Research on Optimization of Formula SAE Truss-Frame

Directory of Open Access Journals (Sweden)

Wang Yu

2017-01-01

Full Text Available The frame, as one of the basic components of a car, plays an important role of connecting components and withstanding various loads. The truss-frame is the commonest form of Formula SAE car in that it has many advantages, such as low cost, simple manufacture, high rigidity and high strength. The development of frame is reviewed. First, the key performance indicators and the mechanical state of the frame under different conditions are discussed. Second, a variety of optimization methods used in the design of frame are described. Finally, the test method of frame are introduced, with the experimental data and the finite element analysis of results being compared. Through analyzing and summarizing the development of the design, optimization and testing technology of the Formula SAE truss-frame, forecasting the future trends, the great theoretical reference is provided for the design and research of the follow-up frame.

An essential role for trimethylguanosine RNA caps in Saccharomyces cerevisiae meiosis and their requirement for splicing of SAE3 and PCH2 meiotic pre-mRNAs.

Science.gov (United States)

Qiu, Zhicheng R; Shuman, Stewart; Schwer, Beate

2011-07-01

Tgs1 is the enzyme that converts m(7)G RNA caps to the 2,2,7-trimethylguanosine (TMG) caps characteristic of spliceosomal snRNAs. Fungi grow vegetatively without TMG caps, thereby raising the question of what cellular transactions, if any, are TMG cap-dependent. Here, we report that Saccharomyces cerevisiae Tgs1 methyltransferase activity is essential for meiosis. tgs1Δ cells are specifically defective in splicing PCH2 and SAE3 meiotic pre-mRNAs. The TMG requirement for SAE3 splicing is alleviated by two intron mutations: a UAUUAAC to UACUAAC change that restores a consensus branchpoint and disruption of a stem-loop encompassing the branchpoint. The TMG requirement for PCH2 splicing is alleviated by a CACUAAC to UACUAAC change restoring a consensus branchpoint and by shortening the PCH2 5' exon. Placing the SAE3 and PCH2 introns within a HIS3 reporter confers Tgs1-dependent histidine prototrophy, signifying that the respective introns are portable determinants of TMG-dependent gene expression. Analysis of in vitro splicing in extracts of TGS1 versus tgs1Δ cells showed that SAE3 intron removal was enfeebled without TMG caps, whereas splicing of ACT1 was unaffected. Our findings illuminate a new mode of tunable splicing, a reliance on TMG caps for an essential developmental RNA transaction, and three genetically distinct meiotic splicing regulons in budding yeast.

USO DA SISTEMATIZAÇÃO DA ASSISTÊNCIA DE ENFERMAGEM (SAE: UMA FERRAMENTA PARA REALIZAÇÃO DA AUDITORIA DE QUALIDADE

Directory of Open Access Journals (Sweden)

Maria Izelta da Silva Santos

2013-04-01

Full Text Available Realizar uma revisao bibliografica acerca da Sistematização da Assistencia de Enfermagem (SAE como instrumento para auditoria de qualidade. Metodos: Foram revisados 12 artigos cientificos sobre auditoria e 10 artigos sobre SAE, bem como dois livros sobre os assuntos abordados. Esta revisao e de carater descritivo. Resultados: Por meio da pesquisa, tornou-se evidente a correlação da SAE com a auditoria. A SAE bem implementada pode contribuir para uma auditoria de qualidade.

Electric Vehicle Communications Standards Testing and Validation - Phase II: SAE J2931/1

Energy Technology Data Exchange (ETDEWEB)

Pratt, Richard M. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Gowri, Krishnan [Pacific Northwest National Lab. (PNNL), Richland, WA (United States)

2013-01-15

Vehicle to grid communication standards enable interoperability among vehicles, charging stations and utility providers and provide the capability to implement charge management. Several standards initiatives by the Society of Automobile Engineers (SAE), International Standards Organization and International Electrotechnical Commission (ISO/IEC), and ZigBee/HomePlug Alliance are developing requirements for communication messages and protocols. Recent work by the Electric Power Research Institute (EPRI) in collaboration with SAE and automobile manufacturers has identified vehicle to grid communication performance requirements and developed a test plan as part of SAE J2931/1 committee work. This laboratory test plan was approved by the SAE J2931/1 committee and included test configurations, test methods, and performance requirements to verify reliability, robustness, repeatability, maximum communication distance, and authentication features of power line carrier (PLC) communication modules at the internet protocol layer level. The goal of the testing effort was to select a communication technology that would enable automobile manufacturers to begin the development and implementation process. The EPRI/Argonne National Laboratory (ANL)/Pacific Northwest National Laboratory (PNNL) testing teams divided the testing so that results for each test could be presented by two teams, performing the tests independently. The PNNL team performed narrowband PLC testing including the Texas Instruments (TI) Concerto, Ariane Controls AC-CPM1, and the MAXIM Tahoe 2 evaluation boards. The scope of testing was limited to measuring the vendor systems communication performance between Electric Vehicle Support Equipment (EVSE) and plug-in electric vehicles (PEV). The testing scope did not address PEV’s CAN bus to PLC or PLC to EVSE (Wi-Fi, cellular, PLC Mains, etc.) communication integration. In particular, no evaluation was performed to delineate the effort needed to translate the IPv6

Kogeja vormistamine eesti keeles : nihkeid SAE perifeerias / Mati Erelt, Helle Metslang

Index Scriptorium Estoniae

Erelt, Mati, 1941-

2008-01-01

Vaadeldakse kogeja vormistamist ning alternatiivsete mallide kasutust ajakirjanduskeeles ajavahemikul 1995-2005. Püütakse välja selgitada toimunud muutused ja nende põhjused. SAE - Euroopa keskmine standard

Kogeja vormistamine eesti keeles : nihkeid SAE perifeerias / Mati Erelt, Helle Metslang

Index Scriptorium Estoniae

Erelt, Mati, 1941-

2008-01-01

Artiklis vaadeldakse kogeja vormistamist ning alternatiivsete mallide kasutust ajakirjanduskeeles ajavahemikul 1995-2005 eesmärgiga välja selgitada toimunud muutused ning nende põhjused. SAE - Euroopa keskmine standard

A model for fracture toughness evaluation of the carburized layer for SAE 5115 steel

OpenAIRE

Sandor, Leonardo Taborda; Ferreira, Itamar

2006-01-01

The purpose of this work is to propose a model for evaluating the fracture toughness along the SAE 5115 steel carburized layer. Due to the small thickness of those layers, it is impossible to machine specimens from those layer in accordance with standards. For simulating the microstructures of the carburized layer in order to get samples for tensile and the fracture toughness testing, specimens of SAE 5115, 5140, 5160, and 52100 steels have been machined, assuming the local influence just the...

Caffeine impairs resection during DNA break repair by reducing the levels of nucleases Sae2 and Dna2

Science.gov (United States)

Tsabar, Michael; Eapen, Vinay V.; Mason, Jennifer M.; Memisoglu, Gonen; Waterman, David P.; Long, Marcus J.; Bishop, Douglas K.; Haber, James E.

2015-01-01

In response to chromosomal double-strand breaks (DSBs), eukaryotic cells activate the DNA damage checkpoint, which is orchestrated by the PI3 kinase-like protein kinases ATR and ATM (Mec1 and Tel1 in budding yeast). Following DSB formation, Mec1 and Tel1 phosphorylate histone H2A on serine 129 (known as γ-H2AX). We used caffeine to inhibit the checkpoint kinases after DSB induction. We show that prolonged phosphorylation of H2A-S129 does not require continuous Mec1 and Tel1 activity. Unexpectedly, caffeine treatment impaired homologous recombination by inhibiting 5′ to 3′ end resection, independent of Mec1 and Tel1 inhibition. Caffeine treatment led to the rapid loss, by proteasomal degradation, of both Sae2, a nuclease that plays a role in early steps of resection, and Dna2, a nuclease that facilitates one of two extensive resection pathways. Sae2's instability is evident in the absence of DNA damage. A similar loss is seen when protein synthesis is inhibited by cycloheximide. Caffeine treatment had similar effects on irradiated HeLa cells, blocking the formation of RPA and Rad51 foci that depend on 5′ to 3′ resection of broken chromosome ends. Our findings provide insight toward the use of caffeine as a DNA damage-sensitizing agent in cancer cells. PMID:26019182

Smoking-Attributable Mortality, Morbidity, and Economic Costs (SAMMEC) - Smoking-Attributable Expenditures (SAE)

Data.gov (United States)

U.S. Department of Health & Human Services — 2005-2009. SAMMEC - Smoking-Attributable Mortality, Morbidity, and Economic Costs. Smoking-attributable expenditures (SAEs) are excess health care expenditures...

Tritium stripping in a nitrogen glovebox using SAES St 198

International Nuclear Information System (INIS)

Klein, J.E.; Wermer, J.R.

1994-01-01

SAES metal getter material St 198 was chosen for glovebox stripper tests to evaluate its effectiveness of removing tritium from a nitrogen atmosphere. The St 198 material is unique from a number of other metal hydride-based getter materials in that it is relatively inert to nitrogen and can thus be used in nitrogen glovebox atmospheres. Six tritium stripper experiments which mock-up the use of a SAES St 198 stripper bed for a full-scale (10,500 liter) nitrogen glovebox have been completed. Experiments consisted of a release of small quantity of protium/deuterium spiked with tritium which were scaled to simulate tritium releases of 0.1 g., 1.0 g., and 10 g. into the glovebox. The tritium spike allows detection using tritium ion chambers. The St 198 stripper system produced a reduction in tritium activity of approximately two orders of magnitude in 24 hours (6--8 atmosphere turn-overs) of stripper operation

Small area estimation (SAE) model: Case study of poverty in West Java Province

Science.gov (United States)

Suhartini, Titin; Sadik, Kusman; Indahwati

2016-02-01

This paper showed the comparative of direct estimation and indirect/Small Area Estimation (SAE) model. Model selection included resolve multicollinearity problem in auxiliary variable, such as choosing only variable non-multicollinearity and implemented principal component (PC). Concern parameters in this paper were the proportion of agricultural venture poor households and agricultural poor households area level in West Java Province. The approach for estimating these parameters could be performed based on direct estimation and SAE. The problem of direct estimation, three area even zero and could not be conducted by directly estimation, because small sample size. The proportion of agricultural venture poor households showed 19.22% and agricultural poor households showed 46.79%. The best model from agricultural venture poor households by choosing only variable non-multicollinearity and the best model from agricultural poor households by implemented PC. The best estimator showed SAE better then direct estimation both of the proportion of agricultural venture poor households and agricultural poor households area level in West Java Province. The solution overcame small sample size and obtained estimation for small area was implemented small area estimation method for evidence higher accuracy and better precision improved direct estimator.

Microstructural characterization of an AISI-SAE 4140 steel without nitridation and nitrided; Caracterizacion microestructural de un acero AISI-SAE 4140 sin nitrurar y nitrurado

Energy Technology Data Exchange (ETDEWEB)

Medina F, A.; Naquid G, C. [Gerencia de Ciencia de Materiales, Depto. de Sintesis y Caracterizacion de Materiales, A.P. 18-1027, 11801 Mexico D.F. (Mexico)

2000-07-01

It was micro structurally characterized an AISI-SAE 4140 steel before and after of nitridation through the nitridation process by plasma post-unloading microwaves through Optical microscopy (OM), Scanning electron microscopy (SEM) by means of secondary electrons and retrodispersed, X-ray diffraction (XRD), Energy dispersion spectra (EDS) and mapping of elements. (Author)

Number of infection events per cell during HIV-1 cell-free infection.

Science.gov (United States)

Ito, Yusuke; Remion, Azaria; Tauzin, Alexandra; Ejima, Keisuke; Nakaoka, Shinji; Iwasa, Yoh; Iwami, Shingo; Mammano, Fabrizio

2017-07-26

HIV-1 accumulates changes in its genome through both recombination and mutation during the course of infection. For recombination to occur, a single cell must be infected by two HIV strains. These coinfection events were experimentally demonstrated to occur more frequently than would be expected for independent infection events and do not follow a random distribution. Previous mathematical modeling approaches demonstrated that differences in target cell susceptibility can explain the non-randomness, both in the context of direct cell-to-cell transmission, and in the context of free virus transmission (Q. Dang et al., Proc. Natl. Acad. Sci. USA 101:632-7, 2004: K. M. Law et al., Cell reports 15:2711-83, 2016). Here, we build on these notions and provide a more detailed and extensive quantitative framework. We developed a novel mathematical model explicitly considering the heterogeneity of target cells and analysed datasets of cell-free HIV-1 single and double infection experiments in cell culture. Particularly, in contrast to the previous studies, we took into account the different susceptibility of the target cells as a continuous distribution. Interestingly, we showed that the number of infection events per cell during cell-free HIV-1 infection follows a negative-binomial distribution, and our model reproduces these datasets.

CARACTHERIZATION OF MECHANICAL PROPERTIES ON SAE 4140 STEEL AFTER DIFFERENT TREATMENTS OF QUENCHING AND TEMPERING

OpenAIRE

LUIS PAULO PIRES GONCALVES RIBEIRO

2006-01-01

Os aços de alta resistência e baixa liga (ARBL) pertencem a uma classe de aços que desperta o interesse de pesquisadores da área metal-mecânica. A pesquisa teve como objetivos estudar a influência dos tratamentos térmicos de têmpera e revenido sobre as propriedades do aço SAE 4140 e avaliar o comportamento do aço SAE 4140 quando submetido ao revenido na faixa de temperatura característica da fragilização. Inicialmente, o material foi tratado termicamente sob di...

SAE for the prediction of road traffic status from taxicab operating data and bus smart card data

Science.gov (United States)

Zhengfeng, Huang; Pengjun, Zheng; Wenjun, Xu; Gang, Ren

Road traffic status is significant for trip decision and traffic management, and thus should be predicted accurately. A contribution is that we consider multi-modal data for traffic status prediction than only using single source data. With the substantial data from Ningbo Passenger Transport Management Sector (NPTMS), we wished to determine whether it was possible to develop Stacked Autoencoders (SAEs) for accurately predicting road traffic status from taxicab operating data and bus smart card data. We show that SAE performed better than linear regression model and Back Propagation (BP) neural network for determining the relationship between road traffic status and those factors. In a 26-month data experiment using SAE, we show that it is possible to develop highly accurate predictions (91% test accuracy) of road traffic status from daily taxicab operating data and bus smart card data.

Retained Austenite in SAE 52100 Steel Post Magnetic Processing and Heat Treatment

Energy Technology Data Exchange (ETDEWEB)

Pappas, Nathaniel R [ORNL; Watkins, Thomas R [ORNL; Cavin, Odis Burl [ORNL; Jaramillo, Roger A [ORNL; Ludtka, Gerard Michael [ORNL

2007-01-01

Steel is an iron-carbon alloy that contains up to 2% carbon by weight. Understanding which phases of iron and carbon form as a function of temperature and percent carbon is important in order to process/manufacture steel with desired properties. Austenite is the face center cubic (fcc) phase of iron that exists between 912 and 1394 C. When hot steel is rapidly quenched in a medium (typically oil or water), austenite transforms into martensite. The goal of the study is to determine the effect of applying a magnetic field on the amount of retained austenite present at room temperature after quenching. Samples of SAE 52100 steel were heat treated then subjected to a magnetic field of varying strength and time, while samples of SAE 1045 steel were heat treated then subjected to a magnetic field of varying strength for a fixed time while being tempered. X-ray diffraction was used to collect quantitative data corresponding to the amount of each phase present post processing. The percentage of retained austenite was then calculated using the American Society of Testing and Materials standard for determining the amount of retained austenite for randomly oriented samples and was plotted as a function of magnetic field intensity, magnetic field apply time, and magnetic field wait time after quenching to determine what relationships exist with the amount of retained austenite present. In the SAE 52100 steel samples, stronger field strengths resulted in lower percentages of retained austenite for fixed apply times. The results were inconclusive when applying a fixed magnetic field strength for varying amounts of time. When applying a magnetic field after waiting a specific amount of time after quenching, the analyses indicate that shorter wait times result in less retained austenite. The SAE 1045 results were inconclusive. The samples showed no retained austenite regardless of magnetic field strength, indicating that tempering removed the retained austenite. It is apparent

Efferocytosis of Pathogen-Infected Cells

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Niloofar Karaji

2017-12-01

Full Text Available The prompt and efficient clearance of unwanted and abnormal cells by phagocytes is termed efferocytosis and is crucial for organism development, maintenance of tissue homeostasis, and regulation of the immune system. Dying cells are recognized by phagocytes through pathways initiated via “find me” signals, recognition via “eat me” signals and down-modulation of regulatory “don’t eat me” signals. Pathogen infection may trigger cell death that drives phagocytic clearance in an immunologically silent, or pro-inflammatory manner, depending on the mode of cell death. In many cases, efferocytosis is a mechanism for eliminating pathogens and pathogen-infected cells; however, some pathogens have subverted this process and use efferocytic mechanisms to avoid innate immune detection and assist phagocyte infection. In parallel, phagocytes can integrate signals received from infected dying cells to elicit the most appropriate effector response against the infecting pathogen. This review focuses on pathogen-induced cell death signals that drive infected cell recognition and uptake by phagocytes, and the outcomes for the infected target cell, the phagocyte, the pathogen and the host.

Tritium stripping in a nitrogen glove box using palladium/zeolite and SAES St 198 trademark

International Nuclear Information System (INIS)

Klien, J.E.; Wermer, J.R.

1995-01-01

Glove box clean-up experiments were conducted in a nitrogen glove box using palladium deposited on zeolite (Pd/z) and a SAES St 198 trademark getter as tritium stripping materials. Protium/deuterium samples spiked with tritium were released into a 620 liter glove box to simulate tritium releases in a 10,500 liter glove box. The Pd/z and the SAES St 198 trademark stripper beds produced a reduction in tritium activity of approximately two to three orders of magnitude and glove box clean-up was limited by a persistent background tritium activity level. Attempts to significantly reduce the glove box activity to lower levels without purging were unsuccessful

40 CFR Appendix D to Subpart B of... - SAE J2810 Standard for Recovery Only Equipment for HFC-134a Refrigerant

Science.gov (United States)

2010-07-01

... removed from, motor vehicle air-conditioning systems. 1. Scope The purpose of this SAE Standard is to... 700 Standard or allow for recycling of the recovered refrigerant to SAE J2788 specifications by using... (R-134a) Recovery/Recycling Equipment and Recovery/Recycling/Recharging for Mobile Air-Conditioning...

La articulación de los SAE y la generación de valor público

OpenAIRE

Suárez Ognio, María del Carmen Leonor; Chumacero Guzmán, Enrique Eloy

2017-01-01

Este trabajo busca identificar el problema que impide la consecución de logros en términos de valor público y trabajar en una de sus causas: la desarticulación de los principales Sistemas Administrativos del Estado (SAE). Este estudio sirve para explicar cómo la teoría de la agencia ha sido usada, consciente o inconscientemente, en el diseño de los SAE, haciendo que actúen como sistemas de control en lugar de ser sistemas para la gestión orientados a generar valor público. Con esto se ha afec...

Sae regulator factor impairs the response to photodynamic inactivation mediated by Toluidine blue in Staphylococcus aureus.

Science.gov (United States)

Gándara, Lautaro; Mamone, Leandro; Dotto, Cristian; Buzzola, Fernanda; Casas, Adriana

2016-12-01

Photodynamic inactivation (PDI) involves the combined use of light and a photosensitizer, which, in the presence of oxygen, originates cytotoxic species capable of inactivating bacteria. Since the emergence of multi-resistant bacterial strains is becoming an increasing public health concern, PDI becomes an attractive choice. The aim of this work was to study the differential susceptibility to Toluidine blue (TB) mediated PDI (TB-PDI) of S. aureus mutants (RN6390 and Newman backgrounds) for different key regulators of virulence factors related to some extent to oxidative stress. Complete bacteria eradication of planktonic cultures of RN6390 S. aureus photosensitized with 13μM TB was obtained upon illumination with a low light dose of 4.2J/cm 2 from a non-coherent light source. Similarly, complete cell death was achieved applying 1.3μM TB and 19J/cm 2 light dose, showing that higher light doses can lead to equal cell death employing low photosensitizer concentrations. Interestingly, RN6390 in planktonic culture responded significantly better to TB-PDI than the Newman strain. We showed that deficiencies in rsbU, mgrA (transcription factors related to stress response) or agr (quorum sensing system involved in copper resistance to oxidative stress) did not modify the response of planktonic S. aureus to PDI. On the other hand, the two component system sae impaired the response to TB-PDI through a mechanism not related to the Eap adhesin. More severe conditions were needed to inactivate S. aureus biofilms (0.5mM TB, 157J/cm 2 laser light). In mutant sae biofilms, strain dependant differential susceptibilities are not noticed. Copyright © 2016 Elsevier B.V. All rights reserved.

Characterization of the loss of SUMO pathway function on cancer cells and tumor proliferation.

Directory of Open Access Journals (Sweden)

Xingyue He

Full Text Available SUMOylation is a post-translational ubiquitin-like protein modification pathway that regulates important cellular processes including chromosome structure, kinetochore function, chromosome segregation, nuclear and sub-nuclear organization, transcription and DNA damage repair. There is increasing evidence that the SUMO pathway is dysregulated in cancer, raising the possibility that modulation of this pathway may have therapeutic potential. To investigate the importance of the SUMO pathway in the context of cancer cell proliferation and tumor growth, we applied lentivirus-based short hairpin RNAs (shRNA to knockdown SUMO pathway genes in human cancer cells. shRNAs for SAE2 and UBC9 reduced SUMO conjugation activity and inhibited proliferation of human cancer cells. To expand upon these observations, we generated doxycycline inducible conditional shRNA cell lines for SAE2 to achieve acute and reversible SAE2 knockdown. Conditional SAE2 knockdown in U2OS and HCT116 cells slowed cell growth in vitro, and SAE2 knockdown induced multiple terminal outcomes including apoptosis, endoreduplication and senescence. Multinucleated cells became senescent and stained positive for the senescence marker, SA-β Gal, and displayed elevated levels of p53 and p21. In an attempt to explain these phenotypes, we confirmed that loss of SUMO pathway activity leads to a loss of SUMOylated Topoisomerase IIα and the appearance of chromatin bridges which can impair proper cytokinesis and lead to multinucleation. Furthermore, knockdown of SAE2 induces disruption of PML nuclear bodies which may further promote apoptosis or senescence. In an in vivo HCT116 xenograft tumor model, conditional SAE2 knockdown strongly impaired tumor growth. These data demonstrate that the SUMO pathway is required for cancer cell proliferation in vitro and tumor growth in vivo, implicating the SUMO pathway as a potential cancer therapeutic target.

Bilayers Polypyrrole Coatings for Corrosion Protection of SAE 4140 Steel

OpenAIRE

Lehr, I.L.; Saidman, S.B.

2014-01-01

In this study polypyrrole (PPy) bilayers films were electrodeposited onto SAE 4140 steel. The inner layer was electropolymerized in the presence of molibdate and nitrate and the outer layer in a solution containing sodium bis (2-ethylhexyl) sulfosuccinate (AOT). The electrosynthesis was done under potentiostatic conditions. The corrosion protection properties of the films were examined in sodium chloride solution by open circuit measurements, linear polarization and electrochemical impedance ...

Microstructural characterization of an AISI-SAE 4140 steel without nitridation and nitrided

International Nuclear Information System (INIS)

Medina F, A.; Naquid G, C.

2000-01-01

It was micro structurally characterized an AISI-SAE 4140 steel before and after of nitridation through the nitridation process by plasma post-unloading microwaves through Optical microscopy (OM), Scanning electron microscopy (SEM) by means of secondary electrons and retrodispersed, X-ray diffraction (XRD), Energy dispersion spectra (EDS) and mapping of elements. (Author)

SAE2.py: a python script to automate parameter studies using SCREAMER with application to magnetic switching on Z

International Nuclear Information System (INIS)

Orndorff-Plunkett, Franklin

2011-01-01

The SCREAMER simulation code is widely used at Sandia National Laboratories for designing and simulating pulsed power accelerator experiments on super power accelerators. A preliminary parameter study of Z with a magnetic switching retrofit illustrates the utility of the automating script for optimizing pulsed power designs. SCREAMER is a circuit based code commonly used in pulsed-power design and requires numerous iterations to find optimal configurations. System optimization using simulations like SCREAMER is by nature inefficient and incomplete when done manually. This is especially the case when the system has many interactive elements whose emergent effects may be unforeseeable and complicated. For increased completeness, efficiency and robustness, investigators should probe a suitably confined parameter space using deterministic, genetic, cultural, ant-colony algorithms or other computational intelligence methods. I have developed SAE2 - a user-friendly, deterministic script that automates the search for optima of pulsed-power designs with SCREAMER. This manual demonstrates how to make input decks for SAE2 and optimize any pulsed-power design that can be modeled using SCREAMER. Application of SAE2 to magnetic switching on model of a potential Z refurbishment illustrates the power of SAE2. With respect to the manual optimization, the automated optimization resulted in 5% greater peak current (10% greater energy) and a 25% increase in safety factor for the most highly stressed element.

Neuraminidase treatment of respiratory syncytial virus-infected cells or virions, but not target cells, enhances cell-cell fusion and infection

International Nuclear Information System (INIS)

Barretto, Naina; Hallak, Louay K.; Peeples, Mark E.

2003-01-01

Respiratory syncytial virus (RSV) infection of HeLa cells induces fusion, but transient expression of the three viral glycoproteins induces fusion poorly, if at all. We found that neuraminidase treatment of RSV-infected cells to remove sialic acid (SA) increases fusion dramatically and that the same treatment of transiently transfected cells expressing the three viral glycoproteins, or even cells expressing the fusion (F) protein alone, results in easily detectable fusion. Neuraminidase treatment of the effector cells, expressing the viral glycoproteins, enhanced fusion while treatment of the target cells did not. Likewise, infectivity was increased by treating virions with neuraminidase, but not by treating target cells. Reduction of charge repulsion by removal of the negatively charged SA is unlikely to explain this effect, since removal of negative charges from either membrane would reduce charge repulsion. Infection with neuraminidase-treated virus remained heparan-sulfate-dependent, indicating that a novel attachment mechanism is not revealed by SA removal. Interestingly, neuraminidase enhancement of RSV infectivity was less pronounced in a virus expressing both the G and the F glycoproteins, compared to virus expressing only the F glycoprotein, possibly suggesting that the G protein sterically hinders access of the neuraminidase to its fusion-enhancing target

Merkel cell polyomavirus infection and Merkel cell carcinoma.

Science.gov (United States)

Liu, Wei; MacDonald, Margo; You, Jianxin

2016-10-01

Merkel cell polyomavirus is the only polyomavirus discovered to date that is associated with a human cancer. MCPyV infection is highly prevalent in the general population. Nearly all healthy adults asymptomatically shed MCPyV from their skin. However, in elderly and immunosuppressed individuals, the infection can lead to a lethal form of skin cancer, Merkel cell carcinoma. In the last few years, new findings have established links between MCPyV infection, host immune response, and Merkel cell carcinoma development. This review discusses these recent discoveries on how MCPyV interacts with host cells to achieve persistent infection and, in the immunocompromised population, contributes to MCC development. Copyright © 2016 Elsevier B.V. All rights reserved.

HCV Infection and B-Cell Lymphomagenesis

Directory of Open Access Journals (Sweden)

Masahiko Ito

2011-01-01

Full Text Available Hepatitis C virus (HCV has been recognized as a major cause of chronic liver diseases worldwide. It has been suggested that HCV infects not only hepatocytes but also mononuclear lymphocytes including B cells that express the CD81 molecule, a putative HCV receptor. HCV infection of B cells is the likely cause of B-cell dysregulation disorders such as mixed cryoglobulinemia, rheumatoid factor production, and B-cell lymphoproliferative disorders that may evolve into non-Hodgkin's lymphoma (NHL. Epidemiological data indicate an association between HCV chronic infection and the occurrence of B-cell NHL, suggesting that chronic HCV infection is associated at least in part with B-cell lymphomagenesis. In this paper, we aim to provide an overview of recent literature, including our own, to elucidate a possible role of HCV chronic infection in B-cell lymphomagenesis.

Multidrug Resistance-Associated Protein 3 (Mrp3/Abcc3/Moat-D) Is Expressed in the SAE Squalus acanthias Shark Embryo–Derived Cell Line

Science.gov (United States)

Kobayashi, Hiroshi; Parton, Angela; Czechanski, Anne; Durkin, Christopher; Kong, Chi-Chon; Barnes, David

2008-01-01

The multidrug resistance-associated protein 3 (MRP3/Mrp3) is a member of the ATP-binding cassette (ABC) protein family of membrane transporters and related proteins that act on a variety of xenobiotic and anionic molecules to transfer these substrates in an ATP-dependent manner. In recent years, useful comparative information regarding evolutionarily conserved structure and transport functions of these proteins has accrued through the use of primitive marine animals such as cartilaginous fish. Until recently, one missing tool in comparative studies with cartilaginous fish was cell culture. We have derived from the embryo of Squalus acanthias, the spiny dogfish shark, the S. acanthias embryo (SAE) mesenchymal stem cell line. This is the first continuously proliferating cell line from a cartilaginous fish. We identified expression of Mrp3 in this cell line, cloned the molecule, and examined molecular and cellular physiological aspects of the protein. Shark Mrp3 is characterized by three membrane-spanning domains and two nucleotide-binding domains. Multiple alignments with other species showed that the shark Mrp3 amino acid sequence was well conserved. The shark sequence was overall 64% identical to human MRP3, 72% identical to chicken Mrp3, and 71% identical to frog and stickleback Mrp3. Highest identity between shark and human amino acid sequence (82%) was seen in the carboxyl-terminal nucleotide-binding domain of the proteins. Cell culture experiments showed that mRNA for the protein was induced as much as 25-fold by peptide growth factors, fetal bovine serum, and lipid nutritional components, with the largest effect mediated by a combination of lipids including unsaturated and saturated fatty acids, cholesterol, and vitamin E. PMID:18284333

Multidrug resistance-associated protein 3 (Mrp3/Abcc3/Moat-D) is expressed in the SAE Squalus acanthias shark embryo-derived cell line.

Science.gov (United States)

Kobayashi, Hiroshi; Parton, Angela; Czechanski, Anne; Durkin, Christopher; Kong, Chi-Chon; Barnes, David

2007-01-01

The multidrug resistance-associated protein 3 (MRP3/Mrp3) is a member of the ATP-binding cassette (ABC) protein family of membrane transporters and related proteins that act on a variety of xenobiotic and anionic molecules to transfer these substrates in an ATP-dependent manner. In recent years, useful comparative information regarding evolutionarily conserved structure and transport functions of these proteins has accrued through the use of primitive marine animals such as cartilaginous fish. Until recently, one missing tool in comparative studies with cartilaginous fish was cell culture. We have derived from the embryo of Squalus acanthias, the spiny dogfish shark, the S. acanthias embryo (SAE) mesenchymal stem cell line. This is the first continuously proliferating cell line from a cartilaginous fish. We identified expression of Mrp3 in this cell line, cloned the molecule, and examined molecular and cellular physiological aspects of the protein. Shark Mrp3 is characterized by three membrane-spanning domains and two nucleotide-binding domains. Multiple alignments with other species showed that the shark Mrp3 amino acid sequence was well conserved. The shark sequence was overall 64% identical to human MRP3, 72% identical to chicken Mrp3, and 71% identical to frog and stickleback Mrp3. Highest identity between shark and human amino acid sequence (82%) was seen in the carboxyl-terminal nucleotide-binding domain of the proteins. Cell culture experiments showed that mRNA for the protein was induced as much as 25-fold by peptide growth factors, fetal bovine serum, and lipid nutritional components, with the largest effect mediated by a combination of lipids including unsaturated and saturated fatty acids, cholesterol, and vitamin E.

A Novel Image Cryptosystem Based on S-AES and Chaotic Map

Directory of Open Access Journals (Sweden)

Bai Lan

2015-01-01

Full Text Available This paper proposes a novel scheme based on simplified advanced encryption standard (S-AES for image encryption. Modified Arnold Map applied as diffusion technique for an image, and the key and dynamic S-box of encryption is generated by PWLCM. The goal is to balance rapidity and security of encryption. Experimental implementation has been done. This light encryption scheme shows resistance against chosen-plaintext attack and is suitable for sensor networks and IoT.

The value of T1-weighted images in the differentiation between MS, white matter lesions, and subcortical arteriosclerotic encephalopathy (SAE)

Energy Technology Data Exchange (ETDEWEB)

Uhlenbrock, D.; Sehlen, S.

1989-07-01

The aim of the study was to define reliable criteria for the differentiation of MR imaging between patients with MS and with 'vascular' white matter lesions/SAE. We examined 35 patients with proven MS according to the Poser criteria and 35 patients with other white matter lesions and/or SAE. The result is that with MR a differentiation can be achieved provided that T1-weighted spin-echo sequences are included and the different pattern of distribution is considered. MS plaques are predominantly located in the subependymal region, vascular white matter lesions are mainly located in the water-shed of the superficial middle cerebral branches and the deep perforating long medullary vessels in the centrum semiovale. Infratentorial lesions are more often seen in MS. Confluence at the lateral ventricles is frequently accompanied by confluent abnormalities around the third ventricle, Sylvian aqueduct, and fourth ventricle, which is uncommon in SAE. In MS many lesions visible on T2-weighted images have a cellular or intracellular composition that renders them visible also on T1-weighted ones as regions with low signal intensity and more or less distinct boundary. 'Vascular' white matter lesions and SAE mainly represent demyelination and can therefore be seen on T2-weighted images, but corresponding low signal intensity lesions on T1-weighted images are uncommon. In some exceptions there are such lesions with low signal representing lacunar infarcts or widened Virchow-Robin-spaces. (orig.).

Oral epithelial cells are susceptible to cell-free and cell-associated HIV-1 infection in vitro

International Nuclear Information System (INIS)

Moore, Jennifer S.; Rahemtulla, Firoz; Kent, Leigh W.; Hall, Stacy D.; Ikizler, Mine R.; Wright, Peter F.; Nguyen, Huan H.; Jackson, Susan

2003-01-01

Epithelial cells lining the oral cavity are exposed to HIV-1 through breast-feeding and oral-genital contact. Genital secretions and breast milk of HIV-1-infected subjects contain both cell-free and cell-associated virus. To determine if oral epithelial cells can be infected with HIV-1 we exposed gingival keratinocytes and adenoid epithelial cells to cell-free virus and HIV-1-infected peripheral blood mononuclear cells and monocytes. Using primary isolates we determined that gingival keratinocytes are susceptible to HIV-1 infection via cell-free CD4-independent infection only. R5 but not X4 viral strains were capable of infecting the keratinocytes. Further, infected cells were able to release infectious virus. In addition, primary epithelial cells isolated from adenoids were also susceptible to infection; both cell-free and cell-associated virus infected these cells. These data have potential implications in the transmission of HIV-1 in the oral cavity

Murid herpesvirus-4 exploits dendritic cells to infect B cells.

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Miguel Gaspar

2011-11-01

Full Text Available Dendritic cells (DCs play a central role in initiating immune responses. Some persistent viruses infect DCs and can disrupt their functions in vitro. However, these viruses remain strongly immunogenic in vivo. Thus what role DC infection plays in the pathogenesis of persistent infections is unclear. Here we show that a persistent, B cell-tropic gamma-herpesvirus, Murid Herpesvirus-4 (MuHV-4, infects DCs early after host entry, before it establishes a substantial infection of B cells. DC-specific virus marking by cre-lox recombination revealed that a significant fraction of the virus latent in B cells had passed through a DC, and a virus attenuated for replication in DCs was impaired in B cell colonization. In vitro MuHV-4 dramatically altered the DC cytoskeleton, suggesting that it manipulates DC migration and shape in order to spread. MuHV-4 therefore uses DCs to colonize B cells.

Alteration of cell cycle progression by Sindbis virus infection

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Yi, Ruirong; Saito, Kengo [Department of Molecular Virology, Graduate School of Medicine, Chiba University Graduate School of Medicine, 1-8-1 Inohana, Chiba 260-8670 (Japan); Isegawa, Naohisa [Laboratory Animal Center, Graduate School of Medicine, Chiba University Graduate School of Medicine, 1-8-1 Inohana, Chiba 260-8670 (Japan); Shirasawa, Hiroshi, E-mail: sirasawa@faculty.chiba-u.jp [Department of Molecular Virology, Graduate School of Medicine, Chiba University Graduate School of Medicine, 1-8-1 Inohana, Chiba 260-8670 (Japan)

2015-07-10

We examined the impact of Sindbis virus (SINV) infection on cell cycle progression in a cancer cell line, HeLa, and a non-cancerous cell line, Vero. Cell cycle analyses showed that SINV infection is able to alter the cell cycle progression in both HeLa and Vero cells, but differently, especially during the early stage of infection. SINV infection affected the expression of several cell cycle regulators (CDK4, CDK6, cyclin E, p21, cyclin A and cyclin B) in HeLa cells and caused HeLa cells to accumulate in S phase during the early stage of infection. Monitoring SINV replication in HeLa and Vero cells expressing cell cycle indicators revealed that SINV which infected HeLa cells during G{sub 1} phase preferred to proliferate during S/G{sub 2} phase, and the average time interval for viral replication was significantly shorter in both HeLa and Vero cells infected during G{sub 1} phase than in cells infected during S/G{sub 2} phase. - Highlights: • SINV infection was able to alter the cell cycle progression of infected cancer cells. • SINV infection can affect the expression of cell cycle regulators. • SINV infection exhibited a preference for the timing of viral replication among the cell cycle phases.

Alteration of cell cycle progression by Sindbis virus infection

International Nuclear Information System (INIS)

Yi, Ruirong; Saito, Kengo; Isegawa, Naohisa; Shirasawa, Hiroshi

2015-01-01

We examined the impact of Sindbis virus (SINV) infection on cell cycle progression in a cancer cell line, HeLa, and a non-cancerous cell line, Vero. Cell cycle analyses showed that SINV infection is able to alter the cell cycle progression in both HeLa and Vero cells, but differently, especially during the early stage of infection. SINV infection affected the expression of several cell cycle regulators (CDK4, CDK6, cyclin E, p21, cyclin A and cyclin B) in HeLa cells and caused HeLa cells to accumulate in S phase during the early stage of infection. Monitoring SINV replication in HeLa and Vero cells expressing cell cycle indicators revealed that SINV which infected HeLa cells during G 1 phase preferred to proliferate during S/G 2 phase, and the average time interval for viral replication was significantly shorter in both HeLa and Vero cells infected during G 1 phase than in cells infected during S/G 2 phase. - Highlights: • SINV infection was able to alter the cell cycle progression of infected cancer cells. • SINV infection can affect the expression of cell cycle regulators. • SINV infection exhibited a preference for the timing of viral replication among the cell cycle phases

SAE 1045 steel/WC-Co/Ni-Cu-Ni/SAE 1045 steel joints prepared by dynamic diffusion bonding: Microelectrochemical studies in 0.6 M NaCl solution

International Nuclear Information System (INIS)

Andreatta, Francesco; Matesanz, Laura; Akita, Adriano H.; Paussa, Luca; Fedrizzi, Lorenzo; Fugivara, Cecilio S.; Gomez de Salazar, Jose M.; Benedetti, Assis V.

2009-01-01

Corrosion of SAE 1045 steel/WC-Co/Ni-Cu-Ni/SAE 1045 steel interfaces was investigated in 0.6 M NaCl solution using an electrochemical microcell, which enables local electrochemical characterization at the micrometer scale. Two pieces of steel, one with a WC-Co coating covered with Ni (12 μm) and Cu (5 μm) layers, and the other with a Ni (15 μm) layer, were welded by dynamic diffusion bonding. A WC-Co coating was applied to the steel by the high velocity oxygen-fuel process, and Ni-Cu and Ni layers by electroplating. Polarization curves were recorded using an electrochemical microcell. Different regions of welded samples were investigated, including steel, cermet coating, and steel/cermet and steel/Ni-Cu-Ni/cermet interfaces. Optical and electronic microscopes were employed to study the corroded regions. Potentiodynamic polarization curves obtained using the microcell revealed that the base metal was more susceptible to corrosion than the cermet. In addition, cermet steel/cermet and steel/Ni-Cu-Ni/cermet joints exhibited different breakdown potentials. Steel was strongly corroded in the regions adjacent to the interfaces, while the cermet was less corroded. Iron oxides/hydroxides and chloride salts were the main corrosion products of steel. After removal of the superficial layer of corrosion products, iron oxides were mainly observed. Chloride ions were detected mainly on a copper-enriched layer placed between two Ni-enriched layers.

Enhancement of surface properties of SAE 1020 by chromium plasma immersion recoil implantation

International Nuclear Information System (INIS)

Ueda, M.; Mello, C.B.; Beloto, A.F.; Rossi, J.O.; Reuther, H.

2007-01-01

SAE 1020 steel is commonly used as concrete reinforcement and small machine parts, but despite its good mechanical properties, as ductility, hardness and wear resistance, it is susceptible to severe corrosion. It is well known that chromium content above 12% in Fe alloys increases their corrosion resistance. In order to obtain this improvement, we studied the introduction of chromium atoms into the matrix of SAE 1020 steel by recoil implantation process using a plasma immersion ion implantation (PIII) system. Potentiodynamic scans showed that the presence of Cr film leads to a gain in the corrosion potential, from -650 mV to -400 mV. After PIII treatment, the corrosion potential increased further to -340 mV, but the corrosion current density presented no significant change. Vickers microhardness tests showed surface hardness increase of up to about 27% for the treated samples. Auger electron spectroscopy showed that, for a 30 nm film, Cr was introduced for about 20 nm into the steel matrix. Tribology tests, of pin-on-disk type, showed that friction coefficient of treated samples was reduced by about 50% and a change in wear mechanism, from adhesive to abrasive mode, occurred

Room temperature fatigue behaviour of a normalized steel SAE 4140 in torsion

International Nuclear Information System (INIS)

Klumpp, S.; Eifler, D.; Macherauch, E.

1990-01-01

Cyclic deformation behaviour of a normalized steel SAE 4140 in shear strain-controlled torsion is characterized by cyclic softening and cyclic hardening. If mean shear stresses are superimposed to an alternating shear stress, cycle-dependent creep occurs, and the number of cycles to failure decreases. In shear strain-controlled torsional loading, mean stresses are observed to relax nearly to zero within a few cycles. Fatigue life is not influenced by mean shear strains. (orig.) [de

Mycobacterium tuberculosis infection induces non-apoptotic cell death of human dendritic cells

LENUS (Irish Health Repository)

Ryan, Ruth CM

2011-10-24

Abstract Background Dendritic cells (DCs) connect innate and adaptive immunity, and are necessary for an efficient CD4+ and CD8+ T cell response after infection with Mycobacterium tuberculosis (Mtb). We previously described the macrophage cell death response to Mtb infection. To investigate the effect of Mtb infection on human DC viability, we infected these phagocytes with different strains of Mtb and assessed viability, as well as DNA fragmentation and caspase activity. In parallel studies, we assessed the impact of infection on DC maturation, cytokine production and bacillary survival. Results Infection of DCs with live Mtb (H37Ra or H37Rv) led to cell death. This cell death proceeded in a caspase-independent manner, and without nuclear fragmentation. In fact, substrate assays demonstrated that Mtb H37Ra-induced cell death progressed without the activation of the executioner caspases, 3\\/7. Although the death pathway was triggered after infection, the DCs successfully underwent maturation and produced a host-protective cytokine profile. Finally, dying infected DCs were permissive for Mtb H37Ra growth. Conclusions Human DCs undergo cell death after infection with live Mtb, in a manner that does not involve executioner caspases, and results in no mycobactericidal effect. Nonetheless, the DC maturation and cytokine profile observed suggests that the infected cells can still contribute to TB immunity.

Coronavirus infection of polarized epithelial cells

NARCIS (Netherlands)

Rossen, J W; Horzinek, M C; Rottier, P J

1995-01-01

Epithelial cells are the first host cells to be infected by incoming c oronaviruses. Recent observations in vitro show that coronaviruses are released from a specific side of these polarized cells, and this polarized release might be important for the spread of the infection in vivo. Mechanisms for

Coincident resection at both ends of random, γ-induced double-strand breaks requires MRX (MRN, Sae2 (Ctp1, and Mre11-nuclease.

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James W Westmoreland

2013-03-01

Full Text Available Resection is an early step in homology-directed recombinational repair (HDRR of DNA double-strand breaks (DSBs. Resection enables strand invasion as well as reannealing following DNA synthesis across a DSB to assure efficient HDRR. While resection of only one end could result in genome instability, it has not been feasible to address events at both ends of a DSB, or to distinguish 1- versus 2-end resections at random, radiation-induced "dirty" DSBs or even enzyme-induced "clean" DSBs. Previously, we quantitatively addressed resection and the role of Mre11/Rad50/Xrs2 complex (MRX at random DSBs in circular chromosomes within budding yeast based on reduced pulsed-field gel electrophoretic mobility ("PFGE-shift". Here, we extend PFGE analysis to a second dimension and demonstrate unique patterns associated with 0-, 1-, and 2-end resections at DSBs, providing opportunities to examine coincidence of resection. In G2-arrested WT, Δrad51 and Δrad52 cells deficient in late stages of HDRR, resection occurs at both ends of γ-DSBs. However, for radiation-induced and I-SceI-induced DSBs, 1-end resections predominate in MRX (MRN null mutants with or without Ku70. Surprisingly, Sae2 (Ctp1/CtIP and Mre11 nuclease-deficient mutants have similar responses, although there is less impact on repair. Thus, we provide direct molecular characterization of coincident resection at random, radiation-induced DSBs and show that rapid and coincident initiation of resection at γ-DSBs requires MRX, Sae2 protein, and Mre11 nuclease. Structural features of MRX complex are consistent with coincident resection being due to an ability to interact with both DSB ends to directly coordinate resection. Interestingly, coincident resection at clean I-SceI-induced breaks is much less dependent on Mre11 nuclease or Sae2, contrary to a strong dependence on MRX complex, suggesting different roles for these functions at "dirty" and clean DSB ends. These approaches apply to resection at

Polypeptide synthesis in alphavirus-infected aedes albopictus cells during the establishment of persistent infection

International Nuclear Information System (INIS)

Richardson, M.A.; Boulton, R.W.; Raghow, R.S.; Dalgarno, L.

1980-01-01

Polypeptide synthesis was examined in mosquito cells during the establishment of a persistent infection with two alphaviruses, Ross River virus (RRV) and Semliki Forest virus (SFV), and in vertebrate cells cytopathically-infected with the same viruses. In Aedes albopictus cells, RRV reached peak titres at 34-48 hours p.i. At 12 hours 85 per cent of cells assayed as infected by infective centre assay; by 48 hours when persistence was established, virus production was reduced and <5 per cent of cells assayed as infected. There was not shutdown of host polypeptide synthesis during infection. Viral polypeptide synthesis was maximal between 10 and 24 hours p.i. The major viral polypeptides labelled were nucleocapsid protein and envelope protein(s).The precursor polypeptide p95 which was prominent in infected BHK cells was not detected in mosquito cells. Similar results were obtained on SFV infection. During the establishment of persistence there was a coordinate decline in the synthesis of RRV polypeptides, reaching undetectable levels by 72 hours p.i. Subculturing persistently-infected cells led to a small increase in viral polypeptide synthesis and virus titre. In contrast, during RRV growth in BHK cells host protein synthesis was severely inhibited and by 9-11 hours p.i. virus-specific polypeptide synthesis represented more than 90 per cent of total protein synthetic activity. (author)

SAES St 909 pilot scale methane cracking tests

International Nuclear Information System (INIS)

Klein, J. E.; Sessions, H. T.

2008-01-01

Pilot scale (0.5 kg) SAES St 909 methane cracking tests were conducted for potential tritium process applications. Up to 1400 hours tests were done at 700 deg.C, 202.7 kPa (1520 torr) with a 0.03 sLPM feed of methane plus impurities, in a 20 vol% hydrogen, balance helium, stream. Carbon dioxide gettered by St 909 can be equated to an equivalent amount of methane gettered, but equating nitrogen to an equivalent amount of methane was nitrogen feed composition dependent. A decreased hydrogen feed increased methane getter rates while a 30 deg.C drop in one furnace zone increased methane emissions by over a factor of 30. The impact of gettered nitrogen can be somewhat minimized if nitrogen feed to the bed has been stopped and sufficient time given to recover the methane cracking rate. (authors)

HIV Cell-to-Cell Spread Results in Earlier Onset of Viral Gene Expression by Multiple Infections per Cell.

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Mikaël Boullé

2016-11-01

Full Text Available Cell-to-cell spread of HIV, a directed mode of viral transmission, has been observed to be more rapid than cell-free infection. However, a mechanism for earlier onset of viral gene expression in cell-to-cell spread was previously uncharacterized. Here we used time-lapse microscopy combined with automated image analysis to quantify the timing of the onset of HIV gene expression in a fluorescent reporter cell line, as well as single cell staining for infection over time in primary cells. We compared cell-to-cell spread of HIV to cell-free infection, and limited both types of transmission to a two-hour window to minimize differences due to virus transit time to the cell. The mean time to detectable onset of viral gene expression in cell-to-cell spread was accelerated by 19% in the reporter cell line and by 35% in peripheral blood mononuclear cells relative to cell-free HIV infection. Neither factors secreted by infected cells, nor contact with infected cells in the absence of transmission, detectably changed onset. We recapitulated the earlier onset by infecting with multiple cell-free viruses per cell. Surprisingly, the acceleration in onset of viral gene expression was not explained by cooperativity between infecting virions. Instead, more rapid onset was consistent with a model where the fastest expressing virus out of the infecting virus pool sets the time for infection independently of the other co-infecting viruses.

Brucella abortus-infected B cells induce osteoclastogenesis.

Science.gov (United States)

Pesce Viglietti, Ayelén Ivana; Arriola Benitez, Paula Constanza; Giambartolomei, Guillermo Hernán; Delpino, María Victoria

2016-09-01

Brucella abortus is an intracellular bacterium that establishes lifelong infections in livestock and humans although the mechanisms of its chronicity are poorly understood. Activated B cells have long lifespan and B. abortus infection activates B cells. Our results indicate that the direct infection of B cells with B. abortus induced matrix metalloproteinase-9 (MMP-9), receptor activator for NF κB ligand (RANKL), tumor necrosis factor (TNF)-α and interleukin (IL)-6 secretion. In addition, supernatants from B. abortus-infected B cells induced bone marrow-derived monocytes to undergo osteoclastogenesis. Using osteoprotegerin, RANKL's decoy receptor, we determined that RANKL is involved in osteoclastogenesis induced by supernatants from B. abortus-infected B cells. The results presented here shed light on how the interactions of B. abortus with B cells may have a role in the pathogenesis of brucellar osteoarticular disease. Copyright © 2016 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

Modeling of the flow behavior of SAE 8620H combing microstructure evolution in hot forming

Science.gov (United States)

Fu, Xiaobin; Wang, Baoyu; Tang, Xuefeng

2017-10-01

With the development of net-shape forming technology, hot forming process is widely applied to manufacturing gear parts, during which, materials suffer severe plastic distortion and microstructure changes continually. In this paper, to understand and model the flow behavior and microstructure evolution, SAE 8620H, a widely used gear steel, is selected as the object and the flow behavior and microstructure evolution are observed by an isothermal hot compression tests at 1273-1373 K with a strain rate of 0.1-10 s-1. Depending on the results of the compression test, a set of internal-state-variable based unified constitutive equations is put forward to describe the flow behavior and microstructure evaluation of SAE 8620H. Moreover, the evaluation of the dislocation density and the fraction of dynamic recrystallization based on the theory of thermal activation is modeled and reincorporated into the constitutive law. The material parameters in the constitutive model are calculated based on the measured flow stress and dynamic recrystallization fraction. The predicted flow stress under different deformation conditions has a good agreement with the measured results.

Demonstration of NK cell-mediated lysis of varicella-zoster virus (VZV)-infected cells: characterization of the effector cells

Energy Technology Data Exchange (ETDEWEB)

Tilden, A.B.; Cauda, R.; Grossi, C.E.; Balch, C.M.; Lakeman, A.D.; Whitley, R.J.

1986-06-01

Infection with varicella-zoster virus (VZV) rendered RAJI cells more susceptible to lysis by non-adherent blood lymphocytes. At an effector to target ratio of 80:1 the mean percentage of /sup 51/Cr release of VZV-infected RAJI cells was 41 +/- 12%, whereas that of uninfected RAJI cells was 15 +/- 6%. The increased susceptibility to lysis was associated with increased effector to target conjugate formation in immunofluorescence binding assays. The effector cells cytotoxic for VZV-infected RAJI cells were predominantly Leu-11a/sup +/ Leu-4/sup -/ granular lymphocytes as demonstrated by fluorescence-activated cell sorting. The effector cell active against VZV-infected RAJI cells appeared similar to those active against herpes simplex virus (HSV)-infected cells, because in cold target competition experiments the lysis of /sup 51/Cr-labeled VZV-infected RAJI cells was efficiently inhibited by either unlabeled VZV-infected RAJI cells (mean 71% inhibition, 2:1 ratio unlabeled to labeled target) or HSV-infected RAJI cells (mean 69% inhibition) but not by uninfected RAJI cells (mean 10% inhibition). In contrast, competition experiments revealed donor heterogeneity in the overlap between effector cells for VZV- or HSV-infected RAJI vs K-562 cells.

Mast cells in viral infections

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Piotr Witczak

2012-04-01

Full Text Available  There are some premises suggesting that mast cells are involved in the mechanisms of anti-virus defense and in viral disease pathomechanisms. Mast cells are particularly numerous at the portals of infections and thus may have immediate and easy contact with the external environment and invading pathogens. These cells express receptors responsible for recognition of virus-derived PAMP molecules, mainly Toll-like receptors (TLR3, TLR7/8 and TLR9, but also RIG-I-like and NOD-like molecules. Furthermore, mast cells generate various mediators, cytokines and chemokines which modulate the intensity of inflammation and regulate the course of innate and adaptive anti-viral immunity. Indirect evidence for the role of mast cells in viral infections is also provided by clinical observations and results of animal studies. Currently, more and more data indicate that mast cells can be infected by some viruses (dengue virus, adenoviruses, hantaviruses, cytomegaloviruses, reoviruses, HIV-1 virus. It is also demonstrated that mast cells can release pre formed mediators as well as synthesize de novo eicosanoids in response to stimulation by viruses. Several data indicate that virus-stimulated mast cells secrete cytokines and chemokines, including interferons as well as chemokines with a key role in NK and Tc lymphocyte influx. Moreover, some information indicates that mast cell stimulation via TLR3, TLR7/8 and TLR9 can affect their adhesion to extracellular matrix proteins and chemotaxis, and influence expression of some membrane molecules. Critical analysis of current data leads to the conclusion that it is not yet possible to make definitive statements about the role of mast cells in innate and acquired defense mechanisms developing in the course of viral infection and/or pathomechanisms of viral diseases.

COMPUTATIONAL FLOW MODELLING OF FORMULA-SAE SIDEPODS FOR OPTIMUM RADIATOR HEAT MANAGEMENT

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C. M. DE SILVA

2011-02-01

Full Text Available Formula SAE vehicles, over the program’s history have showcased a myriad of aerodynamic packages, each claiming specific quantitative and qualitative features. This paper attempts to critique differing aerodynamic sidepod designs and their effect upon radiator heat management. Various features from inlet size, sidepod shape and size, presence of an undertray, suspension cover, gills and chimneys are analysed for their effects. Computational Fluid Dynamics (CFD analyses are performed in the FLUENT environment, with the aid of GAMBIT meshing software and SolidWorks modelling.

Combination of anti-retroviral drugs and radioimmunotherapy specifically kills infected cells from HIV infected individuals

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Dina Tsukrov

2016-09-01

Full Text Available Eliminating virally infected cells is an essential component of any HIV eradication strategy. Radioimmunotherapy (RIT, a clinically established method for killing cells using radiolabeled antibodies, was recently applied to target HIV-1 gp41 antigen expressed on the surface of infect-ed cells. Since gp41 expression by infected cells is likely down-regulated in patients on an-tiretroviral therapy (ART, we evaluated the ability of RIT to kill ART-treated infected cells us-ing both in vitro models and lymphocytes isolated from HIV-infected subjects. Human peripheral blood mononuclear cells (PBMCs were infected with HIV and cultured in the presence of two clinically relevant ART combinations. Scatchard analysis of the 2556 human monoclonal anti-body to HIV gp41 binding to the infected and ART-treated cells demonstrated sufficient residual expression of gp41 on the cell surface to warrant subsequent RIT. This is the first time the quantification of gp41 post-ART is being reported. Cells were then treated with Bismuth-213-labeled 2556 antibody. conjugated to the human monoclonal antibody 2556, which binds to HIV gp41. Cell survival was quantified by Trypan blue and residual viremia by p24 ELISA. Cell surface gp41 expression was assessed by Scatchard analysis. The experiments were repeated using PBMCs isolated from blood specimens obtained from 15 HIV-infected individuals: ten on ART and five ART-naive. We found that 213Bi-2556 killed ART-treated infected PBMCs and reduced viral production to undetectable levels. ART and RIT co-treatment was more effective at reducing viral load in vitro than either therapy alone, indicating that gp41 expression under ART was sufficient to allow 213Bi-2556 to deliver cytocidal doses of radiation to infected cells. This study provides proof of concept that 213Bi-2556 may represent an innovative and effective targeting method for killing HIV-infected cells treated with ART, and supports continued development of 213Bi

Fungal cell gigantism during mammalian infection.

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Oscar Zaragoza

2010-06-01

Full Text Available The interaction between fungal pathogens with the host frequently results in morphological changes, such as hyphae formation. The encapsulated pathogenic fungus Cryptococcus neoformans is not considered a dimorphic fungus, and is predominantly found in host tissues as round yeast cells. However, there is a specific morphological change associated with cryptococcal infection that involves an increase in capsule volume. We now report another morphological change whereby gigantic cells are formed in tissue. The paper reports the phenotypic characterization of giant cells isolated from infected mice and the cellular changes associated with giant cell formation. C. neoformans infection in mice resulted in the appearance of giant cells with cell bodies up to 30 microm in diameter and capsules resistant to stripping with gamma-radiation and organic solvents. The proportion of giant cells ranged from 10 to 80% of the total lung fungal burden, depending on infection time, individual mice, and correlated with the type of immune response. When placed on agar, giant cells budded to produce small daughter cells that traversed the capsule of the mother cell at the speed of 20-50 m/h. Giant cells with dimensions that approximated those in vivo were observed in vitro after prolonged culture in minimal media, and were the oldest in the culture, suggesting that giant cell formation is an aging-dependent phenomenon. Giant cells recovered from mice displayed polyploidy, suggesting a mechanism by which gigantism results from cell cycle progression without cell fission. Giant cell formation was dependent on cAMP, but not on Ras1. Real-time imaging showed that giant cells were engaged, but not engulfed by phagocytic cells. We describe a remarkable new strategy for C. neoformans to evade the immune response by enlarging cell size, and suggest that gigantism results from replication without fission, a phenomenon that may also occur with other fungal pathogens.

Fungal cell gigantism during mammalian infection.

Science.gov (United States)

Zaragoza, Oscar; García-Rodas, Rocío; Nosanchuk, Joshua D; Cuenca-Estrella, Manuel; Rodríguez-Tudela, Juan Luis; Casadevall, Arturo

2010-06-17

The interaction between fungal pathogens with the host frequently results in morphological changes, such as hyphae formation. The encapsulated pathogenic fungus Cryptococcus neoformans is not considered a dimorphic fungus, and is predominantly found in host tissues as round yeast cells. However, there is a specific morphological change associated with cryptococcal infection that involves an increase in capsule volume. We now report another morphological change whereby gigantic cells are formed in tissue. The paper reports the phenotypic characterization of giant cells isolated from infected mice and the cellular changes associated with giant cell formation. C. neoformans infection in mice resulted in the appearance of giant cells with cell bodies up to 30 microm in diameter and capsules resistant to stripping with gamma-radiation and organic solvents. The proportion of giant cells ranged from 10 to 80% of the total lung fungal burden, depending on infection time, individual mice, and correlated with the type of immune response. When placed on agar, giant cells budded to produce small daughter cells that traversed the capsule of the mother cell at the speed of 20-50 m/h. Giant cells with dimensions that approximated those in vivo were observed in vitro after prolonged culture in minimal media, and were the oldest in the culture, suggesting that giant cell formation is an aging-dependent phenomenon. Giant cells recovered from mice displayed polyploidy, suggesting a mechanism by which gigantism results from cell cycle progression without cell fission. Giant cell formation was dependent on cAMP, but not on Ras1. Real-time imaging showed that giant cells were engaged, but not engulfed by phagocytic cells. We describe a remarkable new strategy for C. neoformans to evade the immune response by enlarging cell size, and suggest that gigantism results from replication without fission, a phenomenon that may also occur with other fungal pathogens.

Application of digital image processing to determine the causes of failures in SAE 1018 carbon steel; Aplicacion del procesamiento digital de imagenes para la determinacion de las causas que provocan fallas en el acero al carbono SAE 1018

Energy Technology Data Exchange (ETDEWEB)

Bolanos-Rodriguez, E [Escuela Superior de Tizayuca, Universidad Autonoma del Estado de Hidalgo, Tizayuca, Hidalgo (Mexico)]. E-mail: bola7112@yahoo.com.mx; Gonzalez-Islas, J.C. [Universidad Tecnologica de Tulancingo, Tulancingo, Hidalgo (Mexico)]. E-mail: juanc.gonzalez@utec-tgo.edu.mx; Felipe-Riveron, E.M. [Centro de Investigacion en Computacion, Instituto Politecnico Nacional, Mexico, D.F. (Mexico)]. E-mail: edgardo@cic.ipn.mx

2013-03-15

The research is focused on determining the causes of failures in the SAE 1018 carbon steel that is used in pipes carrying drinking water. Digital processing of images captured by photomicrographs by scanning electron microscopy coupled with the technique of X-ray microanalysis. The results demonstrate that the low quality of the raw materials and the manufacturing process of steel are the reasons for the occurrence of cracks observed in the material. [Spanish] La investigacion se centra en determinar las causas que provocan fallas en el acero al carbono SAE 1018 que es utilizado en tuberias que transportan agua potable. Se emplea el procesamiento digital de imagenes de las microfotografias captadas por medio de microscopia electronica de barrido, unido a la tecnica de microanalisis de rayos X. Los resultados obtenidos demuestran que la baja calidad de las materias primas y el proceso de manufactura del acero son las razones de la ocurrencia de fisuras observadas en el material.

Serviço de assistência especializada (SAE: uma experiência profissional

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Carla Glenda Souza da Silva

Full Text Available O trabalho visa a discutir o processo de implantação e operacionalização de um serviço de assistência especializada (SAE, que objetiva vincular o usuário portador do vírus HIV/Aids à equipe multiprofissional através da descrição de experiência profissional. Alcançou-se a implantação e operacionalização do SAE com a implementação de uma política de assistência voltada para o bem-estar, a humanização e a qualidade de vida do usuário. A implantação do serviço permitiu garantir a resolutividade, a eqüidade e a integralidade dos serviços no Município de Cabedelo, bem como a ampliação do processo de descentralização da atenção, conforme orienta a política institucional do SUS no Brasil, mesmo enfrentando dificuldades de ordem política, econômica e estrutural local. A cidade possui uma população de 47.076 habitantes. Há oitenta e seis casos de Aids notificados no período de jun./85 a fev./05. É um Município predominantemente urbano, litorâneo, portuário, é rota de caminhoneiros, possui um ferry boat, e apresenta elevado índice de prostituição.

Diseño de manguetas delanteras y posteriores de un vehículo monoplaza para la competición en la formula SAE

OpenAIRE

Mejía González, David Santiago

2015-01-01

En este trabajo de investigación se describe y aplica el proceso genérico de desarrollo de nuevos productos para el diseño de las manguetas delanteras y posteriores para un vehículo de competencia F-SAE. Que cumpla con las características de funcionamiento y las normas técnicas que dicta el reglamento emitido por los organizadores de la competencia F-SAE 2014. This research describes and applies the generic process of developing new products for the design of the front and rear uprights fo...

Intracellular Events and Cell Fate in Filovirus Infection

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Elena Ryabchikova

2011-08-01

Full Text Available Marburg and Ebola viruses cause a severe hemorrhagic disease in humans with high fatality rates. Early target cells of filoviruses are monocytes, macrophages, and dendritic cells. The infection spreads to the liver, spleen and later other organs by blood and lymph flow. A hallmark of filovirus infection is the depletion of non-infected lymphocytes; however, the molecular mechanisms leading to the observed bystander lymphocyte apoptosis are poorly understood. Also, there is limited knowledge about the fate of infected cells in filovirus disease. In this review we will explore what is known about the intracellular events leading to virus amplification and cell damage in filovirus infection. Furthermore, we will discuss how cellular dysfunction and cell death may correlate with disease pathogenesis.

Kinetics of liver macrophages (Kupffer cells) in SIV-infected macaques

International Nuclear Information System (INIS)

Ahsan, Muhammad H.; Gill, Amy F.; Alvarez, Xavier; Lackner, Andrew A.; Veazey, Ronald S.

2013-01-01

Since the liver drains antigens from the intestinal tract, and since the intestinal tract is a major site of viral replication, we examined the dynamics of liver macrophages (Kupffer cells) throughout SIV infection. Absolute numbers of Kupffer cells increased in the livers in acute infection, and in animals with AIDS. Significantly higher percentages of proliferating (BrdU+) Kupffer cells were detected in acute infection and in AIDS with similar trends in blood monocytes. Significantly higher percentages of apoptotic (AC3+) Kupffer cells were also found in acute and AIDS stages. However, productively infected cells were not detected in liver of 41/42 animals examined, despite abundant infected cells in gut and lymph nodes of all animals. Increased rates of Kupffer cell proliferation resulting in an increase in Kupffer cells without productive infection indicate SIV infection affects Kupffer cells, but the liver does not appear to be a major site of productive viral replication. - Highlights: • Kupffer cells increase in the liver of SIV-infected macaques. • Increased proliferation and apoptosis of Kupffer cells occurs in SIV infection. • Productively infected cells are rarely detected in the liver. • The liver is not a major site for SIV replication

NKT cell depletion in humans during early HIV infection.

Science.gov (United States)

Fernandez, Caroline S; Kelleher, Anthony D; Finlayson, Robert; Godfrey, Dale I; Kent, Stephen J

2014-08-01

Natural killer T (NKT) cells bridge across innate and adaptive immune responses and have an important role in chronic viral infections such as human immunodeficiency virus (HIV). NKT cells are depleted during chronic HIV infection, but the timing, drivers and implications of this NKT cell depletion are poorly understood. We studied human peripheral blood NKT cell levels, phenotype and function in 31 HIV-infected subjects not on antiretroviral treatment from a mean of 4 months to 2 years after HIV infection. We found that peripheral CD4(+) NKT cells were substantially depleted and dysfunctional by 4 months after HIV infection. The depletion of CD4(+) NKT cells was more marked than the depletion of total CD4(+) T cells. Further, the early depletion of NKT cells correlated with CD4(+) T-cell decline, but not HIV viral levels. Levels of activated CD4(+) T cells correlated with the loss of NKT cells. Our studies suggest that the early loss of NKT cells is associated with subsequent immune destruction during HIV infection.

Alemtuzumab-induced elimination of HIV-1-infected immune cells.

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Ruxrungtham, Kiat; Sirivichayakul, Sunee; Buranapraditkun, Supranee; Krause, Werner

2016-01-01

Currently, there is no drug known that is able to eradicate either HIV or HIV-infected host cells. The effectiveness of all available treatments is based on the prevention of viral replication. We investigated whether the monoclonal, CD52 receptor-targeting antibody, alemtuzumab, which is currently approved for the treatment of multiple sclerosis, is able to eliminate HIV-infected immune cells. In blood samples from healthy donors and from HIV-1-infected subjects who were either treatment-naïve or resistant to HAART, we studied whether the CD52 expression on T cells and their subsets (CD3, CD4, CD8), B cells (CD19), dendritic cells (CD123) and monocytes (CD11c) is retained in HIV-1 infection and whether alemtuzumab is able to eradicate infected cells, using four-colour flow cytometry. We found that CD52 expression on immune cells is retained in HIV-1 infection regardless of CD4 cell count, viral load and treatment status, and is amenable to alemtuzumab-induced depletion. For the first time it could be shown in vitro that HIV-1-infected immune cells can be eliminated by using the monoclonal antibody alemtuzumab.

Analysis and functional annotation of expressed sequence tags from in vitro cell lines of elasmobranchs: Spiny dogfish shark (Squalus acanthias) and little skate (Leucoraja erinacea).

Science.gov (United States)

Parton, Angela; Bayne, Christopher J; Barnes, David W

2010-09-01

Elasmobranchs are the most commonly used experimental models among the jawed, cartilaginous fish (Chondrichthyes). Previously we developed cell lines from embryos of two elasmobranchs, Squalus acanthias the spiny dogfish shark (SAE line), and Leucoraja erinacea the little skate (LEE-1 line). From these lines cDNA libraries were derived and expressed sequence tags (ESTs) generated. From the SAE cell line 4303 unique transcripts were identified, with 1848 of these representing unknown sequences (showing no BLASTX identification). From the LEE-1 cell line, 3660 unique transcripts were identified, and unknown, unique sequences totaled 1333. Gene Ontology (GO) annotation showed that GO assignments for the two cell lines were in general similar. These results suggest that the procedures used to derive the cell lines led to isolation of cell types of the same general embryonic origin from both species. The LEE-1 transcripts included GO categories "envelope" and "oxidoreductase activity" but the SAE transcripts did not. GO analysis of SAE transcripts identified the category "anatomical structure formation" that was not present in LEE-1 cells. Increased organelle compartments may exist within LEE-1 cells compared to SAE cells, and the higher oxidoreductase activity in LEE-1 cells may indicate a role for these cells in responses associated with innate immunity or in steroidogenesis. These EST libraries from elasmobranch cell lines provide information for assembly of genomic sequences and are useful in revealing gene diversity, new genes and molecular markers, as well as in providing means for elucidation of full-length cDNAs and probes for gene array analyses. This is the first study of this type with members of the Chondrichthyes. Copyright 2010 Elsevier Inc. All rights reserved.

Nipah virus infection and glycoprotein targeting in endothelial cells

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Maisner Andrea

2010-11-01

Full Text Available Abstract Background The highly pathogenic Nipah virus (NiV causes fatal respiratory and brain infections in animals and humans. The major hallmark of the infection is a systemic endothelial infection, predominantly in the CNS. Infection of brain endothelial cells allows the virus to overcome the blood-brain-barrier (BBB and to subsequently infect the brain parenchyma. However, the mechanisms of NiV replication in endothelial cells are poorly elucidated. We have shown recently that the bipolar or basolateral expression of the NiV surface glycoproteins F and G in polarized epithelial cell layers is involved in lateral virus spread via cell-to-cell fusion and that correct sorting depends on tyrosine-dependent targeting signals in the cytoplasmic tails of the glycoproteins. Since endothelial cells share many characteristics with epithelial cells in terms of polarization and protein sorting, we wanted to elucidate the role of the NiV glycoprotein targeting signals in endothelial cells. Results As observed in vivo, NiV infection of endothelial cells induced syncytia formation. The further finding that infection increased the transendothelial permeability supports the idea of spread of infection via cell-to-cell fusion and endothelial cell damage as a mechanism to overcome the BBB. We then revealed that both glycoproteins are expressed at lateral cell junctions (bipolar, not only in NiV-infected primary endothelial cells but also upon stable expression in immortalized endothelial cells. Interestingly, mutation of tyrosines 525 and 542/543 in the cytoplasmic tail of the F protein led to an apical redistribution of the protein in endothelial cells whereas tyrosine mutations in the G protein had no effect at all. This fully contrasts the previous results in epithelial cells where tyrosine 525 in the F, and tyrosines 28/29 in the G protein were required for correct targeting. Conclusion We conclude that the NiV glycoprotein distribution is responsible for

Diseño de un prototipo de volante para un vehículo tipo formula SAE mediante técnicas de optimización estructural

OpenAIRE

Picón Mejía, Jaime Andrés

2016-01-01

En este proyecto técnico se presenta una metodología para el diseño y construcción de un prototipo de volante de dirección para un vehículo tipo formula SAE, aplicando técnicas de optimización estructural considerando procesos de manufactura aditiva. In this technical project, a methodology is presented for the design and construction of a prototype steering wheel for a Formula SAE vehicle, using structural optimization techniques considering additive manufacturing processes.

Stem Cell Transplant Patients and Fungal Infections

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... Foodborne, Waterborne, and Environmental Diseases Mycotic Diseases Branch Stem Cell Transplant Patients and Fungal Infections Recommend on Facebook ... Mold . Top of Page Preventing fungal infections in stem cell transplant patients Fungi are difficult to avoid because ...

Establishment of human papillomavirus infection requires cell cycle progression.

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Dohun Pyeon

2009-02-01

Full Text Available Human papillomaviruses (HPVs are DNA viruses associated with major human cancers. As such there is a strong interest in developing new means, such as vaccines and microbicides, to prevent HPV infections. Developing the latter requires a better understanding of the infectious life cycle of HPVs. The HPV infectious life cycle is closely linked to the differentiation state of the stratified epithelium it infects, with progeny virus only made in the terminally differentiating suprabasal compartment. It has long been recognized that HPV must first establish its infection within the basal layer of stratified epithelium, but why this is the case has not been understood. In part this restriction might reflect specificity of expression of entry receptors. However, this hypothesis could not fully explain the differentiation restriction of HPV infection, since many cell types can be infected with HPVs in monolayer cell culture. Here, we used chemical biology approaches to reveal that cell cycle progression through mitosis is critical for HPV infection. Using infectious HPV16 particles containing the intact viral genome, G1-synchronized human keratinocytes as hosts, and early viral gene expression as a readout for infection, we learned that the recipient cell must enter M phase (mitosis for HPV infection to take place. Late M phase inhibitors had no effect on infection, whereas G1, S, G2, and early M phase cell cycle inhibitors efficiently prevented infection. We conclude that host cells need to pass through early prophase for successful onset of transcription of the HPV encapsidated genes. These findings provide one reason why HPVs initially establish infections in the basal compartment of stratified epithelia. Only this compartment of the epithelium contains cells progressing through the cell cycle, and therefore it is only in these cells that HPVs can establish their infection. By defining a major condition for cell susceptibility to HPV infection, these

Poliovirus mutants excreted by a chronically infected hypogammaglobulinemic patient establish persistent infections in human intestinal cells

International Nuclear Information System (INIS)

Labadie, Karine; Pelletier, Isabelle; Saulnier, Aure; Martin, Javier; Colbere-Garapin, Florence

2004-01-01

Immunodeficient patients whose gut is chronically infected by vaccine-derived poliovirus (VDPV) may excrete large amounts of virus for years. To investigate how poliovirus (PV) establishes chronic infections in the gut, we tested whether it is possible to establish persistent VDPV infections in human intestinal Caco-2 cells. Four type 3 VDPV mutants, representative of the viral evolution in the gut of a hypogammaglobulinemic patient over almost 2 years [J. Virol. 74 (2000) 3001], were used to infect both undifferentiated, dividing cells, and differentiated, polarized enterocytes. A VDPV mutant excreted 36 days postvaccination by the patient was lytic in both types of intestinal cell cultures, like the parental Sabin 3 (S3) strain. In contrast, three VDPVs excreted 136, 442, and 637 days postvaccination, established persistent infections both in undifferentiated cells and in enterocytes. Thus, viral determinants selected between day 36 and 136 conferred on VDPV mutants the capacity to infect intestinal cells persistently. The percentage of persistently VDPV-infected cultures was higher in enterocytes than in undifferentiated cells, implicating cellular determinants involved in the differentiation of enterocytes in persistent VDPV infections. The establishment of persistent infections in enterocytes was not due to poor replication of VDPVs in these cells, but was associated with reduced viral adsorption to the cell surface

Novel Treatment of Staphylococcus aureus Device-Related Infections Using Fibrinolytic Agents.

Science.gov (United States)

Hogan, S; O'Gara, J P; O'Neill, E

2018-02-01

Staphylococcal infections involving biofilms represent a significant challenge in the treatment of patients with device-related infections. Staphylococcus aureus biofilms have been shown to be SaeRS regulated and dependent on the coagulase-catalyzed conversion of fibrinogen into fibrin on surfaces coated with human plasma. Here we investigated the treatment of staphylococcal biofilm device-related infections by digesting the fibrin biofilm matrix with and without existing antimicrobials. The fibrinolytic agents plasmin, streptokinase, and nattokinase, and TrypLE, a recombinant trypsin-like protease, were used to digest and treat S. aureus biofilms grown in vitro using in vivo -like static biofilm assays with and without antimicrobials. Cytotoxicity, the potential to induce a cytokine response in whole human blood, and the risk of induction of tolerance to fibrinolytic agents were investigated. A rat model of intravascular catheter infection was established to investigate the efficacy of selected fibrinolytic agents in vivo Under biomimetic conditions, the fibrinolytic agents effectively dispersed established S. aureus biofilms and, in combination with common antistaphylococcal antimicrobials, effectively killed bacterial cells being released from the biofilm. These fibrinolytic agents were not cytotoxic and did not affect the host immune response. The rat model of infection successfully demonstrated the activity of the selected fibrinolytic agents alone and in combination with antimicrobials on established biofilms in vivo TrypLE and nattokinase most successfully removed adherent cells from plasma-coated surfaces and significantly improved the efficacy of existing antimicrobials against S. aureus biofilms in vitro and in vivo These biofilm dispersal agents represent a viable future treatment option for S. aureus device-related infections. Copyright © 2018 American Society for Microbiology.

Summary Report on the SAE 2016 Range Extenders for Electric Vehicles Symposium

Energy Technology Data Exchange (ETDEWEB)

Curran, Scott [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Wagner, Robert M. [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Campbell, Russ [SRA International, Inc., Arlington, VA (United States)

2017-03-01

The SAE 2016 Range Extenders for Electric Vehicles Symposium was a 2-day technical meeting focused on the role of advanced internal combustion engines (ICEs) and other novel energy converter technologies for extending the range of electric vehicles (EVs). The first-of-its-kind symposium was notable for focusing solely on the range extender (REx) technologies and not the EVs. The technical program featured presentations from international leaders from industry, government, national laboratories, and academia. The opening keynote presentations covered a broad range of topics including consumer behavior, policy implications, regulatory considerations, and REx architectures as enablers for advanced technologies. The technical sessions focused on an array of REx technologies including conventional ICEs, as well as less conventional or emerging technologies such as microturbines, fuel cells, low-temperature combustion engines, and aluminum-air batteries. The symposium included two panel sessions. The trend toward increasing vehicle electrification and the changing role of ICEs and other auxiliary power unit technologies for use as REx’s is leading to new research and design development needs. The symposium captured the interest of the industry and research communities in exploring the opportunities and challenges associated with REx’s for EVs. This report includes key takeaways, summarized below, and draft notes for each presentation and panel discussion.

Infection of endothelial cells by common human viruses.

Science.gov (United States)

Friedman, H M

1989-01-01

Common human viruses were evaluated for their ability to replicate in the endothelial cells of human umbilical vein and bovine thoracic aorta in vitro. Infection occurred with most viruses. The susceptibilities of endothelial cells derived from bovine aorta, pulmonary artery, and vena cava were compared. Among the viruses studied, no differences were noted in the ability to grow in endothelial cells from these three large vessels. One virus, herpes simplex virus type 1, was evaluated for its ability to produce persistent infection of endothelial cells. Infection developed and persisted for up to 3 months. After the first week, productive infection was found in less than 1% of cells. Nevertheless, the infection markedly affected the growth and morphology of the endothelial monolayer. Infection with any of several different viruses was noted to alter endothelial cell functions, including adherence of granulocytes, production of colony-stimulating factor, and synthesis of matrix protein. In addition, herpes simplex virus type 1 induced receptors for the Fc portion of IgG and for complement component C3b. These findings indicate that common human viruses can profoundly affect the biology of the endothelium.

Interaction of Staphylococci with Human B cells.

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Tyler K Nygaard

Full Text Available Staphylococcus aureus is a leading cause of human infections worldwide. The pathogen produces numerous molecules that can interfere with recognition and binding by host innate immune cells, an initial step required for the ingestion and subsequent destruction of microbes by phagocytes. To better understand the interaction of this pathogen with human immune cells, we compared the association of S. aureus and S. epidermidis with leukocytes in human blood. We found that a significantly greater proportion of B cells associated with S. epidermidis relative to S. aureus. Complement components and complement receptors were important for the binding of B cells with S. epidermidis. Experiments using staphylococci inactivated by ultraviolet radiation and S. aureus isogenic deletion mutants indicated that S. aureus secretes molecules regulated by the SaeR/S two-component system that interfere with the ability of human B cells to bind this bacterium. We hypothesize that the relative inability of B cells to bind S. aureus contributes to the microbe's success as a human pathogen.

Early events associated with infection of Epstein-Barr virus infection of primary B-cells.

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Sabyasachi Halder

2009-09-01

Full Text Available Epstein Barr virus (EBV is closely associated with the development of a vast number of human cancers. To develop a system for monitoring early cellular and viral events associated with EBV infection a self-recombining BAC containing 172-kb of the Epstein Barr virus genome BAC-EBV designated as MD1 BAC (Chen et al., 2005, J.Virology was used to introduce an expression cassette of green fluorescent protein (GFP by homologous recombination, and the resultant BAC clone, BAC-GFP-EBV was transfected into the HEK 293T epithelial cell line. The resulting recombinant GFP EBV was induced to produce progeny virus by chemical inducer from the stable HEK 293T BAC GFP EBV cell line and the virus was used to immortalize human primary B-cell as monitored by green fluorescence and outgrowth of the primary B cells. The infection, B-cell activation and cell proliferation due to GFP EBV was monitored by the expression of the B-cell surface antigens CD5, CD10, CD19, CD23, CD39, CD40 , CD44 and the intercellular proliferation marker Ki-67 using Flow cytometry. The results show a dramatic increase in Ki-67 which continues to increase by 6-7 days post-infection. Likewise, CD40 signals showed a gradual increase, whereas CD23 signals were increased by 6-12 hours, maximally by 3 days and then decreased. Monitoring the viral gene expression pattern showed an early burst of lytic gene expression. This up-regulation of lytic gene expression prior to latent genes during early infection strongly suggests that EBV infects primary B-cell with an initial burst of lytic gene expression and the resulting progeny virus is competent for infecting new primary B-cells. This process may be critical for establishment of latency prior to cellular transformation. The newly infected primary B-cells can be further analyzed for investigating B cell activation due to EBV infection.

Human innate lymphoid cells (ILCs) in filarial infections.

Science.gov (United States)

Bonne-Année, S; Nutman, T B

2018-02-01

Filarial infections are characteristically chronic and can cause debilitating diseases governed by parasite-induced innate and adaptive immune responses. Filarial parasites traverse or establish niches in the skin (migrating infective larvae), in nonmucosal tissues (adult parasite niche) and in the blood or skin (circulating microfilariae) where they intersect with the host immune response. While several studies have demonstrated that filarial parasites and their antigens can modulate myeloid cells (monocyte, macrophage and dendritic cell subsets), T- and B-lymphocytes and skin resident cell populations, the role of innate lymphoid cells during filarial infections has only recently emerged. Despite the identification and characterization of innate lymphoid cells (ILCs) in murine helminth infections, little is actually known about the role of human ILCs during parasitic infections. The focus of this review will be to highlight the composition of ILCs in the skin, lymphatics and blood; where the host-parasite interaction is well-defined and to examine the role of ILCs during filarial infections. Published 2017. This article is a U.S. Government work and is in the public domain in the USA.

Evidence of the pitting corrosion induced embrittlement of the structural steel SAE 8620

International Nuclear Information System (INIS)

Atanazio Filho, Nelson do Nascimento; Mansur, Tanius Rodrigues; Rabello, Emerson Giovani

2007-01-01

The influence of an aggressive environment (NaCl 3.5% aerated solution) on fatigue crack initiation and crack growth behavior were studied. This study comprised corrosion fatigue tests using specimens of SAE 8620 steel. The decreasing cyclic frequency (60 Hz to 11.7 Hz) effect on corrosion fatigue crack initiation behavior was examined. The tests carried out under rotating-bending loading conditions at 11.7 Hz (700 rpm), showed that pitting corrosion caused by anodic attack was responsible for corrosion fatigue crack initiation (author)

Semen CD4+ T Cells and Macrophages Are Productively Infected at All Stages of SIV infection in Macaques

Science.gov (United States)

Bernard-Stoecklin, Sibylle; Gommet, Céline; Corneau, Aurélien B.; Guenounou, Sabrina; Torres, Claire; Dejucq-Rainsford, Nathalie; Cosma, Antonio; Dereuddre-Bosquet, Nathalie; Le Grand, Roger

2013-01-01

The mucosal events of HIV transmission have been extensively studied, but the role of infected cells present in the genital and rectal secretions, and in the semen, in particular, remains a matter of debate. As a prerequisite to a thorough in vivo investigation of the early transmission events through infected cells, we characterized in detail by multi-parameter flow cytometry the changes in macaque seminal leukocytes during SIVmac251 infection, focusing on T cells, macrophages and dendritic cells. Using immunocytofluorescence targeting SIV proteins and real-time quantitative PCR targeting SIV DNA, we investigated the nature of the infected cells on sorted semen leukocytes from macaques at different stages of infection. Finally, we cocultured semen CD4+ T cells and macrophages with a cell line permissive to SIV infection to assess their infectivity in vitro. We found that primary infection induced strong local inflammation, which was associated with an increase in the number of leukocytes in semen, both factors having the potential to favor cell-associated virus transmission. Semen CD4+ T cells and macrophages were productively infected at all stages of infection and were infectious in vitro. Lymphocytes had a mucosal phenotype and expressed activation (CD69 & HLA-DR) and migration (CCR5, CXCR4, LFA-1) markers. CD69 expression was increased in semen T cells by SIV infection, at all stages of infection. Macrophages predominated at all stages and expressed CD4, CCR5, MAC-1 and LFA-1. Altogether, we demonstrated that semen contains the two major SIV-target cells (CD4+ T cells and macrophages). Both cell types can be productively infected at all stages of SIV infection and are endowed with markers that may facilitate transmission of infection during sexual exposure. PMID:24348253

Infectious mononucleosis accompanied by clonal proliferation of EBV-infected cells and infection of CD8-positive cells.

Science.gov (United States)

Arai, Ayako; Yamaguchi, Takeshi; Komatsu, Honami; Imadome, Ken-Ichi; Kurata, Morito; Nagata, Kaoru; Miura, Osamu

2014-01-01

A 22-year-old male was admitted for a sustained fever of 2 months, lymphadenopathy, and liver dysfunction. Anti-VCA-IgM antibody was positive, with elevated Epstein-Barr virus (EBV)-DNA load in the peripheral blood. Liver biopsy revealed infiltration of CD8-positive and EBV-positive cells. Most peripheral blood mononuclear cells (PBMCs) were also positive for CD8, and showed detectable levels of EBV-DNA. Monoclonal proliferation of EBV-infected cells was detected in the PBMCs by Southern blotting for EBV-terminal repeat (EBV-TR). Although EBV-positive T-cell lymphoproliferative disease (EBV-T-LPD) was suspected, the symptoms spontaneously resolved within 12 months. Anti-VCA-IgM antibody and the clonal band of EBV-TR were negative 1 year after the onset, while anti-EBNA antibody was positive. The final diagnosis was thus confirmed as infectious mononucleosis (IM). Our results indicate that EBV-infected CD8-positive cells and clonal proliferation of EBV-infected cells may be temporally detected in IM. EBV-T-LPDs should be carefully excluded in such cases.

Behaviour of AISI-SAE 8615 steel in ferritic nitrocarburizing using urea-metanol

International Nuclear Information System (INIS)

Barrena, M.I.; Castro, A.

1998-01-01

The present work shows the behaviour of low carbon alloyed steels as AISI-SAE 8615, after a ferritic nitrocarburizing process. Nitrocarburizing takes place at 570 degree centigree for 3 h in an atmosphere by combustion of methanol and urea under different flow rates. Metallographic studies were performed by optical microscopy. Harness profiles were measured and carbon percentages were also analyzed by emission spectrometry in order to determine the extension of the nitrocarburizing process. Optimal flow conditions have been found. The influence of the flow rate on the nitrocarburizing layer thickness has been also studied. (Author) 12 refs

Susceptibility of different leukocyte cell types to Vaccinia virus infection

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Sánchez-Puig Juana M

2004-11-01

Full Text Available Abstract Background Vaccinia virus, the prototype member of the family Poxviridae, was used extensively in the past as the Smallpox vaccine, and is currently considered as a candidate vector for new recombinant vaccines. Vaccinia virus has a wide host range, and is known to infect cultures of a variety of cell lines of mammalian origin. However, little is known about the virus tropism in human leukocyte populations. We report here that various cell types within leukocyte populations have widely different susceptibility to infection with vaccinia virus. Results We have investigated the ability of vaccinia virus to infect human PBLs by using virus recombinants expressing green fluorescent protein (GFP, and monoclonal antibodies specific for PBL subpopulations. Flow cytometry allowed the identification of infected cells within the PBL mixture 1–5 hours after infection. Antibody labeling revealed that different cell populations had very different infection rates. Monocytes showed the highest percentage of infected cells, followed by B lymphocytes and NK cells. In contrast to those cell types, the rate of infection of T lymphocytes was low. Comparison of vaccinia virus strains WR and MVA showed that both strains infected efficiently the monocyte population, although producing different expression levels. Our results suggest that MVA was less efficient than WR in infecting NK cells and B lymphocytes. Overall, both WR and MVA consistently showed a strong preference for the infection of non-T cells. Conclusions When infecting fresh human PBL preparations, vaccinia virus showed a strong bias towards the infection of monocytes, followed by B lymphocytes and NK cells. In contrast, very poor infection of T lymphocytes was detected. These finding may have important implications both in our understanding of poxvirus pathogenesis and in the development of improved smallpox vaccines.

Dendritic cells during Epstein Barr virus infection

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Christian eMunz

2014-06-01

Full Text Available Epstein Barr virus (EBV causes persistent infection in more than 90% of the human adult population and is associated with 2% of all tumors in humans. This -herpesvirus infects primarily human B and epithelial cells, but has been reported to be sensed by dendritic cells (DCs during primary infection. These activated DCs are thought to contribute to innate restriction of EBV infection and initiate EBV specific adaptive immune responses via cross-priming. The respective evidence and their potential importance for EBV specific vaccine development will be discussed in this review.

Novel Aerodynamic Design for Formula SAE Vehicles

Science.gov (United States)

Sentongo, Samuel; Carter, Austin; Cecil, Christopher; Feier, Ioan

2017-11-01

This paper identifies and evaluates the design characteristics of a novel airfoil that harnesses the Magnus Effect, applying a moving-surface boundary-layer control (MSBC) method to a Formula SAE Vehicle. The MSBC minimizes adverse pressure gradient and delays boundary layer separation through the use of a conveyor belt that interacts with the airfoil boundary layer. The MSBC allows dynamic control of the aerodynamic coefficients by variation of the belt speed, minimizing drag in high speed straights and maximizing downforce during vehicle cornering. A conveyer belt wing measuring approximately 0.9 x 0.9m in planform was designed and built to test the mechanical setup for such a MSBC wing. This study follows the relationship between inputted power and outputted surface velocity, with the goal being to maximize speed output vs. power input. The greatest hindrance to maximizing speed output is friction among belts, rollers, and stationary members. The maximum belt speed achieved during testing was 5.9 m/s with a power input of 48.8 W, which corresponds to 45.8 N of downforce based on 2D CFD results. Ongoing progress on this project is presented. United States Air Force Academy.

Immune regulation in Chandipura virus infection: characterization of CD4+ T regulatory cells from infected mice

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Shahir Prajakta

2011-05-01

Full Text Available Abstract Back ground Chandipura virus produces acute infection in mice. During infection drastic reduction of CD4+, CD8+ and CD19 + cell was noticed. Depletion of lymphocytes also noticed in spleen. The reduction may be due to the regulatory mechanism of immune system to prevent the bystander host tissue injury. There are several mechanisms like generation of regulatory cells, activation induced cell death (ACID etc were indicated to control the activation and maintain cellular homeostasis. Role of regulatory cells in homeostasis has been described in several viral diseases. This study was undertaken to characterize CD4+T regulatory cells from the infected mice. Method In this study we purified the CD4+ T cells from Chandipura virus infected susceptible Balb/c mice. CD4+ T regulatory cells were identified by expression of cell surface markers CD25, CD127 and CTLA-4 and intracellular markers Foxp3, IL-10 and TGF-beta. Antigen specificity and ability to suppress the proliferation of other lymphocytes were studied in vitro by purified CD4+CD25+T regulatory cells from infected mice. The proliferation was calculated by proliferation module of Flow Jo software. Expression of death receptors on regulatory cells were studied by flowcytometer. Results The CD4+ T cells isolated from infected mice expressed characteristic markers of regulatory phenotype at all post infective hours tested. The CD4+ T regulatory cells were proliferated when stimulated with Chandipura virus antigen. The regulatory cells did not suppress the proliferation of splenocytes stimulated with anti CD3 antibody when co cultured with them. Interesting observation was, while purification of CD4+ T cells by negative selection, the population of cells negative for CD4 also co purified along with CD4+ T cell. Flow cytometry analysis and light microscopy revealed that CD4 negative cells were of different size and shape (atypical compared to the normal lymphocytes. Greater percentage of

MAIT cells are activated in acute Dengue virus infection and after in vitro Zika virus infection.

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Dominic Paquin-Proulx

2018-01-01

Full Text Available Dengue virus (DENV and Zika virus (ZIKV are members of the Flaviviridae and are predominantly transmitted via mosquito bites. Both viruses are responsible for a growing number of infections in tropical and subtropical regions. DENV infection can cause lethargy with severe morbidity and dengue shock syndrome leading to death in some cases. ZIKV is now linked with Guillain-Barré syndrome and fetal malformations including microcephaly and developmental disorders (congenital Zika syndrome. The protective and pathogenic roles played by the immune response in these infections is unknown. Mucosal-associated invariant T (MAIT cells are a population of innate T cells with potent anti-bacterial activity. MAIT cells have also been postulated to play a role in the immune response to viral infections. In this study, we evaluated MAIT cell frequency, phenotype, and function in samples from subjects with acute and convalescent DENV infection. We found that in acute DENV infection, MAIT cells had elevated co-expression of the activation markers CD38 and HLA-DR and had a poor IFNγ response following bacterial stimulation. Furthermore, we found that MAIT cells can produce IFNγ in response to in vitro infection with ZIKV. This MAIT cell response was independent of MR1, but dependent on IL-12 and IL-18. Our results suggest that MAIT cells may play an important role in the immune response to Flavivirus infections.

Transient Oral Human Cytomegalovirus Infections Indicate Inefficient Viral Spread from Very Few Initially Infected Cells.

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Mayer, Bryan T; Krantz, Elizabeth M; Swan, David; Ferrenberg, James; Simmons, Karen; Selke, Stacy; Huang, Meei-Li; Casper, Corey; Corey, Lawrence; Wald, Anna; Schiffer, Joshua T; Gantt, Soren

2017-06-15

Cytomegalovirus (CMV) is acquired by the oral route in children, and primary infection is associated with abundant mucosal replication, as well as the establishment of latency in myeloid cells that results in lifelong infection. The efficiency of primary CMV infection in humans following oral exposure, however, is unknown. We consistently detected self-limited, low-level oral CMV shedding events, which we termed transient CMV infections, in a prospective birth cohort of 30 highly exposed CMV-uninfected infants. We estimated the likelihood of transient oral CMV infections by comparing their observed frequency to that of established primary infections, characterized by persistent high-level shedding, viremia, and seroconversion. We developed mathematical models of viral dynamics upon initial oral CMV infection and validated them using clinical shedding data. Transient infections comprised 76 to 88% of oral CMV shedding events. For this high percentage of transient infections to occur, we identified two mathematical prerequisites: a very small number of initially infected oral cells (1 to 4) and low viral infectivity (<1.5 new cells infected/cell). These observations indicate that oral CMV infection in infants typically begins with a single virus that spreads inefficiently to neighboring cells. Thus, although the incidence of CMV infection is high during infancy, our data provide a mechanistic framework to explain why multiple CMV exposures are typically required before infection is successfully established. These findings imply that a sufficiently primed immune response could prevent CMV from establishing latent infection in humans and support the achievability of a prophylactic CMV vaccine. IMPORTANCE CMV infects the majority of the world's population and is a major cause of birth defects. Developing a vaccine to prevent CMV infection would be extremely valuable but would be facilitated by a better understanding of how natural human CMV infection is acquired. We

Role of Bruton's tyrosine kinase inhibitors in HIV-1-infected cells.

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Guendel, Irene; Iordanskiy, Sergey; Sampey, Gavin C; Van Duyne, Rachel; Calvert, Valerie; Petricoin, Emanuel; Saifuddin, Mohammed; Kehn-Hall, Kylene; Kashanchi, Fatah

2015-06-01

Many cellular cofactors have been documented to be critical for various stages of viral replication. Using high-throughput proteomic assays, we have previously identified Bruton's tyrosine kinase (BTK) as a host protein that was uniquely upregulated in the plasma membrane of human immunodeficiency virus (HIV-1)-infected T cells. Here, we have further characterized the BTK expression in HIV-1 infection and show that this cellular factor is specifically expressed in infected myeloid cells. Significant upregulation of the phosphorylated form of BTK was observed in infected cells. Using size exclusion chromatography, we found BTK to be virtually absent in the uninfected U937 cells; however, new BTK protein complexes were identified and distributed in both high molecular weight (∼600 kDa) and a small molecular weight complex (∼60-120 kDa) in the infected U1 cells. BTK levels were highest in cells either chronically expressing virus or induced/infected myeloid cells and that BTK translocated to the membrane following induction of the infected cells. BTK knockdown in HIV-1-infected cells using small interfering RNA (siRNA) resulted in selective death of infected, but not uninfected, cells. Using BTK-specific antibody and small-molecule inhibitors including LFM-A13 and a FDA-approved compound, ibrutinib (PCI-32765), we have found that HIV-1-infected cells are sensitive to apoptotic cell death and result in a decrease in virus production. Overall, our data suggests that HIV-1-infected cells are sensitive to treatments targeting BTK expressed in infected cells.

Retroviral infection of non-dividing cells: Old and new perspectives

International Nuclear Information System (INIS)

Yamashita, Masahiro; Emerman, Michael

2006-01-01

The dependence of retroviral replication on cell proliferation was described as early as 1958, although different classes of retroviruses are able to infect non-dividing cells with different efficiencies. For example, the human immunodeficiency virus (HIV) and other lentiviruses infect most non-dividing cells nearly as well as dividing cells, while the gammaretroviruses such as the murine leukemia virus (MLV) cannot infect non-dividing cells, and other retroviruses have intermediate phenotypes. One exception to the ability of HIV to infect non-dividing cells involves resting CD4+ T cells in vitro where there are multiple restrictions. However, recent data show that there is massive infection of non-activated CD4+ T cell during acute infection which suggests that the situation is different in vivo. Finally, much work trying to explain the difference between HIV and MLV in non-dividing cells has focused on describing the ability of HIV to enter the nucleus during interphase. However, we suggest that events in the viral lifecycle other than nuclear import may be more important in determining the ability of a given retrovirus to infect non-dividing cells

T Follicular Helper Cells and B Cell Dysfunction in Aging and HIV-1 Infection.

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Pallikkuth, Suresh; de Armas, Lesley; Rinaldi, Stefano; Pahwa, Savita

2017-01-01

T follicular helper (Tfh) cells are a subset of CD4 T cells that provide critical signals to antigen-primed B cells in germinal centers to undergo proliferation, isotype switching, and somatic hypermutation to generate long-lived plasma cells and memory B cells during an immune response. The quantity and quality of Tfh cells therefore must be tightly controlled to prevent immune dysfunction in the form of autoimmunity and, on the other hand, immune deficiency. Both Tfh and B cell perturbations appear during HIV infection resulting in impaired antibody responses to vaccines such as seasonal trivalent influenza vaccine, also seen in biologic aging. Although many of the HIV-associated defects improve with antiretroviral therapy (ART), excess immune activation and antigen-specific B and T cell responses including Tfh function are still impaired in virologically controlled HIV-infected persons on ART. Interestingly, HIV infected individuals experience increased risk of age-associated pathologies. This review will discuss Tfh and B cell dysfunction in HIV infection and highlight the impact of chronic HIV infection and aging on Tfh-B cell interactions.

Lysis of endogenously infected CD4+ T cell blasts by rIL-2 activated autologous natural killer cells from HIV-infected viremic individuals.

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Manuela Fogli

2008-07-01

Full Text Available Understanding the cellular mechanisms that ensure an appropriate innate immune response against viral pathogens is an important challenge of biomedical research. In vitro studies have shown that natural killer (NK cells purified from healthy donors can kill heterologous cell lines or autologous CD4+ T cell blasts exogenously infected with several strains of HIV-1. However, it is not known whether the deleterious effects of high HIV-1 viremia interferes with the NK cell-mediated cytolysis of autologous, endogenously HIV-1-infected CD4+ T cells. Here, we stimulate primary CD4+ T cells, purified ex vivo from HIV-1-infected viremic patients, with PHA and rIL2 (with or without rIL-7. This experimental procedure allows for the significant expansion and isolation of endogenously infected CD4+ T cell blasts detected by intracellular staining of p24 HIV-1 core antigen. We show that, subsequent to the selective down-modulation of MHC class-I (MHC-I molecules, HIV-1-infected p24(pos blasts become partially susceptible to lysis by rIL-2-activated NK cells, while uninfected p24(neg blasts are spared from killing. This NK cell-mediated killing occurs mainly through the NKG2D activation pathway. However, the degree of NK cell cytolytic activity against autologous, endogenously HIV-1-infected CD4+ T cell blasts that down-modulate HLA-A and -B alleles and against heterologous MHC-I(neg cell lines is particularly low. This phenomenon is associated with the defective surface expression and engagement of natural cytotoxicity receptors (NCRs and with the high frequency of the anergic CD56(neg/CD16(pos subsets of highly dysfunctional NK cells from HIV-1-infected viremic patients. Collectively, our data demonstrate that the chronic viral replication of HIV-1 in infected individuals results in several phenotypic and functional aberrancies that interfere with the NK cell-mediated killing of autologous p24(pos blasts derived from primary T cells.

Networked T cell death following macrophage infection by Mycobacterium tuberculosis.

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Stephen H-F Macdonald

Full Text Available BACKGROUND: Depletion of T cells following infection by Mycobacterium tuberculosis (Mtb impairs disease resolution, and interferes with clinical test performance that relies on cell-mediated immunity. A number of mechanisms contribute to this T cell suppression, such as activation-induced death and trafficking of T cells out of the peripheral circulation and into the diseased lungs. The extent to which Mtb infection of human macrophages affects T cell viability however, is not well characterised. METHODOLOGY/PRINCIPAL FINDINGS: We found that lymphopenia (<1.5 × 10(9 cells/l was prevalent among culture-positive tuberculosis patients, and lymphocyte counts significantly improved post-therapy. We previously reported that Mtb-infected human macrophages resulted in death of infected and uninfected bystander macrophages. In the current study, we sought to examine the influence of infected human alveolar macrophages on T cells. We infected primary human alveolar macrophages (the primary host cell for Mtb or PMA-differentiated THP-1 cells with Mtb H37Ra, then prepared cell-free supernatants. The supernatants of Mtb-infected macrophages caused dose-dependent, caspase-dependent, T cell apoptosis. This toxic effect of infected macrophage secreted factors did not require TNF-α or Fas. The supernatant cytotoxic signal(s were heat-labile and greater than 50 kDa in molecular size. Although ESAT-6 was toxic to T cells, other Mtb-secreted factors tested did not influence T cell viability; nor did macrophage-free Mtb bacilli or broth from Mtb cultures. Furthermore, supernatants from Mycobacterium bovis Bacille de Calmette et Guerin (BCG- infected macrophages also elicited T cell death suggesting that ESAT-6 itself, although cytotoxic, was not the principal mediator of T cell death in our system. CONCLUSIONS: Mtb-Infected macrophages secrete heat-labile factors that are toxic to T cells, and may contribute to the immunosuppression seen in tuberculosis as well as

Mycoplasma agalactiae Induces Cytopathic Effects in Infected Cells Cultured In Vitro.

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Shrilakshmi Hegde

Full Text Available Mycoplasma agalactiae is the etiological agent of the contagious agalactia syndrome in sheep and goats and causes significant economic losses worldwide. Yet the mechanism of pathogenesis is largely unknown. Even whole-genome sequence analysis of its pathogenic type strain did not lead to any conclusions regarding its virulence or pathogenicity factors. Although inflammation and tissue destruction at the local site of M. agalactiae infection are largely considered as effects of the host immune response, the direct effect of the agent on host cells is not completely understood. The aim of this study was to investigate the effect of M. agalactiae infection on the quality and viability of host cells in vitro. Changes in cell morphology including cell elongation, cytoplasm shrinkage and membrane blebbing were observed in infected HeLa cells. Chromatin condensation and increased caspase-3 cleavage in infected HeLa cells 48 h after infection suggests an apoptosis-like phenomenon in M. agalactiae-infected cells. In compliance with these results, decreased viability and cell lysis of M. agalactiae-infected HeLa cells was also observed. Measurement of the amount of LDH released after M. agalactiae infection revealed a time- and dose-dependent increase in HeLa cell lysis. A significant decrease in LDH released after gentamicin treatment of infected cells confirmed the major role of cytadherent M. agalactiae in inducing host cell lysis. This is the first study illustrating M. agalactiae's induction of cytopathic effects in infected HeLa cells. Further detailed investigation of infected host tissue for apoptotic markers might demonstrate the association between M. agalactiae-induced host cell lysis and the tissue destruction observed during M. agalactiae natural infection.

Quantitative comparison of HTLV-1 and HIV-1 cell-to-cell infection with new replication dependent vectors.

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Dmitriy Mazurov

2010-02-01

Full Text Available We have developed an efficient method to quantify cell-to-cell infection with single-cycle, replication dependent reporter vectors. This system was used to examine the mechanisms of infection with HTLV-1 and HIV-1 vectors in lymphocyte cell lines. Effector cells transfected with reporter vector, packaging vector, and Env expression plasmid produced virus-like particles that transduced reporter gene activity into cocultured target cells with zero background. Reporter gene expression was detected exclusively in target cells and required an Env-expression plasmid and a viral packaging vector, which provided essential structural and enzymatic proteins for virus replication. Cell-cell fusion did not contribute to infection, as reporter protein was rarely detected in syncytia. Coculture of transfected Jurkat T cells and target Raji/CD4 B cells enhanced HIV-1 infection two fold and HTLV-1 infection ten thousand fold in comparison with cell-free infection of Raji/CD4 cells. Agents that interfere with actin and tubulin polymerization strongly inhibited HTLV-1 and modestly decreased HIV-1 cell-to-cell infection, an indication that cytoskeletal remodeling was more important for HTLV-1 transmission. Time course studies showed that HTLV-1 transmission occurred very rapidly after cell mixing, whereas slower kinetics of HIV-1 coculture infection implies a different mechanism of infectious transmission. HTLV-1 Tax was demonstrated to play an important role in altering cell-cell interactions that enhance virus infection and replication. Interestingly, superantigen-induced synapses between Jurkat cells and Raji/CD4 cells did not enhance infection for either HTLV-1 or HIV-1. In general, the dependence on cell-to-cell infection was determined by the virus, the effector and target cell types, and by the nature of the cell-cell interaction.

Autoreactive T Cells and Chronic Fungal Infection Drive Esophageal Carcinogenesis

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Zhu, Feng; Willette-Brown, Jami; Song, Na-Young; Lomada, Dakshayani; Song, Yongmei; Xue, Liyan; Gray, Zane; Zhao, Zitong; Davis, Sean R.; Sun, Zhonghe; Zhang, Peilin; Wu, Xiaolin; Zhan, Qimin; Richie, Ellen R.; Hu, Yinling

2018-01-01

SUMMARY Humans with autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED), a T cell–driven autoimmune disease caused by impaired central tolerance, are susceptible to developing chronic fungal infection and esophageal squamous cell carcinoma (ESCC). However, the relationship between autoreactive T cells and chronic fungal infection in ESCC development remains unclear. We find that kinase-dead Ikkα knockin mice develop phenotypes reminiscent of APECED, including impaired central tolerance, autoreactive T cells, chronic fungal infection, and ESCCs expressing specific human ESCC markers. Using this model, we investigated the potential link between ESCC and fungal infection. Autoreactive CD4 T cells permit fungal infection and incite tissue injury and inflammation. Antifungal treatment or depletion of autoreactive CD4 T cells rescues, whereas oral fungal administration promotes, ESCC development. Inhibition of inflammation or EGFR activity decreases fungal burden. Importantly, fungal infection is highly associated with ESCCs in non-autoimmune human patients. Therefore, autoreactive T cells and chronic fungal infection, fostered by inflammation and epithelial injury, promote ESCC development. PMID:28407484

Regulation of NKT Cell Localization in Homeostasis and Infection

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Slauenwhite, Drew; Johnston, Brent

2015-01-01

Natural killer T (NKT) cells are a specialized subset of T lymphocytes that regulate immune responses in the context of autoimmunity, cancer, and microbial infection. Lipid antigens derived from bacteria, parasites, and fungi can be presented by CD1d molecules and recognized by the canonical T cell receptors on NKT cells. Alternatively, NKT cells can be activated through recognition of self-lipids and/or pro-inflammatory cytokines generated during infection. Unlike conventional T cells, only a small subset of NKT cells traffic through the lymph nodes under homeostatic conditions, with the largest NKT cell populations localizing to the liver, lungs, spleen, and bone marrow. This is thought to be mediated by differences in chemokine receptor expression profiles. However, the impact of infection on the tissue localization and function of NKT remains largely unstudied. This review focuses on the mechanisms mediating the establishment of peripheral NKT cell populations during homeostasis and how tissue localization of NKT cells is affected during infection. PMID:26074921

Regulation of NKT Cell Localization in Homeostasis and Infection.

Science.gov (United States)

Slauenwhite, Drew; Johnston, Brent

2015-01-01

Natural killer T (NKT) cells are a specialized subset of T lymphocytes that regulate immune responses in the context of autoimmunity, cancer, and microbial infection. Lipid antigens derived from bacteria, parasites, and fungi can be presented by CD1d molecules and recognized by the canonical T cell receptors on NKT cells. Alternatively, NKT cells can be activated through recognition of self-lipids and/or pro-inflammatory cytokines generated during infection. Unlike conventional T cells, only a small subset of NKT cells traffic through the lymph nodes under homeostatic conditions, with the largest NKT cell populations localizing to the liver, lungs, spleen, and bone marrow. This is thought to be mediated by differences in chemokine receptor expression profiles. However, the impact of infection on the tissue localization and function of NKT remains largely unstudied. This review focuses on the mechanisms mediating the establishment of peripheral NKT cell populations during homeostasis and how tissue localization of NKT cells is affected during infection.

Uma Metodologia para a avaliação dos gradientes de tenacidade à fratura ao longo da camada cementada do aço SAE 5115

OpenAIRE

Sandor,Leonardo Taborda; Ferreira,Itamar

2006-01-01

Este trabalho propõe um modelo para avaliar pontualmente as variações de tenacidade à fratura ao longo da camada cementada de um aço SAE 5115. A pequena espessura dessas camadas impede a retirada de corpos de prova nas dimensões especificadas pelas normas de ensaios de tenacidade à fratura. Assim, para simular uma camada cementada retirou-se corpos-de-prova de tração e de tenacidade à fratura de amostras de aços SAE 5115, 5140, 5160 e 52100 assumindo a influência local apenas da variação do t...

Case depth in SAE 1020 steel using barkhausen noise

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Alessandra Drehmer

2013-01-01

Full Text Available The most widely used thermochemical process for surface hardening of steels is case hardening. Using several different heat treatments, martensitic surface layers were formed on SAE 1020 steel into which carbon had been diffused. Case depths were measured by traditional destructive techniques. Barkhausen noise measurements were made and both the RMS Barkhausen pulse envelope and the fast Fourier transform (FFT were obtained from numerical calculation. The FFT amplitudes, functions of frequency, were associated with distance from the sample surface using the skin depth equation δ = 1/ (πfσµ½ , where f is the frequency of the electromagnetic wave, s is the electrical conductivity, and µ is the magnetic permeability. We define a normalized power index (NPI which can be used to estimate case depths. The NPI is discussed in relation to the sample microstructure and it is shown that the case depth is most easily determined when the magnetic properties of the surface layer and core are substantially different.

Differential pulmonic NK and NKT cell responses in Schistosoma japonicum-infected mice.

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Cha, Hefei; Qin, Wenjuan; Yang, Quan; Xie, Hongyan; Qu, Jiale; Wang, Mei; Chen, Daixiong; Wang, Fang; Dong, Nuo; Chen, Longhua; Huang, Jun

2017-02-01

Natural killer cells (NK cells) and natural killer T cells (NKT cells) play a role in anti-infection, anti-tumor, transplantation immunity, and autoimmune regulation. However, the role of NK and NKT cells during Schistosoma japonicum (S. japonicum) infection has not been widely reported, especially regarding lung infections. The aim of this study was to research the NK and NKT cell response to S. japonicum infection in the lungs of mice. Using immunofluorescent histological analysis, NK and NKT cells were found near pulmonary granulomas. Moreover, flow cytometry revealed that the percentage and number of pulmonic NK cells in S. japonicum-infected mice were significantly increased (P cell number of NKT cells were decreased compared to those of normal mice (P NKT cells was increased after infection (P NKT cells (P cells (P NKT cells significantly increased (P NKT cells (P NKT cell activation during S. japonicum infection.

NK cell-like behavior of Valpha14i NK T cells during MCMV infection.

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Johnna D Wesley

2008-07-01

Full Text Available Immunity to the murine cytomegalovirus (MCMV is critically dependent on the innate response for initial containment of viral replication, resolution of active infection, and proper induction of the adaptive phase of the anti-viral response. In contrast to NK cells, the Valpha14 invariant natural killer T cell response to MCMV has not been examined. We found that Valpha14i NK T cells become activated and produce significant levels of IFN-gamma, but do not proliferate or produce IL-4 following MCMV infection. In vivo treatment with an anti-CD1d mAb and adoptive transfer of Valpha14i NK T cells into MCMV-infected CD1d(-/- mice demonstrate that CD1d is dispensable for Valpha14i NK T cell activation. In contrast, both IFN-alpha/beta and IL-12 are required for optimal activation. Valpha14i NK T cell-derived IFN-gamma is partially dependent on IFN-alpha/beta but highly dependent on IL-12. Valpha14i NK T cells contribute to the immune response to MCMV and amplify NK cell-derived IFN-gamma. Importantly, mortality is increased in CD1d(-/- mice in response to high dose MCMV infection when compared to heterozygote littermate controls. Collectively, these findings illustrate the plasticity of Valpha14i NK T cells that act as effector T cells during bacterial infection, but have NK cell-like behavior during the innate immune response to MCMV infection.

Effects of interferon on cultured cells persistently infected with viruses

Energy Technology Data Exchange (ETDEWEB)

Crespi, M

1986-01-01

The role of interferon (IFN) in viral persistence at the cellular level was investigated. Two types of persistent infections were chosen. The first type was cell lines which contained hepatitis B virus (HBV) DNA (PLC/PRF/5 and Hep 3B cells) uninfected control hepatoma cells, (Mahlavu, HA22T and Hep G2 cells) or simian virus 40 (SV40) DNA (C2, C6, C11 cells) and control uninfected (CV-1 cells). In the second type of infection Vero cells persistently infected with SSPE or Sendai virus were used. The aim of this work was to determine what effect IFN had in these infections in terms of its antiviral and antiproliferative effects; which of the two major IFN-induced pathways, E enzyme or protein kinase were induced; whether there were any differences in sensitivity to IFN between the DNA and RNA virus persistent infections. The anti-viral effect of IFN was examined by its ability to inhibit Sindbis virus replication using a radioimmunoassay system. The antiproliferative effect of IFN was determined by cell counting and /sup 3/H-thymidine incorporation. The activation of the ribonuclease F, determined by the inhibition of /sup 3/H-leucine incorporation after introduction of 2-5 actin into the cells, was variable, being activated in all cell lines with the exception of the PLC/PRF/5, Hep 3B and Hep G2 cells. Major differences between the two DNA persistent infections and the two RNA persistent infections were found. No correlation was found between the presence of HBV or SV40 persistent infections and the sensitivity of the cell lines to IFN. Both the SSPE and Sendai virus persistent infections were resistant to the antiviral and antiproliferative effect of IFN.

Defective Natural Killer cell antiviral capacity in paediatric HBV infection

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Heiberg, Ida Louise; Laura J., Pallett; Winther, Thilde Nordmann

2015-01-01

Natural Killer (NK) cells exhibit dysregulated effector function in adult chronic HBV infection (CHB), which may contribute to virus persistence. The role of NK cells in children infected perinatally with HBV is less studied. Access to a unique cohort enabled the cross-sectional evaluation of NK...... cell frequency, phenotype and function in HBV-infected children relative to uninfected children. We observed a selective defect in NK cell IFN-γ production, with conserved cytolytic function, mirroring the functional dichotomy observed in adult infection. Reduced expression of NKp30 on NK cells...

Invariant NKT cells: regulation and function during viral infection.

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Jennifer A Juno

Full Text Available Natural killer T cells (NKT cells represent a subset of T lymphocytes that express natural killer (NK cell surface markers. A subset of NKT cells, termed invariant NKT cells (iNKT, express a highly restricted T cell receptor (TCR and respond to CD1d-restricted lipid ligands. iNKT cells are now appreciated to play an important role in linking innate and adaptive immune responses and have been implicated in infectious disease, allergy, asthma, autoimmunity, and tumor surveillance. Advances in iNKT identification and purification have allowed for the detailed study of iNKT activity in both humans and mice during a variety of chronic and acute infections. Comparison of iNKT function between non-pathogenic simian immunodeficiency virus (SIV infection models and chronic HIV-infected patients implies a role for iNKT activity in controlling immune activation. In vitro studies of influenza infection have revealed novel effector functions of iNKT cells including IL-22 production and modulation of myeloid-derived suppressor cells, but ex vivo characterization of human iNKT cells during influenza infection are lacking. Similarly, as recent evidence suggests iNKT involvement in dengue virus pathogenesis, iNKT cells may modulate responses to a number of emerging pathogens. This Review will summarize current knowledge of iNKT involvement in responses to viral infections in both human and mouse models and will identify critical gaps in knowledge and opportunities for future study. We will also highlight recent efforts to harness iNKT ligands as vaccine adjuvants capable of improving vaccination-induced cellular immune responses.

Human induced pluripotent stem cell-derived models to investigate human cytomegalovirus infection in neural cells.

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Leonardo D'Aiuto

Full Text Available Human cytomegalovirus (HCMV infection is one of the leading prenatal causes of congenital mental retardation and deformities world-wide. Access to cultured human neuronal lineages, necessary to understand the species specific pathogenic effects of HCMV, has been limited by difficulties in sustaining primary human neuronal cultures. Human induced pluripotent stem (iPS cells now provide an opportunity for such research. We derived iPS cells from human adult fibroblasts and induced neural lineages to investigate their susceptibility to infection with HCMV strain Ad169. Analysis of iPS cells, iPS-derived neural stem cells (NSCs, neural progenitor cells (NPCs and neurons suggests that (i iPS cells are not permissive to HCMV infection, i.e., they do not permit a full viral replication cycle; (ii Neural stem cells have impaired differentiation when infected by HCMV; (iii NPCs are fully permissive for HCMV infection; altered expression of genes related to neural metabolism or neuronal differentiation is also observed; (iv most iPS-derived neurons are not permissive to HCMV infection; and (v infected neurons have impaired calcium influx in response to glutamate.

MuSAE: A European Project for the Diffusion of Energy and Environmental Planning in Small-Medium Sized Municipalities

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Giorgio Baldinelli

2015-12-01

Full Text Available The basic idea of the EU LIFE+ 2011 project MuSAE (“Municipalities Subsidiarity for Actions on Energy”, code LIFE11 ENV/IT/000016 consists of transferring the skills and experience related to energy planning, acquired by the leading beneficiary, the Municipality of Perugia, to three small- or medium-sized Umbrian Municipalities (Marsciano, Umbertide and Lisciano Niccone. This transfer is aimed, among other objectives, at the drafting of the Municipal Energy and Environmental Plan (MEEP and the opening of an energy information office in each partner Municipality, in cooperation with CIRIAF and Umbria Region. The present paper provides a summary of MuSAE activities, analyzing the procedures and modalities of implementation of the various phases of the MEEPs, on the basis of the experience gained over the years through the collaboration with the Municipality of Perugia and adapted to smaller territories such as those represented by the other partner Municipalities. A summary of the dissemination activities and pilot projects is also presented, testifying the first concrete results of the planning activity developed by each administration within the project.

Dynamics of human T-cell lymphotropic virus I (HTLV-I) infection of CD4+ T-cells.

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Katri, Patricia; Ruan, Shigui

2004-11-01

Stilianakis and Seydel (Bull. Math. Biol., 1999) proposed an ODE model that describes the T-cell dynamics of human T-cell lymphotropic virus I (HTLV-I) infection and the development of adult T-cell leukemia (ATL). Their model consists of four components: uninfected healthy CD4+ T-cells, latently infected CD4+ T-cells, actively infected CD4+ T-cells, and ATL cells. Mathematical analysis that completely determines the global dynamics of this model has been done by Wang et al. (Math. Biosci., 2002). In this note, we first modify the parameters of the model to distinguish between contact and infectivity rates. Then we introduce a discrete time delay to the model to describe the time between emission of contagious particles by active CD4+ T-cells and infection of pure cells. Using the results in Culshaw and Ruan (Math. Biosci., 2000) in the analysis of time delay with respect to cell-free viral spread of HIV, we study the effect of time delay on the stability of the endemically infected equilibrium. Numerical simulations are presented to illustrate the results.

Detection of bacteriophage-infected cells of Lactococcus lactis using flow cytometry

DEFF Research Database (Denmark)

Michelsen, Ole; Cuesta-Dominguez, Álvaro; Albrektsen, Bjarne

2007-01-01

Bacteriophage infection in dairy fermentation constitutes a serious problem worldwide. We have studied bacteriophage infection in Lactococcus lactis by using the flow cytometer. The first effect of the infection of the bacterium is a change from cells in chains toward single cells. We interpret...... describe a new method for detection of phage infection in Lactococcus lactis dairy cultures. The method is based on flow cytometric detection of cells with low-density cell walls. The method allows fast and early detection of phage-infected bacteria, independently of which phage has infected the culture...

Role of Bruton’s Tyrosine Kinase inhibitors in HIV-1 infected cells

Science.gov (United States)

Guendel, Irene; Iordanskiy, Sergey; Sampey, Gavin C; Van Duyne, Rachel; Calvert, Valerie; Petricoin, Emanuel; Saifuddin, Mohammed; Kehn-Hall, Kylene; Kashanchi, Fatah

2015-01-01

Many cellular cofactors have been documented to be critical for various stages of viral replication. Using high throughput proteomic assays, we have previously identified Bruton’s tyrosine kinase (BTK) as a host protein that was uniquely up-regulated in the plasma membrane of HIV-1 infected T-cells. Here, we have further characterized the BTK expression in HIV-1 infection and show that this cellular factor is specifically expressed in infected myeloid cells. Significant up-regulation of the phosphorylated form of BTK was observed in infected cells. Using size exclusion chromatography, we found BTK to be virtually absent in the uninfected U937 cells, however new BTK protein complexes were identified and distributed in both high molecular weight (~600 kDa) and a small molecular weight complex (~60–120 kDa) in the infected U1 cells. BTK levels were highest in cells either chronically expressing virus or induced/infected myeloid cells and that BTK translocated to the membrane following induction of the infected cells. BTK knockdown in HIV-1 infected cells using siRNA resulted in selective death of infected, but not uninfected, cells. Using BTK specific antibody and small molecule inhibitors including LFM-A13 and a FDA approved compound, Ibrutinib (PCI – 32765), we have found that HIV-1 infected cells are sensitive to apoptotic cell death and result in a decrease in virus production. Overall, our data suggests that HIV-1 infected cells are sensitive to treatments targeting BTK expressed in infected cells. PMID:25672887

The homeostasis of Plasmodium falciparum-infected red blood cells.

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Jakob M A Mauritz

2009-04-01

Full Text Available The asexual reproduction cycle of Plasmodium falciparum, the parasite responsible for severe malaria, occurs within red blood cells. A merozoite invades a red cell in the circulation, develops and multiplies, and after about 48 hours ruptures the host cell, releasing 15-32 merozoites ready to invade new red blood cells. During this cycle, the parasite increases the host cell permeability so much that when similar permeabilization was simulated on uninfected red cells, lysis occurred before approximately 48 h. So how could infected cells, with a growing parasite inside, prevent lysis before the parasite has completed its developmental cycle? A mathematical model of the homeostasis of infected red cells suggested that it is the wasteful consumption of host cell hemoglobin that prevents early lysis by the progressive reduction in the colloid-osmotic pressure within the host (the colloid-osmotic hypothesis. However, two critical model predictions, that infected cells would swell to near prelytic sphericity and that the hemoglobin concentration would become progressively reduced, remained controversial. In this paper, we are able for the first time to correlate model predictions with recent experimental data in the literature and explore the fine details of the homeostasis of infected red blood cells during five model-defined periods of parasite development. The conclusions suggest that infected red cells do reach proximity to lytic rupture regardless of their actual volume, thus requiring a progressive reduction in their hemoglobin concentration to prevent premature lysis.

T-cell-dependent control of acute Giardia lamblia infections in mice.

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Singer, S M; Nash, T E

2000-01-01

We have studied immune mechanisms responsible for control of acute Giardia lamblia and Giardia muris infections in adult mice. Association of chronic G. lamblia infection with hypogammaglobulinemia and experimental infections of mice with G. muris have led to the hypothesis that antibodies are required to control these infections. We directly tested this hypothesis by infecting B-cell-deficient mice with either G. lamblia or G. muris. Both wild-type mice and B-cell-deficient mice eliminated the vast majority of parasites between 1 and 2 weeks postinfection with G. lamblia. G. muris was also eliminated in both wild-type and B-cell-deficient mice. In contrast, T-cell-deficient and scid mice failed to control G. lamblia infections, as has been shown previously for G. muris. Treatment of wild-type or B-cell-deficient mice with antibodies to CD4 also prevented elimination of G. lamblia, confirming a role for T cells in controlling infections. By infecting mice deficient in either alphabeta- or gammadelta-T-cell receptor (TCR)-expressing T cells, we show that the alphabeta-TCR-expressing T cells are required to control parasites but that the gammadelta-TCR-expressing T cells are not. Finally, infections in mice deficient in production of gamma interferon or interleukin 4 (IL-4) and mice deficient in responding to IL-4 and IL-13 revealed that neither the Th1 nor the Th2 subset is absolutely required for protection from G. lamblia. We conclude that a T-cell-dependent mechanism is essential for controlling acute Giardia infections and that this mechanism is independent of antibody and B cells.

Natural Killer Cell Function and Dysfunction in Hepatitis C Virus Infection

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Kayla A. Holder

2014-01-01

Full Text Available Viruses must continually adapt against dynamic innate and adaptive responses of the host immune system to establish chronic infection. Only a small minority (~20% of those exposed to hepatitis C virus (HCV spontaneously clear infection, leaving approximately 200 million people worldwide chronically infected with HCV. A number of recent research studies suggest that establishment and maintenance of chronic HCV infection involve natural killer (NK cell dysfunction. This relationship is illustrated in vitro by disruption of typical NK cell responses including both cell-mediated cytotoxicity and cytokine production. Expression of a number of activating NK cell receptors in vivo is also affected in chronic HCV infection. Thus, direct in vivo and in vitro evidence of compromised NK function in chronic HCV infection in conjunction with significant epidemiological associations between the outcome of HCV infection and certain combinations of NK cell regulatory receptor and class I human histocompatibility linked antigen (HLA genotypes indicate that NK cells are important in the immune response against HCV infection. In this review, we highlight evidence suggesting that selective impairment of NK cell activity is related to establishment of chronic HCV infection.

Electron Microscopy of Ebola Virus-Infected Cells.

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Noda, Takeshi

2017-01-01

Ebola virus (EBOV) replicates in host cells, where both viral and cellular components show morphological changes during the process of viral replication from entry to budding. These steps in the replication cycle can be studied using electron microscopy (EM), including transmission electron microscopy (TEM) and scanning electron microscopy (SEM), which is one of the most useful methods for visualizing EBOV particles and EBOV-infected cells at the ultrastructural level. This chapter describes conventional methods for EM sample preparation of cultured cells infected with EBOV.

Human neuronal cell protein responses to Nipah virus infection

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Hassan Sharifah

2007-06-01

Full Text Available Abstract Background Nipah virus (NiV, a recently discovered zoonotic virus infects and replicates in several human cell types. Its replication in human neuronal cells, however, is less efficient in comparison to other fully susceptible cells. In the present study, the SK-N-MC human neuronal cell protein response to NiV infection is examined using proteomic approaches. Results Method for separation of the NiV-infected human neuronal cell proteins using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE was established. At least 800 protein spots were resolved of which seven were unique, six were significantly up-regulated and eight were significantly down-regulated. Six of these altered proteins were identified using mass spectrometry (MS and confirmed using MS/MS. The heterogenous nuclear ribonucleoprotein (hnRNP F, guanine nucleotide binding protein (G protein, voltage-dependent anion channel 2 (VDAC2 and cytochrome bc1 were present in abundance in the NiV-infected SK-N-MC cells in contrast to hnRNPs H and H2 that were significantly down-regulated. Conclusion Several human neuronal cell proteins that are differentially expressed following NiV infection are identified. The proteins are associated with various cellular functions and their abundance reflects their significance in the cytopathologic responses to the infection and the regulation of NiV replication. The potential importance of the ratio of hnRNP F, and hnRNPs H and H2 in regulation of NiV replication, the association of the mitochondrial protein with the cytopathologic responses to the infection and induction of apoptosis are highlighted.

Adrenaline-induced mobilization of T cells in HIV-infected patients

DEFF Research Database (Denmark)

Søndergaard, S R; Cozzi-Lepri, A; Ullum, H

2000-01-01

The present study aimed to investigate lymphocyte mobilization from peripheral cell reservoirs in HIV-infected patients. Nine HIV-infected patients on stable highly active anti-retroviral therapy (HAART), eight treatment-naive HIV-infected patients and eight HIV- controls received a 1-h adrenaline...... infusion. The adrenaline infusion induced a three-fold increase in the concentration of lymphocytes in all three groups. All HIV-infected patients mobilized significantly higher numbers of CD8+ cells but less CD4+ cells. All subjects mobilized CD45RA+CD62L+ and CD8+CD28+ cells to a lesser extent than CD45......RO+CD45RA- and CD8+CD28-cells. Furthermore, high numbers of CD8+CD38+ cells were mobilized only in the HIV-infected patients. It was therefore predominantly T cells with an activated phenotype which were mobilized after adrenaline stimulation. It is concluded that the HIV-associated immune defect...

Development of multi-element active aerodynamics for the formula sae car

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Merkel, James Patrick

This thesis focuses on the design, development, and implementation of an active aerodynamics system on 2013 Formula SAE car. The aerodynamics package itself consists of five element front and rear wings as well as an under body diffuser. Five element wings produce significant amounts of drag which is a compromise between the cornering ability of the car and the acceleration capability on straights. The active aerodynamics system allows for the wing angle of attack to dynamically change their configuration on track based on sensory data to optimize the wings for any given scenario. The wings are studied using computational fluid dynamics both in their maximum lift configuration as well as a minimum drag configuration. A control system is then developed using an electro mechanical actuation system to articulate the wings between these two states.

Infections and endothelial cells

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Keller, Tymen T.; Mairuhu, Albert T. A.; de Kruif, Martijn D.; Klein, Saskia K.; Gerdes, Victor E. A.; ten Cate, Hugo; Brandjes, Dees P. M.; Levi, Marcel; van Gorp, Eric C. M.

2003-01-01

Systemic infection by various pathogens interacts with the endothelium and may result in altered coagulation, vasculitis and atherosclerosis. Endothelium plays a role in the initiation and regulation of both coagulation and fibrinolysis. Exposure of endothelial cells may lead to rapid activation of

Identification of a Monocyte Receptor on Herpesvirus-Infected Endothelial Cells

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Etingin, Orli R.; Silverstein, Roy L.; Hajjar, David P.

1991-08-01

The adhesion of circulating blood cells to vascular endothelium may be an initial step in atherosclerosis, inflammation, and wound healing. One mechanism for promoting cell-cell adhesion involves the expression of adhesion molecules on the surface of the target cell. Herpes simplex virus infection of endothelium induces arterial injury and has been implicated in the development of human atherosclerosis. We now demonstrate that HSV-infected endothelial cells express the adhesion molecule GMP140 and that this requires cell surface expression of HSV glycoprotein C and local thrombin generation. Monocyte adhesion to HSV-infected endothelial cells was completely inhibited by anti-GMP140 antibodies but not by antibodies to other adhesion molecules such as VCAM and ELAM-1. The induction of GMP140 expression on HSV-infected endothelium may be an important pathophysiological mechanism in virus-induced cell injury and inflammation.

Lymphocytes and macrophages are infected by Theileria equi, but T cells and B cells are not required to establish infection in vivo.

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Joshua D Ramsay

Full Text Available Theileria equi has a biphasic life cycle in horses, with a period of intraleukocyte development followed by patent erythrocytic parasitemia that causes acute and sometimes fatal hemolytic disease. Unlike Theileria spp. that infect cattle (Theileria parva and Theileria annulata, the intraleukocyte stage (schizont of Theileria equi does not cause uncontrolled host cell proliferation or other significant pathology. Nevertheless, schizont-infected leukocytes are of interest because of their potential to alter host cell function and because immune responses directed against this stage could halt infection and prevent disease. Based on cellular morphology, Theileria equi has been reported to infect lymphocytes in vivo and in vitro, but the specific phenotype of schizont-infected cells has yet to be defined. To resolve this knowledge gap in Theileria equi pathogenesis, peripheral blood mononuclear cells were infected in vitro and the phenotype of infected cells determined using flow cytometry and immunofluorescence microscopy. These experiments demonstrated that the host cell range of Theileria equi was broader than initially reported and included B lymphocytes, T lymphocytes and monocyte/macrophages. To determine if B and T lymphocytes were required to establish infection in vivo, horses affected with severe combined immunodeficiency (SCID, which lack functional B and T lymphocytes, were inoculated with Theileria equi sporozoites. SCID horses developed patent erythrocytic parasitemia, indicating that B and T lymphocytes are not necessary to complete the Theileria equi life cycle in vivo. These findings suggest that the factors mediating Theileria equi leukocyte invasion and intracytoplasmic differentiation are common to several leukocyte subsets and are less restricted than for Theileria annulata and Theileria parva. These data will greatly facilitate future investigation into the relationships between Theileria equi leukocyte tropism and pathogenesis

Differences between Mycobacterium-Host Cell Relationships in Latent Tuberculous Infection of Mice Ex Vivo and Mycobacterial Infection of Mouse Cells In Vitro

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Elena Ufimtseva

2016-01-01

Full Text Available The search for factors that account for the reproduction and survival of mycobacteria, including vaccine strains, in host cells is the priority for studies on tuberculosis. A comparison of BCG-mycobacterial loads in granuloma cells obtained from bone marrow and spleens of mice with latent tuberculous infection and cells from mouse bone marrow and peritoneal macrophage cultures infected with the BCG vaccine in vitro has demonstrated that granuloma macrophages each normally contained a single BCG-Mycobacterium, while those acutely infected in vitro had increased mycobacterial loads and death rates. Mouse granuloma cells were observed to produce the IFNγ, IL-1α, GM-CSF, CD1d, CD25, CD31, СD35, and S100 proteins. None of these activation markers were found in mouse cell cultures infected in vitro or in intact macrophages. Lack of colocalization of lipoarabinomannan-labeled BCG-mycobacteria with the lysosomotropic LysoTracker dye in activated granuloma macrophages suggests that these macrophages were unable to destroy BCG-mycobacteria. However, activated mouse granuloma macrophages could control mycobacterial reproduction in cells both in vivo and in ex vivo culture. By contrast, a considerable increase in the number of BCG-mycobacteria was observed in mouse bone marrow and peritoneal macrophages after BCG infection in vitro, when no expression of the activation-related molecules was detected in these cells.

Infection and Proliferation of Giant Viruses in Amoeba Cells.

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Takemura, Masaharu

2016-01-01

Acanthamoeba polyphaga mimivirus, the first discovered giant virus with genome size and particle size much larger than previously discovered viruses, possesses several genes for translation and CRISPER Cas system-like defense mechanism against virophages, which co-infect amoeba cells with the giant virus and which inhibit giant virus proliferation. Mimiviruses infect amoeba cells by phagocytosis and release their DNA into amoeba cytoplasm through their stargate structure. After infection, giant virion factories (VFs) form in amoeba cytoplasm, followed by DNA replication and particle formation at peripheral regions of VF. Marseilleviruses, the smallest giant viruses, infect amoeba cells by phagocytosis or endocytosis, form larger VF than Mimivirus's VF in amoeba cytoplasm, and replicate their particles. Pandoraviruses found in 2013 have the largest genome size and particle size among all viruses ever found. Pandoraviruses infect amoeba cells by phagocytosis and release their DNA into amoeba cytoplasm through their mouth-like apical pores. The proliferation of Pandoraviruses occurs along with nucleus disruption. New virions form at the periphery of the region formerly occupied by the amoeba cell nucleus.

Potential Cellular Signatures of Viral Infections in Human Hematopoietic Cells

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J. Mikovits

2001-01-01

Full Text Available Expression profiling of cellular genes was performed using a 10,000 cDNA human gene array in order to identify expression changes following chronic infection of human hematopoietic cells with Kapsosi’s Sarcoma -associated Virus (KSHV also known as Human Herpesvirus 8 (HHV8 and Human T cell leukemia virus-1 (HTLV-1. We performed cell-free {\\it in vitro} infection of primary bone marrow derived CD34+ cells using semi-purified HHV8 and a mature IL-2 dependent T cell line, KIT 225, using highly concentrated viral stocks prepared from an infectious molecular clone of HTLV-1. Thirty days post infection, mRNA was isolated from infected cultures and uninfected controls and submitted for microarray analysis. More than 400 genes were differentially expressed more than two-fold following HHV8 infection of primary bone marrow derived CD34+ cells. Of these 400, interferon regulatory factor 4 (IRF4, cyclin B2, TBP-associated factor, eukaryotic elongation factor and pim 2 were up-regulated more than 3.5 fold. In contrast, less than 100 genes were differentially expressed more than two-fold following chronic infection of a mature T cell line with HTLV-1. Of these, only cdc7 was up-regulated more than 3.5 fold. These data may provide insight into cellular signatures of infection useful for diagnosis of infection as well as potential targets for therapeutic intervention.

Sepsis Associated Encephalopathy

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Neera Chaudhry

2014-01-01

Full Text Available Sepsis associated encephalopathy (SAE is a common but poorly understood neurological complication of sepsis. It is characterized by diffuse brain dysfunction secondary to infection elsewhere in the body without overt CNS infection. The pathophysiology of SAE is complex and multifactorial including a number of intertwined mechanisms such as vascular damage, endothelial activation, breakdown of the blood brain barrier, altered brain signaling, brain inflammation, and apoptosis. Clinical presentation of SAE may range from mild symptoms such as malaise and concentration deficits to deep coma. The evaluation of cognitive dysfunction is made difficult by the absence of any specific investigations or biomarkers and the common use of sedation in critically ill patients. SAE thus remains diagnosis of exclusion which can only be made after ruling out other causes of altered mentation in a febrile, critically ill patient by appropriate investigations. In spite of high mortality rate, management of SAE is limited to treatment of the underlying infection and symptomatic treatment for delirium and seizures. It is important to be aware of this condition because SAE may present in early stages of sepsis, even before the diagnostic criteria for sepsis can be met. This review discusses the diagnostic approach to patients with SAE along with its epidemiology, pathophysiology, clinical presentation, and differential diagnosis.

Alterations in the nuclear proteome of HIV-1 infected T-cells

International Nuclear Information System (INIS)

DeBoer, Jason; Jagadish, Teena; Haverland, Nicole A.; Madson, Christian J.; Ciborowski, Pawel; Belshan, Michael

2014-01-01

Virus infection of a cell involves the appropriation of host factors and the innate defensive response of the cell. The identification of proteins critical for virus replication may lead to the development of novel, cell-based inhibitors. In this study we mapped the changes in T-cell nuclei during human immunodeficiency virus type 1 (HIV-1) at 20 hpi. Using a stringent data threshold, a total of 13 and 38 unique proteins were identified in infected and uninfected cells, respectively, across all biological replicates. An additional 15 proteins were found to be differentially regulated between infected and control nuclei. STRING analysis identified four clusters of protein–protein interactions in the data set related to nuclear architecture, RNA regulation, cell division, and cell homeostasis. Immunoblot analysis confirmed the differential expression of several proteins in both C8166-45 and Jurkat E6-1 T-cells. These data provide a map of the response in host cell nuclei upon HIV-1 infection. - Highlights: • We identify changes in the expression of nuclear proteins during HIV-1 infection. • 163 nuclear proteins were found differentially regulated during HIV-1 infection. • Bioinformatic analysis identified several nuclear pathways altered by HIV infection. • Candidate factors were validated in two independent cell lines

Alterations in the nuclear proteome of HIV-1 infected T-cells

Energy Technology Data Exchange (ETDEWEB)

DeBoer, Jason [Department of Medical Microbiology and Immunology, Creighton University, 2500 California Plaza, Omaha, NE 68178 (United States); Jagadish, Teena; Haverland, Nicole A. [Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE 68198 (United States); Madson, Christian J. [Department of Medical Microbiology and Immunology, Creighton University, 2500 California Plaza, Omaha, NE 68178 (United States); Ciborowski, Pawel [Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE 68198 (United States); The Nebraska Center for Virology, University of Nebraska, Lincoln 68583 (United States); Belshan, Michael, E-mail: michaelbelshan@creighton.edu [Department of Medical Microbiology and Immunology, Creighton University, 2500 California Plaza, Omaha, NE 68178 (United States); The Nebraska Center for Virology, University of Nebraska, Lincoln 68583 (United States)

2014-11-15

Virus infection of a cell involves the appropriation of host factors and the innate defensive response of the cell. The identification of proteins critical for virus replication may lead to the development of novel, cell-based inhibitors. In this study we mapped the changes in T-cell nuclei during human immunodeficiency virus type 1 (HIV-1) at 20 hpi. Using a stringent data threshold, a total of 13 and 38 unique proteins were identified in infected and uninfected cells, respectively, across all biological replicates. An additional 15 proteins were found to be differentially regulated between infected and control nuclei. STRING analysis identified four clusters of protein–protein interactions in the data set related to nuclear architecture, RNA regulation, cell division, and cell homeostasis. Immunoblot analysis confirmed the differential expression of several proteins in both C8166-45 and Jurkat E6-1 T-cells. These data provide a map of the response in host cell nuclei upon HIV-1 infection. - Highlights: • We identify changes in the expression of nuclear proteins during HIV-1 infection. • 163 nuclear proteins were found differentially regulated during HIV-1 infection. • Bioinformatic analysis identified several nuclear pathways altered by HIV infection. • Candidate factors were validated in two independent cell lines.

6K2-induced vesicles can move cell to cell during turnip mosaic virus infection

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Romain eGrangeon

2013-12-01

Full Text Available To successfully infect plants, viruses replicate in an initially infected cell and then move to neighboring cells through plasmodesmata (PDs. However, the nature of the viral entity that crosses over the cell barrier into non-infected ones is not clear. The membrane-associated 6K2 protein of turnip mosaic virus (TuMV induces the formation of vesicles involved in the replication and intracellular movement of viral RNA. This study shows that 6K2-induced vesicles trafficked towards the plasma membrane and were associated with plasmodesmata (PD. We demonstrated also that 6K2 moved cell-to-cell into adjoining cells when plants were infected with TuMV. 6K2 was then fused to photo-activable GFP (6K2:PAGFP to visualize how 6K2 move intercellularly during TuMV infection. After activation, 6K2:PAGFP-tagged vesicles moved to the cell periphery and across the cell wall into adjacent cells. These vesicles were shown to contain the viral RNA-dependent RNA polymerase and viral RNA. Symplasmic movement of TuMV may thus be achieved in the form of a membrane-associated viral RNA complex induced by 6K2.

Susceptibility of Primary Human Choroid Plexus Epithelial Cells and Meningeal Cells to Infection by JC Virus.

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O'Hara, Bethany A; Gee, Gretchen V; Atwood, Walter J; Haley, Sheila A

2018-04-15

JC polyomavirus (JCPyV) establishes a lifelong persistence in roughly half the human population worldwide. The cells and tissues that harbor persistent virus in vivo are not known, but renal tubules and other urogenital epithelial cells are likely candidates as virus is shed in the urine of healthy individuals. In an immunosuppressed host, JCPyV can become reactivated and cause progressive multifocal leukoencephalopathy (PML), a fatal demyelinating disease of the central nervous system. Recent observations indicate that JCPyV may productively interact with cells in the choroid plexus and leptomeninges. To further study JCPyV infection in these cells, primary human choroid plexus epithelial cells and meningeal cells were challenged with virus, and their susceptibility to infection was compared to the human glial cell line, SVG-A. We found that JCPyV productively infects both choroid plexus epithelial cells and meningeal cells in vitro Competition with the soluble receptor fragment LSTc reduced virus infection in these cells. Treatment of cells with neuraminidase also inhibited both viral infection and binding. Treatment with the serotonin receptor antagonist, ritanserin, reduced infection in SVG-A and meningeal cells. We also compared the ability of wild-type and sialic acid-binding mutant pseudoviruses to transduce these cells. Wild-type pseudovirus readily transduced all three cell types, but pseudoviruses harboring mutations in the sialic acid-binding pocket of the virus failed to transduce the cells. These data establish a novel role for choroid plexus and meninges in harboring virus that likely contributes not only to meningoencephalopathies but also to PML. IMPORTANCE JCPyV infects greater than half the human population worldwide and causes central nervous system disease in patients with weakened immune systems. Several recent reports have found JCPyV in the choroid plexus and leptomeninges of patients with encephalitis. Due to their role in forming the blood

Microsporidia infection impacts the host cell's cycle and reduces host cell apoptosis

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Higes, Mariano; Sagastume, Soledad; Juarranz, Ángeles; Dias-Almeida, Joyce; Budge, Giles E.; Meana, Aránzazu; Boonham, Neil

2017-01-01

Intracellular parasites can alter the cellular machinery of host cells to create a safe haven for their survival. In this regard, microsporidia are obligate intracellular fungal parasites with extremely reduced genomes and hence, they are strongly dependent on their host for energy and resources. To date, there are few studies into host cell manipulation by microsporidia, most of which have focused on morphological aspects. The microsporidia Nosema apis and Nosema ceranae are worldwide parasites of honey bees, infecting their ventricular epithelial cells. In this work, quantitative gene expression and histology were studied to investigate how these two parasites manipulate their host’s cells at the molecular level. Both these microsporidia provoke infection-induced regulation of genes involved in apoptosis and the cell cycle. The up-regulation of buffy (which encodes a pro-survival protein) and BIRC5 (belonging to the Inhibitor Apoptosis protein family) was observed after infection, shedding light on the pathways that these pathogens use to inhibit host cell apoptosis. Curiously, different routes related to cell cycle were modified after infection by each microsporidia. In the case of N. apis, cyclin B1, dacapo and E2F2 were up-regulated, whereas only cyclin E was up-regulated by N. ceranae, in both cases promoting the G1/S phase transition. This is the first report describing molecular pathways related to parasite-host interactions that are probably intended to ensure the parasite’s survival within the cell. PMID:28152065

Ureaplasma parvum infection alters filamin a dynamics in host cells

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Brown Mary B

2011-04-01

Full Text Available Abstract Background Ureaplasmas are among the most common bacteria isolated from the human urogenital tract. Ureaplasmas can produce asymptomatic infections or disease characterized by an exaggerated inflammatory response. Most investigations have focused on elucidating the pathogenic potential of Ureaplasma species, but little attention has been paid to understanding the mechanisms by which these organisms are capable of establishing asymptomatic infection. Methods We employed differential proteome profiling of bladder tissues from rats experimentally infected with U. parvum in order to identify host cell processes perturbed by colonization with the microbe. Tissues were grouped into four categories: sham inoculated controls, animals that spontaneously cleared infection, asymptomatic urinary tract infection (UTI, and complicated UTI. One protein that was perturbed by infection (filamin A was used to further elucidate the mechanism of U. parvum-induced disruption in human benign prostate cells (BPH-1. BPH-1 cells were evaluated by confocal microscopy, immunoblotting and ELISA. Results Bladder tissue from animals actively colonized with U. parvum displayed significant alterations in actin binding proteins (profilin 1, vinculin, α actinin, and filamin A that regulate both actin polymerization and cell cytoskeletal function pertaining to focal adhesion formation and signal transduction (Fisher's exact test, P U. parvum perturbed the regulation of filamin A. Specifically, infected BPH-1 cells exhibited a significant increase in filamin A phosphorylated at serine2152 (P ≤ 0.01, which correlated with impaired proteolysis of the protein and its normal intracellular distribution. Conclusion Filamin A dynamics were perturbed in both models of infection. Phosphorylation of filamin A occurs in response to various cell signaling cascades that regulate cell motility, differentiation, apoptosis and inflammation. Thus, this phenomenon may be a useful

Effect of particle size on the friction welding of Al2O3 reinforced 6160 Al alloy composite and SAE 1020 steel

International Nuclear Information System (INIS)

Hascalik, Ahmet; Orhan, Nuri

2007-01-01

The aim of this study is to investigate the feasibility of joining Al 2 O 3 reinforced Al alloy composite to SAE 1020 steel by rotational friction welding. The aluminum-based metal matrix composite (MMC) material containing 5, 10 and 15 vol% Al 2 O 3 particles with average particle sizes of 30 and 60 μm was produced by powder metallurgy technique. The integrity of the joints has been investigated by optical and scanning electron microscopy, while the mechanical properties assessment included microhardness and shear tests. Results indicated that Al/Al 2 O 3 composite could be joined to SAE 1020 steel by friction welding. However, it was pointed out that the quality of the joint was effected negatively with the increase in particle size and volume percentage of the oxide particles in the MMC

IL-15 STIMULATED NATURAL KILLER CELLS CLEAR HIV-1 INFECTED CELLS FOLLOWING LATENCY REVERSAL EX VIVO.

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Garrido, Carolina; Abad-Fernandez, Maria; Tuyishime, Marina; Pollara, Justin J; Ferrari, Guido; Soriano-Sarabia, Natalia; Margolis, David M

2018-03-28

Current efforts towards HIV eradication include approaches to augment immune recognition and elimination of persistently infected cells following latency reversal. Natural killer (NK) cells, the main effectors of the innate immune system, recognize and clear targets using different mechanisms than CD8 + T cells, offering an alternative or complementary approach for HIV clearance strategies. We assessed the impact of IL-15 treatment on NK cell function and the potential of stimulated NK cells to clear the HIV reservoir. We measured NK cell receptor expression, antibody-dependent cell-dependent cytotoxicity (ADCC), cytotoxicity, IFN-γ production and antiviral activity in autologous HIV replication systems. All NK cell functions were uniformly improved by IL-15, and more importantly, IL-15-treated NK cells were able to clear latently HIV infected cells after exposure to vorinostat, a clinically relevant latency reversing agent. We also demonstrate that NK cells from HIV infected individuals aviremic on antiretroviral therapy can be efficiently stimulated with IL-15. Our work opens a promising line of investigation towards future immunotherapies to clear persistent HIV infection using NK cells. IMPORTANCE In the search for an HIV cure, strategies to enhance immune function to allow recognition and clearance of HIV infected cells following latency reversal are being evaluated. Natural killer (NK) cells possess characteristics that can be exploited for immunotherapy against persistent HIV infection. We demonstrate that NK cells from HIV-positive donors can be strongly stimulated with IL-15, improving their antiviral and cytotoxic potential, and more importantly, clearing HIV infected cells after latency reversal with a clinically relevant drug. Our results encourage further investigation to design NK cell-based immunotherapies to achieve HIV eradication. Copyright © 2018 American Society for Microbiology.

Benchmark tests for a Formula SAE Student car prototyping

Science.gov (United States)

Mariasiu, Florin

2011-12-01

Aerodynamic characteristics of a vehicle are important elements in its design and construction. A low drag coefficient brings significant fuel savings and increased engine power efficiency. In designing and developing vehicles trough computer simulation process to determine the vehicles aerodynamic characteristics are using dedicated CFD (Computer Fluid Dynamics) software packages. However, the results obtained by this faster and cheaper method, are validated by experiments in wind tunnels tests, which are expensive and were complex testing equipment are used in relatively high costs. Therefore, the emergence and development of new low-cost testing methods to validate CFD simulation results would bring great economic benefits for auto vehicles prototyping process. This paper presents the initial development process of a Formula SAE Student race-car prototype using CFD simulation and also present a measurement system based on low-cost sensors through which CFD simulation results were experimentally validated. CFD software package used for simulation was Solid Works with the FloXpress add-on and experimental measurement system was built using four piezoresistive force sensors FlexiForce type.

Impact of persistent cytomegalovirus infection on human neuroblastoma cell gene expression

International Nuclear Information System (INIS)

Hoever, Gerold; Vogel, Jens-Uwe; Lukashenko, Polina; Hofmann, Wolf-Karsten; Komor, Martina; Doerr, Hans Wilhelm; Cinatl, Jindrich

2005-01-01

In a model of human neuroblastoma (NB) cell lines persistently infected with human cytomegalovirus (HCMV) we previously showed that persistent HCMV infection is associated with an increased malignant phenotype, enhanced drug resistance, and invasive properties. To gain insights into the mechanisms of increased malignancy we analyzed the global changes in cellular gene expression induced by persistent HCMV infection of human neuroblastoma cells by use of high-density oligonucleotide microarrays (HG-U133A, Affymetrix) and RT-PCR. Comparing the gene expression of different NB cell lines with persistently infected cell sub-lines revealed 11 host cell genes regulated in a similar manner throughout all infected samples. Nine of these 11 genes may contribute to the previously observed changes in malignant phenotype of persistently HCMV infected NB cells by influencing invasive growth, apoptosis, angiogenesis, and proliferation. Thus, this work provides the basis for further functional studies

Cell death by pyroptosis drives CD4 T-cell depletion in HIV-1 infection

Science.gov (United States)

Doitsh, Gilad; Galloway, Nicole L. K.; Geng, Xin; Yang, Zhiyuan; Monroe, Kathryn M.; Zepeda, Orlando; Hunt, Peter W.; Hatano, Hiroyu; Sowinski, Stefanie; Muñoz-Arias, Isa; Greene, Warner C.

2014-01-01

The pathway causing CD4 T-cell death in HIV-infected hosts remains poorly understood although apoptosis has been proposed as a key mechanism. We now show that caspase-3-mediated apoptosis accounts for the death of only a small fraction of CD4 T cells corresponding to those that are both activated and productively infected. The remaining over 95% of quiescent lymphoid CD4 T cells die by caspase-1-mediated pyroptosis triggered by abortive viral infection. Pyroptosis corresponds to an intensely inflammatory form of programmed cell death in which cytoplasmic contents and pro-inflammatory cytokines, including IL-1β, are released. This death pathway thus links the two signature events in HIV infection--CD4 T-cell depletion and chronic inflammation--and creates a pathogenic vicious cycle in which dying CD4 T cells release inflammatory signals that attract more cells to die. This cycle can be broken by caspase 1 inhibitors shown to be safe in humans, raising the possibility of a new class of `anti-AIDS' therapeutics targeting the host rather than the virus.

Comportamiento a fatiga del acero sae 4140 usando alta rugosidad superficial y ambiente corrosivo

OpenAIRE

CEBALLOS, WILSON FERNANDO; GÓMEZ, ADOLFO LEÓN; CORONADO, JOHN JAIRO

2010-01-01

En este artículo se presenta el estudio del comportamiento a fatiga del acero SAE 4140 en condiciones diferentes: superficie pulida (acabado espejo), ambiente corrosivo (jugo de caña de azúcar), alta rugosidad superficial (superficie equivalente a la encontrada típicamente en los ejes de molino de caña) y una condición que involucra la combinación de las dos últimas. Las anteriores condiciones están presentes en los ejes de maza de molino de caña de azúcar durante el proceso de molienda. Se r...

COMPORTAMIENTO A FATIGA DEL ACERO SAE 4140 USANDO ALTA RUGOSIDAD SUPERFICIAL Y AMBIENTE CORROSIVO

OpenAIRE

CEBALLOS, WILSON FERNANDO; GÓMEZ, ADOLFO LEÓN; CORONADO, JOHN JAIRO

2010-01-01

En este artículo se presenta el estudio del comportamiento a fatiga del acero SAE 4140 en condiciones diferentes: superficie pulida (acabado espejo), ambiente corrosivo (jugo de caña de azúcar), alta rugosidad superficial (superficie equivalente a la encontrada típicamente en los ejes de molino de caña) y una condición que involucra la combinación de las dos últimas. Las anteriores condiciones están presentes en los ejes de maza de molino de caña de azúcar durante el proceso de molienda. Se r...

Role of Bunyamwera Orthobunyavirus NSs protein in infection of mosquito cells.

Science.gov (United States)

Szemiel, Agnieszka M; Failloux, Anna-Bella; Elliott, Richard M

2012-01-01

Bunyamwera orthobunyavirus is both the prototype and study model of the Bunyaviridae family. The viral NSs protein seems to contribute to the different outcomes of infection in mammalian and mosquito cell lines. However, only limited information is available on the growth of Bunyamwera virus in cultured mosquito cells other than the Aedes albopictus C6/36 line. To determine potential functions of the NSs protein in mosquito cells, replication of wild-type virus and a recombinant NSs deletion mutant was compared in Ae. albopictus C6/36, C7-10 and U4.4 cells, and in Ae. aegypti Ae cells by monitoring N protein production and virus yields at various times post infection. Both viruses established persistent infections, with the exception of NSs deletion mutant in U4.4 cells. The NSs protein was nonessential for growth in C6/36 and C7-10 cells, but was important for productive replication in U4.4 and Ae cells. Fluorescence microscopy studies using recombinant viruses expressing green fluorescent protein allowed observation of three stages of infection, early, acute and late, during which infected cells underwent morphological changes. In the absence of NSs, these changes were less pronounced. An RNAi response efficiently reduced virus replication in U4.4 cells transfected with virus specific dsRNA, but not in C6/36 or C7/10 cells. Lastly, Ae. aegypti mosquitoes were exposed to blood-meal containing either wild-type or NSs deletion virus, and at various times post-feeding, infection and disseminated infection rates were measured. Compared to wild-type virus, infection rates by the mutant virus were lower and more variable. If the NSs deletion virus was able to establish infection, it was detected in salivary glands at 6 days post-infection, 3 days later than wild-type virus. Bunyamwera virus NSs is required for efficient replication in certain mosquito cell lines and in Ae. aegypti mosquitoes.

Apoptosis transcriptional mechanism of feline infectious peritonitis virus infected cells.

Science.gov (United States)

Shuid, Ahmad Naqib; Safi, Nikoo; Haghani, Amin; Mehrbod, Parvaneh; Haron, Mohd Syamsul Reza; Tan, Sheau Wei; Omar, Abdul Rahman

2015-11-01

Apoptosis has been postulated to play an important role during feline infectious peritonitis virus (FIPV) infection; however, its mechanism is not well characterized. This study is focused on apoptosis and transcriptional profiling of FIPV-infected cells following in vitro infection of CRFK cells with FIPV 79-1146 WSU. Flow cytometry was used to determine mode of cell death in first 42 h post infection (hpi). FIPV infected cells underwent early apoptosis at 9 hpi (p apoptosis at 12 hpi (p apoptosis cluster (80 down-regulated and 51 up-regulated) along with increase of apoptosis, p53, p38 MAPK, VEGF and chemokines/cytokines signaling pathways were probably involved in apoptosis process. Six of the de-regulated genes expression (RASSF1, BATF2, MAGEB16, PDCD5, TNFα and TRAF2) and TNFα protein concentration were analyzed by RT-qPCR and ELISA, respectively, at different time-points. Up-regulations of both pro-apoptotic (i.e. PDCD5) and anti-apoptotic (i.e. TRAF2) were detected from first hpi and continuing to deregulate during apoptosis process in the infected cells.

Cells in Dengue Virus Infection In Vivo

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Sansanee Noisakran

2010-01-01

Full Text Available Dengue has been recognized as one of the most important vector-borne emerging infectious diseases globally. Though dengue normally causes a self-limiting infection, some patients may develop a life-threatening illness, dengue hemorrhagic fever (DHF/dengue shock syndrome (DSS. The reason why DHF/DSS occurs in certain individuals is unclear. Studies in the endemic regions suggest that the preexisting antibodies are a risk factor for DHF/DSS. Viremia and thrombocytopenia are the key clinical features of dengue virus infection in patients. The amounts of virus circulating in patients are highly correlated with severe dengue disease, DHF/DSS. Also, the disturbance, mainly a transient depression, of hematological cells is a critical clinical finding in acute dengue patients. However, the cells responsible for the dengue viremia are unresolved in spite of the intensive efforts been made. Dengue virus appears to replicate and proliferate in many adapted cell lines, but these in vitro properties are extremely difficult to be reproduced in primary cells or in vivo. This paper summarizes reports on the permissive cells in vitro and in vivo and suggests a hematological cell lineage for dengue virus infection in vivo, with the hope that a new focus will shed light on further understanding of the complexities of dengue disease.

Apoptosis in HEp-2 cells infected with Ureaplasma diversum.

Science.gov (United States)

Amorim, Aline Teixeira; Marques, Lucas Miranda; Santos, Angelita Maria Oliveira Gusmão; Martins, Hellen Braga; Barbosa, Maysa Santos; Rezende, Izadora Souza; Andrade, Ewerton Ferraz; Campos, Guilherme Barreto; Lobão, Tássia Neves; Cortez, Beatriz Araujo; Monezi, Telma Alvez; Machado-Santelli, Glaucia Maria; Timenetsky, Jorge

2014-09-04

Bacterial pathogens have many strategies for infecting and persisting in host cells. Adhesion, invasion and intracellular life are important features in the biology of mollicutes. The intracellular location of Ureaplasma diversum may trigger disturbances in the host cell. This includes activation or inhibition of pro and anti-apoptotic factors, which facilitate the development of host damage. The aim of the present study was to associate U. diversum infection in HEp-2 cells and apoptosis induction. Cells were infected for 72hs with four U. diversum clinical isolates and an ATCC strain. The U. diversum invasion was analyzed by Confocal Laser Scanning Microscopy and gentamicin invasion assay. The apoptosis was evaluated using pro-apoptotic and anti-apoptotic gene expression, and FITC Annexin V/Dead Cell Apoptosis Kit. The number of internalized ureaplasma in HEp-2 cells increased significantly throughout the infection. The flow cytometry analysis with fluorochromes to detect membrane depolarization and gene expression for caspase 2, 3 and 9 increased in infected cells after 24 hours. However, after 72 hours a considerable decrease of apoptotic cells was observed. The data suggests that apoptosis may be initially induced by some isolates in association with HEp-2 cells, but over time, there was no evidence of apoptosis in the presence of ureaplasma and HEp-2 cells. The initial increase and then decrease in apoptosis could be related to bacterial pathogen-associated molecular pattern (PAMPS). Moreover, the isolates of U. diversum presented differences in the studied parameters for apoptosis. It was also observed that the amount of microorganisms was not proportional to the induction of apoptosis in HEp-2 cells.

TIM-1 Promotes Hepatitis C Virus Cell Attachment and Infection.

Science.gov (United States)

Wang, Jing; Qiao, Luhua; Hou, Zhouhua; Luo, Guangxiang

2017-01-15

Human TIM and TAM family proteins were recently found to serve as phosphatidylserine (PS) receptors which promote infections by many different viruses, including dengue virus, West Nile virus, Ebola virus, Marburg virus, and Zika virus. In the present study, we provide substantial evidence demonstrating that TIM-1 is important for efficient infection by hepatitis C virus (HCV). The knockdown of TIM-1 expression significantly reduced HCV infection but not HCV RNA replication. Likewise, TIM-1 knockout in Huh-7.5 cells remarkably lowered HCV cell attachment and subsequent HCV infection. More significantly, the impairment of HCV infection in the TIM-1 knockout cells could be restored completely by ectopic expression of TIM-1 but not TIM-3 or TIM-4. Additionally, HCV infection and cell attachment were inhibited by PS but not by phosphatidylcholine (PC), demonstrating that TIM-1-mediated enhancement of HCV infection is PS dependent. The exposure of PS on the HCV envelope was confirmed by immunoprecipitation of HCV particles with a PS-specific monoclonal antibody. Collectively, these findings demonstrate that TIM-1 promotes HCV infection by serving as an attachment receptor for binding to PS exposed on the HCV envelope. TIM family proteins were recently found to enhance infections by many different viruses, including several members of the Flaviviridae family. However, their importance in HCV infection has not previously been examined experimentally. The TIM family proteins include three members in humans: TIM-1, TIM-3, and TIM-4. The findings derived from our studies demonstrate that TIM-1, but not TIM-3 or TIM-4, promotes HCV infection by functioning as an HCV attachment factor. Knockout of the TIM-1 gene resulted in a remarkable reduction of HCV cell attachment and infection. PS-containing liposomes blocked HCV cell attachment and subsequent HCV infection. HCV particles could also be precipitated with a PS-specific monoclonal antibody. These findings suggest that TIM-1

In vivo infection of IgG-containing cells by Jembrana disease virus during acute infection

International Nuclear Information System (INIS)

Desport, Moira; Tenaya, I.W. Masa; McLachlan, Alexander; McNab, Tegan J.; Rachmat, Judhi; Hartaningsih, Nining; Wilcox, Graham E.

2009-01-01

Jembrana disease virus (JDV) is an unusual bovine lentivirus which causes a non-follicular proliferation of lymphocytes, a transient immunosuppression and a delayed humoral response in infected Bali cattle in Indonesia. A double-immunofluorescent labeling method was developed to identify the subset of mononuclear cells in which the viral capsid protein could be detected. Viral antigen was present in pleomorphic centroblast-like cells which were identified as IgG-containing cells, including plasma cells, in lymphoid tissues. There was no evidence of infection of CD3 + T-cells or MAC387 + monocytes in tissues but large vacuolated cells with a macrophage-like morphology in the lung were found to contain viral antigen although they could not be shown conclusively to be infected. The tropism of JDV for mature IgG-containing cells may be relevant to understanding the pathogenesis of Jembrana disease, the delayed antibody responses and the genetic composition of this atypical lentivirus.

Dendritic Cells Activate and Mature after Infection with Mycobacterium tuberculosis

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Mamo Gezahagne

2011-07-01

Full Text Available Abstract Background Dendritic cells (DCs can take up an array of different antigens, including microorganisms which they can process and present more effectively than any other antigen presenting cell. However, whether the interaction between the human DC and Mycobacterium tuberculosis represents a defense mechanism by the invaded host, or helping the invader to evade the defense mechanism of the host is still not clearly understood. Findings To analyze the interactions between M. tuberculosis and immune cells, human peripheral blood monocyte-derived immature DCs were infected with M. tuberculosis H37Rv wild type strain and flow cytometry was used to analyse cell surface expression markers. The ability of the M. tuberculosis infected DC to induce T cell proliferation using 5 and 6-carboxyfluorescein diacetate succinimidyl ester (CFSE dilution technique was also investigated. DCs were found to internalize the mycobacteria and show dose dependent infection and necrosis with different multiplicity of infection. Flow cytometry analysis of cell surface expression markers CD40, CD54, CD80, CD83, CD86 and HLA DR in infected DC revealed significant (p M. tuberculosis in comparison to immature DC with no stimulation. Lipopolysaccharide (LPS from Salmonella abortus equi, a known DC maturation agent, was used as a positive control and showed a comparable up regulation of cell surface markers as observed with M. tuberculosis infected DC. It was revealed that the M. tuberculosis infected DC induced T cell proliferation. Conclusion These data clearly demonstrate that M. tuberculosis induces activation and maturation of human monocyte-derived immature DC as well as induces T cell proliferation in vitro.

Unconventional Pro-inflammatory CD4+ T Cell Response in B Cell-Deficient Mice Infected with Trypanosoma cruzi

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Melisa Gorosito Serrán

2017-11-01

Full Text Available Chagas disease, caused by the parasite Trypanosoma cruzi, is endemic in Latin America but has become a global public health concern by migration of infected people. It has been reported that parasite persistence as well as the intensity of the inflammatory immune response are determinants of the clinical manifestations of the disease. Even though inflammation is indispensable for host defense, when deregulated, it can contribute to tissue injury and organ dysfunction. Here, we report the importance of B cells in conditioning T cell response in T. cruzi infection. Mice deficient in mature B cells (muMT mice infected with T. cruzi exhibited an increase in plasma TNF concentration, TNF-producing CD4+ T cells, and mortality. The increase in TNF-producing CD4+ T cells was accompanied by a reduction in IFNγ+CD4+ T cells and a decrease of the frequency of regulatory Foxp3+, IL-10+, and IL17+CD4+ T cells populations. The CD4+ T cell population activated by T. cruzi infection, in absence of mature B cells, had a high frequency of Ly6C+ cells and showed a lower expression of inhibitory molecules such as CTLA-4, PD-1, and LAG3. CD4+ T cells from infected muMT mice presented a high frequency of CD62LhiCD44− cells, which is commonly associated with a naïve phenotype. Through transfer experiments we demonstrated that CD4+ T cells from infected muMT mice were able to condition the CD4+ T cells response from infected wild-type mice. Interestingly, using Blimp-flox/flox-CD23icre mice we observed that in absence of plasmablast/plasma cell T. cruzi-infected mice exhibited a higher number of TNF-producing CD4+ T cells. Our results showed that the absence of B cells during T. cruzi infection affected the T cell response at different levels and generated a favorable scenario for unconventional activation of CD4+ T cell leading to an uncontrolled effector response and inflammation. The product of B cell differentiation, the plasmablast/plasma cells, could be able

Targeted sorting of single virus-infected cells of the coccolithophore Emiliania huxleyi.

Science.gov (United States)

Martínez Martínez, Joaquín; Poulton, Nicole J; Stepanauskas, Ramunas; Sieracki, Michael E; Wilson, William H

2011-01-01

Discriminating infected from healthy cells is the first step to understanding the mechanisms and ecological implications of viral infection. We have developed a method for detecting, sorting, and performing molecular analysis of individual, infected cells of the important microalga Emiliania huxleyi, based on known physiological responses to viral infection. Of three fluorescent dyes tested, FM 1-43 (for detecting membrane blebbing) gave the most unequivocal and earliest separation of cells. Furthermore, we were able to amplify the genomes of single infected cells using Multiple Displacement Amplification. This novel method to reliably discriminate infected from healthy cells in cultures will allow researchers to answer numerous questions regarding the mechanisms and implications of viral infection of E. huxleyi. The method may be transferable to other virus-host systems.

Preferential infection and depletion of Mycobacterium tuberculosis-specific CD4 T cells after HIV-1 infection

NARCIS (Netherlands)

Geldmacher, Christof; Ngwenyama, Njabulo; Schuetz, Alexandra; Petrovas, Constantinos; Reither, Klaus; Heeregrave, Edwin J.; Casazza, Joseph P.; Ambrozak, David R.; Louder, Mark; Ampofo, William; Pollakis, Georgios; Hill, Brenna; Sanga, Erica; Saathoff, Elmar; Maboko, Leonard; Roederer, Mario; Paxton, William A.; Hoelscher, Michael; Koup, Richard A.

2010-01-01

HIV-1 infection results in the progressive loss of CD4 T cells. In this study, we address how different pathogen-specific CD4 T cells are affected by HIV infection and the cellular parameters involved. We found striking differences in the depletion rates between CD4 T cells to two common

Equine infectious anemia virus-infected dendritic cells retain antigen presentation capability

International Nuclear Information System (INIS)

Rivera, Julie A.; McGuire, Travis C.

2005-01-01

To determine if equine monocyte-derived dendritic cells (DC) were susceptible to equine infectious anemia virus (EIAV) infection, ex vivo-generated DC were infected with virus in vitro. EIAV antigen was detected by immunofluorescence 3 days post-infection with maximum antigen being detected on day 4, whereas there was no antigen detected in DC incubated with the same amount of heat-inactivated EIAV. No cytolytic activity was observed after EIAV WSU5 infection of DC. These monocyte-derived DC were more effective than macrophages and B cells in stimulating allogenic T lymphocytes. Both infected macrophages and DC stimulated similar levels of memory CTL responses in mixtures of CD8+ and CD4+ cells as detected with 51 Cr-release assays indicating that EIAV infection of DC did not alter antigen presentation. However, EIAV-infected DC were more effective than infected macrophages when used to stimulate memory CTL in isolated CD8+ cells. The maintenance of antigen processing and presenting function by EIAV-infected DC in vitro suggests that this function is maintained during in vivo infection

Telomerase Activity Impacts on Epstein-Barr Virus Infection of AGS Cells

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Rac, Jürgen; Haas, Florian; Schumacher, Andrina; Middeldorp, Jaap M.; Delecluse, Henri-Jacques; Speck, Roberto F.

2015-01-01

The Epstein-Barr virus (EBV) is transmitted from host-to-host via saliva and is associated with epithelial malignancies including nasopharyngeal carcinoma (NPC) and some forms of gastric carcinoma (GC). Nevertheless, EBV does not transform epithelial cells in vitro where it is rapidly lost from infected primary epithelial cells or epithelial tumor cells. Long-term infection by EBV, however, can be established in hTERT-immortalized nasopharyngeal epithelial cells. Here, we hypothesized that increased telomerase activity in epithelial cells enhances their susceptibility to infection by EBV. Using HONE-1, AGS and HEK293 cells we generated epithelial model cell lines with increased or suppressed telomerase activity by stable ectopic expression of hTERT or of a catalytically inactive, dominant negative hTERT mutant. Infection experiments with recombinant prototypic EBV (rB95.8), recombinant NPC EBV (rM81) with increased epithelial cell tropism compared to B95.8, or recombinant B95.8 EBV with BZLF1-knockout that is not able to undergo lytic replication, revealed that infection frequencies positively correlate with telomerase activity in AGS cells but also partly depend on the cellular background. AGS cells with increased telomerase activity showed increased expression mainly of latent EBV genes, suggesting that increased telomerase activity directly acts on the EBV infection of epithelial cells by facilitating latent EBV gene expression early upon virus inoculation. Thus, our results indicate that infection of epithelial cells by EBV is a very selective process involving, among others, telomerase activity and cellular background to allow for optimized host-to-host transmission via saliva. PMID:25856387

An Early Historical Examination of the Educational Intent of Supervised Agricultural Experiences (SAEs) and Project-Based Learning in Agricultural Education

Science.gov (United States)

Smith, Kasee L.; Rayfield, John

2016-01-01

Project-based learning has been a component of agricultural education since its inception. In light of the current call for additional emphasis of the Supervised Agricultural Experience (SAE) component of agricultural education, there is a need to revisit the roots of project-based learning. This early historical research study was conducted to…

Cell migration is another player of the minute virus of mice infection

Energy Technology Data Exchange (ETDEWEB)

Garcin, Pierre O.; Panté, Nelly, E-mail: pante@zoology.ubc.ca

2014-11-15

The parvovirus minute virus of mice, prototype strain (MVMp), preferentially infects and kills cancer cells. This intrinsic MVMp oncotropism may depend in part on the early stages of MVMp infection. To test this hypothesis, we investigated the early events of MVMp infection in mouse LA9 fibroblasts and a highly invasive mouse mammary tumor cell line derived from polyomavirus middle T antigen-mediated transformation. Using a combination of fluorescence and electron microscopy, we found that various parameters of the cell migration process affect MVMp infection. We show that, after binding to the plasma membrane, MVMp particles rapidly cluster at the leading edge of migrating cells, which exhibit higher levels of MVMp uptake than non-motile cells. Moreover, promoting cell migration on a fibronectin matrix increased MVMp infection, and induction of epithelial–mesenchymal transition allowed MVMp replication in non-permissive epithelial cells. Hence, we propose that cell migration influences the early stages of MVMp infection. - Highlights: • We document early steps of MVMp infection. • We report that a fibronectin matrix promotes MVMp infection. • We show that cellular migration plays a role in MVMp uptake. • We show that epithelial–mesenchymal transition allows MVMp replication.

Cell migration is another player of the minute virus of mice infection

International Nuclear Information System (INIS)

Garcin, Pierre O.; Panté, Nelly

2014-01-01

The parvovirus minute virus of mice, prototype strain (MVMp), preferentially infects and kills cancer cells. This intrinsic MVMp oncotropism may depend in part on the early stages of MVMp infection. To test this hypothesis, we investigated the early events of MVMp infection in mouse LA9 fibroblasts and a highly invasive mouse mammary tumor cell line derived from polyomavirus middle T antigen-mediated transformation. Using a combination of fluorescence and electron microscopy, we found that various parameters of the cell migration process affect MVMp infection. We show that, after binding to the plasma membrane, MVMp particles rapidly cluster at the leading edge of migrating cells, which exhibit higher levels of MVMp uptake than non-motile cells. Moreover, promoting cell migration on a fibronectin matrix increased MVMp infection, and induction of epithelial–mesenchymal transition allowed MVMp replication in non-permissive epithelial cells. Hence, we propose that cell migration influences the early stages of MVMp infection. - Highlights: • We document early steps of MVMp infection. • We report that a fibronectin matrix promotes MVMp infection. • We show that cellular migration plays a role in MVMp uptake. • We show that epithelial–mesenchymal transition allows MVMp replication

A novel single virus infection system reveals that influenza virus preferentially infects cells in g1 phase.

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Ryuta Ueda

Full Text Available BACKGROUND: Influenza virus attaches to sialic acid residues on the surface of host cells via the hemagglutinin (HA, a glycoprotein expressed on the viral envelope, and enters into the cytoplasm by receptor-mediated endocytosis. The viral genome is released and transported in to the nucleus, where transcription and replication take place. However, cellular factors affecting the influenza virus infection such as the cell cycle remain uncharacterized. METHODS/RESULTS: To resolve the influence of cell cycle on influenza virus infection, we performed a single-virus infection analysis using optical tweezers. Using this newly developed single-virus infection system, the fluorescence-labeled influenza virus was trapped on a microchip using a laser (1064 nm at 0.6 W, transported, and released onto individual H292 human lung epithelial cells. Interestingly, the influenza virus attached selectively to cells in the G1-phase. To clarify the molecular differences between cells in G1- and S/G2/M-phase, we performed several physical and chemical assays. Results indicated that: 1 the membranes of cells in G1-phase contained greater amounts of sialic acids (glycoproteins than the membranes of cells in S/G2/M-phase; 2 the membrane stiffness of cells in S/G2/M-phase is more rigid than those in G1-phase by measurement using optical tweezers; and 3 S/G2/M-phase cells contained higher content of Gb3, Gb4 and GlcCer than G1-phase cells by an assay for lipid composition. CONCLUSIONS: A novel single-virus infection system was developed to characterize the difference in influenza virus susceptibility between G1- and S/G2/M-phase cells. Differences in virus binding specificity were associated with alterations in the lipid composition, sialic acid content, and membrane stiffness. This single-virus infection system will be useful for studying the infection mechanisms of other viruses.

CD4 T Cell Responses in Latent and Chronic Viral Infections

Science.gov (United States)

Walton, Senta; Mandaric, Sanja; Oxenius, Annette

2013-01-01

The spectrum of tasks which is fulfilled by CD4 T cells in the setting of viral infections is large, ranging from support of CD8 T cells and humoral immunity to exertion of direct antiviral effector functions. While our knowledge about the differentiation pathways, plasticity, and memory of CD4 T cell responses upon acute infections or immunizations has significantly increased during the past years, much less is still known about CD4 T cell differentiation and their beneficial or pathological functions during persistent viral infections. In this review we summarize current knowledge about the differentiation, direct or indirect antiviral effector functions, and the regulation of virus-specific CD4 T cells in the setting of persistent latent or active chronic viral infections with a particular emphasis on herpes virus infections for the former and chronic lymphocytic choriomeningitis virus infection for the latter. PMID:23717308

Group 1 innate lymphoid cells in Toxoplasma gondii infection.

Science.gov (United States)

Dunay, I R; Diefenbach, A

2018-02-01

Innate lymphoid cells (ILCs) are a group of lymphocytes that carry out important functions in immunity to infections and in organ homeostasis at epithelial barrier surfaces. ILCs are innate immune cells that provide an early source of cytokines to initiate immune responses against pathogens. Cytotoxic ILCs (i.e. conventional (c)NK cells) and several subsets of helper-like ILCs are the major branches of the ILC family. Conventional NK cells and group 1 ILCs share several characteristics such as surface receptors and the ability to produce IFN-γ upon activation, but they differ in their developmental paths and in their dependence on specific transcription factors. Infection of mice with the intracellular parasite Toxoplasma gondii is followed by a strong Th1-mediated immune response. Previous studies indicate that NK1.1 + cells contribute to the production of IFN-γ and TNF and cytotoxicity during acute T. gondii infection. Upon oral infection, the parasite infects intestinal enterocytes, and within the lamina propria, innate immune responses lead to initial parasite control although the infection disseminates widely and persists long-term in immune privileged sites despite adaptive immunity. Upon parasite entry into the small intestine, during the acute stage, ILC1 produce high levels of IFN-γ and TNF protecting barrier surfaces, thus essentially contributing to early parasite control. We will discuss here the role of innate lymphocytes during T. gondii infection in the context of the only recently appreciated diversity of ILC subsets. © 2018 John Wiley & Sons Ltd.

Targeted sorting of single virus-infected cells of the coccolithophore Emiliania huxleyi.

Directory of Open Access Journals (Sweden)

Joaquín Martínez Martínez

Full Text Available Discriminating infected from healthy cells is the first step to understanding the mechanisms and ecological implications of viral infection. We have developed a method for detecting, sorting, and performing molecular analysis of individual, infected cells of the important microalga Emiliania huxleyi, based on known physiological responses to viral infection. Of three fluorescent dyes tested, FM 1-43 (for detecting membrane blebbing gave the most unequivocal and earliest separation of cells. Furthermore, we were able to amplify the genomes of single infected cells using Multiple Displacement Amplification. This novel method to reliably discriminate infected from healthy cells in cultures will allow researchers to answer numerous questions regarding the mechanisms and implications of viral infection of E. huxleyi. The method may be transferable to other virus-host systems.

EBI3 regulates the NK cell response to mouse cytomegalovirus infection

DEFF Research Database (Denmark)

Jensen, Helle; Chen, Shih-Yu; Folkersen, Lasse Westergaard

2017-01-01

Natural killer (NK) cells are key mediators in the control of cytomegalovirus infection. Here, we show that Epstein-Barr virus-induced 3 (EBI3) is expressed by human NK cells after NKG2D or IL-12 plus IL-18 stimulation and by mouse NK cells during mouse cytomegalovirus (MCMV) infection. The induc......Natural killer (NK) cells are key mediators in the control of cytomegalovirus infection. Here, we show that Epstein-Barr virus-induced 3 (EBI3) is expressed by human NK cells after NKG2D or IL-12 plus IL-18 stimulation and by mouse NK cells during mouse cytomegalovirus (MCMV) infection....... The induction of EBI3 protein expression in mouse NK cells is a late activation event. Thus, early activation events of NK cells, such as IFNγ production and CD69 expression, were not affected in EBI3-deficient (Ebi3-/-) C57BL/6 (B6) mice during MCMV infection. Furthermore, comparable levels of early viral...... replication in spleen and liver were observed in MCMV-infected Ebi3-/- and wild-type (WT) B6 mice. Interestingly, the viral load in salivary glands and oral lavage was strongly decreased in the MCMV-infected Ebi3-/- B6 mice, suggesting that EBI3 plays a role in the establishment of MCMV latency. We detected...

T-cell dynamics in healthy and HIV-infected individuals

NARCIS (Netherlands)

Vrisekoop, N.

2007-01-01

This thesis focuses on T-cell dynamics in healthy and both treated and untreated HIV-infected individuals. Although the progressive decline in CD4+ T-cell numbers is the hallmark of HIV infection, the mechanisms behind this depletion remain controversial. Currently the most prevailing ideas include

Enhanced infectivity of bluetongue virus in cell culture by centrifugation.

OpenAIRE

Sundin, D R; Mecham, J O

1989-01-01

The effects of centrifugation of the infection of cell culture with bluetongue virus (BTV) were investigated. Baby hamster kidney cells were infected with BTV with or without centrifugation. Viral antigen was detected by immunofluorescence at 24 h in both centrifuged and noncentrifuged cultures. However, after 24 h of infection, the production of PFU in centrifuged cell cultures was 10- to 20-fold greater than that seen in cultures not centrifuged. In addition, centrifugation enhanced the dir...

A Fluorescent Cell-Based System for Imaging Zika Virus Infection in Real-Time

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Michael J. McFadden

2018-02-01

Full Text Available Zika virus (ZIKV is a re-emerging flavivirus that is transmitted to humans through the bite of an infected mosquito or through sexual contact with an infected partner. ZIKV infection during pregnancy has been associated with numerous fetal abnormalities, including prenatal lethality and microcephaly. However, until recent outbreaks in the Americas, ZIKV has been relatively understudied, and therefore the biology and pathogenesis of ZIKV infection remain incompletely understood. Better methods to study ZIKV infection in live cells could enhance our understanding of the biology of ZIKV and the mechanisms by which ZIKV contributes to fetal abnormalities. To this end, we developed a fluorescent cell-based reporter system allowing for live imaging of ZIKV-infected cells. This system utilizes the protease activity of the ZIKV non-structural proteins 2B and 3 (NS2B-NS3 to specifically mark virus-infected cells. Here, we demonstrate the utility of this fluorescent reporter for identifying cells infected by ZIKV strains of two lineages. Further, we use this system to determine that apoptosis is induced in cells directly infected with ZIKV in a cell-autonomous manner. Ultimately, approaches that can directly track ZIKV-infected cells at the single cell-level have the potential to yield new insights into the host-pathogen interactions that regulate ZIKV infection and pathogenesis.

Identification of XMRV infection-associated microRNAs in four cell types in culture.

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Ketha V K Mohan

Full Text Available INTRODUCTION: XMRV is a gammaretrovirus that was thought to be associated with prostate cancer (PC and chronic fatigue syndrome (CFS in humans until recently. The virus is culturable in various cells of human origin like the lymphocytes, NK cells, neuronal cells, and prostate cell lines. MicroRNAs (miRNA, which regulate gene expression, were so far not identified in cells infected with XMRV in culture. METHODS: Two prostate cell lines (LNCaP and DU145 and two primary cells, Peripheral Blood Lymphocytes [PBL] and Monocyte-derived Macrophages [MDM] were infected with XMRV. Total mRNA was extracted from mock- and virus-infected cells at 6, 24 and 48 hours post infection and evaluated for microRNA profile in a microarray. RESULTS: MicroRNA expression profiles of XMRV-infected continuous prostate cancer cell lines differ from that of virus-infected primary cells (PBL and MDMs. miR-193a-3p and miRPlus-E1245 observed to be specific to XMRV infection in all 4 cell types. While miR-193a-3p levels were down regulated miRPlus-E1245 on the other hand exhibited varied expression profile between the 4 cell types. DISCUSSION: The present study clearly demonstrates that cellular microRNAs are expressed during XMRV infection of human cells and this is the first report demonstrating the regulation of miR193a-3p and miRPlus-E1245 during XMRV infection in four different human cell types.

Follicular Helper T Cells are Essential for the Elimination of Plasmodium Infection

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Damián Pérez-Mazliah

2017-10-01

Full Text Available CD4+ follicular helper T (Tfh cells have been shown to be critical for the activation of germinal center (GC B-cell responses. Similar to other infections, Plasmodium infection activates both GC as well as non-GC B cell responses. Here, we sought to explore whether Tfh cells and GC B cells are required to eliminate a Plasmodium infection. A CD4 T cell-targeted deletion of the gene that encodes Bcl6, the master transcription factor for the Tfh program, resulted in complete disruption of the Tfh response to Plasmodium chabaudi in C57BL/6 mice and consequent disruption of GC responses and IgG responses and the inability to eliminate the otherwise self-resolving chronic P. chabaudi infection. On the other hand, and contrary to previous observations in immunization and viral infection models, Signaling Lymphocyte Activation Molecule (SLAM-Associated Protein (SAP-deficient mice were able to activate Tfh cells, GC B cells, and IgG responses to the parasite. This study demonstrates the critical role for Tfh cells in controlling this systemic infection, and highlights differences in the signals required to activate GC B cell responses to this complex parasite compared with those of protein immunizations and viral infections. Therefore, these data are highly pertinent for designing malaria vaccines able to activate broadly protective B-cell responses.

Cell-mediated immunity in herpes simplex virus-infected mice: functional analysis of lymph node cells during periods of acute and latent infection, with reference to cytotoxic and memory cells.

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Nash, A A; Quartey-Papafio, R; Wildy, P

1980-08-01

The functional characteristics of lymphoid cells were investigated during acute and latent infection of mice with herpes simplex virus (HSV). Cytotoxic T cells were found in the draining lymph node (DLN) 4 days p.i. and had reached maximum activity between 6 and 9 days. After the 12th day and during the period of latent infection (> 20 days) no cytotoxic cell activity was observed. Cytotoxic activity could only be detected when the lymphoid cells had been cultured for a period of 3 days. In general, the cell killing was specific for syngeneic infected target cells, although some killing of uninfected targets was observed. In contrast to the cytotoxic response, DLN cells responding to HSV in a proliferation assay were detected towards the end of the acute phase and at lease up to 9 months thereafter. The significance of these observations for the pathogenesis of HSV is discussed.

Incidence and Predictors of Bacterial infection in Febrile Children with Sickle Cell Disease.

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Morrissey, Benita J; Bycroft, Thomas P; Almossawi, Ofran; Wilkey, Olufunke B; Daniels, Justin G

2015-01-01

Children with sickle cell disease are at increased risk of developing bacteremia and other serious bacterial infections. Fever is a common symptom in sickle cell disease and can also occur with sickle cell crises and viral infections. We aimed to evaluate the incidence and predictors of bacteremia and bacterial infection in children with sickle cell disease presenting with fever to a district hospital and sickle cell center in London. A retrospective analysis was performed on all attendances of children (aged under 16 years) with sickle cell disease presenting with a fever of 38.5 °C or higher over a 1-year period. Confirmed bacterial infection was defined as bacteremia, bacterial meningitis, urinary tract infection (UTI), pneumonia, osteomyelitis or other bacterial infection with positive identification of organism. Children were defined as having a suspected bacterial infection if a bacterial infection was suspected clinically, but no organism was identified. Over a 1-year period there were 88 episodes analyzed in 59 children. Bacteremia occurred in 3.4% of episodes and confirmed bacterial infection in 7.0%. Suspected bacterial infection occurred in 33.0%. One death occurred from Salmonella typhirium septicemia. C-reactive protein (CRP) level and white blood cell (WBC) count were both significantly associated with bacterial infection (p = 0.004 and 0.02, respectively.) In conclusion, bacterial infections continue to be a significant problem in children with sickle cell disease. C-reactive protein was significantly associated with bacterial infections, and could be included in clinical risk criteria for febrile children with sickle cell disease.

Tissue-specific B-cell dysfunction and generalized memory B-cell loss during acute SIV infection.

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Sandrine Peruchon

Full Text Available BACKGROUND: Primary HIV-infected patients display severe and irreversible damage to different blood B-cell subsets which is not restored by highly efficient anti-retroviral therapy (HAART. Because longitudinal investigations of primary HIV-infection is limited by the availability of lymphoid organs, we studied the tissue-specific B-cell dysfunctions in acutely simian immunodeficiency virus (SIV mac251-infected Cynomolgus macaques. METHODS AND FINDINGS: Experiments were performed on three groups of macaques infected for 14, 21 or 28 days and on three groups of animals treated with HAART for two-weeks either initiated at 4 h, 7 or 14 days post-infection (p.i.. We have simultaneously compared changes in B-cell phenotypes and functions and tissue organization of B-cell areas in various lymphoid organs. We showed that SIV induced a steady decline in SIgG-expressing memory (SIgD(-CD27(+ B-cells in spleen and lymph nodes during the first 4 weeks of infection, concomitant to selective homing/sequestration of B-cells to the small intestine and spleen. SIV non-specific Ig production was transiently increased before D14p.i., whereas SIV-specific Ig production was only detectable after D14p.i., coinciding with the presence of CD8(+ T-cells and IgG-expressing plasma cells within germinal centres. Transient B-cell apoptosis on D14p.i. and commitment to terminal differentiation contributed to memory B-cell loss. HAART abrogated B-cell apoptosis, homing to the small intestine and SIV-specific Ig production but had minimal effect on early Ig production, increased B-cell proportions in spleen and loss of memory B-cells. Therefore, virus-B-cell interactions and SIV-induced inflammatory cytokines may differently contribute to early B-cell dysfunction and impaired SIV/HIV-specific antibody response. CONCLUSIONS: These data establish tissue-specific impairments in B-cell trafficking and functions and a generalized and steady memory B-cell loss in secondary lymphoid

Cytotoxic CD4 T Cells: Differentiation, Function, and Application to Dengue Virus Infection.

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Tian, Yuan; Sette, Alessandro; Weiskopf, Daniela

2016-01-01

Dengue virus (DENV) has spread through most tropical and subtropical areas of the world and represents a serious public health problem. The control of DENV infection has not yet been fully successful due to lack of effective therapeutics or vaccines. Nevertheless, a better understanding of the immune responses against DENV infection may reveal new strategies for eliciting and improving antiviral immunity. T cells provide protective immunity against various viral infections by generating effector cells that cooperate to eliminate antigens and memory cells that can survive for long periods with enhanced abilities to control recurring pathogens. Following activation, CD8 T cells can migrate to sites of infection and kill infected cells, whereas CD4 T cells contribute to the elimination of pathogens by trafficking to infected tissues and providing help to innate immune responses, B cells, as well as CD8 T cells. However, it is now evident that CD4 T cells can also perform cytotoxic functions and induce the apoptosis of target cells. Importantly, accumulating studies demonstrate that cytotoxic CD4 T cells develop following DENV infections and may play a crucial role in protecting the host from severe dengue disease. We review our current understanding of the differentiation and function of cytotoxic CD4 T cells, with a focus on DENV infection, and discuss the potential of harnessing these cells for the prevention and treatment of DENV infection and disease.

iNKT Cells and Their potential Lipid Ligands during Viral Infection

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Anunya eOpasawatchai

2015-07-01

Full Text Available Invariant natural killer T (iNKT cells are a unique population of lipid reactive CD1d restricted innate-like T lymphocytes. Despite being a minor population, they serve as an early source of cytokines and promote immunological crosstalk thus bridging innate and adaptive immunity. Diseases ranging from allergy, autoimmunity, and cancer as well as infectious diseases, including viral infection, have been reported to be influenced by iNKT cells. However, it remains unclear how iNKT cells are activated during viral infection, as virus derived lipid antigens have not been reported. Cytokines may activate iNKT cells during infections from influenza and murine cytomegalovirus (MCMV, although CD1d dependent activation is evident in other viral infections. Several viruses, such as dengue virus (DENV, induce CD1d upregulation which correlates with iNKT cell activation. In contrast, Herpes simplex virus type 1 (HSV-1, Human immunodeficiency virus (HIV, Epstein-Barr virus (EBV and Human papiloma virus (HPV promote CD1d downregulation as a strategy to evade iNKT cell recognition. These observations suggest the participation of a CD1d-dependent process in the activation of iNKT cells in response to viral infection. Endogenous lipid ligands, including phospholipids as well as glycosphingolipids, such as glucosylceramide have been proposed to mediate iNKT cell activation. Pro-inflammatory signals produced during viral infection may stimulate iNKT cells through enhanced CD1d dependent endogenous lipid presentation. Furthermore, viral infection may alter lipid composition and inhibit endogenous lipid degradation. Recent advances in this field are reviewed.

Dengue Virus-Infected Dendritic Cells, but Not Monocytes, Activate Natural Killer Cells through a Contact-Dependent Mechanism Involving Adhesion Molecules.

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Costa, Vivian Vasconcelos; Ye, Weijian; Chen, Qingfeng; Teixeira, Mauro Martins; Preiser, Peter; Ooi, Eng Eong; Chen, Jianzhu

2017-08-01

Natural killer (NK) cells play a protective role against dengue virus (DENV) infection, but the cellular and molecular mechanisms are not fully understood. Using an optimized humanized mouse model, we show that human NK cells, through the secretion of gamma interferon (IFN-γ), are critical in the early defense against DENV infection. Depletion of NK cells or neutralization of IFN-γ leads to increased viremia and more severe thrombocytopenia and liver damage in humanized mice. In vitro studies using autologous human NK cells show that DENV-infected monocyte-derived dendritic cells (MDDCs), but not monocytes, activate NK cells in a contact-dependent manner, resulting in upregulation of CD69 and CD25 and secretion of IFN-γ. Blocking adhesion molecules (LFA-1, DNAM-1, CD2, and 2β4) on NK cells abolishes NK cell activation, IFN-γ secretion, and the control of DENV replication. NK cells activated by infected MDDCs also inhibit DENV infection in monocytes. These findings show the essential role of human NK cells in protection against acute DENV infection in vivo , identify adhesion molecules and dendritic cells required for NK cell activation, and delineate the sequence of events for NK cell activation and protection against DENV infection. IMPORTANCE Dengue is a mosquito-transmitted viral disease with a range of symptoms, from mild fever to life-threatening dengue hemorrhagic fever. The diverse disease manifestation is thought to result from a complex interplay between viral and host factors. Using mice engrafted with a human immune system, we show that human NK cells inhibit virus infection through secretion of the cytokine gamma interferon and reduce disease pathogenesis, including depletion of platelets and liver damage. During a natural infection, DENV initially infects dendritic cells in the skin. We find that NK cells interact with infected dendritic cells through physical contact mediated by adhesion molecules and become activated before they can control

Loss of circulating CD4 T cells with B cell helper function during chronic HIV infection.

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Kristin L Boswell

2014-01-01

Full Text Available The interaction between follicular T helper cells (TFH and B cells in the lymph nodes and spleen has a major impact on the development of antigen-specific B cell responses during infection or vaccination. Recent studies described a functional equivalent of these cells among circulating CD4 T cells, referred to as peripheral TFH cells. Here, we characterize the phenotype and in vitro B cell helper activity of peripheral TFH populations, as well as the effect of HIV infection on these populations. In co-culture experiments we confirmed CXCR5+ cells from HIV-uninfected donors provide help to B cells and more specifically, we identified a CCR7(highCXCR5(highCCR6(highPD-1(high CD4 T cell population that secretes IL-21 and enhances isotype-switched immunoglobulin production. This population is significantly decreased in treatment-naïve, HIV-infected individuals and can be recovered after anti-retroviral therapy. We found impaired immunoglobulin production in co-cultures from HIV-infected individuals and found no correlation between the frequency of peripheral TFH cells and memory B cells, or with neutralization activity in untreated HIV infection in our cohort. Furthermore, we found that within the peripheral TFH population, the expression level of TFH-associated genes more closely resembles a memory, non-TFH population, as opposed to a TFH population. Overall, our data identify a heterogeneous population of circulating CD4 T cells that provides in vitro help to B cells, and challenges the origin of these cells as memory TFH cells.

Autophagy protects type II alveolar epithelial cells from Mycobacterium tuberculosis infection

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Guo, Xu-Guang; Ji, Tian-Xing; Xia, Yong; Ma, Yue-Yun

2013-01-01

Highlights: ► We investigated the protective effect of autophagy pathway against MTB infection. ► MTB-infected A549 cells had higher LDH release. ► Inhibition of autophagy signaling significantly enhanced the MTB-induced necrosis. ► Autophagy prevents apoptosis and promotes cell survival in infected cells. -- Abstract: This study was designed to investigate the protective effect of the autophagy signaling pathway against Mycobacterium tuberculosis infection in type II alveolar epithelial cells. An in vitro M. tuberculosis system was established using human A549 cells. Infection-induced changes in the expression of the autophagic marker LC3 were assessed by reverse transcription-PCR and Western blotting. Morphological changes in autophagosomes were detected by transmission electron microscopy (TEM). The function of the autophagy signaling pathway during infection was assessed by measuring the level of cell death and the amount of lactate dehydrogenase (LDH) released in the presence or absence of the inhibitor 3-methyladenine (3-MA). In addition, effects on LDH release were assessed after the siRNA-mediated knockdown of the essential autophagosomal structural membrane protein Atg5. LC3 mRNA expression was significantly reduced in M.tuberculosis-infected A549 cells (16888.76 ± 1576.34 vs. uninfected: 12744.29 ± 1089.37; P < 0.05). TEM revealed M.tuberculosis bacilli-containing compartments that were surrounded by double membranes characteristic of the autophagic process. M.tuberculosis-infected A549 cells released more LDH (1.45 ± 0.12 vs. uninfected: 0.45 ± 0.04; P < 0.05). The inhibition of autophagy signaling significantly enhanced M.tuberculosis-induced necrosis (3-MA: 75 ± 5% vs. untreated: 15 ± 1%; P < 0.05) and LDH release (3-MA: 2.50 ± 0.24 vs. untreated: 0.45 ± 0.04; Atg5 knockdown: 3.19 ± 0.29 vs. untreated: 1.28 ± 0.11; P < 0.05). Our results indicate that autophagy signaling pathway prevents apoptosis in type II alveolar epithelial cells

Tax gene expression and cell cycling but not cell death are selected during HTLV-1 infection in vivo

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Pinatel Christiane

2010-03-01

Full Text Available Abstract Background Adult T cell leukemia results from the malignant transformation of a CD4+ lymphoid clone carrying an integrated HTLV-1 provirus that has undergone several oncogenic events over a 30-60 year period of persistent clonal expansion. Both CD4+ and CD8+ lymphocytes are infected in vivo; their expansion relies on CD4+ cell cycling and on the prevention of CD8+ cell death. Cloned infected CD4+ but not CD8+ T cells from patients without malignancy also add up nuclear and mitotic defects typical of genetic instability related to theexpression of the virus-encoded oncogene tax. HTLV-1 expression is cancer-prone in vitro, but in vivo numerous selection forces act to maintain T cell homeostasis and are possibly involved in clonal selection. Results Here we demonstrate that the HTLV-1 associated CD4+ preleukemic phenotype and the specific patterns of CD4+ and CD8+ clonal expansion are in vivo selected processes. By comparing the effects of recent (1 month experimental infections performed in vitro and those observed in cloned T cells from patients infected for >6-26 years, we found that in chronically HTLV-1 infected individuals, HTLV-1 positive clones are selected for tax expression. In vivo, infected CD4+ cells are positively selected for cell cycling whereas infected CD8+ cells and uninfected CD4+ cells are negatively selected for the same processes. In contrast, the known HTLV-1-dependent prevention of CD8+ T cell death pertains to both in vivo and in vitro infected cells. Conclusions Therefore, virus-cell interactions alone are not sufficient to initiate early leukemogenesis in vivo.

T-cell responses targeting HIV Nef uniquely correlate with infected cell frequencies after long-term antiretroviral therapy.

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Allison S Thomas

2017-09-01

Full Text Available HIV-specific CD8+ T-cell responses limit viral replication in untreated infection. After the initiation of antiretroviral therapy (ART, these responses decay and the infected cell population that remains is commonly considered to be invisible to T-cells. We hypothesized that HIV antigen recognition may persist in ART-treated individuals due to low-level or episodic protein expression. We posited that if persistent recognition were occurring it would be preferentially directed against the early HIV gene products Nef, Tat, and Rev as compared to late gene products, such as Gag, Pol, and Env, which have higher barriers to expression. Using a primary cell model of latency, we observed that a Nef-specific CD8+ T-cell clone exhibited low-level recognition of infected cells prior to reactivation and robust recognition shortly thereafter. A Gag-specific CD8+ T-cell clone failed to recognized infected cells under these conditions, corresponding with a lack of detectable Gag expression. We measured HIV-specific T-cell responses in 96 individuals who had been suppressed on ART for a median of 7 years, and observed a significant, direct correlation between cell-associated HIV DNA levels and magnitudes of IFN-γ-producing Nef/Tat/Rev-specific T-cell responses. This correlation was confirmed in an independent cohort (n = 18. Correlations were not detected between measures of HIV persistence and T-cell responses to other HIV antigens. The correlation with Nef/Tat/Rev-specific T-cells was attributable to Nef-specific responses, the breadth of which also correlated with HIV DNA levels. These results suggest that ongoing Nef expression in ART-treated individuals drives preferential maintenance and/or expansion of T-cells reactive to this protein, implying sensing of infected cells by the immune system. The direct correlation, however, suggests that recognition does not result in efficient elimination of infected cells. These results raise the possibility that

Antigen Requirements for Efficient Priming of CD8+ T Cells by Leishmania major-Infected Dendritic Cells

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Bertholet, Sylvie; Debrabant, Alain; Afrin, Farhat; Caler, Elisabeth; Mendez, Susana; Tabbara, Khaled S.; Belkaid, Yasmine; Sacks, David L.

2005-01-01

CD4+ and CD8+ T-cell responses have been shown to be critical for the development and maintenance of acquired resistance to infections with the protozoan parasite Leishmania major. Monitoring the development of immunodominant or clonally restricted T-cell subsets in response to infection has been difficult, however, due to the paucity of known epitopes. We have analyzed the potential of L. major transgenic parasites, expressing the model antigen ovalbumin (OVA), to be presented by antigen-presenting cells to OVA-specific OT-II CD4+ or OT-I CD8+ T cells. Truncated OVA was expressed in L. major as part of a secreted or nonsecreted chimeric protein with L. donovani 3′ nucleotidase (NT-OVA). Dendritic cells (DC) but not macrophages infected with L. major that secreted NT-OVA could prime OT-I T cells to proliferate and release gamma interferon. A diminished T-cell response was observed when DC were infected with parasites expressing nonsecreted NT-OVA or with heat-killed parasites. Inoculation of mice with transgenic parasites elicited the proliferation of adoptively transferred OT-I T cells and their recruitment to the site of infection in the skin. Together, these results demonstrate the possibility of targeting heterologous antigens to specific cellular compartments in L. major and suggest that proteins secreted or released by L. major in infected DC are a major source of peptides for the generation of parasite-specific CD8+ T cells. The ability of L. major transgenic parasites to activate OT-I CD8+ T cells in vivo will permit the analysis of parasite-driven T-cell expansion, differentiation, and recruitment at the clonal level. PMID:16177338

Roles for Endothelial Cells in Dengue Virus Infection

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Nadine A. Dalrymple

2012-01-01

Full Text Available Dengue viruses cause two severe diseases that alter vascular fluid barrier functions, dengue hemorrhagic fever (DHF and dengue shock syndrome (DSS. The endothelium is the primary fluid barrier of the vasculature and ultimately the effects of dengue virus infection that cause capillary leakage impact endothelial cell (EC barrier functions. The ability of dengue virus to infect the endothelium provides a direct means for dengue to alter capillary permeability, permit virus replication, and induce responses that recruit immune cells to the endothelium. Recent studies focused on dengue virus infection of primary ECs have demonstrated that ECs are efficiently infected, rapidly produce viral progeny, and elicit immune enhancing cytokine responses that may contribute to pathogenesis. Furthermore, infected ECs have also been implicated in enhancing viremia and immunopathogenesis within murine dengue disease models. Thus dengue-infected ECs have the potential to directly contribute to immune enhancement, capillary permeability, viremia, and immune targeting of the endothelium. These effects implicate responses of the infected endothelium in dengue pathogenesis and rationalize therapeutic targeting of the endothelium and EC responses as a means of reducing the severity of dengue virus disease.

Virus specific antigens in mammalian cells infected with herpes simplex virus

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Watson, D. H.; Shedden, W. I. H.; Elliot, A.; Tetsuka, T.; Wildy, P.; Bourgaux-Ramoisy, D.; Gold, E.

1966-01-01

Antisera to specific proteins in herpes simplex infected cells were produced by immunization of rabbits with infected rabbit kidney cells. These antisera were highly virus specific and produced up to twelve lines in immunodiffusion tests against infected cell extracts. Acrylamide electrophoresis and immunoelectrophoresis revealed up to ten virus specific proteins of varying size. ImagesFIG. 1FIG. 2FIG. 3FIG. 4FIG. 5 PMID:4288648

Analysis of gene expression in fetal and adult cells infected with rubella virus

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Adamo, Maria Pilar; Zapata, Marta; Frey, Teryl K.

2008-01-01

Congenital infection with rubella virus (RUB) leads to persistent infection and congenital defects and we showed previously that primary human fetal fibroblasts did not undergo apoptosis when infected with RUB, which could promote fetal virus persistence [Adamo, P., Asis, L., Silveyra, P., Cuffini, C., Pedranti, M., Zapata, M., 2004. Rubella virus does not induce apoptosis in primary human embryo fibroblasts cultures: a possible way of viral persistence in congenital infection. Viral Immunol. 17, 87-100]. To extend this observation, gene chip analysis was performed on a line of primary human fetal fibroblasts (10 weeks gestation) and a line of human adult lung fibroblasts (which underwent apoptosis in response to RUB infection) to compare gene expression in infected and uninfected cells. A total of 632 and 516 genes were upregulated or downregulated in the infected fetal and adult cells respectively in comparison to uninfected cells, however only 52 genes were regulated in both cell types. Although the regulated genes were different, across functional gene categories the patterns of gene regulation were similar. In general, regulation of pro- and anti-apoptotic genes following infection appeared to favor apoptosis in the adult cells and lack of apoptosis in the fetal cells, however there was a greater relative expression of anti-apoptotic genes and reduced expression of pro-apoptotic genes in uninfected fetal cells versus uninfected adult cells and thus the lack of apoptosis in fetal cells following RUB infection was also due to the prevailing background of gene expression that is antagonistic to apoptosis. In support of this hypothesis, it was found that of a battery of five chemicals known to induce apoptosis, two induced apoptosis in the adult cells, but not in fetal cells, and two induced apoptosis more rapidly in the adult cells than in fetal cells (the fifth did not induce apoptosis in either). A robust interferon-stimulated gene response was induced

Early Loss of Splenic Tfh Cells in SIV-Infected Rhesus Macaques.

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Félicien Moukambi

2015-12-01

Full Text Available Follicular T helper cells (Tfh, a subset of CD4 T lymphocytes, provide crucial help to B cells in the production of antigen-specific antibodies. Although several studies have analyzed the dynamics of Tfh cells in peripheral blood and lymph nodes (LNs during Aids, none has yet addressed the impact of SIV infection on the dynamics of Tfh cells in the spleen, the primary organ of B cell activation. We show here a significant decrease in splenic Tfh cells in SIVmac251-infected rhesus macaques (RMs during the acute phase of infection, which persists thereafter. This profound loss is associated with lack of sustained expression of the Tfh-defining transcription factors, Bcl-6 and c-Maf but with higher expression of the repressors KLF2 and Foxo1. In this context of Tfh abortive differentiation and loss, we found decreased percentages of memory B cell subsets and lower titers of SIV-specific IgG. We further demonstrate a drastic remodeling of the lymphoid architecture of the spleen and LNs, which disrupts the crucial cell-cell interactions necessary to maintain memory B cells and Tfh cells. Finally, our data demonstrated the early infection of Tfh cells. Paradoxically, the frequencies of SIV DNA were higher in splenic Tfh cells of RMs progressing more slowly suggesting sanctuaries for SIV in the spleen. Our findings provide important information regarding the impact of HIV/SIV infection on Tfh cells, and provide new clues for future vaccine strategies.

Analysis of differentially expressed proteins in Yersinia enterocolitica-infected HeLa cells.

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Alugubelly, Navatha; Hercik, Kamil; Kibler, Peter; Nanduri, Bindu; Edelmann, Mariola J

2016-05-01

Yersinia enterocolitica is a facultative intracellular pathogen and a causative agent of yersiniosis, which can be contracted by ingestion of contaminated food. Yersinia secretes virulence factors to subvert critical pathways in the host cell. In this study we utilized shotgun label-free proteomics to study differential protein expression in epithelial cells infected with Y.enterocolitica. We identified a total of 551 proteins, amongst which 42 were downregulated (including Prostaglandin E Synthase 3, POH-1 and Karyopherin alpha) and 22 were upregulated (including Rab1 and RhoA) in infected cells. We validated some of these results by western blot analysis of proteins extracted from Caco-2 and HeLa cells. The proteomic dataset was used to identify host canonical pathways and molecular functions modulated by this infection in the host cells. This study constitutes a proteome of Yersinia-infected cells and can support new discoveries in the area of host-pathogen interactions. We describe a proteome of Yersinia enterocolitica-infected HeLa cells, including a description of specific proteins differentially expressed upon infection, molecular functions as well as pathways altered during infection. This proteomic study can lead to a better understanding of Y. enterocolitica pathogenesis in human epithelial cells. Copyright © 2016. Published by Elsevier B.V.

CD154 costimulated ovine primary B cells, a cell culture system that supports productive infection by bovine leukemia virus.

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Van den Broeke, A; Cleuter, Y; Beskorwayne, T; Kerkhofs, P; Szynal, M; Bagnis, C; Burny, A; Griebel, P

2001-02-01

Bovine leukemia virus (BLV) is closely associated with the development of B-cell leukemia and lymphoma in cattle. BLV infection has also been studied extensively in an in vivo ovine model that provides a unique system for studying B-cell leukemogenesis. There is no evidence that BLV can directly infect ovine B cells in vitro, and there are no direct data regarding the oncogenic potential of the viral Tax transactivator in B cells. Therefore, we developed ovine B-cell culture systems to study the interaction between BLV and its natural target, the B cell. In this study, we used murine CD154 (CD40 ligand) and gamma-chain-common cytokines to support the growth of B cells isolated from ovine lymphoid tissues. Integrated provirus, extrachromosomal forms, and viral transcripts were detected in BLV-exposed populations of immature, rapidly dividing surface immunoglobulin M-positive B cells from sheep ileal Peyer's patches and also in activated mature B cells isolated from blood. Conclusive evidence of direct B-cell infection by BLV was obtained through the use of cloned B cells derived from sheep jejunal Peyer's patches. Finally, inoculation of sheep with BLV-infected cultures proved that infectious virus was shed from in vitro-infected B cells. Collectively, these data confirm that a variety of ovine B-cell populations can support productive infection by BLV. The development of ovine B-cell cultures permissive for BLV infection provides a controlled system for investigating B-cell leukemogenic processes and the pathogenesis of BLV infection.

Ebola virus infection induces irregular dendritic cell gene expression.

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Melanson, Vanessa R; Kalina, Warren V; Williams, Priscilla

2015-02-01

Filoviruses subvert the human immune system in part by infecting and replicating in dendritic cells (DCs). Using gene arrays, a phenotypic profile of filovirus infection in human monocyte-derived DCs was assessed. Monocytes from human donors were cultured in GM-CSF and IL-4 and were infected with Ebola virus Kikwit variant for up to 48 h. Extracted DC RNA was analyzed on SuperArray's Dendritic and Antigen Presenting Cell Oligo GEArray and compared to uninfected controls. Infected DCs exhibited increased expression of cytokine, chemokine, antiviral, and anti-apoptotic genes not seen in uninfected controls. Significant increases of intracellular antiviral and MHC I and II genes were also noted in EBOV-infected DCs. However, infected DCs failed to show any significant difference in co-stimulatory T-cell gene expression from uninfected DCs. Moreover, several chemokine genes were activated, but there was sparse expression of chemokine receptors that enabled activated DCs to home to lymph nodes. Overall, statistically significant expression of several intracellular antiviral genes was noted, which may limit viral load but fails to stop replication. EBOV gene expression profiling is of vital importance in understanding pathogenesis and devising novel therapeutic treatments such as small-molecule inhibitors.

Early Trypanosoma cruzi Infection Reprograms Human Epithelial Cells

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María Laura Chiribao

2014-01-01

Full Text Available Trypanosoma cruzi, the causative agent of Chagas disease, has the peculiarity, when compared with other intracellular parasites, that it is able to invade almost any type of cell. This property makes Chagas a complex parasitic disease in terms of prophylaxis and therapeutics. The identification of key host cellular factors that play a role in the T. cruzi invasion is important for the understanding of disease pathogenesis. In Chagas disease, most of the focus is on the response of macrophages and cardiomyocytes, since they are responsible for host defenses and cardiac lesions, respectively. In the present work, we studied the early response to infection of T. cruzi in human epithelial cells, which constitute the first barrier for establishment of infection. These studies identified up to 1700 significantly altered genes regulated by the immediate infection. The global analysis indicates that cells are literally reprogrammed by T. cruzi, which affects cellular stress responses (neutrophil chemotaxis, DNA damage response, a great number of transcription factors (including the majority of NFκB family members, and host metabolism (cholesterol, fatty acids, and phospholipids. These results raise the possibility that early host cell reprogramming is exploited by the parasite to establish the initial infection and posterior systemic dissemination.

The involvement of plasmacytoid cells in HIV infection and pathogenesis.

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Aiello, Alessandra; Giannessi, Flavia; Percario, Zulema A; Affabris, Elisabetta

2018-04-01

Plasmacytoid dendritic cells (pDCs) are a unique dendritic cell subset that are specialized in type I interferon (IFN) production. pDCs are key players in the antiviral immune response and serve as bridge between innate and adaptive immunity. Although pDCs do not represent the main reservoir of the Human Immunodeficiency Virus (HIV), they are a crucial subset in HIV infection as they influence viral transmission, target cell infection and antigen presentation. pDCs act as inflammatory and immunosuppressive cells, thus contributing to HIV disease progression. This review provides a state of art analysis of the interactions between HIV and pDCs and their potential roles in HIV transmission, chronic immune activation and immunosuppression. A thorough understanding of the roles of pDCs in HIV infection will help to improve therapeutic strategies to fight HIV infection, and will further increase our knowledge on this important immune cell subset. Copyright © 2018 Elsevier Ltd. All rights reserved.

Innate Lymphoid Cells in HIV/SIV Infections.

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Shah, Spandan V; Manickam, Cordelia; Ram, Daniel R; Reeves, R Keith

2017-01-01

Over the past several years, new populations of innate lymphocytes have been described in mice and primates that are critical for mucosal homeostasis, microbial regulation, and immune defense. Generally conserved from mice to humans, innate lymphoid cells (ILC) have been divided primarily into three subpopulations based on phenotypic and functional repertoires: ILC1 bear similarities to natural killer cells; ILC2 have overlapping functions with TH2 cells; and ILC3 that share many functions with TH17/TH22 cells. ILC are specifically enriched at mucosal surfaces and are possibly one of the earliest responders during viral infections besides being involved in the homeostasis of gut-associated lymphoid tissue and maintenance of gut epithelial barrier integrity. Burgeoning evidence also suggests that there is an early and sustained abrogation of ILC function and numbers during HIV and pathogenic SIV infections, most notably ILC3 in the gastrointestinal tract, which leads to disruption of the mucosal barrier and dysregulation of the local immune system. A better understanding of the direct or indirect mechanisms of loss and dysfunction will be critical to immunotherapeutics aimed at restoring these cells. Herein, we review the current literature on ILC with a particular emphasis on ILC3 and their role(s) in mucosal immunology and the significance of disrupting the ILC niche during HIV and SIV infections.

Innate Lymphoid Cells in HIV/SIV Infections

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Spandan V. Shah

2017-12-01

Full Text Available Over the past several years, new populations of innate lymphocytes have been described in mice and primates that are critical for mucosal homeostasis, microbial regulation, and immune defense. Generally conserved from mice to humans, innate lymphoid cells (ILC have been divided primarily into three subpopulations based on phenotypic and functional repertoires: ILC1 bear similarities to natural killer cells; ILC2 have overlapping functions with TH2 cells; and ILC3 that share many functions with TH17/TH22 cells. ILC are specifically enriched at mucosal surfaces and are possibly one of the earliest responders during viral infections besides being involved in the homeostasis of gut-associated lymphoid tissue and maintenance of gut epithelial barrier integrity. Burgeoning evidence also suggests that there is an early and sustained abrogation of ILC function and numbers during HIV and pathogenic SIV infections, most notably ILC3 in the gastrointestinal tract, which leads to disruption of the mucosal barrier and dysregulation of the local immune system. A better understanding of the direct or indirect mechanisms of loss and dysfunction will be critical to immunotherapeutics aimed at restoring these cells. Herein, we review the current literature on ILC with a particular emphasis on ILC3 and their role(s in mucosal immunology and the significance of disrupting the ILC niche during HIV and SIV infections.

The CD8 T Cell Response to Respiratory Virus Infections.

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Schmidt, Megan E; Varga, Steven M

2018-01-01

Humans are highly susceptible to infection with respiratory viruses including respiratory syncytial virus (RSV), influenza virus, human metapneumovirus, rhinovirus, coronavirus, and parainfluenza virus. While some viruses simply cause symptoms of the common cold, many respiratory viruses induce severe bronchiolitis, pneumonia, and even death following infection. Despite the immense clinical burden, the majority of the most common pulmonary viruses lack long-lasting efficacious vaccines. Nearly all current vaccination strategies are designed to elicit broadly neutralizing antibodies, which prevent severe disease following a subsequent infection. However, the mucosal antibody response to many respiratory viruses is not long-lasting and declines with age. CD8 T cells are critical for mediating clearance following many acute viral infections in the lung. In addition, memory CD8 T cells are capable of providing protection against secondary infections. Therefore, the combined induction of virus-specific CD8 T cells and antibodies may provide optimal protective immunity. Herein, we review the current literature on CD8 T cell responses induced by respiratory virus infections. Additionally, we explore how this knowledge could be utilized in the development of future vaccines against respiratory viruses, with a special emphasis on RSV vaccination.

Changes in cell adhesion molecule expression on T cells associated with systemic virus infection

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Andersson, E C; Christensen, Jan Pravsgaard; Marker, O

1994-01-01

-4, LFA-1, and ICAM-1, are up-regulated on CD8+ cells, whereas the lymph node homing receptor MEL-14 is down-regulated during the infection; only marginal changes were observed for CD4+ cells. Basically similar but less marked results were obtained in mice infected with Pichinde virus. Further...

DRAM Triggers Lysosomal Membrane Permeabilization and Cell Death in CD4+ T Cells Infected with HIV

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Laforge, Mireille; Limou, Sophie; Harper, Francis; Casartelli, Nicoletta; Rodrigues, Vasco; Silvestre, Ricardo; Haloui, Houda; Zagury, Jean-Francois; Senik, Anna; Estaquier, Jerome

2013-01-01

Productive HIV infection of CD4+ T cells leads to a caspase-independent cell death pathway associated with lysosomal membrane permeabilization (LMP) and cathepsin release, resulting in mitochondrial outer membrane permeabilization (MOMP). Herein, we demonstrate that HIV infection induces damage-regulated autophagy modulator (DRAM) expression in a p53-dependent manner. Knocking down the expression of DRAM and p53 genes with specific siRNAs inhibited autophagy and LMP. However, inhibition of Atg5 and Beclin genes that prevents autophagy had a minor effect on LMP and cell death. The knock down of DRAM gene inhibited cytochrome C release, MOMP and cell death. However, knocking down DRAM, we increased viral infection and production. Our study shows for the first time the involvement of DRAM in host-pathogen interactions, which may represent a mechanism of defense via the elimination of infected cells. PMID:23658518

Increased CD56(bright) NK cells in HIV-HCV co-infection and HCV mono-infection are associated with distinctive alterations of their phenotype.

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Bhardwaj, Suvercha; Ahmad, Fareed; Wedemeyer, Heiner; Cornberg, Marcus; Schulze Zur Wiesch, Julian; van Lunzen, Jan; Sarin, Shiv K; Schmidt, Reinhold E; Meyer-Olson, Dirk

2016-04-18

HIV-HCV co-infection is associated with accelerated progression to hepatic fibrosis, cirrhosis and hepatocellular carcinoma than HCV mono-infection. The contribution of innate immunity during HIV-HCV co-infection has been a relatively under-investigated area. Natural killer (NK) cells are pivotal sentinels of innate immunity against viruses and tumour cells. In this study we evaluated the effect of HIV-HCV co-infection on peripheral blood NK cell subsets with emphasis on the phenotype of CD56(bright) NK cells. Sixty patients were included in the study; HIV mono-infected (n = 12), HCV mono-infected (n = 15), HCV-HIV co-infected (n = 21) and healthy controls (n = 16). PBMCs were isolated and immunophenotyping of NK cells was performed by flowcytometry. We observed an expansion of CD56(bright) NK cell subset in HIV-HCV co-infection as compared to healthy controls and HIV mono-infected group. All the infected groups had an upregulated expression of the activating receptor NKG2D on CD56(bright) NK cells in comparison to healthy controls while not differing amongst themselves. The expression of NKp46 in HIV-HCV co-infected group was significantly upregulated as compared to both HIV as well as HCV mono-infections while NKp30 expression in the HIV-HCV co-infected group significantly differed as compared to HIV mono-infection. The CD56(bright) NK cell subset was activated in HIV-HCV co-infection as assessed by the expression of CD69 as compared to healthy controls but was significantly downregulated in comparison to HIV mono-infection. CD95 expression on CD56(bright) NK cells followed the same pattern where there was an increased expression of CD95 in HIV mono-infection and HIV-HCV co-infection as compared to healthy controls. In contrast to CD69 expression, CD95 expression in HCV mono-infection was decreased when compared to HIV mono-infection and HIV-HCV co-infection. Finally, expression of CXCR3 on CD56(bright) NK cells was increased in HIV-HCV co-infection in comparison

Selective receptor expression restricts Nipah virus infection of endothelial cells

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Diederich Sandra

2008-11-01

Full Text Available Abstract Nipah virus (NiV is a highly pathogenic paramyxovirus that causes severe diseases in animals and humans. Endothelial cell (EC infection is an established hallmark of NiV infection in vivo. Despite systemic virus spread via the vascular system, EC in brain and lung are preferentially infected whereas EC in other organs are less affected. As in vivo, we found differences in the infection of EC in cell culture. Only brain-derived primary or immortalized EC were found to be permissive to NiV infection. Using a replication-independent fusion assay, we could show that the lack of infection in non-brain EC was due to a lack of receptor expression. The NiV entry receptors ephrinB2 (EB2 or ephrinB3 were only expressed in brain endothelia. The finding that EB2 expression in previously non-permissive aortic EC rendered the cells permissive to infection then demonstrated that EB2 is not only necessary but also sufficient to allow the establishment of a productive NiV infection. This strongly suggests that limitations in receptor expression restrict virus entry in certain EC subsets in vivo, and are thus responsible for the differences in EC tropism observed in human and animal NiV infections.

Experimental analysis of residual stresses in pre-straightened SAE 1045 steel

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Diehl, Carla Adriana Theis Soares; Rocha, Alexandre da Silva, E-mail: carla.adriana@ufrgs.br [Universidade Federal do Rio Grande do Sul (UFRGS), Porto Alegre, RS (Brazil). Laboratorio de Formacao de Metais; Epp, Jérémy; Zoch, Hans-Werner [Stiftung Institut für Werkstofftechnik IWT, University of Bremen (Germany)

2017-11-15

This paper aims at analyzing the effects of the roller pre-straightening of wire-rods on residual stress distributions in SAE 1045 steel bars. The combined drawing process is used in industrial production of bars in order to obtain a good surface quality and improved mechanical properties complying with specifications of the final products. In this process, prior to the drawing step, a roller straightening of the steel wire-rod is essential, because it provides the minimum straightness necessary for drawing. Metallographic analysis and hardness test were done for selected samples after different processing steps. Also, residual stress analysis of pre-straightened wire-rods by X-ray diffraction and neutron diffraction were carried out. The hardness tests show higher values near the surface and lower in the center of the wire-rod. Besides, the residual stresses results show a big inhomogeneity from one peripheral position to another and also in the evaluated cross section. (author)

Experimental analysis of residual stresses in pre-straightened SAE 1045 steel

International Nuclear Information System (INIS)

Diehl, Carla Adriana Theis Soares; Rocha, Alexandre da Silva

2017-01-01

This paper aims at analyzing the effects of the roller pre-straightening of wire-rods on residual stress distributions in SAE 1045 steel bars. The combined drawing process is used in industrial production of bars in order to obtain a good surface quality and improved mechanical properties complying with specifications of the final products. In this process, prior to the drawing step, a roller straightening of the steel wire-rod is essential, because it provides the minimum straightness necessary for drawing. Metallographic analysis and hardness test were done for selected samples after different processing steps. Also, residual stress analysis of pre-straightened wire-rods by X-ray diffraction and neutron diffraction were carried out. The hardness tests show higher values near the surface and lower in the center of the wire-rod. Besides, the residual stresses results show a big inhomogeneity from one peripheral position to another and also in the evaluated cross section. (author)

Cytomegalovirus infection induces a stem cell phenotype in human primary glioblastoma cells

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Fornara, O; Bartek, J; Rahbar, A

2016-01-01

Glioblastoma (GBM) is associated with poor prognosis despite aggressive surgical resection, chemotherapy, and radiation therapy. Unfortunately, this standard therapy does not target glioma cancer stem cells (GCSCs), a subpopulation of GBM cells that can give rise to recurrent tumors. GBMs express...... human cytomegalovirus (HCMV) proteins, and previously we found that the level of expression of HCMV immediate-early (IE) protein in GBMs is a prognostic factor for poor patient survival. In this study, we investigated the relation between HCMV infection of GBM cells and the presence of GCSCs. Primary...... GBMs were characterized by their expression of HCMV-IE and GCSCs marker CD133 and by patient survival. The extent to which HCMV infection of primary GBM cells induced a GCSC phenotype was evaluated in vitro. In primary GBMs, a large fraction of CD133-positive cells expressed HCMV-IE, and higher co...

Fiber mediated receptor masking in non-infected bystander cells restricts adenovirus cell killing effect but promotes adenovirus host co-existence.

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Johan Rebetz

Full Text Available The basic concept of conditionally replicating adenoviruses (CRAD as oncolytic agents is that progenies generated from each round of infection will disperse, infect and kill new cancer cells. However, CRAD has only inhibited, but not eradicated tumor growth in xenograft tumor therapy, and CRAD therapy has had only marginal clinical benefit to cancer patients. Here, we found that CRAD propagation and cancer cell survival co-existed for long periods of time when infection was initiated at low multiplicity of infection (MOI, and cancer cell killing was inefficient and slow compared to the assumed cell killing effect upon infection at high MOI. Excessive production of fiber molecules from initial CRAD infection of only 1 to 2% cancer cells and their release prior to the viral particle itself caused a tropism-specific receptor masking in both infected and non-infected bystander cells. Consequently, the non-infected bystander cells were inefficiently bound and infected by CRAD progenies. Further, fiber overproduction with concomitant restriction of adenovirus spread was observed in xenograft cancer therapy models. Besides the CAR-binding Ad4, Ad5, and Ad37, infection with CD46-binding Ad35 and Ad11 also caused receptor masking. Fiber overproduction and its resulting receptor masking thus play a key role in limiting CRAD functionality, but potentially promote adenovirus and host cell co-existence. These findings also give important clues for understanding mechanisms underlying the natural infection course of various adenoviruses.

Epstein-Barr Virus Type 2 Infects T Cells in Healthy Kenyan Children.

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Coleman, Carrie B; Daud, Ibrahim I; Ogolla, Sidney O; Ritchie, Julie A; Smith, Nicholas A; Sumba, Peter O; Dent, Arlene E; Rochford, Rosemary

2017-09-15

The 2 strains of Epstein-Barr virus (EBV), EBV type 1 (EBV-1) and EBV-2, differ in latency genes, suggesting that they use distinct mechanisms to establish latency. We previously reported that EBV-2 infects T cells in vitro. In this study, we tested the possibility that EBV-2 infects T cells in vivo. Purified T-cell fractions isolated from children positive for EBV-1 or EBV-2 and their mothers were examined for the presence of EBV and for EBV type. We detected EBV-2 in all T-cell samples obtained from EBV-2-infected children at 12 months of age, with some children retaining EBV-2-positive T cells through 24 months of age, suggesting that EBV-2 persists in T cells. We were unable to detect EBV-2 in T-cell samples from mothers but could detect EBV-2 in samples of their breast milk and saliva. These data suggest that EBV-2 uses T cells as an additional latency reservoir but that, over time, the frequency of infected T cells may drop below detectable levels. Alternatively, EBV-2 may establish a prolonged transient infection in the T-cell compartment. Collectively, these novel findings demonstrate that EBV-2 infects T cells in vivo and suggest EBV-2 may use the T-cell compartment to establish latency. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.

DRAM triggers lysosomal membrane permeabilization and cell death in CD4(+ T cells infected with HIV.

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Mireille Laforge

Full Text Available Productive HIV infection of CD4(+ T cells leads to a caspase-independent cell death pathway associated with lysosomal membrane permeabilization (LMP and cathepsin release, resulting in mitochondrial outer membrane permeabilization (MOMP. Herein, we demonstrate that HIV infection induces damage-regulated autophagy modulator (DRAM expression in a p53-dependent manner. Knocking down the expression of DRAM and p53 genes with specific siRNAs inhibited autophagy and LMP. However, inhibition of Atg5 and Beclin genes that prevents autophagy had a minor effect on LMP and cell death. The knock down of DRAM gene inhibited cytochrome C release, MOMP and cell death. However, knocking down DRAM, we increased viral infection and production. Our study shows for the first time the involvement of DRAM in host-pathogen interactions, which may represent a mechanism of defense via the elimination of infected cells.

Identification of proteins specific for human herpesvirus 6-infected human T cells

International Nuclear Information System (INIS)

Balachandran, N.; Amelse, R.E.; Zhou, W.W.; Chang, C.K.

1989-01-01

Proteins specific for human herpesvirus 6 (HHV-6)-infected human T cells (HSB-2) were examined by using polyclonal rabbit antibodies and monoclonal antibodies against HHV-6-infected cells and human sera. More than 20 proteins and six glycoproteins specific for HHV-6-infected cells were identified from [ 35 S]methionine- and [ 3 H]glucosamine-labeled total-cell extracts. Polyclonal rabbit antibodies immunoprecipitated 33 [ 35 S]methionine-labeled HHV-6-specific polypeptides with approximate molecular weights ranging from 180,000 to 31,000. In immunoprecipitation and Western immunoblot reactions, a patient's serum also recognized more than 30 HHV-6-specific proteins and seven glycoproteins. In contrast, sera from individuals with high-titered antibodies against other human herpesviruses reacted with fewer HHV-6-infected cell proteins, and only a 135,000-M r polypeptide was prominent. Monoclonal antibodies to HHV-6-infected cells reacted with single and multiple polypeptides specific for virus-infected cells and immunoprecipitated three distinct sets of glycoproteins, which were designated gp105k and gp82k, gp116k, gp64k, and gp54k, and gp102k

Identification of proteins specific for human herpesvirus 6-infected human T cells

International Nuclear Information System (INIS)

Balachandran, N.; Amelse, R.E.; Zhou, W.W.; Chang, C.K.

1989-01-01

Proteins specific for human herpesvirus 6 (HHV-6)-infected human T cells (HSB-2) were examined by using polyclonal rabbit antibodies and monoclonal antibodies against HHV-6-infected cells and human sera. More than 20 proteins and six glycoproteins specific for HHV-6-infected cells were identified from [ 35 S]methionine- and [ 3 H]glucosamine-labeled total-cell extracts. Polyclonal rabbit antibodies immunoprecipitated 33 [ 35 S]methionine-labeled HHV-6-specific polypeptides with approximate molecular weights ranging from 180,000 to 31,000. In immunoprecipitation and Western immunoblot reactions, a patient's serum also recognized more than 30 HHV-6-specific proteins and seven glycoproteins. In contrast, sera from individuals with high-titered antibodies against other human herpes viruses reacted with few HHV-6-infected cell proteins, and only a 135,000-M/sub r/ polypeptide was prominent. Monoclonal antibodies to HHV-6-infected cells reacted with single and multiple polypeptides specific for virus-infected cells and immunoprecipitated three distinct sets of glycoproteins, which were designated gp105K and gp92k, gp116k, gp64k, and gp54k, and gp102k

Altered T cell memory and effector cell development in chronic lymphatic filarial infection that is independent of persistent parasite antigen.

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Cathy Steel

2011-04-01

Full Text Available Chronic lymphatic filarial (LF infection is associated with suppression of parasite-specific T cell responses that persist even following elimination of infection. While several mechanisms have been implicated in mediating this T cell specific downregulation, a role for alterations in the homeostasis of T effector and memory cell populations has not been explored. Using multiparameter flow cytometry, we investigated the role of persistent filarial infection on the maintenance of T cell memory in patients from the filarial-endemic Cook Islands. Compared to filarial-uninfected endemic normals (EN, microfilaria (mf positive infected patients (Inf had a reduced CD4 central memory (T(CM compartment. In addition, Inf patients tended to have more effector memory cells (T(EM and fewer effector cells (T(EFF than did ENs giving significantly smaller T(EFF:T(EM ratios. These contracted T(CM and T(EFF populations were still evident in patients previously mf+ who had cleared their infection (CLInf. Moreover, the density of IL-7Rα, necessary for T memory cell maintenance (but decreased in T effector cells, was significantly higher on memory cells of Inf and CLInf patients, although there was no evidence for decreased IL-7 or increased soluble IL7-Rα, both possible mechanisms for signaling defects in memory cells. However, effector cells that were present in Inf and CLInf patients had lower percentages of HLA-DR suggesting impaired function. These changes in T cell populations appear to reflect chronicity of infection, as filarial-infected children, despite the presence of active infection, did not show alterations in the frequencies of these T cell phenotypes. These data indicate that filarial-infected patients have contracted T(CM compartments and a defect in effector cell development, defects that persist even following clearance of infection. The fact that these global changes in memory and effector cell compartments do not yet occur in infected children

Lymphocytes Negatively Regulate NK Cell Activity via Qa-1b following Viral Infection

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Haifeng C. Xu

2017-11-01

Full Text Available NK cells can reduce anti-viral T cell immunity during chronic viral infections, including infection with the lymphocytic choriomeningitis virus (LCMV. However, regulating factors that maintain the equilibrium between productive T cell and NK cell immunity are poorly understood. Here, we show that a large viral load resulted in inhibition of NK cell activation, which correlated with increased expression of Qa-1b, a ligand for inhibitory NK cell receptors. Qa-1b was predominantly upregulated on B cells following LCMV infection, and this upregulation was dependent on type I interferons. Absence of Qa-1b resulted in increased NK cell-mediated regulation of anti-viral T cells following viral infection. Consequently, anti-viral T cell immunity was reduced in Qa-1b- and NKG2A-deficient mice, resulting in increased viral replication and immunopathology. NK cell depletion restored anti-viral immunity and virus control in the absence of Qa-1b. Taken together, our findings indicate that lymphocytes limit NK cell activity during viral infection in order to promote anti-viral T cell immunity.

Activation of Natural Killer cells during microbial infections

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Amir eHorowitz

2012-01-01

Full Text Available Natural killer (NK cells are large granular lymphocytes that express a diverse array of germline encoded inhibitory and activating receptors for MHC Class I and Class I-like molecules, classical co-stimulatory ligands and cytokines. The ability of NK cells to be very rapidly activated by inflammatory cytokines, to secrete effector cytokines and to kill infected or stressed host cells, suggests that they may be among the very early responders during infection. Recent studies have also identified a small number of pathogen-derived ligands that can bind to NK cell surface receptors and directly induce their activation. Here we review recent studies that have begun to elucidate the various pathways by which viral, bacterial and parasite pathogens activate NK cells. We also consider two emerging themes of NK cell-pathogen interactions, namely their contribution to adaptive immune responses and their potential to take on regulatory and immunomodulatory functions.

Enhanced capacity of DNA repair in human cytomegalovirus-infected cells

International Nuclear Information System (INIS)

Nishiyama, Y.; Rapp, F.

1981-01-01

Plaque formation in Vero cells by UV-irradiated herpes simplex virus was enhanced by infection with human cytomegalovirus (HCMV), UV irradiation, or treatment with methylmethanesulfonate. Preinfection of Vero cells with HCMV enhanced reactivation of UV-irradiated herpes simplex virus more significantly than did treatment with UV or methylmethanesulfonate alone. A similar enhancement by HCMV was observed in human embryonic fibroblasts, but not in xeroderma pigmentosum (XP12BE) cells. It was also found that HCMV infection enhanced hydroxyurea-resistant DNA synthesis induced by UV light or methylmethanesulfonate. Alkaline sucrose gradient sedimentation analysis revealed an enhanced rate of synthesis of all size classes of DNA in UV-irradiated HCMV-infected Vero cells. However, HCMV infection did not induce repairable lesions in cellular DNA and did not significantly inhibit host cell DNA synthesis, unlike UV or methylmethanesulfonate. These results indicate that HCMV enhanced DNA repair capacity in the host cells without producing detectable lesions in cellular DNA and without inhibiting DNA synthesis. This repair appeared to be error proof for UV-damaged herpes simplex virus DNA when tested with herpes simplex virus thymidine kinase-negative mutants

Influenza H5N1 virus infection of polarized human alveolar epithelial cells and lung microvascular endothelial cells

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Yuen Kit M

2009-10-01

Full Text Available Abstract Background Highly pathogenic avian influenza (HPAI H5N1 virus is entrenched in poultry in Asia and Africa and continues to infect humans zoonotically causing acute respiratory disease syndrome and death. There is evidence that the virus may sometimes spread beyond respiratory tract to cause disseminated infection. The primary target cell for HPAI H5N1 virus in human lung is the alveolar epithelial cell. Alveolar epithelium and its adjacent lung microvascular endothelium form host barriers to the initiation of infection and dissemination of influenza H5N1 infection in humans. These are polarized cells and the polarity of influenza virus entry and egress as well as the secretion of cytokines and chemokines from the virus infected cells are likely to be central to the pathogenesis of human H5N1 disease. Aim To study influenza A (H5N1 virus replication and host innate immune responses in polarized primary human alveolar epithelial cells and lung microvascular endothelial cells and its relevance to the pathogenesis of human H5N1 disease. Methods We use an in vitro model of polarized primary human alveolar epithelial cells and lung microvascular endothelial cells grown in transwell culture inserts to compare infection with influenza A subtype H1N1 and H5N1 viruses via the apical or basolateral surfaces. Results We demonstrate that both influenza H1N1 and H5N1 viruses efficiently infect alveolar epithelial cells from both apical and basolateral surface of the epithelium but release of newly formed virus is mainly from the apical side of the epithelium. In contrast, influenza H5N1 virus, but not H1N1 virus, efficiently infected polarized microvascular endothelial cells from both apical and basolateral aspects. This provides a mechanistic explanation for how H5N1 virus may infect the lung from systemic circulation. Epidemiological evidence has implicated ingestion of virus-contaminated foods as the source of infection in some instances and our

microRNA Response to Listeria monocytogenes Infection in Epithelial Cells

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Izar, Benjamin; Mannala, Gopala Krishna; Mraheil, Mobarak Abu; Chakraborty, Trinad; Hain, Torsten

2012-01-01

microRNAs represent a family of very small non-coding RNAs that control several physiologic and pathologic processes, including host immune response and cancer by antagonizing a number of target mRNAs. There is limited knowledge about cell expression and the regulatory role of microRNAs following bacterial infections. We investigated whether infection with a Gram-positive bacterium leads to altered expression of microRNAs involved in the host cell response in epithelial cells. Caco-2 cells were infected with Listeria monocytogenes EGD-e, a mutant strain (ΔinlAB or Δhly) or incubated with purified listeriolysin (LLO). Total RNA was isolated and microRNA and target gene expression was compared to the expression in non-infected cells using microRNA microarrays and qRT-PCR. We identified and validated five microRNAs (miR- 146b, miR-16, let-7a1, miR-145 and miR-155) that were significantly deregulated following listerial infection. We show that expression patterns of particular microRNAs strongly depend on pathogen localization and the presence of bacterial effector proteins. Strikingly, miR-155 which was shown to have an important role in inflammatory responses during infection was induced by wild-type bacteria, by LLO-deficient bacteria and following incubation with purified LLO. It was downregulated following ΔinlAB infection indicating a new potent role for internalins in listerial pathogenicity and miRNA regulation. Concurrently, we observed differences in target transcript expression of the investigated miRNAs. We provide first evidence that L. monocytogenes infection leads to deregulation of a set of microRNAs with important roles in host response. Distinct microRNA expression depends on both LLO and pathogen localization. PMID:22312311

Regulatory NK cells mediated between immunosuppressive monocytes and dysfunctional T cells in chronic HBV infection.

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Li, Haijun; Zhai, Naicui; Wang, Zhongfeng; Song, Hongxiao; Yang, Yang; Cui, An; Li, Tianyang; Wang, Guangyi; Niu, Junqi; Crispe, Ian Nicholas; Su, Lishan; Tu, Zhengkun

2017-09-12

HBV infection represents a major health problem worldwide, but the immunological mechanisms by which HBV causes chronic persistent infection remain only partly understood. Recently, cell subsets with suppressive features have been recognised among monocytes and natural killer (NK) cells. Here we examine the effects of HBV on monocytes and NK cells. Monocytes and NK cells derived from chronic HBV-infected patients and healthy controls were purified and characterised for phenotype, gene expression and cytokines secretion by flow cytometry, quantitative real-time (qRT)-PCR, ELISA and western blotting. Culture and coculture of monocytes and NK cells were used to determine NK cell activation, using intracellular cytokines staining. In chronic HBV infection, monocytes express higher levels of PD-L1, HLA-E, interleukin (IL)-10 and TGF-β, and NK cells express higher levels of PD-1, CD94 and IL-10, compared with healthy individuals. HBV employs hepatitis B surface antigen (HBsAg) to induce suppressive monocytes with HLA-E, PD-L1, IL-10 and TGF-β expression via the MyD88/NFκB signalling pathway. HBV-treated monocytes induce NK cells to produce IL-10, via PD-L1 and HLA-E signals. Such NK cells inhibit autologous T cell activation. Our findings reveal an immunosuppressive cascade, in which HBV generates suppressive monocytes, which initiate regulatory NK cells differentiation resulting in T cell inhibition. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

Anaplasma phagocytophilum Manipulates Host Cell Apoptosis by Different Mechanisms to Establish Infection

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Pilar Alberdi

2016-07-01

Full Text Available Anaplasma phagocytophilum is an emerging zoonotic pathogen that causes human and animal granulocytic anaplasmosis and tick-borne fever of ruminants. This obligate intracellular bacterium evolved to use common strategies to establish infection in both vertebrate hosts and tick vectors. Herein, we discuss the different strategies used by the pathogen to modulate cell apoptosis and establish infection in host cells. In vertebrate neutrophils and human promyelocytic cells HL-60, both pro-apoptotic and anti-apoptotic factors have been reported. Tissue-specific differences in tick response to infection and differential regulation of apoptosis pathways have been observed in adult female midguts and salivary glands in response to infection with A. phagocytophilum. In tick midguts, pathogen inhibits apoptosis through the Janus kinase/signal transducers and activators of transcription (JAK/STAT pathway, while in salivary glands, the intrinsic apoptosis pathways is inhibited but tick cells respond with the activation of the extrinsic apoptosis pathway. In Ixodes scapularis ISE6 cells, bacterial infection down-regulates mitochondrial porin and manipulates protein processing in the endoplasmic reticulum and cell glucose metabolism to inhibit apoptosis and facilitate infection, whereas in IRE/CTVM20 tick cells, inhibition of apoptosis appears to be regulated by lower caspase levels. These results suggest that A. phagocytophilum uses different mechanisms to inhibit apoptosis for infection of both vertebrate and invertebrate hosts.

HIV-1-infected monocyte-derived dendritic cells do not undergo maturation but can elicit IL-10 production and T cell regulation

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Granelli-Piperno, Angela; Golebiowska, Angelika; Trumpfheller, Christine; Siegal, Frederick P.; Steinman, Ralph M.

2004-05-01

Dendritic cells (DCs) undergo maturation during virus infection and thereby become potent stimulators of cell-mediated immunity. HIV-1 replicates in immature DCs, but we now find that infection is not accompanied by many components of maturation in either infected cells or uninfected bystanders. The infected cultures do not develop potent stimulating activity for the mixed leukocyte reaction (MLR), and the DCs producing HIV-1 gag p24 do not express CD83 and DC-lysosome-associated membrane protein maturation markers. If different maturation stimuli are applied to DCs infected with HIV-1, the infected cells selectively fail to mature. When DCs from HIV-1-infected patients are infected and cultured with autologous T cells, IL-10 was produced in 6 of 10 patients. These DC-T cell cocultures could suppress another immune response, the MLR. The regulation was partially IL-10-dependent and correlated in extent with the level of IL-10 produced. Suppressor cells only developed from infected patients, rather than healthy controls, and the DCs had to be exposed to live virus rather than HIV-1 gag peptides or protein. These results indicate that HIV-1-infected DCs have two previously unrecognized means to evade immune responses: maturation can be blocked reducing the efficacy of antigen presentation from infected cells, and T cell-dependent suppression can be induced.

SAMHD1 controls cell cycle status, apoptosis and HIV-1 infection in monocytic THP-1 cells

International Nuclear Information System (INIS)

Bonifati, Serena; Daly, Michele B.; St Gelais, Corine; Kim, Sun Hee; Hollenbaugh, Joseph A.; Shepard, Caitlin; Kennedy, Edward M.; Kim, Dong-Hyun; Schinazi, Raymond F.; Kim, Baek; Wu, Li

2016-01-01

SAMHD1 limits HIV-1 infection in non-dividing myeloid cells by decreasing intracellular dNTP pools. HIV-1 restriction by SAMHD1 in these cells likely prevents activation of antiviral immune responses and modulates viral pathogenesis, thus highlighting a critical role of SAMHD1 in HIV-1 physiopathology. Here, we explored the function of SAMHD1 in regulating cell proliferation, cell cycle progression and apoptosis in monocytic THP-1 cells. Using the CRISPR/Cas9 technology, we generated THP-1 cells with stable SAMHD1 knockout. We found that silencing of SAMHD1 in cycling cells stimulates cell proliferation, redistributes cell cycle population in the G_1/G_0 phase and reduces apoptosis. These alterations correlated with increased dNTP levels and more efficient HIV-1 infection in dividing SAMHD1 knockout cells relative to control. Our results suggest that SAMHD1, through its dNTPase activity, affects cell proliferation, cell cycle distribution and apoptosis, and emphasize a key role of SAMHD1 in the interplay between cell cycle regulation and HIV-1 infection.

SAMHD1 controls cell cycle status, apoptosis and HIV-1 infection in monocytic THP-1 cells

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Bonifati, Serena [Center for Retrovirus Research, Department of Veterinary Biosciences, The Ohio State University, Columbus, OH (United States); Daly, Michele B. [Center for Drug Discovery, Department of Pediatrics, School of Medicine, Emory University, Atlanta, GA (United States); St Gelais, Corine; Kim, Sun Hee [Center for Retrovirus Research, Department of Veterinary Biosciences, The Ohio State University, Columbus, OH (United States); Hollenbaugh, Joseph A.; Shepard, Caitlin [Center for Drug Discovery, Department of Pediatrics, School of Medicine, Emory University, Atlanta, GA (United States); Kennedy, Edward M. [Department of Molecular Genetics and Microbiology, Duke University, Durham, NC (United States); Kim, Dong-Hyun [Department of Pharmacy, School of Pharmacy, Kyung-Hee University, Seoul (Korea, Republic of); Schinazi, Raymond F. [Center for Drug Discovery, Department of Pediatrics, School of Medicine, Emory University, Atlanta, GA (United States); Kim, Baek, E-mail: baek.kim@emory.edu [Center for Drug Discovery, Department of Pediatrics, School of Medicine, Emory University, Atlanta, GA (United States); Department of Pharmacy, School of Pharmacy, Kyung-Hee University, Seoul (Korea, Republic of); Wu, Li, E-mail: wu.840@osu.edu [Center for Retrovirus Research, Department of Veterinary Biosciences, The Ohio State University, Columbus, OH (United States)

2016-08-15

SAMHD1 limits HIV-1 infection in non-dividing myeloid cells by decreasing intracellular dNTP pools. HIV-1 restriction by SAMHD1 in these cells likely prevents activation of antiviral immune responses and modulates viral pathogenesis, thus highlighting a critical role of SAMHD1 in HIV-1 physiopathology. Here, we explored the function of SAMHD1 in regulating cell proliferation, cell cycle progression and apoptosis in monocytic THP-1 cells. Using the CRISPR/Cas9 technology, we generated THP-1 cells with stable SAMHD1 knockout. We found that silencing of SAMHD1 in cycling cells stimulates cell proliferation, redistributes cell cycle population in the G{sub 1}/G{sub 0} phase and reduces apoptosis. These alterations correlated with increased dNTP levels and more efficient HIV-1 infection in dividing SAMHD1 knockout cells relative to control. Our results suggest that SAMHD1, through its dNTPase activity, affects cell proliferation, cell cycle distribution and apoptosis, and emphasize a key role of SAMHD1 in the interplay between cell cycle regulation and HIV-1 infection.

Nipah virus infects specific subsets of porcine peripheral blood mononuclear cells.

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Beata Stachowiak

Full Text Available Nipah virus (NiV, a zoonotic paramyxovirus, is highly contagious in swine, and can cause fatal infections in humans following transmission from the swine host. The main viral targets in both species are the respiratory and central nervous systems, with viremia implicated as a mode of dissemination of NiV throughout the host. The presented work focused on the role of peripheral blood mononuclear cells (PBMC in the viremic spread of the virus in the swine host. B lymphocytes, CD4-CD8-, as well as CD4+CD8- T lymphocytes were not permissive to NiV, and expansion of the CD4+CD8- cells early post infection was consistent with functional humoral response to NiV infection observed in swine. In contrast, significant drop in the CD4+CD8- T cell frequency was observed in piglets which succumbed to the experimental infection, supporting the hypothesis that antibody development is the critical component of the protective immune response. Productive viral replication was detected in monocytes, CD6+CD8+ T lymphocytes and NK cells by recovery of infectious virus in the cell supernatants. Virus replication was supported by detection of the structural N and the non-structural C proteins or by detection of genomic RNA increase in the infected cells. Infection of T cells carrying CD6 marker, a strong ligand for the activated leukocyte cell adhesion molecule ALCAM (CD166 highly expressed on the microvascular endothelial cell of the blood-air and the blood-brain barrier may explain NiV preferential tropism for small blood vessels of the lung and brain.

Lipid Metabolism in Vascular Smooth Muscle Cells Infuenced by HCMV Infection

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Lingfang Li

2016-10-01

Full Text Available Background: The present study was designed to observe the infection of human cytomegalovirus (HCMV to human vascular smooth muscle cells (VSMCs, and the effect of viral infection on lipid metabolism in VSMCs. Methods: The cytopathic effects were observed by inverted microscopy and viral infection were examined by electron microscopy and RT-PCR. The lipid metabolism related gene profiling of VSMCs after HCMV infection was assayed by cDNA assay and the abnormal expression of genes were validated by quantitative RT-PCR. The content of cholesterol in VSMCs after HCMV infection was assayed by cholesterol detection kit. Results: VSMCs showed obvious cytopathic effects after HCMV infection. Intact viral particles could be detected in VSMCs using electron microscope. By use of RT-PCR technology, IE gene of HCMV could be amplified from VSMCs. The expression of cell lipid metabolism related gene profiling showed obvious disorders. The expression levels of HMG-CoA synthase and HMG-CoA reductase after infection increased significantly. The cellular cholesterol content (µmol/106 cells was significantly higher than that of mock infected group at 72h post infection. Conclusion: HCMV can infect VSMCs and the infection can affect cellular lipid metabolism related gene expression, which get involved in the occurrence and development of atherosclerosis (AS.

Milk Oligosaccharides Inhibit Human Rotavirus Infectivity in MA104 Cells.

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Laucirica, Daniel R; Triantis, Vassilis; Schoemaker, Ruud; Estes, Mary K; Ramani, Sasirekha

2017-09-01

Background: Oligosaccharides in milk act as soluble decoy receptors and prevent pathogen adhesion to the infant gut. Milk oligosaccharides reduce infectivity of a porcine rotavirus strain; however, the effects on human rotaviruses are less well understood. Objective: In this study, we determined the effect of specific and abundant milk oligosaccharides on the infectivity of 2 globally dominant human rotavirus strains. Methods: Four milk oligosaccharides-2'-fucosyllactose (2'FL), 3'-sialyllactose (3'SL), 6'-sialyllactose (6'SL), and galacto-oligosaccharides-were tested for their effects on the infectivity of human rotaviruses G1P[8] and G2P[4] through fluorescent focus assays on African green monkey kidney epithelial cells (MA104 cells). Oligosaccharides were added at different time points in the infectivity assays. Infections in the absence of oligosaccharides served as controls. Results: When compared with infections in the absence of glycans, all oligosaccharides substantially reduced the infectivity of both human rotavirus strains in vitro; however, virus strain-specific differences in effects were observed. Compared with control infections, the maximum reduction in G1P[8] infectivity was seen with 2'FL when added after the onset of infection (62% reduction, P rotaviruses in MA104 cells, primarily through an effect on the virus. Although breastfed infants are directly protected, the addition of specific oligosaccharides to infant formula may confer these benefits to formula-fed infants. © 2017 American Society for Nutrition.

Towards a global system of vigilance and surveillance in unrelated donors of haematopoietic progenitor cells for transplantation.

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Shaw, B E; Chapman, J; Fechter, M; Foeken, L; Greinix, H; Hwang, W; Phillips-Johnson, L; Korhonen, M; Lindberg, B; Navarro, W H; Szer, J

2013-11-01

Safety of living donors is critical to the success of blood, tissue and organ transplantation. Structured and robust vigilance and surveillance systems exist as part of some national entities, but historically no global systems are in place to ensure conformity, harmonisation and the recognition of rare adverse events (AEs). The World Health Assembly has recently resolved to require AE/reaction (AE/R) reporting both nationally and globally. The World Marrow Donor Association (WMDA) is an international organisation promoting the safety of unrelated donors and progenitor cell products for use in haematopoietic progenitor cell (HPC) transplantation. To address this issue, we established a system for collecting, collating, analysing, distributing and reacting to serious adverse events and reactions (SAE/R) in unrelated HPC donors. The WMDA successfully instituted this reporting system with 203 SAE/R reported in 2011. The committee generated two rapid reports, reacting to specific SAE/R, resulting in practice changing policies. The system has a robust governance structure, formal feedback to the WMDA membership and transparent information flows to other agencies, specialist physicians and transplant programs and the general public.

Gammaherpesvirus-driven plasma cell differentiation regulates virus reactivation from latently infected B lymphocytes.

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Xiaozhen Liang

2009-11-01

Full Text Available Gammaherpesviruses chronically infect their host and are tightly associated with the development of lymphoproliferative diseases and lymphomas, as well as several other types of cancer. Mechanisms involved in maintaining chronic gammaherpesvirus infections are poorly understood and, in particular, little is known about the mechanisms involved in controlling gammaherpesvirus reactivation from latently infected B cells in vivo. Recent evidence has linked plasma cell differentiation with reactivation of the human gammaherpesviruses EBV and KSHV through induction of the immediate-early viral transcriptional activators by the plasma cell-specific transcription factor XBP-1s. We now extend those findings to document a role for a gammaherpesvirus gene product in regulating plasma cell differentiation and thus virus reactivation. We have previously shown that the murine gammaherpesvirus 68 (MHV68 gene product M2 is dispensable for virus replication in permissive cells, but plays a critical role in virus reactivation from latently infected B cells. Here we show that in mice infected with wild type MHV68, virus infected plasma cells (ca. 8% of virus infected splenocytes at the peak of viral latency account for the majority of reactivation observed upon explant of splenocytes. In contrast, there is an absence of virus infected plasma cells at the peak of latency in mice infected with a M2 null MHV68. Furthermore, we show that the M2 protein can drive plasma cell differentiation in a B lymphoma cell line in the absence of any other MHV68 gene products. Thus, the role of M2 in MHV68 reactivation can be attributed to its ability to manipulate plasma cell differentiation, providing a novel viral strategy to regulate gammaherpesvirus reactivation from latently infected B cells. We postulate that M2 represents a new class of herpesvirus gene products (reactivation conditioners that do not directly participate in virus replication, but rather facilitate virus

Gammaherpesvirus-driven plasma cell differentiation regulates virus reactivation from latently infected B lymphocytes.

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Liang, Xiaozhen; Collins, Christopher M; Mendel, Justin B; Iwakoshi, Neal N; Speck, Samuel H

2009-11-01

Gammaherpesviruses chronically infect their host and are tightly associated with the development of lymphoproliferative diseases and lymphomas, as well as several other types of cancer. Mechanisms involved in maintaining chronic gammaherpesvirus infections are poorly understood and, in particular, little is known about the mechanisms involved in controlling gammaherpesvirus reactivation from latently infected B cells in vivo. Recent evidence has linked plasma cell differentiation with reactivation of the human gammaherpesviruses EBV and KSHV through induction of the immediate-early viral transcriptional activators by the plasma cell-specific transcription factor XBP-1s. We now extend those findings to document a role for a gammaherpesvirus gene product in regulating plasma cell differentiation and thus virus reactivation. We have previously shown that the murine gammaherpesvirus 68 (MHV68) gene product M2 is dispensable for virus replication in permissive cells, but plays a critical role in virus reactivation from latently infected B cells. Here we show that in mice infected with wild type MHV68, virus infected plasma cells (ca. 8% of virus infected splenocytes at the peak of viral latency) account for the majority of reactivation observed upon explant of splenocytes. In contrast, there is an absence of virus infected plasma cells at the peak of latency in mice infected with a M2 null MHV68. Furthermore, we show that the M2 protein can drive plasma cell differentiation in a B lymphoma cell line in the absence of any other MHV68 gene products. Thus, the role of M2 in MHV68 reactivation can be attributed to its ability to manipulate plasma cell differentiation, providing a novel viral strategy to regulate gammaherpesvirus reactivation from latently infected B cells. We postulate that M2 represents a new class of herpesvirus gene products (reactivation conditioners) that do not directly participate in virus replication, but rather facilitate virus reactivation by

Ag85-focused T-cell immune response controls Mycobacterium avium chronic infection.

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Bruno Cerqueira-Rodrigues

Full Text Available CD4+ T cells are essential players for the control of mycobacterial infections. Several mycobacterial antigens have been identified for eliciting a relevant CD4+ T cell mediated-immune response, and numerous studies explored this issue in the context of Mycobacterium tuberculosis infection. Antigen 85 (Ag85, a highly conserved protein across Mycobacterium species, is secreted at the early phase of M. tuberculosis infection leading to the proliferation of Ag85-specific CD4+ T cells. However, in the context of Mycobacterium avium infection, little is known about the expression of this antigen and the elicited immune response. In the current work, we investigated if a T cell receptor (TCR repertoire mostly, but not exclusively, directed at Ag85 is sufficient to mount a protective immune response against M. avium. We show that P25 mice, whose majority of T cells express a transgenic TCR specific for Ag85, control M. avium infection at the same level as wild type (WT mice up to 20 weeks post-infection (wpi. During M. avium infection, Ag85 antigen is easily detected in the liver of 20 wpi mice by immunohistochemistry. In spite of the propensity of P25 CD4+ T cells to produce higher amounts of interferon-gamma (IFNγ upon ex vivo stimulation, no differences in serum IFNγ levels are detected in P25 compared to WT mice, nor enhanced immunopathology is detected in P25 mice. These results indicate that a T cell response dominated by Ag85-specific T cells is appropriate to control M. avium infection with no signs of immunopathology.

Proteome profile of swine testicular cells infected with porcine transmissible gastroenteritis coronavirus.

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Ruili Ma

Full Text Available The interactions occurring between a virus and a host cell during a viral infection are complex. The purpose of this paper was to analyze altered cellular protein levels in porcine transmissible gastroenteritis coronavirus (TGEV-infected swine testicular (ST cells in order to determine potential virus-host interactions. A proteomic approach using isobaric tags for relative and absolute quantitation (iTRAQ-coupled two-dimensional liquid chromatography-tandem mass spectrometry identification was conducted on the TGEV-infected ST cells. The results showed that the 4-plex iTRAQ-based quantitative approach identified 4,112 proteins, 146 of which showed significant changes in expression 48 h after infection. At 64 h post infection, 219 of these proteins showed significant change, further indicating that a larger number of proteomic changes appear to occur during the later stages of infection. Gene ontology analysis of the altered proteins showed enrichment in multiple biological processes, including cell adhesion, response to stress, generation of precursor metabolites and energy, cell motility, protein complex assembly, growth, developmental maturation, immune system process, extracellular matrix organization, locomotion, cell-cell signaling, neurological system process, and cell junction organization. Changes in the expression levels of transforming growth factor beta 1 (TGF-β1, caspase-8, and heat shock protein 90 alpha (HSP90α were also verified by western blot analysis. To our knowledge, this study is the first time the response profile of ST host cells following TGEV infection has been analyzed using iTRAQ technology, and our description of the late proteomic changes that are occurring after the time of vigorous viral production are novel. Therefore, this study provides a solid foundation for further investigation, and will likely help us to better understand the mechanisms of TGEV infection and pathogenesis.

Bioleaching of Gold and Silver from Waste Printed Circuit Boards by Pseudomonas balearica SAE1 Isolated from an e-Waste Recycling Facility.

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Kumar, Anil; Saini, Harvinder Singh; Kumar, Sudhir

2018-02-01

Indigenous bacterial strain Pseudomonas balearica SAE1, tolerant to e-waste toxicity was isolated from an e-waste recycling facility Exigo Recycling Pvt. Ltd., India. Toxicity tolerance of bacterial strain was analyzed using crushed (particle size ≤150 µm) waste computer printed circuit boards (PCBs)/liter (L) of culture medium. The EC 50 value for SAE1 was 325.7 g/L of the e-waste pulp density. Two-step bioleaching was then applied to achieve the dissolution of gold (Au) and silver (Ag) from the e-waste. To maximize precious metal dissolution, factors including pulp density, glycine concentration, pH level, and temperature were optimized. The optimization resulted in 68.5 and 33.8% of Au and Ag dissolution, respectively, at a pH of 9.0, a pulp density of 10 g/L, a temperature of 30 °C, and a glycine concentration of 5 g/L. This is the first study of Au and Ag bioleaching using indigenous e-waste bacteria and its analysis to determine e-waste toxicity tolerance.

Effect of hydrogen charging on the stability of SAE 10B22 steel surface in alkaline solutions

International Nuclear Information System (INIS)

Modiano, S.; Carreno, J.A.; Fugivara, C.S.; Benedetti, A.V.; Mattos, O.R.

2005-01-01

The influence of hydrogen charging into a quenched and tempered boron steel membrane electrode (SAE 10B22) was studied using borate buffer (pH 8.4) and NaOH solutions (pH 12.7), with or without the addition of 0.01 M EDTA. At the hydrogen input side, hydrogen charging influenced cyclic voltammograms increasing the anodic charge of iron(II) hydroxide formation, and decreasing the donor density of passive films. These results suggest that the hydrogen ingress caused instability of metallic surface, increasing the surface area activity

Expansion of murine gammaherpesvirus latently infected B cells requires T follicular help.

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Christopher M Collins

2014-05-01

Full Text Available X linked lymphoproliferative disease (XLP is an inherited immunodeficiency resulting from mutations in the gene encoding the slam associated protein (SAP. One of the defining characteristics of XLP is extreme susceptibility to infection with Epstein-Barr virus (EBV, a gammaherpesvirus belonging to the genus Lymphocryptovirus, often resulting in fatal infectious mononucleosis (FIM. However, infection of SAP deficient mice with the related Murine gammaherpesvirus 68 (MHV68, a gammaherpesvirus in the genus Rhadinovirus, does not recapitulate XLP. Here we show that MHV68 inefficiently establishes latency in B cells in SAP deficient mice due to insufficient CD4 T cell help during the germinal center response. Although MHV68 infected B cells can be found in SAP-deficient mice, significantly fewer of these cells had a germinal center phenotype compared to SAP-sufficient mice. Furthermore, we show that infected germinal center B cells in SAP-deficient mice fail to proliferate. This failure to proliferate resulted in significantly lower viral loads, and likely accounts for the inability of MHV68 to induce a FIM-like syndrome. Finally, inhibiting differentiation of T follicular helper (TFH cells in SAP-sufficient C57Bl/6 mice resulted in decreased B cell latency, and the magnitude of the TFH response directly correlated with the level of infection in B cells. This requirement for CD4 T cell help during the germinal center reaction by MHV68 is in contrast with EBV, which is thought to be capable of bypassing this requirement by expressing viral proteins that mimic signals provided by TFH cells. In conclusion, the outcome of MHV68 infection in mice in the setting of loss of SAP function is distinct from that observed in SAP-deficient patients infected with EBV, and may identify a fundamental difference between the strategies employed by the rhadinoviruses and lymphocryptoviruses to expand B cell latency during the early phase of infection.

RNA expression in a cartilaginous fish cell line reveals ancient 3′ noncoding regions highly conserved in vertebrates

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Forest, David; Nishikawa, Ryuhei; Kobayashi, Hiroshi; Parton, Angela; Bayne, Christopher J.; Barnes, David W.

2007-01-01

We have established a cartilaginous fish cell line [Squalus acanthias embryo cell line (SAE)], a mesenchymal stem cell line derived from the embryo of an elasmobranch, the spiny dogfish shark S. acanthias. Elasmobranchs (sharks and rays) first appeared >400 million years ago, and existing species provide useful models for comparative vertebrate cell biology, physiology, and genomics. Comparative vertebrate genomics among evolutionarily distant organisms can provide sequence conservation information that facilitates identification of critical coding and noncoding regions. Although these genomic analyses are informative, experimental verification of functions of genomic sequences depends heavily on cell culture approaches. Using ESTs defining mRNAs derived from the SAE cell line, we identified lengthy and highly conserved gene-specific nucleotide sequences in the noncoding 3′ UTRs of eight genes involved in the regulation of cell growth and proliferation. Conserved noncoding 3′ mRNA regions detected by using the shark nucleotide sequences as a starting point were found in a range of other vertebrate orders, including bony fish, birds, amphibians, and mammals. Nucleotide identity of shark and human in these regions was remarkably well conserved. Our results indicate that highly conserved gene sequences dating from the appearance of jawed vertebrates and representing potential cis-regulatory elements can be identified through the use of cartilaginous fish as a baseline. Because the expression of genes in the SAE cell line was prerequisite for their identification, this cartilaginous fish culture system also provides a physiologically valid tool to test functional hypotheses on the role of these ancient conserved sequences in comparative cell biology. PMID:17227856

Listeria monocytogenes infection of HD11, chicken macrophage-like cells.

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Jarvis, N A; Donaldson, J R; O'Bryan, C A; Ricke, S C; Crandall, P G

2017-04-01

Listeria monocytogenes can be carried by and infect poultry, although the clinical disease in birds is rare. Escape from macrophage phagocytosis is a key step in pathogenesis for L. monocytogenes. Therefore, we investigated the infection of the chicken macrophage-like cell line HD11 with 2 strains of L. monocytogenes EGD-e and Scott A. After infection, L. monocytogenes was quantified by spread plating and HD11 was quantified with trypan blue exclusion stain before enumeration. The standard macrophage killing protocols require washing the cell monolayers 3 times with PBS, which was found to negatively influence HD11 monolayers. Maximum bacterial densities within macrophages were not different between the 2 Listeria strains. HD11 required more than 11 h to effectively reduce intracellular L. monocytogenes Scott A, and Scott A was more susceptible to HD11 killing than EGD-e. It appears that Listeria infection initially causes attenuation of HD11 growth, and infected HD11 cells do not begin to lyse until at least 11 h post infection. These results suggest that there are subtle strain to strain differences in response to HD11 macrophage phagocytosis. The long lead-time required for HD11 to kill L. monocytogenes cells means that there is sufficient time available for chicken macrophages to circulate in the blood and transfer the intracellular Listeria to multiple tissues. © 2016 Poultry Science Association Inc.

Dendritic cells as Achilles' heel and Trojan horse during varicella zoster virus infection

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Günther eSchönrich

2015-05-01

Full Text Available Varicella zoster virus (VZV, a human alphaherpesvirus, causes varicella and subsequently estab-lishes latency within sensory nerve ganglia. Later in life VZV can reactivate to cause herpes zoster. A reduced frequency of VZV-specific T cells is strongly associated with herpes zoster illustrating that these immune cells are central to control latency. Dendritic cells (DCs are required for the generation of VZV-specific T cells. However, DCs can also be infected in vitro and in vivo allowing VZV to evade the antiviral immune response. Thus, DCs represent the immune systems’ Achilles heel. Uniquely among the human herpesviruses, VZV infects both DCs and T cells, and exploits both as Trojan horses. During primary infection VZV-infected DCs traffic to the draining lymph nodes and tonsils, where the virus is transferred to T cells. VZV-infected T cells subsequently spread infection throughout the body to give the typical varicella skin rash. The delicate interplay between VZV and DCs and its consequences for viral immune evasion and viral dissemination will be discussed in this article.

An Essential Role for Coagulase in Staphylococcus aureus Biofilm Development Reveals New Therapeutic Possibilities for Device-Related Infections.

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Zapotoczna, Marta; McCarthy, Hannah; Rudkin, Justine K; O'Gara, James P; O'Neill, Eoghan

2015-12-15

High-level resistance to antimicrobial drugs is a major factor in the pathogenesis of chronic Staphylococcus aureus biofilm-associated, medical device-related infections. Antimicrobial susceptibility analysis revealed that biofilms grown for ≤ 24 hours on biomaterials conditioned with human plasma under venous shear in iron-free cell culture medium were significantly more susceptible to antistaphylococcal antibiotics. Biofilms formed under these physiologically relevant conditions were regulated by SaeRS and dependent on coagulase-catalyzed conversion of fibrinogen into fibrin. In contrast, SarA-regulated biofilms formed on uncoated polystyrene in nutrient-rich bacteriological medium were mediated by the previously characterized biofilm factors poly-N-acetyl glucosamine, fibronectin-binding proteins, or autolytic activity and were antibiotic resistant. Coagulase-mediated biofilms exhibited increased antimicrobial resistance over time (>48 hours) but were always susceptible to dispersal by the fibrinolytic enzymes plasmin or nattokinase. Biofilms recovered from infected central venous catheters in a rat model of device-related infection were dispersed by nattokinase, supporting the important role of the biofilm phenotype and identifying a potentially new therapeutic approach with antimicrobials and fibrinolytic drugs, particularly during the early stages of device-related infection. © The Author 2015. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Zika virus infection dysregulates human neural stem cell growth and inhibits differentiation into neuroprogenitor cells.

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Devhare, Pradip; Meyer, Keith; Steele, Robert; Ray, Ratna B; Ray, Ranjit

2017-10-12

The current outbreak of Zika virus-associated diseases in South America and its threat to spread to other parts of the world has emerged as a global health emergency. A strong link between Zika virus and microcephaly exists, and the potential mechanisms associated with microcephaly are under intense investigation. In this study, we evaluated the effect of Zika virus infection of Asian and African lineages (PRVABC59 and MR766) in human neural stem cells (hNSCs). These two Zika virus strains displayed distinct infection pattern and growth rates in hNSCs. Zika virus MR766 strain increased serine 139 phosphorylation of histone H2AX (γH2AX), a known early cellular response proteins to DNA damage. On the other hand, PRVABC59 strain upregulated serine 15 phosphorylation of p53, p21 and PUMA expression. MR766-infected cells displayed poly (ADP-ribose) polymerase (PARP) and caspase-3 cleavage. Interestingly, infection of hNSCs by both strains of Zika virus for 24 h, followed by incubation in astrocyte differentiation medium, induced rounding and cell death. However, astrocytes generated from hNSCs by incubation in differentiation medium when infected with Zika virus displayed minimal cytopathic effect at an early time point. Infected hNSCs incubated in astrocyte differentiating medium displayed PARP cleavage within 24-36 h. Together, these results showed that two distinct strains of Zika virus potentiate hNSC growth inhibition by different mechanisms, but both viruses strongly induce death in early differentiating neuroprogenitor cells even at a very low multiplicity of infection. Our observations demonstrate further mechanistic insights for impaired neuronal homeostasis during active Zika virus infection.

Zika virus infection dysregulates human neural stem cell growth and inhibits differentiation into neuroprogenitor cells

Science.gov (United States)

Devhare, Pradip; Meyer, Keith; Steele, Robert; Ray, Ratna B; Ray, Ranjit

2017-01-01

The current outbreak of Zika virus-associated diseases in South America and its threat to spread to other parts of the world has emerged as a global health emergency. A strong link between Zika virus and microcephaly exists, and the potential mechanisms associated with microcephaly are under intense investigation. In this study, we evaluated the effect of Zika virus infection of Asian and African lineages (PRVABC59 and MR766) in human neural stem cells (hNSCs). These two Zika virus strains displayed distinct infection pattern and growth rates in hNSCs. Zika virus MR766 strain increased serine 139 phosphorylation of histone H2AX (γH2AX), a known early cellular response proteins to DNA damage. On the other hand, PRVABC59 strain upregulated serine 15 phosphorylation of p53, p21 and PUMA expression. MR766-infected cells displayed poly (ADP-ribose) polymerase (PARP) and caspase-3 cleavage. Interestingly, infection of hNSCs by both strains of Zika virus for 24 h, followed by incubation in astrocyte differentiation medium, induced rounding and cell death. However, astrocytes generated from hNSCs by incubation in differentiation medium when infected with Zika virus displayed minimal cytopathic effect at an early time point. Infected hNSCs incubated in astrocyte differentiating medium displayed PARP cleavage within 24–36 h. Together, these results showed that two distinct strains of Zika virus potentiate hNSC growth inhibition by different mechanisms, but both viruses strongly induce death in early differentiating neuroprogenitor cells even at a very low multiplicity of infection. Our observations demonstrate further mechanistic insights for impaired neuronal homeostasis during active Zika virus infection. PMID:29022904

Plasmacytoid dendritic cells are crucial in Bifidobacterium adolescentis-mediated inhibition of Yersinia enterocolitica infection.

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Alexandra Wittmann

Full Text Available In industrialized countries bacterial intestinal infections are commonly caused by enteropathogenic Enterobacteriaceae. The interaction of the microbiota with the host immune system determines the adequacy of an appropriate response against pathogens. In this study we addressed whether the probiotic Bifidobacterium adolescentis is protective during intestinal Yersinia enterocolitica infection. Female C57BL/6 mice were fed with B. adolescentis, infected with Yersinia enterocolitica, or B. adolescentis fed and subsequently infected with Yersinia enterocolitica. B. adolescentis fed and Yersinia infected mice were protected from Yersinia infection as indicated by a significantly reduced weight loss and splenic Yersinia load when compared to Yersinia infected mice. Moreover, protection from infection was associated with increased intestinal plasmacytoid dendritic cell and regulatory T-cell frequencies. Plasmacytoid dendritic cell function was investigated using depletion experiments by injecting B. adolescentis fed, Yersinia infected C57BL/6 mice with anti-mouse PDCA-1 antibody, to deplete plasmacytoid dendritic cells, or respective isotype control. The B. adolescentis-mediated protection from Yersinia dissemination to the spleen was abrogated after plasmacytoid dendritic cell depletion indicating a crucial function for pDC in control of intestinal Yersinia infection. We suggest that feeding of B. adolescentis modulates the intestinal immune system in terms of increased plasmacytoid dendritic cell and regulatory T-cell frequencies, which might account for the B. adolescentis-mediated protection from Yersinia enterocolitica infection.

Plasmacytoid dendritic cells are crucial in Bifidobacterium adolescentis-mediated inhibition of Yersinia enterocolitica infection.

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Wittmann, Alexandra; Autenrieth, Ingo B; Frick, Julia-Stefanie

2013-01-01

In industrialized countries bacterial intestinal infections are commonly caused by enteropathogenic Enterobacteriaceae. The interaction of the microbiota with the host immune system determines the adequacy of an appropriate response against pathogens. In this study we addressed whether the probiotic Bifidobacterium adolescentis is protective during intestinal Yersinia enterocolitica infection. Female C57BL/6 mice were fed with B. adolescentis, infected with Yersinia enterocolitica, or B. adolescentis fed and subsequently infected with Yersinia enterocolitica. B. adolescentis fed and Yersinia infected mice were protected from Yersinia infection as indicated by a significantly reduced weight loss and splenic Yersinia load when compared to Yersinia infected mice. Moreover, protection from infection was associated with increased intestinal plasmacytoid dendritic cell and regulatory T-cell frequencies. Plasmacytoid dendritic cell function was investigated using depletion experiments by injecting B. adolescentis fed, Yersinia infected C57BL/6 mice with anti-mouse PDCA-1 antibody, to deplete plasmacytoid dendritic cells, or respective isotype control. The B. adolescentis-mediated protection from Yersinia dissemination to the spleen was abrogated after plasmacytoid dendritic cell depletion indicating a crucial function for pDC in control of intestinal Yersinia infection. We suggest that feeding of B. adolescentis modulates the intestinal immune system in terms of increased plasmacytoid dendritic cell and regulatory T-cell frequencies, which might account for the B. adolescentis-mediated protection from Yersinia enterocolitica infection.

Lysis of cells infected with typhus group rickettsiae by a human cytotoxic T cell clone

International Nuclear Information System (INIS)

Carl, M.; Robbins, F.; Hartzman, R.J.; Dasch, G.A.

1987-01-01

Cytolytic human T cells clones generated in response to the intracellular bacterium Rickettsia typhi were characterized. Growing clones were tested for their ability to proliferate specifically in response to antigens derived from typhus group rickettsiae or to lyse targets infected with R. typhi or Rickettsia prowazekii, as measured by 51 Cr-release from target cells. Two clones were able to lyse targets infected with typhus group rickettsiae. One of these clones was more fully characterized because of its rapid growth characteristics. This cytolytic clone was capable of lysing an autologous infected target as well as a target matched for class I and II histocompatibility leukocyte antigens (HLA). It was not capable, however, of lysing either a target mismatched for both class I and II HLA or a target partially matched for class I HLA. In addition, the clone exhibited specificity in that it was able to lyse an autologous target infected with typhus group rickettsiae, but did not lyse an autologous target infected with an antigenically distinct rickettsial species, Rickettsia tsutsugamushi. These results demonstrate, for the first time, that cells infected with intracellular bacteria can be lysed by human cytotoxic T lymphocytes

Zika virus infection of Hofbauer cells.

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Simoni, Michael K; Jurado, Kellie Ann; Abrahams, Vikki M; Fikrig, Erol; Guller, Seth

2017-02-01

Recent studies have linked antenatal infection with Zika virus (ZIKV) with major adverse fetal and neonatal outcomes, including microcephaly. There is a growing consensus for the existence of a congenital Zika syndrome (CZS). Previous studies have indicated that non-placental macrophages play a key role in the replication of dengue virus (DENV), a closely related flavivirus. As the placenta provides the conduit for vertical transmission of certain viruses, and placental Hofbauer cells (HBCs) are fetal-placental macrophages located adjacent to fetal capillaries, it is not surprising that several recent studies have examined infection of HBCs by ZIKV. In this review, we describe congenital abnormalities associated with ZIKV infection, the role of HBCs in the placental response to infection, and evidence for the susceptibility of HBCs to ZIKV infection. We conclude that HBCs may contribute to the spread of ZIKV in placenta and promote vertical transmission of ZIKV, ultimately compromising fetal and neonatal development and function. Current evidence strongly suggests that further studies are warranted to dissect the specific molecular mechanism through which ZIKV infects HBCs and its potential impact on the development of CZS. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Molecular Responses of Human Retinal Cells to Infection with Dengue Virus.

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Carr, Jillian M; Ashander, Liam M; Calvert, Julie K; Ma, Yuefang; Aloia, Amanda; Bracho, Gustavo G; Chee, Soon-Phaik; Appukuttan, Binoy; Smith, Justine R

2017-01-01

Recent clinical reports indicate that infection with dengue virus (DENV) commonly has ocular manifestations. The most serious threat to vision is dengue retinopathy, including retinal vasculopathy and macular edema. Mechanisms of retinopathy are unstudied, but observations in patients implicate retinal pigment epithelial cells and retinal endothelial cells. Human retinal cells were inoculated with DENV-2 and monitored for up to 72 hours. Epithelial and endothelial cells supported DENV replication and release, but epithelial cells alone demonstrated clear cytopathic effect, and infection was more productive in those cells. Infection induced type I interferon responses from both cells, but this was stronger in epithelial cells. Endothelial cells increased expression of adhesion molecules, with sustained overexpression of vascular adhesion molecule-1. Transcellular impedance decreased for epithelial monolayers, but not endothelial monolayers, coinciding with cytopathic effect. This reduction was accompanied by disorganization of intracellular filamentous-actin and decreased expression of junctional molecules, zonula occludens 1, and catenin- β 1. Changes in endothelial expression of adhesion molecules are consistent with the retinal vasculopathy seen in patients infected with DENV; decreases in epithelial junctional protein expression, paralleling loss of integrity of the epithelium, provide a molecular basis for DENV-associated macular edema. These molecular processes present potential therapeutic targets for vision-threatening dengue retinopathy.

Molecular Responses of Human Retinal Cells to Infection with Dengue Virus

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Jillian M. Carr

2017-01-01

Full Text Available Recent clinical reports indicate that infection with dengue virus (DENV commonly has ocular manifestations. The most serious threat to vision is dengue retinopathy, including retinal vasculopathy and macular edema. Mechanisms of retinopathy are unstudied, but observations in patients implicate retinal pigment epithelial cells and retinal endothelial cells. Human retinal cells were inoculated with DENV-2 and monitored for up to 72 hours. Epithelial and endothelial cells supported DENV replication and release, but epithelial cells alone demonstrated clear cytopathic effect, and infection was more productive in those cells. Infection induced type I interferon responses from both cells, but this was stronger in epithelial cells. Endothelial cells increased expression of adhesion molecules, with sustained overexpression of vascular adhesion molecule-1. Transcellular impedance decreased for epithelial monolayers, but not endothelial monolayers, coinciding with cytopathic effect. This reduction was accompanied by disorganization of intracellular filamentous-actin and decreased expression of junctional molecules, zonula occludens 1, and catenin-β1. Changes in endothelial expression of adhesion molecules are consistent with the retinal vasculopathy seen in patients infected with DENV; decreases in epithelial junctional protein expression, paralleling loss of integrity of the epithelium, provide a molecular basis for DENV-associated macular edema. These molecular processes present potential therapeutic targets for vision-threatening dengue retinopathy.

PKR Activation Favors Infectious Pancreatic Necrosis Virus Replication in Infected Cells

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Amr A.A. Gamil

2016-06-01

Full Text Available The double-stranded RNA-activated protein kinase R (PKR is a Type I interferon (IFN stimulated gene that has important biological and immunological functions. In viral infections, in general, PKR inhibits or promotes viral replication, but PKR-IPNV interaction has not been previously studied. We investigated the involvement of PKR during infectious pancreatic necrosis virus (IPNV infection using a custom-made rabbit antiserum and the PKR inhibitor C16. Reactivity of the antiserum to PKR in CHSE-214 cells was confirmed after IFNα treatment giving an increased protein level. IPNV infection alone did not give increased PKR levels by Western blot, while pre-treatment with PKR inhibitor before IPNV infection gave decreased eukaryotic initiation factor 2-alpha (eIF2α phosphorylation. This suggests that PKR, despite not being upregulated, is involved in eIF2α phosphorylation during IPNV infection. PKR inhibitor pre-treatment resulted in decreased virus titers, extra- and intracellularly, concomitant with reduction of cells with compromised membranes in IPNV-permissive cell lines. These findings suggest that IPNV uses PKR activation to promote virus replication in infected cells.

Longitudinal Analysis of Natural Killer Cells in Dengue Virus-Infected Patients in Comparison to Chikungunya and Chikungunya/Dengue Virus-Infected Patients.

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Caroline Petitdemange

2016-03-01

Full Text Available Dengue virus (DENV is the most prominent arbovirus worldwide, causing major epidemics in South-East Asia, South America and Africa. In 2010, a major DENV-2 outbreak occurred in Gabon with cases of patients co-infected with chikungunya virus (CHIKV. Although the innate immune response is thought to be of primordial importance in the development and outcome of arbovirus-associated pathologies, our knowledge of the role of natural killer (NK cells during DENV-2 infection is in its infancy.We performed the first extensive comparative longitudinal characterization of NK cells in patients infected by DENV-2, CHIKV or both viruses. Hierarchical clustering and principal component analyses were performed to discriminate between CHIKV and DENV-2 infected patients.We observed that both activation and differentiation of NK cells are induced during the acute phase of infection by DENV-2 and CHIKV. Combinatorial analysis however, revealed that both arboviruses induced two different signatures of NK-cell responses, with CHIKV more associated with terminal differentiation, and DENV-2 with inhibitory KIRs. We show also that intracellular production of interferon-γ (IFN-γ by NK cells is strongly stimulated in acute DENV-2 infection, compared to CHIKV.Although specific differences were observed between CHIKV and DENV-2 infections, the significant remodeling of NK cell populations observed here suggests their potential roles in the control of both infections.

B cell follicle sanctuary permits persistent productive simian immunodeficiency virus infection in elite controllers.

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Fukazawa, Yoshinori; Lum, Richard; Okoye, Afam A; Park, Haesun; Matsuda, Kenta; Bae, Jin Young; Hagen, Shoko I; Shoemaker, Rebecca; Deleage, Claire; Lucero, Carissa; Morcock, David; Swanson, Tonya; Legasse, Alfred W; Axthelm, Michael K; Hesselgesser, Joseph; Geleziunas, Romas; Hirsch, Vanessa M; Edlefsen, Paul T; Piatak, Michael; Estes, Jacob D; Lifson, Jeffrey D; Picker, Louis J

2015-02-01

Chronic-phase HIV and simian immunodeficiency virus (SIV) replication is reduced by as much as 10,000-fold in elite controllers (ECs) compared with typical progressors (TPs), but sufficient viral replication persists in EC tissues to allow viral sequence evolution and induce excess immune activation. Here we show that productive SIV infection in rhesus monkey ECs, but not TPs, is markedly restricted to CD4(+) follicular helper T (TFH) cells, suggesting that these EC monkeys' highly effective SIV-specific CD8(+) T cells can effectively clear productive SIV infection from extrafollicular sites, but their relative exclusion from B cell follicles prevents their elimination of productively infected TFH cells. CD8(+) lymphocyte depletion in EC monkeys resulted in a dramatic re-distribution of productive SIV infection to non-TFH cells, with restriction of productive infection to TFH cells resuming upon CD8(+) T cell recovery. Thus, B cell follicles constitute 'sanctuaries' for persistent SIV replication in the presence of potent anti-viral CD8(+) T cell responses, potentially complicating efforts to cure HIV infection with therapeutic vaccination or T cell immunotherapy.

Tumor necrosis factor alpha selectively sensitizes human immunodeficiency virus-infected cells to heat and radiation

International Nuclear Information System (INIS)

Wong, G.H.; McHugh, T.; Weber, R.; Goeddel, D.V.

1991-01-01

We report here that infection of the human T-cell line HUT-78 with human immunodeficiency virus (HIV) increases its sensitivity to heat and radiation toxicity. A possible explanation for this result may be the reduced expression of manganous superoxide dismutase (MnSOD) in HIV-infected cells compared to uninfected cells. Tumor necrosis factor alpha (TNF-alpha) further sensitizes HIV-infected cells but not uninfected cells to heat and radiation. This is consistent with the ability of TNF-alpha to induce the expression of MnSOD in uninfected but not in HIV-infected cells. HIV-infected HUT-78 cell lines engineered to overexpress MnSOD are more resistant to heat and radiation than HIV-infected cells that do not overexpress MnSOD. However, treatment with TNF-alpha still sensitizes these cells to heat and radiation

Cell-mediated immunity to herpes simplex in humans: lymphocyte cytotoxicity measured by 51Cr release from infected cells

International Nuclear Information System (INIS)

Russell, A.S.; Percy, J.S.; Kovithavongs, T.

1975-01-01

We assessed cell-mediated immunity to herpes simplex virus type 1 antigen in patients suffering from recurrent cold sores and in a series of healthy controls. Paradoxically, all those subject to recurrent herpetic infections had, without exception, evidence of cell-mediated immunity to herpes antigens. This was demonstrated by lymphocyte transformation and specific 51 Cr release from infected human amnion cells after incubation with peripheral blood mononuclear cells. Where performed, skin tests with herpes antigen were also positive. In addition, serum from these patients specifically sensitized herpes virus-infected cells to killing by nonimmune, control mononuclear cells. These tests were negative in the control patients except in a few cases, and it is suggested that these latter may be the asymptomatic herpes virus carriers previously recognized or that they may have experienced a genital infection. (U.S.)

A bovine cell line that can be infected by natural sheep scrapie prions.

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Anja M Oelschlegel

Full Text Available Cell culture systems represent a crucial part in basic prion research; yet, cell lines that are susceptible to prions, especially to field isolated prions that were not adapted to rodents, are very rare. The purpose of this study was to identify and characterize a cell line that was susceptible to ruminant-derived prions and to establish a stable prion infection within it. Based on species and tissue of origin as well as PrP expression rate, we pre-selected a total of 33 cell lines that were then challenged with natural and with mouse propagated BSE or scrapie inocula. Here, we report the successful infection of a non-transgenic bovine cell line, a sub-line of the bovine kidney cell line MDBK, with natural sheep scrapie prions. This cell line retained the scrapie infection for more than 200 passages. Selective cloning resulted in cell populations with increased accumulation of PrPres, although this treatment was not mandatory for retaining the infection. The infection remained stable, even under suboptimal culture conditions. The resulting infectivity of the cells was confirmed by mouse bioassay (Tgbov mice, Tgshp mice. We believe that PES cells used together with other prion permissive cell lines will prove a valuable tool for ongoing efforts to understand and defeat prions and prion diseases.

Target cell cyclophilins facilitate human papillomavirus type 16 infection.

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Bienkowska-Haba, Malgorzata; Patel, Hetalkumar D; Sapp, Martin

2009-07-01

Following attachment to primary receptor heparan sulfate proteoglycans (HSPG), human papillomavirus type 16 (HPV16) particles undergo conformational changes affecting the major and minor capsid proteins, L1 and L2, respectively. This results in exposure of the L2 N-terminus, transfer to uptake receptors, and infectious internalization. Here, we report that target cell cyclophilins, peptidyl-prolyl cis/trans isomerases, are required for efficient HPV16 infection. Cell surface cyclophilin B (CyPB) facilitates conformational changes in capsid proteins, resulting in exposure of the L2 N-terminus. Inhibition of CyPB blocked HPV16 infection by inducing noninfectious internalization. Mutation of a putative CyP binding site present in HPV16 L2 yielded exposed L2 N-terminus in the absence of active CyP and bypassed the need for cell surface CyPB. However, this mutant was still sensitive to CyP inhibition and required CyP for completion of infection, probably after internalization. Taken together, these data suggest that CyP is required during two distinct steps of HPV16 infection. Identification of cell surface CyPB will facilitate the study of the complex events preceding internalization and adds a putative drug target for prevention of HPV-induced diseases.

Target cell cyclophilins facilitate human papillomavirus type 16 infection.

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Malgorzata Bienkowska-Haba

2009-07-01

Full Text Available Following attachment to primary receptor heparan sulfate proteoglycans (HSPG, human papillomavirus type 16 (HPV16 particles undergo conformational changes affecting the major and minor capsid proteins, L1 and L2, respectively. This results in exposure of the L2 N-terminus, transfer to uptake receptors, and infectious internalization. Here, we report that target cell cyclophilins, peptidyl-prolyl cis/trans isomerases, are required for efficient HPV16 infection. Cell surface cyclophilin B (CyPB facilitates conformational changes in capsid proteins, resulting in exposure of the L2 N-terminus. Inhibition of CyPB blocked HPV16 infection by inducing noninfectious internalization. Mutation of a putative CyP binding site present in HPV16 L2 yielded exposed L2 N-terminus in the absence of active CyP and bypassed the need for cell surface CyPB. However, this mutant was still sensitive to CyP inhibition and required CyP for completion of infection, probably after internalization. Taken together, these data suggest that CyP is required during two distinct steps of HPV16 infection. Identification of cell surface CyPB will facilitate the study of the complex events preceding internalization and adds a putative drug target for prevention of HPV-induced diseases.

The InSAeS4 Airborne X-Band Interferometric SAR System: A First Assessment on Its Imaging and Topographic Mapping Capabilities

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Stefano Perna

2016-01-01

Full Text Available We present in this work a first assessment of the imaging and topographic mapping capabilities of the InSAeS4 system, which is a single-pass interferometric airborne X-Band Synthetic Aperture Radar (SAR. In particular, we first provide a brief description of the InSAeS4 sensor. Then, we discuss the results of our analysis on the SAR and interferometric SAR products relevant to the first flight-test campaign. More specifically, we have exploited as reference the GPS measurements relevant to nine Corner Reflectors (CRs deployed over the illuminated area during the campaign and a laser scanner Digital Elevation Model (DEM. From the analysis carried out on the CRs we achieved a mean geometric resolution, for the SAR products, of about 0.14 m in azimuth and 0.49 m in range, a positioning misalignment with standard deviation of 0.07 m in range and 0.08 m in azimuth, and a height error with standard deviation of 0.51 m. From the comparison with the laser scanner DEM we estimated a height error with standard deviation of 1.57 m.

Imbalance of placental regulatory T cell and Th17 cell population dynamics in the FIV-infected pregnant cat

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Boudreaux Crystal E

2012-05-01

Full Text Available Abstract Background An appropriate balance in placental regulatory T cells (Tregs, an immunosuppressive cell population, and Th17 cells, a pro-inflammatory cell population, is essential in allowing tolerance of the semi-allogeneic fetus. TGF-β and IL-6 are cytokines that promote differentiation of Tregs and Th17 cells from a common progenitor; aberrant expression of the cytokines may perturb the balance in the two cell populations. We previously reported a pro-inflammatory placental environment with decreased levels of FoxP3, a Treg marker, and increased levels of IL-6 in the placentas of FIV-infected cats at early pregnancy. Thus, we hypothesized that FIV infection in the pregnant cat causes altered placental Treg and Th17 cell populations, possibly resulting in placental inflammation. Methods We examined the effect of FIV infection on Treg and Th17 populations in placentas at early pregnancy using quantitative confocal microscopy to measure FoxP3 or RORγ, a Th17 marker, and qPCR to quantify expression of the key cytokines TGF-β and IL-6. Results FoxP3 and RORγ were positively correlated in FIV-infected placentas at early pregnancy, but not placentas from normal cats, indicating virus-induced alteration in the balance of these cell populations. In control cats the expression of IL-6 and RORγ was positively correlated as predicted, but this relationship was disrupted in infected animals. TGF-β was reduced in infected queens, an occurrence that could dysregulate both Treg and Th17 cell populations. Co-expression analyses revealed a highly significant positive correlation between IL-6 and TGF-β expression in control animals that did not occur in infected animals. Conclusion Collectively, these data point toward potential disruption in the balance of Treg and Th17 cell populations that may contribute to FIV-induced inflammation in the feline placenta.

Skewing to the LFA-3 adhesion pathway by influenza infection of antigen-presenting cells

NARCIS (Netherlands)

van Kemenade, F. J.; Kuijpers, K. C.; de Waal-Malefijt, R.; van Lier, R. A.; Miedema, F.

1993-01-01

The effect of influenza (FLU) infection on heterotypic conjugate formation between antigen-presenting cells and T lymphocytes has been studied with FLU-specific T cell clones and FLU-infected B-lymphoblastoid cells (B-LCL). Conjugate formation between FLU-infected B-LCL (FLU+ B-LCL) and T cells was

Dynamics of NKT-Cell Responses to Chlamydial Infection.

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Shekhar, Sudhanshu; Joyee, Antony George; Yang, Xi

2015-01-01

Natural killer T (NKT) cells have gained great attention owing to their critical functional roles in immunity to various pathogens. In this review, we provide an overview of the current knowledge on the role of NKT cells in host defense against and pathogenesis due to Chlamydia, which is an intracellular bacterial pathogen that poses a threat to the public health worldwide. Accumulating evidence has demonstrated that NKT cells, particularly invariant NKT (iNKT) cells, play a crucial role in host defense against chlamydial infections, especially in C. pneumoniae infection. iNKT cells can promote type-1 protective responses to C. pneumoniae by inducing enhanced production of IL-12 by dendritic cells (DCs), in particular CD8α+ DCs, which promote the differentiation of naive T cells into protective IFN-γ-producing Th1/Tc1 type CD4+/CD8+ T cells. This iNKT-cell-mediated modulation of DC function is largely dependent upon CD40-CD40L interaction, IFN-γ production, and cell-to-cell contact. In addition, iNKT cells modulate the function of natural killer cells. NKT cells may be also involved in the pathogenesis of some chlamydial diseases by inducing different patterns of cytokine production. A better understanding of NKT-cell biology will enable us to rationally design prophylactic and therapeutic tools to combat infectious diseases.

Helicobacter pylori impairs murine dendritic cell responses to infection.

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Ya-Hui Wang

Full Text Available BACKGROUND: Helicobacter pylori, a human pathogen associated with chronic gastritis, peptic ulcer and gastric malignancies, is generally viewed as an extracellular microorganism. Here, we show that H. pylori replicates in murine bone marrow derived-dendritic cells (BMDCs within autophagosomes. METHODOLOGY/PRINCIPAL FINDINGS: A 10-fold increase of CFU is found between 2 h and 6 h p.i. in H. pylori-infected BMDCs. Autophagy is induced around the bacterium and participates at late time points of infection for the clearance of intracellular H. pylori. As a consequence of infection, LC3, LAMP1 and MHC class II molecules are retained within the H. pylori-containing vacuoles and export of MHC class II molecules to cell surface is blocked. However, formalin-fixed H. pylori still maintain this inhibitory activity in BMDC derived from wild type mice, but not in from either TLR4 or TLR2-deficient mice, suggesting the involvement of H. pylori-LPS in this process. TNF-alpha, IL-6 and IL-10 expression was also modulated upon infection showing a TLR2-specific dependent IL-10 secretion. No IL-12 was detected favoring the hypothesis of a down modulation of DC functions during H. pylori infection. Furthermore, antigen-specific T cells proliferation was also impaired upon infection. CONCLUSIONS/SIGNIFICANCE: H. pylori can infect and replicate in BMDCs and thereby affects DC-mediated immune responses. The implication of this new finding is discussed for the biological life cycle of H. pylori in the host.

B-1 cells modulate the murine macrophage response to Leishmania major infection.

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Arcanjo, Angelica F; Nunes, Marise P; Silva-Junior, Elias B; Leandro, Monique; da Rocha, Juliana Dutra Barbosa; Morrot, Alexandre; Decote-Ricardo, Debora; Freire-de-Lima, Celio Geraldo

2017-05-26

To investigate the modulatory effect of B-1 cells on murine peritoneal macrophages infected with Leishmania major ( L. major ) in vitro . Peritoneal macrophages obtained from BALB/c and BALB/c XID mice were infected with L. major and cultured in the presence or absence of B-1 cells obtained from wild-type BALB/c mice. Intracellular amastigotes were counted, and interleukin-10 (IL-10) production was quantified in the cellular supernatants using an enzyme-linked immunosorbent assay. The levels of the lipid mediator prostaglandin E2 (PGE 2 ) were determined using a PGE 2 enzyme immunoassay kit (Cayman Chemical, Ann Arbor, MI), and the number of lipid bodies was quantified in the cytoplasm of infected macrophages in the presence and absence of B-1 cells. Culturing the cells with selective PGE 2 -neutralizing drugs inhibited PGE 2 production and confirmed the role of this lipid mediator in IL-10 production. In contrast, we demonstrated that B-1 cells derived from IL-10 KO mice did not favor the intracellular growth of L. major . We report that B-1 cells promote the growth of L. major amastigotes inside peritoneal murine macrophages. We demonstrated that the modulatory effect was independent of physical contact between the cells, suggesting that soluble factor(s) were released into the cultures. We demonstrated in our co-culture system that B-1 cells trigger IL-10 production by L. major -infected macrophages. Furthermore, the increased secretion of IL-10 was attributed to the presence of the lipid mediator PGE 2 in supernatants of L. major -infected macrophages. The presence of B-1 cells also favors the production of lipid bodies by infected macrophages. In contrast, we failed to obtain the same effect on parasite replication inside L. major -infected macrophages when the B-1 cells were isolated from IL-10 knockout mice. Our results show that elevated levels of PGE 2 and IL-10 produced by B-1 cells increase L. major growth, as indicated by the number of parasites in cell

Caspase-2-dependent dendritic cell death, maturation, and priming of T cells in response to Brucella abortus infection.

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Xinna Li

Full Text Available Smooth virulent Brucella abortus strain 2308 (S2308 causes zoonotic brucellosis in cattle and humans. Rough B. abortus strain RB51, derived from S2308, is a live attenuated cattle vaccine strain licensed in the USA and many other countries. Our previous report indicated that RB51, but not S2308, induces a caspase-2-dependent apoptotic and necrotic macrophage cell death. Dendritic cells (DCs are professional antigen presenting cells critical for bridging innate and adaptive immune responses. In contrast to Brucella-infected macrophages, here we report that S2308 induced higher levels of apoptotic and necrotic cell death in wild type bone marrow-derived DCs (WT BMDCs than RB51. The RB51 and S2308-induced BMDC cell death was regulated by caspase-2, indicated by the minimal cell death in RB51 and S2308-infected BMDCs isolated from caspase-2 knockout mice (Casp2KO BMDCs. More S2308 bacteria were taken up by Casp2KO BMDCs than wild type BMDCs. Higher levels of S2308 and RB51 cells were found in infected Casp2KO BMDCs compared to infected WT BMDCs at different time points. RB51-infected wild type BMDCs were mature and activated as shown by significantly up-regulated expression of CD40, CD80, CD86, MHC-I, and MHC-II. RB51 induced the production of cytokines TNF-α, IL-6, IFN-γ and IL12/IL23p40 in infected BMDCs. RB51-infected WT BMDCs also stimulated the proliferation of CD4(+ and CD8(+ T cells compared to uninfected WT BMDCs. However, the maturation, activation, and cytokine secretion are significantly impaired in Casp2KO BMDCs infected with RB51 or Salmonella (control. S2308-infected WT and Casp2KO BMDCs were not activated and could not induce cytokine production. These results demonstrated that virulent smooth strain S2308 induced more apoptotic and necrotic dendritic cell death than live attenuated rough vaccine strain RB51; however, RB51, but not its parent strain S2308, induced caspase-2-mediated DC maturation, cytokine production, antigen

Infection of Human Fallopian Tube Epithelial Cells with Neisseria gonorrhoeae Protects Cells from Tumor Necrosis Factor Alpha-Induced Apoptosis

Science.gov (United States)

Morales, Priscilla; Reyes, Paz; Vargas, Macarena; Rios, Miguel; Imarai, Mónica; Cardenas, Hugo; Croxatto, Horacio; Orihuela, Pedro; Vargas, Renato; Fuhrer, Juan; Heckels, John E.; Christodoulides, Myron; Velasquez, Luis

2006-01-01

Following infection with Neisseria gonorrhoeae, bacteria may ascend into the Fallopian tubes (FT) and induce salpingitis, a major cause of infertility. In the FT, interactions between mucosal epithelial cells and gonococci are pivotal events in the pathogen's infection cycle and the inflammatory response. In the current study, primary FT epithelial cells were infected in vitro with different multiplicities of infection (MOI) of Pil+ Opa+ gonococci. Bacteria showed a dose-dependent association with cells and induced the secretion of tumor necrosis factor alpha (TNF-α). A significant finding was that gonococcal infection (MOI = 1) induced apoptosis in approximately 30% of cells, whereas increasing numbers of bacteria (MOI = 10 to 100) did not induce apoptosis. Apoptosis was observed in only 11% of cells with associated bacteria, whereas >84% of cells with no adherent bacteria were apoptotic. TNF-α was a key contributor to apoptosis, since (i) culture supernatants from cells infected with gonococci (MOI = 1) induced apoptosis in naïve cultures, suggesting that a soluble factor was responsible; (ii) gonococcal infection-induced apoptosis was inhibited with anti-TNF-α antibodies; and (iii) the addition of exogenous TNF-α induced apoptosis, which was inhibited by the presence of increasing numbers of bacteria (MOI = 10 to 100). These data suggest that TNF-α-mediated apoptosis of FT epithelial cells is likely a primary host defense mechanism to prevent pathogen colonization. However, epithelial cell-associated gonococci have evolved a mechanism to protect the cells from undergoing TNF-α-mediated apoptosis, and this modulation of the host innate response may contribute to establishment of infection. Understanding the antiapoptotic mechanisms used by Neisseria gonorrhoeae will inform the pathogenesis of salpingitis and could suggest new intervention strategies for prevention and treatment of the disease. PMID:16714596

B cell, CD8 + T cell and gamma delta T cell infiltration alters alveolar immune cell homeostasis in HIV-infected Malawian adults [version 2; referees: 1 approved, 2 approved with reservations

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Andrew Mwale

2017-12-01

Full Text Available Background: HIV infection is associated with increased risk to lower respiratory tract infections (LRTI. However, the impact of HIV infection on immune cell populations in the lung is not well defined. We sought to comprehensively characterise the impact of HIV infection on immune cell populations in the lung. Methods: Twenty HIV-uninfected controls and 17 HIV-1 infected ART-naïve adults were recruited from Queen Elizabeth Central Hospital, Malawi. Immunophenotyping of lymphocyte and myeloid cell populations was done on bronchoalveolar lavage fluid and peripheral blood cells. Results: We found that the numbers of CD8 + T cells, B cells and gamma delta T cells were higher in BAL fluid of HIV-infected adults compared to HIV-uninfected controls (all p<0.05. In contrast, there was no difference in the numbers of alveolar CD4 + T cells in HIV-infected adults compared to HIV-uninfected controls (p=0.7065. Intermediate monocytes were the predominant monocyte subset in BAL fluid (HIV-, 63%; HIV+ 81%, while the numbers of classical monocytes was lower in HIV-infected individuals compared to HIV-uninfected adults (1 × 10 5 vs. 2.8 × 10 5 cells/100ml of BAL fluid, p=0.0001. The proportions of alveolar macrophages and myeloid dendritic cells was lower in HIV-infected adults compared to HIV-uninfected controls (all p<0.05. Conclusions: Chronic HIV infection is associated with broad alteration of immune cell populations in the lung, but does not lead to massive depletion of alveolar CD4 + T cells. Disruption of alveolar immune cell homeostasis likely explains in part the susceptibility for LRTIs in HIV-infected adults.

Murine cytomegalovirus infection of neural stem cells alters neurogenesis in the developing brain.

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Manohar B Mutnal

2011-01-01

Full Text Available Congenital cytomegalovirus (CMV brain infection causes serious neuro-developmental sequelae including: mental retardation, cerebral palsy, and sensorineural hearing loss. But, the mechanisms of injury and pathogenesis to the fetal brain are not completely understood. The present study addresses potential pathogenic mechanisms by which this virus injures the CNS using a neonatal mouse model that mirrors congenital brain infection. This investigation focused on, analysis of cell types infected with mouse cytomegalovirus (MCMV and the pattern of injury to the developing brain.We used our MCMV infection model and a multi-color flow cytometry approach to quantify the effect of viral infection on the developing brain, identifying specific target cells and the consequent effect on neurogenesis. In this study, we show that neural stem cells (NSCs and neuronal precursor cells are the principal target cells for MCMV in the developing brain. In addition, viral infection was demonstrated to cause a loss of NSCs expressing CD133 and nestin. We also showed that infection of neonates leads to subsequent abnormal brain development as indicated by loss of CD24(hi cells that incorporated BrdU. This neonatal brain infection was also associated with altered expression of Oct4, a multipotency marker; as well as down regulation of the neurotrophins BDNF and NT3, which are essential to regulate the birth and differentiation of neurons during normal brain development. Finally, we report decreased expression of doublecortin, a marker to identify young neurons, following viral brain infection.MCMV brain infection of newborn mice causes significant loss of NSCs, decreased proliferation of neuronal precursor cells, and marked loss of young neurons.

Antibody response to a T-cell-independent antigen is preserved after splenic artery embolization for trauma.

Science.gov (United States)

Olthof, D C; Lammers, A J J; van Leeuwen, E M M; Hoekstra, J B L; ten Berge, I J M; Goslings, J C

2014-11-01

Splenic artery embolization (SAE) is increasingly being used as a nonoperative management strategy for patients with blunt splenic injury following trauma. The aim of this study was to assess the splenic function of patients who were embolized. A clinical study was performed, with splenic function assessed by examining the antibody response to polysaccharide antigens (pneumococcal 23-valent polysaccharide vaccine), B-cell subsets, and the presence of Howell-Jolly bodies (HJB). The data were compared to those obtained from splenectomized patients and healthy controls (HC) who had been included in a previously conducted study. A total of 30 patients were studied: 5 who had proximal SAE, 7 who had distal SAE, 8 who had a splenectomy, and 10 HC. The median vaccine-specific antibody response of the SAE patients (fold increase, 3.97) did not differ significantly from that of the HC (5.29; P = 0.90); however, the median response of the splenectomized patients (2.30) did differ (P = 0.003). In 2 of the proximally embolized patients and none of the distally embolized patients, the ratio of the IgG antibody level postvaccination compared to that prevaccination was splenic immune function of embolized patients was preserved, and therefore routine vaccination appears not to be indicated. Although the median antibody responses did not differ between the patients who underwent proximal SAE and those who underwent distal SAE, 2 of the 5 proximally embolized patients had insufficient responses to vaccination, whereas none of the distally embolized patients exhibited an insufficient response. Further research should be done to confirm this finding. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Effect of austempering temperature and time on mechanical properties of SAE 9260 steel

Science.gov (United States)

Dalwatkar, Ranjit; Prabhu, N.; Singh, R. K. P.

2018-04-01

This work describes the effect of austempering heat treatment on microstrcuture and mechanical properties of SAE 9260 steel. Steel samples, austenitized at 900 °C for one hour, were isothermally heat treated in the temperature range 300,325 and 350 °C for different times. Microstructural characterization was carried out using optical and scanning electron microscopes. The microstructure of the austempered samples consisted of bainitic ferrite and retained austenite. The volume fraction of retained austenite was determined using X-ray diffraction. Isothermal heat treatment at 350 °C for 20 min, resulted in a retained austenite content of around 38% in the microstructure. Increase in isothermal transformation temperature led to an increase in the fraction of retained austenite. Also, good combination of strength and ductility was obtained in the samples with increased amounts of retained austenite.

T cells for viral infections after allogeneic hematopoietic stem cell transplant.

Science.gov (United States)

Bollard, Catherine M; Heslop, Helen E

2016-06-30

Despite recent advances in the field of allogeneic hematopoietic stem cell transplantation (HSCT), viral infections are still a major complication during the period of immune suppression that follows the procedure. Adoptive transfer of donor-derived virus-specific cytotoxic T cells (VSTs) is a strategy to rapidly restore virus-specific immunity to prevent or treat viral diseases after HSCT. Early proof of principle studies demonstrated that the administration of donor-derived T cells specific for cytomegalovirus or Epstein-Barr virus (EBV) could effectively restore virus-specific immunity and control viral infections. Subsequent studies using different expansion or direct selection techniques have shown that donor-derived VSTs confer protection in vivo after adoptive transfer in 70% to 90% of recipients. Because a major cause of failure is lack of immunity to the infecting virus in a naïve donor, more recent studies have infused closely matched third-party VSTs and reported response rates of 60% to 70%. Current efforts have focused on broadening the applicability of this approach by: (1) extending the number of viral antigens being targeted, (2) simplifying manufacture, (3) exploring strategies for recipients of virus-naïve donor grafts, and (4) developing and optimizing "off the shelf" approaches. © 2016 by The American Society of Hematology.

Dendritic cell maturation, but not type I interferon exposure, restricts infection by HTLV-1, and viral transmission to T-cells.

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Gergès Rizkallah

2017-04-01

Full Text Available Human T lymphotropic Virus type 1 (HTLV-1 is the etiological agent of Adult T cell Leukemia/Lymphoma (ATLL and HTLV-1-Associated Myelopathy/Tropical Spastic Paraparesis (HAM/TSP. Both CD4+ T-cells and dendritic cells (DCs infected with HTLV-1 are found in peripheral blood from HTLV-1 carriers. We previously demonstrated that monocyte-derived IL-4 DCs are more susceptible to HTLV-1 infection than autologous primary T-cells, suggesting that DC infection precedes T-cell infection. However, during blood transmission, breast-feeding or sexual transmission, HTLV-1 may encounter different DC subsets present in the blood, the intestinal or genital mucosa respectively. These different contacts may impact HTLV-1 ability to infect DCs and its subsequent transfer to T-cells. Using in vitro monocyte-derived IL-4 DCs, TGF-β DCs and IFN-α DCs that mimic DCs contacting HTLV-1 in vivo, we show here that despite their increased ability to capture HTLV-1 virions, IFN-α DCs restrict HTLV-1 productive infection. Surprisingly, we then demonstrate that it is not due to the antiviral activity of type-I interferon produced by IFN-α DCs, but that it is likely to be linked to a distinct trafficking route of HTLV-1 in IL-4 DCs vs. IFN-α DCs. Finally, we demonstrate that, in contrast to IL-4 DCs, IFN-α DCs are impaired in their capacity to transfer HTLV-1 to CD4 T-cells, both after viral capture and trans-infection and after their productive infection. In conclusion, the nature of the DCs encountered by HTLV-1 upon primo-infection and the viral trafficking route through the vesicular pathway of these cells determine the efficiency of viral transmission to T-cells, which may condition the fate of infection.

Deficient CD4+ T cell priming and regression of CD8+ T cell functionality in virus-infected mice lacking a normal B cell compartment

DEFF Research Database (Denmark)

Christensen, Jan Pravsgaard; Kauffmann, Susanne Ørding; Thomsen, Allan Randrup

2003-01-01

of virus-specific CD4(+) T cells was markedly impaired in B(-/-) mice infected with either virus strain. Thus, our results indicate that B cells play an important role in antiviral immunity not only as Ab producers, but also in promoting an optimal and sustained T cell response. The T cell defects......In this study, we investigate the state of T cell-mediated immunity in B cell-deficient (B(-/-)) mice infected with two strains of lymphocytic choriomeningitis virus known to differ markedly in their capacity to persist. In B(-/-) C57BL mice infected with the more persisting virus, virus......-specific CD8(+) T cells are initially generated that are qualitatively similar to those in wild-type mice. However, although cell numbers are well sustained over time, the capacity to produce cytokines is rapidly impaired. In similarly infected B(-/-) BALB/c mice, virus-specific CD8(+) T cells are completely...

Merkel Cell Polyomavirus Infection of Animal Dermal Fibroblasts.

Science.gov (United States)

Liu, Wei; Krump, Nathan A; MacDonald, Margo; You, Jianxin

2018-02-15

Merkel cell polyomavirus (MCPyV) is the first polyomavirus to be associated with human cancer. Mechanistic studies attempting to fully elucidate MCPyV's oncogenic mechanisms have been hampered by the lack of animal models for MCPyV infection. In this study, we examined the ability of MCPyV-GFP pseudovirus (containing a green fluorescent protein [GFP] reporter construct), MCPyV recombinant virions, and several MCPyV chimeric viruses to infect dermal fibroblasts isolated from various model animals, including mouse ( Mus musculus ), rabbit ( Oryctolagus cuniculus ), rat ( Rattus norvegicus ), chimpanzee ( Pan troglodytes ), rhesus macaque ( Macaca mulatta ), patas monkey ( Erythrocebus patas ), common woolly monkey ( Lagothrix lagotricha ), red-chested mustached tamarin ( Saguinus labiatus ), and tree shrew ( Tupaia belangeri ). We found that MCPyV-GFP pseudovirus was able to enter the dermal fibroblasts of all species tested. Chimpanzee dermal fibroblasts were the only type that supported vigorous MCPyV gene expression and viral replication, and they did so to a level beyond that of human dermal fibroblasts. We further demonstrated that both human and chimpanzee dermal fibroblasts produce infectious MCPyV virions that can successfully infect new cells. In addition, rat dermal fibroblasts supported robust MCPyV large T antigen expression after infection with an MCPyV chimeric virus in which the entire enhancer region of the MCPyV early promoter has been replaced with the simian virus 40 (SV40) analog. Our results suggest that viral transcription and/or replication events represent the major hurdle for MCPyV cross-species transmission. The capacity of rat dermal fibroblasts to support MCPyV early gene expression suggests that the rat is a candidate model organism for studying viral oncogene function during Merkel cell carcinoma (MCC) oncogenic progression. IMPORTANCE MCPyV plays an important role in the development of a highly aggressive form of skin cancer, Merkel

Persistence of memory B-cell and T-cell responses to the quadrivalent HPV vaccine in HIV-infected children.

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Weinberg, Adriana; Huang, Sharon; Moscicki, Anna-Barbara; Saah, Afred; Levin, Myron J

2018-04-24

To determine the magnitude and persistence of quadrivalent human papillomavirus (HPV)16 and HPV18 B-cell and T-cell memory after three or four doses of quadrivalent HPV vaccine (QHPV) in HIV-infected children. Seventy-four HIV-infected children immunized with four doses and 23 with three doses of QHPV had HPV16 and HPV18 IgG B-cell and IFNγ and IL2 T-cell ELISPOT performed at 2, 3.5 and 4-5 years after the last dose. HPV16 and HPV18 T-cell responses were similar in both treatment groups, with higher responses to HPV16 vs. HPV18. These HPV T-cell responses correlated with HIV disease characteristics at the study visits. Global T-cell function declined over time as measured by nonspecific mitogenic stimulation. B-cell memory was similar across treatment groups and HPV genotypes. There was a decline in HPV-specific B-cell memory over time that reached statistical significance for HPV16 in the four-dose group. B-cell and T-cell memory did not significantly differ after either three or four doses of QHPV in HIV-infected children. The clinical consequences of decreasing global T-cell function and HPV B-cell memory over time in HIV-infected children requires further investigation.

Cytoarchitecture of Zika virus infection in human neuroblastoma and Aedes albopictus cell lines

International Nuclear Information System (INIS)

Offerdahl, Danielle K.; Dorward, David W.; Hansen, Bryan T.; Bloom, Marshall E.

2017-01-01

The Zika virus (ZIKV) pandemic is a global concern due to its role in the development of congenital anomalies of the central nervous system. This mosquito-borne flavivirus alternates between mammalian and mosquito hosts, but information about the biogenesis of ZIKV is limited. Using a human neuroblastoma cell line (SK-N-SH) and an Aedes albopictus mosquito cell line (C6/36), we characterized ZIKV infection by immunofluorescence, transmission electron microscopy (TEM), and electron tomography (ET) to better understand infection in these disparate host cells. ZIKV replicated well in both cell lines, but infected SK-N-SH cells suffered a lytic crisis. Flaviviruses scavenge host cell membranes to serve as replication platforms and ZIKV showed the hallmarks of this process. Via TEM, we identified virus particles and 60–100 nm spherular vesicles. ET revealed these vesicular replication compartments contain smaller 20–30 nm spherular structures. Our studies indicate that SK-N-SH and C6/36 cells are relevant models for viral cytoarchitecture study. - Highlights: •First electron tomography of Zika virus cytoarchitecture. •Comparison of Zika virus infection in human neuroblastoma and mosquito cells. •Ultrastructure of Zika virus infection in human neuroblastoma and mosquito cells.

Cytoarchitecture of Zika virus infection in human neuroblastoma and Aedes albopictus cell lines

Energy Technology Data Exchange (ETDEWEB)

Offerdahl, Danielle K. [Laboratory of Virology, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, MT (United States); Dorward, David W.; Hansen, Bryan T. [Microscopy Unit, Research Technology Branch, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, MT (United States); Bloom, Marshall E., E-mail: mbloom@nih.gov [Laboratory of Virology, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, MT (United States)

2017-01-15

The Zika virus (ZIKV) pandemic is a global concern due to its role in the development of congenital anomalies of the central nervous system. This mosquito-borne flavivirus alternates between mammalian and mosquito hosts, but information about the biogenesis of ZIKV is limited. Using a human neuroblastoma cell line (SK-N-SH) and an Aedes albopictus mosquito cell line (C6/36), we characterized ZIKV infection by immunofluorescence, transmission electron microscopy (TEM), and electron tomography (ET) to better understand infection in these disparate host cells. ZIKV replicated well in both cell lines, but infected SK-N-SH cells suffered a lytic crisis. Flaviviruses scavenge host cell membranes to serve as replication platforms and ZIKV showed the hallmarks of this process. Via TEM, we identified virus particles and 60–100 nm spherular vesicles. ET revealed these vesicular replication compartments contain smaller 20–30 nm spherular structures. Our studies indicate that SK-N-SH and C6/36 cells are relevant models for viral cytoarchitecture study. - Highlights: •First electron tomography of Zika virus cytoarchitecture. •Comparison of Zika virus infection in human neuroblastoma and mosquito cells. •Ultrastructure of Zika virus infection in human neuroblastoma and mosquito cells.

Development of ultrafine ferritic sheaves/plates in SAE 52100 steel for enhancement of strength by controlled thermomechanical processing

Energy Technology Data Exchange (ETDEWEB)

Chakraborty, J. [Department of Metallurgical and Materials Engineering, Indian Institute of Technology, Kharagpur, W.B. 721 302 (India); Scientific Services and Research and Development, Tata Steel, Jamshedpur 831 001, Jharkhand (India); Manna, I., E-mail: imanna@metal.iitkgp.ernet.in [Department of Metallurgical and Materials Engineering, Indian Institute of Technology, Kharagpur, W.B. 721 302 (India); Central Glass and Ceramic Research Institute (CGCRI), Council of Scientific and Industrial Research (CSIR), 196 Raja S C Mullick Road, Jadavpur, Kolkata 700032 (India)

2012-06-30

Highlights: Black-Right-Pointing-Pointer Ultrafine bainite + martensite duplex microstructure developed in SAE 52100 steel. Black-Right-Pointing-Pointer Thermomechanical processing modifies size + morphology of bainitic ferrite. Black-Right-Pointing-Pointer Processing involves hot deformation prior to/during/after austenitizing. Black-Right-Pointing-Pointer Significant improvement in mechanical strength achieved. Black-Right-Pointing-Pointer Similar study on high carbon, low alloy steel not reported in the literature. - Abstract: The present study attempts to tailor the size, morphology and distribution of the ferrite needles/sheaves by thermomechanical processing and develop an ultrafine ferrite + martensite duplex microstructure for enhancement of strength and toughness in SAE 52100 steel. The thermo-mechanical routine included 5% hot deformation before, during or after austenitizing at 950 Degree-Sign C for 15 min followed by austempering at 270 Degree-Sign C for 30 min and subsequent water quenching to room temperature. Optical/electron microscopy along with X-ray diffraction was used to quantitatively monitor the size, morphology and distribution of the phase or phase aggregate. Significant improvement in nanohardness, wear resistance and elastic modulus and was observed in samples subjected to thermomechanical processing, as compared to that following the same austenitizing and austempering routine without hot deformation at any stage. However, improvement in the bulk mechanical property due to the present thermo-mechanical is lower than that obtained in our earlier study comprising cold deformation prior to austenitizing and austempering.

Estimating immunoregulatory gene networks in human herpesvirus type 6-infected T cells

International Nuclear Information System (INIS)

Takaku, Tomoiku; Ohyashiki, Junko H.; Zhang, Yu; Ohyashiki, Kazuma

2005-01-01

The immune response to viral infection involves complex network of dynamic gene and protein interactions. We present here the dynamic gene network of the host immune response during human herpesvirus type 6 (HHV-6) infection in an adult T-cell leukemia cell line. Using a pathway-focused oligonucleotide DNA microarray, we found a possible association between chemokine genes regulating Th1/Th2 balance and genes regulating T-cell proliferation during HHV-6B infection. Gene network analysis using an integrated comprehensive workbench, VoyaGene, revealed that a gene encoding a TEC-family kinase, ITK, might be a putative modulator in the host immune response against HHV-6B infection. We conclude that Th2-dominated inflammatory reaction in host cells may play an important role in HHV-6B-infected T cells, thereby suggesting the possibility that ITK might be a therapeutic target in diseases related to dysregulation of Th1/Th2 balance. This study describes a novel approach to find genes related with the complex host-virus interaction using microarray data employing the Bayesian statistical framework

Detection of mycoplasma infection in circulating tumor cells in patients with hepatocellular carcinoma

Energy Technology Data Exchange (ETDEWEB)

Choi, Hong Seo; Lee, Hyun Min; Kim, Won-Tae; Kim, Min Kyu [Department of Bioscience and Biotechnology, Institute of Bioscience, Sejong University, Seoul (Korea, Republic of); Chang, Hee Jin [Center for Colorectal Cancer, Research Institute and Hospital of National Cancer Center, Goyang-si (Korea, Republic of); Lee, Hye Ran [Department of Internal Medicine, Inje University Ilsan Paik Hospital, Goyang-si (Korea, Republic of); Joh, Jae-Won [Department of Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul (Korea, Republic of); Kim, Dae Shick, E-mail: oncorkim@skku.edu [Department of Pathology, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul (Korea, Republic of); Ryu, Chun Jeih, E-mail: cjryu@sejong.ac.kr [Department of Bioscience and Biotechnology, Institute of Bioscience, Sejong University, Seoul (Korea, Republic of)

2014-04-04

Highlights: • This study generates a monoclonal antibody CA27 against the mycoplasmal p37 protein. • CA27 isolates circulating tumor cells (CTCs) from the blood of liver cancer patients. • Results show the first evidence for mycoplasma infected-CTCs in cancer patients. - Abstract: Many studies have shown that persistent infections of bacteria promote carcinogenesis and metastasis. Infectious agents and their products can modulate cancer progression through the induction of host inflammatory and immune responses. The presence of circulating tumor cells (CTCs) is considered as an important indicator in the metastatic cascade. We unintentionally produced a monoclonal antibody (MAb) CA27 against the mycoplasmal p37 protein in mycoplasma-infected cancer cells during the searching process of novel surface markers of CTCs. Mycoplasma-infected cells were enriched by CA27-conjugated magnetic beads in the peripheral blood mononuclear cells in patients with hepatocellular carcinoma (HCC) and analyzed by confocal microscopy with anti-CD45 and CA27 antibodies. CD45-negative and CA27-positive cells were readily detected in three out of seven patients (range 12–30/8.5 ml blood), indicating that they are mycoplasma-infected circulating epithelial cells. CA27-positive cells had larger size than CD45-positive hematological lineage cells, high nuclear to cytoplasmic ratios and irregular nuclear morphology, which identified them as CTCs. The results show for the first time the existence of mycoplasma-infected CTCs in patients with HCC and suggest a possible correlation between mycoplasma infection and the development of cancer metastasis.

Macrophages are required for dendritic cell uptake of respiratory syncytial virus from an infected epithelium.

Science.gov (United States)

Ugonna, Kelechi; Bingle, Colin D; Plant, Karen; Wilson, Kirsty; Everard, Mark L

2014-01-01

We have previously shown that the respiratory syncytial virus [RSV] can productively infect monocyte derived dendritic cells [MoDC] and remain dormant within the same cells for prolonged periods. It is therefore possible that infected dendritic cells act as a reservoir within the airways of individuals between annual epidemics. In the present study we explored the possibility that sub-epithelial DCs can be infected with RSV from differentiated bronchial epithelium and that in turn RSV from DCs can infect the epithelium. A dual co-culture model was established in which a differentiated primary airway epithelium on an Air Liquid Interface (ALI) was cultured on a transwell insert and MoDCs were subsequently added to the basolateral membrane of the insert. Further experiments were undertaken using a triple co-culture model in which in which macrophages were added to the apical surface of the differentiated epithelium. A modified RSV [rr-RSV] expressing a red fluorescent protein marker of replication was used to infect either the MoDCs or the differentiated epithelium and infection of the reciprocal cell type was assessed using confocal microscopy. Our data shows that primary epithelium became infected when rr-RSV infected MoDCs were introduced onto the basal surface of the transwell insert. MoDCs located beneath the epithelium did not become infected with virus from infected epithelial cells in the dual co-culture model. However when macrophages were present on the apical surface of the primary epithelium infection of the basal MoDCs occurred. Our data suggests that RSV infected dendritic cells readily transmit infection to epithelial cells even when they are located beneath the basal layer. However macrophages appear to be necessary for the transmission of infection from epithelial cells to basal dendritic cells.

Memory Th1 Cells Are Protective in Invasive Staphylococcus aureus Infection

Science.gov (United States)

Lalor, Stephen J.; Leech, John M.; O’Keeffe, Kate M.; Mac Aogáin, Micheál; O’Halloran, Dara P.; Lacey, Keenan A.; Tavakol, Mehri; Hearnden, Claire H.; Fitzgerald-Hughes, Deirdre; Humphreys, Hilary; Fennell, Jérôme P.; van Wamel, Willem J.; Foster, Timothy J.; Geoghegan, Joan A.; Lavelle, Ed C.; Rogers, Thomas R.; McLoughlin, Rachel M.

2015-01-01

Mechanisms of protective immunity to Staphylococcus aureus infection in humans remain elusive. While the importance of cellular immunity has been shown in mice, T cell responses in humans have not been characterised. Using a murine model of recurrent S. aureus peritonitis, we demonstrated that prior exposure to S. aureus enhanced IFNγ responses upon subsequent infection, while adoptive transfer of S. aureus antigen-specific Th1 cells was protective in naïve mice. Translating these findings, we found that S. aureus antigen-specific Th1 cells were also significantly expanded during human S. aureus bloodstream infection (BSI). These Th1 cells were CD45RO+, indicative of a memory phenotype. Thus, exposure to S. aureus induces memory Th1 cells in mice and humans, identifying Th1 cells as potential S. aureus vaccine targets. Consequently, we developed a model vaccine comprising staphylococcal clumping factor A, which we demonstrate to be an effective human T cell antigen, combined with the Th1-driving adjuvant CpG. This novel Th1-inducing vaccine conferred significant protection during S. aureus infection in mice. This study notably advances our understanding of S. aureus cellular immunity, and demonstrates for the first time that a correlate of S. aureus protective immunity identified in mice may be relevant in humans. PMID:26539822

Memory Th1 Cells Are Protective in Invasive Staphylococcus aureus Infection.

LENUS (Irish Health Repository)

Brown, Aisling F

2015-01-01

Mechanisms of protective immunity to Staphylococcus aureus infection in humans remain elusive. While the importance of cellular immunity has been shown in mice, T cell responses in humans have not been characterised. Using a murine model of recurrent S. aureus peritonitis, we demonstrated that prior exposure to S. aureus enhanced IFNγ responses upon subsequent infection, while adoptive transfer of S. aureus antigen-specific Th1 cells was protective in naïve mice. Translating these findings, we found that S. aureus antigen-specific Th1 cells were also significantly expanded during human S. aureus bloodstream infection (BSI). These Th1 cells were CD45RO+, indicative of a memory phenotype. Thus, exposure to S. aureus induces memory Th1 cells in mice and humans, identifying Th1 cells as potential S. aureus vaccine targets. Consequently, we developed a model vaccine comprising staphylococcal clumping factor A, which we demonstrate to be an effective human T cell antigen, combined with the Th1-driving adjuvant CpG. This novel Th1-inducing vaccine conferred significant protection during S. aureus infection in mice. This study notably advances our understanding of S. aureus cellular immunity, and demonstrates for the first time that a correlate of S. aureus protective immunity identified in mice may be relevant in humans.

Exogenous and endogenous hyaluronic acid reduces HIV infection of CD4+ T cells

Science.gov (United States)

Li, Peilin; Fujimoto, Katsuya; Bourguingnon, Lilly; Yukl, Steven; Deeks, Steven; Wong, Joseph K

2014-01-01

Preventing mucosal transmission of HIV is critical to halting the HIV epidemic. Novel approaches to preventing mucosal transmission are needed. Hyaluronic acid (HA) is a major extracellular component of mucosa and the primary ligand for the cell surface receptor CD44. CD44 enhances HIV infection of CD4+ T cells, but the role of HA in this process is not clear. To study this, virions were generated with CD44 (HIVCD44) or without CD44 (HIVmock). Exogenous HA reduced HIV infection of unstimulated CD4+ T cells in a CD44-dependent manner. Conversely, hyaluronidase-mediated reduction of endogenous HA on the cell surface enhanced HIV binding to and infection of unstimulated CD4+ T cells. Exogenous HA treatment reduced activation of protein kinase C alpha via CD44 on CD4+ T cells during infection with HIVCD44. These results reveal new roles for HA during the interaction of HIV with CD4+ T cells that may be relevant to mucosal HIV transmission and could be exploitable as a future strategy to prevent HIV infection. PMID:24957217

Murine and bovine γδ T cells enhance innate immunity against Brucella abortus infections.

Directory of Open Access Journals (Sweden)

Jerod A Skyberg

Full Text Available γδ T cells have been postulated to act as a first line of defense against infectious agents, particularly intracellular pathogens, representing an important link between the innate and adaptive immune responses. Human γδ T cells expand in the blood of brucellosis patients and are active against Brucella in vitro. However, the role of γδ T cells in vivo during experimental brucellosis has not been studied. Here we report TCRδ(-/- mice are more susceptible to B. abortus infection than C57BL/6 mice at one week post-infection as measured by splenic colonization and splenomegaly. An increase in TCRγδ cells was observed in the spleens of B. abortus-infected C57BL/6 mice, which peaked at two weeks post-infection and occurred concomitantly with diminished brucellae. γδ T cells were the major source of IL-17 following infection and also produced IFN-γ. Depletion of γδ T cells from C57BL/6, IL-17Rα(-/-, and GMCSF(-/- mice enhanced susceptibility to B. abortus infection although this susceptibility was unaltered in the mutant mice; however, when γδ T cells were depleted from IFN-γ(-/- mice, enhanced susceptibility was observed. Neutralization of γδ T cells in the absence of TNF-α did not further impair immunity. In the absence of TNF-α or γδ T cells, B. abortus-infected mice showed enhanced IFN-γ, suggesting that they augmented production to compensate for the loss of γδ T cells and/or TNF-α. While the protective role of γδ T cells was TNF-α-dependent, γδ T cells were not the major source of TNF-α and activation of γδ T cells following B. abortus infection was TNF-α-independent. Additionally, bovine TCRγδ cells were found to respond rapidly to B. abortus infection upon co-culture with autologous macrophages and could impair the intramacrophage replication of B. abortus via IFN-γ. Collectively, these results demonstrate γδ T cells are important for early protection to B. abortus infections.

Chronic schistosomiasis during pregnancy epigenetically reprograms T-cell differentiation in offspring of infected mothers.

Science.gov (United States)

Klar, Kathrin; Perchermeier, Sophie; Bhattacharjee, Sonakshi; Harb, Hani; Adler, Thure; Istvanffy, Rouzanna; Loffredo-Verde, Eva; Oostendorp, Robert A; Renz, Harald; Prazeres da Costa, Clarissa

2017-05-01

Schistosomiasis is a nontransplacental helminth infection. Chronic infection during pregnancy suppresses allergic airway responses in offspring. We addressed the question whether in utero exposure to chronic schistosome infection (Reg phase) in mice affects B-cell and T-cell development. Therefore, we focused our analyses on T-cell differentiation capacity induced by epigenetic changes in promoter regions of signature cytokines in offspring. Here, we show that naïve T cells from offspring of schistosome infected female mice had a strong capacity to differentiate into T H 1 cells, whereas T H 2 differentiation was impaired. In accordance, reduced levels of histone acetylation of the IL-4 promoter regions were observed in naïve T cells. To conclude, our mouse model revealed distinct epigenetic changes within the naïve T-cell compartment affecting T H 2 and T H 1 cell differentiation in offspring of mothers with chronic helminth infection. These findings could eventually help understand how helminths alter T-cell driven immune responses induced by allergens, bacterial or viral infections, as well as vaccines. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Quantitative proteomics of Spodoptera frugiperda cells during growth and baculovirus infection.

Directory of Open Access Journals (Sweden)

Nuno Carinhas

Full Text Available Baculovirus infection of Spodoptera frugiperda cells is a system of choice to produce a range of recombinant proteins, vaccines and, potentially, gene therapy vectors. While baculovirus genomes are well characterized, the genome of S. frugiperda is not sequenced and the virus-host molecular interplay is sparsely known. Herein, we describe the application of stable isotope labeling by amino acids in cell culture (SILAC to obtain the first comparative proteome quantitation of S. frugiperda cells during growth and early baculovirus infection. The proteome coverage was maximized by compiling a search database with protein annotations from insect species. Of interest were differentially proteins related to energy metabolism, endoplasmic reticulum and oxidative stress, yet not investigated in the scope of baculovirus infection. Further, the reduced expression of key viral-encoded proteins early in the infection cycle is suggested to be related with decreased viral replication at high cell density culture. These findings have implications for virological research and improvement of baculovirus-based bioprocesses.

Role of dendritic cells infected with human herpesvirus 6 in virus transmission to CD4+ T cells

International Nuclear Information System (INIS)

Takemoto, Masaya; Imasawa, Takayoshi; Yamanishi, Koichi; Mori, Yasuko

2009-01-01

Human herpesvirus 6 (HHV-6) is a ubiquitous betaherpesvirus that predominantly infects and replicates in CD4 + T lymphocytes. However, the mechanism of HHV-6 transmission to T cells from the peripheral mucosa is unknown. Here we found that dendritic cells (DCs) can transmit HHV-6 to T cells, resulting in productive infection. In immature monocyte-derived DCs (MDDCs) infected with HHV-6, viral early and late antigens were expressed, and nucleocapsids containing a DNA core were observed, although few virions were detected in the cytoplasm by electron microscopy, indicating that the maturation of HHV-6 virions may be incomplete in MDDCs. However, HHV-6 transmission from MDDCs to stimulated CD4 + T cells occurred efficiently in coculture of these cells, but not from MDDCs culture supernatants. This transmission was partially inhibited by treating the DCs with a viral DNA synthesis blocker, indicating that viral replication in MDDCs is required for this transmission. Furthermore, myeloid DCs and plasmacytoid DCs infected with HHV-6 could also transmit the virus to stimulated T cells. Thus, DCs may be the first cell population targeted by HHV-6 and could play an important role in the virus' transmission to T cells for their further propagation

HIV-1 Infection Is Associated with Depletion and Functional Impairment of Mycobacterium tuberculosis-Specific CD4 T Cells in Individuals with Latent Tuberculosis Infection.

Science.gov (United States)

Day, Cheryl L; Abrahams, Deborah A; Harris, Levelle D; van Rooyen, Michele; Stone, Lynnett; de Kock, Marwou; Hanekom, Willem A

2017-09-15

Coinfection with HIV is the single greatest risk factor for reactivation of latent Mycobacterium tuberculosis infection (LTBI) and progression to active tuberculosis disease. HIV-associated dysregulation of adaptive immunity by depletion of CD4 Th cells most likely contributes to loss of immune control of LTBI in HIV-infected individuals, although the precise mechanisms whereby HIV infection impedes successful T cell-mediated control of M. tuberculosis have not been well defined. To further delineate mechanisms whereby HIV impairs protective immunity to M. tuberculosis , we evaluated the frequency, phenotype, and functional capacity of M. tuberculosis -specific CD4 T cells in HIV-infected and HIV-uninfected adults with LTBI. HIV infection was associated with a lower total frequency of cytokine-producing M. tuberculosis -specific CD4 T cells, and preferential depletion of a discrete subset of M. tuberculosis -specific IFN-γ + IL-2 - TNF-α + CD4 T cells. M. tuberculosis -specific CD4 T cells in HIV-infected individuals expressed significantly higher levels of Ki67, compared with HIV-uninfected individuals, thus indicating recent activation and turnover of these cells in vivo. The ex vivo proliferative capacity of M. tuberculosis -specific CD4 T cells was markedly impaired in HIV-infected individuals, compared with HIV-uninfected individuals. Moreover, HIV infection was associated with increased M. tuberculosis Ag-induced CD4 T cell death ex vivo, indicating a possible mechanism contributing to impaired proliferative capacity of M. tuberculosis -specific CD4 T cells in HIV-infected individuals. These data provide new insights into the parameters of M. tuberculosis -specific CD4 T cell immunity that are impaired in HIV-infected individuals with LTBI, which may contribute to their increased risk of developing active tuberculosis disease. Copyright © 2017 by The American Association of Immunologists, Inc.

Differential gene expressions of the MAPK signaling pathway in enterovirus 71-infected rhabdomyosarcoma cells

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Weifeng Shi

Full Text Available BACKGROUND: Mitogen-activated protein kinase (MAPK signaling pathway plays an important role in response to viral infection. The aim of this study was to explore the function and mechanism of MAPK signaling pathway in enterovirus 71 (EV71 infection of human rhabdomyosarcoma (RD cells. METHODS: Apoptosis of RD cells was observed using annexin V-FITC/PI binding assay under a fluorescence microscope. Cellular RNA was extracted and transcribed to cDNA. The expressions of 56 genes of MAPK signaling pathway in EV71-infected RD cells at 8 h and 20 h after infection were analyzed by PCR array. The levels of IL-2, IL-4, IL-10, and TNF-α in the supernatant of RD cells infected with EV71 at different time points were measured by ELISA. RESULTS: The viability of RD cells decreased obviously within 48 h after EV71 infection. Compared with the control group, EV71 infection resulted in the significantly enhanced releases of IL-2, IL-4, IL-10 and TNF-α from infected RD cells (p < 0.05. At 8 h after infection, the expressions of c-Jun, c-Fos, IFN-i, MEKK1, MLK3 and NIK genes in EV71-infected RD cells were up-regulated by 2.08-6.12-fold, whereas other 19 genes (e.g. AKT1, AKT2, E2F1, IKK and NF-κB1 exhibited down-regulation. However, at 20 h after infection, those MAPK signaling molecules including MEKK1, ASK1, MLK2, MLK3, NIK, MEK1, MEK2, MEK4, MEK7, ERK1, JNK1 and JNK2 were up-regulated. In addition, the expressions of AKT2, ELK1, c-Jun, c-Fos, NF-κB p65, PI3K and STAT1 were also increased. CONCLUSION: EV71 infection induces the differential gene expressions of MAPK signaling pathway such as ERK, JNK and PI3K/AKT in RD cells, which may be associated with the secretions of inflammatory cytokines and host cell apoptosis.

Interferon-alpha subtype 11 activates NK cells and enables control of retroviral infection.

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Kathrin Gibbert

Full Text Available The innate immune response mediated by cells such as natural killer (NK cells is critical for the rapid containment of virus replication and spread during acute infection. Here, we show that subtype 11 of the type I interferon (IFN family greatly potentiates the antiviral activity of NK cells during retroviral infection. Treatment of mice with IFN-α11 during Friend retrovirus infection (FV significantly reduced viral loads and resulted in long-term protection from virus-induced leukemia. The effect of IFN-α11 on NK cells was direct and signaled through the type I IFN receptor. Furthermore, IFN-α11-mediated activation of NK cells enabled cytolytic killing of FV-infected target cells via the exocytosis pathway. Depletion and adoptive transfer experiments illustrated that NK cells played a major role in successful IFN-α11 therapy. Additional experiments with Mouse Cytomegalovirus infections demonstrated that the therapeutic effect of IFN-α11 is not restricted to retroviruses. The type I IFN subtypes 2 and 5, which bind the same receptor as IFN-α11, did not elicit similar antiviral effects. These results demonstrate a unique and subtype-specific activation of NK cells by IFN-α11.

Flow cytometric monitoring of influenza A virus infection in MDCK cells during vaccine production

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Reichl Udo

2008-04-01

Full Text Available Abstract Background In cell culture-based influenza vaccine production the monitoring of virus titres and cell physiology during infection is of great importance for process characterisation and optimisation. While conventional virus quantification methods give only virus titres in the culture broth, data obtained by fluorescence labelling of intracellular virus proteins provide additional information on infection dynamics. Flow cytometry represents a valuable tool to investigate the influences of cultivation conditions and process variations on virus replication and virus yields. Results In this study, fluorescein-labelled monoclonal antibodies against influenza A virus matrix protein 1 and nucleoprotein were used for monitoring the infection status of adherent Madin-Darby canine kidney cells from bioreactor samples. Monoclonal antibody binding was shown for influenza A virus strains of different subtypes (H1N1, H1N2, H3N8 and host specificity (human, equine, swine. At high multiplicity of infection in a bioreactor, the onset of viral protein accumulation in adherent cells on microcarriers was detected at about 2 to 4 h post infection by flow cytometry. In contrast, a significant increase in titre by hemagglutination assay was detected at the earliest 4 to 6 h post infection. Conclusion It is shown that flow cytometry is a sensitive and robust method for the monitoring of viral infection in fixed cells from bioreactor samples. Therefore, it is a valuable addition to other detection methods of influenza virus infection such as immunotitration and RNA hybridisation. Thousands of individual cells are measured per sample. Thus, the presented method is believed to be quite independent of the concentration of infected cells (multiplicity of infection and total cell concentration in bioreactors. This allows to perform detailed studies on factors relevant for optimization of virus yields in cell cultures. The method could also be used for process

Natural killer cells recognize friend retrovirus-infected erythroid progenitor cells through NKG2D-RAE-1 interactions In Vivo.

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Ogawa, Tatsuya; Tsuji-Kawahara, Sachiyo; Yuasa, Takae; Kinoshita, Saori; Chikaishi, Tomomi; Takamura, Shiki; Matsumura, Haruo; Seya, Tsukasa; Saga, Toshihiko; Miyazawa, Masaaki

2011-06-01

Natural killer (NK) cells function as early effector cells in the innate immune defense against viral infections and also participate in the regulation of normal and malignant hematopoiesis. NK cell activities have been associated with early clearance of viremia in experimental simian immunodeficiency virus and clinical human immunodeficiency virus type 1 (HIV-1) infections. We have previously shown that NK cells function as major cytotoxic effector cells in vaccine-induced immune protection against Friend virus (FV)-induced leukemia, and NK cell depletion totally abrogates the above protective immunity. However, how NK cells recognize retrovirus-infected cells remains largely unclear. The present study demonstrates a correlation between the expression of the products of retinoic acid early transcript-1 (RAE-1) genes in target cells and their susceptibility to killing by NK cells isolated from FV-infected animals. This killing was abrogated by antibodies blocking the NKG2D receptor in vitro. Further, the expression of RAE-1 proteins on erythroblast surfaces increased early after FV inoculation, and administration of an RAE-1-blocking antibody resulted in increased spleen infectious centers and exaggerated pathology, indicating that FV-infected erythroid cells are recognized by NK cells mainly through the NKG2D-RAE-1 interactions in vivo. Enhanced retroviral replication due to host gene-targeting resulted in markedly increased RAE-1 expression in the absence of massive erythroid cell proliferation, indicating a direct role of retroviral replication in RAE-1 upregulation.

Polyploidization on SK-N-MC human neuroblastoma cells infected with herpes simplex virus 1.

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Karalyan, Zaven; Izmailyan, Roza; Karalova, Elena; Abroyan, Liana; Hakobyan, Lina; Avetisyan, Aida; Semerjyan, Zara

2016-01-01

Polyploidization is one of the most dramatic changes occurring within cell genome owing to various reasons including under many viral infections. We examined the impact of herpes simplex virus-1 (HSV-1) on SK-N-MC human neuroblastoma cell line. The infected cells were followed from 6 hours up to 96 hours post infection (hpi). A large number of polyploid cells with giant nuclei was observed under the influence of HSV-1 at 24 hpi with the DNA content of 32c to 64c or more, in comparison with control SK-N-MC cells that were characterized by relatively moderate values of ploidy, i.e. 8с to 16с (where 1c is the haploid amount of nuclear DNA found in normal diploid populations in G0/G1). After 48-96 hpi, the population of polyploid cells with giant nuclei decreased to the benchmark level. The SK-NMC cells infected with HSV-1 for 24 hours were stained with gallocyanine and monitored for cytological features. The infected cells underwent virus induced cellcell and nuclei fusion with the formation of dense nuclei syncytium. The metabolic activity of HSV-1 infected cells was higher in both nuclei and nucleoli when compared to control cells.

Intrinsic Contribution of Perforin to NK-Cell Homeostasis during Mouse Cytomegalovirus Infection

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Maja eArapovic

2016-04-01

Full Text Available In addition to their role as effector cells in virus control, natural killer (NK cells have an immunoregulatory function in shaping the antiviral T-cell response. This function is further pronounced in perforin-deficient mice that show the enhanced NK-cell proliferation and cytokine secretion upon mouse cytomegalovirus (MCMV infection. Here we confirmed that stronger activation and maturation of NK cells in perforin-deficient mice correlates with higher MCMV load. To further characterize the immunoregulatory potential of perforin, we compared the response of NK cells that express or do not express perforin using bone-marrow chimeras. Our results demonstrated that the enhanced proliferation and maturation of NK cells in MCMV-infected bone-marrow chimeras is an intrinsic property of perforin-deficient NK cells. Thus, in addition to confirming that NK-cell proliferation is virus load dependent, our data extend this notion demonstrating that perforin plays an intrinsic role as a feedback mechanism in regulation of NK-cell proliferation during viral infections.

Acute Viral Respiratory Infection Rapidly Induces a CD8+ T Cell Exhaustion-like Phenotype.

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Erickson, John J; Lu, Pengcheng; Wen, Sherry; Hastings, Andrew K; Gilchuk, Pavlo; Joyce, Sebastian; Shyr, Yu; Williams, John V

2015-11-01

Acute viral infections typically generate functional effector CD8(+) T cells (TCD8) that aid in pathogen clearance. However, during acute viral lower respiratory infection, lung TCD8 are functionally impaired and do not optimally control viral replication. T cells also become unresponsive to Ag during chronic infections and cancer via signaling by inhibitory receptors such as programmed cell death-1 (PD-1). PD-1 also contributes to TCD8 impairment during viral lower respiratory infection, but how it regulates TCD8 impairment and the connection between this state and T cell exhaustion during chronic infections are unknown. In this study, we show that PD-1 operates in a cell-intrinsic manner to impair lung TCD8. In light of this, we compared global gene expression profiles of impaired epitope-specific lung TCD8 to functional spleen TCD8 in the same human metapneumovirus-infected mice. These two populations differentially regulate hundreds of genes, including the upregulation of numerous inhibitory receptors by lung TCD8. We then compared the gene expression of TCD8 during human metapneumovirus infection to those in acute or chronic lymphocytic choriomeningitis virus infection. We find that the immunophenotype of lung TCD8 more closely resembles T cell exhaustion late into chronic infection than do functional effector T cells arising early in acute infection. Finally, we demonstrate that trafficking to the infected lung alone is insufficient for TCD8 impairment or inhibitory receptor upregulation, but that viral Ag-induced TCR signaling is also required. Our results indicate that viral Ag in infected lungs rapidly induces an exhaustion-like state in lung TCD8 characterized by progressive functional impairment and upregulation of numerous inhibitory receptors. Copyright © 2015 by The American Association of Immunologists, Inc.

Membrane prteins of herpes simplex infected cells. Immunological and biochemical studies

NARCIS (Netherlands)

Welling-Wester, Sijtske

1981-01-01

As a consequence of infection with herpes simplex virus (HSV), cells exhibit a number of alterations. One of these is expressed as a change in the polypeptide composition of the surface of the infected cells. In this study several methods used for the isolation of these polypeptides expressed on the

[Enhanced lymphocyte proliferation in the presence of epidermal cells of HIV-infected patients in vitro].

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Kappus, R P; Berger, S; Thomas, C A; Ottmann, O G; Ganser, A; Stille, W; Shah, P M

1992-07-01

Clinical observations show that the HIV infection is often associated with affections of the skin. In order to examine the involvement of the epidermal immune system in the HIV infection, we determined accessory cell function of epidermal cells from HIV-1-infected patients. For this we measured the proliferative response of enriched CD(4+)-T-lymphocytes from HIV-infected patients and noninfected controls to stimulation with anti-CD3 and IL-2 in the presence of epidermal cells; the enhancement of the response is dependent on the presence of functionally intact accessory cells. The capacity of epidermal cells to increase the anti-CD3-stimulated T-cell proliferative response was significantly enhanced in HIV patients (CDC III/IVA) as compared with noninfected donors. It is discussed, whether the increased activity of epidermal cells from HIV-infected patients may be responsible for several of the dermal lesions in the course of an HIV infection as due to an enhanced production and release of epidermal cell-derived cytokines.

Functional memory B cells and long-lived plasma cells are generated after a single Plasmodium chabaudi infection in mice.

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Francis Maina Ndungu

2009-12-01

Full Text Available Antibodies have long been shown to play a critical role in naturally acquired immunity to malaria, but it has been suggested that Plasmodium-specific antibodies in humans may not be long lived. The cellular mechanisms underlying B cell and antibody responses are difficult to study in human infections; therefore, we have investigated the kinetics, duration and characteristics of the Plasmodium-specific memory B cell response in an infection of P. chabaudi in mice. Memory B cells and plasma cells specific for the C-terminal region of Merozoite Surface Protein 1 were detectable for more than eight months following primary infection. Furthermore, a classical memory response comprised predominantly of the T-cell dependent isotypes IgG2c, IgG2b and IgG1 was elicited upon rechallenge with the homologous parasite, confirming the generation of functional memory B cells. Using cyclophosphamide treatment to discriminate between long-lived and short-lived plasma cells, we demonstrated long-lived cells secreting Plasmodium-specific IgG in both bone marrow and in spleens of infected mice. The presence of these long-lived cells was independent of the presence of chronic infection, as removal of parasites with anti-malarial drugs had no impact on their numbers. Thus, in this model of malaria, both functional Plasmodium-specific memory B cells and long-lived plasma cells can be generated, suggesting that defects in generating these cell populations may not be the reason for generating short-lived antibody responses.

Role of natural killer cells in innate protection against lethal ebola virus infection.

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Warfield, Kelly L; Perkins, Jeremy G; Swenson, Dana L; Deal, Emily M; Bosio, Catharine M; Aman, M Javad; Yokoyama, Wayne M; Young, Howard A; Bavari, Sina

2004-07-19

Ebola virus is a highly lethal human pathogen and is rapidly driving many wild primate populations toward extinction. Several lines of evidence suggest that innate, nonspecific host factors are potentially critical for survival after Ebola virus infection. Here, we show that nonreplicating Ebola virus-like particles (VLPs), containing the glycoprotein (GP) and matrix protein virus protein (VP)40, administered 1-3 d before Ebola virus infection rapidly induced protective immunity. VLP injection enhanced the numbers of natural killer (NK) cells in lymphoid tissues. In contrast to live Ebola virus, VLP treatment of NK cells enhanced cytokine secretion and cytolytic activity against NK-sensitive targets. Unlike wild-type mice, treatment of NK-deficient or -depleted mice with VLPs had no protective effect against Ebola virus infection and NK cells treated with VLPs protected against Ebola virus infection when adoptively transferred to naive mice. The mechanism of NK cell-mediated protection clearly depended on perforin, but not interferon-gamma secretion. Particles containing only VP40 were sufficient to induce NK cell responses and provide protection from infection in the absence of the viral GP. These findings revealed a decisive role for NK cells during lethal Ebola virus infection. This work should open new doors for better understanding of Ebola virus pathogenesis and direct the development of immunotherapeutics, which target the innate immune system, for treatment of Ebola virus infection.

Foxp3+ regulatory T cells control persistence of viral CNS infection.

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Dajana Reuter

Full Text Available We earlier established a model of a persistent viral CNS infection using two week old immunologically normal (genetically unmodified mice and recombinant measles virus (MV. Using this model infection we investigated the role of regulatory T cells (Tregs as regulators of the immune response in the brain, and assessed whether the persistent CNS infection can be modulated by manipulation of Tregs in the periphery. CD4(+ CD25(+ Foxp3(+ Tregs were expanded or depleted during the persistent phase of the CNS infection, and the consequences for the virus-specific immune response and the extent of persistent infection were analyzed. Virus-specific CD8(+ T cells predominantly recognising the H-2D(b-presented viral hemagglutinin epitope MV-H(22-30 (RIVINREHL were quantified in the brain by pentamer staining. Expansion of Tregs after intraperitoneal (i.p. application of the superagonistic anti-CD28 antibody D665 inducing transient immunosuppression caused increased virus replication and spread in the CNS. In contrast, depletion of Tregs using diphtheria toxin (DT in DEREG (depletion of regulatory T cells-mice induced an increase of virus-specific CD8(+ effector T cells in the brain and caused a reduction of the persistent infection. These data indicate that manipulation of Tregs in the periphery can be utilized to regulate virus persistence in the CNS.

Porcine aminopeptidase N mediated polarized infection by porcine epidemic diarrhea virus in target cells

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Cong, Yingying; Li, Xiaoxue; Bai, Yunyun [College of Veterinary Medicine, Northeast Agricultural University, Harbin 150030 (China); Lv, Xiaonan [College of Veterinary Medicine, Northeast Agricultural University, Harbin 150030 (China); CAS Key Lab for Biomedical Effects of Nanomaterials and Nanosafety, National Center for Nanoscience & Technology of China, Beijing 100090 (China); Herrler, Georg [Institute for Virology, University of Veterinary Medicine, Hannover D-30559 (Germany); Enjuanes, Luis [Department of Molecular and Cell Biology, Centro Nacional de Biotecnología (CNB-CSIC), Campus Universidad Autónoma de Madrid, Cantoblanco, Madrid (Spain); Zhou, Xingdong [College of Veterinary Medicine, Northeast Agricultural University, Harbin 150030 (China); Qu, Bo [Faculty of Life Sciences, Northeast Agricultural University, Harbin 150030 (China); Meng, Fandan [Institute for Virology, University of Veterinary Medicine, Hannover D-30559 (Germany); Cong, Chengcheng [College Animal Husbandry and Veterinary Medicine, Shenyang Agricultural University, Shenyang 110161 (China); Ren, Xiaofeng; Li, Guangxing [College of Veterinary Medicine, Northeast Agricultural University, Harbin 150030 (China)

2015-04-15

Infection of polarized intestinal epithelial cells by porcine epidemic diarrhea virus (PEDV) was characterized. Indirect immunofluorescence assay, real-time PCR, and transmission electron microscopy confirmed PEDV can be successfully propagated in immortalized swine small intestine epithelial cells (IECs). Infection involved porcine aminpeptidase N (pAPN), a reported cellular receptor for PEDV, transient expression of pAPN and siRNA targeted pAPN increased and decreased the infectivity of PEDV in IECs, respectively. Subsequently, polarized entry into and release from both Vero E6 and IECs was analyzed. PEDV entry into polarized cells and pAPN grown on membrane inserts occurs via apical membrane. The progeny virus released into the medium was also quantified which demonstrated that PEDV is preferentially released from the apical membrane. Collectively, our data demonstrate that pAPN, the cellular receptor for PEDV, mediates polarized PEDV infection. These results imply the possibility that PEDV infection may proceed by lateral spread of virus in intestinal epithelial cells. - Highlights: • PEDV infection of polarized intestinal epithelial cells (IECs) was characterized. • Porcine aminpeptidase N (pAPN) facilitated PEDV infection in IECs. • PEDV entry into and release from polarized cell via its apical membrane. • PEDV infection may proceed by lateral spread of virus in IECs.

Porcine aminopeptidase N mediated polarized infection by porcine epidemic diarrhea virus in target cells

International Nuclear Information System (INIS)

Cong, Yingying; Li, Xiaoxue; Bai, Yunyun; Lv, Xiaonan; Herrler, Georg; Enjuanes, Luis; Zhou, Xingdong; Qu, Bo; Meng, Fandan; Cong, Chengcheng; Ren, Xiaofeng; Li, Guangxing

2015-01-01

Infection of polarized intestinal epithelial cells by porcine epidemic diarrhea virus (PEDV) was characterized. Indirect immunofluorescence assay, real-time PCR, and transmission electron microscopy confirmed PEDV can be successfully propagated in immortalized swine small intestine epithelial cells (IECs). Infection involved porcine aminpeptidase N (pAPN), a reported cellular receptor for PEDV, transient expression of pAPN and siRNA targeted pAPN increased and decreased the infectivity of PEDV in IECs, respectively. Subsequently, polarized entry into and release from both Vero E6 and IECs was analyzed. PEDV entry into polarized cells and pAPN grown on membrane inserts occurs via apical membrane. The progeny virus released into the medium was also quantified which demonstrated that PEDV is preferentially released from the apical membrane. Collectively, our data demonstrate that pAPN, the cellular receptor for PEDV, mediates polarized PEDV infection. These results imply the possibility that PEDV infection may proceed by lateral spread of virus in intestinal epithelial cells. - Highlights: • PEDV infection of polarized intestinal epithelial cells (IECs) was characterized. • Porcine aminpeptidase N (pAPN) facilitated PEDV infection in IECs. • PEDV entry into and release from polarized cell via its apical membrane. • PEDV infection may proceed by lateral spread of virus in IECs

Androgen-independent proliferation of LNCaP prostate cancer cells infected by xenotropic murine leukemia virus-related virus

International Nuclear Information System (INIS)

Kakoki, Katsura; Kamiyama, Haruka; Izumida, Mai; Yashima, Yuka; Hayashi, Hideki; Yamamoto, Naoki; Matsuyama, Toshifumi; Igawa, Tsukasa; Sakai, Hideki; Kubo, Yoshinao

2014-01-01

Highlights: • XMRV infection induces androgen-independent growth in LNCaP cells. • XMRV infection reduces expression of androgen receptor. • XMRV promotes appearance of androgen blocker-resistant prostate cancer cells. - Abstract: Xenotropic murine leukemia virus-related virus (XMRV) is a novel gammaretrovirus that was originally isolated from human prostate cancer. It is now believed that XMRV is not the etiologic agent of prostate cancer. An analysis of murine leukemia virus (MLV) infection in various human cell lines revealed that prostate cancer cell lines are preferentially infected by XMRV, and this suggested that XMRV infection may confer some sort of growth advantage to prostate cancer cell lines. To examine this hypothesis, androgen-dependent LNCaP cells were infected with XMRV and tested for changes in certain cell growth properties. We found that XMRV-infected LNCaP cells can proliferate in the absence of the androgen dihydrotestosterone. Moreover, androgen receptor expression is significantly reduced in XMRV-infected LNCaP cells. Such alterations were not observed in uninfected and amphotropic MLV-infected LNCaP cells. This finding explains why prostate cancer cell lines are preferentially infected with XMRV

Induction of T helper 3 regulatory cells by dendritic cells infected with porcine reproductive and respiratory syndrome virus

International Nuclear Information System (INIS)

Silva-Campa, Erika; Flores-Mendoza, Lilian; Resendiz, Monica; Pinelli-Saavedra, Araceli; Mata-Haro, Veronica; Mwangi, Waithaka; Hernandez, Jesus

2009-01-01

Delayed development of virus-specific immune response has been observed in pigs infected with the porcine reproductive and respiratory syndrome virus (PRRSV). Several studies support the hypothesis that the PRRSV is capable of modulating porcine immune system, but the mechanisms involved are yet to be defined. In this study, we evaluated the induction of T regulatory cells by PRRSV-infected dendritic cells (DCs). Our results showed that PRRSV-infected DCs significantly increased Foxp3 + CD25 + T cells, an effect that was reversible by IFN-α treatment, and this outcome was reproducible using two distinct PRRSV strains. Analysis of the expressed cytokines suggested that the induction of Foxp3 + CD25 + T cells is dependent on TGF-β but not IL-10. In addition, a significant up-regulation of Foxp3 mRNA, but not TBX21 or GATA3, was detected. Importantly, our results showed that the induced Foxp3 + CD25 + T cells were able to suppress the proliferation of PHA-stimulated PBMCs. The T cells induced by the PRRSV-infected DCs fit the Foxp3 + CD25 + T helper 3 (Th3) regulatory cell phenotype described in the literature. The induction of this cell phenotype depended, at least in part, on PRRSV viability because IFN-α treatment or virus inactivation reversed these effects. In conclusion, this data supports the hypothesis that the PRRSV succeeds to establish and replicate in porcine cells early post-infection, in part, by inducing Th3 regulatory cells as a mechanism of modulating the porcine immune system.

CCR2+ and CCR5+ CD8+ T cells increase during viral infection and migrate to sites of infection

DEFF Research Database (Denmark)

Nansen, A; Marker, O; Bartholdy, C

2000-01-01

Chemokines and their receptors play a critical role in the selective recruitment of various leukocyte subsets. In this study, we correlated the expression of multiple chemokine and CC chemokine receptor (CCR) genes during the course of intracerebral (i.c.) infection with lymphocytic choriomeningi......Chemokines and their receptors play a critical role in the selective recruitment of various leukocyte subsets. In this study, we correlated the expression of multiple chemokine and CC chemokine receptor (CCR) genes during the course of intracerebral (i.c.) infection with lymphocytic...... a rapidly lethal, T cell-independent encephalitis, and infection resulted in a dramatic early up-regulation of chemokine gene expression. Similar marked up-regulation of chemokine expression was not seen until late after LCMV infection and required the presence of activated T cells. Cerebral CCR gene...... expression was dominated by CCR1, CCR2 and CCR5. However, despite a stronger initial chemokine signal in VSV-infected mice, only LCMV-induced T cell-dependent inflammation was found to be associated with substantially increased expression of CCR genes. Virus-activated CD8+ T cells were found to express CCR2...

Mucosal stromal fibroblasts markedly enhance HIV infection of CD4+ T cells.

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Jason A Neidleman

2017-02-01

Full Text Available Understanding early events of HIV transmission within mucosal tissues is vital for developing effective prevention strategies. Here, we report that primary stromal fibroblasts isolated from endometrium, cervix, foreskin, male urethra, and intestines significantly increase HIV infection of CD4+ T cells-by up to 37-fold for R5-tropic HIV and 100-fold for X4-tropic HIV-without themselves becoming infected. Fibroblasts were more efficient than dendritic cells at trans-infection and mediate this response in the absence of the DC-SIGN and Siglec-1 receptors. In comparison, mucosal epithelial cells secrete antivirals and inhibit HIV infection. These data suggest that breaches in the epithelium allow external or luminal HIV to escape an antiviral environment to access the infection-favorable environment of the stromal fibroblasts, and suggest that resident fibroblasts have a central, but previously unrecognized, role in HIV acquisition at mucosal sites. Inhibiting fibroblast-mediated enhancement of HIV infection should be considered as a novel prevention strategy.

Design optimization of rear uprights for UniMAP Automotive Racing Team Formula SAE racing car

Science.gov (United States)

Azmeer, M.; Basha, M. H.; Hamid, M. F.; Rahman, M. T. A.; Hashim, M. S. M.

2017-10-01

In an automobile, the rear upright are used to provide a physical mounting and links the suspension arms to the hub and wheel assembly. In this work, static structural and shape optimization analysis for rear upright for UniMAP’s Formula SAE racing car had been done using ANSYS software with the objective to reduce weight while maintaining the structural strength of the vehicle upright. During the shape optimization process, the component undergoes 25%, 50% and 75 % weight reduction in order to find the best optimal shape of the upright. The final design of the upright is developed considering the weight reduction, structural integrity and the manufacturability. The final design achieved 21 % weight reduction and is able to withstand several loads.

Chloroquine, an Endocytosis Blocking Agent, Inhibits Zika Virus Infection in Different Cell Models.

Science.gov (United States)

Delvecchio, Rodrigo; Higa, Luiza M; Pezzuto, Paula; Valadão, Ana Luiza; Garcez, Patrícia P; Monteiro, Fábio L; Loiola, Erick C; Dias, André A; Silva, Fábio J M; Aliota, Matthew T; Caine, Elizabeth A; Osorio, Jorge E; Bellio, Maria; O'Connor, David H; Rehen, Stevens; de Aguiar, Renato Santana; Savarino, Andrea; Campanati, Loraine; Tanuri, Amilcar

2016-11-29

Zika virus (ZIKV) infection in utero might lead to microcephaly and other congenital defects. Since no specific therapy is available thus far, there is an urgent need for the discovery of agents capable of inhibiting its viral replication and deleterious effects. Chloroquine is widely used as an antimalarial drug, anti-inflammatory agent, and it also shows antiviral activity against several viruses. Here we show that chloroquine exhibits antiviral activity against ZIKV in Vero cells, human brain microvascular endothelial cells, human neural stem cells, and mouse neurospheres. We demonstrate that chloroquine reduces the number of ZIKV-infected cells in vitro, and inhibits virus production and cell death promoted by ZIKV infection without cytotoxic effects. In addition, chloroquine treatment partially reveres morphological changes induced by ZIKV infection in mouse neurospheres.

Chloroquine, an Endocytosis Blocking Agent, Inhibits Zika Virus Infection in Different Cell Models

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Rodrigo Delvecchio

2016-11-01

Full Text Available Zika virus (ZIKV infection in utero might lead to microcephaly and other congenital defects. Since no specific therapy is available thus far, there is an urgent need for the discovery of agents capable of inhibiting its viral replication and deleterious effects. Chloroquine is widely used as an antimalarial drug, anti-inflammatory agent, and it also shows antiviral activity against several viruses. Here we show that chloroquine exhibits antiviral activity against ZIKV in Vero cells, human brain microvascular endothelial cells, human neural stem cells, and mouse neurospheres. We demonstrate that chloroquine reduces the number of ZIKV-infected cells in vitro, and inhibits virus production and cell death promoted by ZIKV infection without cytotoxic effects. In addition, chloroquine treatment partially reveres morphological changes induced by ZIKV infection in mouse neurospheres.

Chloroquine, an Endocytosis Blocking Agent, Inhibits Zika Virus Infection in Different Cell Models

Science.gov (United States)

Delvecchio, Rodrigo; Higa, Luiza M.; Pezzuto, Paula; Valadão, Ana Luiza; Garcez, Patrícia P.; Monteiro, Fábio L.; Loiola, Erick C.; Dias, André A.; Silva, Fábio J. M.; Aliota, Matthew T.; Caine, Elizabeth A.; Osorio, Jorge E.; Bellio, Maria; O’Connor, David H.; Rehen, Stevens; de Aguiar, Renato Santana; Savarino, Andrea; Campanati, Loraine; Tanuri, Amilcar

2016-01-01

Zika virus (ZIKV) infection in utero might lead to microcephaly and other congenital defects. Since no specific therapy is available thus far, there is an urgent need for the discovery of agents capable of inhibiting its viral replication and deleterious effects. Chloroquine is widely used as an antimalarial drug, anti-inflammatory agent, and it also shows antiviral activity against several viruses. Here we show that chloroquine exhibits antiviral activity against ZIKV in Vero cells, human brain microvascular endothelial cells, human neural stem cells, and mouse neurospheres. We demonstrate that chloroquine reduces the number of ZIKV-infected cells in vitro, and inhibits virus production and cell death promoted by ZIKV infection without cytotoxic effects. In addition, chloroquine treatment partially reveres morphological changes induced by ZIKV infection in mouse neurospheres. PMID:27916837

Comparative proteomics analysis of Spodoptera frugiperda cells during Autographa californica multiple nucleopolyhedrovirus infection.

Science.gov (United States)

Yu, Qian; Xiong, Youhua; Gao, Hang; Liu, Jianliang; Chen, Zhiqiang; Wang, Qin; Wen, Dongling

2015-08-04

Increasing evidence sugggest that in addition of balculovirus controling insect host, host cells also responds to balculovirus infection. However, compared to existing knowledge on virus gene, host cell responses are relatively poorly understood. In this study, Spodoptera frugiperda (Sf9) cells were infected with Autographa californica multiple nucleopolyhedrovirus (AcMNPV). The protein composition and protein changes of Spodoptera frugiperda (Sf9) cells of different infection stages were analysed by isobaric tag for relative and absolute quantification (iTRAQ) techniques. A total of 4004 Sf9 proteins were identified by iTRAQ and 413 proteins were found as more than 1.5-fold changes in abundance. The 413 proteins were categorised according to GO classification for insects and were categorised into: biological process, molecular function and cellular component. The determination of the protein changes in infected Sf9 cells would help to better understanding of host cell responses and facilitate better design of this virus-host cell interaction in pest insect control and other related fields.

Reduction of prion infectivity in packed red blood cells

International Nuclear Information System (INIS)

Morales, Rodrigo; Buytaert-Hoefen, Kimberley A.; Gonzalez-Romero, Dennisse; Castilla, Joaquin; Hansen, Eric T.; Hlavinka, Dennis; Goodrich, Raymond P.; Soto, Claudio

2008-01-01

The link between a new variant form of Creutzfeldt-Jakob disease (vCJD) and the consumption of prion contaminated cattle meat as well as recent findings showing that vCJD can be transmitted by blood transfusion have raised public health concerns. Currently, a reliable test to identify prions in blood samples is not available. The purpose of this study was to evaluate the possibility to remove scrapie prion protein (PrP Sc ) and infectivity from red blood cell (RBC) suspensions by a simple washing procedure using a cell separation and washing device. The extent of prion removal was assessed by Western blot, PMCA and infectivity bioassays. Our results revealed a substantial removal of infectious prions (≥3 logs of infectivity) by all techniques used. These data suggest that a significant amount of infectivity present in RBC preparations can be removed by a simple washing procedure. This technology may lead to increased safety of blood products and reduce the risk of further propagation of prion diseases.

FRET imaging of hemoglobin concentration in Plasmodium falciparum-infected red cells.

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Alessandro Esposito

Full Text Available During its intraerythrocytic asexual reproduction cycle Plasmodium falciparum consumes up to 80% of the host cell hemoglobin, in large excess over its metabolic needs. A model of the homeostasis of falciparum-infected red blood cells suggested an explanation based on the need to reduce the colloid-osmotic pressure within the host cell to prevent its premature lysis. Critical for this hypothesis was that the hemoglobin concentration within the host cell be progressively reduced from the trophozoite stage onwards.The experiments reported here were designed to test this hypothesis by direct measurements of the hemoglobin concentration in live, infected red cells. We developed a novel, non-invasive method to quantify the hemoglobin concentration in single cells, based on Förster resonance energy transfer between hemoglobin molecules and the fluorophore calcein. Fluorescence lifetime imaging allowed the quantitative mapping of the hemoglobin concentration within the cells. The average fluorescence lifetimes of uninfected cohorts was 270+/-30 ps (mean+/-SD; N = 45. In the cytoplasm of infected cells the fluorescence lifetime of calcein ranged from 290+/-20 ps for cells with ring stage parasites to 590+/-13 ps and 1050+/-60 ps for cells with young trophozoites and late stage trophozoite/early schizonts, respectively. This was equivalent to reductions in hemoglobin concentration spanning the range from 7.3 to 2.3 mM, in line with the model predictions. An unexpected ancillary finding was the existence of a microdomain under the host cell membrane with reduced calcein quenching by hemoglobin in cells with mature trophozoite stage parasites.The results support the predictions of the colloid-osmotic hypothesis and provide a better understanding of the homeostasis of malaria-infected red cells. In addition, they revealed the existence of a distinct peripheral microdomain in the host cell with limited access to hemoglobin molecules indicating the

Changes in NK and NKT cells in mesenteric lymph nodes after a Schistosoma japonicum infection.

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Luo, Xueping; Xie, Hongyan; Chen, Dianhui; Yu, Xiuxue; Wu, Fan; Li, Lu; Wu, Changyou; Huang, Jun

2014-03-01

The mesenteric lymph node (MLN) is the main draining lymph node in mouse enterocoelia, which contains many types of immune cells. Among these cells, natural killer (NK) and natural killer T (NKT) cells belong to innate lymphoid cells (ILCs), which have potent activities for controlling a variety of pathogenic infections. In this study, C57BL/6 mice were infected with Schistosoma japonicum for 5-7 weeks. Lymphocytes were isolated from the MLN to detect changes in the phenotype and function of NK and NKT cells using a fluorescence activating cell sorter (FACS). These results demonstrated that a S. japonicum infection could significantly increase the percentage of NK cells in the mouse MLN, (P cell number of both NK and NKT cells. In addition, we found that NK and NKT cells from infected mice expressed higher levels of CD69 compared to normal mice (P NKT cell activation. Moreover, we found that the expression of CD4 was increased in infected MLN NK cells (P NKT cells of infected mice after phorbol 12-myristate 13-acetate (PMA) and ionomycin stimulation (P NKT cells might play roles in modulating the classical T cell response. Finally, our results indicated that the expression of CD94 was decreased in NK cells, suggesting that the downregulation of CD94 expression might served as a mechanism in NK cell activation.

Tonsillar CD56brightNKG2A+ NK cells restrict primary Epstein-Barr virus infection in B cells via IFN-γ.

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Jud, Aurelia; Kotur, Monika; Berger, Christoph; Gysin, Claudine; Nadal, David; Lünemann, Anna

2017-01-24

Natural killer (NK) cells constitute the first line of defense against viruses and cancers cells. Epstein-Barr virus (EBV) was the first human virus to be directly implicated in carcinogenesis, and EBV infection is associated with a broad spectrum of B cell lymphomas. How NK cells restrict EBV-associated oncogenesis is not understood. Here, we investigated the efficacies and mechanisms of distinct NK cell subsets from tonsils, the portal of entry of EBV, in limiting EBV infection in naïve, germinal center-associated and memory B cells. We found that CD56bright and NKG2A expression sufficiently characterizes the potent anti-EBV capacity of tonsillar NK cells. We observed restriction of EBV infection in B cells as early as 18 hours after infection. The restriction was most efficient in naïve B cells and germinal center-associated B cells, the B cell subsets that exhibited highest susceptibility to EBV infection in vitro. IFN-γ release by and partially NKp44 engagement of CD56bright and NKG2A positive NK cells mediated the restriction that eventually inhibited B-cell transformation. Thus, harnessing CD56brightNKG2A+ NK cell function might be promising to improve treatment strategies that target EBV-associated B cell lymphomas.

A three-dimensional comparison of tick-borne flavivirus infection in mammalian and tick cell lines.

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Danielle K Offerdahl

Full Text Available Tick-borne flaviviruses (TBFV are sustained in nature through cycling between mammalian and tick hosts. In this study, we used African green monkey kidney cells (Vero and Ixodes scapularis tick cells (ISE6 to compare virus-induced changes in mammalian and arthropod cells. Using confocal microscopy, transmission electron microscopy (TEM, and electron tomography (ET, we examined viral protein distribution and the ultrastructural changes that occur during TBFV infection. Within host cells, flaviviruses cause complex rearrangement of cellular membranes for the purpose of virus replication. Virus infection was accompanied by a marked expansion in endoplasmic reticulum (ER staining and markers for TBFV replication were localized mainly to the ER in both cell lines. TEM of Vero cells showed membrane-bound vesicles enclosed in a network of dilated, anastomosing ER cisternae. Virions were seen within the ER and were sometimes in paracrystalline arrays. Tubular structures or elongated vesicles were occasionally noted. In acutely and persistently infected ISE6 cells, membrane proliferation and vesicles were also noted; however, the extent of membrane expansion and the abundance of vesicles were lower and no viral particles were observed. Tubular profiles were far more prevalent in persistently infected ISE6 cells than in acutely infected cells. By ET, tubular profiles, in persistently infected tick cells, had a cross-sectional diameter of 60-100 nm, reached up to 800 nm in length, were closed at the ends, and were often arranged in fascicle-like bundles, shrouded with ER membrane. Our experiments provide analysis of viral protein localization within the context of both mammalian and arthropod cell lines as well as both acute and persistent arthropod cell infection. Additionally, we show for the first time 3D flavivirus infection in a vector cell line and the first ET of persistent flavivirus infection.

HIV-1 isolation from infected peripheral blood mononuclear cells.

Science.gov (United States)

Dispinseri, Stefania; Saba, Elisa; Vicenzi, Elisa; Kootstra, Neeltje A; Schuitemaker, Hanneke; Scarlatti, Gabriella

2014-01-01

Human immunodeficiency virus 1 (HIV-1) isolation from peripheral blood mononuclear cells (PBMCs) allows retrieval of replication-competent viral variants. In order to impose the smallest possible selective pressure on the viral isolates, isolation must be carried out in primary cultures of cells and not in tumor derived cell lines. The procedure involves culture of PBMCs from an infected patient with phytohemagglutinin (PHA)-stimulated PBMC from seronegative donors, which provide susceptible target cells for HIV replication. HIV can be isolated from the bulk population of PBMCs or after cloning of the cells to obtain viral biological clones. Viral production is determined with p24 antigen (Ag) detection assays or with reverse transcriptase (RT) activity assay. Once isolated, HIV-1 can be propagated by infecting PHA-stimulated PBMCs from healthy donors. Aliquots from culture with a high production of virus are stored for later use.

High-Throughput Sequencing of MicroRNAs in Adenovirus Type 3 Infected Human Laryngeal Epithelial Cells

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Yuhua Qi

2010-01-01

Full Text Available Adenovirus infection can cause various illnesses depending on the infecting serotype, such as gastroenteritis, conjunctivitis, cystitis, and rash illness, but the infection mechanism is still unknown. MicroRNAs (miRNA have been reported to play essential roles in cell proliferation, cell differentiation, and pathogenesis of human diseases including viral infections. We analyzed the miRNA expression profiles from adenovirus type 3 (AD3 infected Human laryngeal epithelial (Hep2 cells using a SOLiD deep sequencing. 492 precursor miRNAs were identified in the AD3 infected Hep2 cells, and 540 precursor miRNAs were identified in the control. A total of 44 miRNAs demonstrated high expression and 36 miRNAs showed lower expression in the AD3 infected cells than control. The biogenesis of miRNAs has been analyzed, and some of the SOLiD results were confirmed by Quantitative PCR analysis. The present studies may provide a useful clue for the biological function research into AD3 infection.

Detection and quantification of poliovirus infection using FTIR spectroscopy and cell culture

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Lee-Montiel Felipe T

2011-12-01

Full Text Available Abstract Background In a globalized word, prevention of infectious diseases is a major challenge. Rapid detection of viable virus particles in water and other environmental samples is essential to public health risk assessment, homeland security and environmental protection. Current virus detection methods, especially assessing viral infectivity, are complex and time-consuming, making point-of-care detection a challenge. Faster, more sensitive, highly specific methods are needed to quantify potentially hazardous viral pathogens and to determine if suspected materials contain viable viral particles. Fourier transform infrared (FTIR spectroscopy combined with cellular-based sensing, may offer a precise way to detect specific viruses. This approach utilizes infrared light to monitor changes in molecular components of cells by tracking changes in absorbance patterns produced following virus infection. In this work poliovirus (PV1 was used to evaluate the utility of FTIR spectroscopy with cell culture for rapid detection of infective virus particles. Results Buffalo green monkey kidney (BGMK cells infected with different virus titers were studied at 1 - 12 hours post-infection (h.p.i.. A partial least squares (PLS regression method was used to analyze and model cellular responses to different infection titers and times post-infection. The model performs best at 8 h.p.i., resulting in an estimated root mean square error of cross validation (RMSECV of 17 plaque forming units (PFU/ml when using low titers of infection of 10 and 100 PFU/ml. Higher titers, from 103 to 106 PFU/ml, could also be reliably detected. Conclusions This approach to poliovirus detection and quantification using FTIR spectroscopy and cell culture could potentially be extended to compare biochemical cell responses to infection with different viruses. This virus detection method could feasibly be adapted to an automated scheme for use in areas such as water safety monitoring and

Protective Role of Cross-Reactive CD8 T Cells Against Dengue Virus Infection

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Annie Elong Ngono

2016-11-01

Full Text Available Infection with one of the four dengue virus serotypes (DENV1-4 presumably leads to lifelong immunity against the infecting serotype but not against heterotypic reinfection, resulting in a greater risk of developing Dengue Hemorrhagic Fever/Dengue Shock Syndrome (DHF/DSS during secondary infection. Both antibodies and T cell responses have been implicated in DHF/DSS pathogenesis. According to the T cell-based hypothesis termed “original antigenic sin,” secondary DENV infection is dominated by non-protective, cross-reactive T cells that elicit an aberrant immune response. The goal of our study was to compare the roles of serotype-specific and cross-reactive T cells in protection vs. pathogenesis during DENV infection in vivo. Specifically, we utilized IFN-α/βR−/− HLA*B0702 transgenic mice in the context of peptide vaccination with relevant human CD8 T cell epitopes. IFN-α/βR−/− HLA*B0702 transgenic mice were immunized with DENV serotype 2 (DENV2-specific epitopes or variants found in any of the other three serotypes (DENV1, DENV3 or DENV4, followed by challenge with DENV. Although cross-reactive T cell responses were lower than responses elicited by serotype-specific T cells, immunization with either serotype-specific or variant peptide epitopes enhanced viral clearance, demonstrating that both serotype-specific and cross-reactive T cells can contribute to protection in vivo against DENV infection.

Transfusion associated hepatitis B virus infection among sickle cell ...

African Journals Online (AJOL)

Background: Transfusion of blood products is a recognised way of transmitting infections particularly viruses. The extent to which blood transfusion contributes to hepatitis B virus (HBV) infections in transfused patients with sickle cell anaemia (SCA) has been found to be 20% in Lagos, Nigeria. Mamman in Zaria however ...