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  • ItemOpen AccessPublished version Peer-reviewed
    Comparative omics and feeding manipulations in chicken indicate a shift of the endocrine role of visceral fat towards reproduction.
    (Springer nature, 2018-04-26) Bornelöv, Susanne; Seroussi, Eyal; Yosefi, Sara; Benjamini, Sharon; Miyara, Shoval; Ruzal, Mark; Grabherr, Manfred; Rafati, Nima; Molin, Anna-Maja; Pendavis, Ken; Burgess, Shane C; Andersson, Leif; Friedman-Einat, Miriam; Borneloev, Susanne [0000-0001-9276-9981]
    BACKGROUND: The mammalian adipose tissue plays a central role in energy-balance control, whereas the avian visceral fat hardly expresses leptin, the key adipokine in mammals. Therefore, to assess the endocrine role of adipose tissue in birds, we compared the transcriptome and proteome between two metabolically different types of chickens, broilers and layers, bred towards efficient meat and egg production, respectively. RESULTS: Broilers and layer hens, grown up to sexual maturation under free-feeding conditions, differed 4.0-fold in weight and 1.6-fold in ovarian-follicle counts, yet the relative accumulation of visceral fat was comparable. RNA-seq and mass-spectrometry (MS) analyses of visceral fat revealed differentially expressed genes between broilers and layers, 1106 at the mRNA level (FDR ≤ 0.05), and 203 at the protein level (P ≤ 0.05). In broilers, Ingenuity Pathway Analysis revealed activation of the PTEN-pathway, and in layers increased response to external signals. The expression pattern of genes encoding fat-secreted proteins in broilers and layers was characterized in the RNA-seq and MS data, as well as by qPCR on visceral fat under free feeding and 24 h-feed deprivation. This characterization was expanded using available RNA-seq data of tissues from red junglefowl, and of visceral fat from broilers of different types. These comparisons revealed expression of new adipokines and secreted proteins (LCAT, LECT2, SERPINE2, SFTP1, ZP1, ZP3, APOV1, VTG1 and VTG2) at the mRNA and/or protein levels, with dynamic gene expression patterns in the selected chicken lines (except for ZP1; FDR/P ≤ 0.05) and feed deprivation (NAMPT, SFTPA1 and ZP3) (P ≤ 0.05). In contrast, some of the most prominent adipokines in mammals, leptin, TNF, IFNG, and IL6 were expressed at a low level (FPKM/RPKM< 1) and did not show differential mRNA expression neither between broiler and layer lines nor between fed vs. feed-deprived chickens. CONCLUSIONS: Our study revealed that RNA and protein expression in visceral fat changes with selective breeding, suggesting endocrine roles of visceral fat in the selected phenotypes. In comparison to gene expression in visceral fat of mammals, our findings points to a more direct cross talk of the chicken visceral fat with the reproductive system and lower involvement in the regulation of appetite, inflammation and insulin resistance.
  • ItemOpen AccessPublished version Peer-reviewed
    Direct Conversion of Mouse Fibroblasts into Cholangiocyte Progenitor Cells
    (Elsevier, 2018-05-08) Lim, KT; Kim, Jonghun; Hwnag, Seon In; Zhang, Ludi; Han, Heonjong; Bae, Dasom; Kim, Kee-Pyo; Hu, Yi-Ping; Schöler, Hans; Lee, Insuk; Hui, Lijian; Han, Dong Wook; Lim, Kyungtae [0000-0001-6044-2191]
    Disorders of the biliary epithelium, known as cholangiopathies, cause severe and irreversible liver diseases. The limited accessibility of bile duct precludes modeling of several cholangiocyte-mediated diseases. Therefore, novel approaches for obtaining functional cholangiocytes with high purity are needed. Previous work has shown that the combination of Hnf1β and Foxa3 could directly convert mouse fibroblasts into bipotential hepatic stem cell-like cells, termed iHepSCs. However, the efficiency of converting fibroblasts into iHepSCs is low, and these iHepSCs exhibit extremely low differentiation potential into cholangiocytes, thus hindering the translation of iHepSCs to the clinic. Here, we describe that the expression of Hnf1α and Foxa3 dramatically facilitates the robust generation of iHepSCs. Notably, prolonged in vitro culture of Hnf1α- and Foxa3-derived iHepSCs induces a Notch signaling-mediated secondary conversion into cholangiocyte progenitor-like cells that display dramatically enhanced differentiation capacity into mature cholangiocytes. Our study provides a robust two-step approach for obtaining cholangiocyte progenitor-like cells using defined factors.
  • ItemOpen AccessAccepted version Peer-reviewed
    Ultrashort Echo Time Imaging of the Osteochondral Junction in Subjects with Knee Osteoarthritis and Age-matched Healthy Volunteers
    MacKay, JW; Kaggie, Josh; Morgan, Alexandra; Janiczek, Robert; Reid, Scott; McDonnell, Stephen; Khan, Wasim; Graves, Martin; Gilbert, Fiona; McCaskie, Andrew; MacKay, James [0000-0001-7558-3800]; Kaggie, Joshua [0000-0001-6706-3442]; McDonnell, Stephen [0000-0002-3181-4192]; Graves, Martin [0000-0003-4327-3052]; Gilbert, Fiona [0000-0002-0124-9962]; McCaskie, Andrew [0000-0001-6476-0832]
    SYNOPSIS We describe in vivo translation of ultrashort TE (UTE) imaging of the osteochondral junction (OCJ) at the knee in 9 subjects with osteoarthritis (OA) and 4 age-matched healthy volunteers. The OCJ plays an important role in onset and progression of OA. Our study demonstrates that UTE imaging of the OCJ is repeatable and demonstrates OCJ defects in OA subjects but not in healthy volunteers. Areas of OCJ damage commonly co-locate to other osteochondral pathology (bone marrow lesions and cartilage defects). UTE imaging of the OCJ may be a helpful tool for assessing OCJ damage in clinical studies of OA. INTRODUCTION Disruption of the osteochondral junction (OCJ) is thought to play an important role in the onset and progression of osteoarthritis (OA). Using conventional MR imaging, direct visualisation of the OCJ is not possible due to inherent short T1 and T2 relaxation times of the OCJ tissues. However, by achieving echo times (TEs) of < 1 ms, ultrashort echo time (UTE) MR imaging allows direct visualisation of the OCJ. The normal OCJ appears as an area of linear high signal intensity (SI) on UTE images at the bone-cartilage interface. In OA it has been shown that this area of linear high SI can become thinned or absent, compatible with histological findings of OCJ defects(1, 2). These findings have been described in a number of cadaveric MR studies, but there are limited in vivo data available(3-5). The aims of this study were to compare the in vivo appearance of the OCJ on UTE MR imaging between subjects with knee OA and age-matched healthy volunteers, to determine the relationship between OCJ defects and other osteochondral pathology, and to assess test-retest repeatability. METHODS We imaged 9 participants with mild-moderate knee osteoarthritis, characterised by radiographs with medial tibiofemoral compartment predominant disease and Kellgren-Lawrence grades 2-3, and 4 age-matched healthy volunteers. Participants were imaged at baseline and 1 month. MR studies were performed on a 3T system (GE 750, GE Healthcare). The MR protocol consisted of standard clinical sequences (coronal and sagittal intermediate-weighted fat-saturated fast spin echo (FSE) sequences plus a coronal T1-weighted FSE sequence) and a sagittal dual-echo UTE gradient echo sequence acquired using a 3D cones trajectory (research prototype; repetition time 15 ms, TE 0.03/4.5 ms, flip angle 13o, field-of-view 18 x 18 cm, matrix 430 x 430, slice thickness 2 mm, number of averages 1, acquisition time ~ 7.5 minutes). To increase conspicuity of short T2 tissues, we performed weighted digital image subtraction of the longer TE (4.5 ms) from the shorter TE images (0.03 ms)(6). The presence or absence of characteristic linear high SI at the OCJ was scored in 12 regions for each knee, corresponding to tibiofemoral subdivisions commonly used for semi-quantitative scoring. The presence of bone marrow lesions (BML) or cartilage defects in the same regions was also recorded. Assessment was performed by a single musculoskeletal radiologist with 5 years' experience in OA research, blinded to group assignment. We used descriptive statistics to compare the number of regions with OCJ defects in subjects with OA and healthy volunteers, and to assess the frequency with which OCJ defects co-located with BMLs or cartilage defects. Test-retest repeatability was evaluated using kappa statistics. RESULTS Participant characteristics are displayed in table 1. Six out of 9 OA participants (67%) had an OCJ defect in at least one region compared to 0 out of 4 controls (0%). The most commonly involved region was the central medial tibia (4 participants). OCJ defects commonly co-located to BMLs (7 out of 10 OCJ defects, 70%) and cartilage defects (6 out of 10 OCJ defects, 60%). Results are displayed in table 2. Sample images are displayed in figures 1 - 3. The kappa value for test-retest repeatability of OCJ assessment using UTE was 0.83 (95% confidence interval 0.64 to 1). DISCUSSION The appearances of OCJ defects in subjects with OA in vivo are in keeping with abnormalities predicted by cadaveric MR and histology studies(1, 3). The biological plausibility of the findings is enhanced by the frequency of co-location of OCJ damage to other osteochondral pathology (BMLs and cartilage defects). Our findings demonstrate in vivo translation of UTE imaging of the OCJ, and suggest that this is a useful tool for future studies of OA onset and progression. This may include predicting response to intervention, as equine studies have demonstrated that the presence or absence of OCJ damage is an important predictor of response to treatment of cartilage defects(7). Our results demonstrate that UTE imaging of the OCJ is repeatable with kappa values in keeping with 'near-perfect' test-retest repeatability for qualitative assessment(8). Previous in vivo studies have not used age-matched control subjects, therefore it has been unclear whether areas of OCJ damage are related to OA or normal ageing(4). The normal appearance of the OCJ in age-matched control subjects in this study suggests that the OCJ defects are not part of normal ageing, although at present the number of healthy volunteers imaged is small. CONCLUSION In vivo UTE MR imaging of the OCJ is repeatable and demonstrates OCJ defects in subjects with OA. OCJ defects commonly co-locate with other osteochondral pathology.
  • ItemOpen AccessPublished version Peer-reviewed
    Inhibition of β-catenin signalling in dermal fibroblasts enhances hair follicle regeneration during wound healing.
    (The Company of Biologists, 2016-07-15) Rognoni, Emanuel; Gomez, Celine; Pisco, Angela Oliveira; Rawlins, Emma L; Simons, Ben D; Watt, Fiona M; Driskell, Ryan R; Rognoni, Emanuel [0000-0001-6050-2860]; Watt, Fiona M [0000-0001-9151-5154]; Driskell, Ryan R [0000-0001-7673-2564]
    New hair follicles (HFs) do not form in adult mammalian skin unless epidermal Wnt signalling is activated genetically or within large wounds. To understand the postnatal loss of hair forming ability we monitored HF formation at small circular (2 mm) wound sites. At P2, new HFs formed in back skin, but HF formation was markedly decreased by P21. Neonatal tail also formed wound-associated HFs, albeit in smaller numbers. Postnatal loss of HF neogenesis did not correlate with wound closure rate but with a reduction in Lrig1-positive papillary fibroblasts in wounds. Comparative gene expression profiling of back and tail dermis at P1 and dorsal fibroblasts at P2 and P50 showed a correlation between loss of HF formation and decreased expression of genes associated with proliferation and Wnt/β-catenin activity. Between P2 and P50, fibroblast density declined throughout the dermis and clones of fibroblasts became more dispersed. This correlated with a decline in fibroblasts expressing a TOPGFP reporter of Wnt activation. Surprisingly, between P2 and P50 there was no difference in fibroblast proliferation at the wound site but Wnt signalling was highly upregulated in healing dermis of P21 compared with P2 mice. Postnatal β-catenin ablation in fibroblasts promoted HF regeneration in neonatal and adult mouse wounds, whereas β-catenin activation reduced HF regeneration in neonatal wounds. Our data support a model whereby postnatal loss of hair forming ability in wounds reflects elevated dermal Wnt/β-catenin activation in the wound bed, increasing the abundance of fibroblasts that are unable to induce HF formation.
  • ItemOpen AccessAccepted version Peer-reviewed
    A Glial Signature and Wnt7 Signaling Regulate Glioma-Vascular Interactions and Tumor Microenvironment.
    (Elsevier BV, 2018-05-14) Griveau, Amelie; Seano, Giorgio; Shelton, Samuel J; Kupp, Robert; Jahangiri, Arman; Obernier, Kirsten; Krishnan, Shanmugarajan; Lindberg, Olle R; Yuen, Tracy J; Tien, An-Chi; Sabo, Jennifer K; Wang, Nancy; Chen, Ivy; Kloepper, Jonas; Larrouquere, Louis; Ghosh, Mitrajit; Tirosh, Itay; Huillard, Emmanuelle; Alvarez-Buylla, Arturo; Oldham, Michael C; Persson, Anders I; Weiss, William A; Batchelor, Tracy T; Stemmer-Rachamimov, Anat; Suvà, Mario L; Phillips, Joanna J; Aghi, Manish K; Mehta, Shwetal; Jain, Rakesh K; Rowitch, David H; Rowitch, David [0000-0002-0079-0060]
    Gliomas comprise heterogeneous malignant glial and stromal cells. While blood vessel co-option is a potential mechanism to escape anti-angiogenic therapy, the relevance of glial phenotype in this process is unclear. We show that Olig2+ oligodendrocyte precursor-like glioma cells invade by single-cell vessel co-option and preserve the blood-brain barrier (BBB). Conversely, Olig2-negative glioma cells form dense perivascular collections and promote angiogenesis and BBB breakdown, leading to innate immune cell activation. Experimentally, Olig2 promotes Wnt7b expression, a finding that correlates in human glioma profiling. Targeted Wnt7a/7b deletion or pharmacologic Wnt inhibition blocks Olig2+ glioma single-cell vessel co-option and enhances responses to temozolomide. Finally, Olig2 and Wnt7 become upregulated after anti-VEGF treatment in preclinical models and patients. Thus, glial-encoded pathways regulate distinct glioma-vascular microenvironmental interactions.
  • ItemOpen AccessPublished version Peer-reviewed
    Fixation and Spread of Somatic Mutations in Adult Human Colonic Epithelium.
    (Elsevier BV, 2018-06-01) Nicholson, Anna M; Olpe, Cora; Hoyle, Alice; Thorsen, Ann-Sofie; Rus, Teja; Colombé, Mathilde; Brunton-Sim, Roxanne; Kemp, Richard; Marks, Kate; Quirke, Phil; Malhotra, Shalini; Ten Hoopen, Rogier; Ibrahim, Ashraf; Lindskog, Cecilia; Myers, Meagan B; Parsons, Barbara; Tavaré, Simon; Wilkinson, Mark; Morrissey, Edward; Winton, Douglas J; Nicholson, Anna [0000-0001-9128-7024]; Olpe, Cora [0000-0002-6350-018X]; Winton, Douglas [0000-0001-6067-7927]
    We investigated the means and timing by which mutations become fixed in the human colonic epithelium by visualizing somatic clones and mathematical inference. Fixation requires two sequential steps. First, one of approximately seven active stem cells residing within each colonic crypt has to be mutated. Second, the mutated stem cell has to replace neighbors to populate the entire crypt in a process that takes several years. Subsequent clonal expansion due to crypt fission is infrequent for neutral mutations (around 0.7% of all crypts undergo fission in a single year). Pro-oncogenic mutations subvert both stem cell replacement to accelerate fixation and clonal expansion by crypt fission to achieve high mutant allele frequencies with age. The benchmarking of these behaviors allows the advantage associated with different gene-specific mutations to be compared irrespective of the cellular mechanisms by which they are conferred.
  • ItemOpen AccessPublished version Peer-reviewed
    Thromopoietin signaling to chromatin elicits rapid and pervasive epigenome remodeling within poised chromatin architectures.
    (Cold Spring Harbor Laboratory Press) Green, AR; Comoglio, F; Park, H; Schoenfelder, S; Barozzi, I; Bode, D; Fraser, P; Green, Tony [0000-0002-9795-0218]
    Thrombopoietin (TPO) is a critical cytokine regulating hematopoietic stem cell maintenance and differentiation into the megakaryocytic lineage. However, the transcriptional and chromatin dynamics elicited by TPO signaling are poorly understood. Here, we study the immediate early transcriptional and cis-regulatory responses to TPO in hematopoietic stem/progenitor cells (HSPCs) and use this paradigm of cytokine signaling to chromatin to dissect the relation between cis- regulatory activity and chromatin architecture. We show that TPO profoundly alters the transcriptome of HSPCs, with key hematopoietic regulators being transcriptionally repressed within 30 minutes of TPO. By examining cis-regulatory dynamics and chromatin architectures, we demonstrate that these changes are accompanied by rapid and extensive epigenome remodeling of cis-regulatory landscapes that is spatially coordinated within topologically associating domains (TADs). Moreover, TPO-responsive enhancers are spatially clustered and engage in preferential homotypic intra- and inter-TAD interactions that are largely refractory to TPO signaling. By further examining the link between cis-regulatory dynamics and chromatin looping, we show that rapid modulation of cis-regulatory activity is largely independent of chromatin looping dynamics. Finally, we show that, although activated and repressed cis-regulatory elements share remarkably similar DNA sequence compositions, transcription factor binding patterns accurately predict rapid cis-regulatory responses to TPO.
  • ItemOpen AccessPublished version Peer-reviewed
    Predominant Asymmetrical Stem Cell Fate Outcome Limits the Rate of Niche Succession in Human Colonic Crypts.
    (Elsevier BV, 2018-05) Stamp, Craig; Zupanic, Anze; Sachdeva, Ashwin; Stoll, Elizabeth A; Shanley, Daryl P; Mathers, John C; Kirkwood, Thomas BL; Heer, Rakesh; Simons, Benjamin D; Turnbull, Doug M; Greaves, Laura C; Simons, Benjamin [0000-0002-3875-7071]
    Stem cell (SC) dynamics within the human colorectal crypt SC niche remain poorly understood, with previous studies proposing divergent hypotheses on the predominant mode of SC self-renewal and the rate of SC replacement. Here we use age-related mitochondrial oxidative phosphorylation (OXPHOS) defects to trace clonal lineages within human colorectal crypts across the adult life-course. By resolving the frequency and size distribution of OXPHOS-deficient clones, quantitative analysis shows that, in common with mouse, long-term maintenance of the colonic epithelial crypt relies on stochastic SC loss and replacement mediated by competition for limited niche access. We find that the colonic crypt is maintained by ~5 effective SCs. However, with a SC loss/replacement rate estimated to be slower than once per year, our results indicate that the vast majority of individual SC divisions result in asymmetric fate outcome. These findings provide a quantitative platform to detect and study deviations from human colorectal crypt SC niche homeostasis during the process of colorectal carcinogenesis.
  • ItemOpen AccessAccepted version Peer-reviewed
    Clonal analysis of Notch1-expressing cells reveals the existence of unipotent stem cells that retain long-term plasticity in the embryonic mammary gland.
    (Springer Science and Business Media LLC, 2018-06) Lilja, Anna M; Rodilla, Veronica; Huyghe, Mathilde; Hannezo, Edouard; Landragin, Camille; Renaud, Olivier; Leroy, Olivier; Rulands, Steffen; Simons, Benjamin D; Fre, Silvia; Rodilla, Veronica [0000-0003-0512-4179]; Hannezo, Edouard [0000-0001-6005-1561]; Rulands, Steffen [0000-0001-6398-1553]; Simons, Benjamin D [0000-0002-3875-7071]; Fre, Silvia [0000-0002-7209-7636]
    Recent lineage tracing studies have revealed that mammary gland homeostasis relies on unipotent stem cells. However, whether and when lineage restriction occurs during embryonic mammary development, and which signals orchestrate cell fate specification, remain unknown. Using a combination of in vivo clonal analysis with whole mount immunofluorescence and mathematical modelling of clonal dynamics, we found that embryonic multipotent mammary cells become lineage-restricted surprisingly early in development, with evidence for unipotency as early as E12.5 and no statistically discernable bipotency after E15.5. To gain insights into the mechanisms governing the switch from multipotency to unipotency, we used gain-of-function Notch1 mice and demonstrated that Notch activation cell autonomously dictates luminal cell fate specification to both embryonic and basally committed mammary cells. These functional studies have important implications for understanding the signals underlying cell plasticity and serve to clarify how reactivation of embryonic programs in adult cells can lead to cancer.
  • ItemOpen AccessPublished version Peer-reviewed
    The Challenges of First-in-Human Stem Cell Clinical Trials: What Does This Mean for Ethics and Institutional Review Boards?
    (Elsevier BV, 2018-05-08) Barker, Roger A; Carpenter, Melissa K; Forbes, Stuart; Goldman, Steven A; Jamieson, Catriona; Murry, Charles E; Takahashi, Jun; Weir, Gordon; Barker, Roger [0000-0001-8843-7730]
    Stem cell-based clinical interventions are increasingly advancing through preclinical testing and approaching clinical trials. The complexity and diversity of these approaches, and the confusion created by unproven and untested stem cell-based "therapies," create a growing need for a more comprehensive review of these early-stage human trials to ensure they place the patients at minimal risk of adverse events but are also based on solid evidence of preclinical efficacy with a clear scientific rationale for that effect. To address this issue and supplement the independent review process, especially that of the ethics and institutional review boards who may not be experts in stem cell biology, the International Society for Stem Cell Research (ISSCR) has developed a set of practical questions to cover the major issues for which clear evidence-based answers need to be obtained before approving a stem cell-based trial.
  • ItemOpen AccessPublished version Peer-reviewed
    Apathy in Huntington’s Disease: A Review of the Current Conceptualization
    (OMICS Publishing Group, 2018-01-01) Camacho, Marta; Barker, Roger A; Mason, Sarah L; Barker, Roger [0000-0001-8843-7730]; Mason, Sarah [0000-0001-6715-4109]
    Apathy is one of the most common psychiatric symptoms experienced by patients with Huntington’s disease (HD). It appears early, progresses with the disease course and has been shown to contribute significantly to caregiver burden. However, what is understood by apathy in HD is not clearly defined nor the underlying mechanisms responsible for it. In this review, we discuss the concept of apathy in the context of HD and propose that a consensus regarding its conceptualisation and subsequently its diagnostic criteria would significantly benefit the field. In order to undertake this work, we began by reviewing the existing literature on the definition and assessment of apathy in HD, its underlying neurobiological basis and its relationship to other related features such as abulia, anhedonia and alexithymia. In the context of HD, apathy could be described by a loss of or diminished motivation, emotion and goal-directed behaviour that is not best explained by motor or social constraints of the disease. However, there is an urgent need to better understand the characteristics of apathy specifically in HD, how they evolve across the disease course, and how they relate to central dopaminergic pathways. Only by undertaking such work can we hope to better understand this early and disabling aspect of HD.
  • ItemOpen AccessPublished version Peer-reviewed
    The developmental origin of brain tumours: a cellular and molecular framework.
    (The Company of Biologists, 2018-05-14) Azzarelli, Roberta; Simons, Benjamin D; Philpott, Anna; Azzarelli, Roberta [0000-0002-8160-7538]; Philpott, Anna [0000-0003-3789-2463]
    The development of the nervous system relies on the coordinated regulation of stem cell self-renewal and differentiation. The discovery that brain tumours contain a subpopulation of cells with stem/progenitor characteristics that are capable of sustaining tumour growth has emphasized the importance of understanding the cellular dynamics and the molecular pathways regulating neural stem cell behaviour. By focusing on recent work on glioma and medulloblastoma, we review how lineage tracing contributed to dissecting the embryonic origin of brain tumours and how lineage-specific mechanisms that regulate stem cell behaviour in the embryo may be subverted in cancer to achieve uncontrolled proliferation and suppression of differentiation.
  • ItemOpen AccessAccepted version Peer-reviewed
    Sorting apples from oranges in single-cell expression comparisons.
    (Springer Science and Business Media LLC, 2018-04-27) Hamey, Fiona K; Göttgens, Berthold; Hamey, Fiona [0000-0001-7299-2860]; Gottgens, Berthold [0000-0001-6302-5705]
    Two methods for comparing single-cell expression datasets help address the challenge of integrating data across conditions and experiments.
  • ItemOpen AccessAccepted version Peer-reviewed
    Human Pluripotent Stem Cell-Derived Endoderm for Modeling Development and Clinical Applications.
    (Elsevier BV, 2018-04-05) Yiangou, Loukia; Ross, Alexander DB; Goh, Kim Jee; Vallier, Ludovic; Vallier, Ludovic [0000-0002-3848-2602]
    The liver, lung, pancreas, and digestive tract all originate from the endoderm germ layer, and these vital organs are subject to many life-threatening diseases affecting millions of patients. However, primary cells from endodermal organs are often difficult to grow in vitro. Human pluripotent stem cells thus hold great promise for generating endoderm cells and their derivatives as tools for the development of new therapeutics against a variety of global healthcare challenges. Here we describe recent advances in methods for generating endodermal cell types from human pluripotent stem cells and their use for disease modeling and cell-based therapy.
  • ItemOpen AccessAccepted version Peer-reviewed
    A single-cell hematopoietic landscape resolves 8 lineage trajectories and defects in Kit mutant mice.
    (American Society of Hematology, 2018-05-24) Dahlin, Joakim S; Hamey, Fiona K; Pijuan-Sala, Blanca; Shepherd, Mairi; Lau, Winnie WY; Nestorowa, Sonia; Weinreb, Caleb; Wolock, Samuel; Hannah, Rebecca; Diamanti, Evangelia; Kent, David G; Göttgens, Berthold; Wilson, Nicola K; Dahlin, Joakim S [0000-0003-3007-9875]; Hamey, Fiona K [0000-0001-7299-2860]
    Hematopoietic stem and progenitor cells (HSPCs) maintain the adult blood system, and their dysregulation causes a multitude of diseases. However, the differentiation journeys toward specific hematopoietic lineages remain ill defined, and system-wide disease interpretation remains challenging. Here, we have profiled 44 802 mouse bone marrow HSPCs using single-cell RNA sequencing to provide a comprehensive transcriptional landscape with entry points to 8 different blood lineages (lymphoid, megakaryocyte, erythroid, neutrophil, monocyte, eosinophil, mast cell, and basophil progenitors). We identified a common basophil/mast cell bone marrow progenitor and characterized its molecular profile at the single-cell level. Transcriptional profiling of 13 815 HSPCs from the c-Kit mutant (W41/W41) mouse model revealed the absence of a distinct mast cell lineage entry point, together with global shifts in cell type abundance. Proliferative defects were accompanied by reduced Myc expression. Potential compensatory processes included upregulation of the integrated stress response pathway and downregulation of proapoptotic gene expression in erythroid progenitors, thus providing a template of how large-scale single-cell transcriptomic studies can bridge between molecular phenotypes and quantitative population changes.
  • ItemOpen AccessAccepted version Peer-reviewed
    Single-cell transcriptional profiling: a window into embryonic cell-type specification.
    (Springer Science and Business Media LLC, 2018-06) Pijuan-Sala, Blanca; Guibentif, Carolina; Göttgens, Berthold; Göttgens, Berthold [0000-0001-6302-5705]
    During mammalian embryonic development, a single fertilized egg cell will proliferate and differentiate into all the cell lineages and cell types that eventually form the adult organism. Cell lineage diversification involves repeated cell fate choices that ultimately occur at the level of the individual cell rather than at the cell-population level. Improvements in single-cell technologies are transforming our understanding of mammalian development, not only by overcoming the limitations presented by the extremely low cell numbers of early embryos but also by enabling the study of cell fate specification in greater detail. In this Review, we first discuss recent advances in single-cell transcriptomics and imaging and provide a brief outline of current bioinformatics methods available to analyse the resulting data. We then discuss how these techniques have contributed to our understanding of pre-implantation and early post-implantation development and of in vitro pluripotency. Finally, we overview the current challenges facing single-cell research and highlight the latest advances and potential future avenues.
  • ItemOpen AccessAccepted version Peer-reviewed
    UTX-mediated enhancer and chromatin remodeling suppresses myeloid leukemogenesis through noncatalytic inverse regulation of ETS and GATA programs.
    (Nature Publishing Group, 2018-06) Huntly, BJP; Meduri, Eshwar; Yun, Haiyang; Gottgens, Bertie; Vassiliou, George; Huntly, Brian [0000-0003-0312-161X]; Gottgens, Berthold [0000-0001-6302-5705]; Vassiliou, George [0000-0003-4337-8022]
    The H3K27 lysine-specific demethylase UTX is targeted by loss-of-function mutations in multiple cancers. Here, we demonstrate that UTX suppresses myeloid leukemogenesis through non-catalytic functions, a property shared with its catalytically inactive Y-chromosome paralogue, UTY. In keeping with this, we demonstrate concomitant loss/mutation of UTX and UTY in multiple human cancers. Mechanistically, global genomic profiling revealed only minor changes in H3K27Me3, but significant and bidirectional alterations of H3K27Ac and chromatin accessibility, a predominant loss of H3K4Me1 modifications, alterations in ETS and GATA factor binding and altered gene expression upon Utx loss. By integrating proteomic and genomic analyses, we link these changes to UTX regulation of ATP-dependent chromatin remodeling, coordination of the COMPASS complex and enhanced pioneering activity of ETS factors during evolution to AML. Collectively, our findings reveal a dual role for UTX in suppressing acute myeloid leukaemia via repression of oncogenic ETS and upregulation of tumor-suppressive GATA programs
  • ItemOpen AccessPublished version Peer-reviewed
    Association of subchondral bone texture on magnetic resonance imaging with radiographic knee osteoarthritis progression: data from the Osteoarthritis Initiative Bone Ancillary Study.
    (Springer Science and Business Media LLC, 2018-11) MacKay, James W; Kapoor, Geeta; Driban, Jeffrey B; Lo, Grace H; McAlindon, Timothy E; Toms, Andoni P; McCaskie, Andrew W; Gilbert, Fiona J; MacKay, James [0000-0001-7558-3800]; McCaskie, Andrew [0000-0001-6476-0832]; Gilbert, Fiona [0000-0002-0124-9962]
    OBJECTIVES: To assess whether initial or 12-18-month change in magnetic resonance imaging (MRI) subchondral bone texture is predictive of radiographic knee osteoarthritis (OA) progression over 36 months. METHODS: This was a nested case-control study including 122 knees/122 participants in the Osteoarthritis Initiative (OAI) Bone Ancillary Study, who underwent MRI optimised for subchondral bone assessment at either the 30- or 36-month and 48-month OAI visits. Case knees (n = 61) had radiographic OA progression between the 36- and 72-month OAI visits, defined as ≥ 0.7 mm minimum medial tibiofemoral radiographic joint space (minJSW) loss. Control knees (n = 61) without radiographic OA progression were matched (1:1) to cases for age, sex, body mass index and initial medial minJSW. Texture analysis was performed on the medial femoral and tibial subchondral bone. We assessed the association of texture features with radiographic progression by creating a composite texture score using penalised logistic regression and calculating odds ratios. We evaluated the predictive performance of texture features for predicting radiographic progression using c-statistics. RESULTS: Initial (odds ratio [95% confidence interval] = 2.13 [1.41-3.40]) and 12- 18-month change (3.76 [2.04-7.82]) texture scores were significantly associated with radiographic OA progression. Combinations of texture features were significant predictors of radiographic progression using initial (c-statistic [95% confidence interval] = 0.65 [0.64-0.65], p = 0.003) and 12-18-month change (0.68 [0.68-0.68], p < 0.001) data. CONCLUSIONS: Initial and 12-18-month changes in MRI subchondral bone texture score were significantly associated with radiographic progression at 36 months, with better predictive performance for 12-18-month change in texture. These results suggest that texture analysis may be a useful biomarker of subchondral bone in OA. KEY POINTS: • Subchondral bone MRI texture analysis is a promising knee osteoarthritis imaging biomarker. • In this study, subchondral bone texture was associated with knee osteoarthritis progression. • This demonstrates predictive and concurrent validity of MRI subchondral bone texture analysis. • This method may be useful in clinical trials with interventions targeting bone.
  • ItemOpen AccessPublished version Peer-reviewed
    Origin and dynamics of oligodendrocytes in the developing brain: Implications for perinatal white matter injury.
    (Wiley, 2018-02) van Tilborg, Erik; de Theije, Caroline GM; van Hal, Maurik; Wagenaar, Nienke; de Vries, Linda S; Benders, Manon J; Rowitch, David H; Nijboer, Cora H; Rowitch, David [0000-0002-0079-0060]
    Infants born prematurely are at high risk to develop white matter injury (WMI), due to exposure to hypoxic and/or inflammatory insults. Such perinatal insults negatively impact the maturation of oligodendrocytes (OLs), thereby causing deficits in myelination. To elucidate the precise pathophysiology underlying perinatal WMI, it is essential to fully understand the cellular mechanisms contributing to healthy/normal white matter development. OLs are responsible for myelination of axons. During brain development, OLs are generally derived from neuroepithelial zones, where neural stem cells committed to the OL lineage differentiate into OL precursor cells (OPCs). OPCs, in turn, develop into premyelinating OLs and finally mature into myelinating OLs. Recent studies revealed that OPCs develop in multiple waves and form potentially heterogeneous populations. Furthermore, it has been shown that myelination is a dynamic and plastic process with an excess of OPCs being generated and then abolished if not integrated into neural circuits. Myelination patterns between rodents and humans show high spatial and temporal similarity. Therefore, experimental studies on OL biology may provide novel insights into the pathophysiology of WMI in the preterm infant and offers new perspectives on potential treatments for these patients.
  • ItemOpen AccessAccepted version Peer-reviewed
    Contributions of BMPR2 Mutations and Extrinsic Factors to Cellular Phenotypes of Pulmonary Arterial Hypertension Revealed by Induced Pluripotent Stem Cell Modeling.
    (American Thoracic Society, 2018-07-15) Kiskin, Fedir N; Chang, C-Hong; Huang, Christopher JZ; Kwieder, Baraa; Cheung, Christine; Dunmore, Benjamin J; Serrano, Felipe; Sinha, Sanjay; Morrell, Nicholas W; Rana, Amer A; Kiskin, Fedir N [0000-0002-6984-9622]; Rana, Amer A [0000-0002-2330-4643]
    Reduced bone morphogenetic protein receptor 2 (BMPR2) signaling is central to the pathobiology of pulmonary arterial hypertension (PAH). However, the reduced penetrance of BMPR2 mutations in families suggests that other factors are required to establish disease (1). To date, it has proved difficult to elucidate these factors due to a lack of appropriate models. Sa et al. (2017) developed an iPSC-EC model of PAH that recapitulated some of the previously described phenotypes of patient-derived PAECs, as well as appropriate responsiveness to Elafin and FK506 (2). This demonstrated a potential utility of iPSCs in modeling PAECs in PAH. However, other phenotypes such as inner mitochondrial membrane (IMM) hyperpolarization, could not be recapitulated. Therefore, there is a need to better understand the contribution of BMPR2 mutations to PAH-associated phenotypes and the requirement for other factors in this process. Two advantages of iPSCs in disease modeling are their amenability to genome editing and their differentiation into specific cell types under serum-free, chemically-defined conditions. This allows the assessment of the impact of a BMPR2 mutation without the confounding effects of genetic differences between cell lines, and to determine the impact of controlled exposure to extrinsic factors that may influence the acquisition of a diseased state. In addition, no iPSC-smooth muscle cell (SMC) model of PAH has yet been described. We have addressed these issues.