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1.
Arch Virol ; 166(11): 3189-3192, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34524537

RESUMO

Porcine circovirus type 2 (PCV2) is the etiological agent of post-weaning multisystemic wasting syndrome (PMWS). The original prevalent genotype, PCV2a, has been replaced by genotypes 2b and 2d in the swine population worldwide. The Rep protein is critical for viral replication. Comparison of a large number of Rep protein amino acid (aa) sequences showed that three sites distinguish genotype 2b from genotype 2d. In order to analyze the effect of exchanging the amino acids (asparagine and serine) at position 6 in the Rep proteins of PCV2b and PCV2d, two wild-type and two mutant viruses were rescued. Real-time quantitative PCR and a one-step growth curve were used to determine the viral load to assess the replication ability of the rescued viruses. The results showed that there was no significant difference in in vitro performance between the wild-type PCV2b and the mutated virus, while the mutation of PCV2d enhanced viral replication.


Assuntos
Infecções por Circoviridae/virologia , Circovirus/genética , Mutação , Proteínas Virais/genética , Replicação Viral/genética , Animais , Infecções por Circoviridae/veterinária , Circovirus/fisiologia , Suínos , Doenças dos Suínos/virologia , Carga Viral , Proteínas Virais/química , Proteínas Virais/metabolismo
2.
Front Microbiol ; 12: 674907, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34211446

RESUMO

The endoplasmic reticulum (ER) plays an essential role in Ca2+ concentration balance and protein biosynthesis. During infection, the virus needs to complete its life process with the help of ER. At the same time, ER also produces ER stress (ERS), which induces apoptosis to resist virus infection. Our study explored the Ca2+ concentration, ERS, and the apoptosis mechanism after porcine circovirus 2 (PCV2) infection. We show here that PCV2 infection induces the increased cytoplasmic Ca2+ level and PK-15 cell ER swelling. The colocalization of phospholipase C (PLC) and inositol 1,4,5-trisphosphate receptor (IP3R) in the cytoplasm was observed by laser confocal microscopy. Western blot and quantitative polymerase chain reaction experiments confirmed that PLC and IP3R expression levels increased after PCV2 infection, and Ca2+ concentration in the cytoplasm increased after virus infection. These results suggest that PCV2 infection triggers ERS of PK-15 cells via the PLC-IP3R-Ca2+ signaling pathway to promote the release of intracellular Ca2+ and led to cell apoptosis.

3.
Mol Immunol ; 133: 63-66, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33631556

RESUMO

Porcine circovirus type 2 (PCV2), a ubiquitous pathogen that primary cause of postweaning multisystemic wasting syndrome (PMWS), had caused significant morbidity and mortality in swine populations with huge economic losses in the worldwide swine industry. Currently, looking for effective antiviral drugs for PCV2 infection remains an important works. In our study, CRISPR/Cas9 system was used to further detected the key sites of PCV2 replication. We designed 8 single guide RNAs (sgRNA) by targeting essential genes across the genome of PCV2. Western-blot(WB), Cell counting kit-8 for high-throughput sgRNA screening were applied to detect PCV2 replication levels. The results showed that Oc8, O13, O134, NQT and NPS sgRNAs can edit the PCV2 genome efficiently and inhibit PCV2 replication in PK-15 cell; H3 sgRNA cannot edit the PCV2 genome successfully; NAT sgRNA can edit the PCV2 genome efficiently to improve the PCV2 replication in PK-15 cell; O26 sgRNA can edit the PCV2 genome successfully but it is not known yet of its effect on PCV2 replication, besides the Cas9 expression had no effect on cell viability. These data suggest that CRISPR/Cas9 system targeting PCV2 essential genes may serve as a novel therapeutic agent against PCV2 infection in the future.


Assuntos
Sistemas CRISPR-Cas/genética , Infecções por Circoviridae/terapia , Circovirus/genética , Síndrome Definhante Multissistêmico de Suínos Desmamados/terapia , RNA Guia/genética , Animais , Linhagem Celular , Infecções por Circoviridae/veterinária , Infecções por Circoviridae/virologia , Genes Essenciais/genética , Genoma Viral/genética , Glicosilação , Síndrome Definhante Multissistêmico de Suínos Desmamados/virologia , Suínos , Doenças dos Suínos/terapia , Doenças dos Suínos/virologia , Replicação Viral/genética
4.
Front Physiol ; 11: 669, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32695015

RESUMO

Cardiac fibrosis is a common pathological process in multiple cardiovascular diseases, including myocardial infarction (MI). Abnormal cardiac fibroblast (CF) activity is a key event in cardiac fibrosis. Although the Notch signaling pathway has been reported to play a vital role in protection from cardiac fibrosis, the exact mechanisms underlying cardiac fibrosis and protection from it have not yet been elucidated. Similarly, Hif1α and the RhoA/ROCK signaling pathway have been shown to participate in cardiac fibrosis. The RhoA/ROCK signaling pathway has been reported to be an upstream pathway of Hif1α in several pathophysiological processes. In the present study, we aimed to determine the effects of notch3 on CF activity and its relationship with the RhoA/ROCK/Hif1α signaling pathway. Using in vitro experiments, we demonstrated that notch3 inhibited CF proliferation and fibroblast to myofibroblast transition (FMT) and promoted CF apoptosis. A knockdown of notch3 using siRNAs had the exact opposite effect. Next, we found that notch3 regulated CF activity by negative regulation of the RhoA/ROCK/Hif1α signaling pathway. Extending CF-based studies to an in vivo rat MI model, we showed that overexpression of notch3 by the Ad-N3ICD injection attenuated the increase of RhoA, ROCK1, ROCK2, and Hif1α levels following MI and further prevented MI-induced cardiac fibrosis. On the basis of these results, we conclude that notch3 is involved in the regulation of several aspects of CF activity, including proliferation, FMT, and apoptosis, by inhibiting the RhoA/ROCK/Hif1α signaling pathway. These findings are significant to further our understanding of the pathogenesis of cardiac fibrosis and to ultimately identify new therapeutic targets for cardiac fibrosis, potentially based on the RhoA/ROCK/Hif1α signaling pathway.

5.
Biosci Rep ; 40(5)2020 05 29.
Artigo em Inglês | MEDLINE | ID: mdl-32432671

RESUMO

BACKGROUND: Influenza A virus (IAV) has greatly affected public health in recent decades. Accumulating data indicated that host microRNAs (miRNAs) were related to IAV replication. The present study mainly focused on the effects of microRNA-21-3p (miR-21-3p) on H5N1 replication. METHODS: The levels of miR-21-3p, virus structural factors (matrix 1 (M1), nucleoprotein (NP)), type I interferon (IFN) response markers (IFN-ß, IFN-α), IFN-stimulated genes (protein kinase R (PKR), myxovirus resistance A (MxA), 2'-5'-oligoadenylate synthetase 2 (OAS)), and fibroblast growth factor 2 (FGF2) were measured by quantitative real-time polymerase chain reaction (qRT-PCR). The protein levels of M1, NP, and FGF2 were tested by Western blot assay. The virus titer was assessed by tissue culture infective dose 50% (TCID50) assay. The dual-luciferase reporter assay and ribonucleic acid (RNA) immunoprecipitation (RIP) assay were used to verify the interaction between miR-21-3p and FGF2. RESULTS: MiR-21-3p was reduced in H5N1-infected patients and A549 cells. MiR-21-3p overexpression facilitated the levels of M1, NP, TCID50 value, and reduced the levels of IFN-ß, IFN-α, PKR, MxA, and OAS in H5N1-infected A549 cells. FGF2 was verified as a direct target of miR-21-3p. The introduction of FGF2 counteracted miR-21-3p-mediated decrease in the levels of M1, NP, and TCID50 value, as well as reduction in the levels of IFN-ß, IFN-α, PKR, MxA, and OAS in H5N1-infected A549 cells. CONCLUSION: MiR-21-3p down-regulated FGF2 expression to accelerate H5N1 replication and confine IFN response.


Assuntos
Fator 2 de Crescimento de Fibroblastos/metabolismo , Virus da Influenza A Subtipo H5N1/crescimento & desenvolvimento , Influenza Humana/virologia , Interferon Tipo I/metabolismo , MicroRNAs/metabolismo , Replicação Viral , Células A549 , Adulto , Estudos de Casos e Controles , Feminino , Fator 2 de Crescimento de Fibroblastos/genética , Regulação da Expressão Gênica , Interações Hospedeiro-Patógeno , Humanos , Virus da Influenza A Subtipo H5N1/patogenicidade , Influenza Humana/genética , Influenza Humana/metabolismo , Interferon Tipo I/genética , Masculino , MicroRNAs/sangue , MicroRNAs/genética
6.
Vet Med Sci ; 6(1): 76-81, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31621210

RESUMO

PCV2 belongs to the genus Circovirus, family Circoviridae, who is recognized as the causative agents of postweaning multisystemic wasting syndrome. Since being found to China in 2000, it has caused serious damage to the pig industry. In this study, we downloaded 40 PCV2 genome-wide sequences uploaded to GenBank from 2013 to 2018 in Shandong Province, including 23 uploaded by our laboratory. Construction of a genome-wide evolution tree using MEGA V5.0 software. Phylogenetic tree analysis indicated that the genotype of PCV2 in Shandong Province was: three genotypes coexisted (2a, 2b, 2d); among them, PCV2d has become the main genotype in the province due to its number and spread range. Amino acid sequence analysis of different genotypes of ORF2 showed that specific amino acid sites exist in different genotypes, with the most significant range of 81-160; different genotypes of PCV2 can be distinguished at the molecular level. This study found that due to the increase in infections of the PCV2d genotype in recent years, it may replace PCV2b as the dominant base in Shandong.


Assuntos
Infecções por Circoviridae/veterinária , Circovirus/genética , Variação Genética , Genoma Viral , Genótipo , Doenças dos Suínos/virologia , Animais , China , Infecções por Circoviridae/virologia , Análise de Sequência de Proteína/veterinária , Sus scrofa , Suínos
7.
J Virol Methods ; 282: 113774, 2019 Nov 11.
Artigo em Inglês | MEDLINE | ID: mdl-31726113

RESUMO

Porcine Reproductive and Respiratory Syndrome (PRRS), an acute infectious disease caused by the porcine reproductive and respiratory syndrome virus (PRRSV), is one of the most devastating diseases affecting the global swine industry. In order to establish a multiplex real-time PCR method for the simultaneous detection of the classical PRRSV (C-PRRSV) strain, the highly pathogenic PRRSV (HP-PRRSV) strain and NADC30-like PRRSV (NL-PRRSV) strain, we designed specific primers and TaqMan fluorescent probes based on the Nsp2 target gene sequence of these three different PRRSV strains, and designed American-type PRRSV (PRRSV-U) special primers and probes based on the relatively conserved target gene sequence of ORF7. The method established in this study can quickly and accurately detect and differentiate three types of strains of clinical tissue samples, respectively. This method plays a key role in the rapid diagnosis and determination of PRRSV.

8.
Biosci Rep ; 39(6)2019 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-31113873

RESUMO

Background: Some pilot studies already tried to investigate potential associations of leptin (LEP) and LEP receptor (LEPR) variants with coronary artery disease (CAD). However, the results of these studies were not consistent. Thus, we performed the present meta-analysis to explore associations between LEP/LEPR variants and CAD in a larger pooled population.Methods: Systematic literature research of PubMed, Web of Science, Embase and CNKI was performed to identify eligible case-control studies on associations between LEP/LEPR variants and CAD. The initial search was conducted in September 2018 and the latest update was performed in December 2018. Q test and I2 statistic were employed to assess between-study heterogeneities. If probability value(P-value) of Q test was less than 0.1 or I2 was greater than 50%, random-effect models (REMs) would be used to pool the data. Otherwise, fixed-effect models (FEMs) would be applied for synthetic analyses.Results: A total of ten studies published between 2006 and 2018 were eligible for analyses (1989 cases and 2601 controls). Pooled analyses suggested that LEP rs7799039 variant was significantly associated with CAD under over-dominant model (P=0.0007, odds ratio (OR) = 1.36, 95% confidence interval (CI): 1.14-1.63, I2 = 41%, FEM) in overall population, and this significant finding was further confirmed in East Asians in subsequent subgroup analyses. However, no positive findings were observed for LEPR rs1137100 and rs1137101 variants in overall and subgroup analyses.Conclusions: Our meta-analysis suggested that LEP rs7799039 variant might affect individual susceptibility to CAD.


Assuntos
Doença da Artéria Coronariana/genética , Predisposição Genética para Doença , Leptina/genética , Polimorfismo Genético , Receptores para Leptina/genética , Humanos
9.
Medicine (Baltimore) ; 97(38): e12428, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30235722

RESUMO

BACKGROUND: We sought to identify common ion channel single nucleotide polymorphisms (SNPs) associated with the occurrence of sudden cardiac death (SCD) to predict the incidence of SCD in clinical settings. METHODS: This study involved a systematic review and meta-analysis of ion channel SNPs and risk of SCD in adults. We searched public databases for studies published up to September 19, 2017. We examined relationships between SNPs in common ion channel genes and the incidence of SCD. RESULTS: We collected data for 22 trials that included a total of 4149 patients who experienced SCD or had a high risk of SCD and assessed these data in our meta-analysis. An allelic model showed that rs11720524 in SCN5A clearly protected against SCD (odds ratio [OR]: 0.76; 95% confidence interval [95% CI]: 0.67-0.85; P < .001). Subgroup analysis showed that rs11720524 in SCN5A protected against SCD in Europeans and Caucasians but not in Koreans. The allelic model indicated that rs12296050 in KCNQ1 also had significant protective effects against SCD (OR: 0.85; 95% CI: 0.76-0.96; P = .007). Moreover, this model demonstrated that rs2283222 in KCNQ1 had a significant negative relationship with SCD (OR: 0.73; 95% CI: 0.62-0.85; P < .001). Rs12296050 in KCNQ1 protected against SCD in Koreans and Americans. Our results also showed that rs790896 in RYR2 was negatively associated with SCD in a dominant model (OR: 0.66; 95% CI: 0.45-0.97; P = .033). CONCLUSIONS: Rs11720524 in SCN5A is negatively related to SCD in Europeans and Caucasians, and rs12296050 and rs2283222 in KCNQ1 and rs790896 in RYR2 clearly have protective effects against SCD.


Assuntos
Morte Súbita Cardíaca/epidemiologia , Canais Iônicos/genética , Polimorfismo de Nucleotídeo Único/genética , Adulto , Alelos , Grupo com Ancestrais do Continente Asiático/genética , Morte Súbita Cardíaca/etnologia , Morte Súbita Cardíaca/etiologia , Grupo com Ancestrais do Continente Europeu/genética , Feminino , Humanos , Incidência , Canal de Potássio KCNQ1/genética , Masculino , Canal de Sódio Disparado por Voltagem NAV1.5/genética , Estudos Observacionais como Assunto , Valor Preditivo dos Testes , Canal de Liberação de Cálcio do Receptor de Rianodina/genética
10.
Sci Rep ; 8(1): 8215, 2018 May 23.
Artigo em Inglês | MEDLINE | ID: mdl-29795230

RESUMO

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.

11.
Microb Pathog ; 117: 327-334, 2018 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-29496526

RESUMO

Porcine circovirus (PCV) has two potential open reading frames, ORF1 and ORF2. ORF1 is predicted to encode a replication-associated protein (Rep) essential for replication of viral DNA. In some studies, PCV1 replicated more efficiently in PK-15 cells than PCV2 was elucidated. PCV1 compared with PCV2; there is some amino acids' deficiency on Rep protein. To identify whether the above amino acids deletion affects the replication of PCV1 and PCV2, we constructed three double copy clones by overlap extension PCR. The 2PCV2(vV) clone deleted the valine of Rep protein in the backbone of PCV2 genome. The 2PCV2(dSGR) clone inserted serine, glycine and arginine of Rep protein successively in the backbone of PCV2 genome. The 2PCV2(dSGR&vV) clone inserted serine, glycine and arginine as well as deleted the valine of Rep protein in the backbone of PCV2 genome. These clones we constructed with amino acid mutations and parental DNA clones were all transfected in PK-15 cells that free of PCV contamination, and their growth characteristics in vitro were determined and compared, to evaluating the replication of the mutant infectious DNA clones. Our results showed that the double copy infectious clones with amino acid mutations could be rescued in vitro. The 2PCV2(vV) replicated more efficiently than parental viruses 2PCV2 and 2PCV1 but the replicated ability of 2PCV2(dSGR) and 2PCV2(dSGR&vV) is attenuated than parental viruses 2PCV2 and 2PCV1. We can determine the valine is the important amino acid that cause PCV1 replicated more efficiently in PK-15 cells than PCV2 primarily. These findings are benefit for exploring the mechanisms of viral replication in pigs and important implications for PCV2 vaccine development.


Assuntos
Circovirus/genética , Deleção de Sequência , Proteínas Virais/genética , Replicação Viral , Sequência de Aminoácidos , Aminoácidos , Animais , Linhagem Celular , Infecções por Circoviridae/virologia , DNA Viral/genética , Proteínas de Ligação a DNA/genética , Fases de Leitura Aberta/genética , Alinhamento de Sequência , Análise de Sequência de Proteína , Suínos , Vacinas
12.
Sci Rep ; 8(1): 4118, 2018 03 07.
Artigo em Inglês | MEDLINE | ID: mdl-29515183

RESUMO

Epidemiological investigations were conducted on recently emerging porcine reproductive and respiratory syndrome virus (PRRSV) strains in Shandong province in 2014-2015. The proportion of the NADC30 strain identified by ORF7 sequence alignment has been gradually increasing. Three emerging PRRSV strains were successfully isolated, and the complete genomic sequences were determined. Our results indicate the importance of recombinant strains in Shandong province, China. There was a varied degree of recombination of two or three strains (classical, HP-PRRSV and/or NADC30). Moreover, the recombination strains affected the pathogenicity of newly emerged strains.


Assuntos
Genoma Viral , Filogenia , Síndrome Respiratória e Reprodutiva Suína/genética , Vírus da Síndrome Respiratória e Reprodutiva Suína/genética , Recombinação Genética , Suínos/virologia , Animais , Vírus da Síndrome Respiratória e Reprodutiva Suína/isolamento & purificação , Alinhamento de Sequência
13.
Hematology ; 23(8): 478-485, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-29421985

RESUMO

OBJECTIVES: Acute lymphoblastic leukemia (ALL) is the most common cancer before the age of 15 years, seriously endangering the health of children. The main treatment for Childhood ALL was pharmacotherapy. But these drugs have many side effects and some of them could develop drug resistance quickly. Mometasone furoate (MF) is an efficient glucocorticoid for topical treatment of inflammation on the skin, lung and nose. METHODS: In this study, we investigated whether the MF had effects on ALL cells proliferation and migration. RESULTS: The CCK-8 proliferation test showed that the cell viability was the lowest at 25 nM MF treatment and the increased OD value was time-dependent. In transwell assay, the number of CCRF-CEM cells was reduced in MF treated group. We found the expression of anti-apoptotic protein bcl-2 decreased the expression of pro-apoptotic protein caspase3 and bax increased in CCRF-CEM cell line treated with MF. The expression of p-AKT, p-mTOR, p70S6 K, vascular endothelial growth factor and CyclinD1 were decreased in MF treated group. CONCLUSION: This study reveals that MF can inhibit proliferation and invasion/migration and induce apoptosis in Childhood ALL cells, which may be regulated by Phosphatidylinositol 3-kinase signaling pathway. These results suggest MF may be a potential new drug target for clinical ALL treatment.


Assuntos
Leucemia , Furoato de Mometasona/farmacologia , Proteínas de Neoplasias/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Transdução de Sinais/efeitos dos fármacos , Doença Aguda , Adolescente , Linhagem Celular Tumoral , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Recém-Nascido , Leucemia/tratamento farmacológico , Leucemia/metabolismo , Leucemia/patologia , Masculino
14.
Cell Rep ; 19(13): 2809-2822, 2017 06 27.
Artigo em Inglês | MEDLINE | ID: mdl-28658627

RESUMO

Accessory proteins in Frizzled (FZD) receptor complexes are thought to determine ligand selectivity and signaling amplitude. Genetic evidence indicates that specific combinations of accessory proteins and ligands mediate vascular ß-catenin signaling in different CNS structures. In the retina, the tetraspanin TSPAN12 and the ligand norrin (NDP) mediate angiogenesis, and both genes are linked to familial exudative vitreoretinopathy (FEVR), yet the molecular function of TSPAN12 remains poorly understood. Here, we report that TSPAN12 is an essential component of the NDP receptor complex and interacts with FZD4 and NDP via its extracellular loops, consistent with an action as co-receptor that enhances FZD4 ligand selectivity for NDP. FEVR-linked mutations in TSPAN12 prevent the incorporation of TSPAN12 into the NDP receptor complex. In vitro and in Xenopus embryos, TSPAN12 alleviates defects of FZD4 M105V, a mutation that destabilizes the NDP/FZD4 interaction. This study sheds light on the poorly understood function of accessory proteins in FZD signaling.


Assuntos
Proteínas do Olho/genética , Proteínas do Olho/metabolismo , Receptores Frizzled/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Tetraspaninas/metabolismo , Receptores Frizzled/genética , Humanos , Mutação de Sentido Incorreto , Transdução de Sinais
15.
Microb Pathog ; 109: 319-324, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28457899

RESUMO

Porcine reproductive and respiratory syndrome (PRRS), a highly contagious disease, has been constantly causing huge economic losses all over the world. PRRS virus (PRRSV) infection results in immunosuppression and IL-10 up-regulation. The relationship between them is still in dispute. Previous studies demonstrated the protein of PRRSV nucleocapsid (N) protein is able to up-regulate IL-10, yet the underlying molecular mechanisms remain unknown. In this study, the expression kinetics of IL-10 up-regulation induced by PRRSV N protein were analyzed in immortalized porcine alveolar macrophages (PAMs). N protein induced IL-10 expression in a time- and dose-dependent manner. Inhibition experiments of signaling pathways suggested NF-κB and p38 MAPK pathways are both involved in N protein-induced IL-10 up-regulation. Besides, the integrity of N protein is essential for significant IL-10 up-regulation. This research is beneficial for further understanding of the interplay between PRRSV and host immune system.


Assuntos
Interleucina-10/metabolismo , Macrófagos Alveolares/imunologia , NF-kappa B/metabolismo , Proteínas do Nucleocapsídeo/metabolismo , Vírus da Síndrome Respiratória e Reprodutiva Suína/metabolismo , Suínos/imunologia , Regulação para Cima , Animais , Linhagem Celular , Regulação da Expressão Gênica , Imunossupressão , Macrófagos Alveolares/virologia , Proteínas do Nucleocapsídeo/genética , Síndrome Respiratória e Reprodutiva Suína/imunologia , Síndrome Respiratória e Reprodutiva Suína/virologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/genética , Vírus da Síndrome Respiratória e Reprodutiva Suína/imunologia , Transdução de Sinais , Suínos/virologia , Proteínas Virais/genética , Proteínas Virais/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
16.
Res Vet Sci ; 109: 166-168, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27892867

RESUMO

The use of BALB/c mouse as an alternative model to study Haemophilus parasuis (HPS) infections was evaluated, supplying the serotyping scheme by comparing the pathogenicity of different serovar HPS in pigs and mice challenge using statistical analysis. Results showed that the pathogenicity of different serovar HPS in mouse was consistent with in pigs, proving that this model is a viable alternative to pigs. It provides a convenient methodology for determining the virulence of HPS strains.


Assuntos
Infecções por Haemophilus/veterinária , Haemophilus parasuis/fisiologia , Haemophilus parasuis/patogenicidade , Doenças dos Suínos/virologia , Animais , Feminino , Infecções por Haemophilus/virologia , Haemophilus parasuis/classificação , Camundongos , Camundongos Endogâmicos BALB C , Modelos Animais , Sorogrupo , Sorotipagem/veterinária , Suínos , Virulência
17.
Virol J ; 13(1): 185, 2016 Nov 14.
Artigo em Inglês | MEDLINE | ID: mdl-27842600

RESUMO

BACKGROUND: This study aimed at reseaching the immune effect of porcine circovirus type 2 (PCV2) DNA vaccine containing CpG motif on mice. METHODS: A total of 40 6-week-old female BALB/c mice were randomly divided into four groups which were immunized by 18CpG-pVAX1-ORF2, pVAX1-ORF2, pVAX1 and PBS, respectively, and immunized again 2 weeks later. All mice were challenged with 0.2 mL PCV2 cells virulent strain SD (106.0 TCID50/mL) after 4 weeks. Average daily gain, blood antibody levels, microscopic changes and viremia were detected to estimate the effect of DNA vaccine. RESULTS AND DISCUSSION: The results showed that compared to those of the control mice, groups immunized with pVAX1-ORF2 and 18CpG-pVAX1-ORF2 could induce PCV2-specific antibodies. The PCV2-specific antibodies level of 18 CpG-pVAX1-ORF2 groups was higher significantly than other groups and decreased slowly along with time. There was no distinct pathological damage and viremia occurring in mice that inoculated with CpG motif DNA vaccines. The results demonstrated that the DNA vaccine containing 18 CpG could build up resistibility immunity and reduce immune organ damage on mice.


Assuntos
Adjuvantes Imunológicos , Infecções por Circoviridae/prevenção & controle , Circovirus/imunologia , Ilhas de CpG , Vacinas de DNA/imunologia , Vacinas Virais/imunologia , Animais , Anticorpos Antivirais/sangue , Peso Corporal , Infecções por Circoviridae/patologia , Modelos Animais de Doenças , Feminino , Esquemas de Imunização , Camundongos Endogâmicos BALB C , Resultado do Tratamento , Vacinas de DNA/administração & dosagem , Vacinas de DNA/genética , Vacinas Virais/administração & dosagem , Vacinas Virais/genética , Viremia/prevenção & controle
18.
Dev Biol ; 411(2): 257-265, 2016 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-26783883

RESUMO

EFHC1 encodes a ciliary protein that has been linked to Juvenile Myoclonic Epilepsy. In ectodermal explants, derived from Xenopus laevis embryos, the morpholino-mediated down-regulation of EFHC1b inhibited multiciliated cell formation. In those ciliated cells that did form, axoneme but not basal body formation was inhibited. EFHC1b morphant embryos displayed defects in central nervous system (CNS) and neural crest patterning that were rescued by a EFHC1b-GFP chimera. EFHC1b-GFP localized to ciliary axonemes in epidermal, gastrocoele roof plate, and neural tube cells. In X. laevis there is a link between Wnt signaling and multiciliated cell formation. While down-regulation of EFHC1b led to a ~2-fold increase in the activity of the ß-catenin/Wnt-responsive TOPFLASH reporter, EFHC1b-GFP did not inhibit ß-catenin activation of TOPFLASH. Wnt8a RNA levels were increased in EFHC1b morphant ectodermal explants and intact embryos, analyzed prior to the on-set of ciliogenesis. Rescue of the EFHC1b MO's ciliary axonemal phenotypes required the entire protein; in contrast, the EFHC1b morpholino's Wnt8a, CNS, and neural crest phenotypes were rescued by a truncated form of EFHC1b. The EFHC1b morpholino's Wnt8a phenotype was also rescued by the injection of RNAs encoding secreted Wnt inhibitors, suggesting that these phenotypes are due to effects on Wnt signaling, rather than the loss of cilia, an observation of potential relevance to understanding EFHC1's role in human neural development.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Cílios/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Proteínas Wnt/metabolismo , Via de Sinalização Wnt , Proteínas de Xenopus/metabolismo , Xenopus laevis/embriologia , Animais , Genes Reporter , Proteínas de Fluorescência Verde/metabolismo , Imuno-Histoquímica , Hibridização In Situ , Microscopia Confocal , Morfogênese , Mutação , Crista Neural/citologia , Crista Neural/embriologia , Tubo Neural/embriologia , Fenótipo , Estrutura Terciária de Proteína , RNA/metabolismo , Transdução de Sinais , Regulação para Cima , beta Catenina/metabolismo
19.
Viral Immunol ; 28(5): 290-6, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-26046831

RESUMO

Nowadays, adjuvant is still important for boosting immunity and improving resistance in animals. In order to boost the immunity of porcine circovirus type 2 (PCV2) DNA vaccine, CpG motifs were inserted. In this study, the dose-effect was studied, and the immunity of PCV2 DNA vaccines by recombinant open reading frame 2 (ORF2) gene and CpG motifs was evaluated. Three-week-old Changbai piglets were inoculated intramuscularly with 200 µg, 400 µg, and 800 µg DNA vaccines containing 14 and 18 CpG motifs, respectively. Average gain and rectum temperature were recorded everyday during the experiments. Blood was collected from the piglets after vaccination to detect the changes of specific antibodies, interleukin-2, and immune cells every week. Tissues were collected for histopathology and polymerase chain reaction. The results indicated that compared to those of the control piglets, all concentrations of two DNA vaccines could induce PCV2-specific antibodies. A cellular immunity test showed that PCV2-specific lymphocytes proliferated the number of TH, TC, and CD3+ positive T-cells raised in the blood of DNA vaccine immune groups. There was no distinct pathological damage and viremia occurring in pigs that were inoculated with DNA vaccines, but there was some minor pathological damage in the control group. The results demonstrated that CpG motifs as an adjuvant could boost the humoral and cellular immunity of pigs to PCV2, especially in terms of cellular immunity. Comparing two DNA vaccines that were constructed, the one containing 18 CpG motifs was more effective. This is the first report that CpG motifs as an adjuvant insert to the PCV2 DNA vaccine could boost immunity.


Assuntos
Infecções por Circoviridae/imunologia , Infecções por Circoviridae/prevenção & controle , Doenças dos Suínos/imunologia , Vacinas de DNA/imunologia , Vacinas Virais/imunologia , Adjuvantes Imunológicos/farmacologia , Animais , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Formação de Anticorpos/imunologia , Infecções por Circoviridae/veterinária , Circovirus/imunologia , Imunidade Celular/imunologia , Imunidade Humoral/imunologia , Interleucina-2/sangue , Oligodesoxirribonucleotídeos/imunologia , Fases de Leitura Aberta/genética , Distribuição Aleatória , Suínos , Doenças dos Suínos/prevenção & controle , Linfócitos T Citotóxicos/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Vacinação , Viremia/imunologia
20.
Sci Rep ; 5: 10283, 2015 May 27.
Artigo em Inglês | MEDLINE | ID: mdl-26014913

RESUMO

Centrins (Cetns) are highly conserved, widely expressed, and multifunctional Ca(2+)-binding eukaryotic signature proteins best known for their roles in ciliogenesis and as critical components of the global genome nucleotide excision repair system. Two distinct Cetn subtypes, Cetn2-like and Cetn3-like, have been recognized and implicated in a range of cellular processes. In the course of morpholino-based loss of function studies in Xenopus laevis, we have identified a previously unreported Cetn2-specific function, namely in fibroblast growth factor (FGF) mediated signaling, specifically through the regulation of FGF and FGF receptor RNA levels. Cetn2 was found associated with the RNA polymerase II binding sites of the Cetn2-regulated FGF8 and FGFR1a genes, but not at the promoter of a gene (BMP4) whose expression was altered indirectly in Cent2 morphant embryos. These observations point to a previously unexpected role of Cetn2 in the regulation of gene expression and embryonic development.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Fatores de Crescimento de Fibroblastos/metabolismo , Receptores de Fatores de Crescimento de Fibroblastos/metabolismo , Proteínas de Xenopus/metabolismo , Xenopus/genética , Animais , Proteínas de Ligação ao Cálcio/antagonistas & inibidores , Proteínas de Ligação ao Cálcio/genética , Imunoprecipitação da Cromatina , Embrião não Mamífero/metabolismo , Desenvolvimento Embrionário/genética , Fatores de Crescimento de Fibroblastos/genética , Regulação da Expressão Gênica no Desenvolvimento , Mesoderma/metabolismo , Mesoderma/patologia , Microscopia de Fluorescência , Morfolinos/metabolismo , Receptores de Fatores de Crescimento de Fibroblastos/genética , Xenopus/crescimento & desenvolvimento , Proteínas de Xenopus/antagonistas & inibidores , Proteínas de Xenopus/genética
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