Stratification of asthma at the molecular level, especially using accessible biospecimens, could greatly enable patient selection for targeted therapy.
To determine the value of blood analysis to ...identify transcriptional differences between clinically defined asthma and nonasthma groups, identify potential patient subgroups based on gene expression, and explore biological pathways associated with identified differences.
Transcriptomic profiles were generated by microarray analysis of blood from 610 patients with asthma and control participants in the U-BIOPRED (Unbiased Biomarkers in Prediction of Respiratory Disease Outcomes) study. Differentially expressed genes (DEGs) were identified by analysis of variance, including covariates for RNA quality, sex, and clinical site, and Ingenuity Pathway Analysis was applied. Patient subgroups based on DEGs were created by hierarchical clustering and topological data analysis.
A total of 1,693 genes were differentially expressed between patients with severe asthma and participants without asthma. The differences from participants without asthma in the nonsmoking severe asthma and mild/moderate asthma subgroups were significantly related (r = 0.76), with a larger effect size in the severe asthma group. The majority of, but not all, differences were explained by differences in circulating immune cell populations. Pathway analysis showed an increase in chemotaxis, migration, and myeloid cell trafficking in patients with severe asthma, decreased B-lymphocyte development and hematopoietic progenitor cells, and lymphoid organ hypoplasia. Cluster analysis of DEGs led to the creation of subgroups among the patients with severe asthma who differed in molecular responses to oral corticosteroids.
Blood gene expression differences between clinically defined subgroups of patients with asthma and individuals without asthma, as well as subgroups of patients with severe asthma defined by transcript profiles, show the value of blood analysis in stratifying patients with asthma and identifying molecular pathways for further study. Clinical trial registered with www.clinicaltrials.gov (NCT01982162).
Abstract
RNA G-quadruplexes (G4s) are secondary structures proposed to function as regulators of post-transcriptional mRNA localisation and translation. G4s within some neuronal mRNAs are known to ...control distal localisation and local translation, contributing to distinct local proteomes that facilitate the synaptic remodelling attributed to normal cellular function. In this study, we characterise the G4 formation of a (GGN)13 repeat found within the 5′ UTR of the potassium 2-pore domain leak channel Task3 mRNA. Biophysical analyses show that this (GGN)13 repeat forms a parallel G4 in vitro exhibiting the stereotypical potassium specificity of G4s, remaining thermostable under physiological ionic conditions. Through mouse brain tissue G4-RNA immunoprecipitation, we further confirm that Task3 mRNA forms a G4 structure in vivo. The G4 is inhibitory to translation of Task3 in vitro and is overcome through activity of a G4-specific helicase DHX36, increasing K+ leak currents and membrane hyperpolarisation in HEK293 cells. Further, we observe that this G4 is fundamental to ensuring delivery of Task3 mRNA to distal primary cortical neurites. It has been shown that aberrant Task3 expression correlates with neuronal dysfunction, we therefore posit that this G4 is important in regulated local expression of Task3 leak channels that maintain K+ leak within neurons.
Severe asthma is associated with multiple co-morbidities, including gastro-oesophageal reflux disease (GORD) which can contribute to exacerbation frequency and poor quality of life. Since epithelial ...dysfunction is an important feature in asthma, we hypothesised that in severe asthma the bronchial epithelium is more susceptible to the effects of acid reflux.
We developed an
model of GORD using differentiated bronchial epithelial cells (BECs) from normal or severe asthmatic donors exposed to a combination of pepsin, acid pH, and bile acids using a multiple challenge protocol (MCP-PAB). We also analysed bronchial biopsies and undertook RNA-sequencing of bronchial brushings from controls and severe asthmatics without or with GORD.
Exposure of BECs to the MCP-PAB caused structural disruption, increased permeability, IL-33 expression, inflammatory mediator release and changes in gene expression for multiple biological processes. Cultures from severe asthmatics were significantly more affected than those from healthy donors. Analysis of bronchial biopsies confirmed increased IL-33 expression in severe asthmatics with GORD. RNA-sequencing of bronchial brushings from this group identified 15 of the top 37 dysregulated genes found in MCP-PAB treated BECs, including genes involved in oxidative stress responses.
By affecting epithelial permeability, GORD may increase exposure of the airway submucosa to allergens and pathogens, resulting in increased risk of inflammation and exacerbations.
These results suggest the need for research into alternative therapeutic management of GORD in severe asthma.
Background
The COVID-19 pandemic has created pressure on healthcare systems worldwide. Tools that can stratify individuals according to prognosis could allow for more efficient allocation of ...healthcare resources and thus improved patient outcomes. It is currently unclear if blood gene expression signatures derived from patients at the point of admission to hospital could provide useful prognostic information.
Methods
Gene expression of whole blood obtained at the point of admission from a cohort of 78 patients hospitalised with COVID-19 during the first wave was measured by high resolution RNA sequencing. Gene signatures predictive of admission to Intensive Care Unit were identified and tested using machine learning and topological data analysis, TopMD.
Results
The best gene expression signature predictive of ICU admission was defined using topological data analysis with an accuracy: 0.72 and ROC AUC: 0.76. The gene signature was primarily based on differentially activated pathways controlling epidermal growth factor receptor (EGFR) presentation, Peroxisome proliferator-activated receptor alpha (PPAR-α) signalling and Transforming growth factor beta (TGF-β) signalling.
Conclusions
Gene expression signatures from blood taken at the point of admission to hospital predicted ICU admission of treatment naïve patients with COVID-19.
IMPORTANCE: Psychosis is a known risk factor for offending behavior, but little is known about the association between clinical contact with mental health services after an offense and reoffending. ...OBJECTIVE: To examine the association between early contact with mental health services and reoffending after an index offense in individuals with psychosis. DESIGN, SETTING, AND PARTICIPANTS: In this cohort study, individuals diagnosed with psychosis before their index offense from July 1, 2001, to December 31, 2012, and who received a noncustodial sentence were identified by linking health and offending databases in New South Wales, Australia. The incidence of and risk factors for reoffending and time to reoffending within 2 years from the index offense were examined using Cox proportional hazards regression and Kaplan-Meier survival estimates. Specifically, the association between contact with mental health services within 30 days after an offense and reoffending was examined. Data were analyzed from July 1, 2019, to March 5, 2020. EXPOSURES: Hospital admission, emergency department presentation, and contact with community mental health services associated with psychosis. MAIN OUTCOMES AND MEASURES: Reoffending within 2 years of the index offense was compared in individuals with and without clinical contact with mental health services within 30 days after an offense, with adjustment for potential confounders. RESULTS: Of the 7030 offenders with psychosis (4933 male 70.2%; median age at the index offense, 34 interquartile range, 26-42 years), 2605 (37.1%) had clinical contact with mental health services within 30 days after the index offense. The risk of reoffending was significantly lower in those with vs without clinical contact (adjusted hazard ratio AHR, 0.83; 95% CI, 0.76-0.91). The risk of reoffending was 30% less in male offenders with 5 or more clinical contacts compared with male offenders with no clinical contact (AHR, 0.71; 95% CI, 0.59-0.84). Reoffending in both male and female offenders was associated with younger age (eg, AHR for male offenders aged <18 years, 3.31 95% CI, 2.39-4.59; AHR for female offenders aged <18 years, 2.60 95% CI, 1.69-3.99) and offending history (eg, AHR for male offenders with ≥4 prior offenses, 2.28 95% CI, 1.98-2.64; AHR for female offenders with ≥4 prior offenses, 2.22 95% CI, 1.67-2.96). CONCLUSIONS AND RELEVANCE: In this cohort, early and frequent clinical contact with mental health services after an offense in individuals with psychosis was associated with reduced risk of reoffending in this group. More support may be needed for early treatment of those with serious mental illness who are at risk of reoffending.
The worldwide COVID-19 pandemic has claimed millions of lives and has had a profound effect on global life. Understanding the body's immune response to SARS-CoV-2 infection is crucial in improving ...patient management and prognosis. In this study we compared influenza and SARS-CoV-2 infected patient cohorts to identify distinct blood transcript abundances and cellular composition to better understand the natural immune response associated with COVID-19, compared to another viral infection being influenza, and identify a prognostic signature of COVID-19 patient outcome. Clinical characteristics and peripheral blood were acquired upon hospital admission from two well characterised cohorts, a cohort of 88 patients infected with influenza and a cohort of 80 patients infected with SARS-CoV-2 during the first wave of the pandemic and prior to availability of COVID-19 treatments and vaccines. Gene transcript abundances, enriched pathways and cellular composition were compared between cohorts using RNA-seq. A genetic signature between COVID-19 survivors and non-survivors was assessed as a prognostic predictor of COVID-19 outcome. Contrasting immune responses were detected with an innate response elevated in influenza and an adaptive response elevated in COVID-19. Additionally ribosomal, mitochondrial oxidative stress and interferon signalling pathways differentiated the cohorts. An adaptive immune response was associated with COVID-19 survival, while an inflammatory response predicted death. A prognostic transcript signature, associated with circulating immunoglobulins, nucleosome assembly, cytokine production and T cell activation, was able to stratify COVID-19 patients likely to survive or die. This study provides a unique insight into the immune responses of treatment naïve patients with influenza or COVID-19. The comparison of immune response between COVID-19 survivors and non-survivors enables prognostication of COVID-19 patients and may suggest potential therapeutic strategies to improve survival.
There has recently been a huge increase in interest in the formation of stable G-quadruplex structures in mRNAs and their functional significance. In neurons, local translation of mRNA is essential ...for normal neuronal behaviour. It has been discovered that local translation of specific mRNAs encoding some of the best known synaptic proteins is dependent on the presence of a G-quadruplex. The recognition of G-quadruplexes in mRNAs, their transport as repressed complexes and the control of their translation at their subcellular destinations involves a diversity of proteins, including those associated with disease pathologies. This is an exciting field, with rapid improvements to our knowledge and understanding. Here, we discuss some of the recent work on how G-quadruplexes mediate local translation in neurons.
Asthma is a chronic respiratory disease with significant heterogeneity in its clinical presentation and pathobiology. There is need for improved understanding of respiratory lipid metabolism in ...asthma patients and its relation to observable clinical features.
We performed a comprehensive, prospective, cross-sectional analysis of the lipid composition of induced sputum supernatant obtained from asthma patients with a range of disease severities, as well as from healthy controls.
Induced sputum supernatant was collected from 211 adults with asthma and 41 healthy individuals enrolled onto the U-BIOPRED (Unbiased Biomarkers for the Prediction of Respiratory Disease Outcomes) study. Sputum lipidomes were characterized by semiquantitative shotgun mass spectrometry and clustered using topologic data analysis to identify lipid phenotypes.
Shotgun lipidomics of induced sputum supernatant revealed a spectrum of 9 molecular phenotypes, highlighting not just significant differences between the sputum lipidomes of asthma patients and healthy controls, but also within the asthma patient population. Matching clinical, pathobiologic, proteomic, and transcriptomic data helped inform the underlying disease processes. Sputum lipid phenotypes with higher levels of nonendogenous, cell-derived lipids were associated with significantly worse asthma severity, worse lung function, and elevated granulocyte counts.
We propose a novel mechanism of increased lipid loading in the epithelial lining fluid of asthma patients resulting from the secretion of extracellular vesicles by granulocytic inflammatory cells, which could reduce the ability of pulmonary surfactant to lower surface tension in asthmatic small airways, as well as compromise its role as an immune regulator.
The role of IL-17 immunity is well established in patients with inflammatory diseases, such as psoriasis and inflammatory bowel disease, but not in asthmatic patients, in whom further study is ...required.
We sought to undertake a deep phenotyping study of asthmatic patients with upregulated IL-17 immunity.
Whole-genome transcriptomic analysis was performed by using epithelial brushings, bronchial biopsy specimens (91 asthmatic patients and 46 healthy control subjects), and whole blood samples (n = 498) from the Unbiased Biomarkers for the Prediction of Respiratory Disease Outcomes (U-BIOPRED) cohort. Gene signatures induced in vitro by IL-17 and IL-13 in bronchial epithelial cells were used to identify patients with IL-17–high and IL-13–high asthma phenotypes.
Twenty-two of 91 patients were identified with IL-17, and 9 patients were identified with IL-13 gene signatures. The patients with IL-17–high asthma were characterized by risk of frequent exacerbations, airway (sputum and mucosal) neutrophilia, decreased lung microbiota diversity, and urinary biomarker evidence of activation of the thromboxane B2 pathway. In pathway analysis the differentially expressed genes in patients with IL-17-high asthma were shared with those reported as altered in psoriasis lesions and included genes regulating epithelial barrier function and defense mechanisms, such as IL1B, IL6, IL8, and β-defensin.
The IL-17–high asthma phenotype, characterized by bronchial epithelial dysfunction and upregulated antimicrobial and inflammatory response, resembles the immunophenotype of psoriasis, including activation of the thromboxane B2 pathway, which should be considered a biomarker for this phenotype in further studies, including clinical trials targeting IL-17.
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Analysis of induced sputum supernatant is a minimally invasive approach to study the epithelial lining fluid and, thereby, provide insight into normal lung biology and the pathobiology of lung ...diseases. We present here a novel proteomics approach to sputum analysis developed within the U-BIOPRED (unbiased biomarkers predictive of respiratory disease outcomes) international project. We present practical and analytical techniques to optimize the detection of robust biomarkers in proteomic studies. The normal sputum proteome was derived using data-independent HDMSE applied to 40 healthy nonsmoking participants, which provides an essential baseline from which to compare modulation of protein expression in respiratory diseases. The “core” sputum proteome (proteins detected in ≥40% of participants) was composed of 284 proteins, and the extended proteome (proteins detected in ≥3 participants) contained 1666 proteins. Quality control procedures were developed to optimize the accuracy and consistency of measurement of sputum proteins and analyze the distribution of sputum proteins in the healthy population. The analysis showed that quantitation of proteins by HDMSE is influenced by several factors, with some proteins being measured in all participants’ samples and with low measurement variance between samples from the same patient. The measurement of some proteins is highly variable between repeat analyses, susceptible to sample processing effects, or difficult to accurately quantify by mass spectrometry. Other proteins show high interindividual variance. We also highlight that the sputum proteome of healthy individuals is related to sputum neutrophil levels, but not gender or allergic sensitization. We illustrate the importance of design and interpretation of disease biomarker studies considering such protein population and technical measurement variance.
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