B cells are important in the pathogenesis of many, and perhaps all, immune-mediated diseases (IMD... more B cells are important in the pathogenesis of many, and perhaps all, immune-mediated diseases (IMDs). Each B cell expresses a single B cell receptor (BCR) 1 , with the diverse range of BCRs expressed by an individual's total B cell population being termed the "BCR repertoire". Our understanding of the BCR repertoire in the context of IMDs is incomplete, and defining this could reveal new insights into pathogenesis and therapy. We therefore compared the BCR repertoire in systemic lupus erythematosus (SLE), ANCA-associated vasculitis (AAV), Crohn's disease (CD), Behçet's disease (BD), eosinophilic granulomatosis with polyangiitis (EGPA) and IgA vasculitis (IgAV), analysing BCR clonality, and immunoglobulin heavy chain gene (IGHV) and, in particular, isotype usage. An IgA-dominated increased clonality in SLE and CD, together with skewed IGHV gene usage in these and other diseases, suggested a microbial contribution to pathogenesis. Different immunosuppressive treatment had specific and distinct impacts on the repertoire; B cells persisting after rituximab were predominately isotype-switched and clonally expanded, the inverse of those persisting after mycophenolate mofetil. A comparative analysis of Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:
Choosing the best treatment-plan for each individual patient requires accurate forecasts of their... more Choosing the best treatment-plan for each individual patient requires accurate forecasts of their outcome trajectories as a function of the treatment, over time. While large observational data sets constitute rich sources of information to learn from, they also contain biases as treatments are rarely assigned randomly in practice. To provide accurate and unbiased forecasts, we introduce the Disentangled Counterfactual Recurrent Network (DCRN), a novel sequence-tosequence architecture that estimates treatment outcomes over time by learning representations of patient histories that are disentangled into three separate latent factors: a treatment factor, influencing only treatment selection; an outcome factor, influencing only the outcome; and a confounding factor, influencing both. With an architecture that is completely inspired by the causal structure of treatment influence over time, we advance forecast accuracy and disease understanding, as our architecture allows for practitioners to infer which patient features influence which part in a patient's trajectory, contrasting other approaches in this domain. We demonstrate that DCRN outperforms current state-of-the-art methods in forecasting treatment responses, on both real and simulated data.
The endothelial cell has a remarkable ability for sub-specialisation, adapted to the needs of a v... more The endothelial cell has a remarkable ability for sub-specialisation, adapted to the needs of a variety of vascular beds. The role of developmental programming versus the tissue contextual environment for this specialization is not well understood. Here we describe a hierarchy of expression of HOX genes associated with endothelial cell origin and location. In initial microarray studies, differential gene expression was examined in two endothelial cell lines: blood derived outgrowth endothelial cells (BOECs) and pulmonary artery endothelial cells. This suggested shared and differential patterns of HOX gene expression between the two endothelial lines. For example, this included a cluster on chromosome 2 of HOXD1, HOXD3, HOXD4, HOXD8 and HOXD9 that was expressed at a higher level in BOECs. Quantative PCR confirmed the higher expression of these HOXs in BOECs, a pattern that was shared by a variety of microvascular endothelial cell lines. Subsequently, we analysed publically available microarrays from a variety of adult cell and tissue types using the whole ''HOX transcriptome'' of all 39 HOX genes. Using hierarchical clustering analysis the HOX transcriptome was able to discriminate endothelial cells from 61 diverse human cell lines of various origins. In a separate publically available microarray dataset of 53 human endothelial cell lines, the HOX transcriptome additionally organized endothelial cells related to their organ or tissue of origin. Human tissue staining for HOXD8 and HOXD9 confirmed endothelial expression and also supported increased microvascular expression of these HOXs. Together these observations suggest a significant involvement of HOX genes in endothelial cell positional identity.
Background: Although numerous investigations have compared gene expression microarray platforms, ... more Background: Although numerous investigations have compared gene expression microarray platforms, preprocessing methods and batch correction algorithms using constructed spike-in or dilution datasets, there remains a paucity of studies examining the properties of microarray data using diverse biological samples. Most microarray experiments seek to identify subtle differences between samples with variable background noise, a scenario poorly represented by constructed datasets. Thus, microarray users lack important information regarding the complexities introduced in real-world experimental settings. The recent development of a multiplexed, digital technology for nucleic acid measurement enables counting of individual RNA molecules without amplification and, for the first time, permits such a study. Results: Using a set of human leukocyte subset RNA samples, we compared previously acquired microarray expression values with RNA molecule counts determined by the nCounter Analysis System (NanoString Technologies) in selected genes. We found that gene measurements across samples correlated well between the two platforms, particularly for high-variance genes, while genes deemed unexpressed by the nCounter generally had both low expression and low variance on the microarray. Confirming previous findings from spike-in and dilution datasets, this "gold-standard" comparison demonstrated signal compression that varied dramatically by expression level and, to a lesser extent, by dataset. Most importantly, examination of three different cell types revealed that noise levels differed across tissues. Conclusions: Microarray measurements generally correlate with relative RNA molecule counts within optimal ranges but suffer from expression-dependent accuracy bias and precision that varies across datasets. We urge microarray users to consider expression-level effects in signal interpretation and to evaluate noise properties in each dataset independently.
Wolters Kluwer Health may email you for journal alerts and information, but is committed to maint... more Wolters Kluwer Health may email you for journal alerts and information, but is committed to maintaining your privacy and will not share your personal information without your express consent. For more information, please refer to our Privacy Policy. ... Skip Navigation Links Home > February ...
Previous research has generated examples of how genetic and environmental factors can interact to... more Previous research has generated examples of how genetic and environmental factors can interact to create risk for psychopathology. Using a gene-by-environment (G X E) interaction design, we tested whether three polymorphisms in the dopamine transporter gene (DAT1, also referred to as SLC6A3, located at 5p15.33) interacted with maternal parenting style to predict first-onset episodes of depression. Participants were male adolescents (N = 176) recruited from a juvenile detention center in northern Russia. As hypothesized, one of the polymorphisms (rs40184) moderated the effect of perceived maternal rejection on the onset of major depressive disorder, as well as on suicidal ideation. Further, this G X E interaction was specific to depression; it did not predict clinically significant anxiety. These results highlight the need for further research investigating the moderating effects of dopaminergic genes on depression.
Systemic Lupus Erythematosus (SLE) is a chronic autoimmune disorder characterized by broad clinic... more Systemic Lupus Erythematosus (SLE) is a chronic autoimmune disorder characterized by broad clinical manifestations including cardiovascular and renal complications with periodic disease flares and significant morbidity and mortality. One of the main contributing factors to the pathology of SLE is the accumulation and impaired clearance of immune complexes of which the principle components are host auto-antigens and antibodies. The contribution of host lipids to the formation of these autoimmune complexes remains poorly defined. The aim of the present study was to identify and analyze candidate lipid autoantigens and their corresponding anti-lipid antibody responses in a well-defined SLE patient cohort using a combination of immunological and biophysical techniques. Disease monitoring in the SLE cohort was undertaken with serial British Isles Lupus Assessment Group (BILAG) scoring. Correlations between specific lipid/anti-lipid responses were investigated as disease activity developed from active flares to quiescent during a follow up period. We report a significant negative correlation between anti-lipid antibodies for 24S-hydroxycholesterol, cardiolipin and phosphatidylserine with SLE disease activity. Taken together, these data suggest that lipid autoantigens represent a new family of biomarkers that can be employed to monitor disease activity plus the efficacy of therapeutic intervention in SLE.
Autoimmune diseases are common and debilitating, but their severe manifestations could be reduced... more Autoimmune diseases are common and debilitating, but their severe manifestations could be reduced if biomarkers were available to allow individual tailoring of the potentially toxic immunosuppressive therapy required for their control. Gene expression-based biomarkers facilitating individual tailoring of chemotherapy in cancer, but not autoimmunity, have been identified and translated into clinical practice1,2. We show that transcriptional profiling of purified CD8 T cells, which avoids the confounding influences of unseparated cells3,4, identifies two distinct patient subgroups predicting long-term prognosis in two different autoimmune diseases, anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV), a chronic, severe disease characterized by inflammation of medium and small blood vessels5, and systemic lupus erythematosus (SLE), characterized by autoantibodies, immune complex deposition and diverse clinical manifestations ranging from glomerulonephritis to neurological dysfunction6. We show that genes defining the poor prognostic group are enriched for genes of the IL7R pathway, TCR signalling and those expressed by memory T cells. Furthermore, the poor prognostic group is associated with an expanded CD8 T cell memory population. These subgroups, which are also found in the normal population and can be identified by measuring expression of only three genes, raise the prospect of individualized therapy and suggest novel potential therapeutic targets in autoimmunity.
ABSTRACT Autoimmune diseases are common and debilitating, but their severe manifestations could b... more ABSTRACT Autoimmune diseases are common and debilitating, but their severe manifestations could be reduced if biomarkers were available to allow individual tailoring of the potentially toxic immunosuppressive therapy required for their control. Gene expression-based biomarkers have been identified and translated into clinical practice in cancer, but not autoimmunity. We show that transcriptional profiling of purified CD8 T cells, which avoids the confounding influences of unseparated cells, identifies two distinct patient subgroups predicting long-term prognosis in four distinct autoimmune diseases: anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis, systemic lupus erythematosus, ulcerative colitis, and Crohn's disease.While the correlation of this signature with prognosis raises the prospect of individualised therapy in autoimmunity, the biological mechanism underlying its association with relapsing disease may indicate novel therapeutic targets. Acute exposure to foreign antigen results in persistent antigen-specific cellular immunity, but during chronic viral infection persistent antigen exposure may exhaust that response. This exhausted CD8 T-cell phenotype may itself be defined by a distinct transcriptional signature. We show that the same signature seen in chronic viral infection can be seen in chronic self antigen exposure during autoimmunity, with one exception: whereas exhaustion results in viral persistence it predicts favourable outcome in autoimmunity. Provision of adequate CD4 help can prevent or overcome the development of CD8 exhaustion resulting in clearance of chronic viral infection. Consistent with this observation, we show that the CD8 exhaustion signature in autoimmunity is inversely correlated with that of concurrent CD4 help and can be robustly identified in mixed cell populations. This has allowed independent validation of this marker on published datasets, confirming prediction of clinical outcome in a total of 1070 samples from 429 individuals spanning infection (HIV, dengue virus, hepatitis C), vaccination (yellow fever, malaria, influenza), and autoimmunity (type 1 diabetes, ANCA–associated vasculitis, systemic lupus erythematosus, Crohn's disease, and ulcerative colitis).FundingWellcome Trust.
Endothelial cells have a remarkable ability for subspecialisation, adapting to the needs of a var... more Endothelial cells have a remarkable ability for subspecialisation, adapting to the needs of a variety of vascular beds. It is not known how much of this subspecialisation is related to developmental programming versus the environment in which the endothelial cell fi nds itself. In transcriptomic studies of endothelial cells we have noted a hierarchy of HOX gene expression which predicts endothelial cell specifi cation and fate. HOX genes are well described as masterregulators of positional identity, predominately in the developing embryo.
ABSTRACT Background The clinical course of autoimmune and infectious disease varies greatly even ... more ABSTRACT Background The clinical course of autoimmune and infectious disease varies greatly even between individuals with the same condition. An understanding of the molecular basis for this heterogeneity could lead to substantial improvements in both monitoring and treatment. Since persistent antigen is targeted in both autoimmunity and chronic infection, similar pathways could dictate the success of that response. However, whereas a successful response to pathogen results in clearance and recovery from disease, a robust response to self-antigen might drive relapsing autoimmunity. During chronic viral infection, CD8 T cells develop progressive loss of function in a process termed exhaustion, characterised by high expression of inhibitory receptors (such as programmed cell death 1 [PD1]) and low expression of memory markers (such as interleukin-7 receptor). These can serve both as biomarkers of viral progression and as targets for therapy. We sought to determine whether an analogous exhausted state exists during autoimmune responses targeting persistent self-antigen. Methods To identify gene expression signatures associated with clinical outcome, we used weighted gene coexpression network analysis of concurrently sampled CD4 and CD8 T-cell transcriptomes from patients with four distinct autoimmune diseases (23 with systemic lupus erythematosus [SLE], 44 with anti-neutrophil cytoplasmic antibody-associated vasculitis [AAV], and 56 with Crohn's disease or ulcerative colitis) and with chronic viral infection (42 with HIV infection). Association with clinical outcome was independently validated in 1145 samples from 504 individuals with infectious disease (HIV, hepatitis C virus, dengue), during vaccination (malaria, yellow fever, influenza), and autoimmunity (type 1 diabetes, SLE, AAV, Crohn's disease and ulcerative colitis, and idiopathic pulmonary fibrosis). We used an artificial antigen presenting cell to modify provision of co-stimulatory and inhibitory signals (as suggested by the network analysis) to primary human T cells during in-vitro differentiation. In this way we tested whether expression signatures associated with clinical outcome might be modified by manipulating co-stimulatory signals and whether they reflected changes in T-cell function. Findings Network analysis of CD8 T-cell transcriptomes identified a common signature of T-cell exhaustion during responses to chronic viral and self-antigen. However, whereas exhausted responses were associated with poor viral clearance, they predicted a less severe course in multiple autoimmune diseases. In autoimmunity, we found that where CD8 T-cell exhaustion was high a concurrent signature of CD4 T-cell co-stimulation was low. We used this signature to identify CD2 as a co-stimulatory signal preventing the development of T-cell exhaustion during controlled in-vitro assays, recreating expression patterns seen in patients with severe autoimmunity and leading to enhanced T-cell survival during primary and repeated stimulation. Finally, exogenous signals through the inhibitory receptor PD1 restricted development of this robust population. Interpretation Here we show that the process of CD8 T-cell exhaustion is important not only in facilitating viral persistence during chronic infection, but also in shaping the immune response to persistent self-antigen during autoimmunity. The observed correlation between T-cell exhaustion, co-stimulation, and subsequent prognosis increases our understanding of the pathogenesis of severe autoimmune disease and might help stratify patients for personalised therapy. Furthermore, our data suggest that future modulation of T-cell exhaustion may lead to new therapeutic opportunities in autoimmunity. Funding Wellcome Trust, Biomedical Research Centre, National Institute for Health Research.
Journal of the American Society of Nephrology, 2012
The involvement of autoantibodies to human lysosome-associated membrane protein-2 (hLAMP-2) in an... more The involvement of autoantibodies to human lysosome-associated membrane protein-2 (hLAMP-2) in anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis is controversial because of the absence of confirmatory data subsequent to the initial reports of their high prevalence in this disease. We characterized three assays for anti-hLAMP-2 antibodies: ELISA and Western blotting assays using unglycosylated recombinant hLAMP-2 expressed in Escherichia coli, and an indirect immunofluorescence assay using stably transfected ldlD cells that expressed glycosylated full-length hLAMP-2 on the plasma membrane. The assays detected autoantibodies to hLAMP-2 in human sera reproducibly and with comparable sensitivity and the assays gave the same results in 80.5% of the test panel of 40 selected positive and negative sera. In untreated patients at presentation, the frequencies of autoantibodies to LAMP-2 were 89%, 91%, and 80%, respectively, among three groups of patients with ANCA-associated vasculitis from Vienna, Austria (n=19); Groningen, the Netherlands (n=50) and Cambridge, United Kingdom (n=53). Prevalence of LAMP-2 autoantibodies was similar in both those with myeloperoxidase-ANCA and proteinase 3-ANCA. Furthermore, we detected LAMP-2 autoantibodies in two ANCA-negative patients. LAMP-2 autoantibodies rapidly became undetectable after the initiation of immunosuppressive treatment and frequently became detectable again during clinical relapse. We conclude that when robust assays are used, circulating autoantibodies to hLAMP-2 can be detected in most European patients with ANCA-associated vasculitis. Large-scale prospective studies are now needed to determine whether they are pathogenic or merely an epiphenomenon.
Crohn disease (CD) and ulcerative colitis (UC) are increasingly common, chronic forms of inflamma... more Crohn disease (CD) and ulcerative colitis (UC) are increasingly common, chronic forms of inflammatory bowel disease. The behavior of these diseases varies unpredictably among patients. Identification of reliable prognostic biomarkers would enable treatment to be personalized so that patients destined to experience aggressive disease could receive appropriately potent therapies from diagnosis, while those who will experience more indolent disease are not exposed to the risks and side effects of unnecessary immunosuppression. Using transcriptional profiling of circulating T cells isolated from patients with CD and UC, we identified analogous CD8 + T cell transcriptional signatures that divided patients into 2 otherwise indistinguishable subgroups. In both UC and CD, patients in these subgroups subsequently experienced very different disease courses. A substantially higher incidence of frequently relapsing disease was experienced by those patients in the subgroup defined by elevated expression of genes involved in antigen-dependent T cell responses, including signaling initiated by both IL-7 and TCR ligation-pathways previously associated with prognosis in unrelated autoimmune diseases. No equivalent correlation was observed with CD4 + T cell gene expression. This suggests that the course of otherwise distinct autoimmune and inflammatory conditions may be influenced by common pathways and identifies what we believe to be the first biomarker that can predict prognosis in both UC and CD from diagnosis, a major step toward personalized therapy. Conflict of interest: The authors have declared that no conflict of interest exists.
Obese Zucker rats have elevated basal corticosterone levels and an increased stress response sugg... more Obese Zucker rats have elevated basal corticosterone levels and an increased stress response suggestive of an increased activity of the hypothalamic-pituitary-adrenal (HPA) axis. We hypothesized that altered central expression of glucocorticoid receptors (GR), mineralocorticoid receptors (MR), and/or 11β-hydroxysteroid dehydrogenase type 1 (11βHSD1) contribute to these changes. In brains from young adult male rats, in situ hybridization and Western blotting showed that obese rats had normal hippocampal GR mRNA and protein levels. In contrast, in obese rats, 11βHSD1 mRNA levels were reduced in a subpopulation of hippocampal cells in the main neuronal layers (by 37–47%, P < 0.05), whereas 11βHSD1 levels in sparse high-expressing cells did not differ. MR mRNA was decreased in all regions of the hippocampus (by 37–49%, P < 0.05 for CA1–2 and P < 0.01 for dentate gyrus) and in frontal cortex (by 16%, P < 0.05) in obese rats. In whole hippocampal homogenates, however, neither ...
Crohn disease (CD) and ulcerative colitis (UC) are increasingly common, chronic forms of inflamma... more Crohn disease (CD) and ulcerative colitis (UC) are increasingly common, chronic forms of inflammatory bowel disease. The behavior of these diseases varies unpredictably among patients. Identification of reliable prognostic biomarkers would enable treatment to ...
Summary In a recent workshop organized by the JDRF focused on the ‘Identification and Utilization... more Summary In a recent workshop organized by the JDRF focused on the ‘Identification and Utilization of Robust Biomarkers in Type1 Diabetes’, leaders in the field of type 1 diabetes (T1D)/autoimmunity and assay technology came together from academia, government and industry to assess the current state of the field, evaluate available resources/technologies and identify gaps that need to be filled for moving the field of T1D research forward. The highlights of this workshop are discussed in this paper, as well as the proposal for a larger, planned consortium effort, incorporating a JDRF Biomarker Core, to foster collaboration and accelerate progress in this critically needed area of T1D research.
Obese Zucker rats have elevated basal corticosterone levels and an increased stress response sugg... more Obese Zucker rats have elevated basal corticosterone levels and an increased stress response suggestive of an increased activity of the hypothalamic-pituitary-adrenal (HPA) axis. We hypothesized that altered central expression of glucocorticoid receptors (GR), mineralocorticoid receptors (MR), and/or 11-hydroxysteroid dehydrogenase type 1 (11HSD1) contribute to these changes. In brains from young adult male rats, in situ hybridization and Western blotting showed that obese rats had normal hippocampal GR mRNA and protein levels. In contrast, in obese rats, 11HSD1 mRNA levels were reduced in a subpopulation of hippocampal cells in the main neuronal layers (by 37-47%, P < 0.05), whereas 11HSD1 levels in sparse high-expressing cells did not differ. MR mRNA was decreased in all regions of the hippocampus (by 37-49%, P < 0.05 for CA1-2 and P < 0.01 for dentate gyrus) and in frontal cortex (by 16%, P < 0.05) in obese rats. In whole hippocampal homogenates, however, neither the protein concentration of MR by Western blot nor activity of 11HSD1 was measurably different between the phenotypes. To test the functional importance of lower central MR expression, groups of lean and obese rats were given spironolactone before restraint stress. In vehicle-treated animals, obese rats had higher plasma corticosterone levels than lean rats after stress (by ANOVA, P < 0.05). Spironolactone markedly increased the corticosterone response in both groups, but the incremental rise was smaller in the obese rats, so that spironolactone abolished the differences between groups. We conclude that lower levels of MR, but not GR, contribute to the increased HPA activity in the obese Zucker rats and that this seems more influential during stress than in the basal state. This may be exacerbated by impaired local regeneration of corticosterone by 11HSD1. These abnormalities could contribute to the subtle changes in the HPA axis in rodent and human obesity.
ObjectiveTo optimise a strategy for identifying gene expression signatures differentiating system... more ObjectiveTo optimise a strategy for identifying gene expression signatures differentiating systemic lupus erythematosus (SLE) and antineutrophil cytoplasmic antibody-associated vasculitis that provide insight into disease pathogenesis and identify biomarkers.Methods44 vasculitis patients, 13 SLE patients and 25 age and sex-matched controls were enrolled. CD4 and CD8 T cells, B cells, monocytes and neutrophils were isolated from each patient and, together with unseparated peripheral blood mononuclear cells (PBMC), were hybridised to spotted oligonucleotide microarrays.ResultsUsing expression data obtained from purified cells a substantial number of differentially expressed genes were identified that were not detectable in the analysis of PBMC. Analysis of purified T cells identified a SLE-associated, CD4 T-cell signature consistent with type 1 interferon signalling driving the generation and survival of tissue homing T cells and thereby contributing to disease pathogenesis. Moreover,...
B cells are important in the pathogenesis of many, and perhaps all, immune-mediated diseases (IMD... more B cells are important in the pathogenesis of many, and perhaps all, immune-mediated diseases (IMDs). Each B cell expresses a single B cell receptor (BCR) 1 , with the diverse range of BCRs expressed by an individual's total B cell population being termed the "BCR repertoire". Our understanding of the BCR repertoire in the context of IMDs is incomplete, and defining this could reveal new insights into pathogenesis and therapy. We therefore compared the BCR repertoire in systemic lupus erythematosus (SLE), ANCA-associated vasculitis (AAV), Crohn's disease (CD), Behçet's disease (BD), eosinophilic granulomatosis with polyangiitis (EGPA) and IgA vasculitis (IgAV), analysing BCR clonality, and immunoglobulin heavy chain gene (IGHV) and, in particular, isotype usage. An IgA-dominated increased clonality in SLE and CD, together with skewed IGHV gene usage in these and other diseases, suggested a microbial contribution to pathogenesis. Different immunosuppressive treatment had specific and distinct impacts on the repertoire; B cells persisting after rituximab were predominately isotype-switched and clonally expanded, the inverse of those persisting after mycophenolate mofetil. A comparative analysis of Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:
Choosing the best treatment-plan for each individual patient requires accurate forecasts of their... more Choosing the best treatment-plan for each individual patient requires accurate forecasts of their outcome trajectories as a function of the treatment, over time. While large observational data sets constitute rich sources of information to learn from, they also contain biases as treatments are rarely assigned randomly in practice. To provide accurate and unbiased forecasts, we introduce the Disentangled Counterfactual Recurrent Network (DCRN), a novel sequence-tosequence architecture that estimates treatment outcomes over time by learning representations of patient histories that are disentangled into three separate latent factors: a treatment factor, influencing only treatment selection; an outcome factor, influencing only the outcome; and a confounding factor, influencing both. With an architecture that is completely inspired by the causal structure of treatment influence over time, we advance forecast accuracy and disease understanding, as our architecture allows for practitioners to infer which patient features influence which part in a patient's trajectory, contrasting other approaches in this domain. We demonstrate that DCRN outperforms current state-of-the-art methods in forecasting treatment responses, on both real and simulated data.
The endothelial cell has a remarkable ability for sub-specialisation, adapted to the needs of a v... more The endothelial cell has a remarkable ability for sub-specialisation, adapted to the needs of a variety of vascular beds. The role of developmental programming versus the tissue contextual environment for this specialization is not well understood. Here we describe a hierarchy of expression of HOX genes associated with endothelial cell origin and location. In initial microarray studies, differential gene expression was examined in two endothelial cell lines: blood derived outgrowth endothelial cells (BOECs) and pulmonary artery endothelial cells. This suggested shared and differential patterns of HOX gene expression between the two endothelial lines. For example, this included a cluster on chromosome 2 of HOXD1, HOXD3, HOXD4, HOXD8 and HOXD9 that was expressed at a higher level in BOECs. Quantative PCR confirmed the higher expression of these HOXs in BOECs, a pattern that was shared by a variety of microvascular endothelial cell lines. Subsequently, we analysed publically available microarrays from a variety of adult cell and tissue types using the whole ''HOX transcriptome'' of all 39 HOX genes. Using hierarchical clustering analysis the HOX transcriptome was able to discriminate endothelial cells from 61 diverse human cell lines of various origins. In a separate publically available microarray dataset of 53 human endothelial cell lines, the HOX transcriptome additionally organized endothelial cells related to their organ or tissue of origin. Human tissue staining for HOXD8 and HOXD9 confirmed endothelial expression and also supported increased microvascular expression of these HOXs. Together these observations suggest a significant involvement of HOX genes in endothelial cell positional identity.
Background: Although numerous investigations have compared gene expression microarray platforms, ... more Background: Although numerous investigations have compared gene expression microarray platforms, preprocessing methods and batch correction algorithms using constructed spike-in or dilution datasets, there remains a paucity of studies examining the properties of microarray data using diverse biological samples. Most microarray experiments seek to identify subtle differences between samples with variable background noise, a scenario poorly represented by constructed datasets. Thus, microarray users lack important information regarding the complexities introduced in real-world experimental settings. The recent development of a multiplexed, digital technology for nucleic acid measurement enables counting of individual RNA molecules without amplification and, for the first time, permits such a study. Results: Using a set of human leukocyte subset RNA samples, we compared previously acquired microarray expression values with RNA molecule counts determined by the nCounter Analysis System (NanoString Technologies) in selected genes. We found that gene measurements across samples correlated well between the two platforms, particularly for high-variance genes, while genes deemed unexpressed by the nCounter generally had both low expression and low variance on the microarray. Confirming previous findings from spike-in and dilution datasets, this "gold-standard" comparison demonstrated signal compression that varied dramatically by expression level and, to a lesser extent, by dataset. Most importantly, examination of three different cell types revealed that noise levels differed across tissues. Conclusions: Microarray measurements generally correlate with relative RNA molecule counts within optimal ranges but suffer from expression-dependent accuracy bias and precision that varies across datasets. We urge microarray users to consider expression-level effects in signal interpretation and to evaluate noise properties in each dataset independently.
Wolters Kluwer Health may email you for journal alerts and information, but is committed to maint... more Wolters Kluwer Health may email you for journal alerts and information, but is committed to maintaining your privacy and will not share your personal information without your express consent. For more information, please refer to our Privacy Policy. ... Skip Navigation Links Home &amp;gt; February ...
Previous research has generated examples of how genetic and environmental factors can interact to... more Previous research has generated examples of how genetic and environmental factors can interact to create risk for psychopathology. Using a gene-by-environment (G X E) interaction design, we tested whether three polymorphisms in the dopamine transporter gene (DAT1, also referred to as SLC6A3, located at 5p15.33) interacted with maternal parenting style to predict first-onset episodes of depression. Participants were male adolescents (N = 176) recruited from a juvenile detention center in northern Russia. As hypothesized, one of the polymorphisms (rs40184) moderated the effect of perceived maternal rejection on the onset of major depressive disorder, as well as on suicidal ideation. Further, this G X E interaction was specific to depression; it did not predict clinically significant anxiety. These results highlight the need for further research investigating the moderating effects of dopaminergic genes on depression.
Systemic Lupus Erythematosus (SLE) is a chronic autoimmune disorder characterized by broad clinic... more Systemic Lupus Erythematosus (SLE) is a chronic autoimmune disorder characterized by broad clinical manifestations including cardiovascular and renal complications with periodic disease flares and significant morbidity and mortality. One of the main contributing factors to the pathology of SLE is the accumulation and impaired clearance of immune complexes of which the principle components are host auto-antigens and antibodies. The contribution of host lipids to the formation of these autoimmune complexes remains poorly defined. The aim of the present study was to identify and analyze candidate lipid autoantigens and their corresponding anti-lipid antibody responses in a well-defined SLE patient cohort using a combination of immunological and biophysical techniques. Disease monitoring in the SLE cohort was undertaken with serial British Isles Lupus Assessment Group (BILAG) scoring. Correlations between specific lipid/anti-lipid responses were investigated as disease activity developed from active flares to quiescent during a follow up period. We report a significant negative correlation between anti-lipid antibodies for 24S-hydroxycholesterol, cardiolipin and phosphatidylserine with SLE disease activity. Taken together, these data suggest that lipid autoantigens represent a new family of biomarkers that can be employed to monitor disease activity plus the efficacy of therapeutic intervention in SLE.
Autoimmune diseases are common and debilitating, but their severe manifestations could be reduced... more Autoimmune diseases are common and debilitating, but their severe manifestations could be reduced if biomarkers were available to allow individual tailoring of the potentially toxic immunosuppressive therapy required for their control. Gene expression-based biomarkers facilitating individual tailoring of chemotherapy in cancer, but not autoimmunity, have been identified and translated into clinical practice1,2. We show that transcriptional profiling of purified CD8 T cells, which avoids the confounding influences of unseparated cells3,4, identifies two distinct patient subgroups predicting long-term prognosis in two different autoimmune diseases, anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV), a chronic, severe disease characterized by inflammation of medium and small blood vessels5, and systemic lupus erythematosus (SLE), characterized by autoantibodies, immune complex deposition and diverse clinical manifestations ranging from glomerulonephritis to neurological dysfunction6. We show that genes defining the poor prognostic group are enriched for genes of the IL7R pathway, TCR signalling and those expressed by memory T cells. Furthermore, the poor prognostic group is associated with an expanded CD8 T cell memory population. These subgroups, which are also found in the normal population and can be identified by measuring expression of only three genes, raise the prospect of individualized therapy and suggest novel potential therapeutic targets in autoimmunity.
ABSTRACT Autoimmune diseases are common and debilitating, but their severe manifestations could b... more ABSTRACT Autoimmune diseases are common and debilitating, but their severe manifestations could be reduced if biomarkers were available to allow individual tailoring of the potentially toxic immunosuppressive therapy required for their control. Gene expression-based biomarkers have been identified and translated into clinical practice in cancer, but not autoimmunity. We show that transcriptional profiling of purified CD8 T cells, which avoids the confounding influences of unseparated cells, identifies two distinct patient subgroups predicting long-term prognosis in four distinct autoimmune diseases: anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis, systemic lupus erythematosus, ulcerative colitis, and Crohn&#39;s disease.While the correlation of this signature with prognosis raises the prospect of individualised therapy in autoimmunity, the biological mechanism underlying its association with relapsing disease may indicate novel therapeutic targets. Acute exposure to foreign antigen results in persistent antigen-specific cellular immunity, but during chronic viral infection persistent antigen exposure may exhaust that response. This exhausted CD8 T-cell phenotype may itself be defined by a distinct transcriptional signature. We show that the same signature seen in chronic viral infection can be seen in chronic self antigen exposure during autoimmunity, with one exception: whereas exhaustion results in viral persistence it predicts favourable outcome in autoimmunity. Provision of adequate CD4 help can prevent or overcome the development of CD8 exhaustion resulting in clearance of chronic viral infection. Consistent with this observation, we show that the CD8 exhaustion signature in autoimmunity is inversely correlated with that of concurrent CD4 help and can be robustly identified in mixed cell populations. This has allowed independent validation of this marker on published datasets, confirming prediction of clinical outcome in a total of 1070 samples from 429 individuals spanning infection (HIV, dengue virus, hepatitis C), vaccination (yellow fever, malaria, influenza), and autoimmunity (type 1 diabetes, ANCA–associated vasculitis, systemic lupus erythematosus, Crohn&#39;s disease, and ulcerative colitis).FundingWellcome Trust.
Endothelial cells have a remarkable ability for subspecialisation, adapting to the needs of a var... more Endothelial cells have a remarkable ability for subspecialisation, adapting to the needs of a variety of vascular beds. It is not known how much of this subspecialisation is related to developmental programming versus the environment in which the endothelial cell fi nds itself. In transcriptomic studies of endothelial cells we have noted a hierarchy of HOX gene expression which predicts endothelial cell specifi cation and fate. HOX genes are well described as masterregulators of positional identity, predominately in the developing embryo.
ABSTRACT Background The clinical course of autoimmune and infectious disease varies greatly even ... more ABSTRACT Background The clinical course of autoimmune and infectious disease varies greatly even between individuals with the same condition. An understanding of the molecular basis for this heterogeneity could lead to substantial improvements in both monitoring and treatment. Since persistent antigen is targeted in both autoimmunity and chronic infection, similar pathways could dictate the success of that response. However, whereas a successful response to pathogen results in clearance and recovery from disease, a robust response to self-antigen might drive relapsing autoimmunity. During chronic viral infection, CD8 T cells develop progressive loss of function in a process termed exhaustion, characterised by high expression of inhibitory receptors (such as programmed cell death 1 [PD1]) and low expression of memory markers (such as interleukin-7 receptor). These can serve both as biomarkers of viral progression and as targets for therapy. We sought to determine whether an analogous exhausted state exists during autoimmune responses targeting persistent self-antigen. Methods To identify gene expression signatures associated with clinical outcome, we used weighted gene coexpression network analysis of concurrently sampled CD4 and CD8 T-cell transcriptomes from patients with four distinct autoimmune diseases (23 with systemic lupus erythematosus [SLE], 44 with anti-neutrophil cytoplasmic antibody-associated vasculitis [AAV], and 56 with Crohn&#39;s disease or ulcerative colitis) and with chronic viral infection (42 with HIV infection). Association with clinical outcome was independently validated in 1145 samples from 504 individuals with infectious disease (HIV, hepatitis C virus, dengue), during vaccination (malaria, yellow fever, influenza), and autoimmunity (type 1 diabetes, SLE, AAV, Crohn&#39;s disease and ulcerative colitis, and idiopathic pulmonary fibrosis). We used an artificial antigen presenting cell to modify provision of co-stimulatory and inhibitory signals (as suggested by the network analysis) to primary human T cells during in-vitro differentiation. In this way we tested whether expression signatures associated with clinical outcome might be modified by manipulating co-stimulatory signals and whether they reflected changes in T-cell function. Findings Network analysis of CD8 T-cell transcriptomes identified a common signature of T-cell exhaustion during responses to chronic viral and self-antigen. However, whereas exhausted responses were associated with poor viral clearance, they predicted a less severe course in multiple autoimmune diseases. In autoimmunity, we found that where CD8 T-cell exhaustion was high a concurrent signature of CD4 T-cell co-stimulation was low. We used this signature to identify CD2 as a co-stimulatory signal preventing the development of T-cell exhaustion during controlled in-vitro assays, recreating expression patterns seen in patients with severe autoimmunity and leading to enhanced T-cell survival during primary and repeated stimulation. Finally, exogenous signals through the inhibitory receptor PD1 restricted development of this robust population. Interpretation Here we show that the process of CD8 T-cell exhaustion is important not only in facilitating viral persistence during chronic infection, but also in shaping the immune response to persistent self-antigen during autoimmunity. The observed correlation between T-cell exhaustion, co-stimulation, and subsequent prognosis increases our understanding of the pathogenesis of severe autoimmune disease and might help stratify patients for personalised therapy. Furthermore, our data suggest that future modulation of T-cell exhaustion may lead to new therapeutic opportunities in autoimmunity. Funding Wellcome Trust, Biomedical Research Centre, National Institute for Health Research.
Journal of the American Society of Nephrology, 2012
The involvement of autoantibodies to human lysosome-associated membrane protein-2 (hLAMP-2) in an... more The involvement of autoantibodies to human lysosome-associated membrane protein-2 (hLAMP-2) in anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis is controversial because of the absence of confirmatory data subsequent to the initial reports of their high prevalence in this disease. We characterized three assays for anti-hLAMP-2 antibodies: ELISA and Western blotting assays using unglycosylated recombinant hLAMP-2 expressed in Escherichia coli, and an indirect immunofluorescence assay using stably transfected ldlD cells that expressed glycosylated full-length hLAMP-2 on the plasma membrane. The assays detected autoantibodies to hLAMP-2 in human sera reproducibly and with comparable sensitivity and the assays gave the same results in 80.5% of the test panel of 40 selected positive and negative sera. In untreated patients at presentation, the frequencies of autoantibodies to LAMP-2 were 89%, 91%, and 80%, respectively, among three groups of patients with ANCA-associated vasculitis from Vienna, Austria (n=19); Groningen, the Netherlands (n=50) and Cambridge, United Kingdom (n=53). Prevalence of LAMP-2 autoantibodies was similar in both those with myeloperoxidase-ANCA and proteinase 3-ANCA. Furthermore, we detected LAMP-2 autoantibodies in two ANCA-negative patients. LAMP-2 autoantibodies rapidly became undetectable after the initiation of immunosuppressive treatment and frequently became detectable again during clinical relapse. We conclude that when robust assays are used, circulating autoantibodies to hLAMP-2 can be detected in most European patients with ANCA-associated vasculitis. Large-scale prospective studies are now needed to determine whether they are pathogenic or merely an epiphenomenon.
Crohn disease (CD) and ulcerative colitis (UC) are increasingly common, chronic forms of inflamma... more Crohn disease (CD) and ulcerative colitis (UC) are increasingly common, chronic forms of inflammatory bowel disease. The behavior of these diseases varies unpredictably among patients. Identification of reliable prognostic biomarkers would enable treatment to be personalized so that patients destined to experience aggressive disease could receive appropriately potent therapies from diagnosis, while those who will experience more indolent disease are not exposed to the risks and side effects of unnecessary immunosuppression. Using transcriptional profiling of circulating T cells isolated from patients with CD and UC, we identified analogous CD8 + T cell transcriptional signatures that divided patients into 2 otherwise indistinguishable subgroups. In both UC and CD, patients in these subgroups subsequently experienced very different disease courses. A substantially higher incidence of frequently relapsing disease was experienced by those patients in the subgroup defined by elevated expression of genes involved in antigen-dependent T cell responses, including signaling initiated by both IL-7 and TCR ligation-pathways previously associated with prognosis in unrelated autoimmune diseases. No equivalent correlation was observed with CD4 + T cell gene expression. This suggests that the course of otherwise distinct autoimmune and inflammatory conditions may be influenced by common pathways and identifies what we believe to be the first biomarker that can predict prognosis in both UC and CD from diagnosis, a major step toward personalized therapy. Conflict of interest: The authors have declared that no conflict of interest exists.
Obese Zucker rats have elevated basal corticosterone levels and an increased stress response sugg... more Obese Zucker rats have elevated basal corticosterone levels and an increased stress response suggestive of an increased activity of the hypothalamic-pituitary-adrenal (HPA) axis. We hypothesized that altered central expression of glucocorticoid receptors (GR), mineralocorticoid receptors (MR), and/or 11β-hydroxysteroid dehydrogenase type 1 (11βHSD1) contribute to these changes. In brains from young adult male rats, in situ hybridization and Western blotting showed that obese rats had normal hippocampal GR mRNA and protein levels. In contrast, in obese rats, 11βHSD1 mRNA levels were reduced in a subpopulation of hippocampal cells in the main neuronal layers (by 37–47%, P < 0.05), whereas 11βHSD1 levels in sparse high-expressing cells did not differ. MR mRNA was decreased in all regions of the hippocampus (by 37–49%, P < 0.05 for CA1–2 and P < 0.01 for dentate gyrus) and in frontal cortex (by 16%, P < 0.05) in obese rats. In whole hippocampal homogenates, however, neither ...
Crohn disease (CD) and ulcerative colitis (UC) are increasingly common, chronic forms of inflamma... more Crohn disease (CD) and ulcerative colitis (UC) are increasingly common, chronic forms of inflammatory bowel disease. The behavior of these diseases varies unpredictably among patients. Identification of reliable prognostic biomarkers would enable treatment to ...
Summary In a recent workshop organized by the JDRF focused on the ‘Identification and Utilization... more Summary In a recent workshop organized by the JDRF focused on the ‘Identification and Utilization of Robust Biomarkers in Type1 Diabetes’, leaders in the field of type 1 diabetes (T1D)/autoimmunity and assay technology came together from academia, government and industry to assess the current state of the field, evaluate available resources/technologies and identify gaps that need to be filled for moving the field of T1D research forward. The highlights of this workshop are discussed in this paper, as well as the proposal for a larger, planned consortium effort, incorporating a JDRF Biomarker Core, to foster collaboration and accelerate progress in this critically needed area of T1D research.
Obese Zucker rats have elevated basal corticosterone levels and an increased stress response sugg... more Obese Zucker rats have elevated basal corticosterone levels and an increased stress response suggestive of an increased activity of the hypothalamic-pituitary-adrenal (HPA) axis. We hypothesized that altered central expression of glucocorticoid receptors (GR), mineralocorticoid receptors (MR), and/or 11-hydroxysteroid dehydrogenase type 1 (11HSD1) contribute to these changes. In brains from young adult male rats, in situ hybridization and Western blotting showed that obese rats had normal hippocampal GR mRNA and protein levels. In contrast, in obese rats, 11HSD1 mRNA levels were reduced in a subpopulation of hippocampal cells in the main neuronal layers (by 37-47%, P < 0.05), whereas 11HSD1 levels in sparse high-expressing cells did not differ. MR mRNA was decreased in all regions of the hippocampus (by 37-49%, P < 0.05 for CA1-2 and P < 0.01 for dentate gyrus) and in frontal cortex (by 16%, P < 0.05) in obese rats. In whole hippocampal homogenates, however, neither the protein concentration of MR by Western blot nor activity of 11HSD1 was measurably different between the phenotypes. To test the functional importance of lower central MR expression, groups of lean and obese rats were given spironolactone before restraint stress. In vehicle-treated animals, obese rats had higher plasma corticosterone levels than lean rats after stress (by ANOVA, P < 0.05). Spironolactone markedly increased the corticosterone response in both groups, but the incremental rise was smaller in the obese rats, so that spironolactone abolished the differences between groups. We conclude that lower levels of MR, but not GR, contribute to the increased HPA activity in the obese Zucker rats and that this seems more influential during stress than in the basal state. This may be exacerbated by impaired local regeneration of corticosterone by 11HSD1. These abnormalities could contribute to the subtle changes in the HPA axis in rodent and human obesity.
ObjectiveTo optimise a strategy for identifying gene expression signatures differentiating system... more ObjectiveTo optimise a strategy for identifying gene expression signatures differentiating systemic lupus erythematosus (SLE) and antineutrophil cytoplasmic antibody-associated vasculitis that provide insight into disease pathogenesis and identify biomarkers.Methods44 vasculitis patients, 13 SLE patients and 25 age and sex-matched controls were enrolled. CD4 and CD8 T cells, B cells, monocytes and neutrophils were isolated from each patient and, together with unseparated peripheral blood mononuclear cells (PBMC), were hybridised to spotted oligonucleotide microarrays.ResultsUsing expression data obtained from purified cells a substantial number of differentially expressed genes were identified that were not detectable in the analysis of PBMC. Analysis of purified T cells identified a SLE-associated, CD4 T-cell signature consistent with type 1 interferon signalling driving the generation and survival of tissue homing T cells and thereby contributing to disease pathogenesis. Moreover,...
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Papers by Eoin McKinney