291 results on '"Andrew Pekosz"'
Search Results
252. Replication in Cultured C2C12 Muscle Cells Correlates with the Neuroinvasiveness of California Serogroup Bunyaviruses
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Richard S. Davidson, Andrew Pekosz, Kara Stillmock, Neal Nathanson, Francisco Gonzalez-Scarano, Dan Schmeidler, Ingenue Cobbinah, Christian Griot, David M. Lukac, and Maarten Hoek
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Infectivity ,La Crosse virus ,Genotype ,Virulence ,biology ,Muscles ,viruses ,RNA ,Virus Replication ,biology.organism_classification ,Nervous System ,Virology ,Tahyna virus ,Virus ,Cell Line ,Mice ,Titer ,Viral envelope ,Animals ,Serotyping ,Bunyaviridae - Abstract
The neuroinvasiveness of California serogroup bunyaviruses is determined by the ability of the virus to replicate in striated muscle after peripheral inoculation of mice. Neuroinvasiveness was mapped to the medium (M) RNA segment of the virus, which encodes the viral glycoproteins, when reassortants were made between La Crosse/original virus, a neuroinvasive isolate, and Tahyna-181/57 virus, a nonneuroinvasive clone. We have tested the murine muscle cell line C2C12 as a surrogate for myotropism and have found that there is a slight, but reproducible difference in the replication of virus clones bearing the M RNA segment of La Crosse/original virus compared to clones bearing the M RNA segment of Tahyna-181/57 virus, as determined by viral titer, antigen expression, and plaque formation.
- Published
- 1994
253. The influenza C virus CM2 protein can alter intracellular pH, and its transmembrane domain can substitute for that of the influenza A virus M2 protein and support infectious virus production
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Shaun M. Stewart and Andrew Pekosz
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Cytoplasm ,Influenzavirus C ,viruses ,Immunology ,Biology ,medicine.disease_cause ,Virus Replication ,Microbiology ,Antigenic drift ,Cell Line ,Viral Matrix Proteins ,VP40 ,Virology ,Influenza, Human ,Influenza A virus ,medicine ,Animals ,Humans ,Viral matrix protein ,Structure and Assembly ,Antigenic shift ,Hydrogen-Ion Concentration ,Protein Structure, Tertiary ,NS2-3 protease ,Transmembrane domain ,Insect Science ,Influenza C Virus - Abstract
The influenza C virus CM2 protein and a chimeric influenza A virus M2 protein (MCM) containing the CM2 transmembrane domain were assessed for their ability to functionally replace the M2 protein. While all three proteins could alter cytosolic pH to various degrees when expressed from cDNA, only M2 and MCM could at least partially restore infectious virus production to M2-deficient influenza A viruses. The data suggest that while the CM2 ion channel activity is similar to that of M2, sequences in the extracellular and/or cytoplasmic domains play important roles in infectious virus production.
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- 2011
254. Alterations In Permeability And Antiviral Responses In Influenza A Virus Infected Differentiated Human Respiratory Epithelial Cell Cultures
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Venkataramana K. Sidhaye, William A. Fischer, Franco R. D'Alessio, Andrew P. Lane, Landon S. King, and Andrew Pekosz
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medicine.anatomical_structure ,Permeability (electromagnetism) ,Influenza A virus ,medicine ,Biology ,Respiratory system ,medicine.disease_cause ,Virology ,Epithelium - Published
- 2011
255. Influenza Virus-Like Particles Containing M2 Induce Broadly Cross Protective Immunity
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Nico van Rooijen, Bao-Zhong Wang, Andrew Pekosz, Min Chul Kim, Jae-Min Song, Fu-Shi Quan, Kyoung-mi Park, Sang-Moo Kang, Hyun Tak Jin, Richard W. Compans, Molecular cell biology and Immunology, and CCA - Immuno-pathogenesis
- Subjects
Influenza vaccine ,viruses ,Cross Protection ,lcsh:Medicine ,Biology ,medicine.disease_cause ,Antibodies, Viral ,Antigenic drift ,Virus ,Microbiology ,Viral Matrix Proteins ,Mice ,Influenza A Virus, H1N1 Subtype ,Orthomyxoviridae Infections ,Immunity ,Pandemic ,Influenza A virus ,medicine ,Animals ,Humans ,lcsh:Science ,Virology/Vaccines ,Multidisciplinary ,Viral matrix protein ,Strain (biology) ,Macrophages ,lcsh:R ,Virion ,virus diseases ,Dendritic Cells ,Virology ,Influenza Vaccines ,Immunology/Immune Response ,lcsh:Q ,Virology/Host Antiviral Responses ,Research Article - Abstract
Background Current influenza vaccines based on the hemagglutinin protein are strain specific and do not provide good protection against drifted viruses or emergence of new pandemic strains. An influenza vaccine that can confer cross-protection against antigenically different influenza A strains is highly desirable for improving public health. Methodology/Principal Findings To develop a cross protective vaccine, we generated influenza virus-like particles containing the highly conserved M2 protein in a membrane-anchored form (M2 VLPs), and investigated their immunogenicity and breadth of cross protection. Immunization of mice with M2 VLPs induced anti-M2 antibodies binding to virions of various strains, M2 specific T cell responses, and conferred long-lasting cross protection against heterologous and heterosubtypic influenza viruses. M2 immune sera were found to play an important role in providing cross protection against heterosubtypic virus and an antigenically distinct 2009 pandemic H1N1 virus, and depletion of dendritic and macrophage cells abolished this cross protection, providing new insight into cross-protective immune mechanisms. Conclusions/Significance These results suggest that presenting M2 on VLPs in a membrane-anchored form is a promising approach for developing broadly cross protective influenza vaccines.
- Published
- 2011
256. The impact of sex, gender and pregnancy on 2009 H1N1 disease
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Peju Olukoya, Sabra L. Klein, Martha Anker, Andrew Pekosz, and Catherine L. Passaretti
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medicine.medical_specialty ,Pregnancy ,lcsh:QP1-981 ,business.industry ,Mortality rate ,Incidence (epidemiology) ,lcsh:R ,lcsh:Medicine ,Review ,Disease ,medicine.disease ,lcsh:Physiology ,Gender Studies ,Vaccination ,Endocrinology ,Epidemiology ,Immunology ,medicine ,Young adult ,business ,Disease burden ,Demography - Abstract
Children and young adults of reproductive age have emerged as groups that are highly vulnerable to the current 2009 H1N1 pandemic. The sex of an individual is a fundamental factor that can influence exposure, susceptibility and immune responses to influenza. Worldwide, the incidence, disease burden, morbidity and mortality rates following exposure to the 2009 H1N1 influenza virus differ between males and females and are often age-dependent. Pregnancy and differences in the presentation of various risk factors contribute to the worse outcome of infection in women. Vaccination and antiviral treatment efficacy also vary in a sex-dependent manner. Finally, sex-specific genetic and hormonal differences may contribute to the severity of influenza and the clearance of viral infection. The contribution of sex and gender to influenza can only be determined by a greater consideration of these factors in clinical and epidemiological studies and increased research into the biological basis underlying these differences.
- Published
- 2010
257. Characterization Of Live, Attenuated And Wild Type Seasonal H1N1 Influenza Virus Replication In Primary, Differentiated Human Nasal Epithelial Cells
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Landon S. King, William A. Fischer, Ramana Sidhaye, Andrew Pekosz, and Andrew P. Lane
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Primary (chemistry) ,Viral replication ,H1N1 influenza ,Wild type ,Biology ,Virology - Published
- 2010
258. Delivery of woodchuck hepatitis virus-like particle presented influenza M2e by recombinant attenuated Salmonella displaying a delayed lysis phenotype
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Wai Hong Wu, Roy Curtiss, Shamaila Ashraf, Jean Noel Billaud, Keith Ameiss, Andrew Pekosz, David R. Milich, and Wei Kong
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Salmonella typhimurium ,Recombinant Fusion Proteins ,Influenza A Virus, H7N7 Subtype ,Biology ,medicine.disease_cause ,Antibodies, Viral ,Virus ,Article ,Microbiology ,Viral Matrix Proteins ,Mice ,Antigen ,Virus-like particle ,Orthomyxoviridae Infections ,medicine ,Influenza A virus ,Animals ,Hepatitis B Virus, Woodchuck ,Vaccines, Virus-Like Particle ,Alum adjuvant ,Antigens, Viral ,Mice, Inbred BALB C ,General Veterinary ,General Immunology and Microbiology ,Woodchuck hepatitis virus ,Public Health, Environmental and Occupational Health ,biology.organism_classification ,Virology ,Influenza A virus subtype H5N1 ,Infectious Diseases ,Viral replication ,Influenza Vaccines ,Antibody Formation ,Molecular Medicine ,Female - Abstract
The use of live recombinant attenuated Salmonella vaccines (RASV) is a promising approach for controlling infections by multiple pathogens. The highly conserved extracellular domain of the influenza M2 protein (M2e) has been shown to provide broad spectrum protection against multiple influenza subtypes sharing similar M2e sequences. An M2e epitope common to a number of avian influenza subtypes was inserted into the core antigen of woodchuck hepatitis virus and expressed in two different recombinant attenuated Salmonella Typhimurium strains. One strain was attenuated via deletion of the cya and crp genes. The second strain was engineered to exhibit a programmed delayed lysis phenotype. Both strains were able to produce both monomeric fusion proteins and fully assembled core particles. Mice orally immunized with the strain exhibiting delayed lysis induced significantly greater antibody titers than the Δcya Δcrp strain and provided moderate protection against weight loss to a low level challenge with the influenza strain A/WSN/33 modified to express the M2e sequence common to avian viruses. Further studies indicated that the Salmonella expressed core antigen induced comparable antibody levels to the purified core antigen injected with an alum adjuvant and that both are able to reduce viral replication in the lungs. To our knowledge this is the first report demonstrating Salmonella-mediated delivery of influenza virus M2e protein in a mammalian host to induce a protective immune response against viral challenge.
- Published
- 2010
259. Seoul virus suppresses NF-kappaB-mediated inflammatory responses of antigen presenting cells from Norway rats
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Rebecca Y. Au, Sabra L. Klein, Andrew Pekosz, Wei Li, and Anne E. Jedlicka
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Lipopolysaccharides ,Male ,Macrophage ,viruses ,Hantavirus Infections ,TNF ,Antigen-Presenting Cells ,Biology ,Major histocompatibility complex ,Ligands ,Virus Replication ,Article ,Immune system ,Toll-like receptor ,Interferon ,Virology ,medicine ,Animals ,Antigen-presenting cell ,Seoul virus ,Innate immunity ,Inflammation ,Rodent ,Innate immune system ,Macrophages ,Toll-Like Receptors ,NF-kappa B ,Cell Differentiation ,Dendritic Cells ,Rats ,Poly I-C ,Viral replication ,Viral persistence ,Rats, Inbred Lew ,Immunology ,Host-Pathogen Interactions ,biology.protein ,Cytokines ,Tumor necrosis factor alpha ,MHC ,Hantavirus ,medicine.drug - Abstract
Hantavirus infection reduces antiviral defenses, increases regulatory responses, and causes persistent infection in rodent hosts. To address whether hantaviruses alter the maturation and functional activity of antigen presenting cells (APCs), rat bone marrow-derived dendritic cells (BMDCs) and macrophages (BMDMs) were generated and infected with Seoul virus (SEOV) or stimulated with TLR ligands. SEOV infected both DCs and macrophages, but copies of viral RNA, viral antigen, and infectious virus titers were higher in macrophages. The expression of MHCII and CD80, production of IL-6, IL-10, and TNF-α, and expression of Ifnβ were attenuated in SEOV-infected APCs. Stimulation of APCs with poly I:C prior to SEOV infection increased the expression of activation markers and production of inflammatory cytokines and suppressed SEOV replication. Infection of APCs with SEOV suppressed LPS-induced activation and innate immune responses. Hantaviruses reduce the innate immune response potential of APCs derived from a natural host, which may influence persistence of these zoonotic viruses in the environment.
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- 2009
260. SARS Coronavirus Accessory Gene Expression and Function
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Scott R. Schaecher and Andrew Pekosz
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Genetics ,biology ,viruses ,Structural gene ,medicine.disease_cause ,biology.organism_classification ,Genome ,Virology ,Viral replication ,Gene expression ,medicine ,Coronaviridae ,Gene ,Coronavirus ,Subgenomic mRNA - Abstract
Coronavirus genomes are single-stranded positive-sense RNA that are transcribed into a nested set of 3′ coterminal subgenomic RNAs for gene expression. Members of the Coronaviridae express canonical polymerase genes, as well as structural genes, including S, E, M, and N, but also express a highly divergent set of accessory genes whose open reading frames are interspersed among the structural genes within the 3′ one-third of the viral genome. The accessory genes are thought to contain “luxury” functions that are often not required for in-vitro virus replication. The severe acute respiratory syndrome coronavirus (SARS-CoV) expresses eight such accessory genes (ORF3a, -3b, -6, -7a, -7b, -8a, -8b, and -9b), the most of any known coronavirus. This chapter will review our current knowledge of expression, structure, and function of each of the SARS-CoV accessory genes.
- Published
- 2009
261. The cholesterol recognition/interaction amino acid consensus motif of the influenza A virus M2 protein is not required for virus replication but contributes to virulence
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Wai Hong Wu, Erin N. Lalime, Shaun M. Stewart, and Andrew Pekosz
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viruses ,Amino Acid Motifs ,Virulence ,Pathogenesis ,Biology ,medicine.disease_cause ,Virus Replication ,Virus ,Article ,law.invention ,Cell Line ,Viral Matrix Proteins ,03 medical and health sciences ,Mice ,Orthomyxoviridae Infections ,law ,Virology ,Consensus Sequence ,Influenza A virus ,medicine ,Animals ,Humans ,M2 protein ,Cells, Cultured ,030304 developmental biology ,chemistry.chemical_classification ,0303 health sciences ,Mice, Inbred BALB C ,Virus Assembly ,030302 biochemistry & molecular biology ,Epithelial Cells ,CRAC ,In vitro ,Influenza ,3. Good health ,Amino acid ,Trachea ,Cholesterol ,chemistry ,Viral replication ,Recombinant DNA ,Female ,Glycoprotein ,Cholesterol binding - Abstract
Influenza A virus particles assemble and bud from plasma membrane domains enriched with the viral glycoproteins but only a small fraction of the total M2 protein is incorporated into virus particles when compared to the other viral glycoproteins. A membrane proximal cholesterol recognition/interaction amino acid consensus (CRAC) motif was previously identified in M2 and suggested to play a role in protein function. We investigated the importance of the CRAC motif on virus replication by generating recombinant proteins and viruses containing amino acid substitutions in this motif. Alteration or completion of the M2 CRAC motif in two different virus strains caused no changes in virus replication in vitro. Viruses lacking an M2 CRAC motif had decreased morbidity and mortality in the mouse model of infection, suggesting that this motif is a virulence determinant which may facilitate virus replication in vivo but is not required for basic virus replication in tissue culture.
- Published
- 2009
262. Reverse genetics of negative-strand RNA viruses: Closing the circle
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Robert A. Lamb, Andrew Pekosz, and Biao He
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Genetics ,Multidisciplinary ,viruses ,RNA-dependent RNA polymerase ,RNA virus ,Biology ,biology.organism_classification ,Virology ,H5N1 genetic structure ,Virus ,Viral replication ,Viral evolution ,Reassortant Viruses ,Oncovirus - Abstract
The study of viruses and their interactions with host cells and organisms has benefited greatly from the ability to engineer specific mutations into viral genomes, a technique known as reverse genetics. Genome manipulations of DNA viruses, either by transfecting cells with plasmids encoding the viral genome (1) or by heterologous recombination of plasmids bearing viral sequences with the virus genome (2–4), were the first to be performed. Positive-strand RNA virus genome manipulation followed quickly, partly because the viral genome is also mRNA sense. Simply transfecting plasmids, or RNA transcribed from plasmids, containing the poliovirus genome into susceptible cells resulted in the recovery of infectious poliovirus (5, 6). The negative-strand RNA viruses include a number of human and animal pathogens such as influenza A, B, and C viruses, hantaviruses, Lassa virus, rabies virus, Ebola virus, Marburg virus, measles virus, canine distemper virus, rinderpest virus, respiratory syncytial virus, mumps virus, human parainfluenza virus types 1–4, and Nipah virus (which recently emerged in Malaysia, causing respiratory distress and encephalitis in pigs and humans). However, the genomes of the negative-strand RNA viruses have been less amenable to artificial manipulation for several reasons: (i) precise 5′ and 3′ ends are required for replication and packaging of the genomic RNA; (ii) the viral RNA polymerase is essential for transcribing both mRNA and complementary, positive-sense antigenome template RNA; and (iii) both genomic and antigenomic RNAs exist as viral ribonucleoprotein (RNP) complexes (reviewed in ref. 7). The segmented genomes of influenza viruses, bunyaviruses, and arenaviruses allowed some genetic manipulation through the isolation of reassortant viruses, but manipulation of the complete genome of segmented negative-strand RNA viruses has progressed slowly, hampered by the very fact that the genome is segmented.
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- 1999
263. Sialic acid recognition is a key determinant of influenza A virus tropism in murine trachea epithelial cell cultures
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Andrew Pekosz, Pulkit S. Bose, Celeste M. Newby, and Andrew Lutz
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Mutation, Missense ,Hemagglutinin (influenza) ,Glutamic Acid ,Hemagglutinins, Viral ,Virus Attachment ,Sialic acid binding ,Viral Plaque Assay ,medicine.disease_cause ,Virus Replication ,Tropism ,Article ,Cell Line ,03 medical and health sciences ,chemistry.chemical_compound ,Mice ,Dogs ,Virology ,Receptors ,Influenza A virus ,medicine ,Animals ,Hemagglutinin ,Cells, Cultured ,030304 developmental biology ,chemistry.chemical_classification ,0303 health sciences ,Mice, Inbred BALB C ,biology ,030306 microbiology ,Hemagglutination ,Influenza A Virus, H3N2 Subtype ,Epithelial Cells ,Glutamic acid ,Molecular biology ,Influenza ,Sialic acid ,N-Acetylneuraminic Acid ,3. Good health ,Amino acid ,Trachea ,Biochemistry ,chemistry ,Amino Acid Substitution ,Respiratory epithelium ,biology.protein ,N-Acetylneuraminic acid - Abstract
Influenza A virus interacts with specific types of sialic acid during attachment and entry into susceptible cells. The precise amino acids in the hemagglutinin protein that control sialic acid binding specificity and affinity vary among antigenic subtypes. For H3 subtypes, amino acids 226 and 228 are critical for differentiating between alpha2,3- and alpha2,6-linked forms of sialic acid (SA). We demonstrate that position 190 of the HA from A/Udorn/307/72 (H3N2) plays an important role in the recognition of alpha2,3-SA, as changing the residue from a glutamic acid to an aspartic acid led to alteration of red blood cell hemagglutination and a complete loss of replication in differentiated, murine trachea epithelial cell cultures which express only alpha2,3-SA. This amino acid change had a minimal effect on virus replication in MDCK cells, suggesting subtle changes in receptor recognition by the H3 hemagglutinin can lead to significant alterations in cell and species tropism.
- Published
- 2008
264. Emerging viral diseases
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Andrew, Pekosz and Gregory E, Glass
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Birds ,Virus Diseases ,Animals ,Humans ,Global Health ,Antiviral Agents ,Communicable Diseases, Emerging ,Article - Published
- 2008
265. Extending the Cytoplasmic Tail of the Influenza A Virus M2 Protein Leads to Reduced Virus Replication In Vivo but Not In Vitro▿
- Author
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Wai Hong Wu and Andrew Pekosz
- Subjects
viruses ,Immunology ,Orthomyxoviridae ,Virulence ,Recombinant virus ,medicine.disease_cause ,Virus Replication ,Microbiology ,Epitope ,Virus ,Cell Line ,Viral Matrix Proteins ,Mice ,Dogs ,Orthomyxoviridae Infections ,Virology ,Influenza A virus ,medicine ,Animals ,Mice, Inbred BALB C ,Viral matrix protein ,biology ,Body Weight ,biology.organism_classification ,Survival Analysis ,Viral replication ,Insect Science ,Pathogenesis and Immunity - Abstract
A carboxy-terminal epitope tag introduced into the coding region of the A/WSN/33 M2 protein resulted in a recombinant virus (rWSN M2myc) which replicated to titers similar to those of the parental virus (rWSN) in MDCK cells. The rWSN M2myc virus was attenuated in its ability to induce mortality and weight loss after the intranasal inoculation of BALB/c mice, indicating that the M2 cytoplasmic tail plays a role in virus virulence. Mice infected with rWSN M2myc were completely protected from subsequent challenge with rWSN, suggesting that epitope tagging of the M2 protein may be a useful way of attenuating influenza A virus strains.
- Published
- 2007
266. IFN-stimulated gene 15 functions as a critical antiviral molecule against influenza, herpes, and Sindbis viruses
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Deborah J. Lenschow, Andrew Lutz, Natalia Frias-Staheli, Nadia V. Giannakopoulos, Thorsten Wolff, Klaus Peter Knobeloch, Beth Levine, David A. Leib, Ivan Horak, Anna Osiak, Andrew Pekosz, Adolfo García-Sastre, Caroline Lai, Herbert W. Virgin, and Robert E. Schmidt
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Male ,Sindbis virus ,Viral pathogenesis ,viruses ,Orthomyxoviridae ,medicine.disease_cause ,Lymphocytic choriomeningitis ,Herpesviridae ,Virus ,Mice ,medicine ,Animals ,viral pathogenesis ,ubiquitin ,RNA viruses ,DNA viruses ,innate immunity ,Ubiquitins ,Mice, Knockout ,Multidisciplinary ,biology ,Biological Sciences ,medicine.disease ,biology.organism_classification ,Virology ,Herpes simplex virus ,Vesicular stomatitis virus ,Cytokines ,Disease Susceptibility ,Sindbis Virus - Abstract
Type I interferons (IFNs) play an essential role in the host response to viral infection through the induction of numerous IFN-stimulated genes (ISGs), including important antiviral molecules such as PKR, RNase L, Mx, and iNOS. Yet, additional antiviral ISGs likely exist. IFN-stimulated gene 15 (ISG15) is a ubiquitin homolog that is rapidly up-regulated after viral infection, and it conjugates to a wide array of host proteins. Although it has been hypothesized that ISG15 functions as an antiviral molecule, the initial evaluation of ISG15-deficient mice revealed no defects in their responses to vesicular stomatitis virus or lymphocytic choriomeningitis virus, leaving open the important question of whether ISG15 is an antiviral moleculein vivo. Here we demonstrate that ISG15 is critical for the host response to viral infection. ISG15−/−mice are more susceptible to influenza A/WSN/33 and influenza B/Lee/40 virus infections. ISG15−/−mice also exhibited increased susceptibility to both herpes simplex virus type 1 and murine gammaherpesvirus 68 infection and to Sindbis virus infection. The increased susceptibility of ISG15−/−mice to Sindbis virus infection was rescued by expressing wild-type ISG15, but not a mutant form of ISG15 that cannot form conjugates, from the Sindbis virus genome. The demonstration of ISG15 as a novel antiviral molecule with activity against both RNA and DNA viruses provides a target for the development of therapies against important human pathogens.
- Published
- 2007
267. Exposure to Electronic Cigarettes Impairs Pulmonary Anti-Bacterial and Anti-Viral Defenses in a Mouse Model
- Author
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Jinfang Ma, Thomas E. Sussan, Andrew Pekosz, Farhana Hasan, Rajesh K. Thimmulappa, Stephania A. Cormier, Slawo Lomnicki, Nicola Consolini, Kuladeep Sudini, Shyam Biswal, Sachin Gajghate, and Jung Hyun Kim
- Subjects
Male ,Nicotine ,Free Radicals ,lcsh:Medicine ,Inflammation ,medicine.disease_cause ,Mice ,Influenza A Virus, H1N1 Subtype ,Immune system ,Phagocytosis ,In vivo ,medicine ,Animals ,lcsh:Science ,Lung ,Multidisciplinary ,business.industry ,lcsh:R ,Smoking ,Viral Load ,medicine.disease ,3. Good health ,Mice, Inbred C57BL ,Oxidative Stress ,Pneumonia ,Streptococcus pneumoniae ,medicine.anatomical_structure ,Immunology ,lcsh:Q ,Volatilization ,medicine.symptom ,business ,Viral load ,Oxidative stress ,Research Article ,medicine.drug - Abstract
Electronic cigarettes (E-cigs) have experienced sharp increases in popularity over the past five years due to many factors, including aggressive marketing, increased restrictions on conventional cigarettes, and a perception that E-cigs are healthy alternatives to cigarettes. Despite this perception, studies on health effects in humans are extremely limited and in vivo animal models have not been generated. Presently, we determined that E-cig vapor contains 7 x 10(11) free radicals per puff. To determine whether E-cig exposure impacts pulmonary responses in mice, we developed an inhalation chamber for E-cig exposure. Mice that were exposed to E-cig vapor contained serum cotinine concentrations that are comparable to human E-cig users. E-cig exposure for 2 weeks produced a significant increase in oxidative stress and moderate macrophage-mediated inflammation. Since, COPD patients are susceptible to bacterial and viral infections, we tested effects of E-cigs on immune response. Mice that were exposed to E-cig vapor showed significantly impaired pulmonary bacterial clearance, compared to air-exposed mice, following an intranasal infection with Streptococcus pneumonia. This defective bacterial clearance was partially due to reduced phagocytosis by alveolar macrophages from E-cig exposed mice. In response to Influenza A virus infection, E-cig exposed mice displayed increased lung viral titers and enhanced virus-induced illness and mortality. In summary, this study reports a murine model of E-cig exposure and demonstrates that E-cig exposure elicits impaired pulmonary anti-microbial defenses. Hence, E-cig exposure as an alternative to cigarette smoking must be rigorously tested in users for their effects on immune response and susceptibility to bacterial and viral infections.
- Published
- 2015
268. Structure, expression, and intracellular localization of the SARS-CoV accessory proteins 7a and 7b
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Andrew, Pekosz, Scott R, Schaecher, Michael S, Diamond, Daved H, Fremont, Amy C, Sims, and Ralph S, Baric
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Microscopy, Confocal ,Severe Acute Respiratory Syndrome Coronavirus ,Amino Acid Motifs ,Golgi Apparatus ,Biological Transport ,Endoplasmic Reticulum ,Transfection ,Severe Acute Respiratory Syndrome ,Severe Acute Respiratory Distress Syndrome ,Article ,Viral Matrix Proteins ,Open Reading Frames ,Viral Proteins ,Severe acute respiratory syndrome-related coronavirus ,Mutagenesis ,Chlorocebus aethiops ,Vero Cell ,Animals ,Vero Cells ,Severe Acute Respiratory Syndrome Patient - Published
- 2006
269. Production and Characterization of Monoclonal Antibodies Against the Nucleocapsid Protein of SARS-COV
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Lia M. Haynes, Ying Fang, Andrew Pekosz, Eric A. Nelson, and Raymond R. R. Rowland
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medicine.drug_class ,Vero cell ,medicine ,Biology ,Monoclonal antibody ,Virology - Published
- 2006
270. Structure, Expression, and Intracellular Localization of the SARS-CoV Accessory Proteins 7a and 7b
- Author
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Amy C. Sims, Ralph S. Baric, Andrew Pekosz, Michael S. Diamond, Scott R. Schaecher, and Daved H. Fremont
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Open reading frame ,Amino Acid Motifs ,Intracellular localization ,Vero cell ,Mutagenesis (molecular biology technique) ,Transfection ,Severe acute respiratory syndrome coronavirus ,Biology ,Cell biology - Published
- 2006
271. Intracellular Localization of the Severe Acute Respiratory Syndrome Coronavirus Nucleocapsid Protein: Absence of Nucleolar Accumulation during Infection and after Expression as a Recombinant Protein in Vero Cells
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Maureen A. Kerrigan, Vinita Chauhan, Miriam D. Burton, Ying Fang, Andrew Pekosz, and Raymond R. R. Rowland
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Nucleolus ,cells ,viruses ,Immunology ,Amino Acid Motifs ,Molecular Sequence Data ,Nuclear Localization Signals ,Biology ,medicine.disease_cause ,Microbiology ,environment and public health ,Virology ,Chlorocebus aethiops ,medicine ,Coronaviridae ,Animals ,Coronavirus Nucleocapsid Proteins ,Amino Acid Sequence ,Vero Cells ,Coronavirus ,Nucleoplasm ,fungi ,Transfection ,Nucleocapsid Proteins ,biology.organism_classification ,Molecular biology ,Protein subcellular localization prediction ,Recombinant Proteins ,Virus-Cell Interactions ,Severe acute respiratory syndrome-related coronavirus ,Insect Science ,Vero cell ,Nuclear localization sequence - Abstract
The nucleocapsid (N) protein of several members within the order Nidovirales localizes to the nucleolus during infection and after transfection of cells with N genes. However, confocal microscopy of N protein localization in Vero cells infected with the severe acute respiratory syndrome coronavirus (SARS-CoV) or transfected with the SARS-CoV N gene failed to show the presence of N in the nucleoplasm or nucleolus. Amino acids 369 to 389, which contain putative nuclear localization signal (NLS) and nucleolar localization signal motifs, failed to restore nuclear localization to an NLS-minus mutant Rev protein. These data indicate that nuclear localization is not a conserved property among all nidoviruses.
- Published
- 2005
272. Differentiated cultures of primary hamster tracheal airway epithelial cells
- Author
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Regina K. Rowe, Andrew Pekosz, and Steven L. Brody
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Time Factors ,Cellular differentiation ,Blotting, Western ,Cell Culture Techniques ,Hamster ,Fluorescent Antibody Technique ,Biology ,Mucin 5AC ,Article ,Tubulin ,Cricetinae ,medicine ,Electric Impedance ,Animals ,Uteroglobin ,Wound Healing ,Mesocricetus ,Mucins ,Cell Differentiation ,Epithelial Cells ,Cell Biology ,General Medicine ,respiratory system ,biology.organism_classification ,Mucus ,Epithelium ,Cell biology ,Culture Media ,Trachea ,Microscopy, Electron ,medicine.anatomical_structure ,Cell culture ,Influenza A virus ,Matrix Metalloproteinase 7 ,Immunology ,Electrophoresis, Polyacrylamide Gel ,Female ,Stem cell ,Wound healing ,Developmental Biology - Abstract
Primary airway epithelial cell cultures can provide a faithful representation of the in vivo airway while allowing for a controlled nutrient source and isolation from other tissues or immune cells. The methods used have significant differences based on tissue source, cell isolation, culture conditions, and assessment of culture purity. We modified and optimized a method for generating tracheal epithelial cultures from Syrian golden hamsters and characterized the cultures for cell composition and function. Soon after initial plating, the epithelial cells reached a high transepithelial resistance and formed tight junctions. The cells differentiated into a heterogeneous, multicellular culture containing ciliated, secretory, and basal cells after culture at an air-liquid interface (ALI). The secretory cell populations initially consisted of MUC5AC-positive goblet cells and MUC5AC/CCSP double-positive cells, but the makeup changed to predominantly Clara cell secretory protein (CCSP)-positive Clara cells after 14 d. The ciliated cell populations differentiated rapidly after ALI, as judged by the appearance of beta tubulin IV-positive cells. The cultures produced mucus, CCSP, and trypsin-like proteases and were capable of wound repair as judged by increased expression of matrilysin. Our method provides an efficient, high-yield protocol for producing differentiated hamster tracheal epithelial cells that can be used for a variety of in vitro studies including tracheal cell differentiation, airway disease mechanisms, and pathogen-host interactions.
- Published
- 2005
273. The Influenza A Virus M2 Cytoplasmic Tail Is Required for Infectious Virus Production and Efficient Genome Packaging
- Author
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Matthew Mccown and Andrew Pekosz
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viruses ,Immunology ,Orthomyxoviridae ,Viral transformation ,Genome, Viral ,medicine.disease_cause ,Virus Replication ,Microbiology ,Virus ,Viral Matrix Proteins ,VP40 ,Dogs ,Viral entry ,Virology ,Influenza A virus ,medicine ,Animals ,Cells, Cultured ,Sequence Deletion ,biology ,Structure and Assembly ,Viral Core Proteins ,Virus Assembly ,Genetic Complementation Test ,RNA-Binding Proteins ,Nucleocapsid Proteins ,biology.organism_classification ,Protein Structure, Tertiary ,NS2-3 protease ,Nucleoproteins ,Insect Science ,Helper virus ,RNA, Viral - Abstract
The M2integral membrane protein encoded by influenza A virus possesses an ion channel activity that is required for efficient virus entry into host cells. The role of the M2protein cytoplasmic tail in virus replication was examined by generating influenza A viruses encoding M2proteins with truncated C termini. Deletion of 28 amino acids (M2Stop70) resulted in a virus that produced fourfold-fewer particles but >1,000-fold-fewer infectious particles than wild-type virus. Expression of the full-length M2protein intransrestored the replication of the M2truncated virus. Although the M2Stop70 virus particles were similar to wild-type virus in morphology, the M2Stop70 virions contained reduced amounts of viral nucleoprotein and genomic RNA, indicating a defect in vRNP packaging. The data presented indicate the M2cytoplasmic tail plays a role in infectious virus production by coordinating the efficient packaging of genome segments into influenza virus particles.
- Published
- 2005
274. Virus-inducible reporter genes as a tool for detecting and quantifying influenza A virus replication
- Author
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Julie Dyall, Andrew Pekosz, Andrew Lutz, and Paul D. Olivo
- Subjects
medicine.drug_class ,viruses ,Orthomyxoviridae ,Green Fluorescent Proteins ,Gene Expression ,Microbial Sensitivity Tests ,medicine.disease_cause ,Virus Replication ,Virus ,Article ,Cell Line ,Dogs ,Genes, Reporter ,Luciferases, Firefly ,RNA Polymerase I ,Untranslated Regions ,Virology ,Cricetinae ,medicine ,Influenza A virus ,Animals ,Serotyping ,Promoter Regions, Genetic ,Polymerase ,Reporter gene ,biology ,Staining and Labeling ,biology.organism_classification ,Molecular biology ,Nucleoprotein ,Viral replication ,biology.protein ,RNA, Viral ,Antiviral drug - Abstract
The use of influenza A virus-inducible reporter gene segments in detecting influenza A virus replication was investigated. The RNA polymerase I promoter/terminator cassette was used to express RNA transcripts encoding green fluorescence protein or firefly luciferase flanked by the untranslated regions of the influenza A/WSN/33 nucleoprotein (NP) segment. Reporter gene activity was detected after reconstitution of the influenza A virus polymerase complex from cDNA or after virus infection, and was influenza A virus-specific. Reporter gene activity could be detected as early as 6 h post-infection and was virus dose-dependent. Inhibitory effects of antibodies or amantadine could be detected and quantified rapidly, providing a means of not only identifying influenza A virus-specific replication, but also of determining the antigenic subtype as well as antiviral drug susceptibility. Induction of virus-specific reporter genes provides a rapid, sensitive method for detecting virus replication, quantifying virus titers and assessing antiviral sensitivity as well as antigenic subtype.
- Published
- 2004
275. The utility of siRNA transcripts produced by RNA polymerase i in down regulating viral gene expression and replication of negative- and positive-strand RNA viruses
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Andrew Pekosz, Matthew Mccown, and Michael S. Diamond
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Gene Expression Regulation, Viral ,RNA-induced transcriptional silencing ,Transcription, Genetic ,viruses ,Trans-acting siRNA ,RNA-dependent RNA polymerase ,Biology ,Virus Replication ,Cell Line ,Small hairpin RNA ,Viral Matrix Proteins ,Mice ,antivirals ,Transformation, Genetic ,RNA Polymerase I ,Virology ,Cricetinae ,Gene expression ,Animals ,Humans ,RNA, Small Interfering ,RNA ,Molecular biology ,RNA silencing ,Regulatory sequence ,Influenza A virus ,gene expression ,RNA, Viral ,RNA Interference ,West Nile virus - Abstract
Short interfering double-stranded RNAs (siRNAs) expressed under the control of an RNA polymerase I promoter system were used to target gene expression of influenza A and West Nile virus. Decreased RNA and protein expression was induced in a sequence-specific manner—reducing sequence complementarity from 21 to 17 nucleotides abrogated the siRNA effect. Reduced M2 expression resulted in a decrease in total and infectious influenza A virus production. WNV protein expression, genomic RNA, and infectious virus production were all dramatically reduced by siRNAs targeting two distinct viral sequences. The data demonstrate the utility of plasmid-driven siRNAs in regulating the expression of single viral genes, global viral gene expression, as a potential antiviral treatment, and as a genetic tool for viruses whose genomes are difficult to manipulate.
- Published
- 2003
276. Influenza a virus M2 ion channel activity is essential for efficient replication in tissue culture
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Makoto Takeda, Andrew Pekosz, Lawrence H. Pinto, Robert A. Lamb, and Kevin Shuck
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Genes, Viral ,medicine.drug_class ,viruses ,Immunology ,Replication ,Biology ,medicine.disease_cause ,Virus Replication ,Microbiology ,Antiviral Agents ,Virus ,Ion Channels ,Cell Line ,Viral Matrix Proteins ,Dogs ,Viral life cycle ,Virology ,Culture Techniques ,Influenza A virus ,medicine ,Amantadine ,Serine ,Animals ,Humans ,Cell Line, Transformed ,Viral matrix protein ,Viral Core Proteins ,Virus Uncoating ,RNA-Binding Proteins ,Nucleocapsid Proteins ,Nucleoprotein ,Nucleoproteins ,Viral replication ,Insect Science ,Protein Biosynthesis ,Antiviral drug ,Asparagine - Abstract
The amantadine-sensitive ion channel activity of influenza A virus M 2 protein was discovered through understanding the two steps in the virus life cycle that are inhibited by the antiviral drug amantadine: virus uncoating in endosomes and M 2 protein-mediated equilibration of the intralumenal pH of the trans Golgi network. Recently it was reported that influenza virus can undergo multiple cycles of replication without M 2 ion channel activity (T. Watanabe, S. Watanabe, H. Ito, H. Kida, and Y. Kawaoka, J. Virol. 75:5656–5662, 2001). An M 2 protein containing a deletion in the transmembrane (TM) domain (M 2 -del 29–31 ) has no detectable ion channel activity, yet a mutant virus was obtained containing this deletion. Watanabe and colleagues reported that the M 2 -del 29–31 virus replicated as efficiently as wild-type (wt) virus. We have investigated the effect of amantadine on the growth of four influenza viruses: A/WSN/33; N 31 S-M 2 WSN, a mutant in which an asparagine residue at position 31 in the M 2 TM domain was replaced with a serine residue; MUd/WSN, which possesses seven RNA segments from WSN plus the RNA segment 7 derived from A/Udorn/72; and A/Udorn/72. N 31 S-M 2 WSN was amantadine sensitive, whereas A/WSN/33 was amantadine resistant, indicating that the M 2 residue N 31 is the sole determinant of resistance of A/WSN/33 to amantadine. The growth of influenza viruses inhibited by amantadine was compared to the growth of an M 2 -del 29–31 virus. We found that the M 2 -del 29–31 virus was debilitated in growth to an extent similar to that of influenza virus grown in the presence of amantadine. Furthermore, in a test of biological fitness, it was found that wt virus almost completely outgrew M 2 -del 29–31 virus in 4 days after cocultivation of a 100:1 ratio of M 2 -del 29–31 virus to wt virus, respectively. We conclude that the M 2 ion channel protein, which is conserved in all known strains of influenza virus, evolved its function because it contributes to the efficient replication of the virus in a single cycle.
- Published
- 2002
277. The cytoplasmic tails of the influenza virus spike glycoproteins are required for normal genome packaging
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Robert A. Lamb, Jie Zhang, George P. Leser, and Andrew Pekosz
- Subjects
Cytoplasm ,Ultraviolet Rays ,viruses ,Population ,Hemagglutinin (influenza) ,Neuraminidase ,Genome, Viral ,medicine.disease_cause ,Virus ,03 medical and health sciences ,Structure-Activity Relationship ,Dogs ,Virology ,Cricetinae ,Influenza A virus ,medicine ,Centrifugation, Density Gradient ,Animals ,education ,030304 developmental biology ,Infectivity ,chemistry.chemical_classification ,0303 health sciences ,education.field_of_study ,biology ,030306 microbiology ,Virus Assembly ,Virion ,RNA ,Molecular biology ,Microscopy, Electron ,Hemagglutinins ,chemistry ,biology.protein ,Glycoprotein - Abstract
Deletion of the cytoplasmic tails of the influenza A virus spike glycoproteins, hemagglutinin (HA) and neuraminidase (NA), has previously been shown to result in markedly defective virion morphogenesis (Jin et al., 1997, EMBO J. 16, 1236–1247). We have found that influenza A virus preparations lacking the HA and NA cytoplasmic tails (HAt−/NAt−) have a reduced vRNA to protein content, contain an increase in cellular RNA contaminants, and exhibit increased resistance to ultraviolet (UV) inactivation. There is also a direct correlation between abnormal virion morphology and reduced infectivity. The data suggest that the HAt−/NAt− virion population contains a broader range of number of packaged RNA segments than wild-type (wt) virus. Sucrose gradient centrifugation analysis indicated the presence of a subpopulation of virions with pronounced deformation in virion morphology and reduced infectivity. The role of the HA and NA cytoplasmic tails was examined further by using a trans-complementation assay and it was found that expression of wt HA and NA from cDNAs followed by HAt−/NAt− virus infection caused the formation of a pseudotype virus with wt sedimentation properties. Taken together the data indicate that the HA and NA cytoplasmic tails affect not only virion morphology but also proper genome packaging.
- Published
- 2001
278. Influenza Virus Assembly and Lipid Raft Microdomains: a Role for the Cytoplasmic Tails of the Spike Glycoproteins
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Jie Zhang, Robert A. Lamb, and Andrew Pekosz
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Influenzavirus C ,genetic structures ,viruses ,Immunology ,Detergents ,Hemagglutinin (influenza) ,Neuraminidase ,Hemagglutinin Glycoproteins, Influenza Virus ,medicine.disease_cause ,Microbiology ,Virus ,Cell Line ,Viral Matrix Proteins ,Membrane Lipids ,Dig ,Virology ,Influenza A virus ,medicine ,Animals ,Humans ,chemistry.chemical_classification ,Viral matrix protein ,Membrane Glycoproteins ,biology ,Structure and Assembly ,Virus Assembly ,Virion ,Cell Polarity ,Molecular biology ,eye diseases ,chemistry ,Solubility ,Insect Science ,biology.protein ,sense organs ,Glycolipids ,Influenza C Virus ,Glycoprotein ,HeLa Cells - Abstract
Influenza viruses encoding hemagglutinin (HA) and neuraminidase (NA) glycoproteins with deletions in one or both cytoplasmic tails (HAt− or NAt−) have a reduced association with detergent-insoluble glycolipids (DIGs). Mutations which eliminated various combinations of the three palmitoylation sites in HA exhibited reduced amounts of DIG-associated HA in virus-infected cells. The influenza virus matrix (M 1 ) protein was also found to be associated with DIGs, but this association was decreased in cells infected with HAt− or NAt− virus. Regardless of the amount of DIG-associated protein, the HA and NA glycoproteins were targeted primarily to the apical surface of virus-infected, polarized cells. The uncoupling of DIG association and apical transport was augmented by the observation that the influenza A virus M 2 protein as well as the influenza C virus HA-esterase-fusion glycoprotein were not associated with DIGs but were apically targeted. The reduced DIG association of HAt− and NAt− is an intrinsic property of the glycoproteins, as similar reductions in DIG association were observed when the proteins were expressed from cDNA. Examination of purified virions indicated reduced amounts of DIG-associated lipids in the envelope of HAt− and NAt− viruses. The data indicate that deletion of both the HA and NA cytoplasmic tails results in reduced DIG association and changes in both virus polypeptide and lipid composition.
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- 2000
279. Mechanism for proton conduction of the M(2) ion channel of influenza A virus
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Andrew Pekosz, Hui Chun Li, Lawrence H. Pinto, Robert A. Lamb, Christine S. Dudlak, James D. Lear, and Jorgen A. Mould
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Patch-Clamp Techniques ,Hydronium ,Voltage clamp ,Morpholines ,Xenopus ,Analytical chemistry ,Buffers ,Biochemistry ,Ion Channels ,Ion ,Viral Matrix Proteins ,chemistry.chemical_compound ,Proton transport ,Amantadine ,Animals ,Patch clamp ,Deuterium Oxide ,Molecular Biology ,Ion channel ,biology ,Chemistry ,Electric Conductivity ,Water ,Cell Biology ,Hydrogen-Ion Concentration ,Recombinant Proteins ,Membrane ,M2 proton channel ,Alkanesulfonic Acids ,Influenza A virus ,biology.protein ,Oocytes ,Protons - Abstract
The M(2) integral membrane protein of influenza A virus forms a proton-selective ion channel. We investigated the mechanism for proton transport of the M(2) protein in Xenopus oocytes using a two-electrode voltage clamp and in CV-1 cells using the whole cell patch clamp technique. Membrane currents were recorded while manipulating the external solution to alter either the total or free proton concentration or the solvent itself. Membrane conductance decreased by approximately 50% when D(2)O replaced H(2)O as the solvent. From this, we conclude that hydrogen ions do not pass through M(2) as hydronium ions, but instead must interact with titratable groups that line the pore of the channel. M(2) currents measured in solutions of low buffer concentration (15 mM in oocytes and0.15 mM in CV-1 cells) were smaller than those studied in solutions of high buffer concentration. Furthermore, the reversal voltage measured in low buffer was shifted to a more negative voltage than in high buffer. Also, at a given pH, M(2) current amplitude in 15 mM buffer decreased when pH-pK(a) was increased by changing the buffer pK(a). Collectively, these results demonstrate that M(2) currents can be limited by external buffer capacity. The data presented in this study were also used to estimate the maximum single channel current of the M(2) ion channel, which was calculated to be on the order of 1-10 fA.
- Published
- 2000
280. The extracellular domain of La Crosse virus G1 forms oligomers and undergoes pH-dependent conformational changes
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Francisco Gonzalez-Scarano and Andrew Pekosz
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Cytoplasm ,La Crosse virus ,medicine.drug_class ,Protein Conformation ,Succinimides ,Biology ,Monoclonal antibody ,Oligomer ,chemistry.chemical_compound ,Virology ,medicine ,Extracellular ,Centrifugation, Density Gradient ,Humans ,Sequence Deletion ,chemistry.chemical_classification ,Cell Membrane ,Virion ,Hydrogen-Ion Concentration ,Precipitin Tests ,Transmembrane protein ,Monomer ,Cross-Linking Reagents ,chemistry ,Biochemistry ,Solubility ,Biophysics ,Glycoprotein ,Dimerization ,Viral Fusion Proteins - Abstract
The La Crosse virus G1 glycoprotein plays a critical role in virus binding to susceptible cells and in the subsequent fusion of viral and cellular membranes. A soluble form of the G1 glycoprotein (sG1) prepared in a recombinant baculovirus system mimics the cell-binding pattern of La Crosse virus and inhibits La Crosse virus infection (A. Pekosz et al., Virology 214, 339–348, 1995), presumably by competing for a cellular receptor, a finding that implies that sG1 can perform some functions absent G2, the smaller of the two bunyavirus glycoproteins. We have performed experiments to determine whether sG1 is present as an oligomer and whether it undergoes the conformational changes associated with fusion (F. Gonzalez-Scarano, Virology 140, 209–216, 1985). Our results indicate that both sG1 and native G1 undergo similar changes in conformation after exposure to an acidic environment, as detected by reactivity with monoclonal antibodies. Furthermore, using chemical cross-linking, both proteins were detected as oligomers (most likely dimers). Sucrose density gradient analysis of sG1 verified that it was present in monomeric and oligomeric forms. These results demonstrate that the isolated G1 glycoprotein can undergo a pH-dependent change in conformation in the absence of its transmembrane and cytoplasmic tail domains and that the extracellular portion of the glycoprotein can oligomerize.
- Published
- 1996
281. Protective human MAbs to a highly conserved conformational determinant on influenza M2e (38.14)
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Andres Grandea, Ole Olsen, Tom Cox, Mark Renshaw, Philip Hammond, Po-Ying Chan-Hui, Jennifer Mitcham, Witold Cieplak, Shaun Stewart, Michael Grantham, Andrew Pekosz, Masato Hatta, Yoshihiro Kawaoka, and Matthew Moyle
- Subjects
Immunology ,Immunology and Allergy - Abstract
Specific antibody responses to the annual influenza vaccine can protect against infection if the vaccine and the infective strain are antigenically well-matched. However, due to the variability of the influenza genome, emergent strains arise for which the vaccine proves to be ineffective, and moreover, necessitate its yearly reformulation. Efforts have been made into identifying epitopes within more conserved elements of the influenza virus as an alternative attack on a wider range of strains. Through our platform of examining monoclonal antibodies (MAbs) expressed from plasmacytes derived from memory B cells from normal healthy human donors, we have found a previously unknown conformational antigenic determinant within the relatively invariable ectodomain of M2 (M2e) of influenza A. The MAbs bind to virus and cell-surface expressed M2, but not to the corresponding M2e peptide. The M2e region recognized is in almost all strains isolated to date. Furthermore, in a mouse model of infection, the MAbs can protect against lethal challenge. These results suggest that naturally expressed M2 can elicit a broadly cross-reactive and protective antibody response. And, the human recombinant anti-M2e MAbs might serve as therapeutic agents against infection.
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- 2010
282. Immune Response to Seasonal Influenza Vaccination (92.18)
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Amritha Ramakrishnan, Keri Altoff, Andrew Pekosz, and Jay Bream
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Immunology ,Immunology and Allergy - Abstract
The emergence of new pandemic strains of influenza highlights the need to better understand the immune response to vaccination in order to develop novel vaccine strategies. The goal of our study was to identify and assess novel markers of immune activation after vaccination. To this end, 100 healthy adults who received either the seasonal Live Attenuated Vaccine (LAIV) or the Inactivated Vaccine (TIV) were enrolled from 2006-2008. First, we measured the serum antibody titers against vaccine strains of the virus. We found that while TIV induced a robust increase in anti-influenza serum antibody titers, LAIV induced only a modest increase. We next measured the levels of 10 cytokines in the serum of vaccine recipients. Vaccination with TIV resulted in a significant reduction in the levels of TNF-α while LAIV had no effect on any of the measured cytokines. Finally, we assessed the ability of seasonal vaccination to induce neutralizing antibody titers against the pandemic 2009 H1N1 strain. Vaccination with TIV was able to induce a significant increase in cross reactive antibodies against different seasonal H1N1 strains but not against the 2009 pandemic H1N1 strain. In all, these data suggest that 1) different influenza A vaccine formulations induce unique cytokine responses in the periphery and 2) seasonal influenza vaccination does not illicit strong cross-reactive serum antibodies with 2009 H1N1.
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- 2010
283. The RNA Binding Domain of Influenza A Virus NS1 Protein Affects Secretion of Tumor Necrosis Factor Alpha, Interleukin-6, and Interferon in Primary Murine Tracheal Epithelial Cells
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Celeste M. Newby, Leah R. Sabin, and Andrew Pekosz
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Author's Correction ,Cytoplasm ,medicine.medical_treatment ,viruses ,Viral Nonstructural Proteins ,Virus Replication ,medicine.disease_cause ,Mice ,Interferon ,Chlorocebus aethiops ,Influenza A virus ,Cells, Cultured ,Mice, Inbred BALB C ,Microscopy, Confocal ,Virus-Cell Interactions ,Trachea ,Cytokine ,Female ,Tumor necrosis factor alpha ,Binding domain ,medicine.drug ,Orthomyxoviridae ,Immunology ,Mutation, Missense ,Biology ,Microbiology ,Virus ,Proinflammatory cytokine ,Cell Line ,Dogs ,Orthomyxoviridae Infections ,Virology ,medicine ,Animals ,Humans ,Secretion ,Interleukin 6 ,Vero Cells ,Interleukin-6 ,Tumor Necrosis Factor-alpha ,RNA ,Epithelial Cells ,biology.organism_classification ,Survival Analysis ,Molecular biology ,Immunity, Innate ,Protein Structure, Tertiary ,Disease Models, Animal ,Amino Acid Substitution ,Insect Science ,Vero cell ,biology.protein ,Interferons - Abstract
Primary differentiated respiratory epithelial cell cultures closely model the in vivo environment and allow for studies of innate immune responses generated specifically by epithelial cells, the primary cell type infected by human influenza A virus strains. We used primary murine tracheal epithelial cell (mTEC) cultures to investigate antiviral and cytokine responses to influenza A virus infection, focusing on the contribution of the RNA binding domain of the NS1 protein. rWSN NS1 R38A replication is attenuated in mTEC cultures; however, viral antigen is detected predominantly in ciliated cells, similar to wild-type virus. NS1 and NS1 R38A proteins display a primarily cytoplasmic localization in infected mTEC cultures. Increased production of tumor necrosis factor alpha, interleukin-6, and beta interferon is observed during rWSN NS1 R38A infection, and cytokines are secreted in a directional manner. Cytokine pretreatment of mTEC cultures and Vero cells suggest that rWSN NS1 R38A is more sensitive to the presence of antiviral/inflammatory cytokines than wild-type virus. Our results demonstrate that the RNA binding domain is a critical regulator of both cytokine production and cytokine sensitivity during influenza A virus infection of primary tracheal epithelial cells.
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- 2007
284. Bird flu: A virus of our own hatching
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Andrew Pekosz
- Subjects
education.field_of_study ,medicine.medical_specialty ,business.industry ,Public health ,Population ,virus diseases ,Outbreak ,General Medicine ,Public relations ,medicine.disease_cause ,Virology ,Influenza A virus subtype H5N1 ,Book Review ,Avian Influenza A Virus ,Infectious disease (medical specialty) ,Preparedness ,Pandemic ,medicine ,business ,education - Abstract
Are you tired of hearing about the avian influenza A virus (H5N1) and its potential threat to humankind? Are you wondering what is happening with the H5N1 virus threat? These seemingly contradictory questions are being asked frequently as time passes between the first reports of H5N1 infection of humans and its predicted emergence as a pandemic influenza virus. As the H5N1 influenza virus threat continues to simmer but not boil over, it is becoming difficult to keep interest in and focus on the issue of public health preparedness for a global infectious disease outbreak. At this critical time in the development of pandemic influenza preparedness, Michael Greger’s book Bird flu: a virus of our own hatching presents not only a comprehensive and thorough summary of the continuing threat of H5N1 influenza but a critical examination of the initial political and public health efforts that have been developed in response to the spread of human cases of H5N1 infection across Asia, Europe, and Africa. The real strength of Bird flu lies in its comprehensive summary of the H5N1 outbreak. The book is divided into five sections: an introduction to the biology and history of influenza; a discussion of the social and economic factors that are responsible for the increased threat of animal viruses to the human population; a summary of the pandemic preparedness measures that are being put into place in response to the emergence of H5N1; a section on measures the individual can take to prepare for an influenza pandemic; and finally, comments on the prevention of future pandemics. It is the breadth of topics covered that makes this book interesting to a broad range of readers. Perhaps the most impressive aspect of the book is its myriad of carefully documented references and quotes from key scientific, public health, and government officials, which indicates that Greger has gone to the primary sources to gain his information. While these topics are covered in great depth, Greger does have a penchant for turning to the colloquial when describing some of the scientific and public health principles important to understanding the evolving threat of pandemic influenza. Perhaps it is this particular style that will make the book very readable and attractive to nonscientists, but statements such as “some like it hot but not influenza” when referring to the effects of elevated temperature on influenza virus replication or how the body’s cells sometimes “take one for the team and jump on a grenade” when describing the effects of innate immune factors such as interferon on the body’s own cells began to grate a bit on this particular reader. The largest amount of text is dedicated to a discussion of the environmental, economic, and social factors that contributed to the emergence of H5N1 as a threat to humans. Greger gives us a comprehensive overview of influenza virus ecology and the factors that led to increased contact of humans with birds and the viruses that they harbor. While maintaining focus on H5N1 virus, Greger comments extensively on other examples of animal pathogens that became significant human pathogens. This is a key point in the book, as the environment that led to the emergence of the H5N1 virus can also foster the development of other animal pathogens into human pathogens. Greger is not afraid to discuss controversial topics such as the personal stockpiling of influenza antivirals and the role of socioeconomic class in gaining access to the antivirals that are and will be in critical supply when an influenza pandemic occurs. He even includes recipes for homemade hand sanitizer and an interesting strategy for “readministering” Tamiflu, which may be a bit hard for some to swallow (pun intended!) but may be one way to stretch our effective supply of this key antiviral. However, he also includes in-depth discussions on roles for hygiene, quarantine, isolation, and the economic effects of disrupting just-in-time inventory practices and how they may impact our ability to deal with an influenza pandemic. It is the careful, balanced, and comprehensive discussion of these topics that provides the reader with all the relevant facts and makes the book informative but not alarmist in tone. Will avian H5N1 be the cause of the next influenza pandemic or will some other influenza virus strain catch us unawares? Will the next influenza pandemic resemble the 1918, 1957, or 1968 pandemics? Will the next global pandemic result from a new virus unrelated to influenza? Will there even be another pandemic? There are many strong feelings and opinions with respect to these questions, but the reality is, of course, that none of us can predict the future. This should not, however, prevent us from preparing for a global infectious disease outbreak, and Greger’s book certainly provides a comprehensive summary of the many factors that are critical for tackling this kind of a global public health problem and presents them in a way that will be interesting and informative to both scientists and lay persons, irrespective of their familiarity with H5N1.
- Published
- 2007
285. Influenza A virus infection of primary differentiated airway epithelial cell cultures derived from Syrian golden hamsters
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Andrew Pekosz, Regina K. Rowe, and Celeste M. Newby
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Airway epithelium ,Reassortment ,Respiratory Mucosa ,Viral Plaque Assay ,medicine.disease_cause ,Virus Replication ,Virus ,Article ,Microbiology ,Virus tropism ,chemistry.chemical_compound ,Virology ,Cricetinae ,Clara cells ,medicine ,Influenza A virus ,Animals ,Cilia ,Tropism ,Cells, Cultured ,Goblet cell ,biology ,Mesocricetus ,Virus receptor ,Epithelial Cells ,biology.organism_classification ,N-Acetylneuraminic Acid ,Influenza ,Sialic acid ,Virus receptors ,medicine.anatomical_structure ,chemistry ,Receptors, Virus ,Ciliated cells ,Respiratory epithelial cells - Abstract
The ability of several different influenza A virus strains to infect and replicate in primary, differentiated airway epithelial cell cultures from Syrian golden hamsters was investigated. All virus strains tested replicated equivalently in the cultures and displayed a preference for infecting nonciliated cells. This tropism correlated with the expression of both alpha2,3- and alpha2,6-linked sialic acid on the nonciliated cells. In contrast, the ciliated cells did not have detectable alpha2,6-linked sialic acid and expressed only low amounts of alpha2,3-linked sialic acid. In contrast to clinical isolates, laboratory strains of influenza A virus infected a limited number of ciliated cells at late times post-infection. The presence of alpha2,3- and alpha2,6-linked sialic acid residues on the same cell type suggests that Syrian golden hamsters and differentiated airway epithelial cell cultures derived from hamsters may provide a system for studying the reassortment of influenza A virus strains which utilize different forms of sialic acid as a primary virus receptor.
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286. Bioluminescence imaging of vaccinia virus: Effects of interferon on viral replication and spread
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Kathryn E. Luker, Tracey L. Schultz, Andrew Pekosz, Martha Hutchens, and Gary D. Luker
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Diagnostic Imaging ,Male ,Viral Plaque Assay ,viruses ,Vaccinia virus ,Biology ,Virus Replication ,Virus ,Mice ,chemistry.chemical_compound ,Genes, Reporter ,Luciferases, Firefly ,Interferon ,In vivo ,Virology ,Chlorocebus aethiops ,medicine ,Vaccinia ,Animals ,Bioluminescence imaging ,Luciferase ,Lung ,Vero Cells ,Receptors, Interferon ,Mice, Knockout ,Staining and Labeling ,Brain ,Disease Models, Animal ,Liver ,Viral replication ,chemistry ,Luminescent Measurements ,Female ,Interferons ,Spleen ,medicine.drug - Abstract
Whole animal imaging allows viral replication and localization to be monitored in intact animals, which provides significant advantages for determining viral and host factors that determine pathogenesis. To investigate effects of interferons on spatial and temporal progression of vaccinia infection, we generated recombinant viruses that express firefly luciferase or a monomeric orange fluorescent protein. These viruses allow vaccinia infection to be monitored with bioluminescence or fluorescence imaging, respectively. The recombinant viruses were not attenuated in vitro or in vivo relative to a control WR virus. In cell culture, reporters could be detected readily by 4 h post-infection, showing that these viruses can be used as early markers of infection. The magnitude of firefly luciferase activity measured with bioluminescence imaging in vitro and in vivo correlated directly with increasing titers of vaccinia virus, validating imaging data as a marker of viral infection. Replication of vaccinia was significantly greater in mice lacking receptors for type I interferons (IFN I R−/−) compared with wild-type mice, although both genotypes of mice developed focal infections in lungs and brain after intranasal inoculation. IFN I R−/− mice had greater dissemination of virus to liver and spleen than wild-type animals even when mortality occurred at the same time point after infection. Protective effects of type I interferons were mediated primarily through parenchymal cells rather than hematopoietic cells as analyzed by bone marrow transplant experiments. Collectively, our data define a new function for type I interferon signaling in systemic dissemination of vaccinia and validate these reporter viruses for studies of pathogenesis.
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287. Roles for the recycling endosome, Rab8, and Rab11 in hantavirus release from epithelial cells
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Regina K. Rowe, Andrew Pekosz, and Jason W. Suszko
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Orthohantavirus ,Endosome ,viruses ,Andes virus ,Biological Transport, Active ,Endosomes ,Biology ,Article ,symbols.namesake ,Intracellular organelle ,Virology ,Chlorocebus aethiops ,Golgi ,Animals ,Secretion ,RNA, Small Interfering ,Microscopy, Immunoelectron ,Nucleocapsid ,Vero Cells ,Hantavirus ,DNA Primers ,Inclusion Bodies ,Microscopy, Confocal ,Base Sequence ,Virus Assembly ,Intracellular Membranes ,Golgi apparatus ,Nucleocapsid Proteins ,Cell biology ,Cell Compartmentation ,rab GTP-Binding Proteins ,symbols ,RNA Interference ,Rab ,Recycling endosome ,Hantavirus Infection - Abstract
Hantavirus structural proteins are believed to localize to intracellular membranes often identified as Golgi membranes, in virus-infected cells. After virus budding into the Golgi luminal space, virus-containing vesicles are transported to the plasma membrane via trafficking pathways that are not well defined. Using the New World hantavirus, Andes virus, we have investigated the role of various Rab proteins in the release of hantavirus particles from infected cells. Rabs 8 and 11 were found to colocalize with Andes virus proteins in virus infected cells and when expressed from cDNA, implicating the recycling endosome as an organelle important for hantavirus infection. Small interfering RNA-mediated downregulation of Rab11a alone or Rab11a and Rab11b together resulted in a decrease in infectious virus particle secretion from infected cells. Downregulation of Rab8a did not alter infectious virus release but reduction of both isoforms did. These data implicate the recycling endosome and the Rab proteins associated with vesicular transport to or from this intracellular organelle as an important pathway for hantavirus trafficking to the plasma membrane.
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288. Polygenic control of neuroinvasiveness in California serogroup bunyaviruses
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Steven S. Scherer, Kara Stillmock, Michael J. Endres, Neal Nathanson, David M. Lukac, Andrew Pekosz, Christian Griot, Francisco Gonzalez-Scarano, and Dan Schmeidler
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Genes, Viral ,Immunology ,Clone (cell biology) ,Viremia ,Encephalitis Virus, California ,Biology ,Vaccines, Attenuated ,Virus Replication ,Nervous System ,Microbiology ,Virus ,Mice ,Virology ,Reassortant Viruses ,medicine ,Animals ,Recombination, Genetic ,Viral Structural Proteins ,Infectivity ,RNA ,Gene rearrangement ,medicine.disease ,Animals, Suckling ,Viral replication ,Insect Science ,RNA, Viral ,Research Article - Abstract
The pathogenesis of the California serogroup bunyaviruses includes both extraneural and intraneural replicative phases that can be separated experimentally. The present study dissects the viral genetic determinants of extraneural replication. We have previously described two attenuated reassortant clones of California serogroup bunyaviruses which exhibit reduced neuroinvasiveness after subcutaneous inoculation into suckling mice. Clone B1-1a bears an attenuated middle RNA segment (neuroinvasiveness phenotype v alpha v), and clone B.5 bears an attenuated large RNA segment (neuroinvasiveness phenotype alpha vv). We prepared reassortant viruses between these two strains and found that the two attenuated gene segments acted independently and additively, since reassortants bearing two attenuated RNA segments were more attenuated than the parental clones. Reassortants bearing no attenuated RNA segments were much more neuroinvasive than either parental clone, indicating that a neuroinvasive strain can be derived from two attenuated clones. Pathogenesis studies demonstrated that after injection of 10(3) PFU, the attenuated reassortant clones did not replicate in peripheral tissue, failed to reach the brain, and did not cause disease. At a dose of 10(6) PFU, attenuated clones failed to replicate to a significant level in peripheral tissue and produced only a minimal passive plasma viremia during the first 24 h but nevertheless reached high titers in the brain and killed mice. Because of this result, we investigated the possibility that neuroinvasion occurs via retrograde axonal transport, by determining whether sciatic nerve sectioning could protect against virus infection after hind leg footpad inoculation. We found that nerve sectioning had no effect on lethality, ruling out this mode of entry and suggesting that passive viremia is likely to be sufficient for invasion of the central nervous system.
289. Transcriptional profiling of the spleen in progressive visceral leishmaniasis reveals mixed expression of type 1 and type 2 cytokine-responsive genes
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Alvaro G. Hernandez, Peter C. Melby, A. Medina, Mandakini Patel, Omar A. Saldarriaga, Bruno L. Travi, Michael Cappello, Claudia M. Espitia, E. Yaneth Osorio, Mark Band, and Andrew Pekosz
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Transcriptional profiling ,endocrine system ,DNA, Complementary ,animal diseases ,Immunology ,Leishmania donovani ,Hamster ,Microarray ,Interferon-gamma ,Cricetinae ,parasitic diseases ,medicine ,Animals ,Cluster Analysis ,Humans ,RNA, Messenger ,Oligonucleotide Array Sequence Analysis ,Expressed Sequence Tags ,Visceral leishmaniasis ,Regulation of gene expression ,Principal Component Analysis ,Mesocricetus ,biology ,Gene Expression Profiling ,Molecular Sequence Annotation ,Leishmaniasis ,medicine.disease ,biology.organism_classification ,Up-Regulation ,Gene expression profiling ,Gene Ontology ,Gene Expression Regulation ,Disease Progression ,Cytokines ,Leishmaniasis, Visceral ,Interleukin-4 ,Spleen ,Signal Transduction ,Research Article ,Golden hamster - Abstract
Background The Syrian golden hamster (Mesocricetus aureus) has been used as a model to study infections caused by a number of human pathogens. Studies of immunopathogenesis in hamster infection models are challenging because of the limited availability of reagents needed to define cellular and molecular determinants. Results We sequenced a hamster cDNA library and developed a first-generation custom cDNA microarray that included 5131 unique cDNAs enriched for immune response genes. We used this microarray to interrogate the hamster spleen response to Leishmania donovani, an intracellular protozoan that causes visceral leishmaniasis. The hamster model of visceral leishmaniasis is of particular interest because it recapitulates clinical and immunopathological features of human disease, including cachexia, massive splenomegaly, pancytopenia, immunosuppression, and ultimately death. In the microarray a differentially expressed transcript was identified as having at least a 2-fold change in expression between uninfected and infected groups and a False Discovery Rate of
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290. The Influenza C Virus CM2 Protein Can Alter Intracellular pH, and Its Transmembrane Domain Can Substitute for That of the Influenza A Virus M2 Protein and Support Infectious Virus Production.
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Stewart, Shaun M. and Andrew Pekosz
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INFLUENZAVIRUS C - Abstract
An abstract of the article "The Influenza C Virus CM2 Protein Can Alter Intracellular pH, and Its Transmembrane Domain Can Substitute for That of the Influenza A Virus M2 Protein and Support Infectious Virus Production," by Shaunn M. Stewart and Andrew Pekosz is presented.
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- 2012
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291. G-CSF induction early in uropathogenicEscherichia coliinfection of the urinary tract modulates host immunity
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Scott J. Hultgren, Molly A. Ingersoll, Kimberly A. Kline, Hailyn V. Nielsen, Icahn School of Medicine at Mount Sinai [New York] (MSSM), Washington University School of Medicine in St. Louis, Washington University in Saint Louis (WUSTL), This work was supported by NIH Grants R01DK51406 and ORWH SCOR P50DK64540 (with the FDA) (to S.J.H.) and K.A.K. received support from an American Heart Association Postdoctoral Fellowship Award No. 0625736Z., and The authors thank Mark Miller and Andrew Pekosz for many helpful discussions and Swaine Chen, Kelly Wright and Karen Dodson for critical manuscript review and discussion. The authors also acknowledge the Alvin J. Siteman Cancer Center at Washington University School of Medicine and Barnes‐Jewish Hospital in St Louis, MO, for the use of the High Speed Cell Sorter Core, which provided support for the Bio‐Rad Bio‐Plex System and the flow analysis, as well as William Eades, who provided expert advice and help with flow analysis. The Siteman Cancer Center is supported in part by an NCI Cancer Center Support Grant #P30 CA91842.
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Neutrophils ,medicine.medical_treatment ,Interleukin-1beta ,MESH: Urinary Tract Infections ,MESH: Neutrophils ,medicine.disease_cause ,urologic and male genital diseases ,Mice ,MESH: Interleukin-1beta ,Granulocyte Colony-Stimulating Factor ,MESH: Animals ,Chemokine CCL2 ,Escherichia coli Infections ,0303 health sciences ,MESH: Escherichia coli ,female genital diseases and pregnancy complications ,3. Good health ,Granulocyte colony-stimulating factor ,Cytokine ,medicine.anatomical_structure ,Urinary Tract Infections ,[SDV.IMM]Life Sciences [q-bio]/Immunology ,Female ,Urinary system ,Immunology ,Biology ,Granulocyte ,Microbiology ,Article ,03 medical and health sciences ,MESH: Mice, Inbred C57BL ,Virology ,Escherichia coli ,medicine ,Animals ,MESH: Chemokine CCL2 ,MESH: Mice ,Escherichia coli infection ,030304 developmental biology ,MESH: Escherichia coli Infections ,030306 microbiology ,Monocyte ,Chemotaxis ,bacterial infections and mycoses ,Mice, Inbred C57BL ,MESH: Granulocyte Colony-Stimulating Factor ,MESH: Female - Abstract
Correction in:G‐CSF induction early in uropathogenic Escherichia coli infection of the urinary tract modulates host immunity Molly A. Ingersoll Kimberly A. Kline Hailyn V. Nielsen Scott J. Hultgren Volume 12 Issue 3 Cellular Microbiology pages: 411-411; International audience; Uropathogenic Escherichia coli (UPEC), the causative agent of approximately 85% of urinary tract infections (UTI), is a major health concern primarily affecting women. During infection, neutrophils infiltrate the bladder, but the mechanism of recruitment is not well understood. Here, we investigated the role of UPEC-induced cytokine production in neutrophil recruitment and UTI progression. We first examined the kinetics of cytokine expression during UPEC infection of the bladder, and their contribution to neutrophil recruitment. We found that UPEC infection induces expression of several pro-inflammatory cytokines including granulocyte colony-stimulating factor (G-CSF, CSF-3), not previously known to be involved in the host response to UTI. G-CSF induces neutrophil emigration from the bone marrow; these cells are thought to be critical for bacterial clearance during infection. Upon neutralization of G-CSF during UPEC infection, we found fewer circulating neutrophils, decreased neutrophil infiltration into the bladder and, paradoxically, a decreased bacterial burden in the bladder. However, depletion of G-CSF resulted in a corresponding increase in macrophage-activating cytokines, such as monocyte chemotactic protein-1 (MCP-1, CCL-2) and Il-1beta, which may be key in host response to UPEC infection, potentially resolving the paradoxical decreased bacterial burden. Thus, G-CSF acts in a previously unrecognized role to modulate the host inflammatory response during UPEC infection.
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- 2010
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