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1.
Front Immunol ; 10: 272, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30873160

RESUMO

TILRR (Toll-like interleukin-1 receptor regulator), a transcript variant of FREM1, is a novel regulatory component, which stimulates innate immune responses through binding to IL-1R1 (Interleukin-1 receptor, type 1) and TLR (Toll-like receptor) complex. However, it is not known whether TILRR expression influences other genes in the NFκB signal transduction and pro-inflammatory responses. Our previous study identified FREM1 as a novel candidate gene in HIV-1 resistance/susceptibility in the Pumwani Sex worker cohort. In this study, we investigated the effect of TILRR overexpression on expression of genes in the NFκB signaling pathway in vitro. The effect of TILRR on mRNA expression of 84 genes related to NFκB signal transduction pathway was investigated by qRT-PCR. Overexpression of TILRR on pro-inflammatory cytokine/chemokine(s) secretion in cell culture supernatants was analyzed using Bioplex multiplex bead assay. We found that TILRR overexpression significantly influenced expression of many genes in HeLa and VK2/E6E7 cells. Several cytokine/chemokine(s), including IL-6, IL-8 (CXCL8), IP-10, MCP-1, MIP-1ß, and RANTES (CCL5) were significantly increased in the cell culture supernatants following TILRR overexpression. Although how TILRR influences the expression of these genes needs to be further studied, we are the first to show the influence of TILRR on many genes in the NFκB inflammatory pathways. The NFκB inflammatory response pathways are extremely important in microbial infection and pathogenesis, including HIV-1 transmission. Further study of the role of TILRR may identify the novel intervention targets and strategies against HIV infection.

2.
BMC Infect Dis ; 19(1): 117, 2019 Feb 06.
Artigo em Inglês | MEDLINE | ID: mdl-30727970

RESUMO

BACKGROUND: In the spring of 1918, the "War to End All Wars", which would ultimately claim more than 37 million lives, had entered into its final year and would change the global political and economic landscape forever. At the same time, a new global threat was emerging and would become one of the most devastating global health crises in recorded history. MAIN TEXT: The 1918 H1N1 pandemic virus spread across Europe, North America, and Asia over a 12-month period resulting in an estimated 500 million infections and 50-100 million deaths worldwide, of which ~ 50% of these occurred within the fall of 1918 (Emerg Infect Dis 12:15-22, 2006, Bull Hist Med 76:105-115, 2002). However, the molecular factors that contributed to the emergence of, and subsequent public health catastrophe associated with, the 1918 pandemic virus remained largely unknown until 2005, when the characterization of the reconstructed pandemic virus was announced heralding a new era of advanced molecular investigations (Science 310:77-80, 2005). In the century following the emergence of the 1918 pandemic virus we have landed on the Moon, developed the electronic computer (and a global internet), and have eradicated smallpox. In contrast, we have a largely remedial knowledge and understanding of one of the greatest scourges in recorded history. CONCLUSION: Here, we reflect on the 1918 influenza pandemic, including its emergence and subsequent rapid global spread. In addition, we discuss the pathophysiology associated with the 1918 virus and its predilection for the young and healthy, the rise of influenza therapeutic research following the pandemic, and, finally, our level of preparedness for future pandemics.


Assuntos
Antivirais/farmacologia , Vírus da Influenza A Subtipo H1N1/patogenicidade , Influenza Humana/epidemiologia , Influenza Humana/etiologia , Ásia/epidemiologia , Surtos de Doenças , Europa (Continente)/epidemiologia , Saúde Global , História do Século XX , Humanos , Vacinas contra Influenza/farmacologia , Influenza Humana/história , América do Norte/epidemiologia , Saúde Pública
3.
Viruses ; 11(2)2019 01 29.
Artigo em Inglês | MEDLINE | ID: mdl-30699912

RESUMO

Influenza viruses are a threat to global public health resulting in ~500,000 deaths each year. Despite an intensive vaccination program, influenza infections remain a recurrent, yet unsolved public health problem. Secondary bacterial infections frequently complicate influenza infections during seasonal outbreaks and pandemics, resulting in increased morbidity and mortality. Staphylococcus aureus, including methicillin-resistant S. aureus (MRSA), is frequently associated with these co-infections, including the 2009 influenza pandemic. Damage to alveolar epithelium is a major contributor to severe influenza-bacterial co-infections and can result in gas exchange abnormalities, fluid leakage, and respiratory insufficiency. These deleterious manifestations likely involve both pathogen- and host-mediated mechanisms. However, there is a paucity of information regarding the mechanisms (pathogen- and/or host-mediated) underlying influenza-bacterial co-infection pathogenesis. To address this, we characterized the contributions of viral-, bacterial-, and host-mediated factors to the altered structure and function of alveolar epithelial cells during co-infection with a focus on the 2009 pandemic influenza (pdm2009) and MRSA. Here, we characterized pdm2009 and MRSA replication kinetics, temporal host kinome responses, modulation of MRSA virulence factors, and disruption of alveolar barrier integrity in response to pdm2009-MRSA co-infection. Our results suggest that alveolar barrier disruption during co-infection is mediated primarily through host response dysregulation, resulting in loss of alveolar barrier integrity.


Assuntos
Células Epiteliais Alveolares/microbiologia , Células Epiteliais Alveolares/virologia , Barreira Alveolocapilar/fisiopatologia , Influenza Humana/fisiopatologia , Infecções Estafilocócicas/fisiopatologia , Células A549 , Coinfecção/microbiologia , Coinfecção/virologia , Humanos , Vírus da Influenza A Subtipo H1N1 , Influenza Humana/microbiologia , Pulmão/patologia , Staphylococcus aureus Resistente à Meticilina , Análise Serial de Proteínas , Infecções Estafilocócicas/virologia , Fatores de Virulência , Replicação Viral
4.
Viruses ; 10(12)2018 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-30513823

RESUMO

There is an increasing frequency of reports regarding the persistence of the Ebola virus (EBOV) in Ebola virus disease (EVD) survivors. During the 2014⁻2016 West African EVD epidemic, sporadic transmission events resulted in the initiation of new chains of human-to-human transmission. Multiple reports strongly suggest that these re-emergences were linked to persistent EBOV infections and included sexual transmission from EVD survivors. Asymptomatic infection and long-term viral persistence in EVD survivors could result in incidental introductions of the Ebola virus in new geographic regions and raise important national and local public health concerns. Alarmingly, although the persistence of filoviruses and their potential for sexual transmission have been documented since the emergence of such viruses in 1967, there is limited knowledge regarding the events that result in filovirus transmission to, and persistence within, the male reproductive tract. Asymptomatic infection and long-term viral persistence in male EVD survivors could lead to incidental transfer of EBOV to new geographic regions, thereby generating widespread outbreaks that constitute a significant threat to national and global public health. Here, we review filovirus testicular persistence and discuss the current state of knowledge regarding the rates of persistence in male survivors, and mechanisms underlying reproductive tract localization and sexual transmission.

6.
Drugs ; 77(18): 1935-1966, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-29143192

RESUMO

No specific antivirals are currently available for two emerging infectious diseases, Middle East respiratory syndrome (MERS) and severe acute respiratory syndrome (SARS). A literature search was performed covering pathogenesis, clinical features and therapeutics, clinically developed drugs for repurposing and novel drug targets. This review presents current knowledge on the epidemiology, pathogenesis and clinical features of the SARS and MERS coronaviruses. The rationale for and outcomes with treatments used for SARS and MERS is discussed. The main focus of the review is on drug development and the potential that drugs approved for other indications provide for repurposing. The drugs we discuss belong to a wide range of different drug classes, such as cancer therapeutics, antipsychotics, and antimalarials. In addition to their activity against MERS and SARS coronaviruses, many of these approved drugs have broad-spectrum potential and have already been in clinical use for treating other viral infections. A wealth of knowledge is available for these drugs. However, the information in this review is not meant to guide clinical decisions, and any therapeutic described here should only be used in context of a clinical trial. Potential targets for novel antivirals and antibodies are discussed as well as lessons learned from treatment development for other RNA viruses. The article concludes with a discussion of the gaps in our knowledge and areas for future research on emerging coronaviruses.


Assuntos
Antivirais/uso terapêutico , Infecções por Coronavirus/tratamento farmacológico , Síndrome Respiratória Aguda Grave/tratamento farmacológico , Antivirais/química , Infecções por Coronavirus/virologia , Aprovação de Drogas , Humanos , Coronavírus da Síndrome Respiratória do Oriente Médio , Vírus da SARS , Síndrome Respiratória Aguda Grave/virologia
7.
Clin Infect Dis ; 65(8): 1400-1403, 2017 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-28582513

RESUMO

In one patient over time, we found that concentration of Ebola virus RNA in semen during recovery is remarkably higher than blood at peak illness. Virus in semen is replication-competent with no change in viral genome over time. Presence of sense RNA suggests replication in cells present in semen.


Assuntos
Ebolavirus/genética , Doença pelo Vírus Ebola/virologia , Sêmen/virologia , Adulto , Ebolavirus/classificação , Genoma Viral/genética , Humanos , Masculino , RNA Viral/análise , RNA Viral/genética , Carga Viral
8.
Sci Transl Med ; 9(385)2017 04 12.
Artigo em Inglês | MEDLINE | ID: mdl-28404864

RESUMO

The 2013-2015 outbreak of Ebola virus disease in Guinea, Liberia, and Sierra Leone was unprecedented in the number of documented cases, but there have been few published reports on immune responses in clinical cases and their relationships with the course of illness and severity of Ebola virus disease. Symptoms of Ebola virus disease can include severe headache, myalgia, asthenia, fever, fatigue, diarrhea, vomiting, abdominal pain, and hemorrhage. Although experimental treatments are in development, there are no current U.S. Food and Drug Administration-approved vaccines or therapies. We report a detailed study of host gene expression as measured by microarray in daily peripheral blood samples collected from a patient with severe Ebola virus disease. This individual was provided with supportive care without experimental therapies at the National Institutes of Health Clinical Center from before onset of critical illness to recovery. Pearson analysis of daily gene expression signatures revealed marked gene expression changes in peripheral blood leukocytes that correlated with changes in serum and peripheral blood leukocytes, viral load, antibody responses, coagulopathy, multiple organ dysfunction, and then recovery. This study revealed marked shifts in immune and antiviral responses that preceded changes in medical condition, indicating that clearance of replicating Ebola virus from peripheral blood leukocytes is likely important for systemic viral clearance.


Assuntos
Ebolavirus/patogenicidade , Doença pelo Vírus Ebola/virologia , Leucócitos/metabolismo , Surtos de Doenças , Doença pelo Vírus Ebola/sangue , Humanos , Estudos Longitudinais , RNA Viral/sangue , RNA Viral/genética , Replicação Viral/fisiologia
9.
ACS Infect Dis ; 2(11): 787-799, 2016 11 11.
Artigo em Inglês | MEDLINE | ID: mdl-27933782

RESUMO

Outbreaks associated with emerging and re-emerging viral pathogens continue to increase in frequency and are associated with an increasing burden to global health. In light of this, there is a need to integrate basic and clinical research for investigating the connections between molecular and clinical pathogenesis and for therapeutic development strategies. Here, we will discuss this approach with a focus on the emerging viral pathogens Middle East respiratory syndrome coronavirus (MERS-CoV), Ebola virus (EBOV), and monkeypox virus (MPXV) from the context of clinical presentation, immunological and molecular features of the diseases, and OMICS-based analyses of pathogenesis. Furthermore, we will highlight the role of global investigations of host kinases, the kinome, for investigating emerging and re-emerging viral pathogens from the context of characterizing cellular responses and identifying novel therapeutic targets. Lastly, we will address how increased integration of clinical and basic research will assist treatment and prevention efforts for emerging pathogens.


Assuntos
Viroses/genética , Viroses/virologia , Fenômenos Fisiológicos Virais , Animais , Antivirais/farmacologia , Humanos , Viroses/tratamento farmacológico , Viroses/metabolismo , Vírus/efeitos dos fármacos , Vírus/genética , Vírus/patogenicidade
10.
Virology ; 493: 238-46, 2016 06.
Artigo em Inglês | MEDLINE | ID: mdl-27062579

RESUMO

The 1918 influenza pandemic caused ~50 million deaths. Many questions remain regarding the origin, pathogenicity, and mechanisms of human adaptation of this virus. Avian-adapted influenza A viruses preferentially bind α2,3-linked sialic acids (Sia) while human-adapted viruses preferentially bind α2,6-linked Sia. A change in Sia preference from α2,3 to α2,6 is thought to be a requirement for human adaptation of avian influenza viruses. Autopsy data from 1918 cases, however, suggest that factors other than Sia preference played a role in viral binding and entry to human airway cells. Here, we evaluated binding and entry of five 1918 influenza receptor binding domain variants in a primary human airway cell model along with control avian and human influenza viruses. We observed that all five variants bound and entered cells efficiently and that Sia preference did not predict entry of influenza A virus to primary human airway cells evaluated in this model.


Assuntos
Células Epiteliais/virologia , Vírus da Influenza A Subtipo H1N1/metabolismo , Receptores Virais/metabolismo , Ligação Viral , Sítios de Ligação , Brônquios/citologia , Influenza Pandêmica, 1918-1919 , Ácido N-Acetilneuramínico/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Traqueia/citologia , Replicação Viral
11.
Antiviral Res ; 129: 120-129, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-26923881

RESUMO

BACKGROUND: Influenza results in up to 500,000 deaths annually. Seasonal influenza vaccines have an estimated 60% effectiveness, but provide little or no protection against novel subtypes, and may be less protective in high-risk groups. Neuraminidase inhibitors are recommended for the treatment of severe influenza infection, but are not proven to reduce mortality in severe disease. Preclinical models of severe influenza infection that closely correlate to human disease are needed to assess efficacy of new vaccines and therapeutics. METHODS: We developed a nonhuman primate model of influenza and bacterial co-infection that recapitulates severe pneumonia in humans. Animals were infected with influenza A virus via intra-bronchial or small-particle aerosol inoculation, methicillin-resistant Staphylococcus aureus, or co-infected with influenza and methicillin-resistant S. aureus combined. We assessed the severity of disease in animals over the course of our study using tools available to evaluate critically ill human patients including high-resolution computed tomography imaging of the lungs, arterial blood gas analyses, and bronchoalveolar lavage. RESULTS: Using an intra-bronchial route of inoculation we successfully induced severe pneumonia following influenza infection alone and following influenza and bacterial co-infection. Peak illness was observed at day 6 post-influenza infection, manifested by bilateral pulmonary infiltrates and hypoxemia. The timing of radiographic and physiologic manifestations of disease in our model closely match those observed in severe human influenza infection. DISCUSSION: This was the first nonhuman primate study of influenza and bacterial co-infection where high-resolution computed tomography scanning of the lungs was used to quantitatively assess pneumonia over the course of illness and where hypoxemia was correlated with pneumonia severity. With additional validation this model may serve as a pathway for regulatory approval of vaccines and therapeutics for the prevention and treatment of severe influenza pneumonia.


Assuntos
Coinfecção , Vírus da Influenza A , Modelos Animais , Infecções por Orthomyxoviridae/complicações , Pneumonia Estafilocócica/complicações , Pneumonia Viral/complicações , Animais , Humanos , Vírus da Influenza A/patogenicidade , Vacinas contra Influenza , Influenza Humana/complicações , Influenza Humana/microbiologia , Pulmão/microbiologia , Pulmão/patologia , Pulmão/virologia , Macaca mulatta , Masculino , Staphylococcus aureus Resistente à Meticilina/patogenicidade , Tomografia Computadorizada com Tomografia por Emissão de Pósitrons
12.
J Pathol ; 238(1): 85-97, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26383585

RESUMO

To study bacterial co-infection following 1918 H1N1 influenza virus infection, mice were inoculated with the 1918 influenza virus, followed by Streptococcus pneumoniae (SP) 72 h later. Co-infected mice exhibited markedly more severe disease, shortened survival time and more severe lung pathology, including widespread thrombi. Transcriptional profiling revealed activation of coagulation only in co-infected mice, consistent with the extensive thrombogenesis observed. Immunohistochemistry showed extensive expression of tissue factor (F3) and prominent deposition of neutrophil elastase on endothelial and epithelial cells in co-infected mice. Lung sections of SP-positive 1918 autopsy cases showed extensive thrombi and prominent staining for F3 in alveolar macrophages, monocytes, neutrophils, endothelial and epithelial cells, in contrast to co-infection-positive 2009 pandemic H1N1 autopsy cases. This study reveals that a distinctive feature of 1918 influenza virus and SP co-infection in mice and humans is extensive expression of tissue factor and activation of the extrinsic coagulation pathway leading to widespread pulmonary thrombosis.


Assuntos
Coinfecção/complicações , Influenza Humana/microbiologia , Infecções por Orthomyxoviridae/microbiologia , Infecções Pneumocócicas/microbiologia , Embolia Pulmonar/microbiologia , Animais , Coagulação Sanguínea , Modelos Animais de Doenças , Feminino , Humanos , Imuno-Histoquímica , Vírus da Influenza A Subtipo H1N1 , Influenza Pandêmica, 1918-1919 , Influenza Humana/complicações , Influenza Humana/patologia , Camundongos , Camundongos Endogâmicos BALB C , Análise de Sequência com Séries de Oligonucleotídeos , Infecções por Orthomyxoviridae/complicações , Infecções por Orthomyxoviridae/patologia , Infecções Pneumocócicas/complicações , Infecções Pneumocócicas/patologia , Embolia Pulmonar/patologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Streptococcus pneumoniae
13.
Mol Cell Proteomics ; 14(3): 646-57, 2015 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-25573744

RESUMO

The Syrian golden hamster has been increasingly used to study viral hemorrhagic fever (VHF) pathogenesis and countermeasure efficacy. As VHFs are a global health concern, well-characterized animal models are essential for both the development of therapeutics and vaccines as well as for increasing our understanding of the molecular events that underlie viral pathogenesis. However, the paucity of reagents or platforms that are available for studying hamsters at a molecular level limits the ability to extract biological information from this important animal model. As such, there is a need to develop platforms/technologies for characterizing host responses of hamsters at a molecular level. To this end, we developed hamster-specific kinome peptide arrays to characterize the molecular host response of the Syrian golden hamster. After validating the functionality of the arrays using immune agonists of defined signaling mechanisms (lipopolysaccharide (LPS) and tumor necrosis factor (TNF)-α), we characterized the host response in a hamster model of VHF based on Pichinde virus (PICV(1)) infection by performing temporal kinome analysis of lung tissue. Our analysis revealed key roles for vascular endothelial growth factor (VEGF), interleukin (IL) responses, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling, and Toll-like receptor (TLR) signaling in the response to PICV infection. These findings were validated through phosphorylation-specific Western blot analysis. Overall, we have demonstrated that hamster-specific kinome arrays are a robust tool for characterizing the species-specific molecular host response in a VHF model. Further, our results provide key insights into the hamster host response to PICV infection and will inform future studies with high-consequence VHF pathogens.


Assuntos
Febre Hemorrágica Americana/virologia , Pulmão/enzimologia , Vírus Pichinde/fisiologia , Proteínas Quinases/isolamento & purificação , Proteoma/análise , Animais , Modelos Animais de Doenças , Feminino , Febre Hemorrágica Americana/enzimologia , Interleucinas/isolamento & purificação , Pulmão/virologia , Mesocricetus , NF-kappa B/isolamento & purificação , Fosforilação , Transdução de Sinais , Especificidade da Espécie , Receptores Toll-Like/isolamento & purificação , Fator A de Crescimento do Endotélio Vascular/isolamento & purificação
14.
Antimicrob Agents Chemother ; 59(2): 1088-99, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25487801

RESUMO

Middle East respiratory syndrome coronavirus (MERS-CoV) is a lineage C betacoronavirus, and infections with this virus can result in acute respiratory syndrome with renal failure. Globally, MERS-CoV has been responsible for 877 laboratory-confirmed infections, including 317 deaths, since September 2012. As there is a paucity of information regarding the molecular pathogenesis associated with this virus or the identities of novel antiviral drug targets, we performed temporal kinome analysis on human hepatocytes infected with the Erasmus isolate of MERS-CoV with peptide kinome arrays. bioinformatics analysis of our kinome data, including pathway overrepresentation analysis (ORA) and functional network analysis, suggested that extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) and phosphoinositol 3-kinase (PI3K)/serine-threonine kinase (AKT)/mammalian target of rapamycin (mTOR) signaling responses were specifically modulated in response to MERS-CoV infection in vitro throughout the course of infection. The overrepresentation of specific intermediates within these pathways determined by pathway and functional network analysis of our kinome data correlated with similar patterns of phosphorylation determined through Western blot array analysis. In addition, analysis of the effects of specific kinase inhibitors on MERS-CoV infection in tissue culture models confirmed these cellular response observations. Further, we have demonstrated that a subset of licensed kinase inhibitors targeting the ERK/MAPK and PI3K/AKT/mTOR pathways significantly inhibited MERS-CoV replication in vitro whether they were added before or after viral infection. Taken together, our data suggest that ERK/MAPK and PI3K/AKT/mTOR signaling responses play important roles in MERS-CoV infection and may represent novel drug targets for therapeutic intervention strategies.


Assuntos
Infecções por Coronavirus/metabolismo , Hepatócitos/metabolismo , Hepatócitos/virologia , Coronavírus da Síndrome Respiratória do Oriente Médio/fisiologia , Fosfatidilinositol 3-Quinases/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Western Blotting , Linhagem Celular , Biologia Computacional , Infecções por Coronavirus/virologia , Ensaio de Imunoadsorção Enzimática , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Humanos , Fosforilação , Transdução de Sinais/fisiologia
15.
Viruses ; 6(9): 3663-82, 2014 Sep 26.
Artigo em Inglês | MEDLINE | ID: mdl-25256396

RESUMO

Sequence determination of complete or coding-complete genomes of viruses is becoming common practice for supporting the work of epidemiologists, ecologists, virologists, and taxonomists. Sequencing duration and costs are rapidly decreasing, sequencing hardware is under modification for use by non-experts, and software is constantly being improved to simplify sequence data management and analysis. Thus, analysis of virus disease outbreaks on the molecular level is now feasible, including characterization of the evolution of individual virus populations in single patients over time. The increasing accumulation of sequencing data creates a management problem for the curators of commonly used sequence databases and an entry retrieval problem for end users. Therefore, utilizing the data to their fullest potential will require setting nomenclature and annotation standards for virus isolates and associated genomic sequences. The National Center for Biotechnology Information's (NCBI's) RefSeq is a non-redundant, curated database for reference (or type) nucleotide sequence records that supplies source data to numerous other databases. Building on recently proposed templates for filovirus variant naming [ ()////-], we report consensus decisions from a majority of past and currently active filovirus experts on the eight filovirus type variants and isolates to be represented in RefSeq, their final designations, and their associated sequences.


Assuntos
Bases de Dados de Ácidos Nucleicos , Filoviridae/genética , Evolução Molecular , Filoviridae/classificação , Humanos , Seleção Genética
16.
J Virol ; 88(17): 9877-92, 2014 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-24942569

RESUMO

UNLABELLED: Ebola virus (EBOV) causes a severe hemorrhagic disease in humans and nonhuman primates, with a median case fatality rate of 78.4%. Although EBOV is considered a public health concern, there is a relative paucity of information regarding the modulation of the functional host response during infection. We employed temporal kinome analysis to investigate the relative early, intermediate, and late host kinome responses to EBOV infection in human hepatocytes. Pathway overrepresentation analysis and functional network analysis of kinome data revealed that transforming growth factor (TGF-ß)-mediated signaling responses were temporally modulated in response to EBOV infection. Upregulation of TGF-ß signaling in the kinome data sets correlated with the upregulation of TGF-ß secretion from EBOV-infected cells. Kinase inhibitors targeting TGF-ß signaling, or additional cell receptors and downstream signaling pathway intermediates identified from our kinome analysis, also inhibited EBOV replication. Further, the inhibition of select cell signaling intermediates identified from our kinome analysis provided partial protection in a lethal model of EBOV infection. To gain perspective on the cellular consequence of TGF-ß signaling modulation during EBOV infection, we assessed cellular markers associated with upregulation of TGF-ß signaling. We observed upregulation of matrix metalloproteinase 9, N-cadherin, and fibronectin expression with concomitant reductions in the expression of E-cadherin and claudin-1, responses that are standard characteristics of an epithelium-to-mesenchyme-like transition. Additionally, we identified phosphorylation events downstream of TGF-ß that may contribute to this process. From these observations, we propose a model for a broader role of TGF-ß-mediated signaling responses in the pathogenesis of Ebola virus disease. IMPORTANCE: Ebola virus (EBOV), formerly Zaire ebolavirus, causes a severe hemorrhagic disease in humans and nonhuman primates and is the most lethal Ebola virus species, with case fatality rates of up to 90%. Although EBOV is considered a worldwide concern, many questions remain regarding EBOV molecular pathogenesis. As it is appreciated that many cellular processes are regulated through kinase-mediated phosphorylation events, we employed temporal kinome analysis to investigate the functional responses of human hepatocytes to EBOV infection. Administration of kinase inhibitors targeting signaling pathway intermediates identified in our kinome analysis inhibited viral replication in vitro and reduced EBOV pathogenesis in vivo. Further analysis of our data also demonstrated that EBOV infection modulated TGF-ß-mediated signaling responses and promoted "mesenchyme-like" phenotypic changes. Taken together, these results demonstrated that EBOV infection specifically modulates TGF-ß-mediated signaling responses in epithelial cells and may have broader implications in EBOV pathogenesis.


Assuntos
Diferenciação Celular , Ebolavirus/fisiologia , Hepatócitos/fisiologia , Interações Hospedeiro-Patógeno , Mesoderma/crescimento & desenvolvimento , Transdução de Sinais , Fator de Crescimento Transformador beta/metabolismo , Animais , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Doença pelo Vírus Ebola/patologia , Humanos , Camundongos Endogâmicos BALB C
17.
Antimicrob Agents Chemother ; 58(8): 4885-93, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24841273

RESUMO

Outbreaks of emerging infections present health professionals with the unique challenge of trying to select appropriate pharmacologic treatments in the clinic with little time available for drug testing and development. Typically, clinicians are left with general supportive care and often untested convalescent-phase plasma as available treatment options. Repurposing of approved pharmaceutical drugs for new indications presents an attractive alternative to clinicians, researchers, public health agencies, drug developers, and funding agencies. Given the development times and manufacturing requirements for new products, repurposing of existing drugs is likely the only solution for outbreaks due to emerging viruses. In the studies described here, a library of 290 compounds was screened for antiviral activity against Middle East respiratory syndrome coronavirus (MERS-CoV) and severe acute respiratory syndrome coronavirus (SARS-CoV). Selection of compounds for inclusion in the library was dependent on current or previous FDA approval or advanced clinical development. Some drugs that had a well-defined cellular pathway as target were included. In total, 27 compounds with activity against both MERS-CoV and SARS-CoV were identified. The compounds belong to 13 different classes of pharmaceuticals, including inhibitors of estrogen receptors used for cancer treatment and inhibitors of dopamine receptor used as antipsychotics. The drugs identified in these screens provide new targets for in vivo studies as well as incorporation into ongoing clinical studies.


Assuntos
Antivirais/farmacologia , Reposicionamento de Medicamentos , Coronavírus da Síndrome Respiratória do Oriente Médio/efeitos dos fármacos , Vírus da SARS/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/farmacologia , Animais , Antipsicóticos/farmacologia , Cercopithecus aethiops , Aprovação de Drogas , Antagonistas de Estrogênios/farmacologia , Ensaios de Triagem em Larga Escala , Humanos , Concentração Inibidora 50 , Coronavírus da Síndrome Respiratória do Oriente Médio/fisiologia , Vírus da SARS/fisiologia , Células Vero , Replicação Viral/efeitos dos fármacos
18.
Pathog Dis ; 71(2): 213-9, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24687975

RESUMO

Scientists at the National Institute of Allergy and Infectious Diseases Integrated Research Facility at Fort Detrick, Frederick, Maryland, coordinate and facilitate preclinical research on infectious diseases to develop medical countermeasures for high-consequence pathogens. This facility is unique in that it is the only maximum containment laboratory in the world where conventional and molecular medical imaging equipments are incorporated into the design of the facility. This capability provides investigators with unique tools to dissect disease pathogenesis, evaluate the ability of animal models to recapitulate human disease, and test candidate countermeasures. Importantly, advanced molecular imaging has the potential to provide alternative endpoints to lethality. Using these alternative endpoints, investigators can reduce the number of animals used in experiments and evaluate countermeasures in sublethal models. With the incorporation of medical imaging modalities, a clinical laboratory modeled after those existing in hospitals, and a highly trained veterinary medicine team, IRF-Frederick is uniquely suited to advance our understanding of emerging infectious diseases and to facilitate the development of medical countermeasures and clinical care paradigms previously considered impossible.


Assuntos
Pesquisa Biomédica/métodos , Controle de Doenças Transmissíveis/métodos , Doenças Transmissíveis/diagnóstico , Doenças Transmissíveis/terapia , Contenção de Riscos Biológicos , Saúde do Trabalhador , Experimentação Animal , Animais , Técnicas de Laboratório Clínico/métodos , Humanos , Maryland , National Institute of Allergy and Infectious Diseases (U.S.) , Imagem Óptica/métodos , Estados Unidos , Medicina Veterinária/métodos
19.
Pathog Dis ; 71(2): 190-98, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24585711

RESUMO

Currently, there is a paucity of information regarding the molecular pathogenesis for many high-consequence pathogens (HCPs) that pose threats to both national and international public health. In spite of this, investigations of the molecular pathogenesis for many HCPs have been limited to gross pathological changes in animal models or global analysis of gene expression. Further, questions remain regarding the ability of animal models of disease to recapitulate human molecular pathogenesis or act as predictors of therapeutic efficacy. Thus, it is likely that medical countermeasure development for HCPs will rely on identifying therapeutic targets that are uniquely modulated during HCP infection. It is also appreciated that many cellular processes can be regulated independently of changes in transcription or translation through phosphorylation events. Cellular kinases, individually or collectively (the kinome), play critical roles in regulating complex biology, underlie various malignancies, and represent high-priority drug targets. The growing interest in kinases in both basic and translational research has driven efforts to develop technologies that enable characterization of phosphorylation-mediated signal transduction. To this end, enhanced technical capabilities at the IRF-Frederick provide the unique capability for characterizing host responses to HCP insult during the course of infection and identify novel targets for therapeutic intervention.


Assuntos
Doenças Transmissíveis/diagnóstico , Doenças Transmissíveis/tratamento farmacológico , Modelos Animais de Doenças , Interações Hospedeiro-Patógeno , Proteínas Quinases/metabolismo , Processamento de Proteína Pós-Traducional , Proteoma/análise , Animais , Testes Diagnósticos de Rotina/métodos , Descoberta de Drogas/métodos , Regulação da Expressão Gênica , Humanos , National Institute of Allergy and Infectious Diseases (U.S.) , Fosforilação , Estados Unidos
20.
Viruses ; 6(1): 137-50, 2014 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-24402304

RESUMO

The overall threat of a viral pathogen to human populations is largely determined by the modus operandi and velocity of the pathogen that is transmitted among humans. Microorganisms that can spread by aerosol are considered a more challenging enemy than those that require direct body-to-body contact for transmission, due to the potential for infection of numerous people rather than a single individual. Additionally, disease containment is much more difficult to achieve for aerosolized viral pathogens than for pathogens that spread solely via direct person-to-person contact. Thus, aerobiology has become an increasingly necessary component for studying viral pathogens that are naturally or intentionally transmitted by aerosol. The goal of studying aerosol viral pathogens is to improve public health preparedness and medical countermeasure development. Here, we provide a brief overview of the animal biosafety level 4 Aerobiology Core at the NIH/NIAID Integrated Research Facility at Fort Detrick, Maryland, USA.


Assuntos
Aerossóis , Pesquisa Biomédica/métodos , Contenção de Riscos Biológicos/métodos , Exposição por Inalação , Viroses/transmissão , Animais , Modelos Animais de Doenças , Maryland , National Institute of Allergy and Infectious Diseases (U.S.) , National Institutes of Health (U.S.) , Estados Unidos
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