RESUMO
Gardenia jasminoides Ellis is a Chinese herbal medicine with medicinal and economic value, but its mechanism of response to waterlogging stress remains unclear. In this study, the "double pots method" was used to simulate the waterlogging stress of Gardenia jasminoides Ellis to explore its physiological and transcriptomic response mechanism. We found no significant damage to Gardenia jasminoides Ellis membrane lipid during stress. POD played a vital antioxidant role, KEGG enrichment showed that secondary metabolites such as flavonoids might also play an antioxidant role, and PRO played a significant osmotic adjustment. Endogenous hormones regulate the Gardenia jasminoides Ellis's growth and development and play a role in signal transduction. Among them, light waterlogging stress is delayed. At the same time, there were 19631, 23693, and 15045 differentially expressed genes on the 5th, 10d, and 15d of Gardenia jasminoides Ellis under waterlogging stress. These genes were closely associated with the proteasome, endopeptidase, ribosome, MAPK signal transduction, and endogenous hormone signal transduction, plant-pathogen interaction and phenylpropanoid biosynthesis and other physiological and metabolic pathways, which regulate the turnover and transportation of protein, the reinforcement and adhesion of cell walls, the induction of stomatal closure, allergic reactions, defense reactions, leaf movements and others. It also can absorb ultraviolet rays to reduce the generation of oxygen free radicals, change the way of energy utilization and adjust the osmotic pressure of plant cells.
Gardenia jasminoides Ellis é um fitoterápico chinês com valor medicinal e econômico, mas seu mecanismo de resposta ao estresse hídrico permanece obscuro. Neste estudo, o "método de potes duplos" foi usado para simular o estresse hídrico de G. jasminoides Ellis e explorar seu mecanismo de resposta fisiológica e transcriptômica. Não encontramos danos significativos aos lipídios da membrana de G. jasminoides Ellis durante o estresse. POD desempenhou um papel antioxidante vital, o enriquecimento KEGG mostrou que metabólitos secundários, como flavonoides, também podem desempenhar um papel antioxidante e PRO desempenhou um ajuste osmótico significativo. Os hormônios endógenos regulam o crescimento e o desenvolvimento de G. jasminoides Ellis e desempenham um papel na transdução de sinal. Entre eles, o estresse hídrico leve é ââretardado. Ao mesmo tempo, havia 19631, 23693 e 15045 genes diferencialmente expressos de G. jasminoides Ellis nos dias 5, 10 e 15 sob estresse hídrico. Esses genes foram intimamente associados ao proteassoma, endopeptidase, ribossomo, transdução de sinal MAPK e transdução de sinal de hormônio endógeno, interação planta-patógeno e biossíntese de fenilpropanoides e outras vias fisiológicas e metabólicas, que regulam o turnover e transporte de proteínas, o reforço e adesão das paredes celulares, a indução do fechamento estomático, reações alérgicas, reações de defesa, movimentos foliares, entre outros. Também podem absorver os raios ultravioleta para reduzir a geração de radicais livres de oxigênio, alterar a forma de utilização da energia e ajustar a pressão osmótica das células vegetais.
Assuntos
Desidratação , Gardenia/fisiologiaRESUMO
Gardenia jasminoides Ellis is a Chinese herbal medicine with medicinal and economic value, but its mechanism of response to waterlogging stress remains unclear. In this study, the "double pots method" was used to simulate the waterlogging stress of Gardenia jasminoides Ellis to explore its physiological and transcriptomic response mechanism. We found no significant damage to Gardenia jasminoides Ellis membrane lipid during stress. POD played a vital antioxidant role, KEGG enrichment showed that secondary metabolites such as flavonoids might also play an antioxidant role, and PRO played a significant osmotic adjustment. Endogenous hormones regulate the Gardenia jasminoides Ellis's growth and development and play a role in signal transduction. Among them, light waterlogging stress is delayed. At the same time, there were 19631, 23693, and 15045 differentially expressed genes on the 5th, 10d, and 15d of Gardenia jasminoides Ellis under waterlogging stress. These genes were closely associated with the proteasome, endopeptidase, ribosome, MAPK signal transduction, and endogenous hormone signal transduction, plant-pathogen interaction and phenylpropanoid biosynthesis and other physiological and metabolic pathways, which regulate the turnover and transportation of protein, the reinforcement and adhesion of cell walls, the induction of stomatal closure, allergic reactions, defense reactions, leaf movements and others. It also can absorb ultraviolet rays to reduce the generation of oxygen free radicals, change the way of energy utilization and adjust the osmotic pressure of plant cells.
Assuntos
Medicamentos de Ervas Chinesas , Gardenia , Antioxidantes , Endopeptidases , Flavonoides , Frutas , Hormônios , Lipídeos de Membrana , Extratos Vegetais , Folhas de Planta , Complexo de Endopeptidases do Proteassoma , TranscriptomaRESUMO
Carvacrol is a monoterpene that has been linked to neuroprotection in several animal models of neurodegeneration, including ischemia, epilepsy and traumatic neuronal injury. In this study, we investigated the effects of carvacrol (i.p.) upon the neurodegeneration induced by 6-hydroxy-dopamine unilateral intrastriatal injections in mice. We have also used the cylinder test to assess the behavioral effects of carvacrol in that model of Parkinson's disease, and immunoblots to evaluate the levels of caspase-3 and TRPM7, one of major targets of carvacrol. Behavioral testing revealed that carvacrol largely reduced the asymmetrical use of the forelimbs induced by unilateral 6-hydroxy-dopamine. Carvacrol dramatically reduced the loss of tyrosine hydroxylase immunostaining both in the substantia nigra and in the striatum that are typical of the model. Immunoblots for tyrosine hydroxylase confirmed this effect. Caspase-3 levels were very high after toxin injections, but carvacrol appeared to reduce them to control levels. Finally, TRPM7, observed by immunoblots, increased after 6-hydroxy-dopamine, suggesting the involvement of this cation channel in the ensuing neurodegenerative process. The present data suggest that carvacrol promotes a marked neuroprotection in the 6-hydroxy-dopamine model of Parkinson's disease, possibly by its non-specific blocking effect upon TRPM7 channels.
Assuntos
Monoterpenos/farmacologia , Neurônios/efeitos dos fármacos , Neuroproteção/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Doença de Parkinson/tratamento farmacológico , Substância Negra/efeitos dos fármacos , Animais , Cimenos , Modelos Animais de Doenças , Masculino , Camundongos Endogâmicos C57BL , Neurônios/metabolismoRESUMO
Tumor necrosis factor receptor-associated factor 6 (TRAF6) is a crucial adaptor molecule of the interleukin-1 receptor/Toll-like receptor (IL-1/TLR) superfamily, which can trigger downstream signaling cascades involved in innate immunity. The function of TRAF6 has been clarified in mammals but is poorly understood in chicken. In our study, we investigated TRAF6 function in birds, particularly in chicken innate immune responses, by cloning and characterizing chicken TRAF6 (chTRAF6). The full-length coding sequence of chTRAF6 comprised 1638 bp and encoded a 545-amino acid protein, which shares high sequence similarity with TRAF6 of other species and consists of four structurally conserved domains. Quantitative real-time polymerase chain reaction revealed that chTRAF6 was widely expressed in all tested tissues and its expression was induced in chicken embryo fibroblast cells treated with poly(I:C) and poly(dA:dT). Increased expression of chTRAF6 was observed both in vitro and in vivo following infection with Newcastle disease virus in chickens. Taken together, these results suggest that chTRAF6 plays a vital role in host defense against viral infection in chicken.
Assuntos
Proteínas Aviárias/genética , Galinhas/genética , Fator 6 Associado a Receptor de TNF/genética , Animais , Proteínas Aviárias/química , Proteínas Aviárias/metabolismo , Galinhas/imunologia , Galinhas/metabolismo , Resistência à Doença/genética , Resistência à Doença/imunologia , Expressão Gênica , Imunidade Inata/genética , Domínios e Motivos de Interação entre Proteínas , Transdução de Sinais , Fator 6 Associado a Receptor de TNF/química , Fator 6 Associado a Receptor de TNF/metabolismoRESUMO
The aim of this study was to investigate the expression of T-cell immunoglobulin mucin domain molecule-3 (Tim-3) in osteosarcoma tissues, and analyze its effect on cell proliferation and metastasis in an osteosarcoma cell line. Tim-3 mRNA and protein expression in osteosarcoma tissue was detected by reverse transcriptase-polymerase chain reaction and immunohistochemistry, respectively. Additionally, the cell viability, apoptosis rate, and invasive ability of the osteosarcoma cell line MG-63 were tested using the methyl thiazolyl tetrazolium assay, Annexin V-propidium iodide flow cytometry, and a Transwell assay, respectively, following Tim-3 interference using small interfering RNA (siRNA). We also analyzed the expression of Snail, E-cadherin, vimentin, and nuclear factor (NF)-kB in the cells by western blot. We observed that Tim-3 mRNA and protein was significantly overexpressed in osteosarcoma tissues, compared to the adjacent normal tissue (P < 0.01). Moreover, MG-63 cells transfected with the Tim-3 siRNA presented lower cell viability, a greater number of apoptotic cells, and decreased invasive ability (P < 0.01), compared to control cells. Additionally, we observed a decrease in Snail and vimentin expression, an increase in the E-cadherin level, and an increase in NF-kB p65 phosphorylation (P < 0.01) in Tim-3 siRNA-transfected MG-63 cells. Based on these results, we concluded that Tim-3 is highly expressed in osteosarcoma tissue. Moreover, we speculated that interfering in Tim-3 expression could significantly suppress osteosarcoma cell (MG-63) proliferation and metastasis via the NF-kB/Snail signaling pathway and epithelial-mesenchymal transition.
Assuntos
Neoplasias Ósseas/genética , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica , Receptor Celular 2 do Vírus da Hepatite A/genética , Osteossarcoma/genética , Antígenos CD , Apoptose , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/patologia , Caderinas/genética , Caderinas/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Sobrevivência Celular , Colágeno/química , Cultura em Câmaras de Difusão , Combinação de Medicamentos , Receptor Celular 2 do Vírus da Hepatite A/antagonistas & inibidores , Receptor Celular 2 do Vírus da Hepatite A/metabolismo , Humanos , Laminina/química , Metástase Linfática , Invasividade Neoplásica , Osteossarcoma/metabolismo , Osteossarcoma/patologia , Fosforilação , Proteoglicanas/química , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Fatores de Transcrição da Família Snail/genética , Fatores de Transcrição da Família Snail/metabolismo , Fator de Transcrição RelA/genética , Fator de Transcrição RelA/metabolismo , Vimentina/genética , Vimentina/metabolismoRESUMO
With the advent of antibiotic resistance, pathogenic bacteria have become a major threat in cases of neonatal sepsis; however, guidelines for treatment have not yet been standardized. In this study, 15 cases of neonatal Streptococcus agalactiae sepsis from our hospital were retrospectively analyzed. Of these, nine cases showed early-onset and six cases showed late-onset sepsis. Pathogens were characterized by genotyping and antibiotic sensitivity tests on blood cultures. Results demonstrated that in cases with early-onset sepsis, clinical manifestations affected mainly the respiratory tract, while late-onset sepsis was accompanied by intracranial infection. Therefore, we suggest including a cerebrospinal fluid examination when diagnosing neonatal sepsis. Bacterial genotyping indicated the bacteria were mainly type Ib, Ia, and III S. agalactiae. We recommend treatment with penicillin or ampicillin, since bacteria were resistant to clindamycin and tetracycline. In conclusion, our results provide valuable information for the clinical treatment of S. agalactiae sepsis in neonatal infants.
Assuntos
Bacteriemia/diagnóstico , Doenças do Recém-Nascido/diagnóstico , Infecções Estreptocócicas/diagnóstico , Streptococcus agalactiae , Antibacterianos/uso terapêutico , Bacteriemia/tratamento farmacológico , Bacteriemia/microbiologia , Bacteriemia/fisiopatologia , Feminino , Genótipo , Humanos , Recém-Nascido , Doenças do Recém-Nascido/tratamento farmacológico , Doenças do Recém-Nascido/microbiologia , Masculino , Testes de Sensibilidade Microbiana , Prognóstico , Estudos Retrospectivos , Infecções Estreptocócicas/tratamento farmacológico , Infecções Estreptocócicas/microbiologia , Infecções Estreptocócicas/fisiopatologia , Streptococcus agalactiae/genéticaRESUMO
Mycoplasma hyopneumoniae (M. hyopneumoniae) causes porcine enzootic pneumonia (PEP) that significantly affects the pig industry worldwide. Despite the availability of the whole genome sequence, studies on the pathogenesis of this organism have been limited due to the lack of a genetic manipulation system. Therefore, the aim of the current study was to generate a general GFP reporter vector based on a replicating plasmid. Here, we describe the feasibility of GFP reporter expression in M. hyopneumoniae (strain 168L) controlled by the p97 gene promoter of this mycoplasma. An expression plasmid (pMD18-TOgfp) containing the p97 gene promoter, and origin of replication (oriC) of M. hyopneumoniae, tetracycline resistant marker (tetM), and GFP was constructed and used to transform competent M. hyopneumoniae cells. We observed green fluorescence in M. hyopneumoniae transformants under fluorescence microscopy, which indicates that there was expression of the GFP reporter that was driven by the p97 gene promoter. Additionally, an electroporation method for M. hyopneumoniae with an efficiency of approximately 1 x 10(-6) transformants/µg plasmid DNA was optimized and is described herein. In conclusion, our data demonstrate the susceptibility of M. hyopneumoniae to genetic manipulation whereby foreign genes are expressed. This work may encourage the development of genetic tools to manipulate the genome of M. hyopneumoniae for functional genomic analyses.
Assuntos
Vetores Genéticos/genética , Proteínas de Fluorescência Verde/genética , Mycoplasma/genética , Plasmídeos/genética , Proteínas de Fluorescência Verde/metabolismo , Mycoplasma/metabolismo , TransgenesRESUMO
This study aimed to evaluate the effects of exercise training on triglyceride deposition and the expression of musclin and glucose transporter 4 (GLUT4) in a rat model of insulin resistance. Thirty male Sprague-Dawley rats (8 weeks old, weight 160±10 g) were fed a high-fat diet (40% calories from fat) and randomly divided into high-fat control group and swimming intervention group. Rats fed with standard food served as normal control. We found that 8-week swimming intervention significantly decreased body weight (from 516.23±46.27 to 455.43±32.55 g) and visceral fat content (from 39.36±2.50 to 33.02±2.24 g) but increased insulin sensitivity index of the rats fed with a high-fat diet. Moreover, swimming intervention improved serum levels of TG (from 1.40±0.83 to 0.58±0.26 mmol/L) and free fatty acids (from 837.80±164.25 to 556.38±144.77 µEq/L) as well as muscle triglycerides deposition (from 0.55±0.06 to 0.45±0.02 mmol/g) in rats fed a high-fat diet. Compared with rats fed a standard food, musclin expression was significantly elevated, while GLUT4 expression was decreased in the muscles of rats fed a high-fat diet. In sharp contrast, swimming intervention significantly reduced the expression of musclin and increased the expression of GLUT4 in the muscles of rats fed a high-fat diet. In conclusion, increased musclin expression may be associated with insulin resistance in skeletal muscle, and exercise training improves lipid metabolism and insulin sensitivity probably by upregulating GLUT4 and downregulating musclin.
Assuntos
Gorduras na Dieta/administração & dosagem , Transportador de Glucose Tipo 4/metabolismo , Resistência à Insulina/genética , Metabolismo dos Lipídeos/genética , Proteínas Musculares/metabolismo , Animais , Gorduras na Dieta/metabolismo , Regulação da Expressão Gênica , Transportador de Glucose Tipo 4/genética , Resistência à Insulina/fisiologia , Masculino , Proteínas Musculares/genética , Condicionamento Físico Animal , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Tempo , Fatores de TranscriçãoRESUMO
This study aimed to evaluate the effects of exercise training on triglyceride deposition and the expression of musclin and glucose transporter 4 (GLUT4) in a rat model of insulin resistance. Thirty male Sprague-Dawley rats (8 weeks old, weight 160±10 g) were fed a high-fat diet (40% calories from fat) and randomly divided into high-fat control group and swimming intervention group. Rats fed with standard food served as normal control. We found that 8-week swimming intervention significantly decreased body weight (from 516.23±46.27 to 455.43±32.55 g) and visceral fat content (from 39.36±2.50 to 33.02±2.24 g) but increased insulin sensitivity index of the rats fed with a high-fat diet. Moreover, swimming intervention improved serum levels of TG (from 1.40±0.83 to 0.58±0.26 mmol/L) and free fatty acids (from 837.80±164.25 to 556.38±144.77 μEq/L) as well as muscle triglycerides deposition (from 0.55±0.06 to 0.45±0.02 mmol/g) in rats fed a high-fat diet. Compared with rats fed a standard food, musclin expression was significantly elevated, while GLUT4 expression was decreased in the muscles of rats fed a high-fat diet. In sharp contrast, swimming intervention significantly reduced the expression of musclin and increased the expression of GLUT4 in the muscles of rats fed a high-fat diet. In conclusion, increased musclin expression may be associated with insulin resistance in skeletal muscle, and exercise training improves lipid metabolism and insulin sensitivity probably by upregulating GLUT4 and downregulating musclin.
Assuntos
Animais , Masculino , Ratos , Resistência à Insulina/genética , Gorduras na Dieta/administração & dosagem , Transportador de Glucose Tipo 4/metabolismo , Metabolismo dos Lipídeos/genética , Proteínas Musculares/metabolismo , Condicionamento Físico Animal , Fatores de Tempo , Fatores de Transcrição , Resistência à Insulina/fisiologia , Gorduras na Dieta/metabolismo , Distribuição Aleatória , Regulação da Expressão Gênica , Ratos Sprague-Dawley , Transportador de Glucose Tipo 4/genética , Reação em Cadeia da Polimerase em Tempo Real , Proteínas Musculares/genéticaRESUMO
The aim of this study was to investigate the expression of CD44 and its clinical significance in children suffering from hepatoblastoma (HB). CD44 expression was detected with immunohistochemistry staining in 30 samples from hepatoblastoma children and 10 normal liver tissue samples from normal children. The data obtained was statistically analyzed using the chi-square test, using the SPSS (v.11.0) software. The rate of CD44 expression was significantly higher (66.7%) in hepatoblastoma tissues than in normal liver tissues (χ(2) = 4.848, P < 0.05). The rate of CD44 expression was significantly higher in children with stage III or IV hepatoblastoma (83.3%) than that in children with stage I and II hepatoblastoma (χ(2) ï¼ 5.625, P < 0.05) (41.7%). Therefore, CD44 expression might play an important role in the pathogenesis, progression, and prognosis of HB in children.
Assuntos
Hepatoblastoma/metabolismo , Hepatoblastoma/patologia , Receptores de Hialuronatos/metabolismo , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Criança , Progressão da Doença , Feminino , Expressão Gênica , Hepatoblastoma/genética , Humanos , Receptores de Hialuronatos/genética , Imuno-Histoquímica , Neoplasias Hepáticas/genética , Masculino , Estadiamento de Neoplasias , PrognósticoRESUMO
Lipid-associated membrane proteins (LAMPs) are important in the pathogenicity of the Mycoplasma genus of bacteria. We investigated whether Mycoplasma hyopneumoniae LAMPs have pathogenic potential by inducing apoptosis in a St. Jude porcine lung epithelial cell line (SJPL). LAMPs from a pathogenic strain of M. hyopneumoniae (strain 232) were used in the research. Our investigation made use of diamidino-phenylindole (DAPI) and acridine orange/ethidium bromide (AO/EB) staining, terminal dexynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) analysis, and Annexin-V-propidium iodide staining. After LAMP treatment for 24 h, typical changes were induced, chromosomes were concentrated, apoptotic bodies were observed, the 3'-OH groups of cleaved genomes were exposed, and the percentage of apoptotic cells reached 36.5 ± 11.66%. Caspase 3 and caspase 8 were activated and cytochrome c (cyt c) was released from the mitochondria into the cytoplasm; poly ADP ribose polymerase (PARP) was digested into two fragments; p38 mitogen-activated protein kinase (MAPK) was phosphorylated; and the expression of pro-apoptosis protein Bax increased while the anti-apoptosis protein Bcl-2 decreased. LAMPs also stimulated SJPL cells to produce nitric oxide (NO) and superoxide. This study demonstrated that LAMPs from M. hyopneumoniae can induce apoptosis in SJPL cells through the activation of caspase 3, caspase 8, cyt c, Bax, and p38 MAPK, thereby contributing to our understanding of the pathogenesis of M. hyopneumoniae, which should improve the treatment of M. hyopneumoniae infections.
Assuntos
Apoptose , Proteínas de Bactérias/farmacologia , Caspase 3/metabolismo , Células Epiteliais/citologia , Pulmão/citologia , Sistema de Sinalização das MAP Quinases , Mycoplasma hyopneumoniae/metabolismo , Animais , Caspase 8/metabolismo , Morte Celular/efeitos dos fármacos , Linhagem Celular , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Citocromos c/metabolismo , Ativação Enzimática/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/enzimologia , Marcação In Situ das Extremidades Cortadas , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Modelos Biológicos , Óxido Nítrico/metabolismo , Poli(ADP-Ribose) Polimerases/metabolismo , Superóxidos/metabolismo , Sus scrofa , Proteína X Associada a bcl-2/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
We investigated the effect of p38MAPK/AP-1 (activator protein-1) signaling on the apoptosis of osteoblasts induced by high glucose. A lentivirus vector of small hairpin RNA (shRNA) targeting p38MAPK was constructed in vitro. Osteoblasts MC3T3-E1 cultured in vitro were treated with vehicle, high glucose, p38MAPK-shRNA transfection, p38MAPK inhibitor, and unrelated shRNA transfection. Apoptosis, protein levels of p38MAPK, and activities of AP-1 in MC3T3-E1 osteoblasts were measured using TUNEL and flow cytometry, Western blot analysis, and an electrophoretic mobility shift assay. Compared with the vehicle group, high glucose induced apoptosis of MC3T3-E1 osteoblasts and activated p38MAPK and AP-1. p38MAPK-shRNA transfection blocked the effect of high glucose stimulation, and the p38MAPK inhibitor showed similar effects as those observed in p38MAPK transfection. Unrelated shRNA had no effect on these changes in MC3T3-E1 osteoblasts induced by high glucose. Therefore, our results suggest that p38MAPK-shRNA reduce apoptosis of MC3T3-E1 osteoblasts induced by high glucose by inhibiting the p38MAPK-AP-1 signaling pathway.
Assuntos
Apoptose/efeitos dos fármacos , Glucose/farmacologia , Osteoblastos/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Fator de Transcrição AP-1/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Western Blotting , Linhagem Celular , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Citometria de Fluxo , Expressão Gênica/efeitos dos fármacos , Imidazóis/farmacologia , Marcação In Situ das Extremidades Cortadas , Camundongos , Osteoblastos/citologia , Osteoblastos/metabolismo , Ligação Proteica/efeitos dos fármacos , Piridinas/farmacologia , Interferência de RNA , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases p38 Ativadas por Mitógeno/genéticaRESUMO
Tumor necrosis factor receptor-associated factor 3 (TRAF3) is a crucial regulator that suppresses c-Jun N-terminal kinase and non-canonical nuclear factor-kB signaling, but facilitates type I interferon production. To determine TRAF3 function in innate immune responses among birds, particularly chicken, we cloned and characterized the chicken TRAF3 gene (chTRAF3) and detected its tissue expression profile in chicken. We also detected the differential expression of chTRAF3 and its downstream gene interferon-ß (IFN-ß) upon different stimuli in primary chicken embryo fibroblast cells. Two chTRAF3 gene products, chTRAF3-1 and chTRAF3-2, can be produced by alternative splicing. The full-length coding sequence of chTRAF3 (chTRAF3-1) was 1704 base pairs and encoded a protein of 567 amino acids with high identity to TRAF3 homologs from mammals and other birds. The deduced amino acid sequence showed typical characteristics of TRAFs, with a RING finger domain, 2 zf-TRAF motifs, and a MATH domain. Quantitative real-time polymerase chain reaction analysis revealed broad expression of chTRAF3 in all detected tissues, with abundant expression in the spleen, thymus, lung, and small intestine. Expression of chTRAF3 was significantly upregulated in a time- and concentration-dependent manner in chicken embryo fibroblast cells challenged with poly I:C or poly dA-dT. Furthermore, chTRAF3 and IFN-ß mRNA expression from chicken embryo fibroblast cells challenged with Newcastle disease virus F48E9 suffered intense suppression compared with Newcastle disease virus Mukteswar infection. Our results indicate that chTRAF3 plays important roles in defending against both RNA and DNA virus infection.
Assuntos
Proteínas Aviárias/genética , Galinhas/genética , Regulação da Expressão Gênica , Fator 3 Associado a Receptor de TNF/genética , Processamento Alternativo , Motivos de Aminoácidos , Sequência de Aminoácidos , Animais , Proteínas Aviárias/metabolismo , Sequência de Bases , Embrião de Galinha , Galinhas/imunologia , Galinhas/metabolismo , Clonagem Molecular , Evolução Molecular , Imunidade Inata , Dados de Sequência Molecular , Doença de Newcastle/imunologia , Especificidade de Órgãos , Alinhamento de Sequência , Fator 3 Associado a Receptor de TNF/metabolismoRESUMO
The aim of this study was to establish a method for sensitive and rapid diagnosis of Mycoplasma hyopneumoniae in clinical specimens. To this effect, we employed three sets of primers specifically designed for amplification of nucleic acids under isothermal conditions. After optimization of reaction conditions, M. hyopneumoniae could be successfully detected at 63°C in 45 min through use of the loop-mediated isothermal amplification (LAMP) assay. A positive reaction was identified visually as white precipitate and confirmed by gel electrophoresis. The detection limit for this assay was 10 copies/µL, as observed by electrophoretic analysis. The accuracy of the LAMP reaction was confirmed by restriction endonuclease digestion as well as by direct sequencing of the amplified product. This method can specifically detect M. hyopneumoniae; other species with high homology and other bacterial and virus strains gave negative results. To test the utility of this procedure, the LAMP assay was applied to 40 clinical samples collected from swine lung tissues experimentally challenged with M. hyopneumoniae isolates, and compared to the results from a real-time polymerase chain reaction (PCR) assay. A concordance of 100% was observed between the two assays. In conclusion, the results from our study demonstrated that the LAMP assay provided a rapid reaction and was inexpensive to perform, with no need of complex instruments or systems such as Geneamp PCR. The LAMP assay may therefore be applied in routine diagnosis in the clinical laboratory and for in-field detection of M. hyopneumoniae infection.
Assuntos
Genes Bacterianos , Mycoplasma hyopneumoniae/isolamento & purificação , Mycoplasma hyopneumoniae/genética , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Gonadotropin-inhibitory hormone (GnIH) gene is an important gene in reproduction. In this study, we screened single-nucleotide polymorphisms (SNPs) in the chicken GnIH gene among 204 individuals in Erlang mountainous chickens. We then analyzed the associations between polymorphisms of the GnIH gene and 5 egg production traits in chickens. Five SNPs (T3305C, T3310C, G3403C, G3411A, and T3591C) were detected. Associations between polymorphic loci and age at first egg, body weight at first egg, weight at first egg, egg weight in 300 days, and egg production in 300 days were analyzed using analysis of covariance. The results showed that SNP1, SNP3, and SNP4 had large effects on age at first egg, while SNP5 had a large effect on body weight at first egg; of the effect of the TT genotype was significantly higher than that of CT (P < 0.01). Further analysis show that the highest frequency (0.2353) haplotype H1H1 was associated with the latest age at first egg. The H4H5 haplotype had a positive effect on egg production in 300 days and a negative effect on weight at first egg. We observed no association between the H3H3 haplotype and body weight at first egg.
Assuntos
Proteínas Aviárias/genética , Galinhas/genética , Ovos , Estudos de Associação Genética , Polimorfismo de Nucleotídeo Único/genética , Alelos , Animais , Frequência do Gene , Genótipo , Haplótipos/genética , Desequilíbrio de Ligação/genética , Análise de Sequência de DNARESUMO
Dysregulation of microRNAs (miRs) is associated with cancer development and progression and aberrant expression of miR-874 have been found in some types of cancer. However, the expression and function of miR-874 in osteosarcoma remain unclear. The aim of this study was to explore the effects of miR-874 in osteosarcoma tumorigenesis and development. The expression level of miR-874 was quantified by real-time reverse transcription-polymerase chain reaction (RT-PCR) in human osteosarcoma cell lines and tissues. Using a miR-874 mimic, cell proliferation and migration assays were performed in an osteosarcoma cell line and tumorigenicity was observed in vivo in order to determine the effects of miR-874 in osteosarcoma cell lines and tissues. MiR-874 was significantly downregulated in osteosarcoma cell lines and clinical specimens. Decreased miR-874 expression was significantly associated with large tumor size, distant metastasis, and advanced clinical stage, and was an independent predictor of poor survival. Overexpression of miR-874 inhibited cell proliferation, invasion and migration in vitro, promoted cell apoptosis in vitro, and suppressed tumorigenicity in vivo. These findings indicate that miR-874 may act as a tumor suppressor in osteosarcoma and could serve as a novel therapeutic agent for miR-based therapy.
RESUMO
This study aimed to determine the protective effect and mechanism of neutrophil gelatinase-associated lipocalin (NGAL) in rat kidney on ischemia/reperfusion injury (I/R). The rat I/R model was set up by cutting one kidney and clamping the contralateral renal pedicle for 45 min. Male SD rats were randomly divided into sham-operation, I/R and NGAL groups. Hematoxylin-eosin staining was performed to observe the renal pathological changes in the 3 groups; serum creatinine (Scr) and blood urea nitrogen (BUN) determined in blood samples taken from the inferior vena cava 24 h after the reperfusion were measured; TUNEL was used to observe the apoptosis of renal tubular epithelial cells; immunohistochemistry was performed to evaluate the expressions of Bax and activated caspase-3; Western blotting was used to determine the expression changes in apoptotic proteins Fas and Bcl-2. Compared with the I/R group, Scr and BUN of the NGAL group were 63.400 ± 11.908 vs 121.857 ± 17.151 µM and 14.840 ± 2.868 vs 28.557 ± 6.434 mM, respectively. The number of apoptotic tubular epithelial cells was reduced (7.800 ± 1.924 vs 15.400 ± 3.049); the expression of renal tissue Fas mRNA of the NGAL group was decreased (2.34 ± 0.51 vs 6.84 ± 2.34); the expression of the Bax protein was lower (7.440 ± 1.640 vs 15.456 ± 1.955%); the expression of the CC3 protein was decreased (3.171 ± 0.321 vs 7.291 ± 1.059%), while the expression of the Bcl-2 protein increased (6.91 ± 1.64 vs 5.30 ± 1.48), P < 0.05. NGAL had a protective effect towards the renal tubular epithelial cells in I/R, and the effect might have been associated with the reduction in apoptosis and the altered expression of apoptotic proteins, which would thereby reduce tissue damage and protect the kidney.
Assuntos
Proteínas de Fase Aguda/metabolismo , Rim/metabolismo , Lipocalinas/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Traumatismo por Reperfusão/metabolismo , Proteínas de Fase Aguda/genética , Animais , Apoptose , Células Epiteliais/metabolismo , Rim/irrigação sanguínea , Rim/patologia , Lipocalina-2 , Lipocalinas/genética , Masculino , Proteínas Proto-Oncogênicas/genética , Ratos , Ratos Sprague-DawleyRESUMO
The Jiangquhai porcine lean strain (JQHPL) is a new pork meat-type strain that has been developed in recent years from the parent lines Duroc, Fengjing, and Jiangquhai pigs (DurocxFengjing pigxJiangquhai pig). Enzootic pneumonia (EP) in pigs induced by Mycoplasma hyopneumoniae (M. hyopneumoniae) is a chronic respiratory disease of pigs, generating high economic losses in the swine industry. Here, we investigated the degree of resistance to M. hyopneumoniae for the Jiangquhai porcine lean strain and the Duroc x Landrace x Yorkshire (DLY) pigs, which are Western commercial pigs that have been introduced in China. A total of 209 DLY piglets and 221 JQHPL piglets from 19 Landrace x Yorkshire and 22 JQHPL M. hyopneumoniae positive gestating sows with different expected dates of confinement were selected and raised in the same M. hyopneumoniae positive farrowing barn. When the oldest suckling piglets were 37 days old, nasal swabs were collected from all the piglets (ranging from 4 to 37 days old) to detect the M. hyopneumoniae pathogen using n-PCR and M. hyopneumoniae specific SIgA using ELISA. Positive M. hyopneumoniae infection rates in both the strains increased with age; however, positive rates for JQHPL were lower compared to DLY at 14 to 35 days old. The level of the specific SIgA rose rapidly in JQHPL respiratory tracts, particularly in piglets 21 to 35 days in age compared to DLY piglets of the same age; however, the level of the specific SIgA in DLY also marginally increased. In conclusion, JQHPL pigs exhibits higher resistance to M. hyopneumoniae compared to DLY. It is possible that this characteristic is caused by the faster and stronger mucosal immunity phenotype of the JQHPL strain.
Assuntos
Anticorpos Antibacterianos/biossíntese , Imunidade nas Mucosas , Imunoglobulina A/biossíntese , Carne , Mycoplasma hyopneumoniae/imunologia , Pneumonia Suína Micoplasmática/imunologia , Fatores Etários , Animais , Animais Lactentes , Cruzamento , Feminino , Mucosa Nasal/imunologia , Mucosa Nasal/microbiologia , Pneumonia Suína Micoplasmática/microbiologia , Gravidez , SuínosRESUMO
There are conflicting reports associating the TGF-ß1 gene -C509T polymorphism with susceptibility to IgA nephropathy. We investigated this association through a meta-analysis. Case-control studies were searched up to January 2012; the genotype frequencies in the control group were found to be consistent with Hardy-Weinberg equilibrium. Publication bias was tested by funnel plot and the Egger regression test. Eight studies, comprising 1364 cases and 1483 controls, were included. Significant heterogeneity was observed (χ² = 18.29, P = 0.01). Under the random-effects model, the overall odds ratio (OR) was 1.01 [95% confidence interval (95%CI) = 0.74-1.38; P = 0.94]. In the subgroup analysis based on ethnicities, no significant effect was observed in the Asian descent groups (five comparisons, OR = 0.78; 95%CI = 0.53-1.15; moderate heterogeneity between studies). However, an association was observed in the European descent groups (OR = 1.5; 95%CI = 1.15-1.96; P = 0.003; no significant heterogeneity between studies). There was no evidence of publication bias according to funnel plot and the Egger regression test (a = -2.16, P = 0.23). There was heterogeneity between studies and no clear evidence of an association between the TGF-ß1 gene -C509T polymorphism and susceptibility to IgA nephropathy in the worldwide population. Subgroup analysis suggests that the TGF-ß1 gene -C509T polymorphism would not be a risk factor for IgA nephropathy in Asians but might play a role in Europeans. More studies are required for definitive conclusions.