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1.
Transbound Emerg Dis ; 2021 Feb 09.
Artigo em Inglês | MEDLINE | ID: mdl-33559368

RESUMO

The variety and widespread of coronavirus in natural reservoir animals is likely to cause epidemics via interspecific transmission, which has attracted much attention due to frequent coronavirus epidemics in recent decades. Birds are natural reservoir of various viruses, but the existence of coronaviruses in wild birds in central China has been barely studied. Some bird coronaviruses belongs to the genus of Deltacoronavirus. To explore the diversity of bird deltacoronaviruses in central China, we tested fecal samples from 415 wild birds in Hunan Province, China. By RT-PCR detection, we identified eight samples positive for deltacoronaviruses which were all from common magpies, and in four of them, we successfully amplified complete deltacoronavirus genomes distinct from currently known deltacoronavirus, indicating four novel deltacoronavirus stains (HNU1-1, HNU1-2, HNU2 and HNU3) . Comparative analysis on the four genomic sequences showed that these novel magpie deltacoronaviruses shared three different S genes among which the S genes of HNU1-1 and HNU1-2 showed 93.8% amino acid (aa) identity to that of thrush coronavirus HKU12, HNU2 S showed 71.9% aa identity to that of White-eye coronavirus HKU16, and HKU3 S showed 72.4% aa identity to that of sparrow coronavirus HKU17. Recombination analysis showed that frequent recombination events of the S genes occurred among these deltacoronavirus strains. Two novel putative cleavage sites separating the nonstructural proteins in the HNU coronaviruses were found. Bayesian phylogeographic analysis showed that the south coast of China might be a potential origin of bird deltacoronaviruses existing in inland China. In summary, these results suggest that common magpie in China carries diverse deltacoronaviruses with novel genomic features, indicating an important source of environmental coronaviruses closed to human communities, which may provide key information for prevention and control of future coronavirus epidemics.

2.
Huan Jing Ke Xue ; 41(12): 5381-5388, 2020 Dec 08.
Artigo em Chinês | MEDLINE | ID: mdl-33374054

RESUMO

Based on the seasonal changes in the nitrogen and phosphorus concentrations in Hanfeng Lake from March 2017 to February 2018, the nutrient limitation status was evaluated by the stoichiometric molar ratio of nitrogen and phosphorus. The results showed that the average concentrations of TN, DN, and NO3--N were 1.60, 1.25, and 0.91 mg·L-1 in Hanfeng Lake, respectively. The seasonal changes of those indicators were similar, showing the highest concentration in winter and lowest in summer. NO3--N accounted for TN significantly in the water body, and the concentrations of NH4+-N and NO2--N remained at low levels and changed steadily. The average concentrations of TP, DP, and PO43--P were 0.13, 0.09, and 0.06 mg·L-1, respectively. The changes in the concentrations of TP and DP were similar, showing a trend of increasing in spring and summer, and then decreasing in autumn and winter, while the PO43--P concentration showed the trend of fluctuated decrease. TN/TP varied from 11.07 to 56.02, with an average value of 29.23. TN/TP changed seasonally, with the highest value occurring in winter and the lowest value in summer. The conditions of the water body were conducive to growth and reproduction of algae for most of the time during sampling months. The water body was occasionally nitrogen limited and rarely phosphorus limited. The seasonal variation in TN/TP ratio was affected by several factors such as rainfall runoff, fertilizer use, sewage discharge, and aquatic biological activities. Further, protection strategies were proposed for the improvement of the water body in terms of present water quality characteristics in Hanfeng Lake.

4.
Virol Sin ; 2020 Mar 31.
Artigo em Inglês | MEDLINE | ID: mdl-32236817

RESUMO

Porcine adenoviruses (PAdVs) are classified into three species, PAdV-A, PAdV-B, and PAdV-C. The genomes of PAdV-A and PAdV-C have been well characterized. However, the genome of PAdV-B has never been completely sequenced, and the epidemiology of PAdV-B remains unclear. In our study, we have identified a novel strain of PAdV-B, named PAdV-B-HNU1, in porcine samples collected in China by viral metagenomic assay and general PCR. The genome of PAdV-B-HNU1 is 31,743 bp in length and highly similar to that of California sea lion adenovirus 1 (C. sea lion AdV-1), which contains typical mastadenoviral structures and some unique regions at the carboxy-terminal end. Especially, PAdV-B-HNU1 harbors a dUTPase coding region not clustering with other mastadenoviruses except for C. sea lion AdV-1 and a fiber coding region homologous with galectin 4 and 9 of animals. However, the variance of GC contents between PAdV-B-HNU1 (55%) and C. sea lion AdV-1 (36%) indicates their differential evolutionary paths. Further epidemiologic study revealed a high positive rate (51.7%) of PAdV-B-HNU1 in porcine lymph samples, but low positive rates of 10.2% and 16.1% in oral swabs and rectal swabs, respectively. In conclusion, this study characterized a novel representative genome of a lymphotropic PAdV-B with unique evolutionary origin, which contributes to the taxonomical and pathogenic studies of PAdVs.

5.
Arch Virol ; 164(12): 3151-3155, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31616994

RESUMO

Astroviruses (AstVs) have a very wide range of hosts and are associated with enteric and extra-enteric disease in mammals and birds. Cross-species transmission of AstVs has been observed frequently. In the present study, the genome of a novel astrovirus from Amur tigers (Panthera tigris) from a zoo in China was characterized and was found to have the typical genomic features of other mammal AstVs. It showed the highest nucleotide sequence similarity (46.1-87.3% identity) to AstVs from cats, indicating a close phylogenetic relationship and possible cross-species transmission between them. To our knowledge, this is the first identification and characterization of AstV from tigers, and this virus is the third astrovirus identified in hosts of the family Felidae. The results of this study will be helpful for understanding the origin, genetic diversity, and cross-species transmission of AstV.


Assuntos
Animais de Zoológico/virologia , Infecções por Astroviridae/veterinária , Astroviridae/isolamento & purificação , Tigres/virologia , Animais , Astroviridae/classificação , Astroviridae/genética , Infecções por Astroviridae/virologia , Gatos , China , Fezes/virologia , Filogenia , Análise de Sequência de DNA
6.
J Mol Cell Biol ; 11(2): 118-132, 2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-29771336

RESUMO

Fat mass and obesity-associated (FTO) protein is a ferrous ion (Fe2+)/2-oxoglutarate (2-OG)-dependent demethylase preferentially catalyzing m6A sites in RNA. The FTO gene is highly expressed in the hypothalamus with fluctuation in response to various nutritional conditions, which is believed to be involved in the control of whole body metabolism. However, the underlying mechanism in response to different nutritional cues remains poorly understood. Here we show that ketogenic diet-derived ketone body ß-hydroxybutyrate (BHB) transiently increases FTO expression in both mouse hypothalamus and cultured cells. Interestingly, the FTO protein represses Fto promoter activity, which can be offset by BHB. We then demonstrate that FTO binds to its own gene promoter, and Fe2+, but not 2-OG, impedes this binding and increases FTO expression. The BHB-induced occupancy of the promoter by FTO influences the assembly of the basal transcriptional machinery. Importantly, a loss-of-function FTO mutant (I367F), which induces a lean phenotype in FTOI367F mice, exhibits augmented binding and elevated potency to repress the promoter. Furthermore, FTO fails to bind to its own promoter that promotes FTO expression in the hypothalamus of high-fat diet-induced obese and 48-h fasting mice, suggesting a disruption of the stable expression of this gene. Taken together, this study uncovers a new function of FTO as a Fe2+-sensitive transcriptional repressor dictating its own gene switch to form an auto-regulatory loop that may link with the hypothalamic control of body weight.


Assuntos
Dioxigenase FTO Dependente de alfa-Cetoglutarato , Peso Corporal/genética , Dioxigenase FTO Dependente de alfa-Cetoglutarato/genética , Dioxigenase FTO Dependente de alfa-Cetoglutarato/metabolismo , Animais , Linhagem Celular , Regulação da Expressão Gênica , Hipotálamo/metabolismo , Camundongos , Células NIH 3T3 , Obesidade , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
7.
J Proteomics ; 185: 39-50, 2018 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-29953961

RESUMO

Physical exercise has been reported to increase neurotrophin in brain tissues as hippocampus as well as increased neurotrophic level peripherally in blood plasma and might have an effect on/or affect molecular processes of energy metabolism (and homeostasis). In this study, using quantitative proteomic analysis, we obtained a plasma protein profile from the rat with long-term moderate exercise. A total of 752 proteins were identified in the plasma. Among them, 54 proteins were significant up-regulated and 47 proteins were down-regulated in the plasma of exercise group compared with the control group. Bioinformatic analyses showed that these altered proteins are widely involved in multiple biological processes, molecular functions and cellular components, which connect with 11 signaling pathways. Interestingly, 5 up-regulated proteins Rap1b, PTPN11, ARHGDIA, Cdc42 and YWHAE, confirmed by Western blots, are involved in the neurotrophin signaling pathway which shows the lowest P value among the identified pathways. Further analyses showed that the 5 neurotrophin-signaling-pathway-related proteins participate in two important protein-protein interaction networks associated to cell survival and apoptosis, axonal development, synapse formation and plasticity. This study provides an exercise-induced plasma protein profile, suggesting that long-term exercise enhances the proteins involved in neurotrophin signaling pathway which may contribute to health benefit. SIGNIFICANCE: Physical activity contributes to myriad benefits on body health across the lifespan. The changes in plasma proteins after chronic moderate exercise may be used as biomarkers for health and may also play important roles in increase of cardiovascular fitness, enhancement of immune competence, prevention of obesity, decrease of risk for neurological disorders, cancer, stroke, diabetes and other metabolic disorders. Using a TMT-based proteomic method, this study identified 101 altered proteins in the plasma of rats after long-term moderate treadmill running, which may provide novel biomarkers for further investigation of the underlying mechanism of physical exercise. We confirmed that exercise enhances 5 proteins of the neurotrophin signaling pathway that may contribute to health benefits.


Assuntos
Proteínas Sanguíneas/análise , Fatores de Crescimento Neural/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Condicionamento Físico Animal/métodos , Proteômica/métodos , Animais , Proteínas Sanguíneas/química , Proteínas Sanguíneas/metabolismo , Estudos de Casos e Controles , Cromatografia Líquida , Masculino , Neurogênese/fisiologia , Plasma/química , Plasma/metabolismo , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/fisiologia , Espectrometria de Massas em Tandem , Fatores de Tempo
8.
Biochim Biophys Acta Mol Basis Dis ; 1863(6): 1492-1499, 2017 06.
Artigo em Inglês | MEDLINE | ID: mdl-28433711

RESUMO

Voltage-gated sodium channel α-subunit type I (NaV1.1, encoded by SCN1A gene) plays a critical role in the excitability of brain. Downregulation of SCN1A expression is associated with epilepsy, a common neurological disorder characterized by recurrent seizures. Here we reveal a novel role of malate dehydrogenase 2 (MDH2) in the posttranscriptional regulation of SCN1A expression under seizure condition. We identified that MDH2 was an RNA binding protein that could bind two of the four conserved regions in the 3' UTRs of SCN1A. We further showed that knockdown of MDH2 or inactivation of MDH2 activity in HEK-293 cells increased the reporter gene expression through the 3' UTR of SCN1A, and MDH2 overexpression decreased gene expression by affecting mRNA stability. In the hippocampus of seizure mice, the upregulation of MDH2 expression contributed to the decrease of the NaV1.1 levels at posttranscriptional level. In addition, we showed that the H2O2 levels increased in the hippocampus of the seizure mice, and H2O2 could promote the binding of MDH2 to the binding sites of Scn1a gene, whereas ß-mercaptoethanol decreased the binding capability, indicating an important effect of the seizure-induced oxidation on the MDH2-mediated downregulation of Scn1a expression. Taken together, these data suggest that MDH2, functioning as an RNA-binding protein, is involved in the posttranscriptional downregulation of SCN1A expression under seizure condition.


Assuntos
Regiões 3' não Traduzidas , Regulação para Baixo , Malato Desidrogenase/metabolismo , Canal de Sódio Disparado por Voltagem NAV1.1/biossíntese , Proteínas de Ligação a RNA/metabolismo , Convulsões/metabolismo , Animais , Células HEK293 , Hipocampo/metabolismo , Hipocampo/patologia , Humanos , Malato Desidrogenase/genética , Camundongos , Canal de Sódio Disparado por Voltagem NAV1.1/genética , Proteínas de Ligação a RNA/genética , Convulsões/genética , Convulsões/patologia
9.
Neuropharmacology ; 113(Pt A): 480-489, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-27816501

RESUMO

Abnormal expressions of sodium channel SCN1A and SCN3A genes alter neural excitability that are believed to contribute to the pathogenesis of epilepsy, a long-term risk of recurrent seizures. Ketogenic diet (KD), a high-fat and low-carbohydrate treatment for difficult-to-control (refractory) epilepsy in children, has been suggested to reverse gene expression patterns. Here, we reveal a novel role of GAPDH on the posttranscriptional regulation of mouse Scn1a and Scn3a expressions under seizure and KD conditions. We show that GAPDH binds to a conserved region in the 3' UTRs of human and mouse SCN1A and SCN3A genes, which decreases and increases genes' expressions by affecting mRNA stability through SCN1A 3' UTR and SCN3A 3' UTR, respectively. In seizure mice, the upregulation and phosphorylation of GAPDH enhance its binding to the 3' UTR, which lead to downregulation of Scn1a and upregulation of Scn3a. Furthermore, administration of KD generates ß-hydroxybutyric acid which rescues the abnormal expressions of Scn1a and Scn3a by weakening the GAPDH's binding to the element. Taken together, these data suggest that GAPDH-mediated expression regulation of sodium channel genes may be associated with epilepsy and the anticonvulsant action of KD.


Assuntos
Dieta Cetogênica , Gliceraldeído-3-Fosfato Desidrogenases/fisiologia , Canal de Sódio Disparado por Voltagem NAV1.1/genética , Canal de Sódio Disparado por Voltagem NAV1.3/genética , Convulsões/dietoterapia , Convulsões/genética , Canais de Sódio/genética , Animais , Linhagem Celular Tumoral , Dieta Cetogênica/métodos , Células HEK293 , Humanos , Masculino , Camundongos , Canal de Sódio Disparado por Voltagem NAV1.1/biossíntese , Canal de Sódio Disparado por Voltagem NAV1.3/biossíntese , Ligação Proteica/fisiologia , Processamento Pós-Transcricional do RNA/fisiologia , Convulsões/metabolismo , Canais de Sódio/biossíntese
10.
Molecules ; 21(9)2016 Aug 27.
Artigo em Inglês | MEDLINE | ID: mdl-27618892

RESUMO

OBJECTIVE: To screen for influenza virus neuraminidase inhibition and to provide a reference for the clinical treatment of influenza using traditional Chinese medicines (TCM). In this study, 421 crude extracts (solubilized with petroleum ether, ethanol, ethyl acetate, and aqueous solvents) were obtained from 113 TCM. The medicine extracts were then reacted with oseltamivir, using 2'-(4-methylumbelliferyl)-α-D-N-acetylneuraminic acid (MUNANA) as the substrate, to determine influenza virus neuraminidase activity using a standard fluorimetric assay. It was found that Chinese medicine extracts from Pyrola calliantha, Cynanchum wilfordii, Balanophora involucrata and Paeonia delavayi significantly inhibited neuraminidase activity at a concentration of 40 µg/mL. Dose-dependent inhibitory assays also revealed significant inhibition. The IC50 range of the TCM extracts for influenza virus neuraminidase was approximately 12.66-34.85 µg/mL, respectively. Some Chinese medicines have clear anti-influenza viral effects that may play an important role in the treatment of influenza through the inhibition of viral neuraminidase. The results of this study demonstrated that plant medicines can serve as a useful source of neuraminidase (NA) inhibitors and further investigation into the pharmacologic activities of these extracts is warranted.


Assuntos
Medicamentos de Ervas Chinesas/química , Inibidores Enzimáticos/química , Neuraminidase/antagonistas & inibidores , Proteínas Virais/antagonistas & inibidores , Medicamentos de Ervas Chinesas/uso terapêutico , Inibidores Enzimáticos/uso terapêutico , Humanos , Influenza Humana/tratamento farmacológico , Medicina Tradicional Chinesa
11.
Mol Neurobiol ; 51(3): 1053-63, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-24906954

RESUMO

Fragile X syndrome (FXS), a common form of inherited mental retardation, is caused by a loss of expression of the fragile X mental retardation protein (FMRP). FMRP is involved in brain functions by interacting with mRNAs and microRNAs (miRNAs) that selectively control gene expression at translational level. However, little is known about the role of FMRP in regulating miRNA expression. Here, we found a development-dependant dynamic expression of Fmr1 gene (encoding FMRP) in mouse hippocampus with a small peak at postnatal day 7 (P7). MiRNA microarray analysis showed that the levels of 38 miRNAs showed a significant increase with about 15 ~ 250-folds and the levels of 26 miRNAs showed a significant decrease with only about 2 ~ 4-folds in the hippocampus of P7 Fmr1 knockout (KO) mice. The qRT-PCR assay showed that nine of the most increased miRNAs (>100-folds in microarrays) increased about 40 ~ 70-folds and their pre-miRNAs increased about 5 ~ 10-folds, but no significant difference in their pri-miRNA levels was observed, suggesting that the alterations of partial miRNAs are an indirect consequence of FMRP lacking. We further demonstrated that a set of protein-coding mRNAs, potentially targeted by the nine miRNAs, were down-regulated in the hippocampus of Fmr1 KO mice. Finally, luciferase assays demonstrated that miR-34b, miR-340, and miR-148a could down-regulate the reporter gene expression by interacting with the Met 3' UTR. Taken together, these findings suggest that the miRNA expression alterations resulted from the absence of FMRP might contribute to molecular pathology of FXS.


Assuntos
Proteína do X Frágil de Retardo Mental/genética , Síndrome do Cromossomo X Frágil/genética , Síndrome do Cromossomo X Frágil/metabolismo , Perfilação da Expressão Gênica , MicroRNAs/biossíntese , MicroRNAs/genética , Animais , Linhagem Celular Tumoral , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica , Masculino , Camundongos , Camundongos Knockout
12.
Biochim Biophys Acta ; 1849(1): 1-9, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25459751

RESUMO

Increased expression of sodium channel SCN3A, an embryonic-expressed gene, has been identified in epileptic tissues, which is believed to contribute to the development of epilepsy. However, the regulatory mechanism of SCN3A expression under epileptic condition is still unknown. Here we showed a high level of Scn3a mRNA expression in mouse embryonic hippocampus with gradually decreasing to a low level during the postnatal development and a methylation of a specific CpG site (-39C) in the Scn3a promoter was increased in hippocampus during postnatal development, corresponding to the downregulation of Scn3a expression. Furthermore, in vitro methylation and -39C>T mutation of the Scn3a promoter decreased the reporter gene expression, suggesting an important role of the -39C site in regulating gene expression. We then demonstrated that the sequence containing -39C was a MBD2-binding motif and the CpG methylation of the promoter region increased the capability of MBD2's binding to the motif. Knockdown of MBD2 in mouse N1E-115 cells led to the -39C methylation and the downregulation of Scn3a transcription by decreasing the Scn3a promoter activity. In the hippocampus of seizure mice, the expressions of Scn3a and Mbd2 were upregulated after 10-day KA treatment. At the same time point, the -39C site was demethylated and the capability of MBD2's binding to the Scn3a promoter motif was decreased. Taken together, these findings suggest that CpG methylation and MBD2 are involved in altering Scn3a expression during postnatal development and seizure condition.


Assuntos
Proteínas de Ligação a DNA/biossíntese , Hipocampo/crescimento & desenvolvimento , Canal de Sódio Disparado por Voltagem NAV1.3/biossíntese , Convulsões/genética , Animais , Ilhas de CpG/genética , Metilação de DNA/genética , Proteínas de Ligação a DNA/genética , Desenvolvimento Embrionário , Regulação da Expressão Gênica no Desenvolvimento , Hipocampo/patologia , Humanos , Camundongos , Canal de Sódio Disparado por Voltagem NAV1.3/genética , RNA Mensageiro/genética , Convulsões/patologia , Transcrição Genética
13.
Mol Neurobiol ; 50(2): 438-48, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-24436055

RESUMO

Voltage-gated sodium channel α subunit type I (Nav1.1, encoded by SCN1A gene) plays a critical role in the initiation of action potential in the central nervous system. Downregulated expression of SCN1A is believed to be associated with epilepsy. Here, we found that the SCN1A promoter (P1c), located at the 5' untranslated exon 1c, drove the reporter gene expression in human NT2 cells, and a region between nt +53 and +62 downstream of the P1c promoter repressed the promoter activity. Further analyses showed that a scaffolding protein RACK1 (receptor for activated C kinase 1) was involved in binding to this silencer. Knockdown of RACK1 expression in NT2 cells deprived the repressive role of the silencer on the P1c promoter and increased SCN1A transcription, suggesting the potential involvement of RACK1 in negatively regulating SCN1A transcription via interaction with the silencer. Furthermore, we demonstrated that the binding of the protein complex including RACK1 to the SCN1A promoter motif was decreased in neuron-like differentiation of the NT2 cells induced by retinoic acid and resulted in the upregulation of SCN1A transcription. Taken together, this study reports a novel role of RACK1 in regulating SCN1A expression that participates in retinoic acid-induced neuronal differentiation of NT2 cells.


Assuntos
Proteínas de Ligação ao GTP/metabolismo , Mutação/genética , Canal de Sódio Disparado por Voltagem NAV1.1/genética , Proteínas de Neoplasias/metabolismo , Receptores de Superfície Celular/metabolismo , Diferenciação Celular/fisiologia , Células Cultivadas , Expressão Gênica/genética , Humanos , Proteínas do Tecido Nervoso/metabolismo , Regiões Promotoras Genéticas/genética , RNA Mensageiro/genética , Receptores de Quinase C Ativada , Transcrição Genética , Tretinoína/farmacologia
14.
Hum Genet ; 133(6): 801-11, 2014 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-24464349

RESUMO

Mutations in the SCN1A gene-encoding voltage-gated sodium channel α-I subunit (Nav1.1) cause various spectrum of epilepsies including Dravet syndrome (DS), a severe and intractable form. A large number of SCN1A mutations identified from the DS patients lead to the loss of function or truncation of Nav1.1 that result in a haploinsufficiency effects, indicating that the exact expression level of SCN1A should be essential to maintain normal brain function. In this study, we have identified five variants c.*1025T>C, c.*1031A>T, c.*1739C>T, c.*1794C>T and c.*1961C>T in the SCN1A 3' UTR in the patients with DS. The c.*1025T>C, c.*1031A>T and c.*1794C>T are conserved among different species. Of all the five variants, only c.*1794C>T is a novel variant and alters the predicted secondary structure of the 3' UTR. We also show that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) only binds to the 3' UTR sequence containing the mutation allele 1794U but not the wild-type allele 1794C, indicating that the mutation allele forms a new GAPDH-binding site. Functional analyses show that the variant negatively regulates the reporter gene expression by affecting the mRNA stability that is mediated by GAPDH's binding, and this phenomenon could be reversed by shRNA-induced GAPDH knockdown. These findings suggest that GAPDH and the 3'-UTR variant are involved in regulating SCN1A expression at post-transcriptional level, which may provide an important clue for further investigating on the relationship between 3'-UTR variants and SCN1A-related diseases.


Assuntos
Regiões 3' não Traduzidas , Epilepsias Mioclônicas/genética , Gliceraldeído-3-Fosfato Desidrogenases/antagonistas & inibidores , Mutação , Canal de Sódio Disparado por Voltagem NAV1.1/genética , Alelos , Sequência de Bases , Sítios de Ligação , Criança , Epilepsias Mioclônicas/metabolismo , Epilepsias Mioclônicas/patologia , Feminino , Regulação da Expressão Gênica , Gliceraldeído-3-Fosfato Desidrogenases/genética , Gliceraldeído-3-Fosfato Desidrogenases/metabolismo , Haploinsuficiência , Humanos , Dados de Sequência Molecular , Canal de Sódio Disparado por Voltagem NAV1.1/metabolismo , Linhagem , Ligação Proteica , Estabilidade de RNA , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo
15.
Neurosci Bull ; 29(3): 348-54, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23700283

RESUMO

LIM kinase 1 (LIMK1), a cytosolic serine/threonine kinase, regulates actin filament dynamics and reorganization and is involved in neuronal development and brain function. Abnormal expression of LIMK1 is associated with several neurological disorders. In this study, we performed a conservation analysis using Vector NTI (8.0) software. The dualluciferase reporter assay and real-time quantitative RT-PCR were used to assess the protein and mRNA levels of the reporter gene, respectively. We found that a region ranging from nt +884 to +966 in the human LIMK1 3' untranslated region (UTR) was highly conserved in the mouse Limk1 3' UTR and formed a structure containing several loops and stems. Luciferase assay showed that the relative luciferase activity of the mutated construct with the conserved region deleted, pGL4-hLIMK1-3U-M, in SH-SY5Y and HEK-293 cells was only ~60% of that of the wild-type construct pGL4-hLIMK1-3U, indicating that the conserved region is critical for the reporter gene expression. Real-time quantitative RT-PCR analysis demonstrated that the relative Luc2 mRNA levels in SH-SY5Y and HEK293 cells transfected with pGL4-hLIMK1-3U-M decreased to ~50% of that in cells transfected with pGL4-hLIMK1-3U, suggesting an important role of the conserved region in maintaining Luc2 mRNA stability. Our study suggests that the conserved region in the LIMK1 3' UTR is involved in regulating LIMK1 expression at the post-transcriptional level, which may help reveal the mechanism underlying the regulation of LIMK1 expression in the central nervous system and explore the relationship between the 3'-UTR mutant and neurological disorders.


Assuntos
Regiões 3' não Traduzidas/genética , Sistema Nervoso Central/metabolismo , Quinases Lim/metabolismo , Ativação Transcricional/fisiologia , Animais , Linhagem Celular , Regulação da Expressão Gênica , Humanos , Quinases Lim/genética , Camundongos , Fosforilação , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ativação Transcricional/genética , Transfecção/métodos
16.
Zhongguo Shi Yan Xue Ye Xue Za Zhi ; 16(2): 247-53, 2008 Apr.
Artigo em Chinês | MEDLINE | ID: mdl-18426642

RESUMO

This study was purpose to investigate the effects of CD147 on the invasiveness of leukemia cells U937. The experiments were divided into 4 groups: control group, LPS group, CD147mAb group and LPS+CD147 mAb group. Cells were treated by lipopolysaccharide (LPS) or anti-CD147 monoclonal antibody, and the expression of CD147 and MMP-2, -9, the invasive potential of the cells in vitro and ex vivo, as well as the invasion of the implanted tumors in SCID mice were analysed by RT-PCR, FCM, gel zymography and invasion test in vitro respectively. The results showed that the expression of CD147 was elevated by the induction of LPS, and the enhanced expression of CD147 on U937 cells increased the production and secretion of MMP-2 and MMP-9 as measured by reverse transcription-PCR and gel zymography. An increased number of LPS-induced cells invading through a reconstituted basement membrane were observed by invasion assays. These responses were down-regulated after blocking CD147 with anti-CD147 antibody. At 30 days after intravenous injection of LPS pretreated U937 cells to SCID mice human U937 cells were found in the bone marrow and lung of the mice, indicating the invasion of the tumor cells. And overexpressions of CD147, MMP-2 and MMP-9 were found in the lung tissue of the mice injected with LPS-treated but not anti-CD147 antibody treated tumor cells. It is concluded that overexpression of CD147 on U937 cells may increase the secretion and activation of MMP-2 and MMP-9 and thus promote the invasiveness of the tumor cells.


Assuntos
Anticorpos Monoclonais/farmacologia , Basigina/metabolismo , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Invasividade Neoplásica , Animais , Basigina/genética , Basigina/imunologia , Feminino , Humanos , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 9 da Matriz/genética , Camundongos , Camundongos Nus , Camundongos SCID , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Células U937
17.
Ai Zheng ; 26(5): 473-9, 2007 May.
Artigo em Chinês | MEDLINE | ID: mdl-17672935

RESUMO

BACKGROUND & OBJECTIVE: The metastatic potentiality of malignancies is closely associated with their biological dynamic properties, which are affected by intracellular Ca2+ current activity. This study was to investigate the correlation of Ca2+ current features of sub-clonal nasopharyngeal carcinoma (NPC) cell lines 5-8F and 6-10B with different metastatic potentiality to their moving abilities. METHODS: 5-8F cells, with higher metastatic potentiality, and 6-10B cells, with lower metastatic potentiality, were cultured with herbal medicine-containing serum, which holds significant metastasis-inhibiting effect on tumor cells. Cell proliferation was assessed by MTT assay. The expression of nm23-H1 was detected by Western blot. Intracellular Ca2+ current features were detected with patch clamp technique in a whole cell recording way, while cell moving ability was determined by streak culturing assay. RESULTS: The expression of nm23-H1 was significantly lower in 5-8F cells than in 6-10B cells (2.3+/-0.2 vs. 2.9+/-0.4). The Ca2+ release-activated Ca2+ influx current (ICRAC) was significantly lower in 5-8F cells than in 6-10B cells [(-1.39+/-0.36) nA vs. (-0.66+/-0.40) nA, P < 0.05]. The number of cells moved across the streak was significantly higher in 5-8F cells than in 6-10B cells (350+/-3 vs. 246+/-1, P< 0.05). When cultured with herbal medicine-containing serum, no significant difference in proliferation was found between 5-8F cells and 6-10B cells; the expression of nm23-H1 was significantly higher in 5-8F cells than in 6-10B cells(3.9+/-0.1 vs.1.0+/-0.1,P<0.05)û the ICRAC was decreased to (-1.27+/-0.35) nA in 5-8F cells and decreased to (-0.37+/-0.23) nA in 6-10B cell, and the inhibition rate was significantly higher in 5-8F cells than in 6-10B cells [(1.90+/-0.47)% vs. (0.46+/-0.12)%, P < 0.05]û the number of cells moved across the streak was significantly lower in 5-8F cells than in 6-10B cells (94+/-6 vs. 229+/-6, P < 0.05). CONCLUSIONS: There are significant differences in nm23-H1 protein expression, ICRAC level and cell moving ability between 5-8F and 6-10B cells. Medicine intervention could inhibit Ca2+ current and moving ability of 5-8F cells, and meanwhile increase the nm23-H1 activity.


Assuntos
Canais de Cálcio/fisiologia , Movimento Celular/efeitos dos fármacos , Medicamentos de Ervas Chinesas/farmacologia , Nucleosídeo NM23 Difosfato Quinases/metabolismo , Neoplasias Nasofaríngeas , Animais , Antineoplásicos Fitogênicos/farmacologia , Cálcio/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Combinação de Medicamentos , Feminino , Humanos , Masculino , Neoplasias Nasofaríngeas/metabolismo , Neoplasias Nasofaríngeas/patologia , Neoplasias Nasofaríngeas/fisiopatologia , Metástase Neoplásica , Técnicas de Patch-Clamp , Ratos , Ratos Sprague-Dawley
18.
Zhongguo Zhong Xi Yi Jie He Za Zhi ; 26(12): 1086-9, 2006 Dec.
Artigo em Chinês | MEDLINE | ID: mdl-17205820

RESUMO

OBJECTIVE: To investigate the correlation between TCM syndrome type and intracranial aggressive potentiality of untreated nasopharyngeal carcinoma (NPC). METHODS: Sixty untreated NPC patients of different syndrome types were treated conventionally and followed up for over one year. Correlation between the TCM syndrome type differentiated at the first consultation and the intracranial aggressive potentiality of the primary focus of NPC were analyzed. RESULTS: The incidence of intracranial aggression was significantly higher in patients with Qi-Yin deficiency type than that in those with other two syndrome types during the follow-up period (P < 0.01). CONCLUSION: The intracranial aggessive rate in the untreated NPC patients of Qi-Yin deficiency type was higher than in those of either Qi and blood coagulation type or fire-toxin stagnation type.


Assuntos
Neoplasias Encefálicas/secundário , Medicina Tradicional Chinesa , Neoplasias Nasofaríngeas/patologia , Adulto , Diagnóstico Diferencial , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Neoplasias Nasofaríngeas/diagnóstico , Neoplasias Nasofaríngeas/terapia , Invasividade Neoplásica , Síndrome
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