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Microglia play key roles in brain homeostasis as well as responses to neurodegeneration and neuroinflammatory processes caused by physical disease and psychosocial stress. The pig is a physiologically-relevant model species for studying human neurological disorders, many of which are associated with microglial dysfunction. Furthermore, pigs are an important agricultural species, and there is a need to understand how microglial function affects their welfare. As a basis for improved understanding to enhance biomedical and agricultural research, we sought to characterise pig microglial identity at genome-wide scale and conduct inter-species comparisons. We isolated pig hippocampal tissue and microglia from frontal cortex, hippocampus and cerebellum, as well as alveolar macrophages from the lungs and conducted RNA-sequencing (RNAseq). By comparing the transcriptomic profiles between microglia, macrophages, and hippocampal tissue, we derived a set of 365 highly-enriched genes defining the porcine core microglial signature. We found brain regional heterogeneity based on 215 genes showing significant (adjusted p<0.01) regional variations and that cerebellar microglia were most distinct. We compared normalized gene expression for microglia from human, mice and pigs using microglia signature gene lists derived from each species and demonstrated that a core microglial marker gene signature is conserved across species, but that species-specific expression subsets also exist. Importantly, pig and human microglia shared greater similarity than pig and murine microglia. Our data provide a valuable resource defining the pig microglial transcriptome signature that highlights pigs as a useful large animal species bridging between rodents and humans in which to study the role of microglia during homeostasis and disease. Main Points - Defined a pig microglial transcriptome signature comprising 365 genes. - Demonstrated regional variance in the pig microglial transcriptome across the brain. - Revealed greater similarity between pig and human microglia than mouse.
Barbara B Shih; Sarah M Brown; Lucas Lefevre; Neil A Mabbott; Josef Priller; Gerard Thompson; Alistair B Lawrence; Barry W McColl. Defining the pig microglial transcriptome reveals their core signature, regional heterogeneity, and similarity with humans. 2021, 1 .
AMA StyleBarbara B Shih, Sarah M Brown, Lucas Lefevre, Neil A Mabbott, Josef Priller, Gerard Thompson, Alistair B Lawrence, Barry W McColl. Defining the pig microglial transcriptome reveals their core signature, regional heterogeneity, and similarity with humans. . 2021; ():1.
Chicago/Turabian StyleBarbara B Shih; Sarah M Brown; Lucas Lefevre; Neil A Mabbott; Josef Priller; Gerard Thompson; Alistair B Lawrence; Barry W McColl. 2021. "Defining the pig microglial transcriptome reveals their core signature, regional heterogeneity, and similarity with humans." , no. : 1.
In 2019, a novel coronavirus, SARS-CoV-2/nCoV-19, emerged in Wuhan, China, and has been responsible for the current COVID-19 pandemic. The evolutionary origins of the virus remain elusive and understanding its complex mutational signatures could guide vaccine design and development. As part of the international “CoronaHack” in April 2020, we employed a collection of contemporary methodologies to compare the genomic sequences of coronaviruses isolated from human (SARS-CoV-2; n = 163), bat (bat-CoV; n = 215) and pangolin (pangolin-CoV; n = 7) available in public repositories. We have also noted the pangolin-CoV isolate MP789 to bare stronger resemblance to SARS-CoV-2 than other pangolin-CoV. Following de novo gene annotation prediction, analyses of gene–gene similarity network, codon usage bias and variant discovery were undertaken. Strong host-associated divergences were noted in ORF3a, ORF6, ORF7a, ORF8 and S, and in codon usage bias profiles. Last, we have characterised several high impact variants (in-frame insertion/deletion or stop gain) in bat-CoV and pangolin-CoV populations, some of which are found in the same amino acid position and may be highlighting loci of potential functional relevance.
Nicholas J. Dimonaco; Mazdak Salavati; Barbara B. Shih. Computational Analysis of SARS-CoV-2 and SARS-Like Coronavirus Diversity in Human, Bat and Pangolin Populations. Viruses 2020, 13, 49 .
AMA StyleNicholas J. Dimonaco, Mazdak Salavati, Barbara B. Shih. Computational Analysis of SARS-CoV-2 and SARS-Like Coronavirus Diversity in Human, Bat and Pangolin Populations. Viruses. 2020; 13 (1):49.
Chicago/Turabian StyleNicholas J. Dimonaco; Mazdak Salavati; Barbara B. Shih. 2020. "Computational Analysis of SARS-CoV-2 and SARS-Like Coronavirus Diversity in Human, Bat and Pangolin Populations." Viruses 13, no. 1: 49.
Summary Introduction Ageing is associated with increased number of infections, decreased vaccine efficacy and increased systemic inflammation termed inflammageing. These changes are reflected by reduced recall responses to varicella zoster virus (VZV) challenge in the skin of older adults. Vitamin D deficiency is more common in the old and has been associated with frailty and increased inflammation. In addition, vitamin D increases immunoregulatory mechanisms and therefore has the potential to inhibit inflammageing. Objectives We investigated the use of vitamin D3 replacement to enhance cutaneous antigen-specific immunity in older adults (≥65 years). Methods Vitamin D insufficient older adults (n = 18) were administered 6400IU of vitamin D3/day orally for 14 weeks. Antigen-specific immunity to VZV was assessed by clinical score assessment of the injection site and transcriptional analysis of skin biopsies collected from challenged injection sites pre- and post-vitamin D3 replacement. Results We showed that older adults had reduced VZV-specific cutaneous immune response and increased non-specific inflammation as compared to young. Increased non-specific inflammation observed in the skin of older adults negatively correlated with vitamin D sufficiency. We showed that vitamin D3 supplementation significantly increased the response to cutaneous VZV antigen challenge in older adults. This enhancement was associated with a reduction in inflammatory monocyte infiltration with a concomitant enhancement of T cell recruitment to the site of antigen challenge in the skin. Conclusion Vitamin D3 replacement can boost antigen-specific immunity in older adults with sub-optimal vitamin D status.
Emma S Chambers; Milica Vukmanovic-Stejic; Carolin T Turner; Barbara B Shih; Hugh Trahair; Gabriele Pollara; Evdokia Tsaliki; Malcolm Rustin; Tom C Freeman; Neil A Mabbott; Mahdad Noursadeghi; Adrian R Martineau; Arne N Akbar. Vitamin D3 replacement enhances antigen-specific immunity in older adults. Immunotherapy Advances 2020, 1, 1 .
AMA StyleEmma S Chambers, Milica Vukmanovic-Stejic, Carolin T Turner, Barbara B Shih, Hugh Trahair, Gabriele Pollara, Evdokia Tsaliki, Malcolm Rustin, Tom C Freeman, Neil A Mabbott, Mahdad Noursadeghi, Adrian R Martineau, Arne N Akbar. Vitamin D3 replacement enhances antigen-specific immunity in older adults. Immunotherapy Advances. 2020; 1 (1):1.
Chicago/Turabian StyleEmma S Chambers; Milica Vukmanovic-Stejic; Carolin T Turner; Barbara B Shih; Hugh Trahair; Gabriele Pollara; Evdokia Tsaliki; Malcolm Rustin; Tom C Freeman; Neil A Mabbott; Mahdad Noursadeghi; Adrian R Martineau; Arne N Akbar. 2020. "Vitamin D3 replacement enhances antigen-specific immunity in older adults." Immunotherapy Advances 1, no. 1: 1.
In 2019, a novel coronavirus, SARS-CoV-2/nCoV-19, emerged in Wuhan, China, and has been responsible for the current COVID-19 pandemic. The evolutionary origins of the virus remain elusive and understanding its complex mutational signatures could guide vaccine design and development. As part of the international “CoronaHack” in April 2020 (https://www.coronahack.co.uk/), we employed a collection of contemporary methodologies to compare the genomic sequences of coronaviruses isolated from human (SARS-CoV-2;n=163), bat (bat-CoV;n=215) and pangolin (pangolin-CoV;n=7) available in public repositories. Following de novo gene annotation prediction, analysis on gene-gene similarity network, codon usage bias and variant discovery were carried out. Strong host-associated divergences were noted in ORF3a, ORF6, ORF7a, ORF8 and S, and in codon usage bias profiles. Lastly, we have characterised several high impact variants (inframe insertion/deletion or stop gain) in bat-CoV and pangolin-CoV populations, some of which are found in the same amino acid position and may be highlighting loci of potential functional relevance.
Nicholas J. Dimonaco; Mazdak Salavati; Barbara Shih. Hacking the diversity of SARS-CoV-2 and SARS-like coronaviruses in human, bat and pangolin populations. 2020, 1 .
AMA StyleNicholas J. Dimonaco, Mazdak Salavati, Barbara Shih. Hacking the diversity of SARS-CoV-2 and SARS-like coronaviruses in human, bat and pangolin populations. . 2020; ():1.
Chicago/Turabian StyleNicholas J. Dimonaco; Mazdak Salavati; Barbara Shih. 2020. "Hacking the diversity of SARS-CoV-2 and SARS-like coronaviruses in human, bat and pangolin populations." , no. : 1.
The subset of patients who develop critical illness in Covid-19 have extensive inflammation affecting the lungs1 and are strikingly different from other patients: immunosuppressive therapy benefits critically-ill patients, but may harm some non-critical cases.2 Since susceptibility to life-threatening infections and immune-mediated diseases are both strongly heritable traits, we reasoned that host genetic variation may identify mechanistic targets for therapeutic development in Covid-19.3 GenOMICC (Genetics Of Mortality In Critical Care, genomicc.org) is a global collaborative study to understand the genetic basis of critical illness. Here we report the results of a genome-wide association study (GWAS) in 2244 critically-ill Covid-19 patients from 208 UK intensive care units (ICUs), representing >95% of all ICU beds. Ancestry-matched controls were drawn from the UK Biobank population study and results were confirmed in GWAS comparisons with two other population control groups: the 100,000 genomes project and Generation Scotland. We identify and replicate three novel genome-wide significant associations, at chr19p13.3 (rs2109069, p = 3.98 × 10−12), within the gene encoding dipeptidyl peptidase 9 (DPP9), at chr12q24.13 (rs10735079, p =1.65 × 10−8) in a gene cluster encoding antiviral restriction enzyme activators (OAS1, OAS2, OAS3), and at chr21q22.1 (rs2236757, p = 4.99 × 10−8) in the interferon receptor gene IFNAR2. Consistent with our focus on extreme disease in younger patients with less comorbidity, we detect a stronger signal at the known 3p21.31 locus than previous studies (rs73064425, p = 4.77 × 10−30). We identify potential targets for repurposing of licensed medications. Using Mendelian randomisation we found evidence in support of a causal link from low expression of IFNAR2, and high expression of TYK2, to life-threatening disease. Transcriptome-wide association in lung tissue revealed that high expression of the monocyte/macrophage chemotactic receptor CCR2 is associated with severe Covid-19. Our results identify robust genetic signals relating to key host antiviral defence mechanisms, and mediators of inflammatory organ damage in Covid-19. Both mechanisms may be amenable to targeted treatment with existing drugs. Large-scale randomised clinical trials will be essential before any change to clinical practice.
Erola Pairo-Castineira; Sara Clohisey; Lucija Klaric; Andrew Bretherick; Konrad Rawlik; Nicholas Parkinson; Dorota Pasko; Susan Walker; Anne Richmond; Max Head Fourman; Clark D Russell; Andrew Law; James Furniss; Elvina Gountouna; Nicola Wrobel; Loukas Moutsianas; Bo Wang; Alison Meynert; Zhijian Yang; Ranran Zhai; Chenqing Zheng; Fiona Griffiths; Wilna Oosthuyzen; Graeme Grimes; Barbara Shih; Sean Keating; Marie Zechner; Chris Haley; David J. Porteous; Caroline Hayward; Julian Knight; Charlotte Summers; Manu Shankar-Hari; Paul Klenerman; Lance Turtle; Antonia Ho; Charles Hinds; Peter Horby; Alistair Nichol; David Maslove; Lowell Ling; Danny McAuley; Hugh Montgomery; Timothy Walsh; Xia Shen; Kathy Rowan; Angie Fawkes; Lee Murphy; Chris P. Ponting; Albert Tenesa; Mark Caulfield; Richard Scott; Peter J.M. Openshaw; Malcolm G. Semple; Veronique Vitart; James F. Wilson; J. Kenneth Baillie; The GenOMICC Investigators; The ISARIC-4C Investigators; The Covid-19 Human Genetics Initiative. Genetic mechanisms of critical illness in Covid-19. 2020, 1 .
AMA StyleErola Pairo-Castineira, Sara Clohisey, Lucija Klaric, Andrew Bretherick, Konrad Rawlik, Nicholas Parkinson, Dorota Pasko, Susan Walker, Anne Richmond, Max Head Fourman, Clark D Russell, Andrew Law, James Furniss, Elvina Gountouna, Nicola Wrobel, Loukas Moutsianas, Bo Wang, Alison Meynert, Zhijian Yang, Ranran Zhai, Chenqing Zheng, Fiona Griffiths, Wilna Oosthuyzen, Graeme Grimes, Barbara Shih, Sean Keating, Marie Zechner, Chris Haley, David J. Porteous, Caroline Hayward, Julian Knight, Charlotte Summers, Manu Shankar-Hari, Paul Klenerman, Lance Turtle, Antonia Ho, Charles Hinds, Peter Horby, Alistair Nichol, David Maslove, Lowell Ling, Danny McAuley, Hugh Montgomery, Timothy Walsh, Xia Shen, Kathy Rowan, Angie Fawkes, Lee Murphy, Chris P. Ponting, Albert Tenesa, Mark Caulfield, Richard Scott, Peter J.M. Openshaw, Malcolm G. Semple, Veronique Vitart, James F. Wilson, J. Kenneth Baillie, The GenOMICC Investigators, The ISARIC-4C Investigators, The Covid-19 Human Genetics Initiative. Genetic mechanisms of critical illness in Covid-19. . 2020; ():1.
Chicago/Turabian StyleErola Pairo-Castineira; Sara Clohisey; Lucija Klaric; Andrew Bretherick; Konrad Rawlik; Nicholas Parkinson; Dorota Pasko; Susan Walker; Anne Richmond; Max Head Fourman; Clark D Russell; Andrew Law; James Furniss; Elvina Gountouna; Nicola Wrobel; Loukas Moutsianas; Bo Wang; Alison Meynert; Zhijian Yang; Ranran Zhai; Chenqing Zheng; Fiona Griffiths; Wilna Oosthuyzen; Graeme Grimes; Barbara Shih; Sean Keating; Marie Zechner; Chris Haley; David J. Porteous; Caroline Hayward; Julian Knight; Charlotte Summers; Manu Shankar-Hari; Paul Klenerman; Lance Turtle; Antonia Ho; Charles Hinds; Peter Horby; Alistair Nichol; David Maslove; Lowell Ling; Danny McAuley; Hugh Montgomery; Timothy Walsh; Xia Shen; Kathy Rowan; Angie Fawkes; Lee Murphy; Chris P. Ponting; Albert Tenesa; Mark Caulfield; Richard Scott; Peter J.M. Openshaw; Malcolm G. Semple; Veronique Vitart; James F. Wilson; J. Kenneth Baillie; The GenOMICC Investigators; The ISARIC-4C Investigators; The Covid-19 Human Genetics Initiative. 2020. "Genetic mechanisms of critical illness in Covid-19." , no. : 1.
Quantitative and qualitative data derived from the analysis of genomes, genes, proteins or metabolites from tissue or cells are currently generated in huge volumes during biomedical research. Graphia is an open-source platform created for the graph-based analysis of such complex data, e.g. transcriptomics, proteomics, genomics data. The software imports data already defined as a network or a similarity matrix and is designed to rapidly visualise very large graphs in 2D or 3D space, providing a wide range of functionality for graph exploration. An extensive range of analysis algorithms, routines for graph transformation, and options for the visualisation of node and edge attributes are also available. Graphias core is extensible through the deployment of plugins, supporting rapid development of additional computational analyses and features necessary for a given analysis task or data source. A plugin for correlation network analysis is distributed with the core application, to support the generation of correlation graphs from any tabular matrix of continuous or discrete values. This provides a powerful analysis solution for the interpretation of high-dimensional data from many sources. Several use cases of Graphia are described, to showcase its wide range of applications. Graphia runs on all major desktop operating systems and is freely available to download from https://graphia.app/.
Thomas Freeman; Sebastian Horsewell; Anirudh Patir; Josh Harling-Lee; Tim Regan; Barbara B Shih; James Prendergast; David A Hume; Tim Angus. Graphia: A platform for the graph-based visualisation and analysis of complex data. 2020, 1 .
AMA StyleThomas Freeman, Sebastian Horsewell, Anirudh Patir, Josh Harling-Lee, Tim Regan, Barbara B Shih, James Prendergast, David A Hume, Tim Angus. Graphia: A platform for the graph-based visualisation and analysis of complex data. . 2020; ():1.
Chicago/Turabian StyleThomas Freeman; Sebastian Horsewell; Anirudh Patir; Josh Harling-Lee; Tim Regan; Barbara B Shih; James Prendergast; David A Hume; Tim Angus. 2020. "Graphia: A platform for the graph-based visualisation and analysis of complex data." , no. : 1.
Tanning (melanisation and epidermal thickening) is a photoprotective response to solar UVR exposure, but it has been unclear to what degree low-level exposures induce this, or whether this modifies the histological inflammatory response to UVR.
S. J. Felton; B. B. Shih; R. E. B. Watson; R. Kift; A. R. Webb; L. E. Rhodes. Photoprotection conferred by low level summer sunlight exposures against pro-inflammatory UVR insult. Photochemical & Photobiological Sciences 2020, 19, 810 -818.
AMA StyleS. J. Felton, B. B. Shih, R. E. B. Watson, R. Kift, A. R. Webb, L. E. Rhodes. Photoprotection conferred by low level summer sunlight exposures against pro-inflammatory UVR insult. Photochemical & Photobiological Sciences. 2020; 19 (6):810-818.
Chicago/Turabian StyleS. J. Felton; B. B. Shih; R. E. B. Watson; R. Kift; A. R. Webb; L. E. Rhodes. 2020. "Photoprotection conferred by low level summer sunlight exposures against pro-inflammatory UVR insult." Photochemical & Photobiological Sciences 19, no. 6: 810-818.
Ageing results in a decline in immune function. We showed previously that healthy older humans (>65 years old) have reduced antigen-specific cutaneous immunity to varicella zoster virus (VZV) antigen challenge. This was associated with p38 MAP kinase driven inflammation that was induced by mild tissue injury caused by the injection of the antigen itself. Here we show that non-specific injury induced by injection of air or saline into the skin of older adults recruits CCR2+CD14+ monocytes by CCL2 produced by senescent fibroblasts. These monocytes reduced TRM proliferation via secretion of prostaglandin E2 (PGE2). Pre-treatment with a p38-MAPK inhibitor (Losmapimod) in older adults in vivo significantly decreased CCL2 expression, recruitment of monocyte into the skin, COX2 expression and PGE2 production. This enhanced the VZV response in the skin. Therefore, local inflammation arising from interaction between senescent cells and monocytes leads to immune decline in the skin during ageing, a process that can be reversed.SummaryInflammation resulting from tissue injury blocks antigen-specific cutaneous immunity during ageing. Monocytes recruited to the skin inhibit TRM function through COX2-derived prostaglandin E2 production. Blocking inflammation and resulting prostaglandin E2 production with a p38-MAP kinase inhibitor significantly enhances cutaneous antigen-specific responses.
Emma S Chambers; Milica Vukmanovic-Stejic; Barbara B Shih; Hugh Trahair; Priya Subramanian; Oliver P Devine; James Glanville; Derek Gilroy; Malcolm Rustin; Tom C Freeman; Neil A Mabbott; Arne N Akbar. Monocyte-derived Prostaglandin E2 inhibits antigen-specific cutaneous immunity during ageing. 2020, 1 .
AMA StyleEmma S Chambers, Milica Vukmanovic-Stejic, Barbara B Shih, Hugh Trahair, Priya Subramanian, Oliver P Devine, James Glanville, Derek Gilroy, Malcolm Rustin, Tom C Freeman, Neil A Mabbott, Arne N Akbar. Monocyte-derived Prostaglandin E2 inhibits antigen-specific cutaneous immunity during ageing. . 2020; ():1.
Chicago/Turabian StyleEmma S Chambers; Milica Vukmanovic-Stejic; Barbara B Shih; Hugh Trahair; Priya Subramanian; Oliver P Devine; James Glanville; Derek Gilroy; Malcolm Rustin; Tom C Freeman; Neil A Mabbott; Arne N Akbar. 2020. "Monocyte-derived Prostaglandin E2 inhibits antigen-specific cutaneous immunity during ageing." , no. : 1.
RNA-Seq is a powerful transcriptome profiling technology enabling transcript discovery and quantification. Whilst most commonly used for gene-level quantification, the data can be used for the analysis of transcript isoforms. However, when the underlying transcript assemblies are complex, current visualization approaches can be limiting, with splicing events a challenge to interpret. Here, we report on the development of a graph-based visualization method as a complementary approach to understanding transcript diversity from short-read RNA-Seq data. Following the mapping of reads to a reference genome, a read-to-read comparison is performed on all reads mapping to a given gene, producing a weighted similarity matrix between reads. This is used to produce an RNA assembly graph, where nodes represent reads and edges similarity scores between them. The resulting graphs are visualized in 3D space to better appreciate their sometimes large and complex topology, with other information being overlaid on to nodes, e.g. transcript models. Here we demonstrate the utility of this approach, including the unusual structure of these graphs and how they can be used to identify issues in assembly, repetitive sequences within transcripts and splice variants. We believe this approach has the potential to significantly improve our understanding of transcript complexity.
Wan Fahmi Wan Mohamad Nazarie; Barbara Shih; Tim Angus; Mark W Barnett; Sz-Hau Chen; Kim M Summers; Karsten Klein; Geoffrey J Faulkner; Harpreet K Saini; Mick Watson; Stijn Van Dongen; Anton J Enright; Tom C Freeman. Visualization and analysis of RNA-Seq assembly graphs. Nucleic Acids Research 2019, 47, 7262 -7275.
AMA StyleWan Fahmi Wan Mohamad Nazarie, Barbara Shih, Tim Angus, Mark W Barnett, Sz-Hau Chen, Kim M Summers, Karsten Klein, Geoffrey J Faulkner, Harpreet K Saini, Mick Watson, Stijn Van Dongen, Anton J Enright, Tom C Freeman. Visualization and analysis of RNA-Seq assembly graphs. Nucleic Acids Research. 2019; 47 (14):7262-7275.
Chicago/Turabian StyleWan Fahmi Wan Mohamad Nazarie; Barbara Shih; Tim Angus; Mark W Barnett; Sz-Hau Chen; Kim M Summers; Karsten Klein; Geoffrey J Faulkner; Harpreet K Saini; Mick Watson; Stijn Van Dongen; Anton J Enright; Tom C Freeman. 2019. "Visualization and analysis of RNA-Seq assembly graphs." Nucleic Acids Research 47, no. 14: 7262-7275.
The immune composition of the tumor microenvironment regulates processes including angiogenesis, metastasis, and the response to drugs or immunotherapy. To facilitate the characterization of the immune component of tumors from transcriptomics data, a number of immune cell transcriptome signatures have been reported that are made up of lists of marker genes indicative of the presence a given immune cell population. The majority of these gene signatures have been defined through analysis of isolated blood cells. However, blood cells do not reflect the differentiation or activation state of similar cells within tissues, including tumors, and consequently markers derived from blood cells do not necessarily transfer well to tissues. To address this issue, we generated a set of immune gene signatures derived directly from tissue transcriptomics data using a network-based deconvolution approach. We define markers for seven immune cell types, collectively named ImSig, and demonstrate how these markers can be used for the quantitative estimation of the immune-cell content of tumor and non-tumor tissue samples. The utility of ImSig is demonstrated through the stratification of melanoma patients into subgroups of prognostic significance and the identification of immune cells with the use of single-cell RNA-Seq data derived from tumors. Use of ImSig is facilitated by an R package ('imsig').
Ajit J. Nirmal; Tim Regan; Barbara B. Shih; David A. Hume; Andrew H. Sims; Tom C. Freeman. Immune Cell Gene Signatures for Profiling the Microenvironment of Solid Tumors. Cancer Immunology Research 2018, 6, 1388 -1400.
AMA StyleAjit J. Nirmal, Tim Regan, Barbara B. Shih, David A. Hume, Andrew H. Sims, Tom C. Freeman. Immune Cell Gene Signatures for Profiling the Microenvironment of Solid Tumors. Cancer Immunology Research. 2018; 6 (11):1388-1400.
Chicago/Turabian StyleAjit J. Nirmal; Tim Regan; Barbara B. Shih; David A. Hume; Andrew H. Sims; Tom C. Freeman. 2018. "Immune Cell Gene Signatures for Profiling the Microenvironment of Solid Tumors." Cancer Immunology Research 6, no. 11: 1388-1400.
RNA-sequencing (RNA-Seq) is a powerful transcriptome profiling technology enabling transcript discovery and quantification. RNA-Seq data are large, and most commonly used as a source of genelevel quantification measurements, whilst the underlying assemblies of reads, if inspected, are usually viewed as sequence reads mapped on to a reference genome. Whilst sufficient for many needs, when the underlying transcript assemblies are complex, this visualisation approach can be limiting; errors in assembly can be difficult to spot and interpretation of splicing events is challenging.Here we report on the development of a graph-based visualisation method as a complementary approach to understanding transcript diversity and read assembly from short-read RNA-Seq data. Following the mapping of reads to the reference genome, read-to-read comparison is performed on all reads mapping to a given gene, producing a matrix of weighted similarity scores between reads. This is used to produce an RNA assembly graph where nodes represent reads derived from a cDNA and edges similarity scores between reads, above a defined threshold. Visualisation of resulting graphs is performed using Graphia Professional. This tool can render the often large and complex graph topologies that result from DNA/RNA sequence assembly in 3D space and supports info rmatio no verlay on to nodes, e.g. transcript models. We have also implemented an analysis pipeline for the creation of RNA assembly graphs with both a command-line and web-based interface that allows users to create and visualise these data. Here we demonstrate the utility of this approach on RNA-Seq data, including the unusual structure of these graphs and how they can be used to identify issues in assembly, repetitive sequences within transcripts and splice variants. We believe this approach has the potential to significantly improve our understanding of transcript complexity.
Fahmi W. Nazarie; Barbara B Shih; Tim Angus; Mark W. Barnett; Sz-Hau Chen; Kim M. Summers; Karsten Klein; Geoffrey J. Faulkner; Harpreet K. Saini; Mick Watson; Stijn Van Dongen; Anton J. Enright; Tom C. Freeman. Visualisation and analysis of RNA-Seq assembly graphs. 2018, 409573 .
AMA StyleFahmi W. Nazarie, Barbara B Shih, Tim Angus, Mark W. Barnett, Sz-Hau Chen, Kim M. Summers, Karsten Klein, Geoffrey J. Faulkner, Harpreet K. Saini, Mick Watson, Stijn Van Dongen, Anton J. Enright, Tom C. Freeman. Visualisation and analysis of RNA-Seq assembly graphs. . 2018; ():409573.
Chicago/Turabian StyleFahmi W. Nazarie; Barbara B Shih; Tim Angus; Mark W. Barnett; Sz-Hau Chen; Kim M. Summers; Karsten Klein; Geoffrey J. Faulkner; Harpreet K. Saini; Mick Watson; Stijn Van Dongen; Anton J. Enright; Tom C. Freeman. 2018. "Visualisation and analysis of RNA-Seq assembly graphs." , no. : 409573.
Public health guidance recommends limiting sun exposure to sub-sunburn levels, but it is unknown whether these can gain vitamin D (for musculoskeletal health) while avoiding epidermal DNA damage (initiates skin cancer). Well-characterized healthy humans of all skin types (I–VI, lightest to darkest skin) were exposed to a low-dose series of solar simulated UVR of 20%–80% their individual sunburn threshold dose (minimal erythema dose). Significant UVR dose responses were seen for serum 25-hydroxyvitamin D and whole epidermal cyclobutane pyrimidine dimers (CPDs), with as little as 0.2 minimal erythema dose concurrently producing 25-hydroxyvitamin D and CPD. Fractional MEDs generated equivalent levels of whole epidermal CPD and 25-hydroxyvitamin D across all skin types. Crucially, we showed an epidermal gradient of CPD formation strongly correlated with skin darkness (r = 0.74, P< 0.0001), which reflected melanin content and showed increasing protection across the skin types, ranging from darkest skin, where high CPD levels occurred superficially, with none in the germinative basal layer, to lightest skin, where CPD levels were induced evenly across the epidermal depth. People with darker skin can be encouraged to use sub-sunburn UVR-exposure to enhance their vitamin D. In people with lighter skin, basal cell damage occurs concurrent with vitamin D synthesis at exquisitely low UVR levels, providing an explanation for their high skin cancer incidence; greater caution is required.
Barbara B. Shih; Mark D. Farrar; Marcus S. Cooke; Joanne Osman; Abigail K. Langton; Richard Kift; Ann R. Webb; Jacqueline L. Berry; Rachel E.B. Watson; Andy Vail; Frank R. de Gruijl; Lesley E. Rhodes. Fractional Sunburn Threshold UVR Doses Generate Equivalent Vitamin D and DNA Damage in Skin Types I–VI but with Epidermal DNA Damage Gradient Correlated to Skin Darkness. Journal of Investigative Dermatology 2018, 138, 2244 -2252.
AMA StyleBarbara B. Shih, Mark D. Farrar, Marcus S. Cooke, Joanne Osman, Abigail K. Langton, Richard Kift, Ann R. Webb, Jacqueline L. Berry, Rachel E.B. Watson, Andy Vail, Frank R. de Gruijl, Lesley E. Rhodes. Fractional Sunburn Threshold UVR Doses Generate Equivalent Vitamin D and DNA Damage in Skin Types I–VI but with Epidermal DNA Damage Gradient Correlated to Skin Darkness. Journal of Investigative Dermatology. 2018; 138 (10):2244-2252.
Chicago/Turabian StyleBarbara B. Shih; Mark D. Farrar; Marcus S. Cooke; Joanne Osman; Abigail K. Langton; Richard Kift; Ann R. Webb; Jacqueline L. Berry; Rachel E.B. Watson; Andy Vail; Frank R. de Gruijl; Lesley E. Rhodes. 2018. "Fractional Sunburn Threshold UVR Doses Generate Equivalent Vitamin D and DNA Damage in Skin Types I–VI but with Epidermal DNA Damage Gradient Correlated to Skin Darkness." Journal of Investigative Dermatology 138, no. 10: 2244-2252.
Numerous studies have explored the altered transcriptional landscape associated with skin diseases to understand the nature of these disorders. However, data interpretation represents a significant challenge due to a lack of good maker sets for many of the specialized cell types that make up this tissue, whose composition may fundamentally alter during disease. Here we have sought to derive expression signatures that define the various cell types and structures that make up human skin, and demonstrate how they can be used to aid the interpretation of transcriptomic data derived from this organ. Two large normal skin transcriptomic datasets were identified, one RNA-seq (n = 578), the other microarray (n = 165), quality controlled and subjected separately to network-based analyses to identify clusters of robustly co-expressed genes. The biological significance of these clusters was then assigned using a combination of bioinformatics analyses, literature, and expert review. After cross comparison between analyses, 20 gene signatures were defined. These included expression signatures for hair follicles, glands (sebaceous, sweat, apocrine), keratinocytes, melanocytes, endothelia, muscle, adipocytes, immune cells, and a number of pathway systems. Collectively, we have named this resource SkinSig. SkinSig was then used in the analysis of transcriptomic datasets for 18 skin conditions, providing in-context interpretation of these data. For instance, conventional analysis has shown there to be a decrease in keratinization and fatty metabolism with age; we more accurately define these changes to be due to loss of hair follicles and sebaceous glands. SkinSig also highlighted the over-/under-representation of various cell types in skin diseases, reflecting an influx in immune cells in inflammatory disorders and a relative reduction in other cell types. Overall, our analyses demonstrate the value of this new resource in defining the functional profile of skin cell types and appendages, and in improving the interpretation of disease data. © 2016 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.
Barbara B Shih; Ajit Johnson Nirmal; Denis J Headon; Arne Akbar; Neil A Mabbott; Tom C Freeman. Derivation of marker gene signatures from human skin and their use in the interpretation of the transcriptional changes associated with dermatological disorders. The Journal of Pathology 2017, 241, 600 -613.
AMA StyleBarbara B Shih, Ajit Johnson Nirmal, Denis J Headon, Arne Akbar, Neil A Mabbott, Tom C Freeman. Derivation of marker gene signatures from human skin and their use in the interpretation of the transcriptional changes associated with dermatological disorders. The Journal of Pathology. 2017; 241 (5):600-613.
Chicago/Turabian StyleBarbara B Shih; Ajit Johnson Nirmal; Denis J Headon; Arne Akbar; Neil A Mabbott; Tom C Freeman. 2017. "Derivation of marker gene signatures from human skin and their use in the interpretation of the transcriptional changes associated with dermatological disorders." The Journal of Pathology 241, no. 5: 600-613.
The keloid lesion is recognised as a spatially heterogeneous mass both in cellular and acellular composition and biological activity. Here, we have utilised a bioinformatic approach to determine whether this spatial heterogeneity is also evident at the molecular level and to identify key upstream regulators of signalling pathways enriched in the lesion in a spatially-restricted manner. Differentially expressed genes (20% change, p < 0.05) obtained from microarray datasets derived from whole keloid biopsies and in vitro-cultured keloid fibroblasts, both from distinct regions of the keloid lesion (leading edge, centre, and top) have been analysed to show that the TGFβ family plays a significant but spatially dependent role in regulation of keloid gene expression. Furthermore, we have identified additional upstream signalling molecules involved in driving keloid biology and provide information on therapeutic targets whose modulation might be expected to lead to significant therapeutic efficacy.
A Taylor; D Budd; B Shih; O Seifert; A Beaton; T Wright; M Dempsey; F Kelly; J Egerton; R Marshall; N Aston; A Bayat. Transforming Growth Factor Beta Gene Signatures are Spatially Enriched in Keloid Tissue Biopsies and Ex vivo-Cultured Keloid Fibroblasts. Acta Dermato Venereologica 2017, 97, 10 -16.
AMA StyleA Taylor, D Budd, B Shih, O Seifert, A Beaton, T Wright, M Dempsey, F Kelly, J Egerton, R Marshall, N Aston, A Bayat. Transforming Growth Factor Beta Gene Signatures are Spatially Enriched in Keloid Tissue Biopsies and Ex vivo-Cultured Keloid Fibroblasts. Acta Dermato Venereologica. 2017; 97 (1):10-16.
Chicago/Turabian StyleA Taylor; D Budd; B Shih; O Seifert; A Beaton; T Wright; M Dempsey; F Kelly; J Egerton; R Marshall; N Aston; A Bayat. 2017. "Transforming Growth Factor Beta Gene Signatures are Spatially Enriched in Keloid Tissue Biopsies and Ex vivo-Cultured Keloid Fibroblasts." Acta Dermato Venereologica 97, no. 1: 10-16.
Clinical consensus is that debridement is necessary for successful application of dermal skin substitutes (DSS) to chronic wounds. The aim was to identify commonly expressed genes associated with wound healing in untreated acute wounds and chronic wounds treated with wound debridement followed by DSS. Cutaneous biopsies were taken at two time points from untreated acute and chronic wounds and from chronic wounds treated with DSS following debridement. Microarray analysis identified significant differences (p<0.05) related to proliferation (HIPK2, LGR4, FGFR1, SRRT), migration (RHOC, PRPF40A, FGFR1), differentiation (TCF4, COL13A1, GNPTAB, HUWE1, FGFR1), angiogenesis (HIPK2, CASP8), extracellular matrix organisation (VWA1) and apoptosis (BBC3, HIPK2, KLF11, PSME3, MSFD10, TOP2A, MLH1, CASP8, PDIA3, XAF1) when comparing untreated chronic wounds to chronic wounds treated with DSS, with similar expression levels compared to untreated acute wounds. Chronic wounds treated with debridement followed by DSS resemble untreated acute wounds at a genomic level. These novel findings, albeit with limited clinical specimen numbers, strengthen the recommendation to transform chronic into acute wounds prior to application of DSS. This article is protected by copyright. All rights reserved.
Mohammed Ashrafi; Anil Sebastian; Barbara Shih; Nicholas Greaves; Teresa Alonso‐Rasgado; Mohamed Baguneid; Ardeshir Bayat. Whole genome microarray data of chronic wound debridement prior to application of dermal skin substitutes. Wound Repair and Regeneration 2016, 24, 870 -875.
AMA StyleMohammed Ashrafi, Anil Sebastian, Barbara Shih, Nicholas Greaves, Teresa Alonso‐Rasgado, Mohamed Baguneid, Ardeshir Bayat. Whole genome microarray data of chronic wound debridement prior to application of dermal skin substitutes. Wound Repair and Regeneration. 2016; 24 (5):870-875.
Chicago/Turabian StyleMohammed Ashrafi; Anil Sebastian; Barbara Shih; Nicholas Greaves; Teresa Alonso‐Rasgado; Mohamed Baguneid; Ardeshir Bayat. 2016. "Whole genome microarray data of chronic wound debridement prior to application of dermal skin substitutes." Wound Repair and Regeneration 24, no. 5: 870-875.
Barbara B. Shih; Donald Allan; Frank R. De Gruijl; Lesley E. Rhodes. Robust Detection of Minimal Sunburn in Pigmented Skin by 785 nm Laser Speckle Contrast Imaging of Blood Flux. Journal of Investigative Dermatology 2015, 135, 1197 -1199.
AMA StyleBarbara B. Shih, Donald Allan, Frank R. De Gruijl, Lesley E. Rhodes. Robust Detection of Minimal Sunburn in Pigmented Skin by 785 nm Laser Speckle Contrast Imaging of Blood Flux. Journal of Investigative Dermatology. 2015; 135 (4):1197-1199.
Chicago/Turabian StyleBarbara B. Shih; Donald Allan; Frank R. De Gruijl; Lesley E. Rhodes. 2015. "Robust Detection of Minimal Sunburn in Pigmented Skin by 785 nm Laser Speckle Contrast Imaging of Blood Flux." Journal of Investigative Dermatology 135, no. 4: 1197-1199.
Breast capsular contracture formation following silicone implant augmentation/reconstruction is a common complication that remains poorly understood. The aim of this study was to identify potential biomarkers implicated in breast capsular contracture formation by using, for the first time, whole genome arrays. Biopsy samples were taken from 18 patients (23 breast capsules) with Baker Grade I-II (Control) and Baker Grade III-IV (Contracted). Whole genome microarrays were performed and six significantly dysregulated genes were selected for further validation with quantitative reverse transcriptase polymerase chain reaction and immunohistochemistry. Hematoxylin and eosin was also carried out to compare the histological characteristics of control and contracted samples. Microarray results showed that aggrecan, tissue inhibitor of metalloproteinase 4 (TIMP4), and tumor necrosis factor superfamily (ligand) member 11 were significantly down-regulated in contracted capsules; while matrix metallopeptidase 12, serum amyloid A 1, and interleukin 8 (IL8) were significantly up-regulated. The dysregulation of aggrecan, tumor necrosis factor superfamily (ligand) member 11, TIMP4, and IL8 was validated by quantitative reverse transcriptase polymerase chain reaction (p < 0.05). Immunohistochemistry confirmed an increased protein expression for IL8 and matrix metallopeptidase 12 in contracted capsules (p < 0.05), and decreased protein expression of TIMP4 (p < 0.05). This study has shown, for the first time, a number of unique biomarkers of significance in capsular contracture formation. IL8 and TIMP4 may serve as potential key diagnostic, therapeutic, and prognostic biomarkers in capsular contracture formation.
Daniel J. T. Kyle; Alison G. Harvey; Barbara Shih; Kian T. Tan; Iskander H. Chaudhry; Ardeshir Bayat. Identification of molecular phenotypic descriptors of breast capsular contracture formation using informatics analysis of the whole genome transcriptome. Wound Repair and Regeneration 2013, 21, 762 -769.
AMA StyleDaniel J. T. Kyle, Alison G. Harvey, Barbara Shih, Kian T. Tan, Iskander H. Chaudhry, Ardeshir Bayat. Identification of molecular phenotypic descriptors of breast capsular contracture formation using informatics analysis of the whole genome transcriptome. Wound Repair and Regeneration. 2013; 21 (5):762-769.
Chicago/Turabian StyleDaniel J. T. Kyle; Alison G. Harvey; Barbara Shih; Kian T. Tan; Iskander H. Chaudhry; Ardeshir Bayat. 2013. "Identification of molecular phenotypic descriptors of breast capsular contracture formation using informatics analysis of the whole genome transcriptome." Wound Repair and Regeneration 21, no. 5: 762-769.
Chronic wounds are common and lead to significant patient morbidity. A better understanding of their pathogenesis and relevant biomarkers are required. We compared acute and chronic wounds in the same individual using noninvasive imaging including spectrophotometric intracutaneous analysis (SIAscopy) and full-field laser perfusion imaging. Gene expression analysis was also performed on sequential biopsies. Whole genome gene expression microarray analysis (44k), quantitative polymerase chain reaction, and immunohistochemistry were carried out to determine gene expression levels in tissue biopsies. Fifteen Caucasian patients with chronic venous ulcers had biopsies of the wound edges and simultaneously had an acute wound created on their upper arm on days 0, 7, and 14. SIAscopy revealed increased levels of melanin (p < 0.001), reduced levels of collagen (p < 0.001), and hemoglobin (p = 0.022) in chronic wounds. Microarray and subsequent quantitative polymerase chain reaction analysis confirmed an overall differential expression in acute and chronic wounds for several genes. Significantly higher levels of inhibin, beta A (INHBA) expression were confirmed in the dermis of chronic wounds (p < 0.05). Additionally, INHBA and thrombospondin 1 messenger RNA levels significantly correlated with SIAscopy measurements (p < 0.05). This unique study has showed aberrant expression of INHBA in chronic wounds using a sequential biopsy model of chronic vs. acute wounds in the same individual.
Barbara Shih; Muhammad J. Sultan; Iskander H. Chaudhry; Kian T. Tan; Kavan Johal; Arshiya Marstan; Melody Tsai; Mohamed Baguneid; Ardeshir Bayat. Identification of biomarkers in sequential biopsies of patients with chronic wounds receiving simultaneous acute wounds: A genetic, histological, and noninvasive imaging study. Wound Repair and Regeneration 2012, 20, 757 -769.
AMA StyleBarbara Shih, Muhammad J. Sultan, Iskander H. Chaudhry, Kian T. Tan, Kavan Johal, Arshiya Marstan, Melody Tsai, Mohamed Baguneid, Ardeshir Bayat. Identification of biomarkers in sequential biopsies of patients with chronic wounds receiving simultaneous acute wounds: A genetic, histological, and noninvasive imaging study. Wound Repair and Regeneration. 2012; 20 (5):757-769.
Chicago/Turabian StyleBarbara Shih; Muhammad J. Sultan; Iskander H. Chaudhry; Kian T. Tan; Kavan Johal; Arshiya Marstan; Melody Tsai; Mohamed Baguneid; Ardeshir Bayat. 2012. "Identification of biomarkers in sequential biopsies of patients with chronic wounds receiving simultaneous acute wounds: A genetic, histological, and noninvasive imaging study." Wound Repair and Regeneration 20, no. 5: 757-769.
Dupuytren's disease (DD) is a common fibroproliferative disorder affecting the palmar fascia, which may lead to permanent contracture of the affected digit. Profiling studies investigating DD at whole-genomic, transcriptomic and proteomic levels have been carried out, from which large numbers of candidate genes potentially involved in DD have been reported. This review focuses on identifying genes reported by multiple studies or validated by multiple experimental techniques, as well as signalling pathways suggested to contribute to DD. Meta-analysis was also carried out on three microarray datasets. Twenty-one genes were found to be reported as dysregulated in multiple gene expression microarrays, seven of which have been further validated by other experimental methods. Sixty-four genes determined to be dsyregulated by meta-analysis correlate to those reported by published microarray studies. In addition, several pathways have been proposed to be involved in DD by whole-genome or global expression profiling. Further investigation in these genes and pathways, and correlating them to genotypes or environmental factors for DD, may aid in further elucidation of mechanisms involved in DD pathogenesis.
Barbara Shih; S. Watson; Ardeshir Bayat. Whole genome and global expression profiling of Dupuytren's disease: systematic review of current findings and future perspectives. Annals of the Rheumatic Diseases 2012, 71, 1440 -1447.
AMA StyleBarbara Shih, S. Watson, Ardeshir Bayat. Whole genome and global expression profiling of Dupuytren's disease: systematic review of current findings and future perspectives. Annals of the Rheumatic Diseases. 2012; 71 (9):1440-1447.
Chicago/Turabian StyleBarbara Shih; S. Watson; Ardeshir Bayat. 2012. "Whole genome and global expression profiling of Dupuytren's disease: systematic review of current findings and future perspectives." Annals of the Rheumatic Diseases 71, no. 9: 1440-1447.
Dupuytren's disease is a common fibroproliferative disorder with an unknown etiology. Emerging evidence suggests a strong genetic component involved in the manifestation of the disease. This study aims to investigate the potential involvement of copy number variations in Dupuytren's disease pathogenesis. Array-based comparative genomic hybridization (NimbleGen Human CGH 2.1 M) was utilized to compare DNA from (1) nodules versus internal control (patient's blood; n = 4) and (2) nodules (n = 4) versus external control (commercial reference DNA pooled from 10 donors). Analysis was carried out using Nexus 5.1 (BioDiscovery, El Segundo, Calif.) with the inclusion of additional results from previously published array-based comparative genomic hybridization. Copy number variations were considered to be common in Dupuytren's disease if the overlap was statistically significant and they were present in the majority (75 to 87.5 percent when compared with controls) of Dupuytren's disease nodules. The copy number variations loci were also compared with recently published genome wide-association studies. Common copy number variations were further validated using quantitative polymerase chain reaction. DNA from 25 Dupuytren's disease cases and 30 external controls were used in the quantitative polymerase chain reaction validation. In addition, gene expression was compared between Dupuytren's disease nodules and internal controls (transverse palmar fascia; n = 7). Five common copy number variations, on chromosome 17q12, 1p31.1, 20p13, 7p14.1, and 14q11.2, were identified by array-based comparative genomic hybridization. Significantly higher copy numbers of copy number variations at chromosome 7p14.1 and 14q11.2 in Dupuytren's disease were confirmed in quantitative polymerase chain reaction validation. Matrix metalloproteinase-14 and secreted frizzled-related protein 4 (near a polymorphism recently associated with Dupuytren's disease) were significantly up-regulated in nodules. This study demonstrated an association between Dupuytren's disease and copy number variations at chromosomes 7p14.1 and 14q11.2.
Barbara Shih; May Tassabehji; James Stewart Watson; Ardeshir Bayat. DNA Copy Number Variations at Chromosome 7p14.1 and Chromosome 14q11.2 Are Associated with Dupuytrenʼs Disease. Plastic and Reconstructive Surgery 2012, 129, 921 -932.
AMA StyleBarbara Shih, May Tassabehji, James Stewart Watson, Ardeshir Bayat. DNA Copy Number Variations at Chromosome 7p14.1 and Chromosome 14q11.2 Are Associated with Dupuytrenʼs Disease. Plastic and Reconstructive Surgery. 2012; 129 (4):921-932.
Chicago/Turabian StyleBarbara Shih; May Tassabehji; James Stewart Watson; Ardeshir Bayat. 2012. "DNA Copy Number Variations at Chromosome 7p14.1 and Chromosome 14q11.2 Are Associated with Dupuytrenʼs Disease." Plastic and Reconstructive Surgery 129, no. 4: 921-932.