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1.
Sci Adv ; 7(6)2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33547084

RESUMO

The ongoing unprecedented severe acute respiratory syndrome caused by the SARS-CoV-2 outbreak worldwide has highlighted the need for understanding viral-host interactions involved in mechanisms of virulence. Here, we show that the virulence factor Nsp1 protein of SARS-CoV-2 interacts with the host messenger RNA (mRNA) export receptor heterodimer NXF1-NXT1, which is responsible for nuclear export of cellular mRNAs. Nsp1 prevents proper binding of NXF1 to mRNA export adaptors and NXF1 docking at the nuclear pore complex. As a result, a significant number of cellular mRNAs are retained in the nucleus during infection. Increased levels of NXF1 rescues the Nsp1-mediated mRNA export block and inhibits SARS-CoV-2 infection. Thus, antagonizing the Nsp1 inhibitory function on mRNA export may represent a strategy to restoring proper antiviral host gene expression in infected cells.


Assuntos
COVID-19/metabolismo , Expressão Gênica , Interações entre Hospedeiro e Microrganismos/genética , RNA Mensageiro/metabolismo , SARS-CoV-2/metabolismo , Proteínas não Estruturais Virais/metabolismo , Fatores de Virulência/metabolismo , Transporte Ativo do Núcleo Celular/genética , Animais , COVID-19/virologia , Chlorocebus aethiops , Células HEK293 , Humanos , Poro Nuclear/metabolismo , Proteínas de Transporte Nucleocitoplasmático/genética , Proteínas de Transporte Nucleocitoplasmático/metabolismo , RNA Mensageiro/genética , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , SARS-CoV-2/química , Transfecção , Células Vero , Proteínas não Estruturais Virais/genética
2.
Proc Natl Acad Sci U S A ; 117(45): 28344-28354, 2020 11 10.
Artigo em Inglês | MEDLINE | ID: mdl-33097660

RESUMO

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the ongoing coronavirus disease 2019 (COVID-19) pandemic that is a serious global health problem. Evasion of IFN-mediated antiviral signaling is a common defense strategy that pathogenic viruses use to replicate and propagate in their host. In this study, we show that SARS-CoV-2 is able to efficiently block STAT1 and STAT2 nuclear translocation in order to impair transcriptional induction of IFN-stimulated genes (ISGs). Our results demonstrate that the viral accessory protein Orf6 exerts this anti-IFN activity. We found that SARS-CoV-2 Orf6 localizes at the nuclear pore complex (NPC) and directly interacts with Nup98-Rae1 via its C-terminal domain to impair docking of cargo-receptor (karyopherin/importin) complex and disrupt nuclear import. In addition, we show that a methionine-to-arginine substitution at residue 58 impairs Orf6 binding to the Nup98-Rae1 complex and abolishes its IFN antagonistic function. All together our data unravel a mechanism of viral antagonism in which a virus hijacks the Nup98-Rae1 complex to overcome the antiviral action of IFN.


Assuntos
COVID-19/metabolismo , Interferons/metabolismo , Complexo de Proteínas Formadoras de Poros Nucleares/metabolismo , Poro Nuclear/metabolismo , Fator de Transcrição STAT1/metabolismo , Fator de Transcrição STAT2/metabolismo , Proteínas Virais/metabolismo , Transporte Ativo do Núcleo Celular , Animais , Sítios de Ligação , Chlorocebus aethiops , Células HEK293 , Humanos , Proteínas Associadas à Matriz Nuclear/química , Proteínas Associadas à Matriz Nuclear/metabolismo , Proteínas de Transporte Nucleocitoplasmático/química , Proteínas de Transporte Nucleocitoplasmático/metabolismo , Ligação Proteica , Transdução de Sinais , Células Vero
3.
PLoS Pathog ; 16(4): e1008407, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32240278

RESUMO

Influenza A viruses are human pathogens with limited therapeutic options. Therefore, it is crucial to devise strategies for the identification of new classes of antiviral medications. The influenza A virus genome is constituted of 8 RNA segments. Two of these viral RNAs are transcribed into mRNAs that are alternatively spliced. The M1 mRNA encodes the M1 protein but is also alternatively spliced to yield the M2 mRNA during infection. M1 to M2 mRNA splicing occurs at nuclear speckles, and M1 and M2 mRNAs are exported to the cytoplasm for translation. M1 and M2 proteins are critical for viral trafficking, assembly, and budding. Here we show that gene knockout of the cellular protein NS1-BP, a constituent of the M mRNA speckle-export pathway and a binding partner of the virulence factor NS1 protein, inhibits M mRNA nuclear export without altering bulk cellular mRNA export, providing an avenue to preferentially target influenza virus. We performed a high-content, image-based chemical screen using single-molecule RNA-FISH to label viral M mRNAs followed by multistep quantitative approaches to assess cellular mRNA and cell toxicity. We identified inhibitors of viral mRNA biogenesis and nuclear export that exhibited no significant activity towards bulk cellular mRNA at non-cytotoxic concentrations. Among the hits is a small molecule that preferentially inhibits nuclear export of a subset of viral and cellular mRNAs without altering bulk cellular mRNA export. These findings underscore specific nuclear export requirements for viral mRNAs and phenocopy down-regulation of the mRNA export factor UAP56. This RNA export inhibitor impaired replication of diverse influenza A virus strains at non-toxic concentrations. Thus, this screening strategy yielded compounds that alone or in combination may serve as leads to new ways of treating influenza virus infection and are novel tools for studying viral RNA trafficking in the nucleus.


Assuntos
Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Antivirais/farmacologia , Núcleo Celular/virologia , Vírus da Influenza A/metabolismo , Influenza Humana/virologia , RNA Mensageiro/metabolismo , RNA Viral/metabolismo , Avaliação Pré-Clínica de Medicamentos , Humanos , Vírus da Influenza A/genética , RNA Mensageiro/genética , RNA Viral/genética , Replicação Viral/efeitos dos fármacos
4.
Protein Cell ; 10(5): 315-326, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30242641

RESUMO

Many viruses, enveloped or non-enveloped, remodel host membrane structures for their replication, assembly and escape from host cells. Herpesviruses are important human pathogens and cause many diseases. As large enveloped DNA viruses, herpesviruses undergo several complex steps to complete their life cycles and produce infectious progenies. Firstly, herpesvirus assembly initiates in the nucleus, producing nucleocapsids that are too large to cross through the nuclear pores. Nascent nucleocapsids instead bud at the inner nuclear membrane to form primary enveloped virions in the perinuclear space followed by fusion of the primary envelopes with the outer nuclear membrane, to translocate the nucleocapsids into the cytoplasm. Secondly, nucleocapsids obtain a series of tegument proteins in the cytoplasm and bud into vesicles derived from host organelles to acquire viral envelopes. The vesicles are then transported to and fuse with the plasma membrane to release the mature virions to the extracellular space. Therefore, at least two budding and fusion events take place at cellular membrane structures during herpesviruses assembly and egress, which induce membrane deformations. In this review, we describe and discuss how herpesviruses exploit and remodel host membrane structures to assemble and escape from the host cell.


Assuntos
Infecções por Herpesviridae/virologia , Herpesviridae/fisiologia , Interações entre Hospedeiro e Microrganismos/fisiologia , Membrana Nuclear/metabolismo , Nucleocapsídeo/metabolismo , Animais , Humanos , Proteínas Virais/metabolismo , Montagem de Vírus , Liberação de Vírus
5.
Sci Rep ; 5: 13501, 2015 Aug 27.
Artigo em Inglês | MEDLINE | ID: mdl-26310236

RESUMO

The gas flow in shale matrix is of great research interests for optimized shale gas extraction. The gas flow in the nano-scale pore may fall in flow regimes such as viscous flow, slip flow and Knudsen diffusion. A 3-dimensional nano-scale pore network model was developed to simulate dynamic gas flow, and to describe the transient properties of flow regimes. The proposed pore network model accounts for the various size distributions and low connectivity of shale pores. The pore size, pore throat size and coordination number obey normal distribution, and the average values can be obtained from shale reservoir data. The gas flow regimes were simulated using an extracted pore network backbone. The numerical results show that apparent permeability is strongly dependent on pore pressure in the reservoir and pore throat size, which is overestimated by low-pressure laboratory tests. With the decrease of reservoir pressure, viscous flow is weakening, then slip flow and Knudsen diffusion are gradually becoming dominant flow regimes. The fingering phenomenon can be predicted by micro/nano-pore network for gas flow, which provides an effective way to capture heterogeneity of shale gas reservoir.

6.
Cell Microbiol ; 17(11): 1583-93, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-25939747

RESUMO

The matrix protein 1 (M1) is the most abundant structural protein in influenza A virus particles. It oligomerizes to form the matrix layer under the lipid membrane, sustaining stabilization of the morphology of the virion. The present study indicates that M1 forms oligomers based on a fourfold symmetrical oligomerization pattern. Further analysis revealed that the oligomerization pattern of M1 was controlled by a highly conserved region within the C-terminal domain. Two polar residues of this region, serine-183 (S183) and threonine-185 (T185), were identified to be critical for the oligomerization pattern of M1. M1 point mutants suggest that single S183A or T185A substitution could result in the production of morphologically filamentous particles, while double substitutions, M1-S183A/T185A, totally disrupted the fourfold symmetry and resulted in the failure of virus production. These data indicate that the polar groups in these residues are essential to control the oligomerization pattern of M1. Thus, the present study will aid in determining the mechanisms of influenza A virus matrix layer formation during virus morphogenesis.


Assuntos
Vírus da Influenza A/fisiologia , Proteínas da Matriz Viral/metabolismo , Vírion/metabolismo , Montagem de Vírus , Aminoácidos/genética , Animais , Linhagem Celular , Análise Mutacional de DNA , Cães , Humanos , Vírus da Influenza A/genética , Mutação Puntual , Multimerização Proteica , Proteínas da Matriz Viral/genética
7.
J Virol ; 88(13): 7455-63, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24741105

RESUMO

UNLABELLED: The influenza A virus nuclear export protein (NEP) plays crucial roles in the nuclear export of the viral ribonucleoprotein complex through the chromosome region maintenance 1 (CRM1)-mediated cellular protein transport system. However, the detailed mechanism of NEP nucleocytoplasmic trafficking remains incompletely understood. Here, we investigated the subcellular localization of NEP from two strains of H1N1 influenza A virus and found that 2009 swine-origin H1N1 influenza A virus A/California/04/2009 (CA04) NEP displayed a distinct cellular distribution pattern, forming unique nuclear aggregates, compared to A/WSN/33 (H1N1) (WSN) NEP. Characterization of the nucleocytoplasmic transport pathways of these two NEPs showed that they both enter the nucleus by passive diffusion but are exported through the nuclear export receptor CRM1-mediated pathway with different efficiencies. The two identified nuclear export signals (NESs) on the two NEPs functioned similarly despite differences in their amino acid sequences. Using a two-hybrid assay, we confirmed that the CA04 NEP interacts less efficiently with CRM1 and that a threonine residue at position 48 is responsible for the nuclear aggregation. The present study revealed the dissimilarity in subcellular NEP transport processes between the 2009 pandemic (H1N1) influenza A virus CA04 and the laboratory-adapted H1N1 virus WSN and uncovered the mechanism responsible for this difference. IMPORTANCE: Because the efficiency of the nucleocytoplasmic transport of viral components is often correlated with the viral RNA polymerase activity, propagation, and host range of influenza viruses, the present study investigated the subcellular localization of NEP from two strains of H1N1 influenza virus. We found that the NEPs of both A/California/04/2009 (H1N1) (CA04) and A/WSN/33 (H1N1) (WSN) enter the nucleus by passive diffusion but are exported with different efficiencies, which were caused by weaker binding activity between the CA04 NEP and CRM1. The results of the present study revealed characteristics of the nuclear import and export pathways of NEP and the mechanism responsible for the differences in the cellular distribution of NEP between two H1N1 strains.


Assuntos
Transporte Ativo do Núcleo Celular/fisiologia , Vírus da Influenza A Subtipo H1N1/fisiologia , Influenza Humana/metabolismo , Carioferinas/metabolismo , Sinais de Exportação Nuclear/fisiologia , Receptores Citoplasmáticos e Nucleares/metabolismo , Núcleo Celular/metabolismo , Humanos , Vírus da Influenza A Subtipo H1N1/classificação , Influenza Humana/virologia , Microscopia de Fluorescência , Frações Subcelulares , Técnicas do Sistema de Duplo-Híbrido , Replicação Viral/fisiologia
8.
J Virol ; 86(9): 4970-80, 2012 May.
Artigo em Inglês | MEDLINE | ID: mdl-22345439

RESUMO

The nuclear export of the influenza A virus ribonucleoprotein (vRNP) is crucial for virus replication. As a major component of the vRNP, nucleoprotein (NP) alone can also be shuttled out of the nucleus by interacting with chromosome region maintenance 1 (CRM1) and is therefore hypothesized to promote the nuclear export of the vRNP. In the present study, three novel nuclear export signals (NESs) of the NP--NES1, NES2, and NES3--were identified as being responsible for mediating its nuclear export. The nuclear export of NES3 was CRM1 dependent, whereas that of NES1 or NES2 was CRM1 independent. Inactivation of these NESs led to an overall nuclear accumulation of NP. Mutation of all three NP-NESs significantly impaired viral replication. Based on structures of influenza virus NP oligomers, these three hydrophobic NESs are found present on the surface of oligomeric NPs. Functional studies indicated that oligomerization is also required for nuclear export of NP. Together, these results suggest that the nuclear export of NP is important for virus replication and relies on its NESs and oligomerization.


Assuntos
Vírus da Influenza A/metabolismo , Sinais de Exportação Nuclear , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/metabolismo , Proteínas do Core Viral/química , Proteínas do Core Viral/metabolismo , Transporte Ativo do Núcleo Celular , Motivos de Aminoácidos , Sequência de Aminoácidos , Animais , Núcleo Celular/metabolismo , Cães , Expressão Gênica , Humanos , Vírus da Influenza A/genética , Carioferinas/metabolismo , Camundongos , Dados de Sequência Molecular , Mutação , Proteínas do Nucleocapsídeo , Multimerização Proteica , Transporte Proteico , Processamento Pós-Transcricional do RNA , Proteínas de Ligação a RNA/genética , Receptores Citoplasmáticos e Nucleares/metabolismo , Proteínas do Core Viral/genética , Replicação Viral/genética
9.
J Environ Monit ; 13(9): 2443-9, 2011 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-21755071

RESUMO

Air sparging (AS) is one of the groundwater remediation techniques for remediating volatile organic compounds (VOCs) in saturated soil. However, in spite of the success of air sparging as a remediation technique for the cleanup of contaminated soils, to date, the fundamental mechanisms or the physics of air flow through porous media is not well understood. In this study, centrifugal modeling tests were performed to investigate air flow rates and the evolution of the zone of influence during the air sparging under various g-levels. The test results show that with the increase in sparging pressure the mass flow rate of the air sparging volume increases. The air mass flow rate increases linearly with the effective sparging pressure ratio, which is the difference between sparging pressure and hydrostatic pressure normalized with respect to the effective overburden pressure at the sparging point. Also the slope of mass flow rate with effective sparging pressure ratio increases with higher g-levels. This variation of the slope of mass flow rate of air sparging volume versus effective sparging pressure ratio, M, is linear with g-level confirming that the air flow through soil for a given effective sparging pressure ratio only depends on the g-level. The test results also show that with increasing sparging pressure, the zone of influence (ZOI), which consists of the width at the tip of the cone or lateral intrusion and the cone angle, will lead to an increase in both lateral intrusion and the cone angle. With a further increase in air injection pressure, the cone angle reaches a constant value while the lateral intrusion becomes the main contributor to the enlargement of the ZOI. However, beyond a certain value of effective sparging pressure ratio, there is no further enlargement of the ZOI.


Assuntos
Recuperação e Remediação Ambiental/métodos , Água Subterrânea/química , Poluentes Químicos da Água/química , Ar/análise , Movimentos do Ar , Centrifugação , Pressão , Solo/química , Compostos Orgânicos Voláteis/química
10.
Environ Sci Technol ; 44(10): 3883-8, 2010 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-20426462

RESUMO

Air sparging (AS) is one of the most efficient techniques for remediating saturated soils and groundwater contaminated with volatile organic compounds. A series of physical modeling tests for different sizes of porous media under varied injection pressure were conducted to investigate the effect of particle size and air injection pressure on size and shape of the zone of influence (ZOI). The test results show that ZOI can be expressed by two components: the horizontal expansion due to pneumatic fracture or preferential intrusion around the injection point and the angle of ZOI which is the angle between the vertical line and the boundary of ZOI. There exists a limited angle of ZOI for each type of porous media. The measured minimum and maximum air injection pressures in 1g tests are compared with corresponding theoretical values, and it is found that the measured minimum injection pressure is slightly lower than the theoretical value, while the measured maximum injection pressure is much higher than the theoretical maximum injection pressure. Centrifugal test results confirmed nonapplicability of theoretical maximum injection pressure to air sparging design. All of the above provide valuable information for design and theoretical modeling of air sparging for groundwater remediation.


Assuntos
Poluentes Atmosféricos/isolamento & purificação , Recuperação e Remediação Ambiental/métodos , Modelos Teóricos
11.
Vaccine ; 27(21): 2741-5, 2009 May 11.
Artigo em Inglês | MEDLINE | ID: mdl-19428887

RESUMO

An effective vaccine of animals can block transmission of Toxoplasma gondii to humans. In this study, mice have been protected against lethal T. gondii challenge by a prime-boost vaccination strategy using DNA vaccine pVAX/TgSAG1 and recombinant pseudorabies virus rPRV/TgSAG1, both expressing the major immunodominant surface antigen of T. gondii (TgSAG1). High levels of splenocyte proliferative responses and significant levels of IFN-gamma resulted, with strong cytotoxic T lymphocyte (CTL) responses in vitro. After lethal challenge, prime-boost vaccinated mice showed an increased survival time (15.4+/-5.0 days) and a 40% survival rate compared with controls who all died within 11 days of challenge. Results of the present study indicated that this novel immunization strategy is useful in enhancing immune protection in mice against lethal T. gondii infection, which would provide foundation for the development of effective vaccines against T. gondii.


Assuntos
Antígenos de Protozoários/imunologia , Herpesvirus Suídeo 1/imunologia , Imunização Secundária/métodos , Proteínas de Protozoários/imunologia , Toxoplasma/imunologia , Toxoplasmose/prevenção & controle , Vacinas de DNA/genética , Vacinas de DNA/imunologia , Animais , Anticorpos/imunologia , Antígenos de Protozoários/genética , Antígenos de Protozoários/metabolismo , Proliferação de Células , Células Cultivadas , Citocinas/imunologia , Feminino , Herpesvirus Suídeo 1/genética , Herpesvirus Suídeo 1/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/metabolismo , Baço/imunologia , Toxoplasma/genética , Toxoplasma/metabolismo , Toxoplasmose/imunologia , Toxoplasmose/parasitologia , Vacinas de DNA/metabolismo
12.
Acta Trop ; 111(3): 284-8, 2009 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-19467215

RESUMO

Two recombinant plasmids, pVAX/SjFABP and pVAX/mIL-18 containing Schistosoma japonicum 14 kDa fatty acid binding protein (SjFABP) and murine IL-18, were constructed and evaluated for their ability to induce immune responses and to protect against S. japonicum challenge in mice. Mice were intramuscularly immunized twice at three-weekly intervals, and challenged with S. japonicum cercariae at 4 weeks after the last vaccination. All animals vaccinated with pVAX/SjFABP alone or plus pVAX/mIL-18 developed specific anti-SWAP ELISA antibody and T lymphocyte proliferation. Co-injection of pVAX/mIL-18 significantly increased the production of IFN-gamma and IL-2 compared with pVAX/SjFABP alone, indicating that IL-18 enhances the Th1-dominant immune response. The challenge experiment showed that co-injection of plasmid encoding IL-18 significantly enhances protective effect against S. japonicum infection, as demonstrated by worm reduction rates and the hepatic egg reduction rates 45 days post-challenge. These results indicated that IL-18 may become a novel vaccine adjuvant for development of vaccines against schistosomiasis.


Assuntos
Adjuvantes Imunológicos/farmacologia , Proteínas de Ligação a Ácido Graxo/imunologia , Interleucina-18/farmacologia , Vacinas Protozoárias/imunologia , Schistosoma japonicum/imunologia , Esquistossomose Japônica/prevenção & controle , Vacinas de DNA/imunologia , Adjuvantes Imunológicos/administração & dosagem , Animais , Anticorpos Antiprotozoários/sangue , Proliferação de Células , Proteínas de Ligação a Ácido Graxo/genética , Feminino , Imunização Secundária , Injeções Intramusculares , Interferon gama/metabolismo , Interleucina-18/administração & dosagem , Interleucina-2/metabolismo , Fígado/parasitologia , Camundongos , Camundongos Endogâmicos BALB C , Contagem de Ovos de Parasitas , Plasmídeos , Vacinas Protozoárias/genética , Schistosoma japonicum/genética , Esquistossomose Japônica/imunologia , Linfócitos T/imunologia
13.
Trans R Soc Trop Med Hyg ; 103(2): 162-6, 2009 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-18822439

RESUMO

Infection with Toxoplasma gondii is common and usually asymptomatic, but it can have serious consequences in pregnant women if passed to the developing fetus. The aims of this study were to determine the prevalence of toxoplasmosis in pregnant women and to identify the possible risk factors associated with T. gondii infection in China. Of a sample of 235 pregnant women in Changchun, China, 25 (10.6%) were found by ELISA to be positive for IgG and none (0%) for IgM. Major risk factors were found by bivariate and multivariate analysis to include eating raw or undercooked meat, unwashed raw vegetables or fruit, contact with cats, living in rural areas, and low educational standards. In order to lower congenital infection, pregnant women need to be informed about the risk factors for toxoplasmosis.


Assuntos
Complicações Parasitárias na Gravidez/epidemiologia , Toxoplasmose/epidemiologia , Adulto , Animais , Anticorpos Antiprotozoários/sangue , Gatos , China/epidemiologia , Ensaio de Imunoadsorção Enzimática , Fezes/parasitologia , Feminino , Manipulação de Alimentos , Conhecimentos, Atitudes e Prática em Saúde , Humanos , Carne/parasitologia , Gravidez , Complicações Parasitárias na Gravidez/parasitologia , Prevalência , Fatores de Risco , Toxoplasma/imunologia , Toxoplasmose/transmissão , Toxoplasmose Congênita/complicações , Toxoplasmose Congênita/epidemiologia
14.
Microbes Infect ; 10(12-13): 1355-62, 2008 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-18761418

RESUMO

The major immunodominant surface antigen 1 (TgSAG1) of invasive tachyzoites is a vaccine candidate antigen for Toxoplasma gondii. In this study, we developed a recombinant pseudorabies virus (PRV) expressing TgSAG1 (rPRV/SAG1) based on the PRV vaccine strain Bartha K-61 by homologous recombination, in which partial PK and gG genes were deleted. The growth assay of rPRV/SAG1 showed that the recombinant virus can replicate in vitro as efficiently as PRV Bartha K-61, demonstrating that insertion of the TgSAG1 gene in the PK and gG locus of PRV does not affect the replication of PRV. All mice vaccinated with rPRV/SAG1 developed a high level of specific antibody responses against T. gondii lysate antigen (TLA), a strong increase of the splenocyte proliferative response, and significant levels of IFN-gamma and IL-2 production. And the immunization of mice with rPRV/SAG1 elicited strong cytotoxic T lymphocyte (CTL) responses in vitro. These results demonstrate that rPRV/SAG1 could induce significant humoral and cellular Th1 immune responses. Moreover, rPVR/SAG1 immunization induced partial protection (60%) against a lethal challenge with the highly virulent T. gondii RH strain, and neutralizing antibodies against PRV in a BALB/c mouse model. These results suggest that expression of protective antigens of T. gondii in PRV Bartha K-61 is a novel approach towards the development of a vaccine against both animal toxoplasmosis and pseudorabies.


Assuntos
Antígenos de Protozoários , Herpesvirus Suídeo 1 , Proteínas de Protozoários , Pseudorraiva/prevenção & controle , Recombinação Genética , Toxoplasmose Animal/prevenção & controle , Animais , Anticorpos Antiprotozoários/sangue , Anticorpos Antivirais/sangue , Antígenos de Protozoários/genética , Antígenos de Protozoários/imunologia , Antígenos de Protozoários/metabolismo , Herpesvirus Suídeo 1/genética , Herpesvirus Suídeo 1/imunologia , Herpesvirus Suídeo 1/metabolismo , Herpesvirus Suídeo 1/patogenicidade , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos BALB C , Proteínas de Protozoários/genética , Proteínas de Protozoários/imunologia , Proteínas de Protozoários/metabolismo , Vacinas Protozoárias/administração & dosagem , Vacinas Protozoárias/genética , Vacinas Protozoárias/imunologia , Pseudorraiva/imunologia , Pseudorraiva/virologia , Vacinas contra Pseudorraiva/administração & dosagem , Vacinas contra Pseudorraiva/genética , Vacinas contra Pseudorraiva/imunologia , Toxoplasma/genética , Toxoplasma/imunologia , Toxoplasma/metabolismo , Toxoplasma/patogenicidade , Toxoplasmose Animal/imunologia , Toxoplasmose Animal/parasitologia , Vacinação , Vacinas de DNA/administração & dosagem , Vacinas de DNA/genética , Vacinas de DNA/imunologia
15.
Vaccine ; 26(33): 4145-9, 2008 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-18562051

RESUMO

Two recombinant plasmids pVAX/Sj26GST and pVAX/mIL-18 containing Schistosoma japonicum 26kDa GST and murine IL-18 were evaluated for their ability to protect mice against S. japonicum challenge. Mice were given 2 intramuscular immunizations 3 weeks apart, and challenged with S. japonicum cercariae 4 weeks later. Adult worm and egg burdens were determined 48 days post-challenge. All animals vaccinated with pVAX/Sj26GST alone or with pVAX/mIL-18 developed specific anti-SWAP (soluble worm antigen preparation) ELISA antibody and splenocyte proliferation response. Co-injection of pVAX/mIL-18 significantly increased the production of IFN-gamma and IL-12, indicating that IL-18 enhances the Th1-dominant immune response. Challenge experiments showed that worms were reduced in the pVAX/Sj26GST group by 30.1% and by 49.4% in animals given pVAX/mIL-18 additionally. Corresponding hepatic and fecal egg reductions were 44.8% and 53.0%, and 50.6% and 56.6%, respectively. These results indicate that IL-18 may be an effective adjuvant for a schistosomiasis vaccine.


Assuntos
Antígenos de Helmintos/imunologia , Glutationa Transferase/imunologia , Interleucina-18/imunologia , Schistosoma japonicum/imunologia , Vacinas de DNA/imunologia , Adjuvantes Imunológicos/farmacologia , Animais , Anticorpos Anti-Helmínticos/sangue , Antígenos de Helmintos/genética , Proliferação de Células , Fezes/parasitologia , Feminino , Glutationa Transferase/genética , Imunização Secundária , Injeções Intramusculares , Interferon gama/metabolismo , Interleucina-12/metabolismo , Interleucina-18/farmacologia , Fígado/parasitologia , Linfócitos/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Contagem de Ovos de Parasitas , Plasmídeos , Esquistossomose Japônica/prevenção & controle , Baço/imunologia , Vacinas de DNA/genética
16.
Biologicals ; 36(3): 162-7, 2008 May.
Artigo em Inglês | MEDLINE | ID: mdl-18249007

RESUMO

Seoul virus glycoprotein Gn is a major structural protein and candidate antigen of hantavirus that induces a highly immunogenic response for hantavirus vaccine. In this study, a replication-competent recombinant canine adenovirus type-2 expressing Gn was constructed by the in vitro ligation method. The Gn expression cassette, including the human cytomegalovirus (hCMV) promoter/enhancer and the SV40 early mRNA polyadenylation signal, was cloned into the SspI site of the E3 region which is not essential for proliferation of CAV-2. Expression of Gn was confirmed by reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting.


Assuntos
Adenovirus Caninos/metabolismo , Glicoproteínas/metabolismo , Vírus Seoul/metabolismo , Animais , Bioquímica/métodos , Linhagem Celular , Citomegalovirus/genética , Cães , Técnicas Genéticas , Vetores Genéticos , Hantavirus/metabolismo , Camundongos , Modelos Genéticos , Regiões Promotoras Genéticas , Reação em Cadeia da Polimerase Via Transcriptase Reversa
17.
Artigo em Inglês | MEDLINE | ID: mdl-17922312

RESUMO

The main objective of this study is to develop and evaluate the immobilization of Staphylococcal Protein A on magnetic cellulose microspheres (SPA-MCMS) for immunological capture of IgG. After cloning, expression and separation, SPA was immobilized onto MCMS to prepare a magnetic affinity media subject to the purification of IgGs. The binding capacity, binding time, leakage of SPA and its reproducibility were optimized to improve the binding efficiency with an appropriate amount and recovery of IgG. Rabbit IgG was successfully purified from serum in a single-step by SPA-MCMS with an overall recovery of 73.18% and purity of 90.27%. Therefore, this study effectively illustrated the advantages of magnetic microcarriers for rapid and efficient purification of antibodies. The separation media shows a high potential for the future development of affinity isolation and immunodiagnostic application.


Assuntos
Imunoglobulina G/isolamento & purificação , Magnetismo , Microesferas , Microbiologia Industrial , Proteína Estafilocócica A/imunologia
18.
Parasitol Int ; 56(4): 263-8, 2007 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-17587637

RESUMO

Toxoplasma gondii is an obligate intracellular parasite, capable of infecting a variety of mammals and birds. Development of vaccine against T. gondii would be of great medical and veterinary value. In this study, the DNA sequence encoding ROP2 from T. gondii was cloned into the muticopy mycobacterial expression vector, pMV262, under the control of the Bacillus Calmette-Guerin (BCG) hsp60 promoter, and electroporated into BCG. Following selection of kanamycin, the recombinant BCG/pMV262-ROP2 was constructed and the expression of ROP2 was confirmed by Western blotting. The BALB/c mice inoculated with the BCG/pMV262-ROP2 developed specific immune responses against ROP2 protein, and there was an obvious delay in the mortality curve than the control (P<0.05). These results indicated that M. bovis BCG is an adequate vector to express and present antigens of T. gondii, and it may be used to further study the induction of protective immunity in other animals.


Assuntos
Anticorpos Antiprotozoários/sangue , Linfócitos T CD4-Positivos/imunologia , Proteínas de Membrana/imunologia , Mycobacterium bovis/genética , Proteínas de Protozoários/imunologia , Vacinas Protozoárias , Toxoplasma/imunologia , Animais , Clonagem Molecular , Vetores Genéticos , Interferon gama/biossíntese , Interleucina-1/biossíntese , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Mycobacterium bovis/imunologia , Mycobacterium bovis/metabolismo , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Vacinas Protozoárias/administração & dosagem , Vacinas Protozoárias/genética , Vacinas Protozoárias/imunologia , Toxoplasma/genética , Toxoplasma/metabolismo , Toxoplasmose Animal/imunologia , Toxoplasmose Animal/mortalidade , Toxoplasmose Animal/parasitologia , Vacinação , Vacinas de DNA/administração & dosagem , Vacinas de DNA/genética , Vacinas de DNA/imunologia
19.
Cancer Lett ; 254(1): 71-4, 2007 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-17376590

RESUMO

To determine Toxoplasma gondii antibodies in cancer patients, 267 cancer patients were studied using ELISA and higher positivity rates of T. gondii IgG were detected than the control. The positivity rates of T. gondii IgG in nasopharyngeal carcinoma, rectal cancer groups were significantly higher than the other cancer groups, but the differences in IgM positivity rates were not significant, demonstrating that there is a likely association between T. gondii infection and some kinds of cancer, especially nasopharyngeal carcinoma and rectal cancer.


Assuntos
Anticorpos Antiprotozoários/sangue , Neoplasias/sangue , Toxoplasma/imunologia , Toxoplasmose/sangue , Adulto , Idoso , Animais , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Masculino , Pessoa de Meia-Idade , Neoplasias Nasofaríngeas/sangue , Neoplasias Nasofaríngeas/complicações , Neoplasias/complicações , Neoplasias Retais/sangue , Neoplasias Retais/complicações , Toxoplasma/crescimento & desenvolvimento , Toxoplasmose/complicações , Toxoplasmose/parasitologia
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