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BACKGROUND: The low pathogenic H9N2 AIV caused the serious impact on the poultry industry and public safety. Our purpose was to investigate the molecular evolutionary characteristics of the new isolated H9N2 virus and investigate the intracellular target protein of H9N2 AIV replication in sensitive cells. METHODS: AIV A/chicken/Shandong/LY1/2017 (H9N2) was isolated from the cloaca of the healthy chicken in Shandong, and the full-length eight gene segments of this isolated H9N2 AIV were amplified by RT-PCR and analyzed. MDCK cells were used as the target cell model, and VOPBA assay and LC-MS/MS were carried out to identify the virus-binding protein of H9N2 AIV. MDCK cells were pre-treated with the special antibody and siRNA, and treated with H9N2 AIV to detect the virus replication. Additionally, Vimentin-pcDNA3.0 was successfully constructed, and transinfected into MDCK cells, and then H9N2 AIV mRNA was detected with RT-PCR. RESULTS: Phylogenetic analysis revealed that HA, NA, PB2, PB1, PA, NP and M seven genes of the isolated H9N2 AIV were derived from A/Chicken/Shanghai/F/98, while NS gene was derived from A/Duck/Hong Kong/Y439/97. The cleavage site sequence of HA gene of the isolated H9N2 AIV was a PARSSR G pattern, and the left side sequence (224 ~ 229) of receptor binding site was NGQQGR pattern, which were similar to that of A/Chicken/Shanghai/F/98. Following VOPBA assay, we found one protein of about 50KDa binding to H9N2 AIV, and the results of LC-MS/MS analysis proved that vimentin was the vital protein binding to H9N2 AIV. The pre-incubation of the specific antibody and siRNA decreased the viral RNA level in MDCK cells treated with H9N2 AIV. Furthermore, we found that over-expressed vimentin increased H9N2 AIV replication in MDCK cells. CONCLUSIONS: These findings suggested that the isolated H9N2 AIV might be a recent clinical common H9N2 strain, and vimentin protein might be one vital factor for H9N2 AIV replication in MDCK cells, which might be a novel target for design and development of antiviral drug.
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Evolución Molecular , Subtipo H9N2 del Virus de la Influenza A/genética , Filogenia , Vimentina/farmacología , Proteínas Virales/genética , Replicación Viral/efectos de los fármacos , Animales , Pollos/virología , China , Perros , Subtipo H9N2 del Virus de la Influenza A/fisiología , Células de Riñón Canino Madin Darby , Aves de Corral/virología , Enfermedades de las Aves de Corral/virologíaRESUMEN
The bursa of Fabricius (BF) is the acknowledged central humoural immune organ unique to birds and plays a vital role in B lymphocyte development. In addition, the unique molecular immune features of bursal-derived biological peptides involved in B cell development are rarely reported. In this paper, a novel bursal heptapeptide (BP7) with the sequence GGCDGAA was isolated from the BF and was shown to enhance the monoclonal antibody production of a hybridoma. A mouse immunization experiment showed that mice immunized with an AIV antigen and BP7 produced strong antibody responses and cell-mediated immune responses. Additionally, BP7 stimulated increased mRNA levels of sIgM in immature mouse WEHI-231 B cells. Gene microarray results confirmed that BP7 regulated 2465 differentially expressed genes in BP7-treated WEHI-231 cells and induced 13 signalling pathways and various immune-related functional processes. Furthermore, we found that BP7 stimulated WEHI-231 cell autophagy and AMPK-ULK1 phosphorylation and regulated Bcl-2 protein expression. Finally, chicken immunization showed that BP7 enhanced the potential antibody and cytokine responses to the AIV antigen. These results suggested that BP7 might be an active biological factor that functions as a potential immunopotentiator, which provided some novel insights into the molecular mechanisms of the effects of bursal peptides on immune functions and B cell differentiation.
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Proteínas Aviares/genética , Linfocitos B/inmunología , Bolsa de Fabricio/inmunología , Pollos/inmunología , Inmunidad Celular , Activación de Linfocitos/inmunología , Animales , Formación de Anticuerpos , Proteínas Aviares/inmunología , Femenino , Regulación de la Expresión Génica/inmunología , Inmunización , Inmunoglobulina M/inmunología , Ratones , Ratones Endogámicos BALB C , Análisis por Matrices de Proteínas , ARN Mensajero/metabolismoRESUMEN
The bursa of Fabricius (BF) is the key humoral immune organ unique to birds, and is critical for early B-lymphocyte proliferation and differentiation. However, the molecular basis and mechanisms through which the BF regulates B cell development are not fully understood. In this study, we isolated and identified a new bursal peptide (BP8, AGHTKKAP) by RP-HPLC and MALDI-TOF-MS. BP8 promoted colony-forming pre-B formation, bound B cell precursor, regulated B cell development in vitro as well as in vivo, upstream of the EBF-E2A-Pax5 regulatory complex and increased immunoglobulin secretion. These data revealed a bursal-derived multifunctional factor BP8 as a novel biomaterial which is essential for the development of the immune system. This study elucidates further the mechanisms involved in humoral immune system and has implications in treating human diseases.
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Linfocitos B/citología , Bolsa de Fabricio/química , Péptidos/farmacología , Secuencia de Aminoácidos , Animales , Linfocitos B/efectos de los fármacos , Linfocitos B/inmunología , Bolsa de Fabricio/inmunología , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Pollos , Femenino , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Datos de Secuencia Molecular , Péptidos/inmunología , Péptidos/aislamiento & purificaciónRESUMEN
Bursa of Fabricius is the humoral immune system for B cell differentiation and antibody production. Bursopentine (BP5) is a novel immunomodulatory peptide and significantly stimulated an antigen-specific immune response in mice. BP5 was also found to protect LPS-activated murine peritoneal macrophages from oxidative stress. In this study, the effects of BP5 on B cell development were examined. The results suggested that BP5 markedly promoted B cell development by increasing CFU-pre B, and affected the redox homeostasis regulation of B cells. To study the molecular mechanism of effect of bursal-derived BP5, this research utilized 2D-E and MALDI-TOF/TOF to analyze the differentially expressed proteins of BP5-treated WEHI-231 cells. The results showed that BP5 affected the redox homeostasis regulation of WEHI-231 cells and induced alterations in the protein expression profiles related to the oxidoreduction coenzyme metabolic process, precursor metabolites and energy, proteolysis, RNA splicing and translation and cellular process, respectively. BP5 also induced glucose-6-phosphate dehydrogenase (G6PD) activity, an essential anti-oxidant cofactor. We found that the redox homeostasis regulation effect of BP5 was reduced in G6PD-deficient cells. These data suggested that BP5 affected the redox balance toward reducing conditions by promoting the expression of G6PD, which in turn regulated the glutathione redox cycle and other processes.
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Adyuvantes Inmunológicos/farmacología , Linfocitos B/inmunología , Diferenciación Celular/efectos de los fármacos , Oligopéptidos/farmacología , Estrés Oxidativo/efectos de los fármacos , Animales , Linfocitos B/citología , Diferenciación Celular/genética , Diferenciación Celular/inmunología , Línea Celular , Glucosafosfato Deshidrogenasa/genética , Glucosafosfato Deshidrogenasa/inmunología , Homeostasis/efectos de los fármacos , Homeostasis/genética , Homeostasis/inmunología , Lipopolisacáridos/farmacología , Macrófagos Peritoneales/citología , Macrófagos Peritoneales/inmunología , Ratones , Oxidación-Reducción/efectos de los fármacos , Estrés Oxidativo/genética , Estrés Oxidativo/inmunologíaRESUMEN
The bursa of Fabricius, the acknowledged central humoral immune organ, plays a vital role in B lymphocyte differentiation. However, there are few reports of the molecular basis of the mechanism on immune induction and potential antitumor activity of bursal-derived peptides. In this paper, a novel bursal-derived pentapeptide-II (BPP-II, MTLTG) was isolated and exerted immunomodulatory functions on antibody responses in vitro. Gene microarray analyses demonstrated that BPP-II regulated expression of 2478 genes in a mouse-derived hybridoma cell line. Immune-related gene ontology functional procedures were employed for further functional analysis. Furthermore, the majority of BPP-II-regulated pathways were associated with immune responses and tumor processes. Moreover, BPP-II exhibited immunomodulatory effects on antigen-specific immune responses in vivo, including enhancement of avian influenza virus (H9N2 subtype)-specific antibody and cytokine production and modification of T cell immunophenotypes and lymphocyte proliferation. Finally, BPP-II triggered p53 expression and stabilization and selectively inhibited tumor cell proliferation. These data identified the multifunctional factor, BPP-II, as a novel biomaterial representing an important linking between the humoral central immune system and immune induction, including antitumor. Information generated in this study elucidates further the mechanisms involved in humoral immune system and represents the potential basis of effective immunotherapeutic strategies for treating human tumors and immune improvement.
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Factores Inmunológicos/farmacología , Subtipo H9N2 del Virus de la Influenza A/metabolismo , Neoplasias/inmunología , Oligopéptidos/farmacología , Linfocitos T/inmunología , Animales , Anticuerpos Antivirales/inmunología , Bolsa de Fabricio/química , Bolsa de Fabricio/inmunología , Línea Celular Tumoral , Pollos/inmunología , Citocinas/inmunología , Femenino , Humanos , Factores Inmunológicos/química , Factores Inmunológicos/inmunología , Factores Inmunológicos/aislamiento & purificación , Ratones , Ratones Endogámicos BALB C , Neoplasias/tratamiento farmacológico , Neoplasias/patología , Oligopéptidos/química , Oligopéptidos/inmunología , Oligopéptidos/aislamiento & purificación , Proteína p53 Supresora de Tumor/inmunologíaRESUMEN
Constructing ecological security pattern and identifying ecological important areas are the focus of current research on regional ecological security. With Ningbo City as a case study area, we identified ecological sources by remote sensing ecological index, the ecological corridors and pinch point by circuit theory model, and the minimum spanning tree and cuts by graph theory algorithm. The results showed that there were 203 ecological sources in Ningbo, and that the main type of land cover was forest, including a small amount of paddy fields and flooded vegetation. There were 368 ecological corridors with a total length of 573.42 km, being dense in the southwest and sparse in the northeast. There were 91 ecological pinch points, which mainly distributed between coastal areas and closely related ecological sources. According to current situation, we put forward the optimization strategy with 187 primary corridors, 181 secondary corridors, 50 ecological restoration priority areas and 59 long-term ecological restoration areas. The optimization strategy combined with graph theory and circuit theory model would provide a refe-rence for the constructing of ecological security pattern.
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Ecología , Ecosistema , Conservación de los Recursos Naturales , Tecnología de Sensores Remotos , BosquesRESUMEN
In this study, we reported the complete chloroplast genome sequence of Clivia robusta for the first time. The complete chloroplast genome of C. robusta was 157,130 bp in length, containing a large single-copy region (LSC, 85,430 bp), a small single-copy region (SSC, 18,278 bp), and two inverted repeat regions (IRs, 26,711 bp). The overall GC content was 38.01%. The chloroplast genome contained 128 genes in total, including 86 protein-coding, 34 tRNA, and eight rRNA genes. The phylogenetic tree showed that C. robusta formed a monophyletic clade with other Clivia species.
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OBJECTIVE: To detect the gene mutations of beta-myosin heavy chain gene (MYH7) in Chinese pedigrees with hypertrophic cardiomyopathy (HCM), and to analyze the correlation between the genotype and phenotype. METHODS: Exons 3, 5, 7-9, 11-16 and 18-23 of the MYH7 gene were amplified with PCR in three Chinese pedigrees with HCM. The products were sequenced. Sequence alignment between the detected and the standard sequences was performed. RESULTS: A missense mutation of Thr441Met in exon 14 was identified in a pedigree, which was not detected in the controls. Several synonymous mutations of MYH7 gene were detected in the three pedigrees. CONCLUSION: The mutation of Thr441Met, located in the actin binding domain of the globular head, was first identified in Chinese. It probably caused HCM. HCM is a heterogeneous disease. Many factors are involved in the process of its occurrence and development.
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Cardiomiopatía Hipertrófica/genética , Análisis Mutacional de ADN , Mutación , Cadenas Pesadas de Miosina/genética , Linaje , Adulto , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Miosinas Cardíacas , Femenino , Genotipo , Humanos , Masculino , Persona de Mediana Edad , Datos de Secuencia Molecular , Cadenas Pesadas de Miosina/química , FenotipoRESUMEN
OBJECTIVE: To detect gene mutations on beta-myosin heavy chain gene MYH7 in 3 Chinese families with hypertrophic cardiomyopathy (HCM), and to analyze the correlation between genotype and phenotype. METHODS: A denaturing high-performance liquid chromatography (DHPLC) and sequencing mutation screening of the exons (exon3-23) coding for MYH7 gene were performed in 3 Chinese families with HCM. RESULTS: In this study, we identified several mutations in MYH7. A mutation of Thr441Met previously reported in a patient with Laing distal myopathy was first identified in one Chinese pedigree. CONCLUSION: This study illustrated the high frequency of mutation in MYH7 gene in Chinese HCM families. Different mutations and carriers of the MYH7 gene present phenotypic heterogeneity. Mutation screening and analysis in HCM family could therefore facilitate the early HCM diagnosis and would be helpful for the prediction, prevention and early treatment of HCM linked with MYH7 gene mutation.
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Miosinas Cardíacas/genética , Cardiomiopatía Hipertrófica Familiar/genética , Mutación , Cadenas Pesadas de Miosina/genética , Adolescente , Adulto , Pueblo Asiatico/genética , Estudios de Casos y Controles , Niño , Análisis Mutacional de ADN , Exones , Femenino , Genotipo , Humanos , Masculino , Persona de Mediana Edad , Linaje , Fenotipo , Adulto JovenRESUMEN
The complete chloroplast genome of Clivia miniata var. citrina was assembled and subjected to phylogenetic analysis in this study. The complete chloroplast genome of C. miniata var. citrina was 158,112 bp in length, containing a large single-copy region (LSC, 86,202 bp), a small single-copy region (SSC, 18,334 bp), and two inverted repeat regions (IRs, 26,788 bp). The GC content was 37.97%. A total of 130 genes were annotated, including 86 protein-coding genes, 36 tRNA and 8 rRNA genes. Phylogenetic analysis showed that C. miniata var. citrina was the most related with C. miniata and they formed a monophyletic group that was sister to the clade of Hippeastrum, Leucojum, Narcissus and Lysoris.
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Clivia caulescens is an evergreen herbaceous flower with high ornamental value. In this study, we report its complete chloroplast genome sequence. The whole chloroplast genome was 158,149 bp in length, with a large single copy region (LSC, 86,250 bp), a small single copy region (SSC, 18,343 bp), and two inverted repeat regions (IRs, 26,778 bp). The overall GC content was 37.91%. There were 128 genes annotated, including 86 protein-coding genes, 34 tRNA genes, and eight rRNA genes. The phylogenetic tree showed that C. caulescens formed a monophyletic clade with C. miniata, C. miniata var. aurea, and C. gardenii.
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BACKGROUND: The Bursa of Fabricius is an acknowledged central humoral immune organ unique to birds, which provides an ideal research model on the immature B cell development. OBJECTIVE: In this article, our motivation is to study the role on sIgM and establish the molecular basis and functional processes of Bursal Hexapeptide (BHP) in avian immature B cells DT40 cell lines. METHODS: In this article, we detected the expressions of sIgM mRNA with qPCR in DT40 cells with BHP treatment, and investigated the gene expression profiles of BHP-treated DT40 cells, employing microarray analyses. Also, to validate the differentially expressed genes, we performed KEGG pathway and Gene Ontology analysis in the BHP-treated DT40 cells. Finally, we comparatively analyzed the similar regulated genes and their involved immune functional processes between DT40 cell and mouse immature B cell line WEHI231 cell with BHP treatment. RESULTS: Following the proposed framework, we proved that the BHP enhanced the mRNA expression levels of IgM in DT40 cells, and induced 460 upregulated genes and 460 downregulated genes in BHP-treated DT40 cells. The pathway analysis showed that the differentially regulated genes in DT40 cell line with BHP treatment were involved in 12 enrichment pathways, in which Toll-like receptor signaling pathway was the vital pathways, and cytokine-cytokine receptor interaction and Jak-STAT signaling pathway were another two important pathways in BHP-treated DT40 cells. Moreover, BHP induced the immune related biological processes in BHP-treated DT40 cells, including T cell related, cytokine related, lymphocyte related, and innate immune response GO terms. Finally, the comparatively analysis showed that there were two downregulated genes GATA3 and IFNG to be found co-existed among the differentially expressed genes in BHP-treated DT40 cell and WEHI231 cells, which shared some same immune related functional processes in both cell lines. CONCLUSION: After the applying the framework, we proved the inducing roles and the gene expression profiles of BHP on avian immature B cells, and verified some molecular basis from the KEGG and GO analysis. These results provided the insight for mechanism on immature B cell differentiation, and offer the essential direction for the vaccine improvement.
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Oligopéptidos/metabolismo , Células Precursoras de Linfocitos B/metabolismo , Animales , Bolsa de Fabricio , Línea Celular , Pollos , Inmunidad Innata , Inmunoglobulina M/genética , Inmunoglobulina M/metabolismo , Ratones , Oligopéptidos/farmacología , Células Precursoras de Linfocitos B/efectos de los fármacos , Células Precursoras de Linfocitos B/inmunología , ARN Mensajero/metabolismo , Transducción de Señal , Especificidad de la Especie , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunología , Linfocitos T/metabolismoRESUMEN
BACKGROUND: Bursa of Fabricius is the acknowledged central humoral immune organ. The bursal-derived peptides play the important roles on the immature B cell development and antibody production. OBJECTIVES: Here we explored the functions of the new isolated bursal hexapeptide and pentapeptide on the humoral, cellular immune response and antigen presentation to Avian Influenza Virus (AIV) vaccine in mice immunization. METHODS: The bursa extract samples were purified following RP HPLC method, and were analyzed with MS/MS to identify the amino acid sequences. Mice were twice subcutaneously injected with AIV inactivated vaccine plus with two new isolated bursal peptides at three dosages, respectively. On two weeks after the second immunization, sera samples were collected from the immunized mice to measure AIV-specific IgG antibody levels and HI antibody titers. Also, on 7th day after the second immunization, lymphocytes were isolated from the immunized mice to detect T cell subtype and lymphocyte viabilities, and the expressions of co-stimulatory molecule on dendritic cells in the immunized mice. RESULTS: Two new bursal hexapeptide and pentapeptide with amino acid sequences KGNRVY and MPPTH were isolated, respectively. Our investigation proved the strong regulatory roles of bursal hexapeptide on AIV-specific IgG levels and HI antibody titers, and lymphocyte viabilities, and the significant increased T cells subpopulation and expressions of MHCII molecule on dendritic cells in the immunized mice. Moreover, our findings verified the significantly enhanced AIV-specific IgG antibody and HI titers, and the strong increased T cell subpopulation and expressions of CD40 molecule on dendritic cells in the mice immunized with AIV vaccine and bursal pentapeptide. CONCLUSION: We isolated and identified two new hexapeptide and pentapeptide from bursa, and proved that these two bursal peptides effectively induced the AIV-specific antibody, T cell and antigen presentation immune responses, which provided an experimental basis for the further clinical application of the bursal derived active peptide on the vaccine improvement.
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Bolsa de Fabricio/química , Subtipo H9N2 del Virus de la Influenza A/inmunología , Vacunas contra la Influenza/inmunología , Oligopéptidos/química , Animales , Anticuerpos/metabolismo , Formación de Anticuerpos , Bolsa de Fabricio/inmunología , Antígenos CD40/metabolismo , Línea Celular , Supervivencia Celular/efectos de los fármacos , Pollos , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Femenino , Inmunidad Humoral , Gripe Aviar/prevención & control , Gripe Aviar/virología , Ratones Endogámicos BALB C , Oligopéptidos/inmunología , Linfocitos T/citología , Linfocitos T/efectos de los fármacosRESUMEN
The bursa of Fabricius is an acknowledged central humoral immune organ unique to birds, which is vital to B cell differentiation and antibody production. However, the function and mechanism of the biological active peptide isolated from bursa on B cell development and autophagy were less reported. In this study, we isolated a new oligopeptide with nine amino acids Leu-Met-Thr-Phe-Arg-Asn-Glu-Gly-Thr from avian bursa following RP-HPLC, MODIL-TOP-MS, and MS/MS, which was named after BP9. The results of immunization experiments showed that mice injected with 0.01 and 0.05 mg/mL BP9 plus JEV vaccine generated the significant increased antibody levels, compared to those injected with JEV vaccine only. The microarray analysis on the molecular basis of BP9-treated immature B cell showed that vast genes were involved in various immune-related biological processes in BP9-treated WEHI-231 cells, among which the regulation of cytokine production and T cell activation were both major immune-related processes in WEHI-231 cells with BP9 treatment following network analysis. Also, the differentially regulated genes were found to be involved in four significantly enriched pathways in BP9-treated WEHI-231 cells. Finally, we proved that BP9 induced the autophagy formation, regulated the gene and protein expressions related to autophagy in immature B cell, and stimulated AMPK-ULK1 phosphorylation expression. These results suggested that BP9 might be a strong bursal-derived active peptide on antibody response, B cell differentiation, and autophagy in immature B cells, which provided the linking among humoral immunity, B cell differentiation, and autophagy and offered the important reference for the effective immunotherapeutic strategies and immune improvement.
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Anticuerpos Antivirales/sangre , Autofagia , Linfocitos B/inmunología , Bolsa de Fabricio/química , Inmunidad Humoral , Oligopéptidos/inmunología , Animales , Bolsa de Fabricio/inmunología , Diferenciación Celular/inmunología , Línea Celular , Pollos , Femenino , Vacunas contra la Encefalitis Japonesa/inmunología , Ratones , Ratones Endogámicos BALB C , Análisis de Matrices TisularesRESUMEN
There is compelling evidence indicating that reduction of high-density lipoprotein (HDL) level is associated with increased risk of Alzheimer's disease (AD). It is known that the levels of HDL are regulated by cholesteryl ester transfer protein (CETP) and several single nucleotide polymorphisms (SNPs) in the CETP gene have been shown to be associated with the levels of HDL. Therefore, it is assumed that the CETP gene is a reasonable candidate for modifying the susceptibility in AD. In the present study, we investigated the association of four CETP SNPs (D442G, L296Q, Taq1B and I405V) with the risk for sporadic AD in Northern Han-Chinese. One hundred and seven AD cases and 115 age and gender-matched controls were genotyped by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), denaturing high performance liquid chromatography (DHPLC) and DNA sequencing. The frequency of DG genotype (P=0.035) or G allele (P=0.038) for the CETP (D442G) polymorphism was greater in control subjects than in AD patients. The age- and sex-adjusted odds radio for DG vs. DD genotype was 0.202 (95% CI 0.043-0.958, P=0.044). When the sample was stratified by APOE epsilon4 carrier status, the same tendency (P=0.042 for DG genotype, P=0.046 for G allele) was observed in the presence of APOE epsilon4, but not in the absence of APOE epsilon4 (P=0.284 for DG genotype, P=0.298 for G allele). However, these results became not statistically significant after correcting for multiple testing (Bonferroni) because of limited number of our sample. Our current results suggest that G allele of CETP D442G may have a potential protective effect against the development of AD, especially in APOE epsilon4 carriers, in Northern Han-Chinese, possibly through regulating the HDL level in the brain.
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Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/metabolismo , Apolipoproteína E4/genética , Proteínas de Transferencia de Ésteres de Colesterol/genética , Predisposición Genética a la Enfermedad/genética , Polimorfismo Genético/genética , Distribución por Edad , Anciano , Anciano de 80 o más Años , Enfermedad de Alzheimer/etnología , Pueblo Asiatico/etnología , Pueblo Asiatico/genética , Análisis Mutacional de ADN , Femenino , Frecuencia de los Genes/genética , Marcadores Genéticos/genética , Predisposición Genética a la Enfermedad/etnología , Pruebas Genéticas , Genotipo , Heterocigoto , Humanos , Lipoproteínas HDL/metabolismo , Masculino , Distribución por SexoRESUMEN
@#Abstract: Objective To evaluate the value and significance of rifampicin-resistant real-time fluorescence quantitative nucleic acid amplification detection technology (GeneXpert MTB/RIF) in the diagnosis of pulmonary tuberculosis. Methods The clinical data of 228 patients with suspected pulmonary tuberculosis, who admitted to Hebei Chest Hospital from January 2018 to December 2019, were analyzed retrospectively. The sputum was collected for GeneXpert MTB/RIF, sandwich cup liquid-based bacterial acid-fast staining smear microscopy (referred to as “sandwich cup method”) and Loop-Mediated isothermal amplification (referred to as “LAMP method”) and the results were statistically analyzed by SPSS 17.0 software. Results Among the 228 patients with suspected cases, 200 cases were clinically diagnosed as pulmonary tuberculosis and 28 were non-tuberculosis. The positive detection rate of GeneXpert MTB/RIF (81.0%, 162/200) was significantly higher than that of sandwich cup method (62.5%, 125/200) and LAMP method (72.5%,145/200) (χ2=16.885, 4.049, P<0.05). Taking clinical diagnosis as gold standard, the sensitivity of GeneXpert MTB/RIF (80.00%,160/200) was significantly higher than that of sandwich cup method (60.00%, 120/200) and LAMP method (70.50%, 141/200) (χ2=19.048, 4.846, P<0.05). The diagnostic consistency of GeneXpert MTB/RIF (K=0.73) was higher than that of sandwich cup method (K=0.39) and LAMP method (K=0.56). Conclusions The GeneXpert MTB/RIF detection method is rapid and simple, and can diagnose pulmonary tuberculosis rapidly and simultaneously detect rifampicin resistance of Mycobacterium tuberculosis with high sensitivity. It has high clinical value for early diagnosis of pulmonary tuberculosis and guidance of treatment in general and specialized hospitals.
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BACKGROUND: The bursa of Fabricius (BF) is an acknowledged central immune organ, and is important to B cell differentiation. Bursal hexapeptide (BHP) is the recently reported bursalderived peptide, while its inducing function on immune response is uncertain. OBJECTIVES: The main objective of this study was to analyze the immune responses to JEV vaccine in mice induced by BHP plus JEV vaccine, and to detect the signal and biological functions of BHP on immature B cells. METHODS: Mice were immunized with Japanese encephalitis virus (JEV) vaccine and BHP from 0.01 mg/mL to 0.25 mg/mL to detect antibody response and cellular immune response, respectively. The production of IgG, IgG1 and IgG2a specific to JEV in serum from immunized mice were measured by ELISA, and T cell subpopulation from immunized mice were detected with using fluorochrome conjugated mAbs of the corresponding PE-Cys/FITC/PE by flow cytometry. Spleen cells from all immunized mice were harvested after one week of second immunization for lymphocyte proliferation assay. Mouse immature B cell WEHI-231 cell was treated with 0.01µg/mL BHP for 4h, and analyzed the involved biological function and pathway of differentially expressed genes with gene microarray. RESULTS: BHP co-immunization with JEV vaccine generated significant increased antibody levels, neutralizing antibody titers and spleen lymphocyte viability, compared to that of vaccine control. The subpopulations of T cells in spleen lymphocytes were significantly modified in the mice coimmunized with JEV vaccine and BHP. The analysis results of gene expression profiles of WEHI- 231 mouse immature B cells with BHP treatment showed that the regulated genes with BHP treatment were involved various immune related biological functions, including proliferation and activation of lymphocyte and T cell, T cell mediated immunity and regulation of adaptive immune response. Furthermore, BHP stimulated three significant enriched pathways, including amphetamine addiction, long-term potentiation, and RIG-I-like receptor signaling pathway. CONCLUSION: Our results indicated BHP induced significant humoral and cellular immunity to JEV vaccine, and regulated various biological processes and signalling related to immune activation in immature B cells. These results proposed the immunomodulatory function and mechanism of BHP on immune induction, which provided the novel insight on the candidate reagent for immune improvement.
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Formación de Anticuerpos/efectos de los fármacos , Virus de la Encefalitis Japonesa (Especie)/inmunología , Oligopéptidos/farmacología , Células Precursoras de Linfocitos B/efectos de los fármacos , Vacunas Virales/inmunología , Animales , Bolsa de Fabricio/metabolismo , Diferenciación Celular , Supervivencia Celular , Femenino , Inmunidad Celular , Inmunidad Humoral , Inmunoglobulina G/sangre , Ratones Endogámicos BALB C , Oligopéptidos/síntesis química , Oligopéptidos/inmunología , Células Precursoras de Linfocitos B/inmunología , Transducción de Señal , VacunaciónRESUMEN
BACKGROUND: In the recent past, many studies have been focused on extracts of BF and multiple biologically active factors and their effects on humoral immune system in chickens and birds. However, the mechanism of those immunomodulatory peptides on the B lineage cells proliferation and antibody production in chicken is fairly unknown. DT40 cell line, an avian leucosis virus-induced chicken pre-B cell line, expresses immunoglobulin M (IgM) isotype B cell reporter in the plasma membrane. There are many evidences suggesting that DT40 cells are best characterized as a bursal stem cell line. Because of the unique characteristics of DT40 cell line, it has been widely used to observe biological processes of pre-B lymphocyte cell within living cells. METHODS: The chicken B cell line DT40 was cultured in Roswell Park Memorial Institute (RPMI) 1640 medium and cytotoxicity was studied. Also, effect of BP5 on cell proliferation and cell cycle distribution of DT40 cells was studied. Also, the effect of BP5 on sIgM mRNA expression was studied by using real-time PCR. OBJECTIVES: To investigat the effects of Bursopentin (Cys-Lys-Arg-Val-Tyr, BP5) on a chicken promyelocyte cell line DT40, assays of cell proliferation, cell cycle distribution, detection of surface immunoglobulin G (sIgM) mRNA expression and gene microarray analysis were performed. RESULTS: The results showed that BP5 displayed concentration-dependent effects on the proliferation, cell cycle, and sIgM mRNA expression in DT40 cells. And the analysis of expression profiles identified a signature set of 3022 genes (1254 up regulated genes, 1762 down regulated genes), which clearly discriminated the BP5-treated DT40 cells from control with high certainty (P≤0.02). The results of microarray analysis were confirmed by quantitative reverse transcription-polymerase chain reaction for 12 of the differentially expressed genes. CONCLUSION: Theses findings showed the immuno-activity effect of BP5 on B lymphocyte and indicated that BP5 treatment regulated eight signaling pathways, in which Toll-like signaling pathway was the most significant enrichment pathway.
Asunto(s)
Adyuvantes Inmunológicos/farmacología , Pollos/inmunología , Células Precursoras de Granulocitos/efectos de los fármacos , Inmunoglobulina M/biosíntesis , Oligopéptidos/farmacología , Animales , Línea Celular , Proliferación Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Regulación hacia Abajo/efectos de los fármacos , Análisis de Secuencia por Matrices de Oligonucleótidos , ARN Mensajero , Receptores de Antígenos de Linfocitos B/biosíntesis , Regulación hacia Arriba/efectos de los fármacosRESUMEN
OBJECTIVE: To investigate the possible association of IVS5-5G>A polymorphism, positioned in the upstream region of exon 5 of PINK1 gene with the risk for sporadic late onset Parkinson disease (LOPD) in Chinese. METHODS: Intronic regulatory sequence analysis was performed using the web-based in-silico analysis. The authors performed an association study using a case-control series (comprising 382 LOPD patients and 336 controls, Chinese of Han ancestry). Genotyping was performed by PCR-based denaturing high performance liquid chromatography (DHPLC) combined with sequencing analyses. Allele and genotype frequencies were compared by the Chi-square test. RESULTS: In-silico analysis showed that the intronic IVS5-5G>A polymorphism was located within acceptor site of exon 5 and may be the functional single polymorphism (SNP) in the regulatory region with impact on the splicing of PINK1 gene. Those result yielded statistical significant evidence for the association of PINK1 IVS5-5G>A polymorphism with risk for typical PD in Chinese Han population (OR=1.95, 95%CI: 1.29-2.94, P=0.0012). Homozygote of A allele may have increased risk for LOPD (OR=2.45, 95%CI: 1.27-4.72, P=0.009). CONCLUSION: The authors provide the first evidence that the common genetic variation PINK1 IVS5-5G>A may contribute to the risk of LOPD in Chinese population.