Publications by authors named "Mary Herbert"

54 Publications

Deprotection of centromeric cohesin at meiosis II requires APC/C activity but not kinetochore tension.

EMBO J 2021 Apr 1;40(7):e106812. Epub 2021 Mar 1.

Laboratory of Chromosome Biology, Max Planck Institute of Biochemistry, Martinsried, Germany.

Genome haploidization involves sequential loss of cohesin from chromosome arms and centromeres during two meiotic divisions. At centromeres, cohesin's Rec8 subunit is protected from separase cleavage at meiosis I and then deprotected to allow its cleavage at meiosis II. Protection of centromeric cohesin by shugoshin-PP2A seems evolutionarily conserved. However, deprotection has been proposed to rely on spindle forces separating the Rec8 protector from cohesin at metaphase II in mammalian oocytes and on APC/C-dependent destruction of the protector at anaphase II in yeast. Here, we have activated APC/C in the absence of sister kinetochore biorientation at meiosis II in yeast and mouse oocytes, and find that bipolar spindle forces are dispensable for sister centromere separation in both systems. Furthermore, we show that at least in yeast, protection of Rec8 by shugoshin and inhibition of separase by securin are both required for the stability of centromeric cohesin at metaphase II. Our data imply that related mechanisms preserve the integrity of dyad chromosomes during the short metaphase II of yeast and the prolonged metaphase II arrest of mammalian oocytes.
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http://dx.doi.org/10.15252/embj.2020106812DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8013787PMC
April 2021

Mitochondrial genome editing gets precise.

Nature 2020 07;583(7817):521-522

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http://dx.doi.org/10.1038/d41586-020-01974-6DOI Listing
July 2020

Prc1-rich kinetochores are required for error-free acentrosomal spindle bipolarization during meiosis I in mouse oocytes.

Nat Commun 2020 05 27;11(1):2652. Epub 2020 May 27.

Laboratory for Chromosome Segregation, RIKEN Center for Biosystems Dynamics Research (BDR), Kobe, 650-0047, Japan.

Acentrosomal meiosis in oocytes represents a gametogenic challenge, requiring spindle bipolarization without predefined bipolar cues. While much is known about the structures that promote acentrosomal microtubule nucleation, less is known about the structures that mediate spindle bipolarization in mammalian oocytes. Here, we show that in mouse oocytes, kinetochores are required for spindle bipolarization in meiosis I. This process is promoted by oocyte-specific, microtubule-independent enrichment of the antiparallel microtubule crosslinker Prc1 at kinetochores via the Ndc80 complex. In contrast, in meiosis II, cytoplasm that contains upregulated factors including Prc1 supports kinetochore-independent pathways for spindle bipolarization. The kinetochore-dependent mode of spindle bipolarization is required for meiosis I to prevent chromosome segregation errors. Human oocytes, where spindle bipolarization is reportedly error prone, exhibit no detectable kinetochore enrichment of Prc1. This study reveals an oocyte-specific function of kinetochores in acentrosomal spindle bipolarization in mice, and provides insights into the error-prone nature of human oocytes.
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http://dx.doi.org/10.1038/s41467-020-16488-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7253481PMC
May 2020

Oocyte aneuploidy-more tools to tackle an old problem.

Proc Natl Acad Sci U S A 2020 06 19;117(22):11850-11852. Epub 2020 May 19.

Newcastle Fertility Centre, Biosciences Institute, Newcastle University, Newcastle upon Tyne NE1 4EP, United Kingdom

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http://dx.doi.org/10.1073/pnas.2005739117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7275742PMC
June 2020

Tex19.1 inhibits the N-end rule pathway and maintains acetylated SMC3 cohesin and sister chromatid cohesion in oocytes.

J Cell Biol 2020 05;219(5)

Medical Research Council Human Genetics Unit, Medical Research Council Institute of Genetics and Molecular Medicine, University of Edinburgh, Western General Hospital, Crewe Road, Edinburgh, UK.

Age-dependent oocyte aneuploidy, a major cause of Down syndrome, is associated with declining sister chromatid cohesion in postnatal oocytes. Here we show that cohesion in postnatal mouse oocytes is regulated by Tex19.1. We show Tex19.1-/- oocytes have defects maintaining chiasmata, missegregate their chromosomes during meiosis, and transmit aneuploidies to the next generation. Furthermore, we show that mouse Tex19.1 inhibits N-end rule protein degradation mediated by its interacting partner UBR2, and that Ubr2 itself has a previously undescribed role in negatively regulating the acetylated SMC3 subpopulation of cohesin in mitotic somatic cells. Lastly, we show that acetylated SMC3 is associated with meiotic chromosome axes in mouse oocytes, and that this population of cohesin is specifically depleted in the absence of Tex19.1. These findings indicate that Tex19.1 regulates UBR protein activity to maintain acetylated SMC3 and sister chromatid cohesion in postnatal oocytes and prevent aneuploidy from arising in the female germline.
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http://dx.doi.org/10.1083/jcb.201702123DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7199850PMC
May 2020

Reversion after replacement of mitochondrial DNA.

Nature 2019 10 16;574(7778):E8-E11. Epub 2019 Oct 16.

Wellcome Centre for Mitochondrial Research, Institute of Genetic Medicine, Newcastle University, Newcastle upon Tyne, UK.

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http://dx.doi.org/10.1038/s41586-019-1623-3DOI Listing
October 2019

Differentiation of Human Embryonic Stem Cells to Sympathetic Neurons: A Potential Model for Understanding Neuroblastoma Pathogenesis.

Stem Cells Int 2018 1;2018:4391641. Epub 2018 Nov 1.

Wolfson Childhood Cancer Research Centre, Northern Institute for Cancer Research, Newcastle University, UK.

Background And Aims: Previous studies modelling human neural crest differentiation from stem cells have resulted in a low yield of sympathetic neurons. Our aim was to optimise a method for the differentiation of human embryonic stem cells (hESCs) to sympathetic neuron-like cells (SN) to model normal human SNS development.

Results: Using stromal-derived inducing activity (SDIA) of PA6 cells plus BMP4 and B27 supplements, the H9 hESC line was differentiated to neural crest stem-like cells and SN-like cells. After 7 days of PA6 cell coculture, mRNA expression of and neural crest specifier genes and the neural marker () increased. Expression of the pluripotency marker decreased, whereas and expression remained high at levels similar to SHSY5Y and IMR32 neuroblastoma cell lines. A 5-fold increase in the expression of the catecholaminergic marker ) and the noradrenergic marker ) was observed by day 7 of differentiation. Fluorescence-activated cell sorting for the neural crest marker p75, enriched for cells expressing , , , and , was more specific than p75 neural crest stem cell (NCSC) microbeads. On day 28 post p75 sorting, dual immunofluorescence identified sympathetic neurons by PRPH and TH copositivity cells in 20% of the cell population. Noradrenergic sympathetic neurons, identified by copositivity for both PHOX2B and DBH, were present in 9.4% ± 5.5% of cells.

Conclusions: We have optimised a method for noradrenergic SNS development using the H9 hESC line to improve our understanding of normal human SNS development and, in a future work, the pathogenesis of neuroblastoma.
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http://dx.doi.org/10.1155/2018/4391641DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6236576PMC
November 2018

Assessing mitochondrial heteroplasmy using next generation sequencing: A note of caution.

Mitochondrion 2019 05 9;46:302-306. Epub 2018 Aug 9.

Institute of Genetic Medicine, International Centre for Life, Central Parkway, Newcastle upon Tyne NE1 3BZ, UK; The Wellcome Centre for Mitochondrial Research, Newcastle University, Medical School, Framlington Place, Newcastle upon Tyne NE2 4HH, UK. Electronic address:

The mitochondrial genome has recently become the focus of several high-impact next-generation sequencing studies investigating the effect of mutations in disease and assessing the efficacy of mitochondrial replacement therapies. However, these studies have failed to take into consideration the capture of recurring translocations of mitochondrial DNA to the nuclear genome, known as nuclear mitochondrial sequences (NUMTs), continuing to align sequence data to the revised Cambridge reference sequence alone. Here, using different mtDNA enrichment techniques and a variety of tissues, we demonstrate that NUMTs are present in sequence data and that, dependent upon downstream analysis, are at a level which affects variant calling.
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http://dx.doi.org/10.1016/j.mito.2018.08.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6509278PMC
May 2019

Mutation-specific effects in germline transmission of pathogenic mtDNA variants.

Hum Reprod 2018 07;33(7):1331-1341

Department of Genetics and Cell Biology, School for Oncology and Developmental Biology (GROW), Maastricht University, Maastricht, the Netherlands.

Study Question: Does germline selection (besides random genetic drift) play a role during the transmission of heteroplasmic pathogenic mitochondrial DNA (mtDNA) mutations in humans?

Summary Answer: We conclude that inheritance of mtDNA is mutation-specific and governed by a combination of random genetic drift and negative and/or positive selection.

What Is Known Already: mtDNA inherits maternally through a genetic bottleneck, but the underlying mechanisms are largely unknown. Although random genetic drift is recognized as an important mechanism, selection mechanisms are thought to play a role as well.

Study Design, Size, Duration: We determined the mtDNA mutation loads in 160 available oocytes, zygotes, and blastomeres of five carriers of the m.3243A>G mutation, one carrier of the m.8993T>G mutation, and one carrier of the m.14487T>C mutation.

Participants/materials, Setting, Methods: Mutation loads were determined in PGD samples using PCR assays and analysed mathematically to test for random sampling effects. In addition, a meta-analysis has been performed on mutation load transmission data in the literature to confirm the results of the PGD samples.

Main Results And The Role Of Chance: By applying the Kimura distribution, which assumes random mechanisms, we found that mtDNA segregations patterns could be explained by variable bottleneck sizes among all our carriers (moment estimates ranging from 10 to 145). Marked differences in the bottleneck size would determine the probability that a carrier produces offspring with mutations markedly different than her own. We investigated whether bottleneck sizes might also be influenced by non-random mechanisms. We noted a consistent absence of high mutation loads in all our m.3243A>G carriers, indicating non-random events. To test this, we fitted a standard and a truncated Kimura distribution to the m.3243A>G segregation data. A Kimura distribution truncated at 76.5% heteroplasmy has a significantly better fit (P-value = 0.005) than the standard Kimura distribution. For the m.8993T>G mutation, we suspect a skewed mutation load distribution in the offspring. To test this hypothesis, we performed a meta-analysis on published blood mutation levels of offspring-mother (O-M) transmission for the m.3243A>G and m.8993T>G mutations. This analysis revealed some evidence that the O-M ratios for the m.8993T>G mutation are different from zero (P-value <0.001), while for the m.3243A>G mutation there was little evidence that the O-M ratios are non-zero. Lastly, for the m.14487T>G mutation, where the whole range of mutation loads was represented, we found no indications for selective events during its transmission.

Large Scale Data: All data are included in the Results section of this article.

Limitations, Reason For Caution: The availability of human material for the mutations is scarce, requiring additional samples to confirm our findings.

Wider Implications Of The Findings: Our data show that non-random mechanisms are involved during mtDNA segregation. We aimed to provide the mechanisms underlying these selection events. One explanation for selection against high m.3243A>G mutation loads could be, as previously reported, a pronounced oxidative phosphorylation (OXPHOS) deficiency at high mutation loads, which prohibits oogenesis (e.g. progression through meiosis). No maximum mutation loads of the m.8993T>G mutation seem to exist, as the OXPHOS deficiency is less severe, even at levels close to 100%. In contrast, high mutation loads seem to be favoured, probably because they lead to an increased mitochondrial membrane potential (MMP), a hallmark on which healthy mitochondria are being selected. This hypothesis could provide a possible explanation for the skewed segregation pattern observed. Our findings are corroborated by the segregation pattern of the m.14487T>C mutation, which does not affect OXPHOS and MMP significantly, and its transmission is therefore predominantly determined by random genetic drift. Our conclusion is that mutation-specific selection mechanisms occur during mtDNA inheritance, which has implications for PGD and mitochondrial replacement therapy.

Study Funding/competing Interest(s): This work has been funded by GROW-School of Oncology and Developmental Biology. The authors declare no competing interests.
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http://dx.doi.org/10.1093/humrep/dey114DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6551225PMC
July 2018

Progress in mitochondrial replacement therapies.

Nat Rev Mol Cell Biol 2018 01;19(2):71-72

Wellcome Centre for Mitochondrial Research, Institute of Genetic Medicine and Institute of Neurosciences, Newcastle University, Newcastle upon Tyne, UK.

Mitochondrial DNA is maternally inherited, and pathogenic mutations cause a range of life-limiting conditions. Recent studies indicate that transmission of pathogenic mutations may be prevented by reproductive technologies designed to replace the mitochondria in eggs from affected women.
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http://dx.doi.org/10.1038/nrm.2018.3DOI Listing
January 2018

APC/C Enables Removal of Shugoshin-2 from the Arms of Bivalent Chromosomes by Moderating Cyclin-Dependent Kinase Activity.

Curr Biol 2017 May 11;27(10):1462-1476.e5. Epub 2017 May 11.

Department of Biochemistry, University of Oxford, South Parks Road, Oxford OX1 3QU, UK. Electronic address:

In mammalian females, germ cells remain arrested as primordial follicles. Resumption of meiosis is heralded by germinal vesicle breakdown, condensation of chromosomes, and their eventual alignment on metaphase plates. At the first meiotic division, anaphase-promoting complex/cyclosome associated with Cdc20 (APC/C) activates separase and thereby destroys cohesion along chromosome arms. Because cohesion around centromeres is protected by shugoshin-2, sister chromatids remain attached through centromeric/pericentromeric cohesin. We show here that, by promoting proteolysis of cyclins and Cdc25B at the germinal vesicle (GV) stage, APC/C associated with the Cdh1 protein (APC/C) delays the increase in Cdk1 activity, leading to germinal vesicle breakdown (GVBD). More surprisingly, by moderating the rate at which Cdk1 is activated following GVBD, APC/C creates conditions necessary for the removal of shugoshin-2 from chromosome arms by the Aurora B/C kinase, an event crucial for the efficient resolution of chiasmata.
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http://dx.doi.org/10.1016/j.cub.2017.04.023DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5457479PMC
May 2017

How Meiosis Creates the Single-Copy Genome.

Dev Cell 2017 01;40(1):3-4

Institute of Physiological Chemistry, Medical Faculty of TU Dresden, Fiedlerstrasse 42, 01307 Dresden, Germany. Electronic address:

Genome haploidization involves two meiotic divisions following a single round of DNA replication. In this issue of Developmental Cell, Argüello-Miranda et al. (2017) show that production and packaging of the single-copy genome into gametes during the second meiotic division is coordinated by a conserved casein kinase 1.
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http://dx.doi.org/10.1016/j.devcel.2016.12.019DOI Listing
January 2017

Mitochondrial Donation - Clearing the Final Regulatory Hurdle in the United Kingdom.

N Engl J Med 2017 01 28;376(2):171-173. Epub 2016 Dec 28.

From the Wellcome Centre for Mitochondrial Research (M.H., D.T.) and the Newcastle Fertility Centre (M.H.) - both in Newcastle, United Kingdom.

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http://dx.doi.org/10.1056/NEJMcibr1615669DOI Listing
January 2017

Towards clinical application of pronuclear transfer to prevent mitochondrial DNA disease.

Nature 2016 06 8;534(7607):383-6. Epub 2016 Jun 8.

Wellcome Trust Centre for Mitochondrial Research, Institute of Genetic Medicine, Biomedicine West Wing, Centre for Life, Times Square, Newcastle upon Tyne NE1 3BZ, UK.

Mitochondrial DNA (mtDNA) mutations are maternally inherited and are associated with a broad range of debilitating and fatal diseases. Reproductive technologies designed to uncouple the inheritance of mtDNA from nuclear DNA may enable affected women to have a genetically related child with a greatly reduced risk of mtDNA disease. Here we report the first preclinical studies on pronuclear transplantation (PNT). Surprisingly, techniques used in proof-of-concept studies involving abnormally fertilized human zygotes were not well tolerated by normally fertilized zygotes. We have therefore developed an alternative approach based on transplanting pronuclei shortly after completion of meiosis rather than shortly before the first mitotic division. This promotes efficient development to the blastocyst stage with no detectable effect on aneuploidy or gene expression. After optimization, mtDNA carryover was reduced to <2% in the majority (79%) of PNT blastocysts. The importance of reducing carryover to the lowest possible levels is highlighted by a progressive increase in heteroplasmy in a stem cell line derived from a PNT blastocyst with 4% mtDNA carryover. We conclude that PNT has the potential to reduce the risk of mtDNA disease, but it may not guarantee prevention.
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http://dx.doi.org/10.1038/nature18303DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5131843PMC
June 2016

Understanding the dispensary workflow at the Birmingham Free Clinic: a proposed framework for an informatics intervention.

BMC Health Serv Res 2016 Feb 19;16:69. Epub 2016 Feb 19.

Center for Health Informatics for the Underserved, Department of Biomedical Informatics, School of Medicine, University of Pittsburgh, Pittsburgh, PA, USA.

Background: The Birmingham Free Clinic (BFC) in Pittsburgh, Pennsylvania, USA is a free, walk-in clinic that serves medically uninsured populations through the use of volunteer health care providers and an on-site medication dispensary. The introduction of an electronic medical record (EMR) has improved several aspects of clinic workflow. However, pharmacists' tasks involving medication management and dispensing have become more challenging since EMR implementation due to its inability to support workflows between the medical and pharmaceutical services. To inform the design of a systematic intervention, we conducted a needs assessment study to identify workflow challenges and process inefficiencies in the dispensary.

Methods: We used contextual inquiry to document the dispensary workflow and facilitate identification of critical aspects of intervention design specific to the user. Pharmacists were observed according to contextual inquiry guidelines. Graphical models were produced to aid data and process visualization. We created a list of themes describing workflow challenges and asked the pharmacists to rank them in order of significance to narrow the scope of intervention design.

Results: Three pharmacists were observed at the BFC. Observer notes were documented and analyzed to produce 13 themes outlining the primary challenges pharmacists encounter during dispensation at the BFC. The dispensary workflow is labor intensive, redundant, and inefficient when integrated with the clinical service. Observations identified inefficiencies that may benefit from the introduction of informatics interventions including: medication labeling, insufficient process notification, triple documentation, and inventory control.

Conclusions: We propose a system for Prescription Management and General Inventory Control (RxMAGIC). RxMAGIC is a framework designed to mitigate workflow challenges and improve the processes of medication management and inventory control. While RxMAGIC is described in the context of the BFC dispensary, we believe it will be generalizable to pharmacies in other low-resource settings, both domestically and internationally.
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http://dx.doi.org/10.1186/s12913-016-1308-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4759722PMC
February 2016

Research into Policy: A Brief History of Mitochondrial Donation.

Stem Cells 2016 Feb 27;34(2):265-7. Epub 2015 Oct 27.

Wellcome Trust Centre for Mitochondrial Research, Institute of Neuroscience, Newcastle University, Newcastle upon Tyne, United Kingdom.

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http://dx.doi.org/10.1002/stem.2221DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4855617PMC
February 2016

Extreme-Depth Re-sequencing of Mitochondrial DNA Finds No Evidence of Paternal Transmission in Humans.

PLoS Genet 2015 May 14;11(5):e1005040. Epub 2015 May 14.

Wellcome Trust Centre for Mitochondrial Research, Newcastle University, Newcastle-upon-Tyne, United Kingdom; Institute of Genetic Medicine, Newcastle University, Newcastle-upon-Tyne, United Kingdom.

Recent reports have questioned the accepted dogma that mammalian mitochondrial DNA (mtDNA) is strictly maternally inherited. In humans, the argument hinges on detecting a signature of inter-molecular recombination in mtDNA sequences sampled at the population level, inferring a paternal source for the mixed haplotypes. However, interpreting these data is fraught with difficulty, and direct experimental evidence is lacking. Using extreme-high depth mtDNA re-sequencing up to ~1.2 million-fold coverage, we find no evidence that paternal mtDNA haplotypes are transmitted to offspring in humans, thus excluding a simple dilution mechanism for uniparental transmission of mtDNA present in all healthy individuals. Our findings indicate that an active mechanism eliminates paternal mtDNA which likely acts at the molecular level.
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http://dx.doi.org/10.1371/journal.pgen.1005040DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4431825PMC
May 2015

Mitochondrial replacement to prevent the transmission of mitochondrial DNA disease.

EMBO Rep 2015 May 16;16(5):539-40. Epub 2015 Apr 16.

Wellcome Trust Centre for Mitochondrial Research, Institute of Neuroscience, Newcastle University, Newcastle upon Tyne, UK.

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http://dx.doi.org/10.15252/embr.201540354DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4428042PMC
May 2015

Meiosis and maternal aging: insights from aneuploid oocytes and trisomy births.

Cold Spring Harb Perspect Biol 2015 Apr 1;7(4):a017970. Epub 2015 Apr 1.

Wellcome Trust Centre for Mitochondrial Research, Institute of Genetic Medicine, Newcastle University, Newcastle upon Tyne NE1 3BZ, United Kingdom.

In most organisms, genome haploidization requires reciprocal DNA exchanges (crossovers) between replicated parental homologs to form bivalent chromosomes. These are resolved to their four constituent chromatids during two meiotic divisions. In female mammals, bivalents are formed during fetal life and remain intact until shortly before ovulation. Extending this period beyond ∼35 years greatly increases the risk of aneuploidy in human oocytes, resulting in a dramatic increase in infertility, miscarriage, and birth defects, most notably trisomy 21. Bivalent chromosomes are stabilized by cohesion between sister chromatids, which is mediated by the cohesin complex. In mouse oocytes, cohesin becomes depleted from chromosomes during female aging. Consistent with this, premature loss of centromeric cohesion is a major source of aneuploidy in oocytes from older women. Here, we propose a mechanistic framework to reconcile data from genetic studies on human trisomy and oocytes with recent advances in our understanding of the molecular mechanisms of chromosome segregation during meiosis in model organisms.
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http://dx.doi.org/10.1101/cshperspect.a017970DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4382745PMC
April 2015

Concise reviews: Assisted reproductive technologies to prevent transmission of mitochondrial DNA disease.

Stem Cells 2015 Mar;33(3):639-45

Wellcome Trust Centre for Mitochondrial Research, Newcastle University, Newcastle upon Tyne, United Kingdom; Institute of Genetic Medicine, Newcastle University, Newcastle upon Tyne, United Kingdom.

While the fertilized egg inherits its nuclear DNA from both parents, the mitochondrial DNA is strictly maternally inherited. Cells contain multiple copies of mtDNA, each of which encodes 37 genes, which are essential for energy production by oxidative phosphorylation. Mutations can be present in all, or only in some copies of mtDNA. If present above a certain threshold, pathogenic mtDNA mutations can cause a range of debilitating and fatal diseases. Here, we provide an update of currently available options and new techniques under development to reduce the risk of transmitting mtDNA disease from mother to child. Preimplantation genetic diagnosis (PGD), a commonly used technique to detect mutations in nuclear DNA, is currently being offered to determine the mutation load of embryos produced by women who carry mtDNA mutations. The available evidence indicates that cells removed from an eight-cell embryo are predictive of the mutation load in the entire embryo, indicating that PGD provides an effective risk reduction strategy for women who produce embryos with low mutation loads. For those who do not, research is now focused on meiotic nuclear transplantation techniques to uncouple the inheritance of nuclear and mtDNA. These approaches include transplantation of any one of the products or female meiosis (meiosis II spindle, or either of the polar bodies) between oocytes, or the transplantation of pronuclei between fertilized eggs. In all cases, the transferred genetic material arises from a normal meiosis and should therefore, not be confused with cloning. The scientific progress and associated regulatory issues are discussed.
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http://dx.doi.org/10.1002/stem.1887DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4359624PMC
March 2015

Thyroid, typhoid.

Chest 2014 May;145(5):1171

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http://dx.doi.org/10.1378/chest.13-2221DOI Listing
May 2014

The challenges of mitochondrial replacement.

PLoS Genet 2014 Apr 24;10(4):e1004315. Epub 2014 Apr 24.

Wellcome Trust Centre for Mitochondrial Research, Institute for Ageing and Health, Newcastle University, Newcastle upon Tyne, United Kingdom.

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http://dx.doi.org/10.1371/journal.pgen.1004315DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3998882PMC
April 2014

Focusing on the five A's: a comparison of homeless and housed patients' access to and use of pharmacist-provided smoking cessation treatment.

Res Social Adm Pharm 2014 Mar-Apr;10(2):369-77

Division of General Internal Medicine, UPMC Montefiore Hospital, USA.

Introduction: The prevalence of smoking remains high among the medically underserved and could be related to disparities in access to and use of smoking cessation treatments.

Methods: This study implemented and tracked providers' use of the 5 A's intervention for tobacco use (Ask, Assess, Advise, Assist, Arrange) with homeless (n = 260) and housed (n = 226) adults attending a free medical clinic, including referrals to and use of an on-site pharmacist-led smoking cessation service.

Results: Among patients whose tobacco use was Asked about and Assessed (97%), homeless (vs. housed) patients were more likely to smoke (59% vs. 39%; P = 0.008). Among current smokers, there were no homeless-housed disparities in receipt of Advice to quit smoking (84% vs. 78%; P = 0.22) or Arrangement of treatment (36% vs. 31%; P = 0.46). Overall, among patients for whom treatment was Arranged, homeless patients were less likely than housed patients to attend the smoking cessation program (25% vs. 48%; P = 0.04). However, among those that attended any treatment (i.e., were Assisted to quit), homeless and housed patients attended similar numbers of sessions and used pharmacotherapy at similar rates.

Conclusions: Providers may reduce homeless-housed disparities in smoking by offering special Assist(ance) to homeless smokers that reduces barriers to initially accessing treatment services.
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http://dx.doi.org/10.1016/j.sapharm.2013.05.011DOI Listing
November 2014

Therapeutic potential of somatic cell nuclear transfer for degenerative disease caused by mitochondrial DNA mutations.

Sci Rep 2014 Jan 24;4:3844. Epub 2014 Jan 24.

1] Wellcome Centre for Mitochondrial Research, Institute for Ageing and Health, Newcastle University, Newcastle upon Tyne, UK [2] Newcastle Fertility Centre, Centre for Life, Times Square, Newcastle upon Tyne, UK.

Induced pluripotent stem cells (iPSCs) hold much promise in the quest for personalised cell therapies. However, the persistence of founder cell mitochondrial DNA (mtDNA) mutations limits the potential of iPSCs in the development of treatments for mtDNA disease. This problem may be overcome by using oocytes containing healthy mtDNA, to induce somatic cell nuclear reprogramming. However, the extent to which somatic cell mtDNA persists following fusion with human oocytes is unknown. Here we show that human nuclear transfer (NT) embryos contain very low levels of somatic cell mtDNA. In light of a recent report that embryonic stem cells can be derived from human NT embryos, our results highlight the therapeutic potential of NT for mtDNA disease, and underscore the importance of using human oocytes to pursue this goal.
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http://dx.doi.org/10.1038/srep03844DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5379195PMC
January 2014

Nucleosome assembly proteins get SET to defeat the guardian of chromosome cohesion.

PLoS Genet 2013 26;9(9):e1003829. Epub 2013 Sep 26.

Division of Rheumatology, Immunology and Allergy, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts, United States of America.

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http://dx.doi.org/10.1371/journal.pgen.1003829DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3784508PMC
March 2014

Cyclin A2 is required for sister chromatid segregation, but not separase control, in mouse oocyte meiosis.

Cell Rep 2012 Nov 1;2(5):1077-87. Epub 2012 Nov 1.

UPMC Université Paris 06, UMR7622 Laboratoire de Biologie du Développement, 9 quai St. Bernard, Paris 75005, France.

In meiosis, two specialized cell divisions allow the separation of paired chromosomes first, then of sister chromatids. Separase removes the cohesin complex holding sister chromatids together in a stepwise manner from chromosome arms in meiosis I, then from the centromere region in meiosis II. Using mouse oocytes, our study reveals that cyclin A2 promotes entry into meiosis, as well as an additional unexpected role; namely, its requirement for separase-dependent sister chromatid separation in meiosis II. Untimely cyclin A2-associated kinase activity in meiosis I leads to precocious sister separation, whereas inhibition of cyclin A2 in meiosis II prevents it. Accordingly, endogenous cyclin A is localized to kinetochores throughout meiosis II, but not in anaphase I. Additionally, we found that cyclin B1, but not cyclin A2, inhibits separase in meiosis I. These findings indicate that separase-dependent cohesin removal is differentially regulated by cyclin B1 and A2 in mammalian meiosis.
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http://dx.doi.org/10.1016/j.celrep.2012.10.002DOI Listing
November 2012

Production and validation of a good manufacturing practice grade human fibroblast line for supporting human embryonic stem cell derivation and culture.

Stem Cell Res Ther 2012 Mar 28;3(2):12. Epub 2012 Mar 28.

NorthEast England Stem Cell Institute, Centre for Life, Times Square, Newcastle upon Tyne NE1 4EP, UK.

Introduction: The development of reproducible methods for deriving human embryonic stem cell (hESC) lines in compliance with good manufacturing practice (GMP) is essential for the development of hESC-based therapies. Although significant progress has been made toward the development of chemically defined conditions for the maintenance and differentiation of hESCs, efficient derivation of new hESCs requires the use of fibroblast feeder cells. However, GMP-grade feeder cell lines validated for hESC derivation are not readily available.

Methods: We derived a fibroblast cell line (NclFed1A) from human foreskin in compliance with GMP standards. Consent was obtained to use the cells for the production of hESCs and to generate induced pluripotent stem cells (iPSCs). We compared the line with a variety of other cell lines for its ability to support derivation and self-renewal of hESCs.

Results: NclFed1A supports efficient rates (33%) of hESC colony formation after explantation of the inner cell mass (ICM) of human blastocysts. This compared favorably with two mouse embryonic fibroblast (MEF) cell lines. NclFed1A also compared favorably with commercially available foreskin fibroblasts and MEFs in promoting proliferation and pluripotency of a number of existing and widely used hESCs. The ability of NclFed1A to maintain self-renewal remained undiminished for up to 28 population doublings from the master cell bank.

Conclusions: The human fibroblast line Ncl1Fed1A, produced in compliance with GMP standards and qualified for derivation and maintenance of hESCs, is a useful resource for the advancement of progress toward hESC-based therapies in regenerative medicine.
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http://dx.doi.org/10.1186/scrt103DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3392772PMC
March 2012

A novel isolator-based system promotes viability of human embryos during laboratory processing.

PLoS One 2012 29;7(2):e31010. Epub 2012 Feb 29.

Newcastle Fertility Centre, Newcastle upon Tyne, England, United Kingdom.

In vitro fertilisation (IVF) and related technologies are arguably the most challenging of all cell culture applications. The starting material is a single cell from which one aims to produce an embryo capable of establishing a pregnancy eventually leading to a live birth. Laboratory processing during IVF treatment requires open manipulations of gametes and embryos, which typically involves exposure to ambient conditions. To reduce the risk of cellular stress, we have developed a totally enclosed system of interlinked isolator-based workstations designed to maintain oocytes and embryos in a physiological environment throughout the IVF process. Comparison of clinical and laboratory data before and after the introduction of the new system revealed that significantly more embryos developed to the blastocyst stage in the enclosed isolator-based system compared with conventional open-fronted laminar flow hoods. Moreover, blastocysts produced in the isolator-based system contained significantly more cells and their development was accelerated. Consistent with this, the introduction of the enclosed system was accompanied by a significant increase in the clinical pregnancy rate and in the proportion of embryos implanting following transfer to the uterus. The data indicate that protection from ambient conditions promotes improved development of human embryos. Importantly, we found that it was entirely feasible to conduct all IVF-related procedures in the isolator-based workstations.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0031010PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3290619PMC
July 2012