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The accessory protease transmembrane protease serine 2 (TMPRSS2) enhances severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) uptake into ACE2-expressing cells, although how increased entry impacts downstream viral and host processes remains unclear. To investigate this in more detail, we performed infection assays in engineered cells promoting ACE2-mediated entry with and without TMPRSS2 coexpression. Electron microscopy and inhibitor experiments indicated TMPRSS2-mediated cell entry was associated with increased virion internalization into endosomes, and partially dependent upon clathrin-mediated endocytosis. TMPRSS2 increased panvariant uptake efficiency and enhanced early rates of virus replication, transcription, and secretion, with variant-specific profiles observed. On the host side, transcriptional profiling confirmed the magnitude of infection-induced antiviral and proinflammatory responses were linked to uptake efficiency, with TMPRSS2-assisted entry boosting early antiviral responses. In addition, TMPRSS2-enhanced infections increased rates of cytopathology, apoptosis, and necrosis and modulated virus secretion kinetics in a variant-specific manner. On the virus side, convergent signatures of cell-uptake-dependent innate immune induction were recorded in viral genomes, manifesting as switches in dominant coupled Nsp3 residues whose frequencies were correlated to the magnitude of the cellular response to infection. Experimentally, we demonstrated that selected Nsp3 mutations conferred enhanced interferon antagonism. More broadly, we show that TMPRSS2 orthologues from evolutionarily diverse mammals facilitate panvariant enhancement of cell uptake. In summary, our study uncovers previously unreported associations, linking cell entry efficiency to innate immune activation kinetics, cell death rates, virus secretion dynamics, and convergent selection of viral mutations. These data expand our understanding of TMPRSS2's role in the SARS-CoV-2 life cycle and confirm its broader significance in zoonotic reservoirs and animal models.
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COVID-19 , Imunidade Inata , SARS-CoV-2 , Serina Endopeptidases , Internalização do Vírus , SARS-CoV-2/imunologia , SARS-CoV-2/fisiologia , SARS-CoV-2/metabolismo , Humanos , Serina Endopeptidases/metabolismo , Serina Endopeptidases/genética , COVID-19/virologia , COVID-19/imunologia , COVID-19/metabolismo , Enzima de Conversão de Angiotensina 2/metabolismo , Enzima de Conversão de Angiotensina 2/genética , Replicação Viral , Animais , Endocitose , Células HEK293 , Chlorocebus aethiops , CitologiaRESUMO
Hepatitis C virus (HCV) infection progresses to chronicity in the majority of infected individuals. Its high intra-host genetic variability enables HCV to evade the continuous selection pressure exerted by the host, contributing to persistent infection. Utilizing a cell culture-adapted HCV population (p100pop) which exhibits increased replicative capacity in various liver cell lines, this study investigated virus and host determinants that underlie enhanced viral fitness. Characterization of a panel of molecular p100 clones revealed that cell culture adaptive mutations optimize a range of virus-host interactions, resulting in expanded cell tropism, altered dependence on the cellular co-factor micro-RNA 122 and increased rates of virus spread. On the host side, comparative transcriptional profiling of hepatoma cells infected either with p100pop or its progenitor virus revealed that enhanced replicative fitness correlated with activation of endoplasmic reticulum stress signaling and the unfolded protein response. In contrast, infection of primary human hepatocytes with p100pop led to a mild attenuation of virion production which correlated with a greater induction of cell-intrinsic antiviral defense responses. In summary, long-term passage experiments in cells where selective pressure from innate immunity is lacking improves multiple virus-host interactions, enhancing HCV replicative fitness. However, this study further indicates that HCV has evolved to replicate at low levels in primary human hepatocytes to minimize innate immune activation, highlighting that an optimal balance between replicative fitness and innate immune induction is key to establish persistence. IMPORTANCE: Hepatitis C virus (HCV) infection remains a global health burden with 58 million people currently chronically infected. However, the detailed molecular mechanisms that underly persistence are incompletely defined. We utilized a long-term cell culture-adapted HCV, exhibiting enhanced replicative fitness in different human liver cell lines, in order to identify molecular principles by which HCV optimizes its replication fitness. Our experimental data revealed that cell culture adaptive mutations confer changes in the host response and usage of various host factors. The latter allows functional flexibility at different stages of the viral replication cycle. However, increased replicative fitness resulted in an increased activation of the innate immune system, which likely poses boundary for functional variation in authentic hepatocytes, explaining the observed attenuation of the adapted virus population in primary hepatocytes.
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Aptidão Genética , Hepacivirus , Hepatócitos , Interações entre Hospedeiro e Microrganismos , Imunidade Inata , Mutação , Humanos , Células Cultivadas , Estresse do Retículo Endoplasmático , Aptidão Genética/genética , Aptidão Genética/imunologia , Hepacivirus/genética , Hepacivirus/crescimento & desenvolvimento , Hepacivirus/imunologia , Hepacivirus/fisiologia , Hepatite C/imunologia , Hepatite C/virologia , Hepatócitos/imunologia , Hepatócitos/virologia , Interações entre Hospedeiro e Microrganismos/imunologia , MicroRNAs/metabolismo , Inoculações Seriadas , Resposta a Proteínas não Dobradas , Tropismo Viral , Vírion/crescimento & desenvolvimento , Vírion/metabolismo , Replicação Viral/genética , Replicação Viral/imunologiaRESUMO
BACKGROUND AND AIMS: HEV is estimated to be responsible for 70,000 deaths annually, yet therapy options remain limited. In the pursuit of effective antiviral therapies, targeting viral entry holds promise and has proven effective for other viruses. However, the precise mechanisms and host factors required during HEV entry remain unclear. Cellular proteases have emerged as host factors required for viral surface protein activation and productive cell entry by many viruses. Hence, we investigated the functional requirement and therapeutic potential of cellular protease during HEV infection. APPROACH AND RESULTS: Using our established HEV cell culture model and subgenomic HEV replicons, we found that blocking lysosomal cathepsins (CTS) with small molecule inhibitors impedes HEV infection without affecting replication. Most importantly, the pan-cathepsin inhibitor K11777 suppressed HEV infections with an EC 50 of ~0.02 nM. Inhibition by K11777, devoid of notable toxicity in hepatoma cells, was also observed in HepaRG and primary human hepatocytes. Furthermore, through time-of-addition and RNAscope experiments, we confirmed that HEV entry is blocked by inhibition of cathepsins. Cathepsin L (CTSL) knockout cells were less permissive to HEV, suggesting that CTSL is critical for HEV infection. Finally, we observed cleavage of the glycosylated ORF2 protein and virus particles by recombinant CTSL. CONCLUSIONS: In summary, our study highlights the pivotal role of lysosomal cathepsins, especially CTSL, in the HEV entry process. The profound anti-HEV efficacy of the pan-cathepsin inhibitor K11777, especially with its notable safety profile in primary cells, further underscores its potential as a therapeutic candidate.
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Libraries composed of licensed drugs represent a vast repertoire of molecules modulating physiological processes in humans, providing unique opportunities for the discovery of host-targeting antivirals. We screened the Repurposing, Focused Rescue, and Accelerated Medchem (ReFRAME) repurposing library with approximately 12,000 molecules for broad-spectrum coronavirus antivirals and discovered 134 compounds inhibiting an alphacoronavirus and mapping to 58 molecular target categories. Dominant targets included the 5-hydroxytryptamine receptor, the dopamine receptor, and cyclin-dependent kinases. Gene knock-out of the drugs' host targets including cathepsin B and L (CTSB/L; VBY-825), the aryl hydrocarbon receptor (AHR; Phortress), the farnesyl-diphosphate farnesyltransferase 1 (FDFT1; P-3622), and the kelch-like ECH-associated protein 1 (KEAP1; Omaveloxolone), significantly modulated HCoV-229E infection, providing evidence that these compounds inhibited the virus through acting on their respective host targets. Counter-screening of all 134 primary compound candidates with SARS-CoV-2 and validation in primary cells identified Phortress, an AHR activating ligand, P-3622-targeting FDFT1, and Omaveloxolone, which activates the NFE2-like bZIP transcription factor 2 (NFE2L2) by liberating it from its endogenous inhibitor KEAP1, as antiviral candidates for both an Alpha- and a Betacoronavirus. This study provides an overview of HCoV-229E repurposing candidates and reveals novel potentially druggable viral host dependency factors hijacked by diverse coronaviruses.
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Coronavirus Humano 229E , Infecções por Coronavirus , Tiazóis , Triterpenos , Humanos , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Reposicionamento de Medicamentos , Fator 2 Relacionado a NF-E2/metabolismo , Coronavirus Humano 229E/metabolismo , Antivirais/farmacologia , Antivirais/uso terapêuticoRESUMO
BACKGROUND AND AIMS: Severe acute respiratory syndrome coronavirus (SARS-CoV-2) preferentially infects the respiratory tract; however, several studies have implicated a multi-organ involvement. Hepatic dysfunctions caused by SARS-CoV-2 infection have been increasingly recognized and described to correlate with disease severity. To elucidate molecular factors that could contribute towards hepatic infection, we concentrated on microRNAs (miRNAs), a class of small non-coding RNAs that modulate various cellular processes and which are reported to be differentially regulated during liver injury. We aimed to study the infection of primary human hepatocytes (PHH) with SARS-CoV-2 and to evaluate the potential of miRNAs for modulating viral infection. METHODS: We analysed liver autopsies from a coronavirus disease 19 (COVID-19)-positive cohort for the presence of viral RNA using Nanopore sequencing. PHH were used for the infection with SARS-CoV-2. The candidate miRNAs targeting angiotensin converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2) were identified using in silico approaches. To discover the potential regulatory mechanism, transfection experiments, qRT-PCRs, western blots and luciferase reporter assays were performed. RESULTS: We could detect SARS-CoV-2 RNA in COVID-19-positive liver autopsies. We show that PHH express ACE2 and TMPRSS2 and can be readily infected with SARS-CoV-2, resulting in robust replication. Transfection of selected miRNA mimics reduced SARS-CoV-2 receptor expression and SARS-CoV-2 burden in PHH. In silico and biochemical analyses supported a potential direct binding of miR-141-3p to the SARS-CoV-2 genome. CONCLUSION: We confirm that PHH are susceptible to SARS-CoV-2 infection and demonstrate selected miRNAs targeting SARS-CoV-2 entry factors and/or the viral genome reduce viral loads. These data provide novel insights into hepatic susceptibility to SARS-CoV-2 and associated dysfunctions in COVID-19.
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C-type lectin domain-containing proteins (CTLDcps) shape host responses to pathogens and infectious disease outcomes. Previously, we identified the murine CTLDcp Cd302 as restriction factor, limiting hepatitis C virus (HCV) infection of murine hepatocytes. In this study, we investigated in detail the human orthologue's ability to restrict HCV infection in human liver cells. CD302 overexpression in Huh-7.5 cells potently inhibited infection of diverse HCV chimeras representing seven genotypes. Transcriptional profiling revealed abundant CD302 mRNA expression in human hepatocytes, the natural cellular target of HCV. Knockdown of endogenously expressed CD302 modestly enhanced HCV infection of Huh-7.5 cells and primary human hepatocytes. Functional analysis of naturally occurring CD302 transcript variants and engineered CD302 mutants showed that the C-type lectin-like domain (CTLD) is essential for HCV restriction, whereas the cytoplasmic domain (CPD) is dispensable. Coding single nucleotide polymorphisms occurring in human populations and mapping to different domains of CD302 did not influence the capacity of CD302 to restrict HCV. Assessment of the anti-HCV phenotype at different life cycle stages indicated that CD302 preferentially targets the viral entry step. In contrast to the murine orthologue, overexpression of human CD302 did not modulate downstream expression of nuclear receptor-controlled genes. Ectopic CD302 expression restricted infection of liver tropic hepatitis E virus (HEV), while it did not affect infection rates of two respiratory viruses, including respiratory syncytial virus (RSV) and the alpha coronavirus HVCoV-229E. Together, these findings suggest that CD302 contributes to liver cell-intrinsic defense against HCV and might mediate broader antiviral defenses against additional hepatotropic viruses. IMPORTANCE The liver represents an immunoprivileged organ characterized by enhanced resistance to immune responses. However, the importance of liver cell-endogenous, noncytolytic innate immune responses in pathogen control is not well defined. Although the role of myeloid cell-expressed CTLDcps in host responses to viruses has been characterized in detail, we have little information about their potential functions in the liver and their relevance for immune responses in this organ. Human hepatocytes endogenously express the CTLDcp CD302. Here, we provide evidence that CD302 limits HCV infection of human liver cells, likely by inhibiting a viral cell entry step. We confirm that the dominant liver-expressed transcript variant, as well as naturally occurring coding variants of CD302, maintain the capacity to restrict HCV. We further show that the CTLD of the protein is critical for the anti-HCV activity and that overexpressed CD302 limits HEV infection. Thus, CD302 likely contributes to human liver-intrinsic antiviral defenses.
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Hepacivirus , Hepatite C , Lectinas Tipo C , Receptores de Superfície Celular , Antivirais/metabolismo , Hepacivirus/fisiologia , Hepatite C/imunologia , Hepatócitos/imunologia , Hepatócitos/virologia , Humanos , Lectinas Tipo C/genética , Lectinas Tipo C/metabolismo , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Replicação ViralRESUMO
A novel application of the Theil-Sen robust regression method for determining the temporal trends in the concentration of heavy metals in UK ambient air over the period 2005-2020 is presented and compared to other regression methods. We have demonstrated improvements over non-robust methods of regression, proving the ability to tease out trends that are small with respect to the variability of the concentration measurement. The method is used to identify, in general, large and significant trends in the concentrations of Ni, As, Pb and V over the period 2005-2020, either across the UK as a whole or at groupings of site classifications in the UK. These trends have been compared to trends in emission data determined in the same manner. Although the results for most metals provide confidence that the UK metal network of monitoring sites is successful in appropriately capturing changes in emissions, a key finding of this work is the disagreement between trends in measured concentrations and emissions for Cu, Mn and Ni, for which we suggest improvements in future network design. The results also indicate that UK emission data for V should be reviewed, as we propose that the rate of reduction of V emissions is likely to have been overestimated.
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Poluentes Atmosféricos , Metais Pesados , Poluentes Atmosféricos/análise , Monitoramento Ambiental/métodos , Metais Pesados/análise , Análise de Regressão , Reino UnidoRESUMO
Transcriptional profiling provides global snapshots of virus-mediated cellular reprogramming, which can simultaneously encompass pro- and antiviral components. To determine early transcriptional signatures associated with HCV infection of authentic target cells, we performed ex vivo infections of adult primary human hepatocytes (PHHs) from seven donors. Longitudinal sampling identified minimal gene dysregulation at six hours post infection (hpi). In contrast, at 72 hpi, massive increases in the breadth and magnitude of HCV-induced gene dysregulation were apparent, affecting gene classes associated with diverse biological processes. Comparison with HCV-induced transcriptional dysregulation in Huh-7.5 cells identified limited overlap between the two systems. Of note, in PHHs, HCV infection initiated broad upregulation of canonical interferon (IFN)-mediated defense programs, limiting viral RNA replication and abrogating virion release. We further find that constitutive expression of IRF1 in PHHs maintains a steady-state antiviral program in the absence of infection, which can additionally reduce HCV RNA translation and replication. We also detected infection-induced downregulation of â¼90 genes encoding components of the EIF2 translation initiation complex and ribosomal subunits in PHHs, consistent with a signature of translational shutoff. As HCV polyprotein translation occurs independently of the EIF2 complex, this process is likely pro-viral: only translation initiation of host transcripts is arrested. The combination of antiviral intrinsic and inducible immunity, balanced against pro-viral programs, including translational arrest, maintains HCV replication at a low-level in PHHs. This may ultimately keep HCV under the radar of extra-hepatocyte immune surveillance while initial infection is established, promoting tolerance, preventing clearance and facilitating progression to chronicity.IMPORTANCEAcute HCV infections are often asymptomatic and therefore frequently undiagnosed. We endeavored to recreate this understudied phase of HCV infection using explanted PHHs and monitored host responses to initial infection. We detected temporally distinct virus-induced perturbations in the transcriptional landscape, which were initially narrow but massively amplified in breadth and magnitude over time. At 72 hpi, we detected dysregulation of diverse gene programs, concurrently promoting both virus clearance and virus persistence. On the one hand, baseline expression of IRF1 combined with infection-induced upregulation of IFN-mediated effector genes suppresses virus propagation. On the other, we detect transcriptional signatures of host translational inhibition, which likely reduces processing of IFN-regulated gene transcripts and facilitates virus survival. Together, our data provide important insights into constitutive and virus-induced transcriptional programs in PHHs, and identifies simultaneous antagonistic dysregulation of pro-and anti-viral programs which may facilitate host tolerance and promote viral persistence.
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Continuous release of pollutants into the environment poses serious threats to environmental sustainability and human health. For trace-level analysis of pollutants, layered double hydroxide (LDH) is an attractive option to impart enhanced sorption capability and sensitivity toward pollutants because of its unique layered structure, tunable interior architecture, high anion-exchange capacities, and high porosity (e.g., Zn/Cr LDH/DABCO-IL, Ni/Al LDH, CS-Ni/Fe LDH, SDS-Fe3O4@SiO2@Mg-Al LDH, Boeh/Mg/Al LDH/pC, and Fe@NiAl LDH). In concert with the well-defined analytical methodologies (e.g., HPLC and GC), the LDH materials can be employed to detect trace-level targets (e.g., as low as â¼ 20 fg/L for phenols) in aqueous environments. This review highlights LDH as a promising material for pre-treatment of a variety of organic and inorganic target pollutants in complex real matrices. Challenges and future requirements for research into LDH-based analytical methods are also discussed.
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Poluentes Ambientais , Humanos , Hidróxidos/química , Dióxido de SilícioRESUMO
Humans face threats from air pollutants present in both indoor and outdoor environments. The emerging role of plants in remediating the atmospheric environment is now being actively investigated as a possible solution for this problem. Foliar surfaces of plants (e.g., the leaves of cotton) can absorb a variety of airborne pollutants (e.g., formaldehyde, benzene, trimethylamine, and xylene), thereby reducing their concentrations in indoor environments. Recently, theoretical and experimental studies have been conducted to offer better insights into the interactions between plants and the surrounding air. In our research, an overview on the role of plants in reducing air pollution (often referred to as phytoremediation) is provided based on a comprehensive literature survey. The major issues for plant-based research for the reduction of air pollution in both outdoor and indoor environments are discussed in depth along with future challenges. Analysis of the existing data confirms the effectiveness of phytoremediation in terms of the absorption and purification of pollutants (e.g., by the leaves and roots of plants and trees), while being controlled by different variables (e.g., pore characteristics and planting patterns). Although most lab-scale studies have shown that plants can effectively absorb pollutants, it is important for such studies to reflect the real-world conditions, especially with the influence of human activities. Under such conditions, pollutants are to be replenished continually while the plant surface area to ambient atmosphere volume ratio vastly decreases (e.g., relative to lab-based experiments). The replication of such experimental conditions is the key challenge in this field of research. This review is expected to offer valuable insights into the innate ability of various plants in removing diverse pollutants (such as formaldehyde, benzene, and particulate matter) under different environmental settings.
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Poluentes Atmosféricos , Poluição do Ar em Ambientes Fechados , Poluição do Ar , Poluentes Atmosféricos/análise , Poluição do Ar/análise , Poluição do Ar/prevenção & controle , Poluição do Ar em Ambientes Fechados/análise , Humanos , Material Particulado/análise , PlantasRESUMO
This Feature highlights the role of metrology, the science of measurement, in maintaining the infrastructure we all rely on for accurate chemical measurements. In particular, the recent change to the definition of the mole, the unit of chemistry, is explained.
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Sistema Internacional de UnidadesRESUMO
A strategy is proposed for the design of wall envelopes to improve unsteady thermal performance in non-air-conditioned buildings and to reduce energy costs in air-conditioned buildings. The thermophysical properties of building materials (e.g., burnt bricks, mud bricks, laterite stone, cinder concrete, and expanded polystyrene) were measured experimentally using a thermal analyzer. A total of 28 combinations for composite walls were designed with expanded polystyrene as an insulation material based on seven criteria and were subjected to 8 different external surface heat transfer coefficients, which were tested for unsteady thermal performance parameters and air-conditioning cost-saving potential. In this paper, unsteady thermal transmittance obtained from admittance method has been employed to compute cost saving potential of air-conditioning for the various wall envelopes. The use of C-H5 design at a 2 m/s wind speed was found to increase the decrement lag of burnt brick, mud brick, laterite stone, and cinder concrete composite wall envelopes by 48.1%, 49.0%, 59.5%, and 47.0%, respectively, relative to the common wall design (C-H1) in non-air-conditioned buildings. The laterite with a C-H5 design offers the highest annual energy cost savings (1.71 $/m2 at 2 m/s), the highest life cycle cost savings (18.32 $/m2 at 2 m/s), and the lowest payback period (4.03 yrs at 2 m/s) in all tested building materials for air-conditioned buildings. The overall results of this study are expected to open new paths to deliver simple design strategies for energy-efficient buildings.
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Ar Condicionado , Materiais de Construção , Temperatura AltaRESUMO
Metrology remains a uniquely important endeavour. A sign of its success and robustness as an infra-technology is that it usually goes unnoticed. This means that it is in danger of being under-valued and under-appreciated. The sure-footing that metrology provides to the quality infrastructure will be especially important as the world grapples with the aftereffects of the COVID-19 pandemic, rebuilding global economies and also re-focusing on addressing global grand challenges and exploiting emerging technologies. In this context it is important and timely to re-examine the concept of metrology and how it relates to the quality infrastructure that it serves, but differs to measurement in general. The concept of metrology as 'measuring measurement' is proposed, emphasising the characteristic meta-thought associated with the discipline that distinguishes it from routine measurement.
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BACKGROUND & AIMS: HCV is a positive-strand RNA virus that primarily infects human hepatocytes. Recent studies have reported that C19orf66 is expressed as an interferon (IFN)-stimulated gene; however, the intrinsic regulation of this gene within the liver as well as its antiviral effects against HCV remain elusive. METHODS: Expression of C19orf66 was quantified in both liver biopsies and primary human hepatocytes, with or without HCV infection. Mechanistic studies of the potent anti-HCV phenotype mediated by C19orf66 were conducted using state-of-the-art virological, biochemical and genetic approaches, as well as correlative light and electron microscopy and transcriptome and proteome analysis. RESULTS: Upregulation of C19orf66 mRNA was observed in both primary human hepatocytes upon HCV infection and in the livers of patients with chronic hepatitis C (CHC). In addition, pegIFNα/ribavirin therapy induced C19orf66 expression in patients with CHC. Transcriptomic profiling and whole cell proteomics of hepatoma cells ectopically expressing C19orf66 revealed no induction of other antiviral genes. Expression of C19orf66 restricted HCV infection, whereas CRIPSPR/Cas9 mediated knockout of C19orf66 attenuated IFN-mediated suppression of HCV replication. Co-immunoprecipitation followed by mass spectrometry identified a stress granule protein-dominated interactome of C19orf66. Studies with subgenomic HCV replicons and an expression system revealed that C19orf66 expression impairs HCV-induced elevation of phosphatidylinositol-4-phosphate, alters the morphology of the viral replication organelle (termed the membranous web) and thereby targets viral RNA replication. CONCLUSION: C19orf66 is an IFN-stimulated gene, which is upregulated in hepatocytes within the first hours post IFN treatment or HCV infection in vivo. The encoded protein possesses specific antiviral activity against HCV and targets the formation of the membranous web. Our study identifies C19orf66 as an IFN-inducible restriction factor with a novel antiviral mechanism that specifically targets HCV replication. LAY SUMMARY: Interferon-stimulated genes are thought to be important to for antiviral immune responses to HCV. Herein, we analysed C19orf66, an interferon-stimulated gene, which appears to inhibit HCV replication. It prevents the HCV-induced elevation of phosphatidylinositol-4-phosphate and alters the morphology of HCV's replication organelle.
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Antivirais/uso terapêutico , Hepacivirus/genética , Hepatite C Crônica/tratamento farmacológico , Hepatite C Crônica/metabolismo , Interferons/uso terapêutico , Organelas/virologia , Proteínas de Ligação a RNA/metabolismo , Compartimentos de Replicação Viral/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos , Adulto , Linhagem Celular Tumoral , Feminino , Técnicas de Inativação de Genes , Genótipo , Células HEK293 , Hepatite C Crônica/patologia , Hepatite C Crônica/virologia , Hepatócitos/metabolismo , Humanos , Fígado/patologia , Masculino , Pessoa de Meia-Idade , Organelas/efeitos dos fármacos , Organelas/metabolismo , RNA Viral/metabolismo , Proteínas de Ligação a RNA/genética , Replicon/efeitos dos fármacos , Replicon/genética , Ribavirina/uso terapêutico , Resultado do Tratamento , Replicação Viral/genéticaRESUMO
OBJECTIVES: We aimed to identify existing outcome measures for functional neurological disorder (FND), to inform the development of recommendations and to guide future research on FND outcomes. METHODS: A systematic review was conducted to identify existing FND-specific outcome measures and the most common measurement domains and measures in previous treatment studies. Searches of Embase, MEDLINE and PsycINFO were conducted between January 1965 and June 2019. The findings were discussed during two international meetings of the FND-Core Outcome Measures group. RESULTS: Five FND-specific measures were identified-three clinician-rated and two patient-rated-but their measurement properties have not been rigorously evaluated. No single measure was identified for use across the range of FND symptoms in adults. Across randomised controlled trials (k=40) and observational treatment studies (k=40), outcome measures most often assessed core FND symptom change. Other domains measured commonly were additional physical and psychological symptoms, life impact (ie, quality of life, disability and general functioning) and health economics/cost-utility (eg, healthcare resource use and quality-adjusted life years). CONCLUSIONS: There are few well-validated FND-specific outcome measures. Thus, at present, we recommend that existing outcome measures, known to be reliable, valid and responsive in FND or closely related populations, are used to capture key outcome domains. Increased consistency in outcome measurement will facilitate comparison of treatment effects across FND symptom types and treatment modalities. Future work needs to more rigorously validate outcome measures used in this population.
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Doenças do Sistema Nervoso/diagnóstico , Doenças do Sistema Nervoso/terapia , Avaliação de Resultados em Cuidados de Saúde , HumanosRESUMO
A dualistic mind-body understanding of functional neurological disorders (FNDs), also known as conversion disorders, has led to the view that the cause of the symptom should be either psychological (psychogenic) or physical (neurogenic-"organic"). One of the most influential psychological approaches is the Freudian model of conversion, which suggests that FNDs arise from a defense process in which emotional stress is converted into physical symptoms. This conversion theory has been challenged in recent years, accompanied by a shift in emphasis toward neuropathophysiological models of FND and away from historical psychological concepts. In this review, the authors consider the contemporary relevance of the conversion model from the neuroscientific perspective to reconcile the role of both psychological and biological factors in FND. A narrative review of recent neuroscientific findings pertaining to the conversion model of FND, encompassing neuroimaging, cognitive psychology, biological markers, and epigenetic studies, was performed. Research on the role of psychological stressors is discussed. Neurobiological mechanisms of repression of traumatic memories and their translation into physical symptoms are then explored. Finally, the role of physical symptoms as a potential protective defense mechanism against social stressors is considered. The authors argue that the conversion concept is consistent with recent neuroscientific research findings, and the model allows psychological and neurobiological concepts to be reconciled within a single account of FND that begins to resolve the dualistic mind-body dichotomy.
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Transtorno Conversivo , Doenças do Sistema Nervoso , Transtornos Psicofisiológicos , Transtorno Conversivo/etiologia , Transtorno Conversivo/fisiopatologia , Transtorno Conversivo/psicologia , Humanos , Doenças do Sistema Nervoso/etiologia , Doenças do Sistema Nervoso/fisiopatologia , Doenças do Sistema Nervoso/psicologia , Transtornos Psicofisiológicos/etiologia , Transtornos Psicofisiológicos/fisiopatologia , Transtornos Psicofisiológicos/psicologiaRESUMO
Hepatitis C virus (HCV) displays a restricted host species tropism and only humans and chimpanzees are susceptible to infection. A robust immunocompetent animal model is still lacking, hampering mechanistic analysis of virus pathogenesis, immune control, and prophylactic vaccine development. The closest homolog of HCV is the equine nonprimate hepacivirus (NPHV), which shares similar features with HCV and thus represents an animal model to study hepacivirus infections in their natural hosts. We aimed to dissect equine immune responses after experimental NPHV infection and conducted challenge experiments to investigate immune protection against secondary NPHV infections. Horses were i.v. injected with NPHV containing plasma. Flow cytometric analysis was used to monitor immune cell frequencies and activation status. All infected horses became viremic after 1 or 2 wk and viremia could be detected in two horses for several weeks followed by a delayed seroconversion and viral clearance. Histopathological examinations of liver biopsies revealed mild, periportally accentuated infiltrations of lymphocytes, macrophages, and plasma cells with some horses displaying subclinical signs of hepatitis. Following viral challenge, an activation of equine immune responses was observed. Importantly, after a primary NPHV infection, horses were protected against rechallenge with the homologous as well as a distinct isolate with only minute amounts of circulating virus being detectable.
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Hepacivirus/fisiologia , Hepatite C/veterinária , Doenças dos Cavalos/imunologia , Animais , Anticorpos Antivirais/imunologia , Modelos Animais de Doenças , Hepacivirus/classificação , Hepacivirus/genética , Hepatite C/imunologia , Hepatite C/prevenção & controle , Hepatite C/virologia , Doenças dos Cavalos/prevenção & controle , Doenças dos Cavalos/virologia , Cavalos , Humanos , Filogenia , Linfócitos T/imunologiaRESUMO
Chronic pain is exacerbated by maladaptive cognition such as pain catastrophizing (PC). Biomarkers of PC mechanisms may aid precision medicine for chronic pain. Here, we investigate EEG biomarkers using mass univariate and multivariate (machine learning) approaches. We test theoretical notions that PC results from a combination of augmented aversive-value encoding ("magnification") and persistent expectations of pain ("rumination"). Healthy individuals with high or low levels of PC underwent an experimental pain model involving nociceptive laser stimuli preceded by cues predicting forthcoming pain intensity. Analysis of EEG acquired during the cue and laser stimulation provided event-related potentials (ERPs) identifying spatially and temporally-extended neural representations associated with pain catastrophizing. Specifically, differential neural responses to cues predicting high vs. low intensity pain (i.e. aversive value encoding) were larger in the high PC group, largely originating from mid-cingulate and superior parietal cortex. Multivariate spatiotemporal EEG patterns evoked from cues with high aversive value selectively and significantly differentiated the high PC from low PC group (64.6% classification accuracy). Regression analyses revealed that neural patterns classifying groups could be partially predicted (R2â¯=â¯28%) from those neural patterns classifying the aversive value of cues. In contrast, behavioural and EEG analyses did not provide evidence that PC modifies more persistent effects of prior expectation on pain perception and nociceptive responses. These findings support the hypothesis of magnification of aversive value encoding but not persistent expression of expectation in pain catastrophizers. Multivariate patterns of aversive value encoding provide promising biomarkers of maladaptive cognitive responses to chronic pain that have future potential for psychological treatment development and clinical stratification.
Assuntos
Encéfalo/fisiopatologia , Catastrofização/fisiopatologia , Adulto , Antecipação Psicológica/fisiologia , Mapeamento Encefálico/métodos , Sinais (Psicologia) , Eletroencefalografia/métodos , Potenciais Evocados/fisiologia , Feminino , Humanos , Masculino , Dor/psicologia , Percepção da Dor/fisiologia , Processamento de Sinais Assistido por Computador , Adulto JovemRESUMO
BACKGROUND & AIMS: Hepatitis C virus (HCV) infection causes chronic liver disease. Antivirals have been developed and cure infection. However, resistance can emerge and salvage therapies with alternative modes of action could be useful. Several licensed drugs have emerged as HCV entry inhibitors and are thus candidates for drug repurposing. We aimed to dissect their mode of action, identify improved derivatives and determine their viral targets. METHODS: HCV entry inhibition was tested for a panel of structurally related compounds, using chimeric viruses representing diverse genotypes, in addition to viruses containing previously determined resistance mutations. Chemical modeling and synthesis identified improved derivatives, while generation of susceptible and non-susceptible chimeric viruses pinpointed E1 determinants of compound sensitivity. RESULTS: Molecules of the diphenylpiperazine, diphenylpiperidine, phenothiazine, thioxanthene, and cycloheptenepiperidine chemotypes inhibit HCV infection by interfering with membrane fusion. These molecules and a novel p-methoxy-flunarizine derivative with improved efficacy preferentially inhibit genotype 2 viral strains. Viral residues within a central hydrophobic region of E1 (residues 290-312) control susceptibility. At the same time, viral features in this region also govern pH-dependence of viral membrane fusion. CONCLUSIONS: Small molecules from different chemotypes related to flunarizine preferentially inhibit HCV genotype 2 membrane fusion. A hydrophobic region proximal to the putative fusion loop controls sensitivity to these drugs and the pH range of membrane fusion. An algorithm considering viral features in this region predicts viral sensitivity to membrane fusion inhibitors. Resistance to flunarizine correlates with more relaxed pH requirements for fusion. LAY SUMMARY: This study describes diverse compounds that act as HCV membrane fusion inhibitors. It defines viral properties that determine sensitivity to these molecules and thus provides information to identify patients that may benefit from treatment with membrane fusion inhibitors.