Anatomy and Neuroscience - Research Publications

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    Exposure to chorioamnionitis alters the monocyte transcriptional response to the neonatal pathogen Staphylococcus epidermidis
    de Jong, E ; Hancock, DG ; Wells, C ; Richmond, P ; Simmer, K ; Burgner, D ; Strunk, T ; Currie, AJ (WILEY, 2018-09-01)
    Preterm infants are uniquely susceptible to late-onset sepsis that is frequently caused by the skin commensal Staphylococcus epidermidis. Innate immune responses, particularly from monocytes, are a key protective mechanism. Impaired cytokine production by preterm infant monocytes is well described, but few studies have comprehensively assessed the corresponding monocyte transcriptional response. Innate immune responses in preterm infants may be modulated by inflammation such as prenatal exposure to histologic chorioamnionitis which complicates 40-70% of preterm pregnancies. Chorioamnionitis alters the risk of late-onset sepsis, but its effect on monocyte function is largely unknown. Here, we aimed to determine the impact of exposure to chorioamnionitis on the proportions and phenotype of cord blood monocytes using flow cytometry, as well as their transcriptional response to live S. epidermidis. RNA-seq was performed on purified cord blood monocytes from very preterm infants (<32 weeks gestation, with and without chorioamnionitis-exposure) and term infants (37-40 weeks), pre- and postchallenge with live S. epidermidis. Preterm monocytes from infants without chorioamnionitis-exposure did not exhibit an intrinsically deficient transcriptional response to S. epidermidis compared to term infants. In contrast, chorioamnionitis-exposure was associated with hypo-responsive transcriptional phenotype regarding a subset of genes involved in antigen presentation and adaptive immunity. Overall, our findings suggest that prenatal exposure to inflammation may alter the risk of sepsis in preterm infants partly by modulation of monocyte responses to pathogens.
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    Group A Streptococcus M1T1 Intracellular Infection of Primary Tonsil Epithelial Cells Dampens Levels of Secreted IL-8 Through the Action of SpyCEP
    Soderholm, AT ; Barnett, TC ; Korn, O ; Rivera-Hernandez, T ; Seymour, LM ; Schulz, BL ; Nizet, V ; Wells, CA ; Sweet, MJ ; Walker, MJ (FRONTIERS MEDIA SA, 2018-05-17)
    Streptococcus pyogenes (Group A Streptococcus; GAS) commonly causes pharyngitis in children and adults, with severe invasive disease and immune sequelae being an infrequent consequence. The ability of GAS to invade the host and establish infection likely involves subversion of host immune defenses. However, the signaling pathways and innate immune responses of epithelial cells to GAS are not well-understood. In this study, we utilized RNAseq to characterize the inflammatory responses of primary human tonsil epithelial (TEpi) cells to infection with the laboratory-adapted M6 strain JRS4 and the M1T1 clinical isolate 5448. Both strains induced the expression of genes encoding a wide range of inflammatory mediators, including IL-8. Pathway analysis revealed differentially expressed genes between mock and JRS4- or 5448-infected TEpi cells were enriched in transcription factor networks that regulate IL-8 expression, such as AP-1, ATF-2, and NFAT. While JRS4 infection resulted in high levels of secreted IL-8, 5448 infection did not, suggesting that 5448 may post-transcriptionally dampen IL-8 production. Infection with 5448ΔcepA, an isogenic mutant lacking the IL-8 protease SpyCEP, resulted in IL-8 secretion levels comparable to JRS4 infection. Complementation of 5448ΔcepA and JRS4 with a plasmid encoding 5448-derived SpyCEP significantly reduced IL-8 secretion by TEpi cells. Our results suggest that intracellular infection with the pathogenic GAS M1T1 clone induces a strong pro-inflammatory response in primary tonsil epithelial cells, but modulates this host response by selectively degrading the neutrophil-recruiting chemokine IL-8 to benefit infection.
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    The Impact of APP on Alzheimer-like Pathogenesis and Gene Expression in Down Syndrome iPSC-Derived Neurons
    Ovchinnikov, DA ; Korn, O ; Virshup, I ; Wells, CA ; Wolvetang, EJ (CELL PRESS, 2018-07-10)
    Early-onset Alzheimer disease (AD)-like pathology in Down syndrome is commonly attributed to an increased dosage of the amyloid precursor protein (APP) gene. To test this in an isogenic human model, we deleted the supernumerary copy of the APP gene in trisomic Down syndrome induced pluripotent stem cells or upregulated APP expression in euploid human pluripotent stem cells using CRISPRa. Cortical neuronal differentiation shows that an increased APP gene dosage is responsible for increased β-amyloid production, altered Aβ42/40 ratio, and deposition of the pyroglutamate (E3)-containing amyloid aggregates, but not for several tau-related AD phenotypes or increased apoptosis. Transcriptome comparisons demonstrate that APP has a widespread and temporally modulated impact on neuronal gene expression. Collectively, these data reveal an important role for APP in the amyloidogenic aspects of AD but challenge the idea that increased APP levels are solely responsible for increasing specific phosphorylated forms of tau or enhanced neuronal cell death in Down syndrome-associated AD pathogenesis.
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    Age-specific biological and molecular profiling distinguishes paediatric from adult acute myeloid leukaemias
    Chaudhury, S ; O'Connor, C ; Canete, A ; Bittencourt-Silvestre, J ; Sarrou, E ; Prendergast, A ; Choi, J ; Johnston, P ; Wells, CA ; Gibson, B ; Keeshan, K (NATURE PUBLISHING GROUP, 2018-12-11)
    Acute myeloid leukaemia (AML) affects children and adults of all ages. AML remains one of the major causes of death in children with cancer and for children with AML relapse is the most common cause of death. Here, by modelling AML in vivo we demonstrate that AML is discriminated by the age of the cell of origin. Young cells give rise to myeloid, lymphoid or mixed phenotype acute leukaemia, whereas adult cells give rise exclusively to AML, with a shorter latency. Unlike adult, young AML cells do not remodel the bone marrow stroma. Transcriptional analysis distinguishes young AML by the upregulation of immune pathways. Analysis of human paediatric AML samples recapitulates a paediatric immune cell interaction gene signature, highlighting two genes, RGS10 and FAM26F as prognostically significant. This work advances our understanding of paediatric AML biology, and provides murine models that offer the potential for developing paediatric specific therapeutic strategies.
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    Stemformatics: visualize and download curated stem cell data
    Choi, J ; Pacheco, CM ; Mosbergen, R ; Korn, O ; Chen, T ; Nagpal, I ; Englart, S ; Angel, PW ; Wells, CA (OXFORD UNIV PRESS, 2019-01-08)
    Stemformatics is an established gene expression data portal containing over 420 public gene expression datasets derived from microarray, RNA sequencing and single cell profiling technologies. Developed for the stem cell community, it has a major focus on pluripotency, tissue stem cells, and staged differentiation. Stemformatics includes curated 'collections' of data relevant to cell reprogramming, as well as hematopoiesis and leukaemia. Rather than simply rehosting datasets as they appear in public repositories, Stemformatics uses a stringent set of quality control metrics and its own pipelines to process handpicked datasets from raw files. This means that about 30% of datasets processed by Stemformatics fail the quality control metrics and never make it to the portal, ensuring that Stemformatics data are of high quality and have been processed in a consistent manner. Stemformatics provides easy-to-use and intuitive tools for biologists to visually explore the data, including interactive gene expression profiles, principal component analysis plots and hierarchical clusters, among others. The addition of tools that facilitate cross-dataset comparisons provides users with snapshots of gene expression in multiple cell and tissues, assisting the identification of cell-type restricted genes, or potential housekeeping genes. Stemformatics is freely available at stemformatics.org.
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    Toll-Like Receptors Drive Specific Patterns of Tolerance and Training on Restimulation of Macrophages
    Butcher, SK ; O'Carroll, CE ; Wells, CA ; Carmody, RJ (FRONTIERS MEDIA SA, 2018-05-14)
    Tolerance is a long-recognized property of macrophages that leads to an altered response to repeated or chronic exposure to endotoxin. The physiological role of tolerance is to limit the potential damage to host tissue that may otherwise result from prolonged production of pro-inflammatory cytokines. Tolerance is induced by all toll-like receptor (TLR) ligands tested to date, however, tolerance induced by the TLR4 ligand lipopolysaccharide (LPS) is by far the best studied. LPS tolerance involves a global transcriptional shift from a pro-inflammatory response toward one characterized by the expression of anti-inflammatory and pro-resolution factors. Although largely reversible, LPS-tolerance leads to a hybrid macrophage activation state that is pro-inflammatory in nature, but possesses distinct regulatory anti-inflammatory features. Remarkably, a comparative transcriptomic analysis of tolerance induced by different TLR ligands has not previously been reported. Here, we describe the transcriptomic profiles of mouse macrophages tolerized with ligands for TLR2, TLR3, TLR4 and TLR 9. While we identified TLR-specific transcriptional profiles in macrophages tolerized with each ligand, tolerance induced by TLR4 represented an archetype pattern, such that each gene tolerized by any of the TLRs tested was also found to be tolerized by TLR4. Pro-inflammatory cytokines are not universally suppressed in all tolerant cells, but distinct patterns of cytokine expression distinguished TLR-specific tolerance. Analysis of gene regulatory regions revealed specific DNA sequence motifs associated with distinct states of TLR tolerance, implicating previously identified as well as novel transcriptional regulators of tolerance in macrophages. These data provide a basis for the future exploitation of TLR-specific tolerant states to achieve therapeutic re-programming of the innate immune response.
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    Transcriptional Profiling of Stem Cells: Moving from Descriptive to Predictive Paradigms
    Wells, CA ; Choi, J (CELL PRESS, 2019-08-13)
    Transcriptional profiling is a powerful tool commonly used to benchmark stem cells and their differentiated progeny. As the wealth of stem cell data builds in public repositories, we highlight common data traps, and review approaches to combine and mine this data for new cell classification and cell prediction tools. We touch on future trends for stem cell profiling, such as single-cell profiling, long-read sequencing, and improved methods for measuring molecular modifications on chromatin and RNA that bring new challenges and opportunities for stem cell analysis.
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    Shared activity patterns arising at genetic susceptibility loci reveal underlying genomic and cellular architecture of human disease
    Baillie, JK ; Bretherick, A ; Haley, CS ; Clohisey, S ; Grays, A ; Neyton, LPA ; Barrett, J ; Stahl, EA ; Tenesa, A ; Andersson, R ; Brown, JB ; Faulkner, GJ ; Lizio, M ; Schaefer, U ; Daub, C ; Itoh, M ; Kondo, N ; Lassmann, T ; Kawai, J ; Mole, D ; Bajic, VB ; Heutink, P ; Rehli, M ; Kawaji, H ; Sandelin, A ; Suzuki, H ; Satsangi, J ; Wells, CA ; Hacohen, N ; Freeman, TC ; Hayashizaki, Y ; Carninci, P ; Forrest, ARR ; Hume, DA ; Bergmann, S (PUBLIC LIBRARY SCIENCE, 2018-03-01)
    Genetic variants underlying complex traits, including disease susceptibility, are enriched within the transcriptional regulatory elements, promoters and enhancers. There is emerging evidence that regulatory elements associated with particular traits or diseases share similar patterns of transcriptional activity. Accordingly, shared transcriptional activity (coexpression) may help prioritise loci associated with a given trait, and help to identify underlying biological processes. Using cap analysis of gene expression (CAGE) profiles of promoter- and enhancer-derived RNAs across 1824 human samples, we have analysed coexpression of RNAs originating from trait-associated regulatory regions using a novel quantitative method (network density analysis; NDA). For most traits studied, phenotype-associated variants in regulatory regions were linked to tightly-coexpressed networks that are likely to share important functional characteristics. Coexpression provides a new signal, independent of phenotype association, to enable fine mapping of causative variants. The NDA coexpression approach identifies new genetic variants associated with specific traits, including an association between the regulation of the OCT1 cation transporter and genetic variants underlying circulating cholesterol levels. NDA strongly implicates particular cell types and tissues in disease pathogenesis. For example, distinct groupings of disease-associated regulatory regions implicate two distinct biological processes in the pathogenesis of ulcerative colitis; a further two separate processes are implicated in Crohn's disease. Thus, our functional analysis of genetic predisposition to disease defines new distinct disease endotypes. We predict that patients with a preponderance of susceptibility variants in each group are likely to respond differently to pharmacological therapy. Together, these findings enable a deeper biological understanding of the causal basis of complex traits.
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    Dynamic interplay of innate and adaptive immunity during sterile retinal inflammation: Insights from the transcriptome
    Natoli, R ; Mason, E ; Jiao, H ; Chuah, A ; Patel, H ; Fernando, N ; Valter, K ; Wells, C ; Provis, J ; Rutar, M (Frontiers Media, 2018-02-24)
    The pathogenesis of many retinal degenerations, such as age-related macular degeneration (AMD), is punctuated by an ill-defined network of sterile inflammatory responses. The delineation of innate and adaptive immune milieu amongst the broad leukocyte infiltrate, and the gene networks which construct these responses, are poorly described in the eye. Using photo-oxidative damage in a rodent model of subretinal inflammation, we employed a novel RNA-sequencing framework to map the global gene network signature of retinal leukocytes. This revealed a previously uncharted interplay of adaptive immunity during subretinal inflammation, including prolonged enrichment of myeloid and lymphocyte migration, antigen presentation, and the alternative arm of the complement cascade involving Factor B. We demonstrate Factor B-deficient mice are protected against macrophage infiltration and subretinal inflammation. Suppressing the drivers of retinal leukocyte proliferation, or their capacity to elicit complement responses, may help preserve retinal structure and function during sterile inflammation in diseases such as AMD.