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On 23 March 2020, the UK Government announced a nationwide lockdown in response to the COVID-19 epidemic, with people banned from leaving their homes except for essential journeys and exercise., Impact of the UK lockdown on early COPD https://bit.ly/3laMsmi

Theranostics, the combination of diagnosis and therapy, has long held promise as a means to achieving personalised precision cancer treatments. However, despite its potential, theranostics has yet to realise significant clinical translation, largely due the complexity and overriding toxicity concerns of existing theranostic nanoparticle strategies. Methods: Here, we present an alternative nanoparticle-free theranostic approach based on simultaneous Raman spectroscopy and photodynamic therapy (PDT) in an integrated clinical platform for cancer theranostics. Results: We detail the compatibility of Raman spectroscopy and PDT for cancer theranostics, whereby Raman spectroscopic diagnosis can be performed on PDT photosensitiser-positive cells and tissues without inadvertent photosensitiser activation/photobleaching or impaired diagnostic capacity. We further demonstrate that our theranostic platform enables in vivo tumour diagnosis, treatment, and post-treatment molecular monitoring in real-time. Conclusion: This system thus achieves effective theranostic performance, providing a promising new avenue towards the clinical realisation of theranostics.

Hannah R Whittaker,1 Jeanne M Pimenta,2 Deborah Jarvis,1 Steven J Kiddle,3 Jennifer K Quint1 1Respiratory Epidemiology, Occupational Medicine and Public Health, National Heart and Lung Institute, Imperial College London, London, UK; 2Epidemiology (Value Evidence and Outcomes), GlaxoSmithKline, R&D, Uxbridge, UK; 3MRC Biostatistics Unit, University of Cambridge, Cambridge, UKCorrespondence: Hannah R WhittakerImperial College, Emmanuel Kaye Building, National Heart and Lung Institute, London, UKEmail h.whittaker@imperial.ac.ukBackground: Estimates for lung function decline in chronic obstructive pulmonary disease (COPD) have differed by study setting and have not been described in a UK primary care population.Purpose: To describe rates of FEV1 and FVC decline in COPD and investigate characteristics associated with accelerated decline.Patients and Methods: Current/ex-smoking COPD patients (35 years+) who had at least 2 FEV1 or FVC measurements ≥ 6 months apart were included using Clinical Practice Research Datalink. Patients were followed up for a maximum of 13 years. Accelerated rate of lung function decline was defined as the fastest quartile of decline using mixed linear regression, and association with baseline characteristics was investigated using logistic regression.Results: A total of 72,683 and 50,649 COPD patients had at least 2 FEV1 or FVC measurements, respectively. Median rates of FEV1 and FVC changes or decline were − 18.1mL/year (IQR: − 31.6 to − 6.0) and − 22.7mL/year (IQR: − 39.9 to − 6.7), respectively. Older age, high socioeconomic status, being underweight, high mMRC dyspnoea and frequent AECOPD or severe AECOPD were associated with an accelerated rate of FEV1 and FVC decline. Current smoking, mild airflow obstruction and inhaled corticosteroid treatment were additionally associated with accelerated FEV1 decline whilst women, sputum production and severe airflow obstruction were associated with accelerated FVC decline.Conclusion: Rate of FEV1 and FVC decline was similar and showed similar heterogeneity. Whilst FEV1 and FVC shared associations with baseline characteristics, a few differences highlighted the importance of both lung function measures in COPD progression. We identified important characteristics that should be monitored for disease progression.Keywords: pulmonary disease, chronic obstructive, spirometry, lung function

BACKGROUND: Human respiratory syncytial virus (RSV) is classified into antigenic subgroups A and B. Thirteen genotypes have been defined for RSV-A and 20 for RSV-B, without any consensus on genotype definition. METHODS: We evaluated clustering of RSV sequences published in GenBank until February 2018 to define genotypes by using maximum likelihood and Bayesian phylogenetic analyses and average p-distances. RESULTS: We compared the patterns of sequence clustering of complete genomes; the three surface glycoproteins genes (SH, G, and F, single and concatenated); the ectodomain and the 2nd hypervariable region of G gene. Although complete genome analysis achieved the best resolution, the F, G, and G-ectodomain phylogenies showed similar topologies with statistical support comparable to complete genome. Based on the widespread geographic representation and large number of available G-ectodomain sequences, this region was chosen as the minimum region suitable for RSV genotyping. A genotype was defined as a monophyletic cluster of sequences with high statistical support (≥80% bootstrap and ≥0.8 posterior probability), with an intragenotype p-distance ≤0.03 for both subgroups and an intergenotype p-distance ≥0.09 for RSV-A and ≥0.05 for RSV-B. In this work, the number of genotypes was reduced from 13 to three for RSV-A (GA1-GA3) and from 20 to seven for RSV-B (GB1-GB7). Within these, two additional levels of classification were defined: subgenotypes and lineages. Signature amino acid substitutions to complement this classification were also identified. CONCLUSIONS: We propose an objective protocol for RSV genotyping suitable for adoption as an international standard to support the global expansion of RSV molecular surveillance. ispartof: INFLUENZA AND OTHER RESPIRATORY VIRUSES vol:14 issue:3 pages:274-285 ispartof: location:England status: published

Extracellular vesicles (EVs) represent a promising cell-free alternative for treatment of cardiovascular diseases. Nevertheless, the lack of standardised and reproducible isolation methods capable of recovering pure, intact EVs presents a significant obstacle. Additionally, there is significant interest in investigating the interactions of EVs with different cardiac cell types. Here we established a robust technique for the production and isolation of EVs harvested from an enriched (>97% purity) population of human induced pluripotent stem cell (iPSC)-derived cardiomyocytes (CMs) with size exclusion chromatography. Utilizing an advanced fluorescence labelling strategy, we then investigated the interplay of the CM-EVs with the three major cellular components of the myocardium (fibroblasts, cardiomyocytes and endothelial cells) and identified that cardiac endothelial cells show preferential uptake of these EVs. Overall, our findings provide a great opportunity to overcome the translational hurdles associated with the isolation of intact, non-aggregated human iPSC-CM EVs at high purity. Furthermore, understanding in detail the interaction of the secreted EVs with their surrounding cells in the heart may open promising new avenues in the field of EV engineering for targeted delivery in cardiac regeneration.

Background Respiratory viral infections are typically more severe in older adults. Older adults are more vulnerable to infection and do not respond effectively to vaccines due to a combination of immunosenescence, so-called inflamm-ageing, and accumulation of comorbidities. Although age-related changes in immune responses have been described, the causes of this enhanced respiratory disease in older adults remain poorly understood. We therefore performed volunteer challenge with respiratory syncytial virus (RSV) in groups of younger and older adult volunteers. The aim of this study was to establish the safety and tolerability of this model and define age-related clinical, virological, and immunological outcomes. Methods In this human infection challenge pilot study, adults aged 18–55 years and 60–75 years were assessed for enrolment using protocol-defined inclusion and exclusion criteria. Symptoms were documented by self-completed diaries and viral load determined by quantitative PCR of nasal lavage. Peripheral blood B cell frequencies were measured by enzyme-linked immunospot and antibodies against pre-fusion and post-fusion, NP, and G proteins in the blood and upper respiratory tract were measured. The study was registered with ClinicalTrials.gov, NCT03728413. Findings 381 adults aged 60–75 years (older cohort) and 19 adults aged 18–55 years (young cohort) were assessed for enrolment using protocol-defined inclusion and exclusion criteria between Nov 12, 2018, and Feb 26, 2020. 12 healthy volunteers aged 60–75 years and 21 aged 18–55 years were inoculated intranasally with RSV Memphis-37. Nine (67%) of the 12 older volunteers became infected, developing mild-to-moderate upper respiratory tract symptoms that resolved without serious adverse events or sequelae. Viral load peaked on day 6 post-inoculation and symptoms peaked between days 6 and 8. Increases in circulating IgG-positive and IgA-positive antigen-specific plasmablasts, serum neutralising antibodies, and pre-F specific IgG were similar younger and older adults. However, in contrast to young participants, secretory IgA titres in older volunteers failed to increase during infection and, unlike serum IgG, did not correlate with protection. Interpretation Better understanding of age-related differences in clinical outcomes and immune correlates of protection can overcome reduction in vaccine efficacy with advancing age. We identify correlates of protection in older adults, revealing previously unrecognised factors which might have implications for targeted vaccine discovery and drug development in this vulnerable group. Funding Medical Research Council and GlaxoSmithKline EMINENT Consortium.

Rationale Type 2 innate lymphoid cells (ILC2s) are significant sources of type 2 cytokines, which are implicated in the pathogenesis of asthma and asthma exacerbations. The role of ILC2s in virus-induced asthma exacerbations is not well-characterized. Objectives To characterize pulmonary ILC responses following experimental rhinovirus challenge in patients with moderate asthma and healthy subjects. Methods Patients with moderate asthma and healthy subjects were inoculated with rhinovirus-16, and underwent bronchoscopy at baseline, day 3 and day 8 post-inoculation. Pulmonary ILC1s and ILC2s were quantified in bronchoalveolar lavage (BAL) using flow cytometry. The ratio of BAL ILC2:ILC1 was assessed to determine their relative contributions to the clinical and immune response to rhinovirus challenge. Measurements and Main Results At baseline, ILC2s were significantly higher in patients with asthma than healthy subjects. At day 8, ILC2s significantly increased from baseline in both groups, which was significantly higher in asthma than in healthy subjects (all comparisons P

BACKGROUND. Respiratory syncytial virus (RSV) is an important cause of acute pulmonary disease and one of the last remaining major infections of childhood for which there is no vaccine. CD4+ T cells play a key role in antiviral immunity, but they have been little studied in the human lung.METHODS. Healthy adult volunteers were inoculated i.n. with RSV A Memphis 37. CD4+ T cells in blood and the lower airway were analyzed by flow cytometry and immunohistochemistry. Bronchial soluble mediators were measured using quantitative PCR and MesoScale Discovery. Epitope mapping was performed by IFN-γ ELISpot screening, confirmed by in vitro MHC binding.RESULTS. Activated CD4+ T cell frequencies in bronchoalveolar lavage correlated strongly with local C-X-C motif chemokine 10 levels. Thirty-nine epitopes were identified, predominantly toward the 3′ end of the viral genome. Five novel MHC II tetramers were made using an immunodominant EFYQSTCSAVSKGYL (F-EFY) epitope restricted to HLA-DR4, -DR9, and -DR11 (combined allelic frequency: 15% in Europeans) and G-DDF restricted to HLA-DPA1*01:03/DPB1*02:01 and -DPA1*01:03/DPB1*04:01 (allelic frequency: 55%). Tetramer labeling revealed enrichment of resident memory CD4+ T (Trm) cells in the lower airway; these Trm cells displayed progressive differentiation, downregulation of costimulatory molecules, and elevated CXCR3 expression as infection evolved.CONCLUSIONS. Human infection challenge provides a unique opportunity to study the breadth of specificity and dynamics of RSV-specific T-cell responses in the target organ, allowing the precise investigation of Trm recognizing novel viral antigens over time. The new tools that we describe enable precise tracking of RSV-specific CD4+ cells, potentially accelerating the development of effective vaccines.TRIAL REGISTRATION. ClinicalTrials.gov NCT02755948.FUNDING. Medical Research Council, Wellcome Trust, National Institute for Health Research.

ADP-ribosylation is integral to a diverse range of cellular processes such as DNA repair, chromatin regulation and RNA processing. However, proteome-wide investigation of its cellular functions has been limited due to numerous technical challenges including the complexity of the poly(ADP-ribose) (PAR) chains, low abundance of the modification and lack of sensitive enrichment methods. We herein show that an adenosine analogue with a terminal alkyne functionality at position 2 of the adenine (2-alkyne adenosine or 2YnAd) is suitable for selective enrichment, fluorescence detection and mass spectrometry proteomics analysis of the candidate ADP-ribosylome in mammalian cells. Although similar labelling profiles were observed via fluorescence imaging for 2YnAd and 6YnAd, a previously reported clickable NAD+ precursor, quantitative mass spectrometry analysis of the two probes in MDA-MB-231 breast cancer cells revealed a significant increase in protein coverage of the 2YnAd probe. To facilitate global enrichment of ADP-ribosylated proteins, we developed a dual metabolic labelling approach that involves simultaneous treatment of live cells with both 2YnAd and 6YnAd. By combining this dual metabolic labelling strategy with highly sensitive tandem mass tag (TMT) isobaric mass spectrometry and hierarchical Bayesian analysis, we have quantified the responses of thousands of endogenous proteins to clinical PARP inhibitors Olaparib and Rucaparib.

INTRODUCTION: Public health measures to reduce the transmission of COVID-19 have required various changes in life-style, including loss or risk to employment and social isolation. We wished to assess how these measured effected 30-45 year old smokers at risk of COPD participating in the BLF Early COPD cohort study METHODS: At enrolment, participants were aged 30-45 years, tobacco smokers with >10 pack-year smoking history, FEV1=>80% predicted and a BMI < 35kg/m2. Participants were seen face-to-face in clinic pre-COVID. During lock-down, they were posted questionnaires and contacted by telephone. The last clinic visit took place on the 12 March 2020, remote visits took place between 16 April and 28 Sep. 260 individuals at six sites (25 Belfast, 38 Birmingham, 25 Edinburgh, 101 London, 27 Manchester and 44 Nottingham) were asked about smoking habits. The MRC chronic bronchitis questionnaire, COPD Assessment test (CAT), Leicester cough questionnaire, Hospital Anxiety and Depression questionnaire were completed in writing by the participant and returned by post or photographed and returned by email. At enrolment, the post-BD FEV1 was 3.81 (SD 0.8) litres, 101% (11) of GLI predicted. Comparisons were made by paired t-tests and chi-squared tests. RESULTS: Level of anxiety increased from 6.74 (SD 4.4) to 7.37 (SD 4.7, n=233; p=0.010) during lock-down; depression scores increased from 4.30 (3.9) to 5.14 (SD 4.1; n=233; p

BackgroundAsthma is a chronic airway disease driven by complex genetic–environmental interactions. The role of epigenetic modifications in bronchial epithelial cells (BECs) in asthma is poorly understood.MethodsWe piloted genome-wide profiling of the enhancer-associated histone modification H3K27ac in BECs from people with asthma (n = 4) and healthy controls (n = 3).ResultsWe identified n = 4,321 (FDR < 0.05) regions exhibiting differential H3K27ac enrichment between asthma and health, clustering at genes associated predominately with epithelial processes (EMT). We identified initial evidence of asthma-associated Super-Enhancers encompassing genes encoding transcription factors (TP63) and enzymes regulating lipid metabolism (PTGS1). We integrated published datasets to identify epithelium-specific transcription factors associated with H3K27ac in asthma (TP73) and identify initial relationships between asthma-associated changes in H3K27ac and transcriptional profiles. Finally, we investigated the potential of CRISPR-based approaches to functionally evaluate H3K27ac-asthma landscape in vitro by identifying guide-RNAs capable of targeting acetylation to asthma DERs and inducing gene expression (TLR3).ConclusionOur small pilot study validates genome-wide approaches for deciphering epigenetic mechanisms underlying asthma pathogenesis in the airways.

Background Rapid diagnostic tests are commonly used by hospital laboratories in England to detect rotavirus (RV), and results are used to inform clinical management and support national surveillance of the infant rotavirus immunisation programme since 2013. In 2017, the Public Health England (PHE) national reference laboratory for enteric viruses observed that the presence of RV could not be confirmed by PCR in a proportion of RV-positive samples referred for confirmatory detection. Aim We aimed to compare the positivity rate of detection methods used by hospital laboratories with the PHE confirmatory test rate. Methods Rotavirus specimens testing positive at local hospital laboratories were re-tested at the PHE national reference laboratory using a PCR test. Confirmatory results were compared to original results from the PHE laboratory information management system. Results Hospital laboratories screened 70.1% (2,608/3,721) of RV samples using immunochromatographic assay (IC) or rapid tests, 15.5% (578/3,721) using enzyme immunoassays (EIA) and 14.4% (535/3,721) using PCR. Overall, 1,011/3,721 (27.2%) locally RV-positive samples referred to PHE in 2016 and 2017 failed RV detection using the PHE reference laboratory PCR test. Confirmation rates were 66.9% (1,746/2,608) for the IC tests, 87.4% (505/578) for the EIA and 86.4% (465/535) for the PCR assays. Seasonal confirmation rate discrepancies were also evident for IC tests. Conclusions This report highlights high false positive rates with the most commonly used RV screening tests and emphasises the importance of implementing verified confirmatory tests for RV detections. This has implications for clinical diagnosis and national surveillance.

INTRODUCTION Even with intimate exposure to a virus, some people fail to become infected. Variable transmission partly depends on the dose and duration of exposure but is also governed by the immune status of the host, such as the presence of specific protective antibodies or T cells. However, for some infections, the reasons for erratic transmission are largely unknown. For example, respiratory syncytial virus (RSV) can repeatedly reinfect individuals throughout their lives despite the presence of specific immunity. Additionally, antibodies and T cells have limited efficacy against newly emergent pathogens with pandemic potential. However, the intrinsic and innate mechanisms underlying protection when people are exposed to these viruses are poorly understood. RATIONALE We reasoned that the prior state of the respiratory mucosa’s innate defenses may contribute to the variable outcome of RSV inoculation. By performing experimental challenge of adult volunteers, we were able to measure variations in the status of the nasal mucosa before inoculation and in mucosal responses during the presymptomatic phase of infection. Neither of these phases is easily observable during natural spontaneous transmission. Our observations could then be validated using specific interventional studies in a well-established mouse model of RSV infection. RESULTS After nasal administration of RSV, 57% of inoculated volunteers became infected. The uptake of infection was poorly explained by specific B or T cell immunity. However, transcriptomic profiling of the nasal tissue before inoculation demonstrated a neutrophilic inflammatory signal in those destined to develop symptomatic infection, and this was associated with suppression of an early interleukin-17 (IL-17)–dominated immune response during the presymptomatic period. This was followed by symptomatic infection associated with the expression of proinflammatory cytokines. By contrast, those who resisted infection showed a transient boost in mucosal markers of innate immune activation for several days after viral administration and no subsequent viral replication. In mice, chemokine-induced neutrophil recruitment to the airway before viral exposure transiently enhanced viral replication immediately after inoculation. As with human subjects, this was associated with decreased expression of some innate mediators in the respiratory tract, which was then followed by enhanced disease driven by CD8+ T cells. CONCLUSION Although reinfection with a respiratory virus can be partially explained by the waning of antibody titers and T cell numbers, neutrophilic inflammation in the airway at the time of pathogen exposure also predisposes individuals to symptomatic infection. After exposure, the response of the mucosa diverges: A mild and transient increase in nasal inflammatory mediators is accompanied by termination of viral infection, whereas failure to mount this response is followed by viral success. The state of innate immune preparedness of the respiratory mucosa is thus a major determinant of susceptibility to viral challenge. Our results show the importance of understanding the mucosal microenvironment in studies of respiratory infection and highlight targets for local intervention, which may enhance protection against a range of respiratory pathogens.

Raman spectroscopy enables nondestructive, label-free imaging with unprecedented molecular contrast, but is limited by slow data acquisition, largely preventing high-throughput imaging applications. Here, we present a comprehensive framework for higher-throughput molecular imaging via deep-learning-enabled Raman spectroscopy, termed DeepeR, trained on a large data set of hyperspectral Raman images, with over 1.5 million spectra (400 h of acquisition) in total. We first perform denoising and reconstruction of low signal-to-noise ratio Raman molecular signatures via deep learning, with a 10× improvement in the mean-squared error over common Raman filtering methods. Next, we develop a neural network for robust 2-4× spatial super-resolution of hyperspectral Raman images that preserve molecular cellular information. Combining these approaches, we achieve Raman imaging speed-ups of up to 40-90×, enabling good-quality cellular imaging with a high-resolution, high signal-to-noise ratio in under 1 min. We further demonstrate Raman imaging speed-up of 160×, useful for lower resolution imaging applications such as the rapid screening of large areas or for spectral pathology. Finally, transfer learning is applied to extend DeepeR from cell to tissue-scale imaging. DeepeR provides a foundation that will enable a host of higher-throughput Raman spectroscopy and molecular imaging applications across biomedicine. ispartof: ANALYTICAL CHEMISTRY vol:93 issue:48 pages:15850-15860 ispartof: location:United States status: published

It has been demonstrated that some commonly used Extracellular Vesicle (EV) isolation techniques can lead to substantial contamination with non-EV factors. Whilst it has been established that this impacts the identification of biomarkers, the impact on apparent EV bioactivity has not been explored. Extracellular vesicles have been implicated as critical mediators of therapeutic human mesenchymal stem cell (hMSC) paracrine signalling. Isolated hMSC-EVs have been used to treat multiple in vitro and in vivo models of tissue damage. However, the relative contributions of EVs and non-EV factors have not been directly compared. The dependence of hMSC paracrine signalling on EVs was first established by ultrafiltration of hMSC-conditioned medium to deplete EVs, which led to a loss of signalling activity. Here, we show that this method also causes depletion of non-EV factors, and that when this is prevented proangiogenic signalling activity is fully restored in vitro. Subsequently, we used size-exclusion chromatography (SEC) to separate EVs and soluble proteins to directly and quantitatively compare their relative contributions to signalling. Non-EV factors were found to be necessary and sufficient for the stimulation of angiogenesis and wound healing in vitro. EVs in isolation were found to be capable of potentiating signalling only when isolated by a low-purity method, or when used at comparatively high concentrations. These results indicate a potential for contaminating soluble factors to artefactually increase the apparent bioactivity of EV isolates and could have implications for future studies on the biological roles of EVs.

The global decline in prevalence of lymphatic filariasis has been one of the major successes of the WHO’s NTD programme. The recommended strategy of intensive, community-wide mass drug administration, aims to break localised transmission by either reducing the prevalence of microfilaria positive infections to below 1%, or antigen positive infections to below 2%. After the threshold is reached, and mass drug administration is stopped, geographically defined evaluation units must pass Transmission Assessment Surveys to demonstrate that transmission has been interrupted. In this study, we use an empirically parameterised stochastic transmission model to investigate the appropriateness of 1% microfilaria-positive prevalence as a stopping threshold, and statistically evaluate how well various monitoring prevalence-thresholds predict elimination or disease resurgence in the future by calculating their predictive value. Our results support the 1% filaremia prevalence target as appropriate stopping criteria. However, because at low prevalence-levels random events dominate the transmission dynamics, we find single prevalence measurements have poor predictive power for predicting resurgence, which suggests alternative criteria for restarting MDA may be beneficial., Author summary The disease lymphatic filariasis is caused by parasitic worms that are spread by mosquito vectors, and is endemic across much of Sub-Saharan Africa, Asia, South and Central America, with over 893 million people in 49 countries considered to be at-risk. In 1997 the Global Programme to Eliminate Lymphatic Filariasis was established by the WHO with the aim of eliminating the disease as a public health problem by 2020 and supplies recommended guidelines for national programs to implement. The recommended control strategy is to annually administer deworming drugs to the population at-risk until stopping criteria based on disease prevalence are met, after which the disease prevalence is periodically measured to determine if transmission is ongoing. In our study, we simulate the disease transmission in a synthetic population, both during a mass drug administration program and after it is stopped, to statistically evaluate the effectiveness of the current guidelines. We focus on the ability of prevalence measurement post-MDA cessation to predict bounce-back or elimination by estimating the predictive value of antigenemia and filaremia surveys.

Cell cryopreservation plays a key role in the development of reproducible and cost-effective cell-based therapies. Trehalose accumulated in freezing- and desiccation-tolerant organisms in nature has been sought as an attractive nontoxic cryoprotectant. Herein, we report a coincubation method for very rapid and efficient delivery of membrane-impermeable trehalose into ovine erythrocytes through reversible membrane permeabilization using pH-responsive, comb-like pseudopeptides. The pseudopeptidic polymers containing relatively long alkyl side chains were synthesized to mimic membrane-anchoring fusogenic proteins. The intracellular trehalose delivery efficiency was optimized by manipulating the side chain length, degree of substitution, and concentration of the pseudopeptides with different hydrophobic alkyl side chains, the pH, temperature, and time of incubation, as well as the polymer-to-cell ratio and the concentration of extracellular trehalose. Treatment of erythrocytes with the comb-like pseudopeptides for only 15 min yielded an intracellular trehalose concentration of 177.9 ± 8.6 mM, which resulted in 90.3 ± 0.7% survival after freeze-thaw. The very rapid and efficient delivery was found to be attributed to the reversible, pronounced membrane curvature change as a result of strong membrane insertion of the comb-like pseudopeptides. The pseudopeptides can enable efficient intracellular delivery of not only trehalose for improved cell cryopreservation but also other membrane-impermeable cargos.

Background The most commonly used diagnostic tool for soil-transmitted helminths (STH) is the Kato-Katz (KK) thick smear technique. However, numerous studies have suggested that the sensitivity of KK can be problematic, especially in low prevalence and low intensity settings. An emerging alternative is quantitative polymerase chain reaction (qPCR). Methods In this study, both KK and qPCR were conducted on stool samples from 648 participants in an STH epidemiology study conducted in the delta region of Myanmar in June 2016. Results Prevalence of any STH was 20.68% by KK and 45.06% by qPCR. Prevalence of each individual STH was also higher by qPCR than KK, the biggest difference was for hookworm with an approximately 4-fold increase between the two diagnostic techniques. Prevalence of Ancylostoma ceylanicum, a parasite predominately found in dogs, was 4.63%, indicating that there is the possibility of zoonotic transmission in the study setting. In individuals with moderate to high intensity infections there is evidence for a linear relationship between eggs per gram (EPG) of faeces, derived from KK, and DNA copy number, derived from qPCR which is particularly strong for Ascaris lumbricoides. Conclusions The use of qPCR in low prevalence settings is important to accurately assess the epidemiological situation and plan control strategies for the ‘end game’. However, more work is required to accurately assess STH intensity from qPCR results and to reduce the cost of qPCR so that is widely accessible in STH endemic countries.

ObjectiveTo characterise the clinical features of patients admitted to hospital with coronavirus disease 2019 (covid-19) in the United Kingdom during the growth phase of the first wave of this outbreak who were enrolled in the International Severe Acute Respiratory and emerging Infections Consortium (ISARIC) World Health Organization (WHO) Clinical Characterisation Protocol UK (CCP-UK) study, and to explore risk factors associated with mortality in hospital.DesignProspective observational cohort study with rapid data gathering and near real time analysis.Setting208 acute care hospitals in England, Wales, and Scotland between 6 February and 19 April 2020. A case report form developed by ISARIC and WHO was used to collect clinical data. A minimal follow-up time of two weeks (to 3 May 2020) allowed most patients to complete their hospital admission.Participants20 133 hospital inpatients with covid-19.Main outcome measuresAdmission to critical care (high dependency unit or intensive care unit) and mortality in hospital.ResultsThe median age of patients admitted to hospital with covid-19, or with a diagnosis of covid-19 made in hospital, was 73 years (interquartile range 58-82, range 0-104). More men were admitted than women (men 60%, n=12 068; women 40%, n=8065). The median duration of symptoms before admission was 4 days (interquartile range 1-8). The commonest comorbidities were chronic cardiac disease (31%, 5469/17 702), uncomplicated diabetes (21%, 3650/17 599), non-asthmatic chronic pulmonary disease (18%, 3128/17 634), and chronic kidney disease (16%, 2830/17 506); 23% (4161/18 525) had no reported major comorbidity. Overall, 41% (8199/20 133) of patients were discharged alive, 26% (5165/20 133) died, and 34% (6769/20 133) continued to receive care at the reporting date. 17% (3001/18 183) required admission to high dependency or intensive care units; of these, 28% (826/3001) were discharged alive, 32% (958/3001) died, and 41% (1217/3001) continued to receive care at the reporting date. Of those receiving mechanical ventilation, 17% (276/1658) were discharged alive, 37% (618/1658) died, and 46% (764/1658) remained in hospital. Increasing age, male sex, and comorbidities including chronic cardiac disease, non-asthmatic chronic pulmonary disease, chronic kidney disease, liver disease and obesity were associated with higher mortality in hospital.ConclusionsISARIC WHO CCP-UK is a large prospective cohort study of patients in hospital with covid-19. The study continues to enrol at the time of this report. In study participants, mortality was high, independent risk factors were increasing age, male sex, and chronic comorbidity, including obesity. This study has shown the importance of pandemic preparedness and the need to maintain readiness to launch research studies in response to outbreaks.Study registrationISRCTN66726260.

Extracellular vesicles (EVs) are secreted by the vast majority of cells and are being intensively studied due to their emerging involvement in a variety of cellular communication processes. However, the study of their cellular uptake and fate has been hampered by difficulty in imaging EVs against the cellular background. Here, we show that EVs combined with hydrophobic gold nanoclusters (AuNCs) can self-assemble into supraparticles, offering an excellent labeling strategy for high-resolution electron microscopic imaging in vitro. We have tracked and visualized the reuptake of breast cancer cell-derived EV AuNC supraparticles into their parent cells, from early endocytosis to lysosomal degradation, using focused ion beam-scanning electron microscopy (FIB-SEM). The presence of gold within the EVs and lysosomes was confirmed via DF-STEM EDX analysis of lift-out sections. The demonstrated formation of AuNC EV supraparticles will facilitate future applications in EV imaging as well as the EV-assisted cellular delivery of AuNCs.

Peptide– and protein–nanoparticle conjugates have emerged as powerful tools for biomedical applications, enabling the treatment, diagnosis, and prevention of disease. In this review, we focus on the key roles played by peptides and proteins in improving, controlling, and defining the performance of nanotechnologies. Within this framework, we provide a comprehensive overview of the key sequences and structures utilised to provide biological and physical stability to nano-constructs, direct particles to their target and influence their cellular and tissue distribution, induce and control biological responses, and form polypeptide self-assembled nanoparticles. In doing so, we highlight the great advances made by the field, as well as the challenges still faced in achieving the clinical translation of peptide- and protein-functionalised nano-drug delivery vehicles, imaging species, and active therapeutics.

Nooreen Shaikh,1,2 Malcolm Johnson,3 David A Hall,4 Kian Fan Chung,1,2 John H Riley,3 Sally Worsley,5 Pankaj K Bhavsar1,2 1Experimental Studies, National Heart and Lung Institute, Imperial College London, 2Biomedical Research Unit, Royal Brompton and Harefield NHS Trust, London, 3Respiratory Global Franchise, GlaxoSmithKline, Uxbridge, 4Fibrosis and Lung Injury Development Planning Unit, GlaxoSmithKline, Stevenage, 5Respiratory Research & Development, GlaxoSmithKline, Uxbridge, UK Background: Intracellular mechanisms of action of umeclidinium (UMEC), a long-acting muscarinic receptor antagonist, and vilanterol (VI), a long-acting β2-adrenoceptor (β2R) agonist, were investigated in target cells: human airway smooth-muscle cells (ASMCs). Materials and methods: ASMCs from tracheas of healthy lung-transplant donors were treated with VI, UMEC, UMEC and VI combined, or control compounds (salmeterol, propranolol, ICI 118.551, or methacholine [MCh]). Cyclic adenosine monophosphate (cAMP) was measured using an enzyme-linked immunosorbent assay, intracellular free calcium ([Ca2+]i) using a fluorescence assay, and regulator of G-protein signaling 2 (RGS2) messenger RNA using real-time quantitative polymerase chain reaction. Results: VI and salmeterol (10–12–10–6 M) induced cAMP production from ASMCs in a concentration-dependent manner, which was greater for VI at all concentrations. β2R antagonism by propranolol or ICI 118.551 (10–12–10–4 M) resulted in concentration-dependent inhibition of VI-induced cAMP production, and ICI 118.551 was more potent. MCh (5×10–6 M, 30 minutes) attenuated VI-induced cAMP production (P

Pulmonary arterial hypertension (PAH), at one time a largely overlooked disease, is now the subject of intense study in many academic and biotech groups. The availability of new treatments has increased awareness of the condition. This in turn has driven a change in the demographics of PAH, with an increase in the mean age at diagnosis. The diagnosis of PAH in more elderly patients has highlighted the need for careful phenotyping of patients and for further studies to understand how best to manage pulmonary hypertension associated with, for example, left heart disease. The breadth and depth of expertise focused on unravelling the molecular pathology of PAH has yielded novel insights, including the role of growth factors, inflammation and metabolic remodelling. The description of the genetic architecture of PAH is accelerating in parallel, with novel variants, such as those reported in potassium two-pore domain channel subfamily K member 3 (KCNK3), adding to the list of more established mutations in genes associated with bone morphogenetic protein receptor type 2 (BMPR2) signalling. These insights have supported a paradigm shift in treatment strategies away from simply addressing the imbalance of vasoactive mediators observed in PAH towards tackling more directly the structural remodelling of the pulmonary vasculature. Here, we summarize the changing clinical and molecular landscape of PAH. We highlight novel drug therapies that are in various stages of clinical development, targeting for example cell proliferation, metabolic, inflammatory/immune and BMPR2 dysfunction, and the challenges around developing these treatments. We argue that advances in the treatment of PAH will come through deep molecular phenotyping with the integration of clinical, genomic, transcriptomic, proteomic and metabolomic information in large populations of patients through international collaboration. This approach provides the best opportunity for identifying key signalling pathways, both as potential drug targets and as biomarkers for patient selection. The expectation is that together these will enable the prioritization of potential therapies in development and the evolution of personalized medicine for PAH.

Applying machine learning of complex motion phenotypes obtained from cardiac MR images allows more accurate prediction of patient outcomes in pulmonary hypertension., Purpose To determine if patient survival and mechanisms of right ventricular failure in pulmonary hypertension could be predicted by using supervised machine learning of three-dimensional patterns of systolic cardiac motion. Materials and Methods The study was approved by a research ethics committee, and participants gave written informed consent. Two hundred fifty-six patients (143 women; mean age ± standard deviation, 63 years ± 17) with newly diagnosed pulmonary hypertension underwent cardiac magnetic resonance (MR) imaging, right-sided heart catheterization, and 6-minute walk testing with a median follow-up of 4.0 years. Semiautomated segmentation of short-axis cine images was used to create a three-dimensional model of right ventricular motion. Supervised principal components analysis was used to identify patterns of systolic motion that were most strongly predictive of survival. Survival prediction was assessed by using difference in median survival time and area under the curve with time-dependent receiver operating characteristic analysis for 1-year survival. Results At the end of follow-up, 36% of patients (93 of 256) died, and one underwent lung transplantation. Poor outcome was predicted by a loss of effective contraction in the septum and free wall, coupled with reduced basal longitudinal motion. When added to conventional imaging and hemodynamic, functional, and clinical markers, three-dimensional cardiac motion improved survival prediction (area under the receiver operating characteristic curve, 0.73 vs 0.60, respectively; P < .001) and provided greater differentiation according to difference in median survival time between high- and low-risk groups (13.8 vs 10.7 years, respectively; P < .001). Conclusion A machine-learning survival model that uses three-dimensional cardiac motion predicts outcome independent of conventional risk factors in patients with newly diagnosed pulmonary hypertension. Online supplemental material is available for this article.

Highlights • This paper compares two mathematical models describing the transmission dynamics of schistosome infection and the impact of mass drug administration. • The models differ structurally in a number of ways, including the dynamics of the intermediate snail host and the treatment of adult worms within the human host. • The models are validated against data taken from a mass-drug administration trial in Mozambique. • The differences between the model predictions and the data are discussed in the context of the structural differences between the models., The predictions of two mathematical models describing the transmission dynamics of schistosome infection and the impact of mass drug administration are compared. The models differ in their description of the dynamics of the parasites within the host population and in their representation of the stages of the parasite lifecycle outside of the host. Key parameters are estimated from data collected in northern Mozambique from 2011 to 2015. This type of data set is valuable for model validation as treatment prior to the study was minimal. Predictions from both models are compared with each other and with epidemiological observations. Both models have difficulty matching both the intensity and prevalence of disease in the datasets and are only partially successful at predicting the impact of treatment. The models also differ from each other in their predictions, both quantitatively and qualitatively, of the long-term impact of 10 years’ school-based mass drug administration. We trace the dynamical differences back to basic assumptions about worm aggregation, force of infection and the dynamics of the parasite in the snail population in the two models and suggest data which could discriminate between them. We also discuss limitations with the datasets used and ways in which data collection could be improved.

Objectives: To characterize peak cardiac troponin levels, in patients presenting with acute myocardial infarction (AMI), according to their comorbid condition and determine the influence of peak troponin (cTn) levels on mortality. Methods: We included patients with the first admission for AMI in the United Kingdom. We used linear regression to estimate the association between eight common co-morbidities {diabetes mellitus(DM), previous angina, peripheral arterial disease(PAD), previous myocardial infarction(MI), chronic kidney disease(CKD), cerebrovascular disease(CVD), chronic heart failure(CHF), and chronic obstructive pulmonary disease(COPD)} and peak cTn. Peak cTn levels were adjusted for age, sex, smoking status and co-morbidities. Logistic regression and restricted cubic spline models were employed to investigate the association between peak cTn and 180-day mortality for each co-morbidity. Results: 330,367 patients with AMI were identified. Adjusted peak cTn levels were significantly higher in patients with CKD[adjusted % difference in peak cTnT for CKD=42%, 95%CI 13.1 to 78.4] and significantly lower for patients with COPD, previous angina, previous MI and CHF when compared to patients without the respective co-morbidities (reference group) [cTnI;COPD=-21.7%,95%CI -29.1 to -13.4;previous angina=-24.2%, 95%CI -29.6 to -8.3;previous MI=-13.5%, 95%CI -20.6 to -5.9;CHF=-28% 95%CI -37.2 to -17.6]. Risk of 180-day mortality in most of the co-morbidities did not change substantially after adjusting for peak cTn. In general, cTnI had a stronger association with mortality than cTnT. Conclusions: In this nationwide analyses of patients presenting with acute myocardial infarction, co-morbidities substantially influenced systemic concentrations of peak cTn. Comorbid illness is a significant predictor of mortality regardless of peak cTn levels and should be taken into consideration while interpreting cTn both as a diagnostic and prognostic biomarker.

Rate of FEV1 decline in COPD is heterogeneous and the extent to which inhaled corticosteroids (ICS) influence the rate of decline is unclear. The majority of previous reviews have investigated specific ICS and non-ICS inhalers and have consisted of randomised control trials (RCTs), which have specific inclusion and exclusion criteria and short follow up times. We aimed to investigate the association between change in FEV1 and ICS-containing medications in COPD patients over longer follow up times. MEDLINE and EMBASE were searched and literature comparing change in FEV1 in COPD patients taking ICS-containing medications with patients taking non-ICS-containing medications were identified. Titles, abstract, and full texts were screened and information extracted using the PICO checklist. Risk of bias was assessed using the Cochrane Risk of Bias tool and a descriptive synthesis of the literature was carried out due to the heterogeneity of included studies. 17 studies met our inclusion criteria. We found that the difference in change in FEV1 in people using ICS and non-ICS containing medications depended on the study follow-up time. Shorter follow-up studies (1 year or less) were more likely to report an increase in FEV1 from baseline in both patients on ICS and in patients on non-ICS-containing medications, with the majority of these studies showing a greater increase in FEV1 in patients on ICS-containing medications. Longer follow-up studies (greater than 1 year) were more likely to report a decline in FEV1 from baseline in patients on ICS and in patients on non-ICS containing medications but differences between rates of FEV1 of decline were not significant. Further studies are needed to better understand changes in FEV1 when ICS-containing medications are prescribed and to determine whether ICS-containing medications influence rate of decline in FEV1 in the long term. Results from inclusive trials and observational patient cohorts may provide information more generalisable to a population of COPD patients.

Deep-eutectic solvents and room temperature ionic liquids are increasingly recognised as appropriate materials for use as active pharmaceutical ingredients and formulation additives. Aqueous mixtures of choline and geranate (CAGE), in particular, have been shown to offer promising biomedical properties but understanding the thermophysical behaviour of these mixtures remains limited. Here, we develop interaction potentials for use in the SAFT-γ Mie group-contribution approach, to study the thermodynamic properties and phase behaviour of aqueous mixtures of choline geranate and geranic acid. The determination of the interaction parameters between chemical functional groups is carried out in a sequential fashion, characterising each group based on those previously developed. The parameters of the groups relevant to geranic acid are estimated using experimental fluid phase-equilibrium data such as vapour pressure and saturated-liquid density of simple pure components (n-alkenes, branched alkenes and carboxylic acids) and the phase equilibrium data of mixtures (aqueous solutions of branched alkenes and of carboxylic acids). Geranate is represented by further incorporating the anionic carboxylate group, COO−, which is characterised using aqueous solution data of sodium carboxylate salts, assuming full dissociation of the salt in water. Choline is described by incorporating the cationic quaternary ammonium group, N+, using data for choline chloride solutions. The osmotic pressure of aqueous mixtures of CAGE at several concentrations is predicted and compared to experimental data obtained as part of our work to assess the accuracy of the modelling platform. The SAFT-γ Mie approach is shown to be predictive, providing a good description of the measured data for a wide range of mixtures and properties. Furthermore, the new group-interaction parameters needed to represent CAGE extend the set of functional groups of the group-contribution approach, and can be used in a transferable way to predict the properties of systems beyond those studied in the current work.

ObjectivesTo characterise peak cardiac troponin levels, in patients presenting with acute myocardial infarction (AMI), according to their comorbid condition and determine the influence of peak cardiac troponin (cTn) levels on mortality.MethodsWe included patients with the first admission for AMI in the UK. We used linear regression to estimate the association between eight common comorbidities (diabetes mellitus, previous angina, peripheral arterial disease, previous myocardial infarction (MI), chronic kidney disease (CKD), cerebrovascular disease, chronic heart failure (CHF) and chronic obstructive pulmonary disease (COPD)) and peak cTn. Peak cTn levels were adjusted for age, sex, smoking status and comorbidities. Logistic regression and restricted cubic spline models were employed to investigate the association between peak cTn and 180-day mortality for each comorbidity.Results330 367 patients with ST elevation myocardial infarction and non-ST elevation myocardial infarction were identified. Adjusted peak cTn levels were significantly higher in patients with CKD (adjusted % difference in peak cTnT for CKD=42%, 95% CI 13.1 to 78.4) and significantly lower for patients with COPD, previous angina, previous MI and CHF when compared with patients without the respective comorbidities (reference group) (cTnI; COPD=−21.7%, 95% CI −29.1 to −13.4; previous angina=−24.2%, 95% CI −29.6 to −8.3; previous MI=−13.5%, 95% CI −20.6 to −5.9; CHF=−28%, 95% CI −37.2 to −17.6). Risk of 180-day mortality in most of the comorbidities did not change substantially after adjusting for peak cTn. In general, cTnI had a stronger association with mortality than cTnT.ConclusionsIn this nationwide analysis of patients presenting with AMI, comorbidities substantially influenced systemic concentrations of peak cTn. Comorbid illness is a significant predictor of mortality regardless of peak cTn levels and should be taken into consideration while interpreting cTn both as a diagnostic and prognostic biomarker.

Capturing the dynamical properties of time series concisely as interpretable feature vectors can enable efficient clustering and classification for time-series applications across science and industry. Selecting an appropriate feature-based representation of time series for a given application can be achieved through systematic comparison across a comprehensive time-series feature library, such as those in the hctsa toolbox. However, this approach is computationally expensive and involves evaluating many similar features, limiting the widespread adoption of feature-based representations of time series for real-world applications. In this work, we introduce a method to infer small sets of time-series features that (i) exhibit strong classification performance across a given collection of time-series problems, and (ii) are minimally redundant. Applying our method to a set of 93 time-series classification datasets (containing over 147 000 time series) and using a filtered version of the hctsa feature library (4791 features), we introduce a set of 22 CAnonical Timeseries CHaracteristics, catch22, tailored to the dynamics typically encountered in time-series data-mining tasks. This dimensionality reduction, from 4791 to 22, is associated with an approximately 1000-fold reduction in computation time and near linear scaling with time-series length, despite an average reduction in classification accuracy of just 7%. catch22 captures a diverse and interpretable signature of time series in terms of their properties, including linear and non-linear autocorrelation, successive differences, value distributions and outliers, and fluctuation scaling properties. We provide an efficient implementation of catch22, accessible from many programming environments, that facilitates feature-based time-series analysis for scientific, industrial, financial and medical applications using a common language of interpretable time-series properties.

The gut microbiome is an important factor in human health. It is affected by what we eat, what medicines we take, and what infections we acquire. In turn, it affects the way we absorb nutrients and whether we have excessive intestinal inflammation. Intestinal worms may have an important impact on the composition of the gut microbiome. Without a complete understanding of the impact of mass deworming programs on the microbiome, it is impossible to accurately calculate the cost-effectiveness of such public health interventions and to guard against any possible deleterious side effects. Our research examines this question in a “real-world” setting, using a longitudinal cohort, in which individuals with and without worm infections are treated with deworming medication and followed up at both three weeks and three months posttreatment. We quantify the impact of roundworms and hookworms on gut microbial composition, suggesting that the impact is small, but that treatment of hookworm infection results in significant changes. This work points to the need for follow-up studies to further examine the impact of hookworm on the gut microbiota and determine the health consequences of the observed changes., Murine studies suggest that the presence of some species of intestinal helminths is associated with changes in host microbiota composition and diversity. However, studies in humans have produced varied conclusions, and the impact appears to vary widely depending on the helminth species present. To demonstrate how molecular approaches to the human gut microbiome can provide insights into the complex interplay among disparate organisms, DNA was extracted from cryopreserved stools collected from residents of 5 rural Kenyan villages prior to and 3 weeks and 3 months following albendazole (ALB) therapy. Samples were analyzed by quantitative PCR (qPCR) for the presence of 8 species of intestinal parasites and by MiSeq 16S rRNA gene sequencing. Based on pretreatment results, the presence of neither Ascaris lumbricoides nor Necator americanus infection significantly altered the overall diversity of the microbiota in comparison with age-matched controls. Following ALB therapy and clearance of soil-transmitted helminths (STH), there were significant increases in the proportion of the microbiota made up by Clostridiales (P = 0.0002; average fold change, 0.57) and reductions in the proportion made up by Enterobacteriales (P = 0.0004; average fold change, −0.58). There was a significant posttreatment decrease in Chao1 richness, even among individuals who were uninfected pretreatment, suggesting that antimicrobial effects must be considered in any posttreatment setting. Nevertheless, the helminth-associated changes in Clostridiales and Enterobacteriales suggest that clearance of STH, and of N. americanus in particular, alters the gut microbiota.

Hannah R Whittaker,1 Hana Müllerova,2 Deborah Jarvis,1 Neil C Barnes,2 Paul W Jones,2 Chris H Compton,2 Steven J Kiddle,3 Jennifer K Quint11National Heart and Lung Institute, Imperial College London, London, UK; 2Respiratory Epidemiology, GlaxoSmithKline R&D, Uxbridge, UK; 3MRC Biostatistics Unit, University of Cambridge, Cambridge, UKBackground: Inhaled corticosteroid (ICS)-containing medications slow rate of decline of FEV1. Blood eosinophil (EOS) levels are associated with the degree of exacerbation reduction with ICS.Purpose: We investigated whether FEV1 decline differs between patients with and without ICS, stratified by blood EOS level.Patients and methods: The UK Clinical Practice Research Datalink (primary care records) and Hospital Episode Statistics (hospital records) were used to identify COPD patients aged 35 years or older, who were current or ex-smokers with ≥2 FEV1 measurements ≥6 months apart. Prevalent ICS use and the nearest EOS count to start of follow-up were identified. Patients were classified at baseline as higher stratum EOS (≥150 cell/μL) on ICS; higher stratum EOS not on ICS; lower stratum EOS (

Background: Viruses play an important role in asthma exacerbations and are detected by Toll-like receptors (TLRs). Better characterization of mucosal innate immunity to viral triggers may help understand dysregulated host responses in asthma. Aims & Objectives: A synthetic analogue of single-stranded RNA (ssRNA) and TLR7/8 agonist resiquimod (R848) was administered in vivo to study the effect of allergy and asthma on nasal mucosal innate immune responses. Methods: Nasal spray with saline and R848 was administered to healthy non-allergic (n=12), allergic rhinitis (n=12) and allergic asthma (n=11) participants. Immune mediators from nasal and blood samples, nasal mucosal gene expression and peripheral differential cell counts were measured. Results: R848 was well tolerated with no evidence of systemic immune activation. R848 significantly induced nasal mucosal IFN-a2a, IFN-?, pro-inflammatory cytokines (TNF-a, IL-2, IL-12p70) and chemokines (CXCL10, CCL2, CCL3, CCL4 and CCL13) compared to saline. Participants with allergic rhinitis and asthma had increased IFN-a2a, CCL3 and CCL13 relative to healthy participants, whilst those with asthma alone had increased gene expression of interferon stimulated genes DDX58, MX1 and IFIT3. Nasal R848 administration was associated with a decrease in blood eosinophils at 4h and decrease in peripheral lymphocytes at 24h, a finding restricted to participants with allergic rhinitis and asthma. Conclusions: These results confirm the suitability of nasal delivery of R848 as a non-invasive tool to assess mucosal innate immunity and highlights a key role for asthma in determining host responses to viral RNA analogues.

Peripheral nerve decoding algorithms form an important component of closed-loop bioelectronic medicines devices. For any decoding method, meaningful properties need to be extracted from the peripheral nerve signal as the first step. Simple measures such as signal amplitude and features of the Fourier power spectrum are most typically used, leaving open whether important information is encoded in more subtle properties of the dynamics. We here propose a feature-based analysis method that identifies changes in firing characteristics across recording sections by unsupervised dimensionality reduction in a high-dimensional feature-space and selects single efficiently implementable estimators for each characteristic to be used as the basis for a better decoding in future bioelectronic medicines devices.

Invasive group A streptococcal (GAS) disease is uncommon but carries a high case-fatality rate relative to other infectious diseases. Given the ubiquity of mild GAS infections, it remains unclear why healthy individuals will occasionally develop life-threatening infections, raising the possibility of host genetic predisposition. Here, we present the results of a case-control study including 43 invasive GAS cases and 1,540 controls. Using HLA imputation and linear mixed-models, we find each copy of theHLA-DQA1*01:03 allele associates with a two-fold increased risk of disease (odds ratio 2.3, 95% confidence interval 1.3-4.4,P=0.009), an association which persists with classical HLA typing of a subset of cases and analysis with an alternative large control dataset with validated HLA data. Moreover, we propose the association is driven by the allele itself rather than the background haplotype. Overall this finding provides impetus for further investigation of the immunogenetic basis of this devastating bacterial disease.

Background Of over 90 serotypes of Streptococcus pneumoniae, only seven were included in the first pneumococcal conjugate vaccine (PCV). While PCV reduced the disease incidence, in part because of a herd immunity effect, a replacement effect was observed whereby disease was increasingly caused by serotypes not included in the vaccine. Dynamic transmission models can account for these effects to describe post-vaccination scenarios, whereas economic evaluations can enable decision-makers to compare vaccines of increasing valency for implementation. Aim The aim of this review was to examine epidemiological and economic models and their assumptions for their potential contributions to future research and immunisation policy. Sources Pubmed, Scopus, Ovid, ISI Web of Knowledge, Centre of Reviews and Dissemination (CRD) databases were searched. Content Twenty-three dynamic transmission models and 21 economic models were retrieved and reviewed. Published models employed various templates, revealing several key uncertainties regarding the biology and epidemiology of pneumococcal infection. While models suggested that PCVs will reduce the burden of disease, the extent to which they are predicted to do so depended on various assumptions regarding features of pneumococcal infection and epidemiology that governed PCV cost-effectiveness as well. Such features include the duration of protection and competitive interactions between serotypes, which are unclear at present, but which directly relate to herd immunity and serotype replacement. Implications Economic evaluations are not typically based on transmission dynamic models and hence omit indirect herd immunity effects. The two tools could be used in conjunction to inform decision-makers on vaccine implementation, but so far there have been few attempts to build economic evaluations on transmission dynamic models, and none in this field. Future directions for research could include studies to evaluate key parameters for the models involving herd immunity, serotype competition and the natural history of infection.

Organic solvent nanofiltration (OSN) technology is a membrane process for molecular separation in harsh organic media. However, despite having well-documented potential applications, development hurdles have hindered the widespread uptake of OSN technology. One of the most promising areas of application is as an iterative synthesis platform, for instance for oligonucleotides or peptides, where a thorough purification step is required after each synthesis cycle, preferably in the same working solvent. In this work, we report a process development study for liquid-phase oligonucleotide synthesis (LPOS) using OSN technology. Oligonucleotide (oligo) based drugs are being advanced as a new generation of therapeutics functioning at the protein expression level. Currently, over 100 oligo based drugs are undergoing clinical trials, suggesting that it will soon be necessary to produce oligos at a scale of metric tons per year. However, there are as yet no synthesis platforms that can manufacture oligos at >10 kg batch scale. With the process developed here, we have successfully carried out eight iterative cycles of chain extension and synthesized 5-mer and 9-mer 2′-O-methyl oligoribonucleotide phosphorothioates, all in liquid phase media. This paper discusses the key challenges, both anticipated and unexpected, faced during development of this process and suggests solutions to reduce the development period. An economic analysis has been carried out, highlighting the potential competitiveness of the LPOS-OSN process and the necessity for a solvent recovery unit.

BACKGROUND: Exacerbations of asthma and COPD are triggered by rhinoviruses. Uncontrolled inflammatory pathways, pathogenic bacterial burden and impaired antiviral immunity are thought to be important factors in disease severity and duration. Macrolides including azithromycin are often used to treat the above diseases, but exhibit variable levels of efficacy. Inhaled corticosteroids are also readily used in treatment, but may lack specificity. Ideally, new treatment alternatives should suppress unwanted inflammation, but spare beneficial antiviral immunity. METHODS: In the present study, we screened 225 novel macrolides and tested them for enhanced antiviral activity against rhinovirus, as well as anti-inflammatory activity and activity against Gram-positive and Gram-negative bacteria. Primary bronchial epithelial cells were grown from 10 asthmatic individuals and the effects of macrolides on rhinovirus replication were also examined. Another 30 structurally similar macrolides were also examined. RESULTS: The oleandomycin derivative Mac5, compared with azithromycin, showed superior induction (up to 5-fold, EC50 = 5-11 μM) of rhinovirus-induced type I IFNβ, type III IFNλ1 and type III IFNλ2/3 mRNA and the IFN-stimulated genes viperin and MxA, yet had no effect on IL-6 and IL-8 mRNA. Mac5 also suppressed rhinovirus replication at 48 h, proving antiviral activity. Mac5 showed antibacterial activity against Gram-positive Streptococcus pneumoniae; however, it did not have any antibacterial properties compared with azithromycin when used against Gram-negative Escherichia coli (as a model organism) and also the respiratory pathogens Pseudomonas aeruginosa and non-typeable Haemophilus influenzae. Further non-toxic Mac5 derivatives were identified with various anti-inflammatory, antiviral and antibacterial activities. CONCLUSIONS: The data support the idea that macrolides have antiviral properties through a mechanism that is yet to be ascertained. We also provide evidence that macrolides can be developed with anti-inflammatory, antibacterial and antiviral activity and show surprising versatility depending on the clinical need.

BACKGROUND: Depression, a condition commonly comorbid with multiple sclerosis (MS), is associated more generally with elevated inflammatory markers and hippocampal pathology. We hypothesized that neuroinflammation in the hippocampus is responsible for depression associated with MS. We characterized the relationship between depressive symptoms and hippocampal microglial activation in patients with MS using the 18-kDa translocator protein radioligand [(18)F]PBR111. To evaluate pathophysiologic mechanisms, we explored the relationships between hippocampal neuroinflammation, depressive symptoms, and hippocampal functional connectivities defined by resting-state functional magnetic resonance imaging.METHODS: The Beck Depression Inventory (BDI) was administered to 11 patients with MS and 22 healthy control subjects before scanning with positron emission tomography and functional magnetic resonance imaging. We tested for higher [(18)F]PBR111 uptake in the hippocampus of patients with MS relative to healthy control subjects and examined the correlations between [(18)F]PBR111 uptake, BDI scores, and hippocampal functional connectivities in the patients with MS.RESULTS: Patients with MS had an increased hippocampal [(18)F]PBR111 distribution volume ratio relative to healthy control subjects (p = .024), and the hippocampal distribution volume ratio was strongly correlated with the BDI score in patients with MS (r = .86, p = .006). Hippocampal functional connectivities to the subgenual cingulate and prefrontal and parietal regions correlated with BDI scores and [(18)F]PBR111 distribution volume ratio.CONCLUSIONS: Our results provide evidence that hippocampal microglial activation in MS impairs the brain functional connectivities in regions contributing to maintenance of a normal affective state. Our results suggest a rationale for the responsiveness of depression in some patients with MS to effective control of brain neuroinflammation. Our findings also lend support to further investigation of the role of inflammatory processes in the pathogenesis of depression more generally.

The epilepsies affect around 65 million people worldwide and have a substantial missing heritability component. We report a genome-wide mega-analysis involving 15,212 individuals with epilepsy and 29,677 controls, which reveals 16 genome-wide significant loci, of which 11 are novel. Using various prioritization criteria, we pinpoint the 21 most likely epilepsy genes at these loci, with the majority in genetic generalized epilepsies. These genes have diverse biological functions, including coding for ion-channel subunits, transcription factors and a vitamin-B6 metabolism enzyme. Converging evidence shows that the common variants associated with epilepsy play a role in epigenetic regulation of gene expression in the brain. The results show an enrichment for monogenic epilepsy genes as well as known targets of antiepileptic drugs. Using SNP-based heritability analyses we disentangle both the unique and overlapping genetic basis to seven different epilepsy subtypes. Together, these findings provide leads for epilepsy therapies based on underlying pathophysiology., SCOPUS: ar.j, info:eu-repo/semantics/published

Aims We sought to identify metabolic pathways associated with right ventricular (RV) adaptation to pulmonary hypertension (PH). We evaluated candidate metabolites, previously associated with survival in pulmonary arterial hypertension, and used automated image segmentation and parametric mapping to model their relationship to adverse patterns of remodelling and wall stress. Methods and results In 312 PH subjects (47.1% female, mean age 60.8 ± 15.9 years), of which 182 (50.5% female, mean age 58.6 ± 16.8 years) had metabolomics, we modelled the relationship between the RV phenotype, haemodynamic state, and metabolite levels. Atlas-based segmentation and co-registration of cardiac magnetic resonance imaging was used to create a quantitative 3D model of RV geometry and function—including maps of regional wall stress. Increasing mean pulmonary artery pressure was associated with hypertrophy of the basal free wall (β = 0.29) and reduced relative wall thickness (β = −0.38), indicative of eccentric remodelling. Wall stress was an independent predictor of all-cause mortality (hazard ratio = 1.27, P = 0.04). Six metabolites were significantly associated with elevated wall stress (β = 0.28–0.34) including increased levels of tRNA-specific modified nucleosides and fatty acid acylcarnitines, and decreased levels (β = −0.40) of sulfated androgen. Conclusion Using computational image phenotyping, we identify metabolic profiles, reporting on energy metabolism and cellular stress-response, which are associated with adaptive RV mechanisms to PH.

GATA-3 plays a critical role in regulating the expression of the cytokines interleukin (IL)-4, IL-5, and IL-13 from T helper-2 (Th2) cells and therefore is a key mediator of allergic diseases. Corticosteroids are highly effective in suppressing allergic inflammation, but their effects on GATA-3 are unknown. We investigated the effect of the corticosteroid fluticasone propionate on GATA-3 regulation in human T-lymphocytes in vitro and in vivo.In a T lymphocyte cell line (HuT-78) and peripheral blood mononuclear cells stimulated by anti-CD3 and anti-CD28 in vitro we demonstrated that fluticasone inhibits nuclear translocation of GATA-3 and expression of Th2 cytokines via a mechanism independent of nuclear factor-kappaB and is due, in part, to competition between GATA-3 and the ligand-activated glucocorticoid receptor for nuclear transport through the nuclear importer importin-alpha. In addition, fluticasone induces the expression of mitogen-activated protein kinase (MAPK) phosphatase-1 (MKP-1), the endogenous inhibitor of p38 MAPK, which is necessary for GATA-3 nuclear translocation. These inhibitory effects of fluticasone are rapid, potent, and prolonged. We also demonstrated that inhaled fluticasone inhibits GATA-3 nuclear translocation in peripheral blood lymphocytes of patients with asthma in vivo.Corticosteroids have a potent inhibitory effect on GATA-3 via two interacting mechanisms that potently suppress Th2 cytokine expression. This novel mechanism of action of corticosteroids may account for the striking clinical efficacy of corticosteroids in the treatment of allergic diseases.

Bioelectronic Medicines that modulate the activity patterns on peripheral nerves have promise as a new way of treating diverse medical conditions from epilepsy to rheumatism. Progress in the field builds upon time consuming and expensive experiments in living organisms. To reduce experimentation load and allow for a faster, more detailed analysis of peripheral nerve stimulation and recording, computational models incorporating experimental insights will be of great help. We present a peripheral nerve simulator that combines biophysical axon models and numerically solved and idealised extracellular space models in one environment. We modelled the extracellular space as a three-dimensional resistive continuum governed by the electro-quasistatic approximation of the Maxwell equations. Potential distributions were precomputed in finite element models for different media (homogeneous, nerve in saline, nerve in cuff) and imported into our simulator. Axons, on the other hand, were modelled more abstractly as one-dimensional chains of compartments. Unmyelinated fibres were based on the Hodgkin-Huxley model; for myelinated fibres, we adapted the model proposed by McIntyre et al. in 2002 to smaller diameters. To obtain realistic axon shapes, an iterative algorithm positioned fibres along the nerve with a variable tortuosity fit to imaged trajectories. We validated our model with data from the stimulated rat vagus nerve. Simulation results predicted that tortuosity alters recorded signal shapes and increases stimulation thresholds. The model we developed can easily be adapted to different nerves, and may be of use for Bioelectronic Medicine research in the future.

Rationale: Previous studies have suggested that acute exacerbations of COPD may be associated with increased risk of myocardial infarction and ischaemic stroke. Objective: We aimed to quantify the increased risks of myocardial infarction and ischemic stroke risk associated with both moderate and severe acute exacerbation, and to investigate factors which may modify these risks. Methods: We performed a self-controlled case-series to investigate the rates of myocardial infarction and ischemic stroke following acute exacerbation compared to stable time, within individuals. The participants were 5,696 adults with COPD with a first myocardial infarction (n = 2,850) or ischemic stroke (n = 3,010) and at least one acute exacerbation from the UK Clinical Practice Research Datalink with linked Hospital Episodes Statistics data. Results: The risks of both myocardial infarction and ischemic stroke were increased in the 91 days following a acute exacerbation. The risks were greater following a severe exacerbation (incidence rate ratio (IRR) of 2.58 (95% CI 2.26 to 2.95) for myocardial infarction; and IRR 1.97 (95% CI 1.66-2.33) for ischemic stroke) than after a moderate exacerbation (IRR 1.58 (95% CI 1.46-1.71) for myocardial infarction; and IRR 1.45 (95% CI 1.33-1.57) for ischemic stroke. The relative risks of myocardial infarction and ischemic stroke associated with acute exacerbation were lower among those with more frequent exacerbations (IRR 1.42 (95% CI 1.24-1.62) vs 1.69 (95% CI 1.50-1.91) for myocardial infarction; and IRR 1.30 (95% CI 1.15-1.48) vs 1.68 (95% CI 1.50-1.89) for ischemic stroke). Higher GOLD stage associated with a lower rate of myocardial infarction (IRR 1.98 (95% CI 1.61-2.05) vs 1.69 (95% CI 1.45-1.98)) but not for ischemic stroke. Aspirin use at baseline was associated with a lower risk of ischemic stroke (IRR 1.28 (95% CI 1.10-1.50) vs 1.63 (95% CI 1.47-1.80)), but not with myocardial infarction. Conclusions: Acute exacerbations of COPD are associated with an increased risk of myocardial infarction and ischemic stroke within 28 days of their onset. Several patient characteristics were identified which are associated with these events.

Extracellular vesicles (EVs), sub-micron vectors used in intercellular communication, have demonstrated great promise as natural drug delivery systems. Recent reports have detailed impressive in vivo results from the administration of EVs pre-loaded with therapeutic cargo, including small molecules, nanoparticles, proteins and oligonucleotides. These results have sparked intensive research interest across a huge range of disease models. There are, however, enduring limitations that have restricted widespread clinical and pharmaceutical adoption. In this perspective, we discuss these practical and biological concerns, critically compare the relative merit of EVs and synthetic drug delivery systems, and highlight the need for a more comprehensive understanding of in vivo transport and delivery. Within this framework, we seek to establish key areas in which EVs can gain a competitive advantage in order to provide the tangible added value required for widespread translation.

Rationale: Acute exacerbations of chronic obstructive pulmonary disease (AECOPDs) are important adverse events in the natural history of chronic obstructive pulmonary disease (COPD). Objectives: To investigate the natural history of AECOPDs over 10 years of follow-up. Methods: We identified 99,574 patients with COPD from January 1, 2004, to March 31, 2015, from the UK Clinical Practice Research Datalink. We defined moderate AECOPDs as those managed outside hospital and severe as those requiring hospitalization. During the baseline period (first year of follow-up), patients were grouped according to the number and severity of AECOPDs and then followed for a maximum of 10 years (mean, 4.9 yr). We investigated the effect of baseline AECOPD number and severity on risk of further events and death. Measurements and Main Results: Around one-quarter of the patients with COPD did not exacerbate during follow-up. Compared with no AECOPDs in the baseline period, AECOPD number predicted the future long-term rate of AECOPDs in a graduated fashion, ranging from hazard ratio (HR) of 1.71 (1.66–1.77) for one event to HR of 3.41 (3.27–3.56) for five or more events. Two or more moderate AECOPDs were also associated with an increased risk of death in a graduated fashion, ranging from HR of 1.10 (1.03–1.18) for two moderate AECOPDs to HR of 1.57 (1.45–1.70) for five or more moderate AECOPDs, compared with those with no AECOPDs at baseline. Severe AECOPDs were associated with an even higher risk of death (HR, 1.79; 1.65–1.94). Conclusions: A large proportion of patients with COPD do not exacerbate over a maximum 10 years of follow-up. AECOPD frequency in a single year predicts long-term AECOPD rate. Increasing frequency and severity of AECOPDs is associated with risk of death and highlights the importance of preventing AECOPDs.

Due to the discovery of RNAi, oligonucleotides (oligos) have re-emerged as a major pharmaceutical target that may soon be required in ton quantities. However, it is questionable whether solid-phase oligo synthesis (SPOS) methods can provide a scalable synthesis. Liquid-phase oligo synthesis (LPOS) is intrinsically scalable and amenable to standard industrial batch synthesis techniques. However, most reported LPOS strategies rely upon at least one precipitation per chain extension cycle to separate the growing oligonucleotide from reaction debris. Precipitation can be difficult to develop and control on an industrial scale and, because many precipitations would be required to prepare a therapeutic oligonucleotide, we contend that this approach is not viable for large-scale industrial preparation. We are developing an LPOS synthetic strategy for 2'-methyl RNA phosphorothioate that is more amenable to standard batch production techniques, using organic solvent nanofiltration (OSN) as the critical scalable separation technology. We report the first LPOS-OSN preparation of a 2'-Me RNA phosphorothioate 9-mer, using commercial phosphoramidite monomers, and monitoring all reactions by HPLC, (31)P NMR spectroscopy and MS.

Head injury survivors can develop neurodegeneration associated with persistent neuroinflammation, but whether the latter is harmful or beneficial is unclear. Scott et al. report that minocycline reduces neuroinflammation months and years after injury but increases a blood marker of neurodegeneration, suggesting that persistent neuroinflammation has reparative effects long after injury., Survivors of a traumatic brain injury can deteriorate years later, developing brain atrophy and dementia. Traumatic brain injury triggers chronic microglial activation, but it is unclear whether this is harmful or beneficial. A successful chronic-phase treatment for traumatic brain injury might be to target microglia. In experimental models, the antibiotic minocycline inhibits microglial activation. We investigated the effect of minocycline on microglial activation and neurodegeneration using PET, MRI, and measurement of the axonal protein neurofilament light in plasma. Microglial activation was assessed using 11C-PBR28 PET. The relationships of microglial activation to measures of brain injury, and the effects of minocycline on disease progression, were assessed using structural and diffusion MRI, plasma neurofilament light, and cognitive assessment. Fifteen patients at least 6 months after a moderate-to-severe traumatic brain injury received either minocycline 100 mg orally twice daily or no drug, for 12 weeks. At baseline, 11C-PBR28 binding in patients was increased compared to controls in cerebral white matter and thalamus, and plasma neurofilament light levels were elevated. MRI measures of white matter damage were highest in areas of greater 11C-PBR28 binding. Minocycline reduced 11C-PBR28 binding (mean Δwhite matter binding = −23.30%, 95% confidence interval −40.9 to −5.64%, P = 0.018), but increased plasma neurofilament light levels. Faster rates of brain atrophy were found in patients with higher baseline neurofilament light levels. In this experimental medicine study, minocycline after traumatic brain injury reduced chronic microglial activation while increasing a marker of neurodegeneration. These findings suggest that microglial activation has a reparative effect in the chronic phase of traumatic brain injury.

MOTIVATION: Left ventricular (LV) hypertrophy is a strong predictor of cardiovascular outcomes, but its genetic regulation remains largely unexplained. Conventional phenotyping relies on manual calculation of LV mass and wall thickness, but advanced cardiac image analysis presents an opportunity for high-throughput mapping of genotype-phenotype associations in three dimensions (3D). RESULTS: High-resolution cardiac magnetic resonance images were automatically segmented in 1,124 healthy volunteers to create a 3D shape model of the heart. Mass univariate regression was used to plot a 3D effect-size map for the association between wall thickness and a set of predictors at each vertex in the mesh. The vertices where a significant effect exists were determined by applying threshold-free cluster enhancement to boost areas of signal with spatial contiguity. Experiments on simulated phenotypic signals and SNP replication show that this approach offers a substantial gain in statistical power for cardiac genotype-phenotype associations while providing good control of the false discovery rate. This framework models the effects of genetic variation throughout the heart and can be automatically applied to large population cohorts. AVAILABILITY: The proposed approach has been coded in an R package freely available at https://doi.org/10.5281/zenodo.834610 together with the clinical data used in this work., 14 pages, 11 figures. Version accepted by Bioinformatics (Sept 2017). Includes Supplementary Materials

Rhinoviruses (RVs) are the pathogens most often responsible for the common cold, and are a frequent cause of exacerbations in asthma, chronic obstructive pulmonary disease and cystic fibrosis. Here we report the discovery of IMP-1088, a picomolar dual inhibitor of the human N-myristoyltransferases NMT1 and NMT2, and use it to demonstrate that pharmacological inhibition of host-cell N-myristoylation rapidly and completely prevents rhinoviral replication without inducing cytotoxicity. The identification of cooperative binding between weak-binding fragments led to rapid inhibitor optimization through fragment reconstruction, structure-guided fragment linking and conformational control over linker geometry. We show that inhibition of the co-translational myristoylation of a specific virus-encoded protein (VP0) by IMP-1088 potently blocks a key step in viral capsid assembly, to deliver a low nanomolar antiviral activity against multiple RV strains, poliovirus and foot and-mouth disease virus, and protection of cells against virus-induced killing, highlighting the potential of host myristoylation as a drug target in picornaviral infections.