follow-up studies will need to be undertaken to examine the long-term safety of the procedure. Alarmingly, there are currently numerous examples where adjunct treatments are used in the absence of evidence-based medicine and often at an additional fee. In some cases, when RCTs have shown the technique to be ineffective, it is eventually withdrawn from the clinic. In this paper, we discuss some of the adjunct treatments currently being offered globally in IVF laboratories, including embryo glue and adherence compounds, sperm DNA fragmentation, time-lapse imaging, preimplantation genetic screening, mitochondria DNA load measurement and assisted hatching. We examine the evidence for their safety and efficacy in increasing LBRs. We conclude that robust studies are needed to confirm the safety and efficacy of any adjunct treatment or test before they are offered routinely to IVF patients.
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http://dx.doi.org/10.1093/humrep/dex004DOI Listing
March 2017

Detecting mosaicism in trophectoderm biopsies.

Hum Reprod 2017 03;32(3):712-713

GENOMA-Molecular Genetics Laboratories, Via di Castel Giubileo, 1100138Rome, Italy.

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http://dx.doi.org/10.1093/humrep/dew346DOI Listing
March 2017

Novel technologies emerging for preimplantation genetic diagnosis and preimplantation genetic testing for aneuploidy.

Authors:
Karen Sermon

Expert Rev Mol Diagn 2017 01 1;17(1):71-82. Epub 2016 Dec 1.

a Research Group Reproduction and Genetics , Vrije Universiteit Brussel , Brussels , Belgium.

Introduction: Preimplantation genetic diagnosis (PGD) was introduced as an alternative to prenatal diagnosis: embryos cultured in vitro were analysed for a monogenic disease and only disease-free embryos were transferred to the mother, to avoid the termination of pregnancy with an affected foetus. It soon transpired that human embryos show a great deal of acquired chromosomal abnormalities, thought to explain the low success rate of IVF - hence preimplantation genetic testing for aneuploidy (PGT-A) was developed to select euploid embryos for transfer. Areas covered: PGD has followed the tremendous evolution in genetic analysis, with only a slight delay due to adaptations for diagnosis on small samples. Currently, next generation sequencing combining chromosome with single-base pair analysis is on the verge of becoming the golden standard in PGD and PGT-A. Papers highlighting the different steps in the evolution of PGD/PGT-A were selected. Expert commentary: Different methodologies used in PGD/PGT-A with their pros and cons are discussed.
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http://dx.doi.org/10.1080/14737159.2017.1262261DOI Listing
January 2017

A High Proliferation Rate is Critical for Reproducible and Standardized Embryoid Body Formation from Laminin-521-Based Human Pluripotent Stem Cell Cultures.

Stem Cell Rev Rep 2016 Dec;12(6):721-730

Research Group Reproduction and Genetics, Faculty of Medicine and Pharmacy, Vrije Universiteit Brussel, Laarbeeklaan 103, 1090, Brussels, Belgium.

When aiming for homogenous embryoid body (EB) differentiation, the use of equal-sized EBs is required to avoid a size-induced differentiation bias. In this study we developed an efficient and standardized EB formation protocol for human pluripotent stem cells (hPSC) cultured in a laminin-521-based xeno-free system. As the cell proliferation rate of the cells growing on laminin-521 strongly affected the efficiency of aggregate formation, we found that recently passaged cells, as well as the addition of ROCK inhibitor, were essential for reproducible EB formation from hPSC single-cell suspensions. EBs could be obtained in a variety of differentiation media, in 96-well round-bottom plates and in hanging drops. Gene expression studies on differentially sized EBs from three individual human embryonic stem cell lines demonstrated that the medium used for differentiation influenced the differentiation outcome to a much greater extent than the number of cells used for the initial EB formation. Our findings give a new insight into factors that influence the EB formation and differentiation process. This optimized method allows us to easily manipulate EB formation and provide an excellent starting point for downstream EB-based differentiation protocols.
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http://dx.doi.org/10.1007/s12015-016-9679-zDOI Listing
December 2016

The why, the how and the when of PGS 2.0: current practices and expert opinions of fertility specialists, molecular biologists, and embryologists.

Mol Hum Reprod 2016 08 2;22(8):845-57. Epub 2016 Jun 2.

Department of Reproductive Medicine, Maastricht University Medical Center, P. Debyelaan 25, 6229 HX Maastricht, The Netherlands Department of Clinical Genetics, Maastricht University Medical Center, P. Debyelaan 25, 6229 HX Maastricht, The Netherlands.

Study Question: We wanted to probe the opinions and current practices on preimplantation genetic screening (PGS), and more specifically on PGS in its newest form: PGS 2.0?

Study Finding: Consensus is lacking on which patient groups, if any at all, can benefit from PGS 2.0 and, a fortiori, whether all IVF patients should be offered PGS.

What Is Known Already: It is clear from all experts that PGS 2.0 can be defined as biopsy at the blastocyst stage followed by comprehensive chromosome screening and possibly combined with vitrification. Most agree that mosaicism is less of an issue at the blastocyst stage than at the cleavage stage but whether mosaicism is no issue at all at the blastocyst stage is currently called into question.

Study Design, Samples/materials, Methods: A questionnaire was developed on the three major aspects of PGS 2.0: the Why, with general questions such as PGS 2.0 indications; the How, specifically on genetic analysis methods; the When, on the ideal method and timing of embryo biopsy. Thirty-five colleagues have been selected to address these questions on the basis of their experience with PGS, and demonstrated by peer-reviewed publications, presentations at meetings and participation in the discussion. The first group of experts who were asked about 'The Why' comprised fertility experts, the second group of molecular biologists were asked about 'The How' and the third group of embryologists were asked about 'The When'. Furthermore, the geographical distribution of the experts has been taken into account. Thirty have filled in the questionnaire as well as actively participated in the redaction of the current paper.

Main Results And The Role Of Chance: The 30 participants were from Europe (Belgium, Germany, Greece, Italy, Netherlands, Spain, UK) and the USA. Array comparative genome hybridization is the most widely used method amongst the participants, but it is slowly being replaced by massive parallel sequencing. Most participants offering PGS 2.0 to their patients prefer blastocyst biopsy. The high efficiency of vitrification of blastocysts has added a layer of complexity to the discussion, and it is not clear whether PGS in combination with vitrification, PGS alone, or vitrification alone, followed by serial thawing and eSET will be the favoured approach. The opinions range from in favour of the introduction of PGS 2.0 for all IVF patients, over the proposal to use PGS as a tool to rank embryos according to their implantation potential, to scepticism towards PGS pending a positive outcome of robust, reliable and large-scale RCTs in distinct patient groups.

Limitations, Reasons For Caution: Care was taken to obtain a wide spectrum of views from carefully chosen experts. However, not all invited experts agreed to participate, which explains a lack of geographical coverage in some areas, for example China. This paper is a collation of current practices and opinions, and it was outside the scope of this study to bring a scientific, once-and-for-all solution to the ongoing debate.

Wider Implications Of The Findings: This paper is unique in that it brings together opinions on PGS 2.0 from all different perspectives and gives an overview of currently applied technologies as well as potential future developments. It will be a useful reference for fertility specialists with an expertise outside reproductive genetics.

Large Scale Data: none.

Study Funding And Competing Interests: No specific funding was obtained to conduct this questionnaire.
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http://dx.doi.org/10.1093/molehr/gaw034DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4986417PMC
August 2016

Preimplantation genetic screening 2.0: the theory.

Mol Hum Reprod 2016 08 2;22(8):839-44. Epub 2016 Jun 2.

Research Group Reproduction and Genetics, Vrije Universiteit Brussel, Laarbeeklaan 101, Brussels 1090, Belgium.

During the last few years a new generation of preimplantation genetic screening (PGS) has been introduced. In this paper, an overview of the different aspects of this so-called PGS 2.0 with respect to the why (what are the indications), the when (which developmental stage, i.e. which material should be studied) and the how (which molecular technique should be used) is given. With respect to the aims it is clear that PGS 2.0 can be used for a variety of indications. However, the beneficial effect of PGS 2.0 has not been proved yet in RCTs. It is clear that cleavage stage is not the optimal stage for biopsy. Almost all advocates of PGS 2.0 prefer trophectoderm biopsy. There are many new methods that allow the study of complete aneuploidy with respect to one or more of the 24 chromosomes. Because of the improved vitrification methods, selection of fresh embryos for transfer is more and more often replaced by frozen embryo transfer. The main goal of PGS has always been the improvement of IVF success. However, success is defined by different authors in many different ways. This makes it very difficult to compare the outcomes of different studies. In conclusion, the introduction of PGS 2.0 will depend on the success of the new biopsy strategies in combination with the analysis of all 24 chromosomes. It remains to be seen which approach will be the most successful and for which specific groups of patients.
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http://dx.doi.org/10.1093/molehr/gaw033DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4986416PMC
August 2016

Higher-Density Culture in Human Embryonic Stem Cells Results in DNA Damage and Genome Instability.

Stem Cell Reports 2016 Mar 25;6(3):330-41. Epub 2016 Feb 25.

Research Group Reproduction and Genetics, Faculty of Medicine and Pharmacy, Vrije Universiteit Brussel (VUB), Laarbeeklaan 103, 1090 Brussels, Belgium. Electronic address:

Human embryonic stem cells (hESC) show great promise for clinical and research applications, but their well-known proneness to genomic instability hampers the development to their full potential. Here, we demonstrate that medium acidification linked to culture density is the main cause of DNA damage and genomic alterations in hESC grown on feeder layers, and this even in the short time span of a single passage. In line with this, we show that increasing the frequency of the medium refreshments minimizes the levels of DNA damage and genetic instability. Also, we show that cells cultured on laminin-521 do not present this increase in DNA damage when grown at high density, although the (long-term) impact on their genomic stability remains to be elucidated. Our results explain the high levels of genome instability observed over the years by many laboratories worldwide, and show that the development of optimal culture conditions is key to solving this problem.
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http://dx.doi.org/10.1016/j.stemcr.2016.01.015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4788786PMC
March 2016

The Role of D4Z4-Encoded Proteins in the Osteogenic Differentiation of Mesenchymal Stromal Cells Isolated from Bone Marrow.

Stem Cells Dev 2015 Nov 26;24(22):2674-86. Epub 2015 Aug 26.

1 Laboratory of Molecular Biology, University of Mons , Mons, Belgium .

Facioscapulohumeral muscular dystrophy (FSHD) is associated with an activation of the double homeobox 4 (DUX4) gene, which we previously identified within the D4Z4 repeated elements in the 4q35 subtelomeric region. The pathological DUX4 mRNA is derived from the most distal D4Z4 unit and extends unexpectedly within the flanking pLAM region, which provides an intron and polyadenylation signal. The conditions that are required to develop FSHD are a permissive allele providing the polyadenylation signal and hypomethylation of the D4Z4 repeat array compared with the healthy muscle. The DUX4 protein is a 52-kDa transcription factor that initiates a large gene deregulation cascade leading to muscle atrophy, inflammation, differentiation defects, and oxidative stress, which are the key features of FSHD. DUX4 is a retrogene that is normally expressed in germline cells and is submitted to repeat-induced silencing in adult tissues. Since DUX4 mRNAs have been detected in human embryonic and induced pluripotent stem cells, we investigated whether they could also be expressed in human mesenchymal stromal cells (hMSCs). We found that DUX4 mRNAs were induced during the differentiation of hMSCs into osteoblasts and that this process involved DUX4 and new longer protein forms (58 and 70 kDa). A DUX4 mRNA with a more distant 5' start site was characterized that presented a 60-codon reading frame extension and encoded the 58-kDa protein. Transfections of hMSCs with an antisense oligonucleotide targeting DUX4 mRNAs decreased both the 52- and 58-kDa protein levels and confirmed their identity. Gain- and loss-of-function experiments in hMSCs suggested these DUX4 proteins had opposite roles in osteogenic differentiation as evidenced by the alkaline phosphatase activity and calcium deposition. Differentiation was delayed by the 58-kDa DUX4 expression and it was increased by 52-kDa DUX4. These data indicate a role for DUX4 protein forms in the osteogenic differentiation of hMSCs.
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http://dx.doi.org/10.1089/scd.2014.0575DOI Listing
November 2015

Concurrent whole-genome haplotyping and copy-number profiling of single cells.

Am J Hum Genet 2015 Jun 14;96(6):894-912. Epub 2015 May 14.

Centre for Human Genetics, University Hospital Leuven, Department of Human Genetics, KU Leuven, 3000 Leuven, Belgium; Single-cell Genomics Centre, Welcome Trust Sanger Institute, Hinxton CB10 1SA, UK. Electronic address:

Methods for haplotyping and DNA copy-number typing of single cells are paramount for studying genomic heterogeneity and enabling genetic diagnosis. Before analyzing the DNA of a single cell by microarray or next-generation sequencing, a whole-genome amplification (WGA) process is required, but it substantially distorts the frequency and composition of the cell's alleles. As a consequence, haplotyping methods suffer from error-prone discrete SNP genotypes (AA, AB, BB) and DNA copy-number profiling remains difficult because true DNA copy-number aberrations have to be discriminated from WGA artifacts. Here, we developed a single-cell genome analysis method that reconstructs genome-wide haplotype architectures as well as the copy-number and segregational origin of those haplotypes by employing phased parental genotypes and deciphering WGA-distorted SNP B-allele fractions via a process we coin haplarithmisis. We demonstrate that the method can be applied as a generic method for preimplantation genetic diagnosis on single cells biopsied from human embryos, enabling diagnosis of disease alleles genome wide as well as numerical and structural chromosomal anomalies. Moreover, meiotic segregation errors can be distinguished from mitotic ones.
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http://dx.doi.org/10.1016/j.ajhg.2015.04.011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4473724PMC
June 2015

Human embryonic stem cells show low-grade microsatellite instability.

Mol Hum Reprod 2014 Oct 31;20(10):981-9. Epub 2014 Jul 31.

Research Group Reproduction and Genetics, Faculty of Medicine and Pharmacy, Vrije Universiteit Brussel (VUB), Laarbeeklaan 103, 1090 Brussels, Belgium

It is well known that human embryonic stem cells (hESCs) frequently acquire recurrent chromosomal abnormalities, very reminiscent of those found in cancerous cells. Given the parallels between cancer and stem cell biology, we set out to investigate the occurrence of a common form of genome instability in tumors, namely microsatellite instability (MSI), in hESCs. MSI is caused by a deficiency in mismatch repair (MMR) genes, which leads to the accumulation of mutations during DNA replication. In this study, we analyzed up to 122 microsatellites in a total of 10 hESC lines, for 1-11 different passages, ranging from passage 7 to passage 334. In two lines, this revealed that two microsatellites had altered allelic patterns. Small-pool PCR for several microsatellites and testing of the Bethesda panel microsatellites (commonly used in cancer studies) revealed that, whilst MSI is common in all tested lines, it occurs at a very low and variable frequency, ranging from ∼1 to 20% of the total number of alleles. In cancerous cells, MSI leads to multiple large shifts in allele sizes within the majority of the cells, while hESCs show small changes in a minority of the cells. Since these genetic alterations do not consistently take over the culture, we assume that they are not concurrent with a selective advantage as it is in tumors. Finally, the MMR genes showed a very variable gene expression that could not be correlated with the variable (low) levels of MSI in the different hESC lines.
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http://dx.doi.org/10.1093/molehr/gau059DOI Listing
October 2014

Current issues in medically assisted reproduction and genetics in Europe: research, clinical practice, ethics, legal issues and policy.

Hum Reprod 2014 Aug 8;29(8):1603-9. Epub 2014 Jul 8.

Department of Biology and Medical Genetics, Charles University-2 Faculty of Medicine and University Hospital Motol, Prague, Czech Republic.

Study Question: How has the interface between genetics and assisted reproduction technology (ART) evolved since 2005?

Summary Answer: The interface between ART and genetics has become more entwined as we increase our understanding about the genetics of infertility and we are able to perform more comprehensive genetic testing.

What Is Known Already: In March 2005, a group of experts from the European Society of Human Genetics and European Society of Human Reproduction and Embryology met to discuss the interface between genetics and ART and published an extended background paper, recommendations and two Editorials.

Study Design, Size, Duration: An interdisciplinary workshop was held, involving representatives of both professional societies and experts from the European Union Eurogentest2 Coordination Action Project.

Participants/materials, Setting, Methods: In March 2012, a group of experts from the European Society of Human Genetics, the European Society of Human Reproduction and Embryology and the EuroGentest2 Coordination Action Project met to discuss developments at the interface between clinical genetics and ART.

Main Results And The Role Of Chance: As more genetic causes of reproductive failure are now recognized and an increasing number of patients undergo testing of their genome prior to conception, either in regular health care or in the context of direct-to-consumer testing, the need for genetic counselling and PGD may increase. Preimplantation genetic screening (PGS) thus far does not have evidence from RCTs to substantiate that the technique is both effective and efficient. Whole genome sequencing may create greater challenges both in the technological and interpretational domains, and requires further reflection about the ethics of genetic testing in ART and PGD/PGS. Diagnostic laboratories should be reporting their results according to internationally accepted accreditation standards (ISO 15189). Further studies are needed in order to address issues related to the impact of ART on epigenetic reprogramming of the early embryo.

Limitations, Reasons For Caution: The legal landscape regarding assisted reproduction is evolving, but still remains very heterogeneous and often contradictory. The lack of legal harmonization and uneven access to infertility treatment and PGD/PGS fosters considerable cross-border reproductive care in Europe, and beyond.

Wider Implications Of The Findings: This continually evolving field requires communication between the clinical genetics and IVF teams and patients to ensure that they are fully informed and can make well-considered choices.

Study Funding/competing Interests: Funding was received from ESHRE, ESHG and EuroGentest2 European Union Coordination Action project (FP7 - HEALTH-F4-2010-26146) to support attendance at this meeting.
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http://dx.doi.org/10.1093/humrep/deu130DOI Listing
August 2014

Current issues in medically assisted reproduction and genetics in Europe: research, clinical practice, ethics, legal issues and policy. European Society of Human Genetics and European Society of Human Reproduction and Embryology.

Eur J Hum Genet 2013 Nov;21 Suppl 2:S1-21

UCL Centre for PG&D, Institute for Womens Health, University College London, London, UK.

In March 2005, a group of experts from the European Society of Human Genetics and European Society of Human Reproduction and Embryology met to discuss the interface between genetics and assisted reproductive technology (ART), and published an extended background paper, recommendations and two Editorials. Seven years later, in March 2012, a follow-up interdisciplinary workshop was held, involving representatives of both professional societies, including experts from the European Union Eurogentest2 Coordination Action Project. The main goal of this meeting was to discuss developments at the interface between clinical genetics and ARTs. As more genetic causes of reproductive failure are now recognised and an increasing number of patients undergo testing of their genome before conception, either in regular health care or in the context of direct-to-consumer testing, the need for genetic counselling and preimplantation genetic diagnosis (PGD) may increase. Preimplantation genetic screening (PGS) thus far does not have evidence from randomised clinical trials to substantiate that the technique is both effective and efficient. Whole-genome sequencing may create greater challenges both in the technological and interpretational domains, and requires further reflection about the ethics of genetic testing in ART and PGD/PGS. Diagnostic laboratories should be reporting their results according to internationally accepted accreditation standards (International Standards Organisation - ISO 15189). Further studies are needed in order to address issues related to the impact of ART on epigenetic reprogramming of the early embryo. The legal landscape regarding assisted reproduction is evolving but still remains very heterogeneous and often contradictory. The lack of legal harmonisation and uneven access to infertility treatment and PGD/PGS fosters considerable cross-border reproductive care in Europe and beyond. The aim of this paper is to complement previous publications and provide an update of selected topics that have evolved since 2005.
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http://dx.doi.org/10.1038/ejhg.2013.219DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3831061PMC
November 2013

Role of BMP signaling in pancreatic progenitor differentiation from human embryonic stem cells.

Stem Cell Rev Rep 2013 Oct;9(5):569-77

Cell Differentiation Unit, Diabetes Research Center, Vrije Universiteit Brussel (VUB), Laarbeeklaan 103, 1090, Brussels, Belgium.

Transplantation of pancreatic progenitors derived from human embryonic stem cells (hESCs) is a promising way to treat diabetes. Strategies to obtain the required cell mass would rely on the up scaling of current differentiation protocols, or the proliferation of committed progenitors. We aimed at finding conditions that maintain a proliferating pancreatic progenitor pool and we assessed the role of BMP4 signaling in this process. hESCs were differentiated into PDX1 positive pancreatic progenitor stage following our established protocol with few modifications, and then the progenitor cells were passaged in a defined proliferation medium (PM). During passage, the effect of BMP4 signaling on the differentiation and proliferation of pancreatic progenitors was examined by RT-PCR and immunofluorescence analysis. We found that PDX1 positive pancreatic progenitors proliferated and gained NKX6.1 expression in the PM, whereas they failed to express NKX6.1 if BMP signaling was inhibited with Noggin. In this latter condition, part of the progenitors rather generated pro-endocrine cells denoted by NGN3 and synaptophysin expression. On the contrary, addition of BMP4 to the PM promoted the early derivation of PDX1 and NKX6.1 coexpressing pancreatic progenitors. Our findings are in line with mouse pancreas development, and indicate that BMP4 signaling is required for the derivation and maintenance of hESC-derived PDX1+NKX6.1+ pancreatic progenitors. These results are instructive for guiding the development of an efficient pancreas differentiation protocol in view of diabetes cell replacement therapy.
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http://dx.doi.org/10.1007/s12015-013-9435-6DOI Listing
October 2013

Human embryonic stem cells commonly display large mitochondrial DNA deletions.

Nat Biotechnol 2013 Jan;31(1):20-3

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http://dx.doi.org/10.1038/nbt.2473DOI Listing
January 2013

FGF signaling via MAPK is required early and improves Activin A-induced definitive endoderm formation from human embryonic stem cells.

Biochem Biophys Res Commun 2012 Sep 29;426(3):380-5. Epub 2012 Aug 29.

Cell Differentiation Unit, Diabetes Research Center, Vrije Universiteit Brussel (VUB), Laarbeeklaan 103, 1090 Brussels, Belgium.

Considering their unlimited proliferation and pluripotency properties, human embryonic stem cells (hESCs) constitute a promising resource applicable for cell replacement therapy. To facilitate this clinical translation, it is critical to study and understand the early stage of hESCs differentiation wherein germ layers are defined. In this study, we examined the role of FGF signaling in Activin A-induced definitive endoderm (DE) differentiation in the absence of supplemented animal serum. We found that activated FGF/MAPK signaling is required at the early time point of Activin A-induced DE formation. In addition, FGF activation increased the number of DE cells compared to Activin A alone. These DE cells could further differentiate into PDX1 and NKX6.1 positive pancreatic progenitors in vitro. We conclude that Activin A combined with FGF/MAPK signaling efficiently induce DE cells in the absence of serum. These findings improve our understanding of human endoderm formation, and constitute a step forward in the generation of clinical grade hESCs progenies for cell therapy.
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http://dx.doi.org/10.1016/j.bbrc.2012.08.098DOI Listing
September 2012

Establishment of hESC lines from the inner cell mass of blastocyst-stage embryos and single blastomeres of 4-cell stage embryos.

Methods Mol Biol 2012 ;873:81-112

Research Group Reproduction and Genetics, Vrije Universiteit Brussel, Brussels, Belgium.

More than 600 human embryonic stem cell (hESC) lines have been reported today at the human European Embryonic Stem Cell Registry ( http://www.hescreg.eu/ ). Despite these high numbers, there are currently no general protocols for derivation, culture, and characterization of hESC. Moreover, data on the culture of the embryo used for the derivation (medium, day of ICM isolation) are usually not available but can have an impact on the derivation rate. We present here the protocols for derivation, culture and characterization as we applied them for the 22 hESC lines (named VUB-hESC) in our laboratory.
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http://dx.doi.org/10.1007/978-1-61779-794-1_6DOI Listing
August 2012

Transplantation of human embryonic stem cell-derived pancreatic endoderm reveals a site-specific survival, growth, and differentiation.

Cell Transplant 2013 ;22(5):821-30

Cell Differentiation Unit, Diabetes Research Center, Vrije Universiteit Brussel, Brussels, Belgium.

Development of β-cells from human embryonic stem cells (hESCs) could compensate for the shortage of islet donors required for diabetes therapy. Although pancreatic progenitors have been derived from hESCs using various protocols, no fully functional b-cells could be generated in vitro. We evaluated the in vivo growth and differentiation of PDX1+ pancreatic endoderm cells obtained from hESCs. Here we show site-specific survival and differentiation when comparing cells grafted in the epididymal fat pad or the subcutaneous space of NOD/SCID mice after 12 weeks follow-up. Subcutaneous grafts persisted and expressed PDX1 at all time points analyzed, showed PDX1 and NKX6.1 coexpression after 6 weeks, and contained NGN3+ cells after 12 weeks.These findings suggest that further specification along the pancreatic lineage occured at the subcutaneous site.In sharp contrast, in the fat pad grafts only a minority of PDX1+ cells remained after 2 weeks, and no further pancreatic differentiation was observed later on. In addition, contaminating mesenchymal cells present in the implants further developed into cartilage tissue after 6 weeks implantation in the fat pad, but not in the subcutaneous space. These findings indicate that the in vivo microenvironment plays a critical role in the further differentiation of transplanted pancreatic endoderm cells.
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http://dx.doi.org/10.3727/096368912X636812DOI Listing
December 2013

A specific increase in inositol 1,4,5-trisphosphate 3-kinase B expression upon differentiation of human embryonic stem cells.

Cell Signal 2012 Jul 13;24(7):1461-70. Epub 2012 Mar 13.

Research Laboratory on Human Reproduction, Université Libre de Bruxelles, Brussels, Belgium.

Human embryonic stem cells (hESCs) are of great hope for regenerative medicine due to their dual pluripotency and self-renewal properties. We report a comparison of inositol phosphate (InsP(s)) production in undifferentiated, differentiated hESCs and in two cancer cell lines, Ntera2 cells, a human embryonal carcinoma cell (hECC) line and HeLa cells. To evaluate the potential impact of InsP(s) in differentiation, hESCs were spontaneously differentiated in culture for two weeks. The distribution of the different InsP(s) was affected upon differentiation: the level of highly phosphorylated InsP(s) was decreased. In contrast, the total level of phosphoinositides (PI) was increased. Using real time quantitative PCR (qPCR), the mRNA expression of several enzymes of the metabolism of InsP(s) was determined: a specific increase in inositol 1,4,5-trisphosphate 3-kinase A and B (ITPKA and ITPKB) was observed upon hESCs spontaneous differentiation. Ins(1,4,5)P(3) 3-kinase activity, undetectable in undifferentiated hESCs, increased upon differentiation. The same observation was made by Western blotting using an antibody directed against human ITPKB. This is the first report showing the potential implication of soluble InsP(s) in hESCs and possible function of isoenzymes of the inositol trisphosphate 3-kinase family in differentiation.
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http://dx.doi.org/10.1016/j.cellsig.2012.03.006DOI Listing
July 2012

Screening ethnically diverse human embryonic stem cells identifies a chromosome 20 minimal amplicon conferring growth advantage.

Authors:
Katherine Amps Peter W Andrews George Anyfantis Lyle Armstrong Stuart Avery Hossein Baharvand Julie Baker Duncan Baker Maria B Munoz Stephen Beil Nissim Benvenisty Dalit Ben-Yosef Juan-Carlos Biancotti Alexis Bosman Romulo Martin Brena Daniel Brison Gunilla Caisander María V Camarasa Jieming Chen Eric Chiao Young Min Choi Andre B H Choo Daniel Collins Alan Colman Jeremy M Crook George Q Daley Anne Dalton Paul A De Sousa Chris Denning Janet Downie Petr Dvorak Karen D Montgomery Anis Feki Angela Ford Victoria Fox Ana M Fraga Tzvia Frumkin Lin Ge Paul J Gokhale Tamar Golan-Lev Hamid Gourabi Michal Gropp Guangxiu Lu Ales Hampl Katie Harron Lyn Healy Wishva Herath Frida Holm Outi Hovatta Johan Hyllner Maneesha S Inamdar Astrid Kresentia Irwanto Tetsuya Ishii Marisa Jaconi Ying Jin Susan Kimber Sergey Kiselev Barbara B Knowles Oded Kopper Valeri Kukharenko Anver Kuliev Maria A Lagarkova Peter W Laird Majlinda Lako Andrew L Laslett Neta Lavon Dong Ryul Lee Jeoung Eun Lee Chunliang Li Linda S Lim Tenneille E Ludwig Yu Ma Edna Maltby Ileana Mateizel Yoav Mayshar Maria Mileikovsky Stephen L Minger Takamichi Miyazaki Shin Yong Moon Harry Moore Christine Mummery Andras Nagy Norio Nakatsuji Kavita Narwani Steve K W Oh Sun Kyung Oh Cia Olson Timo Otonkoski Fei Pan In-Hyun Park Steve Pells Martin F Pera Lygia V Pereira Ouyang Qi Grace Selva Raj Benjamin Reubinoff Alan Robins Paul Robson Janet Rossant Ghasem H Salekdeh Thomas C Schulz Karen Sermon Jameelah Sheik Mohamed Hui Shen Eric Sherrer Kuldip Sidhu Shirani Sivarajah Heli Skottman Claudia Spits Glyn N Stacey Raimund Strehl Nick Strelchenko Hirofumi Suemori Bowen Sun Riitta Suuronen Kazutoshi Takahashi Timo Tuuri Parvathy Venu Yuri Verlinsky Dorien Ward-van Oostwaard Daniel J Weisenberger Yue Wu Shinya Yamanaka Lorraine Young Qi Zhou

Nat Biotechnol 2011 Nov 27;29(12):1132-44. Epub 2011 Nov 27.

Centre for Stem Cell Biology, Department of Biomedical Science, The University of Sheffield, Sheffield, UK.

The International Stem Cell Initiative analyzed 125 human embryonic stem (ES) cell lines and 11 induced pluripotent stem (iPS) cell lines, from 38 laboratories worldwide, for genetic changes occurring during culture. Most lines were analyzed at an early and late passage. Single-nucleotide polymorphism (SNP) analysis revealed that they included representatives of most major ethnic groups. Most lines remained karyotypically normal, but there was a progressive tendency to acquire changes on prolonged culture, commonly affecting chromosomes 1, 12, 17 and 20. DNA methylation patterns changed haphazardly with no link to time in culture. Structural variants, determined from the SNP arrays, also appeared sporadically. No common variants related to culture were observed on chromosomes 1, 12 and 17, but a minimal amplicon in chromosome 20q11.21, including three genes expressed in human ES cells, ID1, BCL2L1 and HM13, occurred in >20% of the lines. Of these genes, BCL2L1 is a strong candidate for driving culture adaptation of ES cells.
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http://dx.doi.org/10.1038/nbt.2051DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3454460PMC
November 2011

Current controversies in prenatal diagnosis 1: is stem cell therapy ready for human fetuses?

Prenat Diagn 2011 Mar 3;31(3):228-30. Epub 2011 Jan 3.

Department of Anatomy and Embryology, Leiden University Medical Center, Leiden, The Netherlands.

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http://dx.doi.org/10.1002/pd.2657DOI Listing
March 2011