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3,700 | Acute Phase Proteins in Marine Mammals: State of Art, Perspectives and Challenges | The term “acute phase response” (APR) is referred to a nonspecific and complex reaction of an organism that occurs shortly after any tissue damage, such as infection, trauma, neoplasia, inflammation, and stress. The APR can be identified and monitored with some laboratory tests, such as the concentration of several plasma proteins, the acute phase proteins (APPs). The APPs are components of the non-specific innate immune response, and their plasma concentration is proportional to the severity and/or the extent of tissue damage. The evaluation of health status of marine mammals is difficult because the classical clinical signs of illness used for human and domestic animals are difficult to recognize and understand. For this reason, in the past years, several efforts were done to identify laboratory markers of disease in these animals. The APPs have demonstrated their role as early markers of inflammation in veterinary medicine, thus several APPs were tested in marine mammals, such as C-reactive protein (CRP), serum amyloid-A (SAA), and Haptoglobin (Hp). However, the difficulty to extrapolate the knowledge about APPs in one species to another, the lack of specie-specific reagents, the absence of data about negative APPs have hampered their extent use in marine mammals. Herein, the state of art of APPs in marine mammals is reviewed, with particular attention to pre-analytical and analytical factors that should be taken into account in validation and interpretation of APPs assays. Moreover, the current application, potential utility and the future developments of APPs in marine mammals is highlighted and discussed. |
3,701 | The role of interleukin-1 in general pathology | Interleukin-1, an inflammatory cytokine, is considered to have diverse physiological functions and pathological significances and play an important role in health and disease. In this decade, interleukin-1 family members have been expanding and evidence is accumulating that highlights the importance of interleukin-1 in linking innate immunity with a broad spectrum of diseases beyond inflammatory diseases. In this review, we look back on the definition of “inflammation” in traditional general pathology and discuss new insights into interleukin-1 in view of its history and the molecular bases of diseases, as well as current progress in therapeutics. |
3,702 | Use of Big Data and Machine Learning Methods in the Monitoring and Evaluation of Digital Health Programs in India: An Exploratory Protocol | BACKGROUND: Digital health programs, which encompass the subsectors of health information technology, mobile health, electronic health, telehealth, and telemedicine, have the potential to generate “big data.” OBJECTIVE: Our aim is to evaluate two digital health programs in India—the maternal mobile messaging service (Kilkari) and the mobile training resource for frontline health workers (Mobile Academy). We illustrate possible applications of machine learning for public health practitioners that can be applied to generate evidence on program effectiveness and improve implementation. Kilkari is an outbound service that delivers weekly gestational age–appropriate audio messages about pregnancy, childbirth, and childcare directly to families on their mobile phones, starting from the second trimester of pregnancy until the child is one year old. Mobile Academy is an Interactive Voice Response audio training course for accredited social health activists (ASHAs) in India. METHODS: Study participants include pregnant and postpartum women (Kilkari) as well as frontline health workers (Mobile Academy) across 13 states in India. Data elements are drawn from system-generated databases used in the routine implementation of programs to provide users with health information. We explain the structure and elements of the extracted data and the proposed process for their linkage. We then outline the various steps to be undertaken to evaluate and select final algorithms for identifying gaps in data quality, poor user performance, predictors for call receipt, user listening levels, and linkages between early listening and continued engagement. RESULTS: The project has obtained the necessary approvals for the use of data in accordance with global standards for handling personal data. The results are expected to be published in August/September 2019. CONCLUSIONS: Rigorous evaluations of digital health programs are limited, and few have included applications of machine learning. By describing the steps to be undertaken in the application of machine learning approaches to the analysis of routine system-generated data, we aim to demystify the use of machine learning not only in evaluating digital health education programs but in improving their performance. Where articles on analysis offer an explanation of the final model selected, here we aim to emphasize the process, thereby illustrating to program implementors and evaluators with limited exposure to machine learning its relevance and potential use within the context of broader program implementation and evaluation. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/11456 |
3,703 | iProt-Sub: a comprehensive package for accurately mapping and predicting protease-specific substrates and cleavage sites | Regulation of proteolysis plays a critical role in a myriad of important cellular processes. The key to better understanding the mechanisms that control this process is to identify the specific substrates that each protease targets. To address this, we have developed iProt-Sub, a powerful bioinformatics tool for the accurate prediction of protease-specific substrates and their cleavage sites. Importantly, iProt-Sub represents a significantly advanced version of its successful predecessor, PROSPER. It provides optimized cleavage site prediction models with better prediction performance and coverage for more species-specific proteases (4 major protease families and 38 different proteases). iProt-Sub integrates heterogeneous sequence and structural features and uses a two-step feature selection procedure to further remove redundant and irrelevant features in an effort to improve the cleavage site prediction accuracy. Features used by iProt-Sub are encoded by 11 different sequence encoding schemes, including local amino acid sequence profile, secondary structure, solvent accessibility and native disorder, which will allow a more accurate representation of the protease specificity of approximately 38 proteases and training of the prediction models. Benchmarking experiments using cross-validation and independent tests showed that iProt-Sub is able to achieve a better performance than several existing generic tools. We anticipate that iProt-Sub will be a powerful tool for proteome-wide prediction of protease-specific substrates and their cleavage sites, and will facilitate hypothesis-driven functional interrogation of protease-specific substrate cleavage and proteolytic events. |
3,704 | Forecasting national and regional influenza-like illness for the USA | Health planners use forecasts of key metrics associated with influenza-like illness (ILI); near-term weekly incidence, week of season onset, week of peak, and intensity of peak. Here, we describe our participation in a weekly prospective ILI forecasting challenge for the United States for the 2016-17 season and subsequent evaluation of our performance. We implemented a metapopulation model framework with 32 model variants. Variants differed from each other in their assumptions about: the force-of-infection (FOI); use of uninformative priors; the use of discounted historical data for not-yet-observed time points; and the treatment of regions as either independent or coupled. Individual model variants were chosen subjectively as the basis for our weekly forecasts; however, a subset of coupled models were only available part way through the season. Most frequently, during the 2016-17 season, we chose; FOI variants with both school vacations and humidity terms; uninformative priors; the inclusion of discounted historical data for not-yet-observed time points; and coupled regions (when available). Our near-term weekly forecasts substantially over-estimated incidence early in the season when coupled models were not available. However, our forecast accuracy improved in absolute terms and relative to other teams once coupled solutions were available. In retrospective analysis, we found that the 2016-17 season was not typical: on average, coupled models performed better when fit without historically augmented data. Also, we tested a simple ensemble model for the 2016-17 season and found that it underperformed our subjective choice for all forecast targets. In this study, we were able to improve accuracy during a prospective forecasting exercise by coupling dynamics between regions. Although reduction of forecast subjectivity should be a long-term goal, some degree of human intervention is likely to improve forecast accuracy in the medium-term in parallel with the systematic consideration of more sophisticated ensemble approaches. |
3,705 | ADAM17-dependent signaling is required for oncogenic human papillomavirus entry platform assembly | Oncogenic human papillomaviruses (HPV) are small DNA viruses that infect keratinocytes. After HPV binding to cell surface receptors, a cascade of molecular interactions mediates the infectious cellular internalization of virus particles. Aside from the virus itself, important molecular players involved in virus entry include the tetraspanin CD151 and the epidermal growth factor receptor (EGFR). To date, it is unknown how these components are coordinated in space and time. Here, we studied plasma membrane dynamics of CD151 and EGFR and the HPV16 capsid during the early phase of infection. We find that the proteinase ADAM17 activates the extracellular signal-regulated kinases (ERK1/2) pathway by the shedding of growth factors which triggers the formation of an endocytic entry platform. Infectious endocytic entry platforms carrying virus particles consist of two-fold larger CD151 domains containing the EGFR. Our finding clearly dissects initial virus binding from ADAM17-dependent assembly of a HPV/CD151/EGFR entry platform. |
3,706 | Development of a Novel Ex-vivo 3D Model to Screen Amoebicidal Activity on Infected Tissue | Amoebiasis is a parasitic disease that causes thousands of deaths every year, its adverse effects and resistance to conventional treatments have led to the search of new treatment options, as well as the development of novel screening methods. In this work, we implemented a 3D model of intestine and liver slices from hamsters that were infected ex vivo with virulent E. histolytica trophozoites. Results show preserved histology in both uninfected tissues as well as ulcerations, destruction of the epithelial cells, and inflammatory reaction in intestine slices and formation of micro abscesses, and the presence of amoebae in the sinusoidal spaces and in the interior of central veins in liver slices. The three chemically synthetized compounds T-001, T-011, and T-016, which act as amoebicides in vitro, were active in both infected tissues, as they decreased the number of trophozoites, and provoked death by disintegration of the amoeba, similar to metronidazole. However, compound T-011 induced signs of cytotoxicity to liver slices. Our results suggest that ex vivo cultures of precision-cut intestinal and liver slices represent a reliable 3D approach to evaluate novel amoebicidal compounds, and to simultaneously detect their toxicity, while reducing the number of experimental animals commonly required by other model systems. |
3,707 | Preclinical septic shock research: why we need an animal ICU | Animal experiments are widely used in preclinical medical research with the goal of disease modeling and exploration of novel therapeutic approaches. In the context of sepsis and septic shock, the translation into clinical practice has been disappointing. Classical animal models of septic shock usually involve one-sex-one-age animal models, mostly in mice or rats, contrasting with the heterogeneous population of septic shock patients. Many other factors limit the reliability of preclinical models and may contribute to preclinical research failure in critical care, including the host specificity of several pathogens, the fact that laboratory animals are raised in pathogen-free facilities and that organ support techniques are either absent or minimal. Advanced animal models have been developed with the aim of improving the clinical translatability of experimental findings. So-called animal ICUs refer to the preclinical investigation of adult or even aged animals of either sex, using—in case of rats and mice—miniaturized equipment allowing for reproducing an ICU environment at a small animal scale and integrating chronic comorbidities to more closely reflect the clinical conditions studied. Strength and limitations of preclinical animal models designed to decipher the mechanisms involved in septic cardiomyopathy are discussed. This article reviews the current status and the challenges of setting up an animal ICU. |
3,708 | The MAP3K7-mTOR Axis Promotes the Proliferation and Malignancy of Hepatocellular Carcinoma Cells | Targeted therapy is currently limited for patients with hepatocellular carcinoma (HCC) due to the lack of suitable targets. Kinases play pivotal roles in many cellular biological processes, whereas dysregulation of kinases may lead to various diseases, particularly cancer. However, the role of kinases in HCC malignancy remains unclear. In this study, we employed a kinome small interfering RNA (siRNA) library, comprising 710 kinase-related genes, to screen whether any kinases were essential for cell proliferation in various HCC cell lines. Through a kinome siRNA library screening, we found that MAP3K7 was a crucial gene for HCC cell proliferation. Pharmacological or genetic ablation of MAP3K7 diminished the growth, migration, and invasion of HCC cells, including primary HCC cells. Stable knockdown of MAP3K7 attenuated tumor formation in a spheroid cell culture model and tumor xenograft mouse model. In addition, silencing MAP3K7 reduced the phosphorylation and expression of mammalian target of rapamycin (mTOR) in HCC cells. MAP3K7 expression was positively correlated with mTOR expression in tumors of patients with HCC. Higher co-expression of MAP3K7 and mTOR was significantly associated with poor prognosis of HCC. Taken together, our results revealed that the MAP3K7-mTOR axis might promote tumorigenesis and malignancy, which provides a potential marker or therapeutic target for HCC patients. |
3,709 | Immune checkpoint blockade and CAR-T cell therapy in hematologic malignancies | Harnessing the power of the immune system to recognize and eliminate cancer cells is a longtime exploration. In the past decade, monoclonal antibody (mAb)-based immune checkpoint blockade (ICB) and chimeric antigen receptor T (CAR-T) cell therapy have proven to be safe and effective in hematologic malignancies. Despite the unprecedented success of ICB and CAR-T therapy, only a subset of patients can benefit partially due to immune dysfunction and lack of appropriate targets. Here, we review the preclinical and clinical advances of CTLA-4 and PD-L1/PD-1-based ICB and CD19-specific CAR-T cell therapy in hematologic malignancies. We also discuss the basic research and ongoing clinical trials on emerging immune checkpoints (Galectin-9/Tim-3, CD70/CD27, LAG-3, and LILRBs) and on new targets for CAR-T cell therapy (CD22, CD33, CD123, BCMA, CD38, and CD138) for the treatment of hematologic malignancies. |
3,710 | Host–Microbe Interactions and Gut Health in Poultry—Focus on Innate Responses | Commercial poultry are continually exposed to, frequently pathogenic, microorganisms, usually via mucosal surfaces such as the intestinal mucosa. Thus, understanding host–microbe interactions is vital. Many of these microorganisms may have no or limited contact with the host, while most of those interacting more meaningfully with the host will be dealt with by the innate immune response. Fundamentally, poultry have evolved to have immune responses that are generally appropriate and adequate for their acquired microbiomes, although this is challenged by commercial production practices. Innate immune cells and their functions, encompassing inflammatory responses, create the context for neutralising the stimulus and initiating resolution. Dysregulated inflammatory responses can be detrimental but, being a highly conserved biological process, inflammation is critical for host defence. Heterogeneity and functional plasticity of innate immune cells is underappreciated and offers the potential for (gut) health interventions, perhaps including exogenous opportunities to influence immune cell metabolism and thus function. New approaches could focus on identifying and enhancing decisive but less harmful immune processes, improving the efficiency of innate immune cells (e.g., targeted, efficient microbial killing) and promoting phenotypes that drive resolution of inflammation. Breeding strategies and suitable exogenous interventions offer potential solutions to enhance poultry gut health, performance and welfare. |
3,711 | High sensitivity methods to quantify chloroquine and its metabolite in human blood samples using LC–MS/MS | AIM: Chloroquine is an antimalarial drug used in the treatment of Plasmodium vivax malaria. Three methods to quantify chloroquine and its metabolite in blood matrices were developed and validated. METHODOLOGY & RESULTS: Different high-throughput extraction techniques were used to recover the drugs from whole blood (50 μl), plasma (100 μl) and dried blood spots (15 μl as punched discs) followed by quantification with LC–MS/MS. The intra- and inter-batch precisions were below 15%, and thus meet regulatory acceptance criteria. CONCLUSION: The developed methods demonstrated satisfactory validation performance with high sensitivity and selectivity. The assays used simple and easy to automate extraction techniques. All methods were reliable with robust performance and demonstrated to be suitable to implement into high-throughput routine analysis of clinical pharmacokinetic samples. |
3,712 | Efficient Production of Human Norovirus-Specific IgY in Egg Yolks by Vaccination of Hens with a Recombinant Vesicular Stomatitis Virus Expressing VP1 Protein | Human norovirus (HuNoV) is responsible for more than 95% of outbreaks of acute nonbacterial gastroenteritis worldwide. Despite major efforts, there are no vaccines or effective therapeutic interventions against this virus. Chicken immunoglobulin Y (IgY)-based passive immunization has been shown to be an effective strategy to prevent and treat many enteric viral diseases. Here, we developed a highly efficient bioreactor to generate high titers of HuNoV-specific IgY in chicken yolks using a recombinant vesicular stomatitis virus expressing HuNoV capsid protein (rVSV-VP1) as an antigen. We first demonstrated that HuNoV VP1 protein was highly expressed in chicken cells infected by rVSV-VP1. Subsequently, we found that White Leghorn hens immunized intramuscularly with rVSV-VP1 triggered a high level of HuNoV-specific yolk IgY antibodies. The purified yolk IgY was efficiently recognized by HuNoV virus-like particles (VLPs). Importantly, HuNoV-specific IgY efficiently blocked the binding of HuNoV VLPs to all three types (A, B, and O) of histo-blood group antigens (HBGAs), the attachment factors for HuNoV. In addition, the receptor blocking activity of IgY remained stable at temperature below 70 °C and at pH ranging from 4 to 9. Thus, immunization of hens with VSV-VP1 could be a cost-effective and practical strategy for large-scale production of anti-HuNoV IgY antibodies for potential use as prophylactic and therapeutic treatment against HuNoV infection. |
3,713 | Extracellular Vesicles and Ebola Virus: A New Mechanism of Immune Evasion | Ebola virus (EBOV) disease can result in a range of symptoms anywhere from virtually asymptomatic to severe hemorrhagic fever during acute infection. Additionally, spans of asymptomatic persistence in recovering survivors is possible, during which transmission of the virus may occur. In acute infection, substantial cytokine storm and bystander lymphocyte apoptosis take place, resulting in uncontrolled, systemic inflammation in affected individuals. Recently, studies have demonstrated the presence of EBOV proteins VP40, glycoprotein (GP), and nucleoprotein (NP) packaged into extracellular vesicles (EVs) during infection. EVs containing EBOV proteins have been shown to induce apoptosis in recipient immune cells, as well as contain pro-inflammatory cytokines. In this manuscript, we review the current field of knowledge on EBOV EVs including the mechanisms of their biogenesis, their cargo and their effects in recipient cells. Furthermore, we discuss some of the effects that may be induced by EBOV EVs that have not yet been characterized and highlight the remaining questions and future directions. |
3,714 | Animals as Reservoir for Human Norovirus | Norovirus is the most common cause of non-bacterial gastroenteritis and is a burden worldwide. The increasing norovirus diversity is currently categorized into at least 10 genogroups which are further classified into more than 40 genotypes. In addition to humans, norovirus can infect a broad range of hosts including livestock, pets, and wild animals, e.g., marine mammals and bats. Little is known about norovirus infections in most non-human hosts, but the close genetic relatedness between some animal and human noroviruses coupled with lack of understanding where newly appearing human norovirus genotypes and variants are emerging from has led to the hypothesis that norovirus may not be host restricted and might be able to jump the species barrier. We have systematically reviewed the literature to describe the diversity, prevalence, and geographic distribution of noroviruses found in animals, and the pathology associated with infection. We further discuss the evidence that exists for or against interspecies transmission including surveillance data and data from in vitro and in vivo experiments. |
3,715 | Coxsackievirus B3 Responds to Polyamine Depletion via Enhancement of 2A and 3C Protease Activity | Polyamines are small positively-charged molecules abundant in eukaryotic cells that are crucial to RNA virus replication. In eukaryotic cells, polyamines facilitate processes such as transcription, translation, and DNA replication, and viruses similarly rely on polyamines to facilitate transcription and translation. Whether polyamines function at additional stages in viral replication remains poorly understood. Picornaviruses, including Coxsackievirus B3 (CVB3), are sensitive to polyamine depletion both in vitro and in vivo; however, precisely how polyamine function in picornavirus infection has not been described. Here, we describe CVB3 mutants that arise with passage in polyamine-depleted conditions. We observe mutations in the 2A and 3C proteases, and we find that these mutant proteases confer resistance to polyamine depletion. Using a split luciferase reporter system to measure protease activity, we determined that polyamines facilitate viral protease activity. We further observe that the 2A and 3C protease mutations enhance reporter protease activity in polyamine-depleted conditions. Finally, we find that these mutations promote cleavage of cellular eIF4G during infection of polyamine-depleted cells. In sum, our results suggest that polyamines are crucial to protease function during picornavirus infection. Further, these data highlight viral proteases as potential antiviral targets and highlight how CVB3 may overcome polyamine-depleting antiviral therapies. |
3,716 | Influenza virus-related critical illness: prevention, diagnosis, treatment | Annual seasonal influenza epidemics of variable severity result in significant morbidity and mortality in the United States (U.S.) and worldwide. In temperate climate countries, including the U.S., influenza activity peaks during the winter months. Annual influenza vaccination is recommended for all persons in the U.S. aged 6 months and older, and among those at increased risk for influenza-related complications in other parts of the world (e.g. young children, elderly). Observational studies have reported effectiveness of influenza vaccination to reduce the risks of severe disease requiring hospitalization, intensive care unit admission, and death. A diagnosis of influenza should be considered in critically ill patients admitted with complications such as exacerbation of underlying chronic comorbidities, community-acquired pneumonia, and respiratory failure during influenza season. Molecular tests are recommended for influenza testing of respiratory specimens in hospitalized patients. Antigen detection assays are not recommended in critically ill patients because of lower sensitivity; negative results of these tests should not be used to make clinical decisions, and respiratory specimens should be tested for influenza by molecular assays. Because critically ill patients with lower respiratory tract disease may have cleared influenza virus in the upper respiratory tract, but have prolonged influenza viral replication in the lower respiratory tract, an endotracheal aspirate (preferentially) or bronchoalveolar lavage fluid specimen (if collected for other diagnostic purposes) should be tested by molecular assay for detection of influenza viruses. Observational studies have reported that antiviral treatment of critically ill adult influenza patients with a neuraminidase inhibitor is associated with survival benefit. Since earlier initiation of antiviral treatment is associated with the greatest clinical benefit, standard-dose oseltamivir (75 mg twice daily in adults) for enteric administration is recommended as soon as possible as it is well absorbed in critically ill patients. Based upon observational data that suggest harms, adjunctive corticosteroid treatment is currently not recommended for children or adults hospitalized with influenza, including critically ill patients, unless clinically indicated for another reason, such as treatment of asthma or COPD exacerbation, or septic shock. A number of pharmaceutical agents are in development for treatment of severe influenza. |
3,717 | A Highly Active Bidentate Magnesium Catalyst for Amine‐Borane Dehydrocoupling: Kinetic and Mechanistic Studies | A magnesium complex (1) featuring a bidentate aminopyridinato ligand is a remarkably selective catalyst for the dehydrocoupling of amine‐boranes. This reaction proceeds to completion with low catalyst loadings (1 mol %) under mild conditions (60 °C), exceeding previously reported s‐block systems in terms of selectivity, rate, and turnover number (TON). Mechanistic studies by in situ NMR analysis reveals the reaction to be first order in both catalyst and substrate. A reaction mechanism is proposed to account for these findings, with the high TON of the catalyst attributed to the bidentate nature of the ligand, which allows for reversible deprotonation of the substrate and regeneration of 1 as a stable resting state. |
3,718 | Applications of Nanodiamonds in the Detection and Therapy of Infectious Diseases | We are constantly exposed to infectious diseases, and they cause millions of deaths per year. The World Health Organization (WHO) estimates that antibiotic resistance could cause 10 million deaths per year by 2050. Multidrug-resistant bacteria are the cause of infection in at least one in three people suffering from septicemia. While antibiotics are powerful agents against infectious diseases, the alarming increase in antibiotic resistance is of great concern. Alternatives are desperately needed, and nanotechnology provides a great opportunity to develop novel approaches for the treatment of infectious diseases. One of the most important factors in the prognosis of an infection caused by an antibiotic resistant bacteria is an early and rigorous diagnosis, jointly with the use of novel therapeutic systems that can specifically target the pathogen and limit the selection of resistant strains. Nanodiamonds can be used as antimicrobial agents due to some of their properties including size, shape, and biocompatibility, which make them highly suitable for the development of efficient and tailored nanotherapies, including vaccines or drug delivery systems. In this review, we discuss the beneficial findings made in the nanodiamonds field, focusing on diagnosis and treatment of infectious diseases. We also highlight the innovative platform that nanodiamonds confer for vaccine improvement, drug delivery, and shuttle systems, as well as their role in the generation of faster and more sensitive clinical diagnosis. |
3,719 | Improved phylogenomic sampling of free-living nematodes enhances resolution of higher-level nematode phylogeny | BACKGROUND: Nematodes are among the most diverse and abundant metazoans on Earth, but research on them has been biased toward parasitic taxa and model organisms. Free-living nematodes, particularly from the clades Enoplia and Dorylaimia, have been underrepresented in genome-scale phylogenetic analyses to date, leading to poor resolution of deep relationships within the phylum. RESULTS: We supplemented publicly available data by sequencing transcriptomes of nine free-living nematodes and two important outgroups and conducted a phylum-wide phylogenomic analysis including a total of 108 nematodes. Analysis of a dataset generated using a conservative orthology inference strategy resulted in a matrix with a high proportion of missing data and moderate to weak support for branching within and placement of Enoplia. A less conservative orthology inference approach recovered more genes and resulted in higher support for the deepest splits within Nematoda, recovering Enoplia as the sister taxon to the rest of Nematoda. Relationships within major clades were similar to those found in previously published studies based on 18S rDNA. CONCLUSIONS: Expanded transcriptome sequencing of free-living nematodes has contributed to better resolution among deep nematode lineages, though the dataset is still strongly biased toward parasites. Inclusion of more free-living nematodes in future phylogenomic analyses will allow a clearer understanding of many interesting aspects of nematode evolution, such as morphological and molecular adaptations to parasitism and whether nematodes originated in a marine or terrestrial environment. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12862-019-1444-x) contains supplementary material, which is available to authorized users. |
3,720 | Protein quality control in the nucleolus safeguards recovery of epigenetic regulators after heat shock | Maintenance of epigenetic modifiers is of utmost importance to preserve the epigenome and consequently appropriate cellular functioning. Here, we analyzed Polycomb group protein (PcG) complex integrity in response to heat shock (HS). Upon HS, various Polycomb Repressive Complex (PRC)1 and PRC2 subunits, including CBX proteins, but also other chromatin regulators, are found to accumulate in the nucleolus. In parallel, binding of PRC1/2 to target genes is strongly reduced, coinciding with a dramatic loss of H2AK119ub and H3K27me3 marks. Nucleolar-accumulated CBX proteins are immobile, but remarkably both CBX protein accumulation and loss of PRC1/2 epigenetic marks are reversible. This post-heat shock recovery of pan-nuclear CBX protein localization and reinstallation of epigenetic marks is HSP70 dependent. Our findings demonstrate that the nucleolus is an essential protein quality control center, which is indispensable for recovery of epigenetic regulators and maintenance of the epigenome after heat shock. |
3,721 | Ambulatory and stationary healthcare use in survivors of ARDS during the first year after discharge from ICU: findings from the DACAPO cohort | BACKGROUND: For many survivors of acute respiratory distress syndrome (ARDS), the process from discharge from intensive care unit (ICU) to recovery is long and difficult. However, healthcare use after discharge from ICU has received only little attention by research. This study sets out to investigate the extent of ambulatory and stationary healthcare use among survivors of ARDS in Germany (multicenter DACAPO cohort) and to analyze predictors of stationary healthcare use. RESULTS: A total of 396 survivors of ARDS provided data at 1 year after discharge from ICU. Fifty percent of 1-year survivors were hospitalized for 48 days or longer after discharge from ICU, with 10% spending more than six out of 12 months in stationary care. The duration of hospitalization increased significantly by the length of the initial ICU stay. All participants reported at least one outpatient visit (including visits to general practitioners), and 50% contacted four or more different medical specialties within the first year after discharge from ICU. CONCLUSIONS: For most of the patients, the first year after ARDS is characterized by an extensive amount of healthcare utilization, especially with regard to stationary health care. These findings shed light on the substantial morbidity of patients after ARDS and contribute to a better understanding of the situation of patients following discharge from ICU. |
3,722 | Novel insights into endogenous RNA viral elements in Ixodes scapularis and other arbovirus vector genomes | Many emerging arboviruses are not transmitted by traditional mosquito vectors, but by lesser-studied arthropods such as ticks, midges, and sand flies. Small RNA (sRNA) silencing pathways are the main antiviral defence mechanism for arthropods, which lack adaptive immunity. Non-retroviral integrated RNA virus sequences (NIRVS) are one potential source of sRNAs which comprise these pathways. NIRVS are remnants of past germline RNA viral infections, where viral cDNA integrates into the host genome and is vertically transmitted. In Aedes mosquitoes, NIRVS are widespread and produce PIWI-interacting RNAs (piRNAs). These are hypothesised to target incoming viral transcripts to modulate viral titre, perhaps rendering the organism a more efficient arbovirus vector. To explore the NIRVS landscape in alternative arbovirus vectors, we validated the NIRVS landscape in Aedes spp. and then identified novel NIRVS in six medically relevant arthropods and also in Drosophila melanogaster. We identified novel NIRVS in Phlebotomus papatasi, Culicoides sonorensis, Rhipicephalus microplus, Anopheles gambiae, Culex quinquefasciatus, and Ixodes scapularis. Due to their unexpected abundance, we further characterised NIRVS in the blacklegged tick I. scapularis (n = 143). Interestingly, NIRVS are not enriched in R. microplus, another hard tick, suggesting this is an Ixodes-specific adaptation. I. scapularis NIRVS are enriched in bunya- and orthomyxo-like sequences, reflecting that ticks are a dominant host for these virus groups. Unlike in mosquitoes, I. scapularis NIRVS are more commonly derived from the non-structural region (replicase) of negative-sense viruses, as opposed to structural regions (e.g. glycoprotein). Like other arthropods, I. scapularis NIRVS preferentially integrate into genomic piRNA clusters, and serve as a template for primary piRNA production in the commonly used embryonic I. scapularis ISE6 cell line. Interestingly, we identified a two-fold enrichment of non-long terminal repeat (non-LTR) retrotransposons, in genomic proximity to NIRVS, contrasting with studeis in Ae. aegypti, where LTR retrotransposons are instead associated with NIRVS formation. We characterised NIRVS phylogeny and integration patterns in the important vector, I. scapularis, revealing they are distinct from those in Aedes spp. Future studies will explore the possible antiviral mechanism conferred by NIRVS to I. scapularis,which may help the transmission of pathogenic arboviruses. Finally, this study explored NIRVS as an untapped wealth of viral diversity in arthropods. |
3,723 | A shared protocol for porcine surfactant use in pediatric acute respiratory distress syndrome: a feasibility study | BACKGROUND: Pediatric ARDS still represents a difficult challenge in Pediatric Intensive Care Units (PICU). Among different treatments proposed, exogenous surfactant showed conflicting results. Aim of this multicenter retrospective observational study was to evaluate whether poractant alfa use in pediatric ARDS might improve gas exchange in children less than 2 years old, according to a shared protocol. METHODS: The study was carried out in fourteen Italian PICUs after dissemination of a standardized protocol for surfactant administration within the Italian PICU network. The protocol provides the administration of surfactant (50 mg/kg) divided in two doses: the first dose is used as a bronchoalveolar lavage while the second as supplementation. Blood gas exchange variations before and after surfactant use were recorded. RESULTS: Sixty-nine children, age 0–24 months, affected by Acute Respiratory Distress Syndrome treated with exogenous porcine surfactant were enrolled. Data collection consisted of patient demographics, respiratory variables and arterial blood gas analysis. The most frequent reasons for PICU admission were acute respiratory failure, mainly bronchiolitis and pneumonia, and septic shock. Fifty-four children (78.3%) had severe ARDS (define by oxygen arterial pressure and inspired oxygen fraction ratio (P/F) < 100), 15 (21.7%) had moderate ARDS (100 < P/F < 200). PO(2), P/F, Oxygenation Index (OI) and pH showed a significant improvement after surfactant use with respect to baseline (p < 0.001 at each included time-point for each parameter). No significant difference in blood gas variations were observed among four different subgroups of diseases (bronchiolitis, pneumonia, septic shock and others). Overall, 11 children died (15.9%) and among these, 10 (90.9%) had complex chronic conditions. Two children (18.2%) died while being treated with Extracorporeal Membrane Oxygenation (ECMO). Mortality for severe pARDS was 20.4%. CONCLUSION: The use of porcine Surfactant improves oxygenation, P/F ratio, OI and pH in a population of children with moderate or severe pARDS caused by multiple diseases. A shared protocol seems to be a good option to obtain the same criteria of enrollment among different PICUs and define a unique way of use and administration of the drug for future studies. |
3,724 | Development of a Rhinovirus Inoculum Using a Reverse Genetics Approach | BACKGROUND: Experimental inoculation is an important tool for common cold and asthma research. Producing rhinovirus (RV) inocula from nasal secretions has required prolonged observation of the virus donor to exclude extraneous pathogens. We produced a RV-A16 inoculum using reverse genetics and determined the dose necessary to cause moderate colds in seronegative volunteers. METHODS: The consensus sequence of RV-A16 from a previous inoculum was cloned, and inoculum virus was produced using reverse genetics techniques. After safety testing, volunteers were inoculated with either RV-A16 (n = 26) or placebo (n = 10), Jackson cold scores were recorded, and nasal secretions were tested for shedding of RV-A16 ribonucleic acid. RESULTS: The reverse genetics process produced infectious virus that was neutralized by specific antisera and had a mutation rate similar to conventional virus growth techniques. The 1000 median tissue culture infectious dose (TCID(50)) dose produced moderate colds in most individuals with effects similar to that of a previously tested conventional RV-A16 inoculum. CONCLUSIONS: Reverse genetics techniques produced a RV-A16 inoculum that can cause clinical colds in seronegative volunteers, and they also serve as a stable source of virus for laboratory use. The recombinant production procedures eliminate the need to derive seed virus from nasal secretions, thus precluding introduction of extraneous pathogens through this route. |
3,725 | Reversible ADP-ribosylation of RNA | ADP-ribosylation is a reversible chemical modification catalysed by ADP-ribosyltransferases such as PARPs that utilize nicotinamide adenine dinucleotide (NAD(+)) as a cofactor to transfer monomer or polymers of ADP-ribose nucleotide onto macromolecular targets such as proteins and DNA. ADP-ribosylation plays an important role in several biological processes such as DNA repair, transcription, chromatin remodelling, host-virus interactions, cellular stress response and many more. Using biochemical methods we identify RNA as a novel target of reversible mono-ADP-ribosylation. We demonstrate that the human PARPs - PARP10, PARP11 and PARP15 as well as a highly diverged PARP homologue TRPT1, ADP-ribosylate phosphorylated ends of RNA. We further reveal that ADP-ribosylation of RNA mediated by PARP10 and TRPT1 can be efficiently reversed by several cellular ADP-ribosylhydrolases (PARG, TARG1, MACROD1, MACROD2 and ARH3), as well as by MACROD-like hydrolases from VEEV and SARS viruses. Finally, we show that TRPT1 and MACROD homologues in bacteria possess activities equivalent to the human proteins. Our data suggest that RNA ADP-ribosylation may represent a widespread and physiologically relevant form of reversible ADP-ribosylation signalling. |
3,726 | Junín virus induces autophagy in human A549 cells | Autophagy, a highly regulated degradative process that promotes cellular homeostasis, is increasingly recognised as a fundamental component of the cellular response against viral infection. In this study, we investigated the role of autophagy during Junín virus (JUNV) multiplication using human A549 cells. We found that JUNV infection induces an increment of the LC3-II/LC3-I ratio, an accumulation of punctate pattern in RFP-LC3-transfected cells and the colocalisation of viral nucleoprotein and LC3 protein, suggesting autophagosome formation. JUNV infection also induced the degradation of the autophagy receptor p62, suggesting that complete autophagic flux was triggered. In addition, we showed that inhibition of autophagy with bafilomycin A1 or 3-methyladenine significantly reduces viral multiplication. Moreover, viral yield was increased when autophagy was induced using rapamycin. Furthermore, JUNV infection induced the colocalisation of p62, ATG16, RAB5, RAB7A and LAMP1 with the autophagosomal LC3 protein. That suggests that phagosomes undergo the maturation process during viral infection. Finally, we demonstrated that siRNA experiments targeting essential autophagy genes (ATG5, ATG7 and Beclin 1) reduce viral protein synthesis and viral yield. Overall, our results indicate that JUNV activates host autophagy machinery enhancing its multiplication. |
3,727 | High antistreptolysin O titer is associated with coronary artery lesions in patients with Kawasaki disease | PURPOSE: In Kawasaki disease (KD) patients, coronary artery complications, incomplete and refractory types occur more frequently in patients with streptococcal or other bacterial/viral infections. Recently, we observed a higher incidence of coronary lesions in KD patients with high anti-streptolysin O (ASO) titer. Therefore, we hypothesized that KD patients diagnosed with concurrent streptococcal infection have poor prognoses, with respect to treatment response and development of coronary artery lesions. METHODS: A retrospective review was performed in 723 patients with KD who were admitted to 2 major hospitals between June 2010 and September 2017. RESULTS: Among 723 patients with KD, 11 initially showed an elevated ASO titer (>320 IU/mL) or elevated follow-up ASO titer after treatment. Of these patients, 5 showed no response to the first intravenous immunoglobulin treatment, 3 had abnormalities of the coronary arteries. This is a significantly higher proportion of patients with a high ASO titer (n=3, 27.3%) than those with a normal ASO titer (n=53 [7.4%], P=0.047). A severe clinical course was seen in 81.8% of patients in the high ASO group versus 14.5% of patients in the normal ASO group. CONCLUSION: It is not certain whether acute streptococcal infection may cause KD, but this study revealed that KD with high ASO titers showed higher rates of severe clinical course. It may be helpful to analyze concurrent streptococcal infection in patients with a severe clinical course. |
3,728 | Microbial Proteases Applications | The use of chemicals around the globe in different industries has increased tremendously, affecting the health of people. The modern world intends to replace these noxious chemicals with environmental friendly products for the betterment of life on the planet. Establishing enzymatic processes in spite of chemical processes has been a prime objective of scientists. Various enzymes, specifically microbial proteases, are the most essentially used in different corporate sectors, such as textile, detergent, leather, feed, waste, and others. Proteases with respect to physiological and commercial roles hold a pivotal position. As they are performing synthetic and degradative functions, proteases are found ubiquitously, such as in plants, animals, and microbes. Among different producers of proteases, Bacillus sp. are mostly commercially exploited microbes for proteases. Proteases are successfully considered as an alternative to chemicals and an eco-friendly indicator for nature or the surroundings. The evolutionary relationship among acidic, neutral, and alkaline proteases has been analyzed based on their protein sequences, but there remains a lack of information that regulates the diversity in their specificity. Researchers are looking for microbial proteases as they can tolerate harsh conditions, ways to prevent autoproteolytic activity, stability in optimum pH, and substrate specificity. The current review focuses on the comparison among different proteases and the current problems faced during production and application at the industrial level. Deciphering these issues would enable us to promote microbial proteases economically and commercially around the world. |
3,729 | Selective use of primate CD4 receptors by HIV-1 | Individuals chronically infected with HIV-1 harbor complex viral populations within their bloodstreams. Recently, it has come to light that when these people infect others, the new infection is typically established by only one or a small number of virions from within this complex viral swarm. An important goal is to characterize the biological properties of HIV-1 virions that seed and exist early in new human infections because these are potentially the only viruses against which a prophylactic HIV-1 vaccine would need to elicit protection. This includes understanding how the Envelope (Env) protein of these virions interacts with the T-cell receptor CD4, which supports attachment and entry of HIV-1 into target cells. We examined early HIV-1 isolates for their ability to infect cells via the CD4 receptor of 15 different primate species. Primates were the original source of HIV-1 and now serve as valuable animal models for studying HIV-1. We find that most primary isolates of HIV-1 from the blood, including early isolates, are highly selective and enter cells through some primate CD4 receptor orthologs but not others. This phenotype is remarkably consistent, regardless of route of transmission, viral subtype, or time of isolation post infection. We show that the weak CD4 binding affinity of blood-derived HIV-1 isolates is what makes them sensitive to the small sequence differences in CD4 from one primate species to the next. To substantiate this, we engineered an early HIV-1 Env to have high, medium, or low binding affinity to CD4, and we show that it loses the ability to enter cells via the CD4 receptor of many primate species as the binding affinity gets weaker. Based on the phenotype of selective use of primate CD4, we find that weak CD4 binding appears to be a nearly universal property of HIV-1 circulating in the bloodstream. Therefore, weak binding to CD4 must be a selected and important property in the biology of HIV-1 in the body. We identify six primate species that encode CD4 receptors that fully support the entry of early HIV-1 isolates despite their low binding affinity for CD4. These findings will help inform long-standing efforts to model HIV-1 transmission and early disease in primates. |
3,730 | The Group B Streptococcal surface antigen I/II protein, BspC, interacts with host vimentin to promote adherence to brain endothelium and inflammation during the pathogenesis of meningitis | Streptococcus agalactiae (Group B Streptococcus, GBS) normally colonizes healthy adults but can cause invasive disease, such as meningitis, in the newborn. To gain access to the central nervous system, GBS must interact with and penetrate brain or meningeal blood vessels; however, the exact mechanisms are still being elucidated. Here, we investigate the contribution of BspC, an antigen I/II family adhesin, to the pathogenesis of GBS meningitis. Disruption of the bspC gene reduced GBS adherence to human cerebral microvascular endothelial cells (hCMEC), while heterologous expression of BspC in non-adherent Lactococcus lactis conferred bacterial attachment. In a murine model of hematogenous meningitis, mice infected with ΔbspC mutants exhibited lower mortality as well as decreased brain bacterial counts and inflammatory infiltrate compared to mice infected with WT GBS strains. Further, BspC was both necessary and sufficient to induce neutrophil chemokine expression. We determined that BspC interacts with the host cytoskeleton component vimentin and confirmed this interaction using a bacterial two-hybrid assay, microscale thermophoresis, immunofluorescent staining, and imaging flow cytometry. Vimentin null mice were protected from WT GBS infection and also exhibited less inflammatory cytokine production in brain tissue. These results suggest that BspC and the vimentin interaction is critical for the pathogenesis of GBS meningitis. |
3,731 | Complement Receptor 1 availability on red blood cell surface modulates Plasmodium vivax invasion of human reticulocytes | Plasmodium vivax parasites preferentially invade reticulocyte cells in a multistep process that is still poorly understood. In this study, we used ex vivo invasion assays and population genetic analyses to investigate the involvement of complement receptor 1 (CR1) in P. vivax invasion. First, we observed that P. vivax invasion of reticulocytes was consistently reduced when CR1 surface expression was reduced through enzymatic cleavage, in the presence of naturally low-CR1-expressing cells compared with high-CR1-expressing cells, and with the addition of soluble CR1, a known inhibitor of P. falciparum invasion. Immuno-precipitation experiments with P. vivax Reticulocyte Binding Proteins showed no evidence of complex formation. In addition, analysis of CR1 genetic data for worldwide human populations with different exposure to malaria parasites show significantly higher frequency of CR1 alleles associated with low receptor expression on the surface of RBCs and higher linkage disequilibrium in human populations exposed to P. vivax malaria compared with unexposed populations. These results are consistent with a positive selection of low-CR1-expressing alleles in vivax-endemic areas. Collectively, our findings demonstrate that CR1 availability on the surface of RBCs modulates P. vivax invasion. The identification of new molecular interactions is crucial to guiding the rational development of new therapeutic interventions against vivax malaria. |
3,732 | Assessing dengue control in Tokyo, 2014 | BACKGROUND: In summer 2014, an autochthonous outbreak of dengue occurred in Tokyo, Japan, in which Yoyogi Park acted as the focal area of transmission. Recognizing the outbreak, concerted efforts were made to control viral spread, which included mosquito control, public announcement of the outbreak, and a total ban on entering the park. We sought to assess the effectiveness of these control measures. METHODOLOGY/PRINCIPAL FINDINGS: We used a mathematical model to describe the transmission dynamics. Using dates of exposure and illness onset, we categorized cases into three groups according to the availability of these datasets. The infection process was parametrically modeled by generation, and convolution of the infection process and the incubation period was fitted to the data. By estimating the effective reproduction number, we determined that the effect of dengue risk communication together with mosquito control from 28 August 2014 was insufficiently large to lower the reproduction number to below 1. However, once Yoyogi Park was closed on 4 September, the value of the effective reproduction number began to fall below 1, and the associated relative reduction in the effective reproduction number was estimated to be 20%–60%. The mean incubation period was an estimated 5.8 days. CONCLUSIONS/SIGNIFICANCE: Regardless of the assumed number of generations of cases, the combined effect of mosquito control, risk communication, and park closure appeared to be successful in interrupting the chain of dengue transmission in Tokyo. |
3,733 | GRASP depletion–mediated Golgi destruction decreases cell adhesion and migration via the reduction of α5β1 integrin | The Golgi apparatus is a membrane-bound organelle that serves as the center for trafficking and processing of proteins and lipids. To perform these functions, the Golgi forms a multilayer stacked structure held by GRASP55 and GRASP65 trans-oligomers and perhaps their binding partners. Depletion of GRASP proteins disrupts Golgi stack formation and impairs critical functions of the Golgi, such as accurate protein glycosylation and sorting. However, how Golgi destruction affects other cellular activities is so far unknown. Here, we report that depletion of GRASP proteins reduces cell attachment and migration. Interestingly, GRASP depletion reduces the protein level of α5β1 integrin, the major cell adhesion molecule at the surface of HeLa and MDA-MB-231 cells, due to decreased integrin protein synthesis. GRASP depletion also increases cell growth and total protein synthesis. These new findings enrich our understanding on the role of the Golgi in cell physiology and provide a potential target for treating protein-trafficking disorders. |
3,734 | Development and evaluation of a new real‐time RT‐PCR assay for detecting the latest H9N2 influenza viruses capable of causing human infection | The H9N2 subtype of avian influenza A viruses (AIV) has spread among domestic poultry and wild birds worldwide. H9N2 AIV is sporadically transmitted to humans from avian species. A total of 42 laboratory‐confirmed cases of non‐fatal human infection with the Eurasian Y280 and G1 lineages have been reported in China, Hong Kong, Bangladesh and Egypt since 1997. H9N2 AIV infections in poultry have become endemic in Asia and the Middle East and are a major source of viral internal genes for other AIV subtypes, such that continuous monitoring of H9N2 AIV is recommended. In this study, a new, one‐step, real‐time RT‐PCR assay was developed to detect two major Eurasian H9 lineages of AIV capable of causing human infection. The sensitivity of this assay was determined using in vitro‐transcribed RNA, and the detection limit was approximately 3 copies/reaction. In this assay, no cross‐reactivity was observed against RNA from H1–15 subtypes of influenza A viruses, influenza B viruses and other viral respiratory pathogens. In addition, this assay could detect the H9 hemagglutinin (HA) gene from artificially reconstituted clinical samples spiked with H9N2 virus without any non‐specific reactions. Therefore, this assay is highly sensitive and specific for H9 HA detection. The assay is useful both for diagnostic purposes in cases of suspected human infection with influenza H9N2 viruses and for the surveillance of both avian and human influenza viruses. |
3,735 | Protective effects of the Francisella tularensis ΔpdpC mutant against its virulent parental strain SCHU P9 in Cynomolgus macaques | Tularemia is a severe infectious zoonotic disease caused by Francisella tularensis. Although F. tularensis is considered to be a potential biological weapon due to its high infectivity and mortality rate, no vaccine has been currently licensed. Recently, we reported that F. tularensis SCHU P9 derived ΔpdpC strain lacking the pathogenicity determinant protein C gene conferred stable and good protection in a mouse lethal model. In this study, the protective effect of ΔpdpC was evaluated using a monkey lethal model. Two cynomolgus macaques (Macaca fascicularis) intratracheally challenged with the virulent strain SCHU P9 were euthanized on 7 and 11 days post-challenge after the development of severe clinical signs. The bacterial replication in alveolar macrophages and type II epithelial cells in the lungs would cause severe pneumonia accompanied by necrosis. Conversely, two animals subcutaneously immunized with ΔpdpC survived 3 weeks after SCHU P9 challenge. Though one of the two animals developed mild symptoms of tularemia, bacterial replication was limited in the respiratory organs, which may be due to a high level of humoral and cellular immune responses against F. tularensis. These results suggest that the ΔpdpC mutant would be a safe and promising candidate as a live attenuated tularemia vaccine. |
3,736 | Protocol for development of a risk assessment tool for planning and management of religious mass-gathering events of India—a health system-strengthening initiative | BACKGROUND: Religious mass gatherings (MGs) have always been an integral part of our society. At the outset, mass-gathering events provide challenging settings to plan a suitable emergency public health response. Published studies basically talk about retrospective reviews, case studies of the public health preparedness, or health care provided at individual events. Developing an understanding of the variables associated with MGs is the first step for public health managers. Risk assessment (RA) is a crucial part of pre-event planning as it helps foresee potential risks. Based on RA, one can develop preventive measures and ensure that the infrastructure to control the potential problems is in place. This study is an attempt to systemize RA process during MG events in a country that is culturally rich but with poor resources to handle such events. A RA tool will be developed for planning and management of religious MG events of India. METHODS/DESIGN: Various strategies will be used to develop the risk assessment tool (RA tool). Extensive review of literature clubbed with key informant interviews will be done in order to identify the risk variables and decide the domains and items of the tool. Further, this tool will be developed as a mobile-based application. The feasibility of the mobile-based RA tool will be tested in real-time MG event in one part of the country. Concurrently in the same event, a community survey of residents and visitors will be done in order to assess public perceptions of public health and environmental risks associated with MG events. DISCUSSION: The findings of this study will provide insights into the public health and environmental concerns that need to be considered if preventive strategies and intervention programs are to be designed for MG events. A “RA Tool,” which can be used in the planning and management of MG events by the public health managers will strengthen the existing health systems preparedness plans for MGs. |
3,737 | Broadly resistant HIV-1 against CD4-binding site neutralizing antibodies | Recently identified broadly neutralizing antibodies (bnAbs) show great potential for clinical interventions against HIV-1 infection. However, resistant strains may impose substantial challenges. Here, we report on the identification and characterization of a panel of HIV-1 strains with broad and potent resistance against a large number of bnAbs, particularly those targeting the CD4-binding site (CD4bs). Site-directed mutagenesis revealed that several key epitope mutations facilitate resistance and are located in the inner domain, loop D, and β23/loop V5/β24 of HIV-1 gp120. The resistance is largely correlated with binding affinity of antibodies to the envelope trimers expressed on the cell surface. Our results therefore demonstrate the existence of broadly resistant HIV-1 strains against CD4bs neutralizing antibodies. Treatment strategies based on the CD4bs bnAbs must overcome such resistance to achieve optimal clinical outcomes. |
3,738 | Identification of a Novel Non-desmoglein Autoantigen in Pemphigus Vulgaris | Pemphigus vulgaris (PV) is an autoimmune bullous disease of the skin and mucous membranes characterized by the presence of circulating and tissue-bound autoantibodies against keratinocyte cell surface antigens, specifically desmoglein (Dsg) 1 and 3. The pathogenic role of anti-Dsg antibodies is well-established, while the mechanism of blister formation is only partly defined. We have applied a previously developed method for the efficient immortalization of IgG+ memory B cells to identify novel target antigens in PV. A human monoclonal antibody reactive with a hitherto unreported non-Dsg antigen was isolated. Immunoprecipitation and immunoblotting studies with keratinocyte extracts indicated α-catenin as the putative antigen, then confirmed by immunoblotting on the recombinant protein. Four of ten PV sera reacted with recombinant α-catenin. Although the isolated human monoclonal antibody was per se unable to dissociate keratinocyte monolayers and also to synergize with a pathogenic antibody in vitro, further studies are warranted to assess its possible in vivo contribution in the multifactorial pathogenesis and heterogeneous manifestations of PV disease. |
3,739 | Aims, Study Design, and Enrollment Results From the Assessing Predictors of Infant Respiratory Syncytial Virus Effects and Severity Study | BACKGROUND: The majority of infants hospitalized with primary respiratory syncytial virus (RSV) infection have no obvious risk factors for severe disease. OBJECTIVE: The aim of this study (Assessing Predictors of Infant RSV Effects and Severity, AsPIRES) was to identify factors associated with severe disease in full-term healthy infants younger than 10 months with primary RSV infection. METHODS: RSV infected infants were enrolled from 3 cohorts during consecutive winters from August 2012 to April 2016 in Rochester, New York. A birth cohort was prospectively enrolled and followed through their first winter for development of RSV infection. An outpatient supplemental cohort was enrolled in the emergency department or pediatric offices, and a hospital cohort was enrolled on admission with RSV infection. RSV was diagnosed by reverse transcriptase-polymerase chain reaction. Demographic and clinical data were recorded and samples collected for assays: buccal swab (cytomegalovirus polymerase chain reaction, PCR), nasal swab (RSV qualitative PCR, complete viral gene sequence, 16S ribosomal ribonucleic acid [RNA] amplicon microbiota analysis), nasal wash (chemokine and cytokine assays), nasal brush (nasal respiratory epithelial cell gene expression using RNA sequencing [RNAseq]), and 2 to 3 ml of heparinized blood (flow cytometry, RNAseq analysis of purified cluster of differentiation [CD]4+, CD8+, B cells and natural killer cells, and RSV-specific antibody). Cord blood (RSV-specific antibody) was also collected for the birth cohort. Univariate and multivariate logistic regression will be used for analysis of data using a continuous Global Respiratory Severity Score (GRSS) as the outcome variable. Novel statistical methods will be developed for integration of the large complex datasets. RESULTS: A total of 453 infants were enrolled into the 3 cohorts; 226 in the birth cohort, 60 in the supplemental cohort, and 78 in the hospital cohort. A total of 126 birth cohort infants remained in the study and were evaluated for 150 respiratory illnesses. Of the 60 RSV positive infants in the supplemental cohort, 42 completed the study, whereas all 78 of the RSV positive hospital cohort infants completed the study. A GRSS was calculated for each RSV-infected infant and is being used to analyze each of the complex datasets by correlation with disease severity in univariate and multivariate methods. CONCLUSIONS: The AsPIRES study will provide insights into the complex pathogenesis of RSV infection in healthy full-term infants with primary RSV infection. The analysis will allow assessment of multiple factors potentially influencing the severity of RSV infection including the level of RSV specific antibodies, the innate immune response of nasal epithelial cells, the adaptive response by various lymphocyte subsets, the resident airway microbiota, and viral factors. Results of this study will inform disease interventions such as vaccines and antiviral therapies. |
3,740 | ABCB1 protects bat cells from DNA damage induced by genotoxic compounds | Bats are unusual mammals, with the ability to fly, and long lifespans. In addition, bats have a low incidence of cancer, but the mechanisms underlying this phenomenon remain elusive. Here we discovered that bat cells are more resistant than human and mouse cells to DNA damage induced by genotoxic drugs. We found that bat cells accumulate less chemical than human and mouse cells, and efficient drug efflux mediated by the ABC transporter ABCB1 underlies this improved response to genotoxic reagents. Inhibition of ABCB1 triggers an accumulation of doxorubicin, DNA damage, and cell death. ABCB1 is expressed at higher levels in several cell lines and tissues derived from bats compared to humans. Furthermore, increased drug efflux and high expression of ABCB1 are conserved across multiple bat species. Our findings suggest that enhanced efflux protects bat cells from DNA damage induced by genotoxic compounds, which may contribute to their low cancer incidence. |
3,741 | Identification and Molecular Characterisation of Bovine Parainfluenza Virus-3 and Bovine Respiratory Syncytial Virus - First Report from Turkey | INTRODUCTION: Bovine parainfluenza virus-3 (BPIV3) and bovine respiratory syncytial virus (BRSV) are the cause of respiratory disease in cattle worldwide. With other pathogens, they cause bovine respiratory disease complex (BRDC) in ruminants. The aim of the study was the detection and molecular characterisation of BPIV3 and BRSV from nasal swabs and lung samples of cows in and around the Erzurum region of eastern Turkey. MATERIAL AND METHODS: In total, 155 samples were collected. Of animals used in the study 92 were males and 63 females. The age of the animals was between 9 months and 5 years, mean 1.4 years. Most males were in the fattening period and being raised in open sheds; females were in the lactating period and kept in free stall barns. All samples were tested for the presence of viral genes using RT-PCR. Gene-specific primers in a molecular method (RT-PCR) identified BRSV (fusion gene) and BPIV3 (matrix gene) strains at the genus level. RESULTS: RNA from BRSV and BPIV3 was detected in two (1.29%) and three (1.93%) samples, respectively, one of each of which was sequenced and the sequences were aligned with reference virus strains. Phylogenetic analyses clustered the strains in genotype C/BPIV3 and subgroup III/BRSV. CONCLUSION: The results indicate that BRSV and BPIV3 contribute to bovine respiratory disease cases in Turkey. This is the first report on their detection and molecular characterisation in ruminants in Turkey. |
3,742 | Comparison of the immune response to vaccination with pigeon circovirus recombinant capsid protein (PiCV rCP) in pigeons uninfected and subclinically infected with PiCV | Infections with immunosuppressive pigeon circovirus (PiCV) pose the most severe health problem to the global pigeon breeding. The vaccination with immunogenic PiCV recombinant capsid protein (PiCV rCP) is a potential tool for disease control. Because of the high prevalence of PiCV asymptomatic infections, the subclinically infected pigeons will be vaccinated in practice. The aim of this study was to answer a question if vaccination of asymptomatic, infected with PiCV pigeons induces a similar immune response to PiCV rCP as in uninfected birds. One hundred and twenty 6-week-old carrier pigeons were divided into 4 groups (2 groups of naturally infected and uninfected with PiCV individuals). Birds from groups V and V1 were vaccinated twice with PiCV rCP mixed with an adjuvant, whereas pigeons from groups C and C1 were immunized with an adjuvant only. The expression of genes encoding IFN-γ, CD4, and CD8 T lymphocyte receptors; the number of anti-PiCV rCP IgY-secreting B cells (SBC) and anti-PiCV rCP IgY were evaluated 2, 21, 39 and 46 days post vaccination (dpv). Study results showed that the expression of CD8 and IFN-γ genes was higher in both groups of infected pigeons than in the uninfected birds, irrespective of vaccination. In the uninfected birds, the expression of these genes was insignificantly higher in the vaccinated pigeons. The anti-PiCV rCP IgY-SBC were detected on 2 and 23 dpv and seroconversion was noted on 23 and 39 dpv in V and V1 groups, respectively. In the light of the results obtained, it could be concluded that pigeon circovirus recombinant capsid protein elicits the immune response in both naturally infected and uninfected pigeons, but its rate varies depending on PiCV infectious status. The infection with PiCV masks the potential cellular immune response to the vaccination with PiCV rCP and leads to the suppression of humoral immunity. |
3,743 | “In sickness and in health” – how neutrophil extracellular trap (NET) works in infections, selected diseases and pregnancy | The discovery of the NET network (neutrophil extracellular trap) has revolutionized the perception of defense mechanisms used by neutrophils in infections and non-infectious states, as this mechanism proves the complexity of the ways in which neutrophils can act in the organism. The paper describes the NET network and its participation in bacterial, viral, fungal and parasitic infections, both in a positive and a negative aspect. In addition, attention was paid to the participation of NETs in the course of autoimmune diseases, cancer, as well as its impact on pregnancy and fertility in mammals. |
3,744 | 3′,8″-Dimerization Enhances the Antioxidant Capacity of Flavonoids: Evidence from Acacetin and Isoginkgetin | To probe the effect of 3′,8″-dimerization on antioxidant flavonoids, acacetin and its 3′,8″-dimer isoginkgetin were comparatively analyzed using three antioxidant assays, namely, the ·O(2)(−) scavenging assay, the Cu(2+) reducing assay, and the 2,2′-azino bis(3-ethylbenzothiazolin-6-sulfonic acid) radical scavenging assay. In these assays, acacetin had consistently higher IC(50) values than isoginkgetin. Subsequently, the acacetin was incubated with 4-methoxy-2,2,6,6-tetramethylpiperidine-1-oxy radicals (4-methoxy-TEMPO) and then analyzed by ultra-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight tandem mass spectrometry (UHPLC−ESI−Q−TOF−MS) technology. The results of the UHPLC−ESI−Q−TOF−MS analysis suggested the presence of a dimer with m/z 565, 550, 413, 389, 374, 345, 330, and 283 peaks. By comparison, standard isoginkgetin yielded peaks at m/z 565, 533, 518, 489, 401, 389, 374, and 151 in the mass spectra. Based on these experimental data, MS interpretation, and the relevant literature, we concluded that isoginkgetin had higher electron transfer potential than its monomer because of the 3′,8″-dimerization. Additionally, acacetin can produce a dimer during its antioxidant process; however, the dimer is not isoginkgetin. |
3,745 | HBV Immune-Therapy: From Molecular Mechanisms to Clinical Applications | Chronic hepatitis B virus (HBV) infection represents a worldwide public health concern with approximately 250 million people chronically infected and at risk of developing liver cirrhosis and hepatocellular carcinoma. Nucleos(t)ide analogues (NUC) are the most widely used therapies for HBV infection, but they often require long-lasting administration to avoid the risk of HBV reactivation at withdrawal. Therefore, there is an urgent need to develop novel treatments to shorten the duration of NUC therapy by accelerating virus control, and to complement the effect of available anti-viral therapies. In chronic HBV infection, virus-specific T cells are functionally defective, and this exhaustion state is a key determinant of virus persistence. Reconstitution of an efficient anti-viral T cell response may thus represent a rational strategy to treat chronic HBV patients. In this perspective, the enhancement of adaptive immune responses by a checkpoint inhibitor blockade, specific T cell vaccines, lymphocyte metabolism targeting, and autologous T cell engineering, including chimeric antigen receptor (CAR) and TCR-redirected T cells, constitutes a promising immune modulatory approach for a therapeutic restoration of protective immunity. The advances of the emerging immune-based therapies in the setting of the HBV research field will be outlined. |
3,746 | Respiratory syncytial virus in hematopoietic cell transplant recipients and patients with hematologic malignancies | In the USA and other western nations, respiratory syncytial virus is one of the most commonly encountered respiratory viruses among patients who have been diagnosed with a hematologic malignancy or who have undergone a stem cell transplant. Multiple studies have been performed to evaluate the complications associated with respiratory syncytial virus infections. Other studies have evaluated therapeutic agents and strategies in which these agents can be used. There have also been numerous reports of outbreaks in bone marrow transplant units and oncology wards, where infection control measures have been invaluable in controlling the spread of disease. However, despite these novel approaches, respiratory syncytial virus continues to be potentially fatal in immunocompromised populations. In this review, we discuss the incidence of respiratory syncytial viral infections, risk factors associated with progression from upper respiratory tract infection to lower respiratory tract infection, other complications and outcomes (including mortality), management strategies, and prevention strategies in patients with a hematologic malignancy and in hematopoietic cell transplant recipients. |
3,747 | Assessing Zika Virus Transmission Within Households During an Outbreak in Martinique, 2015–2016 | Since 2015, Zika virus (ZIKV) has caused large epidemics in the Americas. Households are natural targets for control interventions, but quantification of the contribution of household transmission to overall spread is needed to guide policy. We developed a modeling framework to evaluate this contribution and key epidemic features of the ZIKV epidemic in Martinique in 2015–2016 from the joint analysis of a household transmission study (n = 68 households), a study among symptomatic pregnant women (n = 281), and seroprevalence surveys of blood donors (n = 457). We estimated that the probability of mosquito-mediated within-household transmission (from an infected member to a susceptible one) was 21% (95% credible interval (CrI): 5, 51), and the overall probability of infection from outside the household (i.e., in the community) was 39% (95% CrI: 27, 50). Overall, 50% (95% CrI: 43, 58) of the population was infected, with 22% (95% CrI: 5, 46) of infections acquired in households and 40% (95% CrI: 23, 56) being asymptomatic. The probability of presenting with Zika-like symptoms due to another cause was 16% (95% CrI: 10, 23). This study characterized the contribution of household transmission in ZIKV epidemics, demonstrating the benefits of integrating multiple data sets to gain more insight into epidemic dynamics. |
3,748 | Cross-Species Genome-Wide Analysis Reveals Molecular and Functional Diversity of the Unconventional Interferon-ω Subtype | Innate immune interferons (IFNs), particularly type I IFNs, are primary mediators regulating animal antiviral, antitumor, and cell-proliferative activity. These antiviral cytokines have evolved remarkable molecular and functional diversity to confront ever-evolving viral threats and physiological regulation. We have annotated IFN gene families across 110 animal genomes, and showed that IFN genes, after originating in jawed fishes, had several significant evolutionary surges in vertebrate species of amphibians, bats and ungulates, particularly pigs and cattle. For example, pigs have the largest but still expanding type I IFN family consisting of nearly 60 IFN-coding genes that encode seven IFN subtypes including multigene subtypes of IFN-α, -δ, and -ω. Whereas, subtypes such as IFN-α and -β have been widely studied in many species, the unconventional subtypes such as IFN-ω have barely been investigated. We have cross-species defined the IFN evolution, and shown that unconventional IFN subtypes particularly the IFN-ω subtype have evolved several novel features including: (1) being a signature multi-gene subtype expanding primarily in mammals such as bats and ungulates, (2) emerging isoforms that have superior antiviral potency than typical IFN-α, (3) highly cross-species antiviral (but little anti-proliferative) activity exerted in cells of humans and other mammalian species, and (4) demonstrating potential novel molecular and functional properties. This study focused on IFN-ω to investigate the immunogenetic evolution and functional diversity of unconventional IFN subtypes, which may further IFN-based novel antiviral design pertinent to their cross-species high antiviral and novel activities. |
3,749 | A novel group of avian astroviruses from Neotropical passerine birds broaden the diversity and host range of Astroviridae | Metagenomics is helping to expand the known diversity of viruses, especially of those with poorly studied hosts in remote areas. The Neotropical region harbors a considerable diversity of avian species that may play a role as both host and short-distance vectors of unknown viruses. Viral metagenomics of cloacal swabs from 50 Neotropical birds collected in French Guiana revealed the presence of four complete astrovirus genomes. They constitute an early diverging novel monophyletic clade within the Avastrovirus phylogeny, representing a putative new astrovirus species (provisionally designated as Avastrovirus 5) according to the International Committee on Taxonomy of Viruses (ICTV) classification criteria. Their genomic organization shares some characteristics with Avastrovirus but also with Mamastrovirus. The pan-astrovirus RT-PCR analysis of the cloacal samples of 406 wild Neotropical birds showed a community-level prevalence of 4.9% (5.1% in passerines, the highest described so far in this order of birds). By screening birds of a remote region, we expanded the known host range of astroviruses to the avian families Cardinalidae, Conopophagidae, Furnariidae, Thamnophilidae, Turdidae and Tyrannidae. Our results provide important first insights into the unexplored viral communities, the ecology, epidemiology and features of host-pathogen interactions that shape the evolution of avastroviruses in a remote Neotropical rainforest. |
3,750 | The interleukin-4/PPARγ signaling axis promotes oligodendrocyte differentiation and remyelination after brain injury | The repair of white matter damage is of paramount importance for functional recovery after brain injuries. Here, we report that interleukin-4 (IL-4) promotes oligodendrocyte regeneration and remyelination. IL-4 receptor expression was detected in a variety of glial cells after ischemic brain injury, including oligodendrocyte lineage cells. IL-4 deficiency in knockout mice resulted in greater deterioration of white matter over 14 d after stroke. Consistent with these findings, intranasal delivery of IL-4 nanoparticles after stroke improved white matter integrity and attenuated long-term sensorimotor and cognitive deficits in wild-type mice, as revealed by histological immunostaining, electron microscopy, diffusion tensor imaging, and electrophysiology. The selective effect of IL-4 on remyelination was verified in an ex vivo organotypic model of demyelination. By leveraging primary oligodendrocyte progenitor cells (OPCs), microglia-depleted mice, and conditional OPC-specific peroxisome proliferator-activated receptor gamma (PPARγ) knockout mice, we discovered a direct salutary effect of IL-4 on oligodendrocyte differentiation that was mediated by the PPARγ axis. Our findings reveal a new regenerative role of IL-4 in the central nervous system (CNS), which lies beyond its known immunoregulatory functions on microglia/macrophages or peripheral lymphocytes. Therefore, intranasal IL-4 delivery may represent a novel therapeutic strategy to improve white matter integrity in stroke and other brain injuries. |
3,751 | 2HybridTools, a handy software to facilitate clone identification and mutation mapping from yeast two-hybrid screening | Yeast Two-Hybrid (Y2H) and reverse Two-Hybrid (RY2H) are powerful protein–protein interaction screening methods that rely on the interaction of bait and prey proteins fused to DNA binding (DB) and activation domains (AD), respectively. Y2H allows identification of protein interaction partners using screening libraries, while RY2H is used to determine residues critical to a given protein–protein interaction by exploiting site-directed mutagenesis. Currently, both these techniques still rely on sequencing of positive clones using conventional Sanger sequencing. For Y2H, a screen can yield several positives; the identification of such clones is further complicated by the fact that sequencing products usually contain vector sequence. For RY2H, obtaining a complete sequence is required to identify the full range of residues involved in protein–protein interactions. However, with Sanger sequencing limited to 500–800 nucleotides, sequencing is usually carried from both ends for clones greater than this length. Analysis of such RY2H data thus requires assembly of sequencing products combined with trimming of vector sequences and of low-quality bases at the beginning and ends of sequencing products. Further, RY2H analysis requires collation of mutations that abrogate a DB/AD interaction. Here, we present 2HybridTools, a Java program with a user-friendly interface that allows addressing all these issues inherent to both Y2H and RY2H. Specifically, for Y2H, 2HybridTools enables automated identification of positive clones, while for RY2H, 2HybridTools provides detailed mutation reports as a basis for further investigation of given protein–protein interactions. |
3,752 | Influenza Before the 1890 and 1918 Pandemics in the US Army and at the US Military Academy | Influenza rates for the US Army and West Point cadets showed that seasonal influenza was not necessarily an annual event, and there was little influenzal illness in the decade before 1918 except for 1911 and 1916. Annual records from 1862–1918 also indicated a similar paucity of influenzal illness before 1890. |
3,753 | Small Extracellular Vesicles Are Key Regulators of Non-cell Autonomous Intercellular Communication in Senescence via the Interferon Protein IFITM3 | Senescence is a cellular phenotype present in health and disease, characterized by a stable cell-cycle arrest and an inflammatory response called senescence-associated secretory phenotype (SASP). The SASP is important in influencing the behavior of neighboring cells and altering the microenvironment; yet, this role has been mainly attributed to soluble factors. Here, we show that both the soluble factors and small extracellular vesicles (sEVs) are capable of transmitting paracrine senescence to nearby cells. Analysis of individual cells internalizing sEVs, using a Cre-reporter system, show a positive correlation between sEV uptake and senescence activation. We find an increase in the number of multivesicular bodies during senescence in vivo. sEV protein characterization by mass spectrometry (MS) followed by a functional siRNA screen identify interferon-induced transmembrane protein 3 (IFITM3) as being partially responsible for transmitting senescence to normal cells. We find that sEVs contribute to paracrine senescence. |
3,754 | Nonhuman primate species as models of human bacterial sepsis | Sepsis involves a disordered host response to systemic infection leading to high morbidity and mortality. Despite intense research, targeted sepsis therapies beyond antibiotics have remained elusive. The cornerstone of sepsis research is the development of animal models to mimic human bacterial infections and test novel pharmacologic targets. Nonhuman primates (NHPs) have served as an attractive, but expensive, animal to model human bacterial infections due to their nearly identical cardiopulmonary anatomy and physiology, as well as host response to infection. Several NHP species have provided substantial insight into sepsis-mediated inflammation, endothelial dysfunction, acute lung injury, and multi-organ failure. The use of NHPs has usually focused on translating therapies from early preclinical models to human clinical trials. However, despite successful sepsis interventions in NHP models, there are still no FDA-approved sepsis therapies. This review highlights major NHP models of bacterial sepsis and their relevance to clinical medicine. |
3,755 | African Swine Fever Virus Armenia/07 Virulent Strain Controls Interferon Beta Production through the cGAS-STING Pathway | African swine fever virus (ASFV) is a complex, cytoplasmic double-stranded DNA (dsDNA) virus that is currently expanding throughout the world. Currently, circulating virulent genotype II Armenia/07-like viruses cause fatal disease in pigs and wild boar, whereas attenuated strains induce infections with various levels of chronic illness. Sensing cytosolic dsDNA, mainly by the key DNA sensor cyclic GMP-AMP synthase (cGAS), leads to the synthesis of type I interferon and involves signaling through STING, TBK1, and IRF3. After phosphorylation, STING translocates from the endoplasmic reticulum to the Golgi compartment and to the perinuclear region, acting as an indispensable adaptor connecting the cytosolic detection of DNA to the TBK1-IRF3 signaling pathway. We demonstrate here that attenuated NH/P68, but not virulent Armenia/07, activates the cGAS-STING-IRF3 cascade very early during infection, inducing STING phosphorylation and trafficking through a mechanism involving cGAMP. Both TBK1 and IRF3 are subsequently activated and, in response to this, a high level of beta interferon (IFN-β) was produced during NH/P68 infection; in contrast, Armenia/07 infection generated IFN-β levels below those of uninfected cells. Our results show that virulent Armenia/07 ASFV controls the cGAS-STING pathway, but these mechanisms are not at play when porcine macrophages are infected with attenuated NH/P68 ASFV. These findings show for the first time the involvement of the cGAS-STING-IRF3 route in ASFV infection, where IFN-β production or inhibition was found after infection by attenuated or virulent ASFV strains, respectively, thus reinforcing the idea that ASFV virulence versus attenuation may be a phenomenon grounded in ASFV-mediated innate immune modulation where the cGAS-STING pathway might play an important role. IMPORTANCE African swine fever, a devastating disease for domestic pigs and wild boar, is currently spreading in Europe, Russia, and China, becoming a global threat with huge economic and ecological consequences. One interesting aspect of ASFV biology is the molecular mechanism leading to high virulence of some strains compared to more attenuated strains, which produce subclinical infections. In this work, we show that the presently circulating virulent Armenia/07 virus blocks the synthesis of IFN-β, a key mediator between the innate and adaptive immune response. Armenia/07 inhibits the cGAS-STING pathway by impairing STING activation during infection. In contrast, the cGAS-STING pathway is efficiently activated during NH/P68 attenuated strain infection, leading to the production of large amounts of IFN-β. Our results show for the first time the relationship between the cGAS-STING pathway and ASFV virulence, contributing to uncover the molecular mechanisms of ASFV virulence and to the rational development of ASFV vaccines. |
3,756 | Engineering for Success: Approaches to Improve Chimeric Antigen Receptor T Cell Therapy for Solid Tumors | While impressive clinical responses have been observed using chimeric antigen receptor (CAR) T cells targeting CD19+ hematologic malignancies, limited clinical benefit has been observed using CAR T cells for a variety of solid tumors. Results of clinical studies have highlighted several obstacles which CAR T cells face in the context of solid tumors, including insufficient homing to tumor sites, lack of expansion and persistence, encountering a highly immunosuppressive tumor microenvironment, and heterogeneous antigen expression. In this review, we review clinical outcomes and discuss strategies to improve the antitumor activity of CAR T cells for solid tumors. |
3,757 | Transcription attenuation-derived small RNA rnTrpL regulates tryptophan biosynthesis gene expression in trans | Ribosome-mediated transcription attenuation is a basic posttranscriptional regulation mechanism in bacteria. Liberated attenuator RNAs arising in this process are generally considered nonfunctional. In Sinorhizobium meliloti, the tryptophan (Trp) biosynthesis genes are organized into three operons, trpE(G), ppiD-trpDC-moaC-moeA, and trpFBA-accD-folC, of which only the first one, trpE(G), contains a short ORF (trpL) in the 5′-UTR and is regulated by transcription attenuation. Under conditions of Trp sufficiency, transcription is terminated between trpL and trpE(G), and a small attenuator RNA, rnTrpL, is produced. Here, we show that rnTrpL base-pairs with trpD and destabilizes the polycistronic trpDC mRNA, indicating rnTrpL-mediated downregulation of the trpDC operon in trans. Although all three trp operons are regulated in response to Trp availability, only in the two operons trpE(G) and trpDC the Trp-mediated regulation is controlled by rnTrpL. Together, our data show that the trp attenuator coordinates trpE(G) and trpDC expression posttranscriptionally by two fundamentally different mechanisms: ribosome-mediated transcription attenuation in cis and base-pairing in trans. Also, we present evidence that rnTrpL-mediated regulation of trpDC genes expression in trans is conserved in Agrobacterium and Bradyrhizobium, suggesting that the small attenuator RNAs may have additional conserved functions in the control of bacterial gene expression. |
3,758 | Global health diplomacy, health and human security: The ascendancy of enlightened self-interest | The political, social, economic, and security implications of health-related issues such as emerging infectious diseases or the epidemic of Non Communicable Diseases offer a rare opportunity for professionals in foreign policy and international relations to engage with the health arena and at the same time for global health experts to enter into and intersect with the domain of diplomacy. The aim of this review is to understand and explore the concepts of global health diplomacy (GHD), health security, and human security. For this narrative review, a literature search was done in PubMed, Scopus, and EBSCO for the “global health diplomacy,” “health security,” and “human security,” and full-texts were reviewed. The recent outbreaks of Ebola in West Africa and Zika in South America are pertinent examples of the nature of the human security crisis and the imminent and severe threat posed to human life across the globe as a result of these epidemics. The Commission on Human Security defines human security as the protection of the vital core of all human lives from critical and pervasive threats. We highlight the ways in which health has now become an issue of national security/global concern and also how GHD can aid in the development of new bilateral or multilateral agreements to safeguard the health and security of people in our globalized world. The paper provides a prospective about, and overview of, health and human security that essentially emphasizes the growing interlinkages between global health, diplomacy, and foreign policy. |
3,759 | Microarray analysis of infectious bronchitis virus infection of chicken primary dendritic cells | BACKGROUND: Avian infectious bronchitis virus (IBV) is a major respiratory disease-causing agent in birds that leads to significant losses. Dendritic cells (DCs) are specialised cells responsible for sampling antigens and presenting them to T cells, which also play an essential role in recognising and neutralising viruses. Recent studies have suggested that non-coding RNAs may regulate the functional program of DCs. Expression of host non-coding RNAs changes markedly during infectious bronchitis virus infection, but their role in regulating host immune function has not been explored. Here, microarrays of mRNAs, miRNAs, and lncRNAs were globally performed to analyse how avian DCs respond to IBV. RESULTS: First, we found that IBV stimulation did not enhance the maturation ability of avian DCs. Interestingly, inactivated IBV was better able than IBV to induce DC maturation and activate lymphocytes. We identified 1093 up-regulated and 845 down-regulated mRNAs in IBV-infected avian DCs. Gene Ontology analysis suggested that cellular macromolecule and protein location (GO-BP) and transcription factor binding (GO-MF) were abundant in IBV-stimulated avian DCs. Meanwhile, pathway analysis indicated that the oxidative phosphorylation and leukocyte transendothelial migration signalling pathways might be activated in the IBV group. Moreover, alteration of microRNAs (miRNAs) and long non-coding RNAs (lncRNAs) was detected in IBV-stimulated avian DCs. In total, 19 significantly altered (7 up and 12 down) miRNAs and 101 (75 up and 26 down) lncRNAs were identified in the IBV-treated group. Further analysis showed that the actin cytoskeleton and MAPK signal pathway were related to the target genes of IBV-stimulated miRNAs. Finally, our study identified 2 TF-microRNA and 53 TF–microRNA–mRNA interactions involving 1 TF, 2 miRNAs, and 53 mRNAs in IBV-stimulated avian DCs. CONCLUSIONS: Our research suggests a new mechanism to explain why IBV actively blocks innate responses needed for inducing immune gene expression and also provides insight into the pathogenic mechanisms of avian IBV. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12864-019-5940-6) contains supplementary material, which is available to authorized users. |
3,760 | TIM-1 serves as a receptor for Ebola virus in vivo, enhancing viremia and pathogenesis | BACKGROUND: T cell immunoglobulin mucin domain-1 (TIM-1) is a phosphatidylserine (PS) receptor, mediating filovirus entry into cells through interactions with PS on virions. TIM-1 expression has been implicated in Ebola virus (EBOV) pathogenesis; however, it remains unclear whether this is due to TIM-1 serving as a filovirus receptor in vivo or, as others have suggested, TIM-1 induces a cytokine storm elicited by T cell/virion interactions. Here, we use a BSL2 model virus that expresses EBOV glycoprotein to demonstrate the importance of TIM-1 as a virus receptor late during in vivo infection. METHODOLOGY/PRINCIPAL FINDINGS: Infectious, GFP-expressing recombinant vesicular stomatitis virus encoding either full length EBOV glycoprotein (EBOV GP/rVSV) or mucin domain deleted EBOV glycoprotein (EBOV GPΔO/rVSV) was used to assess the role of TIM-1 during in vivo infection. GFP-expressing rVSV encoding its native glycoprotein G (G/rVSV) served as a control. TIM-1-sufficient or TIM-1-deficient BALB/c interferon α/β receptor(-/-) mice were challenged with these viruses. While G/rVSV caused profound morbidity and mortality in both mouse strains, TIM-1-deficient mice had significantly better survival than TIM-1-expressing mice following EBOV GP/rVSV or EBOV GPΔO/rVSV challenge. EBOV GP/rVSV or EBOV GPΔO/rVSV in spleen of infected animals was high and unaffected by expression of TIM-1. However, infectious virus in serum, liver, kidney and adrenal gland was reduced late in infection in the TIM-1-deficient mice, suggesting that virus entry via this receptor contributes to virus load. Consistent with higher virus loads, proinflammatory chemokines trended higher in organs from infected TIM-1-sufficient mice compared to the TIM-1-deficient mice, but proinflammatory cytokines were more modestly affected. To assess the role of T cells in EBOV GP/rVSV pathogenesis, T cells were depleted in TIM-1-sufficient and -deficient mice and the mice were challenged with virus. Depletion of T cells did not alter the pathogenic consequences of virus infection. CONCLUSIONS: Our studies provide evidence that at late times during EBOV GP/rVSV infection, TIM-1 increased virus load and associated mortality, consistent with an important role of this receptor in virus entry. This work suggests that inhibitors which block TIM-1/virus interaction may serve as effective antivirals, reducing virus load at late times during EBOV infection. |
3,761 | Changes in Serum Amyloid A (SAA) Concentration in Arabian Endurance Horses During First Training Season | Sport training leads to adaptation to physical effort that is reflected by the changes in blood parameters. In equine endurance athletes, blood testing is accepted as a support in training, however, only the changes before versus after exercise in creatine phosphokinase activity (CPK) and basic blood parameters are usually measured. This study is the first longitudinal investigation of the changes in routinely measured blood parameters and, additionally, serum amyloid A (SAA), during seven months, in Arabian horses introduced to endurance training and competing in events for young horses. It has been determined that CPK, aspartate aminotransferase (AST), packed cell volume (PCV), hemoglobin concentration, red blood cell count (RBC), and concentration of total serum protein (TSP) slightly increased after training sessions and competitions in similar manner. The increase in white blood cell (WBC) count was higher after competitions and SAA increased only after competitions. Total protein concentration was the only parameter that increased with training during a 7-month program. SAA indicated only in the case of heavy effort, and, it thus may be helpful in the monitoring of training in young horses. In an optimal program, its concentration should not increase after a training session but only after heavy effort, which should not be repeated too often. |
3,762 | Mapping the drivers of within-host pathogen evolution using massive data sets | Differences among hosts, resulting from genetic variation in the immune system or heterogeneity in drug treatment, can impact within-host pathogen evolution. Genetic association studies can potentially identify such interactions. However, extensive and correlated genetic population structure in hosts and pathogens presents a substantial risk of confounding analyses. Moreover, the multiple testing burden of interaction scanning can potentially limit power. We present a Bayesian approach for detecting host influences on pathogen evolution that exploits vast existing data sets of pathogen diversity to improve power and control for stratification. The approach models key processes, including recombination and selection, and identifies regions of the pathogen genome affected by host factors. Our simulations and empirical analysis of drug-induced selection on the HIV-1 genome show that the method recovers known associations and has superior precision-recall characteristics compared to other approaches. We build a high-resolution map of HLA-induced selection in the HIV-1 genome, identifying novel epitope-allele combinations. |
3,763 | Efficacy of Houttuynia eye drops for the treatment of vernal keratoconjunctivitis: A systemic review and meta-analysis protocol | BACKGROUND: Vernal keratoconjunctivitis (VKC) is a common eye disease and can result in permanent decrease or loss of vision. Houttuynia eye drops (HED) is used for the treatment of VKC. However, the clinical evidence of HED has not been well concluded. Herein, we described a proposed systemic review and meta-analysis to evaluate the clinical efficacy of HED for the treatment of VKC. METHODS: Six electronic databases (Medline, Embase, the Cochrane database, Chinese National Knowledge Infrastructure, Wanfang database, and Chinese Biology and Medicine database) will be searched for randomized controlled trials (RCTs) which evaluating the clinical efficacy of HED for the treatment of VKC. Studies meet the eligibility criteria will be included. Data of the included studies will be extracted and the quality will also be evaluated. Data synthesis will be performed using RevMan software. Sensitivity analysis and publication bias will also be investigated. RESULTS: This study will provide high-quality systemic review and synthesis of RCTs on efficacy of HED for the treatment of VKC. CONCLUSION: This systemic review and meta-analysis will conclude the efficacy of HED for the treatment of VKC. REGISTRATION: PROSPERO CRD42019124737 |
3,764 | Virucidal activity of Garcinia parvifolia leaf extracts in animal cell culture | BACKGROUND: Garcinia species contain bioactive compounds such as flavonoids, xanthones, triterpernoids, and benzophenones with antibacterial, antifungal, anti-inflammatory, and antioxidant activities. In addition, many of these compounds show interesting biological properties such as anti-human immunodeficiency virus activity. Garcinia parvifolia is used in traditional medicine. Currently, the antiviral activity of G. parvifolia is not known. METHODS: This study was conducted to determine the effects of ethyl acetate (45 L Ea), ethanol (45 L Et), and hexane (45 L H) leaf extracts of G. parvifolia on the infectivity of pseudorabies virus (PrV) in Vero cells. The antiviral effects of the extracts were determined by cytopathic effect (CPE), inhibition, attachment, and virucidal assays. RESULTS: The 50% cytotoxicity concentration (CC(50)) values obtained were 237.5, 555.0, and < 1.25 μg/mL for 45 L Ea, 45 L Et, and 45 L H, respectively. The 45 L Ea showed the greatest viral inhibition potency of 75% at 125 μg/mL. Both 45 L Ea and 45 l Et caused 100% residual viral inhibition at 250 μg/mL. The selectivity index values for 45 L Ea, 45 L Et, and 45 L H were 2.65, 1.75, and 0.10 showing that 45 L Ea had the greatest antiviral activity among the three extracts. CONCLUSION: This study showed that ethyl acetate is the best solvent to be used to obtain extract from G. parvifolia leaves with potent antiviral activities. |
3,765 | Establishment and development of national community-based collaborative innovation demonstration areas to achieve the control target of hepatitis B in China | BACKGROUND: The major infectious diseases of hepatitis B has constituted an acute public health challenge in China. An effective and affordable HBV control model is urgently needed. A national project of Community-based Collaborative Innovation HBV (CCI-HBV) demonstration areas has optimized the existing community healthcare resources and obtained initial results in HBV control. METHODS: Based on the existing community healthcare network, CCI-HBV project combined the community health management and health contract signing service for long-staying residents in hepatitis B screening. Moreover, HBV field research strategy was popularized in CCI-HBV areas. After screening, patients with seropositive results were enrolled in corresponding cohorts and received treatment at an early stage. And the uninfected people received medical supports including health education through new media, behavior intervention and HBV vaccinations. In this process, a cloud-based National Information Platform (NIP) was established to collect and store residents’ epidemiological data. In addition, a special quality control team was set up for CCI project. RESULTS: After two rounds of screening, HBsAg positive rate dropped from 5.05% (with 5,173,003 people screened) to 4.57% (with 3,819,675 people screened), while the rate of new HBV infections was 0.28 per 100 person-years in the fixed cohorts of 2,584,322 people. The quality control team completed PPS sampling simultaneously and established the serum sample database with 2,800,000 serum samples for unified testing. CONCLUSIONS: CCI-HBV project has established a large-scale field research to conduct whole-population screening and intervention. We analyzed the HBsAg prevalence and new infection rate of HBV in the fixed population for the epidemic trend and intervention effect. The purpose of CCI-HBV project is to establish and evaluate a practical model of grid management and field strategy, to realize the new goal to control hepatitis B in China. To provide policymakers with a feasible model, our results are directly applicable. TRIAL REGISTRATION: The project was funded by the Major Projects of Science Research for the 11th and 12th five-year plans of China, entitled “The prevention and control of AIDS, viral hepatitis and other major infectious diseases”, Grant Nos. 2009ZX10004901, 2011ZX10004901, 2013ZX10004904, 2014ZX10004007 and 2014ZX10004008. |
3,766 | Slow-Binding Inhibition of Tyrosinase by Ecklonia cava Phlorotannins | Tyrosinase inhibitors improve skin whitening by inhibiting the formation of melanin precursors in the skin. The inhibitory activity of seven phlorotannins (1–7), triphlorethol A (1), eckol (2), 2-phloroeckol (3), phlorofucofuroeckol A (4), 2-O-(2,4,6-trihydroxyphenyl)-6,6′-bieckol (5), 6,8′-bieckol (6), and 8,8′-bieckol (7), from Ecklonia cava was tested against tyrosinase, which converts tyrosine into dihydroxyphenylalanine. Compounds 3 and 5 had IC(50) values of 7.0 ± 0.2 and 8.8 ± 0.1 μM, respectively, in competitive mode, with K(i) values of 8.2 ± 1.1 and 5.8 ± 0.8 μM. Both compounds showed the characteristics of slow-binding inhibitors over the time course of the enzyme reaction. Compound 3 had a single-step binding mechanism and compound 5 a two-step-binding mechanism. With stable AutoDock scores of −6.59 and −6.68 kcal/mol, respectively, compounds 3 and 5 both interacted with His85 and Asn260 at the active site. |
3,767 | Selective and competitive inhibition of kynurenine aminotransferase 2 by glycyrrhizic acid and its analogues | The enzyme kynurenine aminotransferase (KAT) catalyses the conversion of kynurenine (KYN) to kynurenic acid (KYNA). Although the isozymes KAT1–4 have been identified, KYNA is mainly produced by KAT2 in brain tissues. KNYA is an antagonist of N-methyl-D-aspartate and α-7-nicotinic acetylcholine receptors, and accumulation of KYNA in the brain has been associated with the pathology of schizophrenia. Therefore, KAT2 could be exploited as a therapeutic target for the management of schizophrenia. Although currently available KAT2 inhibitors irreversibly bind to pyridoxal 5′-phosphate (PLP), inhibition via this mechanism may cause adverse side effects because of the presence of other PLP-dependent enzymes. Therefore, we identified novel selective KAT2 inhibitors by screening approximately 13,000 molecules. Among these, glycyrrhizic acid (GL) and its analogues, glycyrrhetinic acid (GA) and carbenoxolone (CBX), were identified as KAT2 inhibitors. These compounds were highly selective for KAT2 and competed with its substrate KYN, but had no effects on the other 3 KAT isozymes. Furthermore, we demonstrated that in complex structures that were predicted in docking calculations, GL, GA and CBX were located on the same surface as the aromatic ring of KYN. These results indicate that GL and its analogues are highly selective and competitive inhibitors of KAT2. |
3,768 | Using “outbreak science” to strengthen the use of models during epidemics | Infectious disease modeling has played a prominent role in recent outbreaks, yet integrating these analyses into public health decision-making has been challenging. We recommend establishing ‘outbreak science’ as an inter-disciplinary field to improve applied epidemic modeling. |
3,769 | High basal heat-shock protein expression in bats confers resistance to cellular heat/oxidative stress | Bats, unique among mammals with powered flight, have many species with the longest size-proportionate lifespan of all mammals. Evolutionary adaptations would have been required to survive the elevated body temperatures during flight. Heat shock protein (HSP), highly conserved master regulators of cell stress, expression was examined across tissues and various cell lines in bats. Basal expression level of major HSPs (HSP70 and HSP90) is significantly higher in two different bat species compared to other mammals. This HSP expression could be a bat-unique, key factor to modulate cellular stress and death. Consequently, bat cells survive prolonged heat treatment, along with other stress stimuli, in a HSP-dependent manner, whereas other mammalian cells succumbed. This suggests HSP expression in bats could be an important adaption to intrinsic metabolic stresses like flight and therefore an important model to study stress resilience and longevity in general. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s12192-019-01013-y) contains supplementary material, which is available to authorized users. |
3,770 | Reverse Engineering Provides Insights on the Evolution of Subgroups A to E Avian Sarcoma and Leukosis Virus Receptor Specificity † | The initial step of retrovirus entry—the interaction between the virus envelope glycoprotein trimer and a cellular receptor—is complex, involving multiple, noncontiguous determinants in both proteins that specify receptor choice, binding affinity and the ability to trigger conformational changes in the viral glycoproteins. Despite the complexity of this interaction, retroviruses have the ability to evolve the structure of their envelope glycoproteins to use a different cellular protein as receptors. The highly homologous subgroup A to E Avian Sarcoma and Leukosis Virus (ASLV) glycoproteins belong to the group of class 1 viral fusion proteins with a two-step triggering mechanism that allows experimental access to intermediate structures during the fusion process. We and others have taken advantage of replication-competent ASLVs and exploited genetic selection strategies to force the ASLVs to naturally evolve and acquire envelope glycoprotein mutations to escape the pressure on virus entry and still yield a functional replicating virus. This approach allows for the simultaneous selection of multiple mutations in multiple functional domains of the envelope glycoprotein that may be required to yield a functional virus. Here, we review the ASLV family and experimental system and the reverse engineering approaches used to understand the evolution of ASLV receptor usage. |
3,771 | Investigation of Fugitive Aerosols Released into the Environment during High-Flow Therapy | Background: Nebulised medical aerosols are designed to deliver drugs to the lungs to aid in the treatment of respiratory diseases. However, an unintended consequence is the potential for fugitive emissions during patient treatment, which may pose a risk factor in both clinical and homecare settings. Methods: The current study examined the potential for fugitive emissions, using albuterol sulphate as a tracer aerosol during high-flow therapy. A nasal cannula was connected to a head model or alternatively, a interface was connected to a tracheostomy tube in combination with a simulated adult and paediatric breathing profile. Two aerodynamic particle sizers (APS) recorded time-series aerosol concentrations and size distributions at two different distances relative to the simulated patient. Results: The results showed that the quantity and characteristics of the fugitive emissions were influenced by the interface type, patient type and supplemental gas-flow rate. There was a trend in the adult scenarios; as the flow rate increased, the fugitive emissions and the mass median aerodynamic diameter (MMAD) of the aerosol both decreased. The fugitive emissions were comparable when using the adult breathing profiles for the nasal cannula and tracheostomy interfaces; however, there was a noticeable distinction between the two interfaces when compared for the paediatric breathing profiles. The highest recorded aerosol concentration was 0.370 ± 0.046 mg m(−3) from the tracheostomy interface during simulated paediatric breathing with a gas-flow rate of 20 L/min. The averaged MMAD across all combinations ranged from 1.248 to 1.793 µm by the APS at a distance of 0.8 m away from the patient interface. Conclusions: Overall, the results highlight the potential for secondary inhalation of fugitive emissions released during simulated aerosol treatment with concurrent high-flow therapy. The findings will help in developing policy and best practice for risk mitigation from fugitive emissions. |
3,772 | Biological Function and Application of Picornaviral 2B Protein: A New Target for Antiviral Drug Development | Picornaviruses are associated with acute and chronic diseases. The clinical manifestations of infections are often mild, but infections may also lead to respiratory symptoms, gastroenteritis, myocarditis, meningitis, hepatitis, and poliomyelitis, with serious impacts on human health and economic losses in animal husbandry. Thus far, research on picornaviruses has mainly focused on structural proteins such as VP1, whereas the non-structural protein 2B, which plays vital roles in the life cycle of the viruses and exhibits a viroporin or viroporin-like activity, has been overlooked. Viroporins are viral proteins containing at least one amphipathic α-helical structure, which oligomerizes to form transmembrane hydrophilic pores. In this review, we mainly summarize recent research data on the viroporin or viroporin-like activity of 2B proteins, which affects the biological function of the membrane, regulates cell death, and affects the host immune response. Considering these mechanisms, the potential application of the 2B protein as a candidate target for antiviral drug development is discussed, along with research challenges and prospects toward realizing a novel treatment strategy for picornavirus infections. |
3,773 | Safety and Immunogenicity of a Novel Recombinant Simian Adenovirus ChAdOx2 as a Vectored Vaccine | Adenovirus vectored vaccines are a highly effective strategy to induce cellular immune responses which are particularly effective against intracellular pathogens. Recombinant simian adenovirus vectors were developed to circumvent the limitations imposed by the use of human adenoviruses due to widespread seroprevalence of neutralising antibodies. We have constructed a replication deficient simian adenovirus-vectored vaccine (ChAdOx2) expressing 4 genes from the Mycobacterium avium subspecies paratuberculosis (AhpC, Gsd, p12 and mpa). Safety and T-cell immunogenicity results of the first clinical use of the ChAdOx2 vector are presented here. The trial was conducted using a ‘three-plus-three’ dose escalation study design. We demonstrate the vaccine is safe, well tolerated and immunogenic. |
3,774 | Serological Evidence of Influenza D Virus Circulation Among Cattle and Small Ruminants in France | Influenza D virus (IDV) has first been identified in 2011 in the USA and was shown to mainly circulate in cattle. While IDV is associated with mild respiratory signs, its prevalence is still unknown. In the present study we show that IDV has been circulating throughout France in cattle and small ruminants, with 47.2% and 1.5% seropositivity, respectively. The high prevalence and moderate pathogenicity of IDV in cattle suggest that it may play an initiating role in the bovine respiratory disease complex. |
3,775 | Cytolytic Perforin as an Adjuvant to Enhance the Immunogenicity of DNA Vaccines | DNA vaccines present one of the most cost-effective platforms to develop global vaccines, which have been tested for nearly three decades in preclinical and clinical settings with some success in the clinic. However, one of the major challenges for the development of DNA vaccines is their poor immunogenicity in humans, which has led to refinements in DNA delivery, dosage in prime/boost regimens and the inclusion of adjuvants to enhance their immunogenicity. In this review, we focus on adjuvants that can enhance the immunogenicity of DNA encoded antigens and highlight the development of a novel cytolytic DNA platform encoding a truncated mouse perforin. The application of this innovative DNA technology has considerable potential in the development of effective vaccines. |
3,776 | Seoul Virus Tropism and Pathology in Naturally Infected Feeder Rats | Seoul virus (SEOV) is a zoonotic orthohantavirus carried by black and brown rats, and can cause hemorrhagic fever with renal syndrome in humans. Human cases of SEOV virus infection have most recently been reported in the USA, United Kingdom, France and the Netherlands and were primarily associated with contact with pet rats and feeder rats. Infection of rats results in an asymptomatic but persistent infection. Little is known about the cell tropism of SEOV in its reservoir and most available data is based on experimental infection studies in which rats were inoculated via a route which does not recapitulate virus transmission in nature. Here we report the histopathological analysis of SEOV cell tropism in key target organs following natural infection of a cohort of feeder rats, comprising 19 adults and 11 juveniles. All adult rats in this study were positive for SEOV specific antibodies and viral RNA in their tissues. One juvenile rat was seropositive, but negative in the rRT-PCR. Of the 19 adult rats of which subsequently additional organs were tested, SEOV RNA was detected in all lungs, followed by kidney (79%) and liver (74%). Histopathologic changes associated with SEOV infection were primarily found in the liver, consistent with a pathological diagnosis of a mild hepatitis. In conclusion, natural SEOV infection results in mild inflammation of the liver in the absence of clinical disease. |
3,777 | Current and Novel Approaches in Influenza Management | Influenza is a disease that poses a significant health burden worldwide. Vaccination is the best way to prevent influenza virus infections. However, conventional vaccines are only effective for a short period of time due to the propensity of influenza viruses to undergo antigenic drift and antigenic shift. The efficacy of these vaccines is uncertain from year-to-year due to potential mismatch between the circulating viruses and vaccine strains, and mutations arising due to egg adaptation. Subsequently, the inability to store these vaccines long-term and vaccine shortages are challenges that need to be overcome. Conventional vaccines also have variable efficacies for certain populations, including the young, old, and immunocompromised. This warrants for diverse efficacious vaccine developmental approaches, involving both active and passive immunization. As opposed to active immunization platforms (requiring the use of whole or portions of pathogens as vaccines), the rapidly developing passive immunization involves administration of either pathogen-specific or broadly acting antibodies against a kind or class of pathogens as a treatment to corresponding acute infection. Several antibodies with broadly acting capacities have been discovered that may serve as means to suppress influenza viral infection and allow the process of natural immunity to engage opsonized pathogens whilst boosting immune system by antibody-dependent mechanisms that bridge the innate and adaptive arms. By that; passive immunotherapeutics approach assumes a robust tool that could aid control of influenza viruses. In this review, we comment on some improvements in influenza management and promising vaccine development platforms with an emphasis on the protective capacity of passive immunotherapeutics especially when coupled with the use of antivirals in the management of influenza infection. |
3,778 | Animal Virus Ecology and Evolution Are Shaped by the Virus Host-Body Infiltration and Colonization Pattern | The current classification of animal viruses is largely based on the virus molecular world. Less attention is given to why and how virus fitness results from the success of virus transmission. Virus transmission reflects the infection-shedding-transmission dynamics, and with it, the organ system involvement and other, macroscopic dimensions of the host environment. This study describes the transmission ecology of the world main livestock viruses, 36 in total, a mix of RNA, DNA and retroviruses. Following an iterative process, the viruses are virtually ranked in an outer- to inner-body fashion, by organ system, on ecological grounds. Also portrayed are the shifts in virus host tropism and virus genome. The synthesis of the findings reveals a predictive virus evolution framework, based on the outer- to inner-body changes in the interplay of host environment-transmission modes-organ system involvement-host cell infection cycle-virus genome. Outer-body viruses opportunistically respond to the variation in the external environment. For example, respiratory and enteric viruses tend to be associated with poultry and pig mass rearing. Ruminant and equine viruses tend to be more deep-rooted and host-specific, and also establish themselves in the vital inner-body systems. It is concluded that the framework may assist the study of new emerging viruses and pandemic risks. |
3,779 | Computational Approaches and Challenges to Developing Universal Influenza Vaccines | The traditional design of effective vaccines for rapidly-evolving pathogens, such as influenza A virus, has failed to provide broad spectrum and long-lasting protection. With low cost whole genome sequencing technology and powerful computing capabilities, novel computational approaches have demonstrated the potential to facilitate the design of a universal influenza vaccine. However, few studies have integrated computational optimization in the design and discovery of new vaccines. Understanding the potential of computational vaccine design is necessary before these approaches can be implemented on a broad scale. This review summarizes some promising computational approaches under current development, including computationally optimized broadly reactive antigens with consensus sequences, phylogenetic model-based ancestral sequence reconstruction, and immunomics to compute conserved cross-reactive T-cell epitopes. Interactions between virus-host-environment determine the evolvability of the influenza population. We propose that with the development of novel technologies that allow the integration of data sources such as protein structural modeling, host antibody repertoire analysis and advanced phylodynamic modeling, computational approaches will be crucial for the development of a long-lasting universal influenza vaccine. Taken together, computational approaches are powerful and promising tools for the development of a universal influenza vaccine with durable and broad protection. |
3,780 | Novel Picobirnaviruses in Respiratory and Alimentary Tracts of Cattle and Monkeys with Large Intra- and Inter-Host Diversity | Picobirnaviruses (PBVs) are mostly found in animal alimentary samples. In this study, among 576 respiratory specimens from 476 mammals and 100 chickens, genogroup I PBVs were detected in three cattle and three monkeys, and a genogroup II PBV-positive sample was collected from one cattle specimen. More than one PBV sequence type was observed in two and one genogroup I PBV-positive samples from cattle and monkeys, respectively. Twenty-four complete/near-complete segments 2 (nine from respiratory and 15 from alimentary samples) from the cattle and monkey genogroup I PBVs and one complete segment 2 from the cattle genogroup II PBV were sequenced. Similar to other studies, the cattle PBVs also showed a high diversity. In contrast, the monkey PBVs observed in this study were clustered into three distinct clades. Within each clade, all the sequences showed >99% amino acid identities. This unique phenomenon is probably due to the fact that monkeys in our locality reside in separated troops with minimal inter-troop contact. |
3,781 | A Comparison of Plasmid DNA and mRNA as Vaccine Technologies | This review provides a comparison of the theoretical issues and experimental findings for plasmid DNA and mRNA vaccine technologies. While both have been under development since the 1990s, in recent years, significant excitement has turned to mRNA despite the licensure of several veterinary DNA vaccines. Both have required efforts to increase their potency either via manipulating the plasmid DNA and the mRNA directly or through the addition of adjuvants or immunomodulators as well as delivery systems and formulations. The greater inherent inflammatory nature of the mRNA vaccines is discussed for both its potential immunological utility for vaccines and for the potential toxicity. The status of the clinical trials of mRNA vaccines is described along with a comparison to DNA vaccines, specifically the immunogenicity of both licensed veterinary DNA vaccines and select DNA vaccine candidates in human clinical trials. |
3,782 | Recent Advances in Droplet-based Microfluidic Technologies for Biochemistry and Molecular Biology | Recently, droplet-based microfluidic systems have been widely used in various biochemical and molecular biological assays. Since this platform technique allows manipulation of large amounts of data and also provides absolute accuracy in comparison to conventional bioanalytical approaches, over the last decade a range of basic biochemical and molecular biological operations have been transferred to drop-based microfluidic formats. In this review, we introduce recent advances and examples of droplet-based microfluidic techniques that have been applied in biochemistry and molecular biology research including genomics, proteomics and cellomics. Their advantages and weaknesses in various applications are also comprehensively discussed here. The purpose of this review is to provide a new point of view and current status in droplet-based microfluidics to biochemists and molecular biologists. We hope that this review will accelerate communications between researchers who are working in droplet-based microfluidics, biochemistry and molecular biology. |
3,783 | IFITM3 Clusters on Virus Containing Endosomes and Lysosomes Early in the Influenza A Infection of Human Airway Epithelial Cells | Interferon-induced transmembrane proteins (IFITMs) have been shown to strongly affect influenza A virus (IAV) infectivity in tissue culture. Moreover, polymorphisms in IFITM3 have been associated with the severity of the disease in humans. IFITM3 appears to act early in the infection, but its mechanism of action and potential interactions with incoming IAV structures are not yet defined. Here, we visualized endogenous IFITM3 interactions with IAV in the human lung epithelial cell line A549 and in primary human airway epithelial cells employing stimulated emission depletion super-resolution microscopy. By applying an iterative approach for the cluster definition and computational cluster analysis, we found that IFITM3 reorganizes into clusters as IAV infection progresses. IFITM3 cluster formation started at 2-3 h post infection and increased over time to finally coat IAV-containing endosomal vesicles. This IAV-induced phenotype was due to the endosomal recruitment of IFITM3 rather than to an overall increase in the IFITM3 abundance. While the IAV-induced IFITM3 clustering and localization to endosomal vesicles was comparable in primary human airway epithelial cells and the human lung epithelial cell line A549, the endogenous IFITM3 signal was higher in primary cells. Moreover, we observed IFITM3 signals adjacent to IAV-containing recycling endosomes. |
3,784 | Porcine Interferon Complex and Co-Evolution with Increasing Viral Pressure after Domestication | Consisting of nearly 60 functional genes, porcine interferon (IFN)-complex represents an evolutionary surge of IFN evolution in domestic ungulate species. To compare with humans and mice, each of these species contains about 20 IFN functional genes, which are better characterized using the conventional IFN-α/β subtypes as examples. Porcine IFN-complex thus represents an optimal model for studying IFN evolution that resulted from increasing viral pressure during domestication and industrialization. We hypothesize and justify that porcine IFN-complex may extend its functionality in antiviral and immunomodulatory activity due to its superior molecular diversity. Furthermore, these unconventional IFNs could even confer some functional and signaling novelty beyond that of the well-studied IFN-α/β subtypes. Investigations into porcine IFN-complex will further our understanding of IFN biology and promote IFN-based therapeutic designs to confront swine viral diseases. |
3,785 | Assessing the Potential Interactions between Cellular miRNA and Arboviral Genomic RNA in the Yellow Fever Mosquito, Aedes aegypti | Although the role of exogenous small interfering RNA (siRNA) and P-element induced wimpy testis (PIWI)-interacting RNA (piRNA) pathways in mosquito antiviral immunity is increasingly better understood, there is still little knowledge regarding the role of mosquito cellular microRNA (miRNA). Identifying direct interactions between the mosquito miRNAs and the RNA genome of arboviruses and choosing the relevant miRNA candidates to explore resulting antiviral mechanisms are critical. Here, we carried out genomic analyses to identify Aedes aegypti miRNAs that potentially interact with various lineages and genotypes of chikungunya, dengue, and Zika viruses. By using prediction tools with distinct algorithms, several miRNA binding sites were commonly found within different genotypes/and or lineages of each arbovirus. We further analyzed those miRNAs that could target more than one arbovirus, required a low energy threshold to form miRNA-viralRNA (vRNA) complexes, and predicted potential RNA structures using RNAhybrid software. We predicted miRNA candidates that might participate in regulating arboviral replication in Ae. aegypti. Even without any experimental validation, which should be done as a next step, this study can shed further light on the role of miRNA in mosquito innate immunity and targets for future studies. |
3,786 | Hypsugopoxvirus: A Novel Poxvirus Isolated from Hypsugo savii in Italy | Interest in bat-related viruses has increased considerably during the last decade, leading to the discovery of a rising number of new viruses in several bat species. Poxviridae are a large, diverse family of DNA viruses that can infect a wide range of vertebrates and invertebrates. To date, only a few documented detections of poxviruses have been described in bat populations on three different continents (America, Africa, and Australia). These viruses are phylogenetically dissimilar and have diverse clinical impacts on their hosts. Herein, we report the isolation, nearly complete genome sequencing, and annotation of a novel poxvirus detected from an insectivorous bat (Hypsugo savii) in Northern Italy. The virus is tentatively named Hypsugopoxvirus (HYPV) after the bat species from which it was isolated. The nearly complete genome size is 166,600 nt and it encodes 161 genes. Genome analyses suggest that HYPV belongs to the Chordopoxvirinae subfamily, with the highest nucleotide identity (85%) to Eptesipoxvirus (EPTV) detected from a microbat Eptesicus fuscus in WA, USA, in 2011. To date, HYPV represents the first poxvirus detected in bats in Europe; thus, its viral ecology and disease associations should be investigated further. |
3,787 | Novel Synthetic DNA Immunogens Targeting Latent Expressed Antigens of Epstein–Barr Virus Elicit Potent Cellular Responses and Inhibit Tumor Growth | Infectious diseases are linked to 15%–20% of cancers worldwide. Among them, Epstein–Barr virus (EBV) is an oncogenic herpesvirus that chronically infects over 90% of the adult population, with over 200,000 cases of cancer and 150,000 cancer-related deaths attributed to it yearly. Acute EBV infection can present as infectious mononucleosis, and lead to the future onset of multiple cancers, including Burkitt lymphoma, Hodgkin lymphoma, nasopharyngeal carcinoma, and gastric carcinoma. Many of these cancers express latent viral genes, including Epstein–Barr virus nuclear antigen 1 (EBNA1) and latent membrane proteins 1 and 2 (LMP1 and LMP2). Previous attempts to create potent immunogens against EBV have been reported but generated mixed success. We designed novel Synthetic Consensus (SynCon) DNA vaccines against EBNA1, LMP1 and LMP2 to improve on the immune potency targeting important antigens expressed in latently infected cells. These EBV tumor antigens are hypothesized to be useful targets for potential immunotherapy of EBV-driven cancers. We optimized the genetic sequences for these three antigens, studied them for expression, and examined their immune profiles in vivo. We observed that these immunogens generated unique profiles based on which antigen was delivered as the vaccine target. EBNA1vax and LMP2Avax generated the most robust T cell immunity. Interestingly, LMP1vax was a very weak immunogen, generating very low levels of CD8 T cell immunity both as a standalone vaccine and as part of a trivalent vaccine cocktail. LMP2Avax was able to drive immunity that impacted EBV-antigen-positive tumor growth. These studies suggest that engineered EBV latent protein vaccines deserve additional study as potential agents for immunotherapy of EBV-driven cancers. |
3,788 | The microbiota protects from viral-induced neurologic damage through microglia-intrinsic TLR signaling | Symbiotic microbes impact the function and development of the central nervous system (CNS); however, little is known about the contribution of the microbiota during viral-induced neurologic damage. We identify that commensals aid in host defense following infection with a neurotropic virus through enhancing microglia function. Germfree mice or animals that receive antibiotics are unable to control viral replication within the brain leading to increased paralysis. Microglia derived from germfree or antibiotic-treated animals cannot stimulate viral-specific immunity and microglia depletion leads to worsened demyelination. Oral administration of toll-like receptor (TLR) ligands to virally infected germfree mice limits neurologic damage. Homeostatic activation of microglia is dependent on intrinsic signaling through TLR4, as disruption of TLR4 within microglia, but not the entire CNS (excluding microglia), leads to increased viral-induced clinical disease. This work demonstrates that gut immune-stimulatory products can influence microglia function to prevent CNS damage following viral infection. |
3,789 | Bioinformatic Study of Transcriptome Changes in the Mice Lumbar Spinal Cord After the 30-Day Spaceflight and Subsequent 7-Day Readaptation on Earth: New Insights Into Molecular Mechanisms of the Hypogravity Motor Syndrome | The hypogravity motor syndrome (HMS) is one of the deleterious impacts of weightlessness on the human body in orbital space missions. There is a hypothesis that disorders of musculoskeletal system as part of HMS arise in consequence of changes in spinal motor neurons. The study was aimed at bioinformatic analysis of transcriptome changes in lumbar spinal cords of mice after a 30-day spaceflight aboard biosatellite Bion-M1 (space group, S) and subsequent 7-day readaptation to the Earth’s gravity (recovery group, R) when compared with control mice (C group) housed in simulated biosatellite conditions on the Earth. Gene ontology and human phenotype ontology databases were used to detect biological processes, molecular functions, cellular components, and human phenotypes associated with HMS. Our results suggest resemblance of molecular changes developing in space orbit and during the postflight recovery to terrestrial neuromuscular disorders. Remarkably, more prominent transcriptome changes were revealed in R vs. S and R vs. C comparisons that are possibly related to the 7-day recovery period in the Earth’s gravity condition. These data may assist with establishment of HMS pathogenesis and proposing effective preventive and therapeutic options. |
3,790 | Identification of Ebola Virus Inhibitors Targeting GP2 Using Principles of Molecular Mimicry | A key step in the Ebola virus (EBOV) replication cycle involves conformational changes in viral glycoprotein 2 (GP2) which facilitate host-viral membrane fusion and subsequent release of the viral genome. Ebola GP2 plays a critical role in virus entry and has similarities in mechanism and structure to the HIV gp41 protein for which inhibitors have been successfully developed. In this work, a putative binding pocket for the C-terminal heptad repeat in the N-terminal heptad repeat trimer was targeted for identification of small molecules that arrest EBOV-host membrane fusion. Two computational structure-based virtual screens of ∼1.7 M compounds were performed (DOCK program) against a GP2 five-helix bundle, resulting in 165 commercially available compounds purchased for experimental testing. Based on assessment of inhibitory activity, cytotoxicity, and target specificity, four promising candidates emerged with 50% inhibitory concentration values in the 3 to 26 μM range. Molecular dynamics simulations of the two most potent candidates in their DOCK-predicted binding poses indicate that the majority of favorable interactions involve seven highly conserved residues that can be used to guide further inhibitor development and refinement targeting EBOV. IMPORTANCE The most recent Ebola virus disease outbreak, from 2014 to 2016, resulted in approximately 28,000 individuals becoming infected, which led to over 12,000 causalities worldwide. The particularly high pathogenicity of the virus makes paramount the identification and development of promising lead compounds to serve as inhibitors of Ebola infection. To limit viral load, the virus-host membrane fusion event can be targeted through the inhibition of the class I fusion glycoprotein of Ebolavirus. In the current work, several promising small-molecule inhibitors that target the glycoprotein GP2 were identified through systematic application of structure-based computational and experimental drug design procedures. |
3,791 | Avian Influenza A (H7N9) and related Internet search query data in China | The use of Internet-based systems for infectious disease surveillance has been increasingly explored in recent years. However, few studies have used Internet search query or social media data to monitor spatial and temporal trends of avian influenza in China. This study investigated the potential of using search query and social media data in detecting and monitoring avian influenza A (H7N9) cases in humans in China. We collected weekly data on laboratory-confirmed H7N9 cases in humans, as well as H7N9-related Baidu Search Index (BSI) and Weibo Posting Index (WPI) data in China from 2013 to 2017, to explore the spatial and temporal trends of H7N9 cases and H7N9-related Internet search queries. Our findings showed a positive relationship of H7N9 cases with BSI and WPI search queries spatially and temporally. The outbreak threshold time and peak time of H7N9-related BSI and WPI searches preceded H7N9 cases in most years. Seasonal autoregressive integrated moving average (SARIMA) models with BSI (β = 0.008, p < 0.001) and WPI (β = 0.002, p = 0.036) were used to predict the number of H7N9 cases. Regression tree model analysis showed that the average H7N9 cases increased by over 2.4-fold (26.8/11) when BSI for H7N9 was > = 11524. Both BSI and WPI data could be used as indicators to develop an early warning system for H7N9 outbreaks in the future. |
3,792 | Plasma and tissue angiotensin‐converting enzyme 2 activity and plasma equilibrium concentrations of angiotensin peptides in dogs with heart disease | BACKGROUND: Angiotensin‐converting enzyme 2 (ACE2) is a homologue of angiotensin‐converting enzyme (ACE) and produces angiotensin peptides (APs), such as angiotensin 1‐9 and 1‐7 that are vasodilatory and natriuretic, and act to counterbalance angiotensin II. HYPOTHESIS: Evidence of ACE2 can be found in tissues and plasma of dogs. Equilibrium concentrations of renin angiotensin aldosterone system (RAAS) APs differ in dogs with heart disease compared to healthy dogs and recombinant human ACE2 (rhACE2) alters relative concentrations of APs. ANIMALS: Forty‐nine dogs with and 34 dogs without heart disease. METHODS: Immunohistochemistry and assays for tissue and plasma ACE2 activity and equilibrium concentrations of plasma RAAS APs were performed. RESULTS: Immunolabeling for ACE2 was present in kidney and myocardial tissue. Median plasma ACE2 activity was significantly increased in dogs with congestive heart failure (CHF; 6.9 mU/mg; interquartile range [IQR], 5.1‐12.1) as compared to control (2.2 mU/mg; IQR, 1.8‐3.0; P = .0003). Plasma equilibrium analysis of RAAS APs identified significant increases in the median concentrations of beneficial APs, such as angiotensin 1‐7, in dogs with CHF (486.7 pg/mL; IQR, 214.2‐1168) as compared to those with preclinical disease (41.0 pg/mL; IQR, 27.4‐45.1; P < .0001) or control (11.4 pg/mL; IQR, 7.1‐25.3; P = .01). Incubation of plasma samples from dogs with CHF with rhACE2 increased beneficial APs, such as angiotensin 1‐9 (preincubation, 10.3 pg/mL; IQR, 4.4‐37.2; postincubation, 2431 pg/mL; IQR, 1355‐3037; P = .02), while simultaneously decreasing maladaptive APs, such as angiotensin II (preincubation, 53.4 pg/mL; IQR, 28.6‐226.4; postincubation, 2.4 pg/mL; IQR, 0.50‐5.8; P = .02). CONCLUSIONS AND CLINICAL IMPORTANCE: Recognition of the ACE2 system expands the conventional view of the RAAS in the dog and represents an important potential therapeutic target. |
3,793 | Exploratory cohort study to determine if dry cow vaccination with a Salmonella Newport bacterin can protect dairy calves against oral Salmonella challenge | BACKGROUND: Salmonellosis is a major cause of morbidity and mortality in neonatal calves, often occurring before preventative vaccines can be administered. HYPOTHESIS/OBJECTIVE: To evaluate the protective effect on calves of colostrum from cows vaccinated with a commercially available Salmonella Newport bacterin against a Salmonella Typhimurium challenge. ANIMALS: Twenty Holstein bull calves from a university dairy farm. METHODS: Nonrandomized placebo‐controlled trial in which colostrum was harvested from 30 cows that received 2 doses of either Salmonella bacterin or saline before calving. Colostrum collected from each group was pooled and fed to 2 groups of 10 calves at birth. At approximately 2 weeks of age, calves were challenged with Salmonella Typhimurium. Clinical, hematologic, microbiological, and postmortem findings were compared between the 2 groups. RESULTS: No differences in mortality, clinical findings, hematology results, blood and fecal cultures, or necropsy findings between the 2 groups were observed. Vaccinated cows had higher colostral titers, and calves fed this colostrum had higher serum titers (mean difference, 0.429; mean [SE], 0.852 [0.02] for vaccinated versus 0.423 [0.02] for control calves). CONCLUSIONS AND CLINICAL IMPORTANCE: Transfer of colostral immunoglobulins from Salmonella enterica serotype Newport bacterin to neonatal calves was not sufficient to decrease mortality, clinical signs, sepsis, intestinal damage, or fecal shedding when exposed to a highly pathogenic Salmonella isolate. A large‐scale randomized controlled clinical trial is needed to evaluate the efficacy of this bacterin when administered in the dry period for prevention of salmonellosis in neonatal calves. |
3,794 | Discovery of All Three Types in Cartilaginous Fishes Enables Phylogenetic Resolution of the Origins and Evolution of Interferons | Interferons orchestrate host antiviral responses in jawed vertebrates. They are categorized into three classes; IFN1 and IFN3 are the primary antiviral cytokine lineages, while IFN2 responds to a broader variety of pathogens. The evolutionary relationships within and between these three classes have proven difficult to resolve. Here, we reassess interferon evolution, considering key phylogenetic pitfalls including taxon sampling, alignment quality, model adequacy, and outgroup choice. We reveal that cartilaginous fishes, and hence the jawed vertebrate ancestor, possess(ed) orthologs of all three interferon classes. We show that IFN3 groups sister to IFN1, resolve the origins of the human IFN3 lineages, and find that intronless IFN3s emerged at least three times. IFN2 genes are highly conserved, except for IFN-γ-rel, which we confirm resulted from a teleost-specific duplication. Our analyses show that IFN1 phylogeny is highly sensitive to phylogenetic error. By accounting for this, we describe a new backbone IFN1 phylogeny that implies several IFN1 genes existed in the jawed vertebrate ancestor. One of these is represented by the intronless IFN1s of tetrapods, including mammalian-like repertoires of reptile IFN1s and a subset of amphibian IFN1s, in addition to newly-identified intron-containing shark IFN1 genes. IFN-f, previously only found in teleosts, likely represents another ancestral jawed vertebrate IFN1 family member, suggesting the current classification of fish IFN1s into two groups based on the number of cysteines may need revision. The providence of the remaining fish IFN1s and the coelacanth IFN1s proved difficult to resolve, but they may also be ancestral jawed vertebrate IFN1 lineages. Finally, a large group of amphibian-specific IFN1s falls sister to all other IFN1s and was likely also present in the jawed vertebrate ancestor. Our results verify that intronless IFN1s have evolved multiple times in amphibians and indicate that no one-to-one orthology exists between mammal and reptile IFN1s. Our data also imply that diversification of the multiple IFN1s present in the jawed vertebrate ancestor has occurred through a rapid birth-death process, consistent with functional maintenance over a 450-million-year host-pathogen arms race. In summary, this study reveals a new model of interferon evolution important to our understanding of jawed vertebrate antiviral immunity. |
3,795 | Exosomes Derived From Septic Mouse Serum Modulate Immune Responses via Exosome-Associated Cytokines | Sepsis is a life-threatening condition caused by an immune response triggered by infection, and highly elevated cytokine/chemokine levels in the blood play crucial roles in the progression of sepsis. Serum exosomes are nanovesicles that have multiple biological functions, playing roles in antigen presentation, intercellular signal communication, inflammatory response and immune surveillance. However, the biological functions and related molecular bases remain to be elucidated. In this study, we investigated the profiles of cytokines/chemokines harbored in the exosomes of septic mice and explored the mechanisms of immunomodulation on T cells treated with exosomes harvested from septic mice. Blood cytokines/chemokines existed in both the soluble form and in the insoluble exosomal form; the profiles of the cytokines/chemokines in these two forms displayed different dynamics in the blood of mice challenged with LPS. Exosomes from septic mice induced the differentiation of Th1/Th2 cells, which was blocked by specific antibodies targeting IL-12 and IL-4. In addition, these exosomes significantly augmented the proliferation and migration of T lymphocytes. Furthermore, preadministration of exosomes by intravenous injection restrained the inflammatory response, attenuated lung and liver tissue damage, and prolonged the survival of cecal ligation and puncture (CLP) mice. Our results indicate that exosomes enriched with cytokines/chemokines play critical roles in T cell differentiation, proliferation and chemotaxis during the sepsis process and have a protective effect on cecal ligation and puncture (CLP) mice. Thus, these findings not only strengthen our understanding of the role of sepsis via exosomes but also provide potential targets for therapeutic applications. |
3,796 | Peptide-conjugate antisense based splice-correction for Duchenne muscular dystrophy and other neuromuscular diseases | Duchenne muscular dystrophy (DMD) is an X-linked disorder characterized by progressive muscle degeneration, caused by the absence of dystrophin. Exon skipping by antisense oligonucleotides (ASOs) has recently gained recognition as therapeutic approach in DMD. Conjugation of a peptide to the phosphorodiamidate morpholino backbone (PMO) of ASOs generated the peptide-conjugated PMOs (PPMOs) that exhibit a dramatically improved pharmacokinetic profile. When tested in animal models, PPMOs demonstrate effective exon skipping in target muscles and prolonged duration of dystrophin restoration after a treatment regime. Herein we summarize the main pathophysiological features of DMD and the emergence of PPMOs as promising exon skipping agents aiming to rescue defective gene expression in DMD and other neuromuscular diseases. The listed PPMO laboratory findings correspond to latest trends in the field and highlight the obstacles that must be overcome prior to translating the animal-based research into clinical trials tailored to the needs of patients suffering from neuromuscular diseases. |
3,797 | Highly Efficient and Practical N-Heterocyclic Carbene Organocatalyzed Chemoselective N(1)/C(3)-Functionalization of Isatins with Green Chemistry Principles | [Image: see text] Ecofriendly N-heterocyclic carbene (NHC) organocatalysis can control the N(1)-functionalization (aza-Michael addition) and C(3)-functionalization (Morita–Baylis–Hillman reaction, MBH) of isatins in the absence of (1) a protecting group, (2) a stoichiometric reagent, and (3) heat energy. The challengeable N(1)-functionalization of N-unsubstituted isatins into N-substituted (NS) isatins was realized through 10 mol % NHC and 10 mol % 1,8-diazabicyclo[5.4.0]undec-7-ene catalysts within 10 min with up to 98% isolation yield. The subsequent MBH adducts of as-synthesized NS-isatins (N(1)/C(3)-functionalization) was perfectly acquired in 10 mol % NHC and 10 mol % 1,4-diazabicyclo[2.2.2]octane catalysis within 30 min with superiority to C(3)/N(1)-functionalization (MBH/aza-Michael). For guiding the application to a versatile druggable isatin library, the NHC catalysis was compared with reported functionalization of isatins in view of green chemistry principles including solvent scoring of ACS GCI pharmaceutical roundtable, E-factor, atom economy, and so on. |
3,798 | Hepatitis E in southern Vietnam: Seroepidemiology in humans and molecular epidemiology in pigs | Viral pathogens account for a significant proportion of the burden of emerging infectious diseases in humans. The Wellcome Trust-Vietnamese Initiative on Zoonotic Infections (WT-VIZIONS) is aiming to understand the circulation of viral zoonotic pathogens in animals that pose a potential risk to human health. Evidence suggests that human exposure and infections with hepatitis E virus (HEV) genotypes (GT) 3 and 4 results from zoonotic transmission. Hypothesising that HEV GT3 and GT4 are circulating in the Vietnamese pig population and can be transmitted to humans, we aimed to estimate the seroprevalence of HEV exposure in a population of farmers and the general population. We additionally performed sequence analysis of HEV in pig populations in the same region to address knowledge gaps regarding HEV circulation and to evaluate if pigs were a potential source of HEV exposure. We found a high prevalence of HEV GT3 viral RNA in pigs (19.1% in faecal samples and 8.2% in rectal swabs) and a high HEV seroprevalence in pig farmers (16.0%) and a hospital-attending population (31.7%) in southern Vietnam. The hospital population was recruited as a general-population proxy even though this particular population subgroup may introduce bias. The detection of HEV RNA in pigs indicates that HEV may be a zoonotic disease risk in this location, although a larger sample size is required to infer an association between HEV positivity in pigs and seroprevalence in humans. |
3,799 | Haploidentical Stem Cell Transplantation in Children with Benign Disorders: Improved Survival and Cost-Effective Care Over 15 Years from a Single Center in India | We present our experience in haploidentical stem cell transplantation (haplo SCT) in children with benign disorders. We performed a retrospective study where children aged up to 18 years diagnosed to have benign disorders and underwent haplo SCT from 2002 to September 2017 were included. Of the 54 children, the most common indications were Fanconi anaemia 12 (22%), severe aplastic anaemia 8 (14%) and primary immune deficiency disorders (PID) 25 (46%). Post-transplant cyclophosphamide (PTCy) was used in 41 (75.9%) and ex vivo T depletion in 13 (24.1%). Engraftment rates were 70% with acute graft versus host disease in 36% and cytomegalovirus reactivation in 55% children. There was a statistically significant difference found between survival with siblings as donors as compared to parents (p value 0.018). Overall survival was 60% which is the 1-year survival, with 68% survival among those with PIDs. Cytokine release syndrome was noted in 12/41 (29%) of children who received T replete graft and PTCy. In children over 6 months of age, PTCy at a cost of INR 1200 provides cost effective T cell depletion comparable with TCR α/β depletion priced at INR 1200,000. Haplo SCT is feasible option for cure in children with benign disorder. |
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