Deletion of the Polycomb-Group Protein EZH2 Leads to Compromised Self-Renewal and Differentiation Defects in Human Embryonic Stem Cells

Through the histone methyltransferase EZH2, the Polycomb complex PRC2 mediates H3K27me3 and is associated with transcriptional repression. PRC2 regulates cell-fate decisions in model organisms; however, its role in regulating cell differentiation during human embryogenesis is unknown. Here, we report the characterization of $\small \textit{EZH2}$-deficient human embryonic stem cells (hESCs). H3K27me3 was lost upon $\small \textit{EZH2}$ deletion, identifying an essential requirement for EZH2 in methylating H3K27 in hESCs, in contrast to its non-essential role in mouse ESCs. Developmental regulators were derepressed in $\small \textit{EZH2}$-deficient hESCs, and single-cell analysis revealed an unexpected acquisition of lineage-restricted transcriptional programs. $\small \textit{EZH2}$-deficient hESCs show strongly reduced self-renewal and proliferation, thereby identifying a more severe phenotype compared to mouse ESCs. $\small \textit{EZH2}$-deficient hESCs can initiate differentiation toward developmental lineages; however, they cannot fully differentiate into mature specialized tissues. Thus, $\small \textit{EZH2}$ is required for stable ESC self-renewal, regulation of transcriptional programs, and for late-stage differentiation in this model of early human development.Wellcome Trust (Grant ID: WT093736), Biotechnology and Biological Sciences Research Council (Grant ID: BBS/E/B/000C0402), Medical Research Council (DTG Studentships, Grant ID: MR/J003808/1

Neuromyelitis optica spectrum disorder related tonic spasms responsive to lacosamide

Paroxysmal tonic spasms [PTS] are common in patients with neuromyelitis optica spectrum disorder (NMOSD).1 2 In patients with demyelinating disease, PTS can significantly reduce the quality of life, limit activities of daily living and the rehabilitative process following an acute relapse 3. As in patients with multiple sclerosis (MS), paroxysmal tonic spasms in NMOSD usually respond well to treatment with carbamazepine.2 However, the optimal treatment in patients where carbamazepine is contraindicated or poorly tolerated is unclear. We describe a patient with NMOSD with severe paroxysmal tonic spasms who did not tolerate carbamazepine but was successfully treated with lacosamide (Vimpat)

Molecular profiling of aged neural progenitors identifies Dbx2 as a candidate regulator of age-associated neurogenic decline.

Adult neurogenesis declines with aging due to the depletion and functional impairment of neural stem/progenitor cells (NSPCs). An improved understanding of the underlying mechanisms that drive age-associated neurogenic deficiency could lead to the development of strategies to alleviate cognitive impairment and facilitate neuroregeneration. An essential step towards this aim is to investigate the molecular changes that occur in NSPC aging on a genomewide scale. In this study, we compare the transcriptional, histone methylation and DNA methylation signatures of NSPCs derived from the subventricular zone (SVZ) of young adult (3 months old) and aged (18 months old) mice. Surprisingly, the transcriptional and epigenomic profiles of SVZ-derived NSPCs are largely unchanged in aged cells. Despite the global similarities, we detect robust age-dependent changes at several hundred genes and regulatory elements, thereby identifying putative regulators of neurogenic decline. Within this list, the homeobox gene Dbx2 is upregulated in vitro and in vivo, and its promoter region has altered histone and DNA methylation levels, in aged NSPCs. Using functional in vitro assays, we show that elevated Dbx2 expression in young adult NSPCs promotes age-related phenotypes, including the reduced proliferation of NSPC cultures and the altered transcript levels of age-associated regulators of NSPC proliferation and differentiation. Depleting Dbx2 in aged NSPCs caused the reverse gene expression changes. Taken together, these results provide new insights into the molecular programmes that are affected during mouse NSPC aging, and uncover a new functional role for Dbx2 in promoting age-related neurogenic decline.This work was supported by grants to P.J.R.-G. from the 6 Wellcome Trust (WT093736) and the BBSRC (BB/P013406/1, BB/M022285/1), by funding from 7 Sapienza University of Rome (G.L, S.B., E.C) and by a grant from the Spanish Ministry of 8 Economy to P.B. (BFU2016-75412-R, co-financed by FEDER). The Babraham Institute Biological 9 Services Unit is supported by Campus Capability Grant funding from the BBSRC

Interictal MEG abnormalities to guide intracranial electrode implantation and predict surgical outcome

Intracranial EEG (iEEG) is the gold standard technique for epileptogenic zone (EZ) localisation, but requires a hypothesis of which tissue is epileptogenic, guided by qualitative analysis of seizure semiology and other imaging modalities such as magnetoencephalography (MEG). We hypothesised that if quantifiable MEG band power abnormalities were sampled by iEEG, then patients' post-resection seizure outcome were better. Thirty-two individuals with neocortical epilepsy underwent MEG and iEEG recordings as part of pre-surgical evaluation. Interictal MEG band power abnormalities were derived using 70 healthy controls as a normative baseline. MEG abnormality maps were compared to electrode implantation, with the spatial overlap of iEEG electrodes and MEG abnormalities recorded. Finally, we assessed if the implantation of electrodes in abnormal tissue, and resection of the strongest abnormalities determined by MEG and iEEG explained surgical outcome. Intracranial electrodes were implanted in brain tissue with the most abnormal MEG findings in individuals that were seizure-free post-resection (T=3.9, p=0.003). The overlap between MEG abnormalities and iEEG electrodes distinguished outcome groups moderately well (AUC=0.68). In isolation, the resection of the strongest MEG and iEEG abnormalities separated surgical outcome groups well (AUC=0.71, AUC=0.74 respectively). A model incorporating all three features separated outcome groups best (AUC=0.80). Intracranial EEG is a key tool to delineate the EZ and help render patients seizure-free after resection. We showed that data-driven abnormalities derived from interictal MEG recordings have clinical value and may help guide electrode placement in individuals with neocortical epilepsy. Finally, our predictive model of post-operative seizure-freedom, which leverages both MEG and iEEG recordings, may aid patient counselling of expected outcome.Comment: 22 pages, 6 figure

Interictal magnetoencephalography abnormalities to guide intracranial electrode implantation and predict surgical outcome

Intracranial EEG is the gold standard technique for epileptogenic zone localization but requires a preconceived hypothesis of the location of the epileptogenic tissue. This placement is guided by qualitative interpretations of seizure semiology, MRI, EEG and other imaging modalities, such as magnetoencephalography. Quantitative abnormality mapping using magnetoencephalography has recently been shown to have potential clinical value. We hypothesized that if quantifiable magnetoencephalography abnormalities were sampled by intracranial EEG, then patients’ post-resection seizure outcome may be better. Thirty-two individuals with refractory neocortical epilepsy underwent magnetoencephalography and subsequent intracranial EEG recordings as part of presurgical evaluation. Eyes-closed resting-state interictal magnetoencephalography band power abnormality maps were derived from 70 healthy controls as a normative baseline. Magnetoencephalography abnormality maps were compared to intracranial EEG electrode implantation, with the spatial overlap of intracranial EEG electrode placement and cerebral magnetoencephalography abnormalities recorded. Finally, we assessed if the implantation of electrodes in abnormal tissue and subsequent resection of the strongest abnormalities determined by magnetoencephalography and intracranial EEG corresponded to surgical success. We used the area under the receiver operating characteristic curve as a measure of effect size. Intracranial electrodes were implanted in brain tissue with the most abnormal magnetoencephalography findings—in individuals that were seizure-free postoperatively (T = 3.9, P = 0.001) but not in those who did not become seizure-free. The overlap between magnetoencephalography abnormalities and electrode placement distinguished surgical outcome groups moderately well (area under the receiver operating characteristic curve = 0.68). In isolation, the resection of the strongest abnormalities as defined by magnetoencephalography and intracranial EEG separated surgical outcome groups well, area under the receiver operating characteristic curve = 0.71 and area under the receiver operating characteristic curve = 0.74, respectively. A model incorporating all three features separated surgical outcome groups best (area under the receiver operating characteristic curve = 0.80). Intracranial EEG is a key tool to delineate the epileptogenic zone and help render individuals seizure-free postoperatively. We showed that data-driven abnormality maps derived from resting-state magnetoencephalography recordings demonstrate clinical value and may help guide electrode placement in individuals with neocortical epilepsy. Additionally, our predictive model of postoperative seizure freedom, which leverages both magnetoencephalography and intracranial EEG recordings, could aid patient counselling of expected outcome

Critique of the review of 'Water fluoridation for the prevention of dental caries' published by the Cochrane Collaboration in 2015

The Cochrane Review on water fluoridation for the prevention of dental caries was published in 2015 and attracted considerable interest and comment, especially in countries with extensive water fluoridation programmes. The Review had two objectives: (i) to evaluate the effects of water fluoridation (artificial or natural) on the prevention of dental caries, and (ii) to evaluate the effects of water fluoridation (artificial or natural) on dental fluorosis. The authors concluded, inter alia, that there was very little contemporary evidence, meeting the Review's inclusion criteria, that evaluated the effectiveness of water fluoridation for the prevention of dental caries. The purpose of this critique is to examine the conduct of the above Review, and to put it into context in the wider body of evidence regarding the effectiveness of water fluoridation. While the overall conclusion that water fluoridation is effective in caries prevention agrees with previous reviews, many important public health questions could not be answered by the Review because of the restrictive criteria used to judge adequacy of study design and risk of bias. The potential benefits of using wider criteria in order to achieve a fuller understanding of the effectiveness of water fluoridation are discussed

Comprehensive Cell Surface Protein Profiling Identifies Specific Markers of Human Naive and Primed Pluripotent States

Human pluripotent stem cells (PSCs) exist in naive and primed states and provide important models to investigate the earliest stages of human development. Naive cells can be obtained through primed-to-naive resetting, but there are no reliable methods to prospectively isolate unmodified naive cells during this process. Here we report comprehensive profiling of cell surface proteins by flow cytometry in naive and primed human PSCs. Several naive-specific, but not primed-specific, proteins were also expressed by pluripotent cells in the human preimplantation embryo. The upregulation of naive-specific cell surface proteins during primed-to-naive resetting enabled the isolation and characterization of live naive cells and intermediate cell populations. This analysis revealed distinct transcriptional and X chromosome inactivation changes associated with the early and late stages of naive cell formation. Thus, identification of state-specific proteins provides a robust set of molecular markers to define the human PSC state and allows new insights into the molecular events leading to naive cell resetting.Imaging was performed at the Live Cell Imaging Facility/Nikon Center of Excellence, Department of Biosciences and Nutrition, Karolinska Institutet, Huddinge, Sweden, supported by grants from the Knut and Alice Wallenberg Foundation, the Swedish Research Council, the Centre for Innovative Medicine, and the Jonasson donation to the School of Technology and Health, Royal Institute of Technology, Sweden. We would like to acknowledge the MedH Flow Cytometry facility at Karolinska Institutet, supported by grants from Karolinska Institutet and the Stockholm County Council. We thank Céline Vallot and Claire Rougeulle at the Université Paris Diderot for providing X chromosome SNP coordinates. We are grateful to Rudolph Jaenisch at the Whitehead Institute for Biomedical Research for providing WIBR3 cells and Austin Smith at the WT–MRC Cambridge Stem Cell Institute for providing H9 NK2 and FiPS cells. We thank all couples who donated embryos to this study. S.P., A.P.R., J.P.S., and F.L. are supported by grants from the Swedish Research Council (2013-2570), Ragnar Söderberg Foundation (M67/13), Swedish Foundation for Strategic Research (ICA-5), Knut and Alice Wallenberg Foundation (4-1205/2016 and 4-148/2017), and Centre for Innovative Medicine and by a Lau fellowship. R.W. is an ISAC Shared Resource Laboratory Emerging Leader. A.J.C. is supported by an MRC DTG Studentship (MR/J003808/1). P.J.R.G. is supported by the Wellcome Trust (WT093736) and BBSRC (BBS/ E/B/000C0402)

Developmental Defects of Enamel in Primary Teeth and Association with Early Life Course Events: A Study of 6--36 Month old Children in Manyara, Tanzania.

Children with low birth weight show an increased prevalence of developmental defects of enamel in the primary dentition that subsequently may predispose to early childhood caries (ECC).Focusing 6--36 months old, the purpose of this study was to assess the frequency of enamel defects in the primary dentition and identify influences of early life course factors; socio-demographics, birth weight, child's early illness episodes and mothers' perceived size of the child at birth, whilst controlling for more recent life course events in terms of current breastfeeding and oral hygiene. A cross-sectional study was conducted in the high fluoride area of Manyara, northern Tanzania including 1221 child-mother pairs who attended Reproductive and Child Health (RCH) clinics for immunization and/or growth monitoring. After the primary caregivers had completed face to face interviews at the health care facility, children underwent oral clinical examination whereby ECC and developmental defects of enamel were recorded using field criteria. All erupted teeth were examined and the enamel defects were assessed on buccal surfaces according to the modified DDE Index. The prevalence of enamel defects was 33.3%. Diffuse opacities were the most common defects identified (23.1%), followed by hypoplasia (7.6%) and demarcated opacities (5.0%). The most frequently affected teeth were the upper central incisors (29.0% - 30.5%), whereas lower central incisors (4.3% to 4.5%) were least frequently affected. Multiple logistic regression analysis, adjusting for confounding the factors revealed that having normal birth weight (equal or more than 2500 g) associated with lower odds of having enamel hypoplasia [OR 0.22 (95% CI 0.1-0.7)]. No statistically significant association occurred between birth weight and diffuse opacities, demarcated opacities or combined DDE. Children with the history of low birth weight were more likely than their normal birth weight counterparts to present with enamel hypoplasia. In view of the frequent occurrence of enamel defects and the fact that hypoplasia may constitute a risk factor for future ECC, enamel defects should be included as a dental health indicator in epidemiological studies of children in northern Tanzania

A regression analysis of gene expression in ES cells reveals two gene classes that are significantly different in epigenetic patterns

<p>Abstract</p> <p>Background</p> <p>To understand the gene regulatory system that governs the self-renewal and pluripotency of embryonic stem cells (ESCs) is an important step for promoting regenerative medicine. In it, the role of several core transcription factors (TFs), such as Oct4, Sox2 and Nanog, has been intensively investigated, details of their involvement in the genome-wide gene regulation are still not well clarified.</p> <p>Methods</p> <p>We constructed a predictive model of genome-wide gene expression in mouse ESCs from publicly available ChIP-seq data of 12 core TFs. The tag sequences were remapped on the genome by various alignment tools. Then, the binding density of each TF is calculated from the genome-wide bona fide TF binding sites. The TF-binding data was combined with the data of several epigenetic states (DNA methylation, several histone modifications, and CpG island) of promoter regions. These data as well as the ordinary peak intensity data were used as predictors of a simple linear regression model that predicts absolute gene expression. We also developed a pipeline for analyzing the effects of predictors and their interactions.</p> <p>Results</p> <p>Through our analysis, we identified two classes of genes that are either well explained or inefficiently explained by our model. The latter class seems to be genes that are not directly regulated by the core TFs. The regulatory regions of these gene classes show apparently distinct patterns of DNA methylation, histone modifications, existence of CpG islands, and gene ontology terms, suggesting the relative importance of epigenetic effects. Furthermore, we identified statistically significant TF interactions correlated with the epigenetic modification patterns.</p> <p>Conclusions</p> <p>Here, we proposed an improved prediction method in explaining the ESC-specific gene expression. Our study implies that the majority of genes are more or less directly regulated by the core TFs. In addition, our result is consistent with the general idea of relative importance of epigenetic effects in ESCs.</p