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Graphene-like materials in two dimensions hold great promise for energy storage and transformation applications owing to their distinctive features, such as lightweight composition, porous geometry, etc. Among these materials, a recently discovered unit known as g-B5N3 has demonstrated high performance in energy storage and transformation. In our efforts to enhance its applicability in adsorbing energy gases, we propose a novel composite structure by decorating Li atoms on the surface of pristine g-B5N3. The electronic properties of this composite have been comprehensively investigated using a first-principles method. Our findings reveal that the added Li atoms can be securely anchored on the g-B5N3 with an adsorption energy of -3.01 eV. Furthermore, the Li atom transfers its partial 2s electrons to the g-B5N3, exhibiting considerable electropositivity. These metallic sites effectively polarize the adsorbed H2 molecules, enhancing the mutual electrostatic interactions. Each primitive cell of Li-doped g-B5N3 can adsorb up to 13 H2 molecules, resulting in a storage capacity up to 6.3 wt %. This capacity significantly surpasses the goal of 4.5 wt % set by the U.S. Department of Energy. Furthermore, the typical adsorption energy of -0.209 eV per molecule of H2 aligns with the energy range suitable for reversible hydrogen storage. This study underscores the potential of Li-doped g-B5N3 for energy gas adsorption, shedding light on further advancements in this field.
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Background: Diabetes and thyroid dysfunction are two closely related endocrine diseases. Increasing evidences show that gut microbiota plays an important role in both glucose metabolism and thyroid homeostasis. Meanwhile, copy number variation (CNV) of host salivary α-amylase gene (AMY1) has been shown to correlate with glucose homeostasis. Hence, we aim to characterize the gut microbiota and CNV of AMY1 in type 2 diabetes (T2D) patients with or without subclinical hypothyroidism (SCH). Methods: High-throughput sequencing was used to analyze the gut microbiota of euthyroid T2D patients, T2D patients with SCH and healthy controls. Highly sensitive droplet digital PCR was used to measure AMY1 CN. Results: Our results revealed that T2D patients have lower gut microbial diversity, no matter with or without SCH. The characteristic taxa of T2D patients were Coriobacteriales, Coriobacteriaceae, Peptostreptococcaceae, Pseudomonadaceae, Collinsella, Pseudomonas and Romboutsia. Meanwhile, Escherichia/Shigella, Lactobacillus_Oris, Parabacteroides Distasonis_ATCC_8503, Acetanaerobacterium, Lactonifactor, uncultured bacterium of Acetanaerobacterium were enriched in T2D patients with SCH. Moreover, serum levels of free triiodothyronine (FT3) and free thyroxine (FT4) in T2D patients were both negatively correlated with richness of gut microbiota. A number of specific taxa were also associated with clinical parameters at the phylum and genus level. In contrast, no correlation was found between AMY1 CN and T2D or T2D_SCH. Conclusion: This study identified characteristic bacterial taxa in gut microbiota of T2D patients with or without SCH, as well as the taxa associated with clinical indices in T2D patients. These results might be exploited in the prevention, diagnosis and treatment of endocrine disorders in the future.
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Diabetes Mellitus Tipo 2 , Microbioma Gastrointestinal , Hipotireoidismo , Humanos , Microbioma Gastrointestinal/genética , Diabetes Mellitus Tipo 2/complicações , Variações do Número de Cópias de DNARESUMO
The monkeypox epidemic has attracted global attention to poxviruses. The cytoplasmic replication of poxviruses requires extensive protein synthesis, challenging the capacity of the endoplasmic reticulum (ER). However, the role of the ER in the life cycle of poxviruses is unclear. In this study, we demonstrate that infection with the lumpy skin disease virus (LSDV), a member of the poxvirus family, causes ER stress in vivo and in vitro, further facilitating the activation of the unfolded protein response (UPR). Although UPR activation aids in the restoration of the cellular environment, its significance in the LSDV life cycle remains unclear. Furthermore, the significance of ER imbalance for viral replication is also unknown. We show that LSDV replication is hampered by an unbalanced ER environment. In addition, we verify that the LSDV replication depends on the activation of PERK-eIF2α and IRE1-XBP1 signaling cascades rather than ATF6, implying that global translation and reduced XBP1 cleavage are deleterious to LSDV replication. Taken together, these findings indicate that LSDV is involved in the repression of global translational signaling, ER chaperone transcription, and ATF6 cleavage from the Golgi into the nucleus, thereby maintaining cell homeostasis; moreover, PERK and IRE1 activation contribute to LSDV replication. Our findings suggest that targeting UPR elements may be applied in response to infection from LSDV or even other poxviruses, such as monkeypox.
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Vírus da Doença Nodular Cutânea , Mpox , Animais , Bovinos , Humanos , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Vírus da Doença Nodular Cutânea/metabolismo , Mpox/metabolismo , Transdução de Sinais , Resposta a Proteínas não Dobradas , Estresse do Retículo Endoplasmático/fisiologia , Retículo Endoplasmático/metabolismo , Fator 6 Ativador da Transcrição/metabolismoRESUMO
Antibody development is the integral process of generating and characterizing an antibody. It commences by inoculating the antigen of interest into laboratory animals, allowing the immune system develops large quantities of antibodies. This was aimed at developing antibodies against the virion of Goatpox and Sheeppox virus vaccines. The ability of Goatpox and Sheeppox vaccines was assessed. Regarding this study, the antibody titers against both Goatpox and Sheeppox viruses was increased in the same manner. The amount of IgG was determined to be 2.29 µg/µl and 2.18 µg/µl against virions of Goatpox virus and Sheeppox respectively. The purified IgG was analyzed by SDS-PAGE. Different bands of the purified antibodies were clearly visualized, and the molecular weight of IgG was estimated to be 67 kDa and 25 kDa. Additionally, antigen/antibody binding was confirmed by Western blot using GTPV A27 antigen. No significant differences in antibody titers were observed between the two groups (p < 0, 05).
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BACKGROUND: Orf virus (ORFV) is a member of the genus Parapoxvirus and family Poxviridae. The virus has a worldwide distribution and infects sheep, goats, humans, and wild animals. However, due to the complex structure of the poxvirus, the underlying mechanism of the entry and infection by ORFV remains largely unknown. ORFV ORF047 encodes a protein named L1R. Poxviral L1R serves as the receptor-binding protein and blocks virus binding and entry independently of glycosaminoglycans (GAGs). The study aimed to identify the host interaction partners of ORFV ORF047. METHODS: Yeast two-hybrid cDNA library of sheep testicular cells was applied to screen the host targets with ORF047 as the bait. ORF047 was cloned into a pBT3-N vector and expressed in the NMY51 yeast strain. Then, the expression of bait proteins was validated by Western blot analysis. RESULTS: Sheep SERP1and PABPC4 were identified as host target proteins of ORFV ORF047, and a Co-IP assay further verified their interaction. CONCLUSIONS: New host cell proteins SERP1and PABPC4 were found to interact with ORFV ORF047 and might involve viral mRNA translation and replication.
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Interações entre Hospedeiro e Microrganismos , Vírus do Orf/metabolismo , Proteínas do Envelope Viral/metabolismo , Animais , Células Cultivadas , Masculino , Proteínas de Membrana/metabolismo , Vírus do Orf/química , Vírus do Orf/genética , Ligação Proteica , Ovinos/virologia , Testículo/citologia , Proteínas do Envelope Viral/análise , Proteínas do Envelope Viral/genéticaRESUMO
The Japanese encephalitis virus (JEV) is a leading cause of mosquito-borne viral encephalitis worldwide. Clinical symptoms other than encephalitis, on the other hand, are substantially more prevalent with JEV infection, demonstrating the relevance of peripheral pathophysiology. We studied the peripheral immunopathogenesis of JEV using IFNAR deficient (IFNAR-/-) mice infected with the SA14-14-2 strain under the BSL-2. The body weight and survival rate of infected-IFNAR-/-mice decreased significantly. Infected-IFNAR-/-mice's liver and spleen demonstrated obvious tissue damage and inflammatory cell infiltration. There was also extensive viral replication in the organs. IFN-α/ß protein expression was dramatically elevated in peripheral tissues and serum, although the related interferon-stimulated genes (ISGs) remained low in the spleen and liver of infected-IFNAR-/-animals. Consistently, the differentially expressed genes (DEGs) analysis using RNA-sequencing of spleens showed inflammatory cytokines upregulation, such as IL-6, TNF-α, and MCP-1, and IFN-γ associated cytokine storm. The infiltration of macrophages and neutrophils in the spleen and liver of SA14-14-2-infected IFNAR-/- mice was dramatically elevated. However, there was no significant difference in tissue damage, viral multiplication, or the production of IFNα/ß and inflammatory cytokines in the brain. Infection with the JEV SA14-14-2 strain resulted in a lethal peripheral inflammatory response and organ damage without encephalitis in IFNAR-/- mice. Our findings may help shed light on the peripheral immunopathogenesis associated with clinical JEV infection and aid in developing treatment options.
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Altered gut microbial ecology contributes to the development of metabolic diseases including obesity. However, studies based on different populations have generated conflicting results due to diet, environment, methodologies, etc. The aim of our study was to explore the association between gut microbiota and BMI in Chinese college students. The 16S next-generation sequencing (NGS) was used to test the gut microbiota of nine lean, nine overweight/obesity and ten normal-weight male college students. The differences in gut microbiota distribution among three groups were compared, and the relationship between the richness, diversity, composition of gut microbiota and BMI were analysed. The predominant phyla Bacteroidetes and Firmicutes were further confirmed by real-time PCR. Metagenomic biomarker discovery was conducted by linear discriminant analysis (LDA) effect size (LEfSe). NGS revealed that gut microbiota composition was different among three groups, but there was no difference in the abundance ratio of Firmicutes:Bacteroidetes. Several bacterial taxa were in linear relationship with BMI (positive relationship: uncultured bacterium (Bacteroides genus); negative relationship: Porphyromonadaceae, Acidaminococcaceae, Rikenellaceae, Desulfovibrionaceae, Blautia, Anaerotruncus, Parabacteroides, Alistipes). Moreover, gut microbiota diversity decreased with the increase in BMI. And LEfSe analysis indicated that Blautia, Anaerotruncus and its uncultured species were significantly enriched in the lean group (LDA score ≥ 3), Parasuterella and its uncultured species were significantly enriched in the overweight/obese groups (LDA score ≥ 3). In general, gut microbiota composition and microbial diversity were associated with BMI in Chinese male college students. Our results might enrich the understanding between gut microbiota and obesity.
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Bactérias/genética , Índice de Massa Corporal , Microbioma Gastrointestinal/genética , Sequenciamento de Nucleotídeos em Larga Escala , Sobrepeso/microbiologia , Povo Asiático , Biodiversidade , China , Humanos , Masculino , Adulto JovemRESUMO
Objective To elucidate the regulating effect of orf virus (ORFV) encoded ORF128 on NF-κB signaling pathway during the infection of HEK293T cells with ORFV and the underlying mechanism. Methods The ORF128 DNA sequences from ORFV/QH02/2010 strain were constructed into eukaryotic expression vectors pCMV-tag2B and pEGFP-N1. During viral infection of cells, the level of ORF128 mRNA was detected by reverse transcription PCR and the subcellular localization of ORF128 protein by laser confocal microscopy. A dual luciferase reporter assay system was used to analyze the regulating effect of ORF128 on NF-κB signaling pathway, and Western blot analysis to detect the nuclear translocation of NF-κBp65 and the phosphorylation of IκBα protein (p-IκBα). Results ORF128 protein was expressed at the early stage and localized in the cell nuclear during ORFV infection, and it inhibited the expression of NF-κB reporter luciferase activity. The expression of the protein blocked the nuclear translocation of NF-κBp65 and the degradation of p-IκBα. Conclusion ORF128 inhibits the activation of NF-κB signaling pathway by blocking the nuclear translocation of NF-κBp65 and the degradation of p-IκBα.
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Transdução de Sinais , Células HEK293 , Humanos , Proteínas I-kappa B , Inibidor de NF-kappaB alfa , NF-kappa B , Fases de Leitura Aberta , FosforilaçãoRESUMO
Activation of the DNA-dependent innate immune pathway plays a pivotal role in the host defense against poxvirus. Cyclic GMP-AMP synthase (cGAS) is a key cytosolic DNA sensor that produces the cyclic dinucleotide cGMP-AMP (cGAMP) upon activation, which triggers stimulator of interferon genes (STING), leading to type I Interferons (IFNs) production and an antiviral response. Ectromelia virus (ECTV) has emerged as a valuable model for investigating the host-Orthopoxvirus relationship. However, the role of cGas-Sting pathway in response to ECTV is not clearly understood. Here, we showed that murine cells (L929 and RAW264.7) mount type I IFN responses to ECTV that are dependent upon cGas, Sting, TANK binding kinase 1 (Tbk1), and interferon regulatory factor 3 (Irf3) signaling. Disruption of cGas or Sting expression in mouse macrophages blocked the type I IFN production and facilitated ECTV replication. Consistently, mice deficient in cGas or Sting exhibited lower type I IFN levels and higher viral loads, and are more susceptible to mousepox. Collectively, our study indicates that the cGas-Sting pathway is critical for sensing of ECTV infection, inducing the type I IFN production, and controlling ECTV replication.
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Vírus da Ectromelia/imunologia , Ectromelia Infecciosa/imunologia , Ectromelia Infecciosa/metabolismo , Imunidade Inata , Proteínas de Membrana/metabolismo , Nucleotidiltransferases/metabolismo , Transdução de Sinais , Animais , Chlorocebus aethiops , Ectromelia Infecciosa/virologia , Interações Hospedeiro-Patógeno , Humanos , Fator Regulador 3 de Interferon/metabolismo , Interferon Tipo I/biossíntese , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Células NIH 3T3 , Fosforilação , Proteínas Serina-Treonina Quinases/metabolismo , Células RAW 264.7 , Células Vero , Replicação ViralRESUMO
The ectromelia virus (ECTV) is a mouse specific Orthopoxvirus that causes lethal infection in some mouse strains. ECTV infection of these mouse strains has been used as a valuable model for understanding the interplay between Orthopoxvirus species and their hosts, including variola virus in humans. Although poxviruses encode numerous proteins required for DNA and RNA synthesis, and are less dependent on host functions than other DNA viruses, a detailed understanding of the host factors required for the replication of poxviruses is lacking. Heat shock protein 70 (Hsp70) isoforms have been reported to serve various roles in the replication cycle of numerous viruses. In the present study, microarray and reverse transcriptionquantitative polymerase chain reaction analysis were conducted to investigate the host gene expression profiles following ECTV infection in mice and cell cultures. The results indicated that one Hsp70 isoform, Hsp70 member 1B (Hspa1b), was highly upregulated during ECTV infection in vitro and in vivo. Subsequently, overexpression of Hspa1b protein and small interfering RNAmediated gene silencing of Hspa1b revealed that Hspa1b is required for efficient replication of ECTV. Furthermore, the results demonstrated that ECTV replication may be significantly suppressed by two chemical Hspa1b inhibitors: Quercetin and VER155008. In conclusion, the present study clearly demonstrated that ECTV infection upregulates the expression of Hspa1b in order to promote its replication. The dependence on Hsp70 may be used as a novel therapeutic target for the treatment of Orthopoxvirus infection.
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Vírus da Ectromelia/fisiologia , Ectromelia Infecciosa/genética , Ectromelia Infecciosa/virologia , Proteínas de Choque Térmico HSP70/genética , Interações Hospedeiro-Patógeno , Camundongos/virologia , Replicação Viral , Animais , Replicação do DNA , Masculino , Camundongos Endogâmicos BALB C , Regulação para CimaRESUMO
Asp-Glu-Ala-Asp (DEAD)-box polypeptide 5 (DDX5), also called p68, is a prototypical member of the large ATP-dependent RNA helicases family and is known to participate in all aspects of RNA metabolism ranging from transcription to translation, RNA decay, and miRNA processing. The roles of DDX5 in cell cycle regulation, tumorigenesis, apoptosis, cancer development, adipogenesis, Wnt-ß-catenin signaling, and viral infection have been established. Several RNA viruses have been reported to hijack DDX5 to facilitate various steps of their replication cycles. Furthermore, DDX5 can be bounded by the viral proteins of some viruses with unknown functions. Interestingly, an antiviral function of DDX5 has been reported during hepatitis B virus and myxoma virus infection. Thus, the precise roles of this apparently multifaceted protein remain largely obscure. Here, we provide a rapid and critical overview of the structure and functions of DDX5 with a particular emphasis on its role during virus infection.
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RNA Helicases DEAD-box/química , RNA Helicases DEAD-box/metabolismo , Vírus de DNA/efeitos dos fármacos , Vírus de RNA/fisiologia , Adipogenia , Animais , Antivirais/farmacologia , Ciclo Celular , RNA Helicases DEAD-box/farmacologia , Humanos , Modelos Moleculares , Conformação Proteica , Replicação ViralRESUMO
The classical swine fever virus C-strain vaccine (C-strain vaccine) plays a vital role in preventing and controlling the spread of classical swine fever (CSF). However, the protective mechanisms of C-strain vaccine and cellular immunity conferred by T cell receptors (TCRs) are less well defined. We aimed to analyse the association between the complementarity determining region 3 (CDR3) spectratype of αßTCR in CD4+ T cells and C-strain vaccine; and to find conserved CDR3 amino acid motifs in specific TCR α- and ß-chains. We found that the CDR3 spectratype showed dynamic changes correlating with C-strain vaccine immunisation and that TCR AV5S/8-3S/8-4S/14/38 and BV4S/6S/7S/15S/30 gene families showed clonal expansion in immunised pigs. The sequences of CDR3 from these clonally expanded T cells indicated a high frequency of the 'KLX' motif in the TCR α chain and the 'GGX' motif in ß chain, and Jα39, Jα43, Jß2.5 and Jß2.3 genes were also found in high frequency. To the best of our knowledge, this is the first report describing the dynamic changes of αßTCRs and conserved CDR3 amino acid motifs in CD4+ T cells from C-strain vaccine-immunised pigs, which will provide a basis for the development of high-efficiency epitope vaccines.
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Linfócitos T CD4-Positivos/imunologia , Vírus da Febre Suína Clássica/imunologia , Regiões Determinantes de Complementaridade/metabolismo , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Vacinas Virais/imunologia , Animais , Regiões Determinantes de Complementaridade/genética , Família Multigênica , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Análise de Sequência de DNA , Suínos , Vacinas Virais/administração & dosagemRESUMO
Ectromelia virus (ECTV) is the causative agent of mousepox, which has devastating effects in laboratory-mouse colonies and causes economic loss in biomedical research. More importantly, ECTV has been extensively used as an excellent model for studies of the pathogenesis and immunobiology of human smallpox. A rapid and sensitive SYBR Green I-based real-time PCR assay was developed and used for the detection and quantitation of orthopoxvirus by using ECTV in this study. Primers targeted to the highly conserved region of major core protein P4b gene of orthopoxvirus were designed and the standard plasmid was constructed. This assay was able to detect a minimum of 10 copies of standard DNA and 5 TCID50 units of ECTV. In addition, no cross-reactions were observed with two DNA viruses, such as herpes simplex virus and swine pseudorabies virus, and one RNA virus, vesicular stomatitis virus. Furthermore, intra- and inter-assay variability data showed that this method had a highly reproducibility and reliability. Moreover, the current assay was faster and had a higher sensitivity for detection of ECTV genomic DNA in cell cultured and clinical test samples. Therefore, the high sensitivity and reproducibility of this SYBR Green real-time PCR approach is a more effective method than the conventional PCR for ECTV diagnosis and quantitation.
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Vírus da Ectromelia/isolamento & purificação , Compostos Orgânicos/química , Orthopoxvirus/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real/métodos , Animais , Benzotiazóis , Chlorocebus aethiops , Diaminas , Ectromelia Infecciosa/virologia , Limite de Detecção , Masculino , Camundongos Endogâmicos C57BL , Quinolinas , Padrões de Referência , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Células VeroRESUMO
Contagious ecthyma is a highly contagious disease with worldwide distribution, which is caused by the Orf virus (ORFV) belonging to the Parapoxvirus. To study the alteration of host gene expression in response to ORFV infection at the transcriptional level, several young small-tailed Han sheep were inoculated with ORFV, and their oral mucosa tissue samples (T0, T3, T7 and T15) were collected on day 0, 3, 7 and 15 after ORFV infection respectively. RNA-seq transcriptome comparisons were performed, showing that 1928, 3219 and 2646 differentially expressed genes (DEGs) were identified among T3 vs. T0, T7 vs. T0, and T15 vs. T0 respectively. Gene Ontology (GO) analyses of the DEGs from these comparisons, revealed that ORFV might provoke vigorous immune response of the host cells during the early stage of infection. Moreover, GO and network analysis showed that positive and negative regulative mechanisms of apoptosis were integrated in the host cells through up or down-regulating the expression level of DEGs involved in apoptotic pathways, in order to reach a homeostasis of oral mucosa tissues during the exposure to ORFV infection. In conclusion, our study for the first time describes the direct effects of ORFV on the global host gene expression of its host using high-throughput RNA sequencing, which provides a resource for future characterizing the interaction mechanism between the mammalian host and ORFV.
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Ectima Contagioso/virologia , Mucosa Bucal/virologia , Vírus do Orf/genética , Transcriptoma , Animais , Apoptose , Vírus do Orf/isolamento & purificação , OvinosRESUMO
The T cell receptor (TCR) is a complex heterodimer that recognizes fragments of antigens as peptides and binds to major histocompatibility complex molecules. The TCR α and ß chains possess three hypervariable regions termed complementarity determining regions (CDR1, 2 and 3). CDR3 is responsible for recognizing processed antigen peptides. Immunoscope spectratyping is a simple technique for analyzing CDR3 polymorphisms and sequence length diversity, in order to investigate T cell function and the pattern of TCR utilization. The present study employed this technique to analyze CDR3 polymorphisms and the sequence length diversity of TCR α and ß chains in porcine CD4+ and CD8+ T cells. Polymerase chain reaction products of 19 TCR α variable regions (AV) and 20 TCR ß variable regions (BV) gene families obtained from the CD4+ and CD8+ T cells revealed a clear band following separation by 1.5% agarose gel electrophoresis, and each family exhibited >8 bands following separation by 6% sequencing gel electrophoresis. CDR3 spectratyping of all identified TCR AV and BV gene families in the sorted CD4+ and CD8+ T cells by GeneScan, demonstrated a standard Gaussian distribution with >8 peaks. CDR3 in CD4+ and CD8+ T cells demonstrated different expression patterns. The majority of CDR3 recombined in frame and the results revealed that there were 10 and 14 amino acid discrepancies between the longest and shortest CDR3 lengths in specific TCR AV and TCR BV gene families, respectively. The results demonstrated that CDR3 polymorphism and length diversity demonstrated different expression and utilization patterns in CD4+ and CD8+ T cells. These results may facilitate future research investigating the porcine TCR CDR3 gene repertoire as well as the functional complexity and specificity of the TCR molecule.
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Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Regiões Determinantes de Complementaridade/genética , Variação Genética , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Feminino , Expressão Gênica , Frequência do Gene , Família Multigênica , Análise de Sequência de DNA , SuínosRESUMO
Ectromelia virus (ECTV), the causative agent of mousepox, has emerged as a valuable model for investigating the host-Orthopoxvirus relationship as it relates to pathogenesis and the immune response. ECTV is a mouse-specific virus and causes high mortality in susceptible mice strains, including BALB/c and C3H, whereas C57BL/6 and 129 strains are resistant to the disease. To understand the host genetic factors in different mouse strains during the ECTV infection, we carried out a microarray analysis of spleen tissues derived from BALB/c and C57BL/6 mice, respectively, at 3 and 10 days after ECTV infection. Differential Expression of Genes (DEGs) analyses revealed distinct differences in the gene profiles of susceptible and resistant mice. The susceptible BALB/c mice generated more DEGs than the resistant C57BL/6 mice. Additionally, gene ontology and KEGG pathway analysis showed the DEGs of susceptible mice were involved in innate immunity, apoptosis, metabolism, and cancer-related pathways, while the DEGs of resistant mice were largely involved in MAPK signaling and leukocyte transendothelial migration. Furthermore, the BALB/c mice showed a strong induction of interferon-induced genes, which, however, were weaker in the C57BL/6 mice. Collectively, the differential transcriptome profiles of susceptible and resistant mouse strains with ECTV infection will be crucial for further uncovering the molecular mechanisms of the host-Orthopoxvirus interaction.
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Resistência à Doença/genética , Ectromelia Infecciosa/genética , Interações Hospedeiro-Parasita/genética , Transcriptoma/genética , Animais , Suscetibilidade a Doenças/virologia , Vírus da Ectromelia/patogenicidade , Ectromelia Infecciosa/patologia , Ectromelia Infecciosa/virologia , Regulação da Expressão Gênica , Imunidade Inata/genética , Interferons/genética , Camundongos , Baço/metabolismo , Baço/virologiaRESUMO
Toll-like receptors (TLRs) are a large family of germ-line encoded pattern recognition receptors (PRRs) that recognize pathogen-associated molecular patterns and evoke the relevant innate immune responses. TLR8 is a member of several endosome nucleic acid-sensing TLRs; however little attention has been paid to murine TLR8 (mTLR8) compared with other endosome nucleic acid-sensing TLRs. In the present study, mTLR8 was cloned using reverse transcription-polymerase chain reaction from murine peripheral blood mononuclear cells and its function in regulating innate immune response was characterized. The open reading frame of mTLR8 consists of 3,099 bps and encodes 1,032 amino acids. It contains typical leucine-rich repeats, a transmembrane domain and a Toll/interleukin-1 receptor domain, and it shares a high level of identity with other mammalian species. The expression of mTLR8 has been widely observed in different tissues, and higher expression levels of mTLR8 have mainly been detected in the heart, spleen and lung. Overexpression of mTLR8 is required for the activation of transcription factor nuclear factor-κB and the production of tumor necrosis factor-α. However, mTLR8 is not able to activate interferon regulatory factor 3 or activator protein 1, nor can it induce interferon-α in HEK293T cells. These results indicate that mTLR8, as an important PRR, is indeed functional and is vital role in the activation of innate immune responses. This study may aid in determining the molecular basis of the interactions between mTLR8 and pathogens.
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Receptor 8 Toll-Like/genética , Sequência de Aminoácidos , Animais , Clonagem Molecular , Cristalografia por Raios X , Perfilação da Expressão Gênica , Camundongos Endogâmicos C57BL , Dados de Sequência Molecular , NF-kappa B/metabolismo , Filogenia , Estrutura Secundária de Proteína , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Análise de Sequência de Proteína , Receptor 8 Toll-Like/química , Receptor 8 Toll-Like/metabolismo , Fator de Necrose Tumoral alfa/metabolismoRESUMO
OBJECTIVE: To study the structural characteristics of the mouse DNA-dependent activator of interferon-regulatory factors (DAI) and its related molecular mechanism in anti-viral innate immune responses and signal transduction. METHODS: The coding sequence of mouse DAI gene was amplified from splenic mononuclear cells by reverse transcription-PCR, and the genetic evolution and molecular structure of the mouse DAI gene were analyzed by bioinformatics softwares. After mouse DAI was stimulated by poly(dA-dT) and poly(dG-dC), the nuclear factor κB (NF-κB) and interferon-beta (IFN-ß) promoter-driven luciferase activity were detected by dual-luciferase reporter assay system. RESULTS: The open reading frame (ORF) of the cloned mouse DAI sequence was 1236 bp, encoding 411 amino acids, which exhibited identity with the corresponding sequences of cattle, pig, rat and other mammals ranging from 60%, 63.1%, 84%, and it contained two Z-DNA domains (Zα and Zß), DNA binding region (D3) and signaling domain (SD). The stimulation of poly (dA-dT) increased the expressions of mouse DAI activated transcription factors NF-κB and IFN-ß promoter. However, the stimulation of poly(dG-dC) only induced the activation of NF-κB but not IFN-ß promoter. CONCLUSION: Mouse DAI as an important cytosolic DNA sensor, is responsible for the recognition of A/T or G/C-rich DNA derived from DNA virus. It may play an important role in anti-viral innate immune responses.
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DNA/metabolismo , Glicoproteínas/genética , Glicoproteínas/metabolismo , Camundongos/genética , Animais , Bovinos , Feminino , Glicoproteínas/química , Humanos , Masculino , Mamíferos/classificação , Mamíferos/genética , Camundongos/metabolismo , Camundongos Endogâmicos C57BL , Estrutura Terciária de Proteína , Proteínas de Ligação a RNA , Ratos , SuínosRESUMO
In this paper, we investigated the enhancement of adjuvant effects of porcine IL-2 (pIL-2) by packaging it into a solid lipid nanoparticle (SLN) delivery system. SLN-pIL-2 was prepared using hydrogenated castor oil and Polylactide-co-glycolide by double emulsion solvent evaporation methods (w/o/w). In animal trials, BALB/c mice were immunized with inactivated foot and mouth disease virus (FMDV) antigen combined with the SLN-pIL-2 adjuvant on days 0 and 14. Antibody titer, splenocyte proliferation, and secretion of IFN-γ and IL-4 cytokines were determined. Our results showed that SLN-pIL-2 could significantly enhance FMDV-specific antibody level compared with recombinant pIL-2 alone (p<0.05). In addition, SLN-pIL-2 significantly increased the proliferative responses of antigen-specific spleen cells. Furthermore, SLN-pIL-2 induced the secretion of IFN-γ at a level higher than that induced by recombinant pIL-2 alone. Our results indicate that packaging recombinant pIL-2 in SLNs can be an effective way of boosting the effectiveness of pIL-2 as an adjuvant to enhance immune responses of vaccines.
Assuntos
Adjuvantes Imunológicos/farmacologia , Interleucina-2/farmacologia , Lipídeos/imunologia , Nanopartículas/uso terapêutico , Suínos/imunologia , Adjuvantes Imunológicos/administração & dosagem , Animais , Anticorpos Antivirais/sangue , Proliferação de Células , Feminino , Febre Aftosa/imunologia , Febre Aftosa/prevenção & controle , Vírus da Febre Aftosa/imunologia , Imunização , Interleucina-2/administração & dosagem , Camundongos , Camundongos Endogâmicos BALB C , Microscopia Eletrônica de Transmissão , Nanopartículas/ultraestrutura , Distribuição AleatóriaRESUMO
In this study, the interactions of classical swine fever virus (CSFV) C-strain and the virulent GSLZ strain with mouse bone marrow-derived immature dendritic cells (BM-imDCs) were investigated for the first time. Both the C-strain and the virulent GSLZ strain could effectively infect and replicate in mouse BM-imDCs. C-strain-infected BM-imDCs showed a greatly enhanced degree of maturation, and could effectively promote the expansion and proliferation of allogeneic naive T cells. The C-strain induced a stronger Th1 response. Infection with the virulent GSLZ strain had no obvious influence on cell maturation or lymphocyte proliferation, and failed to induce any obvious immune response. The results of this study provided initial information for research of the immunologic mechanisms of CSFV using mouse DCs as the model cells.