Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 61
Filtrar
Mais filtros

Base de dados
País/Região como assunto
Tipo de documento
Intervalo de ano de publicação
1.
Nucleic Acids Res ; 47(2): 634-647, 2019 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-30407533

RESUMO

In Trypanosoma brucei, genes are arranged in Polycistronic Transcription Units (PTUs), which are demarcated by transcription start and stop sites. Transcription start sites are also binding sites of Origin Recognition Complex 1 (ORC1). This spatial coincidence implies that transcription and replication in trypanosomes must occur in a highly ordered and cooperative manner. Interestingly, a previously published genetic screen identified the T. brucei MCM-BP, which interacts with subunits of MCM helicase, as a protein whose downregulation results in the loss of transcriptional silencing at subtelomeric loci. Here, I show that TbMCM-BP is required for DNA replication and transcription. TbMCM-BP depletion causes a significant reduction of replicating cells in S phase and genome-wide impairments of replication origin activation. Moreover, levels of sense and antisense transcripts increase at boundaries of PTUs in the absence of TbMCM-BP. TbMCM-BP is also important for transcriptional repression of the specialized subtelomeric PTUs, the Bloodstream-form Expression-Sites (BESs), which house the major antigenic determinant (the Variant Surface Glycoprotein, VSG gene) as well as TbORC1 binding sites. Overall, this study reveals that TbMCM-BP, a replication initiation protein, also guides the initiation, termination and directionality of transcription.


Assuntos
Replicação do DNA , Proteínas de Protozoários/fisiologia , Transcrição Gênica , Trypanosoma brucei brucei/genética , Dano ao DNA , DNA de Protozoário/biossíntese , Regulação da Expressão Gênica , Genoma de Protozoário , RNA Antissenso/biossíntese , Regiões Terminadoras Genéticas , Sítio de Iniciação de Transcrição , Trypanosoma brucei brucei/metabolismo
2.
PLoS Genet ; 12(1): e1005762, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26796638

RESUMO

Trypanosoma brucei is a protozoan parasite that lacks many transcription factors found in other eukaryotes, such as those whose binding demarcates enhancers. T. brucei retains histone variants and modifications, however, and it is hypothesized that it relies on epigenetic marks to define transcription-related boundaries. The histone H3 variant (H3.V) and an alternate nucleotide, base J (ß-D-glucosyl-hydroxymethyluracil), are two chromatin marks found at both transcription termination sites (TTSs) and telomeres. Here, we report that the absence of both base J and H3.V result in transcription readthrough and the appearance of antisense transcripts near TTSs. Additionally, we find that maintaining the transcriptional silencing of pol I-transcribed telomeric Variant Surface Glycoprotein (VSG) genes appears to be dependent on deposition of H3.V alone. Our study reveals that gene expression depends on different epigenetic cues depending on chromosomal location and on the transcribing polymerase. This work provides insight into how these signals may have evolved into the more nuanced and fine-tuned gene regulatory mechanisms observed in other model systems.


Assuntos
Histonas/genética , Transcrição Gênica , Trypanosoma brucei brucei/genética , Glicoproteínas Variantes de Superfície de Trypanosoma/genética , Cromatina/genética , Regulação da Expressão Gênica , Glucosídeos/genética , Regiões Promotoras Genéticas , RNA Polimerase I/genética , Telômero/genética , Uracila/análogos & derivados
3.
PLoS Biol ; 13(12): e1002316, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26646171

RESUMO

Trypanosoma brucei, the causative agent of African sleeping sickness, is transmitted to its mammalian host by the tsetse. In the fly, the parasite's surface is covered with invariant procyclin, while in the mammal it resides extracellularly in its bloodstream form (BF) and is densely covered with highly immunogenic Variant Surface Glycoprotein (VSG). In the BF, the parasite varies this highly immunogenic surface VSG using a repertoire of ~2500 distinct VSG genes. Recent reports in mammalian systems point to a role for histone acetyl-lysine recognizing bromodomain proteins in the maintenance of stem cell fate, leading us to hypothesize that bromodomain proteins may maintain the BF cell fate in trypanosomes. Using small-molecule inhibitors and genetic mutants for individual bromodomain proteins, we performed RNA-seq experiments that revealed changes in the transcriptome similar to those seen in cells differentiating from the BF to the insect stage. This was recapitulated at the protein level by the appearance of insect-stage proteins on the cell surface. Furthermore, bromodomain inhibition disrupts two major BF-specific immune evasion mechanisms that trypanosomes harness to evade mammalian host antibody responses. First, monoallelic expression of the antigenically varied VSG is disrupted. Second, rapid internalization of antibodies bound to VSG on the surface of the trypanosome is blocked. Thus, our studies reveal a role for trypanosome bromodomain proteins in maintaining bloodstream stage identity and immune evasion. Importantly, bromodomain inhibition leads to a decrease in virulence in a mouse model of infection, establishing these proteins as potential therapeutic drug targets for trypanosomiasis. Our 1.25Å resolution crystal structure of a trypanosome bromodomain in complex with I-BET151 reveals a novel binding mode of the inhibitor, which serves as a promising starting point for rational drug design.


Assuntos
Modelos Moleculares , Proteínas de Protozoários/metabolismo , Fatores de Transcrição/metabolismo , Trypanosoma brucei brucei/fisiologia , Substituição de Aminoácidos , Animais , Sítios de Ligação , Linhagem Celular , Técnicas de Silenciamento de Genes , Técnicas de Inativação de Genes , Evasão da Resposta Imune , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Mutantes/química , Proteínas Mutantes/metabolismo , Conformação Proteica , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Proteínas de Protozoários/antagonistas & inibidores , Proteínas de Protozoários/química , Proteínas de Protozoários/genética , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Análise de Sobrevida , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/química , Fatores de Transcrição/genética , Tripanossomicidas/farmacologia , Tripanossomicidas/uso terapêutico , Trypanosoma brucei brucei/efeitos dos fármacos , Trypanosoma brucei brucei/imunologia , Trypanosoma brucei brucei/patogenicidade , Tripanossomíase Africana/tratamento farmacológico , Tripanossomíase Africana/parasitologia , Tripanossomíase Africana/fisiopatologia , Virulência
4.
J Epidemiol ; 26(5): 233-41, 2016 May 05.
Artigo em Inglês | MEDLINE | ID: mdl-26853101

RESUMO

BACKGROUND: Prompt treatment affects prognosis and survival after acute myocardial infarction (AMI) onset. This study evaluated the awareness of early symptoms of AMI and knowledge of appropriate responses on symptom occurrence, along with related factors. METHODS: Participants' knowledge of the early symptoms of and responses to AMI onset were investigated using a random digit dialing survey. We included 9600 residents of 16 metropolitan cities and provinces in Korea. RESULTS: The proportions of respondents who were aware of early symptoms of AMI ranged from 32.9% (arm or shoulder pain) to 79.1% (chest pain and discomfort). Of the respondents, 67.0% would call an ambulance if someone showed signs of AMI, 88.7% knew ≥1 symptom, 10.9% knew all five symptoms, and 3.1% had excellent knowledge (correct identification of all five AMI symptoms, not answering "Yes" to the trap question, and correctly identifying calling an ambulance as the appropriate response when someone is exhibiting AMI symptoms). The odds ratio (OR) for having excellent knowledge was significantly higher for those who graduated college or higher (OR 3.42; 95% confidence interval [CI], 1.09-10.76) than for those with less than a primary school education, as well as for subjects with AMI advertisement exposure (OR 1.49; 95% CI, 1.10-2.02) and with knowledge of AMI (OR 1.63; 95% CI, 1.16-2.27). The 60- to 79-year-old group had significantly lower OR for excellent knowledge than the 20- to 39-year-old group (OR 0.53; 95% CI, 0.28-0.99). CONCLUSIONS: Awareness of AMI symptoms and the appropriate action to take after symptom onset in South Korea was poor. Therefore, educational and promotional strategies to increase the overall awareness in the general public, especially in the elderly and those with low education levels, are needed.


Assuntos
Diagnóstico Precoce , Tratamento de Emergência , Conhecimentos, Atitudes e Prática em Saúde , Infarto do Miocárdio/diagnóstico , Infarto do Miocárdio/terapia , Adulto , Fatores Etários , Idoso , Escolaridade , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , República da Coreia , Inquéritos e Questionários , Telefone , Adulto Jovem
5.
J Korean Med Sci ; 31(11): 1703-1710, 2016 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-27709846

RESUMO

The aim of this study was to investigate time trends in the public awareness of stroke and its predicting factors. The target population was 9,600 community-dwelling adults, aged 19-79 years, in 16 metropolitan cities and provinces in Korea. The survey samples in 2012 and 2014 were selected separately (entirely different sets of subjects) using a proportionate quota sampling method. Information concerning knowledge of stroke and demographics was collected by trained telephone interviewers using random digit dialing. After excluding subjects with a non-response or refusal to answer any question, the analyses included 8,191 subjects in 2012 and 8,127 subjects in 2014. Respondents' awareness of stroke warning signs (numbness or weakness, difficulty speaking or understanding speech, dizziness, visual impairment, and severe headache) was highest for difficulty speaking or understanding speech (80.9% in 2012 and 86.4% in 2014). There were significant increases in the proportion of respondents understanding the appropriate action (i.e., calling an ambulance) at the time of stroke occurrence (59.6% to 67.1%), and in the proportion aware of the general need for prompt treatment (86.7% to 89.8%). In multivariable logistic regression analysis, older age, higher education level, higher household income, current non-smoking, exposure to stroke-related public relations materials, and experience of stroke education were significantly associated with both high knowledge of stroke warning signs and awareness of the need for prompt treatment. Between 2012 and 2014, the public's awareness of stroke increased significantly. More specialized interventions, including public relations materials and education, should focus on subgroups who have lower stroke knowledge.


Assuntos
Conhecimentos, Atitudes e Prática em Saúde , Acidente Vascular Cerebral/patologia , Adulto , Fatores Etários , Idoso , Feminino , Humanos , Entrevistas como Assunto , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Razão de Chances , República da Coreia , Fumar , Classe Social , Inquéritos e Questionários , Telefone , Adulto Jovem
6.
Biotechnol Lett ; 37(8): 1637-44, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25851950

RESUMO

OBJECTIVE: To examine the role of a gene encoding flavin-containing monooxygenase (cFMO) from Corynebacterium glutamicum ATCC13032 when cloned and expressed in Escherichia coli for the production of indigo pigments. RESULTS: The blue pigments produced by recombinant E. coli were identified as indigo and indirubin. The cFMO was purified as a fused form with maltose-binding protein (MBP). The enzyme was optimal at 25 °C and pH 8. From absorption spectrum analysis, the cFMO was classified as a flavoprotein. FMO activity was strongly inhibited by 1 mM Cu(2+) and recovered by adding 1-10 mM EDTA. The enzyme catalyzed the oxidation of TMA, thiourea, and cysteamine, but not glutathione or cysteine. MBP-cFMO had an indole oxygenase activity through oxygenation of indole to indoxyl. The recombinant E. coli produced 685 mg indigo l(-1) and 103 mg indirubin l(-1) from 2.5 g L-tryptophan l(-1). CONCLUSION: The results suggest the cFMO can be used for the microbial production of both indigo and indirubin.


Assuntos
Corynebacterium glutamicum/enzimologia , Índigo Carmim/metabolismo , Oxigenases/metabolismo , Clonagem Molecular , Cobre/metabolismo , Corynebacterium glutamicum/genética , Inibidores Enzimáticos/metabolismo , Estabilidade Enzimática , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Concentração de Íons de Hidrogênio , Indóis/metabolismo , Oxigenases/química , Oxigenases/genética , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Análise Espectral , Temperatura
7.
Mol Microbiol ; 87(1): 196-210, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23216794

RESUMO

Binding of the Origin Recognition Complex (ORC) to replication origins is essential for initiation of DNA replication, but ORC has non-essential functions outside of DNA replication, including in heterochromatic gene silencing and telomere maintenance. Trypanosoma brucei, a protozoan parasite that causes human African trypanosomiasis, uses antigenic variation as a major virulence mechanism to evade the host's immune attack by expressing its major surface antigen, the Variant Surface Glycoprotein (VSG), in a monoallelic manner. An Orc1/Cdc6 homologue has been identified in T. brucei, but its role in DNA replication has not been directly confirmed and its potential involvement in VSG repression or switching has not been thoroughly investigated. In this study, we show that TbOrc1 is essential for nuclear DNA replication in mammalian-infectious bloodstream and tsetse procyclic forms (BF and PF). Depletion of TbOrc1 resulted in derepression of telomere-linked silent VSGs in both BF and PF, and increased VSG switching particularly through the in situ transcriptional switching mechanism. TbOrc1 associates with telomere repeats but appears to do so independently of two known T. brucei telomere proteins, TbRAP1 and TbTRF. We conclude that TbOrc1 has conserved functions in DNA replication and is also required to control telomere-linked VSG expression and VSG switching.


Assuntos
Inativação Gênica , Complexo de Reconhecimento de Origem/genética , Trypanosoma brucei brucei/genética , Glicoproteínas Variantes de Superfície de Trypanosoma/genética , Variação Antigênica , Replicação do DNA , DNA de Protozoário/biossíntese , DNA de Protozoário/genética , Genes de Protozoários , Glicoproteínas de Membrana/genética , Complexo de Reconhecimento de Origem/metabolismo , Regiões Promotoras Genéticas , Trypanosoma brucei brucei/metabolismo
8.
Appl Microbiol Biotechnol ; 98(5): 2145-54, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23893328

RESUMO

A novel acetylalginate esterase (AcAlgE) gene was cloned and characterized from the genomic DNA library of Sphingomonas sp. MJ-3. A putative gene encoding AcAlgE protein of 292-residue precursor protein with 20-amino acid signal peptide was identified in the alg operon. The deduced AcAlgE protein has GDSL-like consensus motif and shares a highest sequence identity (51%) with GDSL family lipolytic protein from Pseudoxanthomonas suwonensis. Enzymatic assays with bacterial acetylalginate as the substrate showed that the recombinant AcAlgE protein possesses deacetylation activity. The optimal temperature and pH for the AcAlgE were 22 °C and pH 6.5 (citrate buffer), respectively. The recombinant AcAlgE protein catalyzed deacetylation of acetylalginate with release of acetate. The resulting de-acetylated alginate was readily degraded by alginate lyases, indicating that the recombinant AcAlgE enhanced the subsequent degradation of acetylalginate by alginate lyases. The recombinant AcAlgE can play an important role in the degradation of acetylated alginate such as mucoidal acetylalginate in cystic fibrosis patient.


Assuntos
Alginatos/metabolismo , Esterases/metabolismo , Sphingomonas/enzimologia , Acetatos/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Clonagem Molecular , Estabilidade Enzimática , Esterases/química , Esterases/genética , Esterases/isolamento & purificação , Expressão Gênica , Ácido Glucurônico/metabolismo , Ácidos Hexurônicos/metabolismo , Concentração de Íons de Hidrogênio , Dados de Sequência Molecular , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Sphingomonas/genética , Temperatura
9.
J Korean Med Sci ; 28(1): 16-24, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23341707

RESUMO

This study attempted to calculate and investigate the incidence of hospitalized acute myocardial infarction (AMI) and stroke in Korea. Using the National Health Insurance claim data, we investigated patients whose main diagnostic codes included AMI or stroke during 2006 to 2010. As a result, we found out that the number of AMI hospitalized patients had decreased since 2006 and amounted to 15,893 in 2010; and that the number of those with stroke had decreased since 2006 and amounted to 73,501 in 2010. The age-standardized incidence rate of hospitalized AMI, after adjustment for readmission, was 41.6 cases per 100,000-population in 2006, and had decreased to 29.4 cases in 2010 (for trend P < 0.001). In the case of stroke was estimated at 172.8 cases per 100,000-population in 2006, and had decreased to 135.1 cases in 2010 (for trend P < 0.001). In conclusion, the age-standardized incidence rates of both hospitalized AMI and stroke in Korea had decreased continuously during 2006 to 2010. We consider this decreasing trend due to the active use of pharmaceuticals, early vascular intervention, and the national cardio-cerebrovascular disease care project as the primary and secondary prevention efforts.


Assuntos
Hospitalização/tendências , Infarto do Miocárdio/epidemiologia , Acidente Vascular Cerebral/epidemiologia , Doença Aguda , Adolescente , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Feminino , Humanos , Incidência , Lactente , Recém-Nascido , Masculino , Pessoa de Meia-Idade , Readmissão do Paciente , República da Coreia/epidemiologia , Fatores Sexuais , Adulto Jovem
10.
Nat Commun ; 14(1): 4390, 2023 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-37474515

RESUMO

Replication Protein A (RPA) is a broadly conserved complex comprised of the RPA1, 2 and 3 subunits. RPA protects the exposed single-stranded DNA (ssDNA) during DNA replication and repair. Using structural modeling, we discover an inhibitor, JC-229, that targets RPA1 in Trypanosoma brucei, the causative parasite of African trypanosomiasis. The inhibitor is highly toxic to T. brucei cells, while mildly toxic to human cells. JC-229 treatment mimics the effects of TbRPA1 depletion, including DNA replication inhibition and DNA damage accumulation. In-vitro ssDNA-binding assays demonstrate that JC-229 inhibits the activity of TbRPA1, but not the human ortholog. Indeed, despite the high sequence identity with T. cruzi and Leishmania RPA1, JC-229 only impacts the ssDNA-binding activity of TbRPA1. Site-directed mutagenesis confirms that the DNA-Binding Domain A (DBD-A) in TbRPA1 contains a JC-229 binding pocket. Residue Serine 105 determines specific binding and inhibition of TbRPA1 but not T. cruzi and Leishmania RPA1. Our data suggest a path toward developing and testing highly specific inhibitors for the treatment of African trypanosomiasis.


Assuntos
Doença de Chagas , Trypanosoma brucei brucei , Tripanossomíase Africana , Animais , Humanos , Trypanosoma brucei brucei/genética , Proteína de Replicação A/metabolismo , Replicação do DNA , DNA de Cadeia Simples/genética , Ligação Proteica
11.
PLoS Pathog ; 6(7): e1000992, 2010 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-20628569

RESUMO

Homologous recombination (HR) mediates one of the major mechanisms of trypanosome antigenic variation by placing a different variant surface glycoprotein (VSG) gene under the control of the active expression site (ES). It is believed that the majority of VSG switching events occur by duplicative gene conversion, but only a few DNA repair genes that are central to HR have been assigned a role in this process. Gene conversion events that are associated with crossover are rarely seen in VSG switching, similar to mitotic HR. In other organisms, TOPO3alpha (Top3 in yeasts), a type IA topoisomerase, is part of a complex that is involved in the suppression of crossovers. We therefore asked whether a related mechanism might suppress VSG recombination. Using a set of reliable recombination and switching assays that could score individual switching mechanisms, we discovered that TOPO3alpha function is conserved in Trypanosoma brucei and that TOPO3alpha plays a critical role in antigenic switching. Switching frequency increased 10-40-fold in the absence of TOPO3alpha and this hyper-switching phenotype required RAD51. Moreover, the preference of 70-bp repeats for VSG recombination was mitigated, while homology regions elsewhere in ES were highly favored, in the absence of TOPO3alpha. Our data suggest that TOPO3alpha may remove undesirable recombination intermediates constantly arising between active and silent ESs, thereby balancing ES integrity against VSG recombination.


Assuntos
Variação Antigênica/genética , DNA Topoisomerases Tipo I/fisiologia , Conversão Gênica , Trypanosoma brucei brucei/genética , Glicoproteínas Variantes de Superfície de Trypanosoma/genética , Proteínas de Bactérias/genética , Proteínas Fúngicas , Humanos , Recombinação Genética , Trypanosoma brucei brucei/imunologia
12.
Appl Microbiol Biotechnol ; 95(6): 1643-53, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-22805784

RESUMO

A gene for a polyMG-specific alginate lyase possessing a novel structure was identified and cloned from Stenotrophomas maltophilia KJ-2 by using PCR with homologous nucleotide sequences-based primers. The recombinant alginate lyase consisting of 475 amino acids was purified on Ni-Sepharose column and exhibited the highest activity at pH 8 and 40 °C. Interestingly, the recombinant alginate lyase was expected to have a similar catalytic active site of chondroitin B lyase but did not show chondroitin lyase activity. In the test of substrate specificity, the recombinant alginate lyase preferentially degraded the glycosidic bond of polyMG-block than polyM-block and polyG-block. The chemical structures of the degraded alginate oligosaccharides were elucidated to have mannuronate (M) at the reducing end on the basis of NMR analysis, supporting that KJ-2 polyMG-specific alginate lyase preferably degraded the glycosidic bond in M-G linkage than that in G-M linkage. The KJ-2 polyMG-specific alginate lyase can be used in combination with other alginate lyases for a synergistic saccharification of alginate.


Assuntos
Alginatos/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/isolamento & purificação , Clonagem Molecular , Polissacarídeo-Liases/química , Polissacarídeo-Liases/isolamento & purificação , Stenotrophomonas maltophilia/enzimologia , Alginatos/química , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Estabilidade Enzimática , Ácido Glucurônico/química , Ácido Glucurônico/metabolismo , Ácidos Hexurônicos/química , Ácidos Hexurônicos/metabolismo , Concentração de Íons de Hidrogênio , Dados de Sequência Molecular , Polissacarídeo-Liases/genética , Polissacarídeo-Liases/metabolismo , Alinhamento de Sequência , Stenotrophomonas maltophilia/química , Stenotrophomonas maltophilia/genética , Stenotrophomonas maltophilia/metabolismo , Especificidade por Substrato
13.
Can J Physiol Pharmacol ; 90(7): 863-72, 2012 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-22702717

RESUMO

Mepivacaine is an aminoamide-linked local anesthetic with an intermediate duration that intrinsically produces vasoconstriction both in vivo and in vitro. The aims of this in-vitro study were to examine the direct effect of mepivacaine in isolated rat aortic rings and to determine the associated cellular mechanism with a particular focus on endothelium-derived vasodilators, which modulate vascular tone. In the aortic rings with or without endothelium, cumulative mepivacaine concentration-response curves were generated in the presence or absence of the following antagonists: N(ω)-nitro-L-arginine methyl ester [L-NAME], indomethacin, fluconazole, methylene blue, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one [ODQ], verapamil, and calcium-free Krebs solution. Mepivacaine produced vasoconstriction at low concentrations (1 × 10(-3) and 3 × 10(-3) mol/L) followed by vasodilation at a high concentration (1 × 10(-2) mol/L). The mepivacaine-induced contraction was higher in endothelium-denuded aortae than in endothelium-intact aortae. Pretreatment with L-NAME, ODQ, and methylene blue enhanced mepivacaine-induced contraction in the endothelium-intact rings, whereas fluconazole had no effect. Indomethacin slightly attenuated mepivacaine-induced contraction, whereas verapamil and calcium-free Krebs solution more strongly attenuated this contraction. The vasoconstriction induced by mepivacaine is attenuated mainly by the endothelial nitric oxide - cyclic guanosine monophosphate pathway. In addition, mepivacaine-induced contraction involves cyclooxygenase pathway activation and extracellular calcium influx via voltage-operated calcium channels.


Assuntos
Anestésicos Locais/farmacologia , Aorta/efeitos dos fármacos , Endotélio Vascular/efeitos dos fármacos , Mepivacaína/farmacologia , Músculo Liso Vascular/efeitos dos fármacos , Vasoconstrição/efeitos dos fármacos , Animais , Aorta/metabolismo , Cálcio/metabolismo , Canais de Cálcio/metabolismo , Células Cultivadas , GMP Cíclico/metabolismo , Endotélio Vascular/metabolismo , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Masculino , Contração Muscular/efeitos dos fármacos , Músculo Liso Vascular/metabolismo , Óxido Nítrico/metabolismo , Prostaglandina-Endoperóxido Sintases/metabolismo , Ratos , Ratos Sprague-Dawley , Vasodilatação/efeitos dos fármacos
14.
Phytother Res ; 26(4): 605-12, 2012 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-21971692

RESUMO

The ethanol extract of the flower of P. vulgaris var. lilacina (EEPV) has been used traditionally as an antiinflammatory agent in many countries. Inducers of heme oxygenase-1 (HO-1) reduce high mobility group box 1 (HMGB1), a late phase cytokine, in sepsis. Although EEPV has been used as an antiinflammatory agent, no report is available as to whether it modifies HMGB1 in sepsis due to HO-1 induction. It was found that EEPV increased HO-1 protein expression in RAW 264.7 cells, which was significantly inhibited by LY294002, but not PD98059, SB203580 or SP600125. In addition, EEPV activated NF-E2-related factor (Nrf2) to move from the cytosol to the nucleus, and EEPV-induced HO-1 and activation of ARE-luciferase activity were significantly reduced by siNrf2 transfection and LY294002 but not SB203508. EEPV also significantly inhibited NF-κB luciferase activity, and decreased both iNOS/NO and COX-2/PGE(2) production in lipopolysaccharide (LPS)-stimulated macrophages which was reversed by siHO-1 RNA transfection. Importantly, EEPV inhibited HMGB1 release in LPS-activated macrophages in a PI3K-sensitive manner and reduced serum HMGB1 level and lung HMGB1 expression in cecal ligation and puncture (CLP)-induced septic mice. It is concluded that EEPV induces HO-1 expression through PI3K/Nrf2 signal pathways, which may be beneficial for the treatment of sepsis due to a reduction of HMGB1 release.


Assuntos
Ceco/lesões , Proteína HMGB1/antagonistas & inibidores , Heme Oxigenase-1/metabolismo , Proteínas de Membrana/metabolismo , Extratos Vegetais/farmacologia , Prunella/química , Animais , Antracenos/farmacologia , Ceco/patologia , Linhagem Celular , Sobrevivência Celular , Cromonas/farmacologia , Ciclo-Oxigenase 2/metabolismo , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Ativação Enzimática , Indução Enzimática , Etanol , Flavonoides/farmacologia , Flores/química , Vetores Genéticos , Proteína HMGB1/sangue , Heme Oxigenase-1/antagonistas & inibidores , Imidazóis/farmacologia , Lipopolissacarídeos/efeitos adversos , Pulmão/patologia , Macrófagos/efeitos dos fármacos , Macrófagos/enzimologia , Masculino , Proteínas de Membrana/antagonistas & inibidores , Camundongos , Camundongos Endogâmicos BALB C , Morfolinas/farmacologia , Fator 2 Relacionado a NF-E2/metabolismo , NF-kappa B/metabolismo , Óxido Nítrico Sintase Tipo II/antagonistas & inibidores , Óxido Nítrico Sintase Tipo II/metabolismo , Transporte Proteico , Piridinas/farmacologia , Sepse/patologia , Transdução de Sinais , Transfecção
15.
Can J Microbiol ; 57(12): 1032-41, 2011 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-22145710

RESUMO

An alginate lyase gene of a newly isolated Pseudomonas sp. strain KS-408 was cloned by using PCR with the specific primers designed from homologous nucleotide sequences. A partial protein sequence of KS-408 alginate lyase was homology-modeled on the basis of the crystal structure of A1-III alginate lyase from Sphingomonas sp. strain A1. The proposed 3-D structure of KS-408 alginate lyase shows that Asn-198, His-199, Arg-246, and Tyr-253 residues are conserved for the catalytic active site. The recombinant KS-408-1F (with signal peptide) and KS-408-2F (without signal peptide) alginate lyases with the (His)(6) tag consist of 393 (44.5 kDa) and 372 (42.4 kDa) amino acids with isoelectric points of 8.64 and 8.46, respectively. The purified recombinant KS-408 alginate lyase was very stable when it was incubated at 40 °C for 30 min. Alginate oligosaccharides produced by the KS-408-2F alginate lyase were purified on a Bio-Gel P2 column and analyzed by thin-layer chromatography, fast-protein liquid chromatography, and electrospray ionization mass spectrometry. (1)H NMR data showed that the KS-408-2F alginate lyase cleaved the glycosidic linkages between two mannuronates (mannuronate-ß(1-4)-mannuronate) or mannuronate and guluronate (mannuronate-ß(1-4)-guluronate), indicating that the KS-408 alginate lyase is a polyM-specific lyase.


Assuntos
Polissacarídeo-Liases/química , Polissacarídeo-Liases/genética , Pseudomonas/enzimologia , Pseudomonas/genética , Alginatos/química , Alginatos/metabolismo , Animais , Ativação Enzimática/efeitos dos fármacos , Ácido Glucurônico/química , Ácido Glucurônico/metabolismo , Glicosídeos/metabolismo , Ácidos Hexurônicos/química , Ácidos Hexurônicos/metabolismo , Concentração de Íons de Hidrogênio , Metais/farmacologia , Modelos Moleculares , Dados de Sequência Molecular , Ressonância Magnética Nuclear Biomolecular , Polissacarídeo-Liases/isolamento & purificação , Polissacarídeo-Liases/metabolismo , Estrutura Terciária de Proteína , RNA Ribossômico 16S/genética , Proteínas Recombinantes/metabolismo , Sphingomonas/enzimologia , Sphingomonas/genética
16.
Bioprocess Biosyst Eng ; 34(1): 113-9, 2011 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-20640446

RESUMO

The marine alginate lyase from Streptomyces sp. ALG-5, which specifically degrades poly-G block of alginate, was functionally expressed as a His-tagged form with an Escherichia coli expression system. The recombinant alginate lyase expressed with pColdI at 15 °C exhibited the highest alginate-degrading activity. The recombinant alginate lyase was efficiently immobilized onto two types of magnetic nanoparticles, superparamagnetic iron oxide nanoparticle, and hybrid magnetic silica nanoparticle, based on the affinity between His-tag and Ni(2+) that displayed on the surfaces of nanoparticles. An alginate oligosaccharide mixture consisting of dimer and trimer was prepared by the immobilized alginate lyase. The immobilized enzymes were re-used repeatedly more than 10 times after magnetic separation.


Assuntos
Enzimas Imobilizadas/metabolismo , Escherichia coli/enzimologia , Polissacarídeo-Liases/biossíntese , Proteínas Recombinantes de Fusão/biossíntese , Streptomyces/enzimologia , Alginatos/metabolismo , Clonagem Molecular , Enzimas Imobilizadas/química , Escherichia coli/genética , Magnetismo , Nanopartículas/química , Polissacarídeo-Liases/genética , Polissacarídeo-Liases/isolamento & purificação , Proteínas Recombinantes de Fusão/isolamento & purificação
17.
Front Cell Dev Biol ; 9: 744878, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34722526

RESUMO

In Trypanosoma brucei, genes are assembled in polycistronic transcription units (PTUs). Boundaries of PTUs are designated transcription start sites and transcription termination sites (TTSs). Messenger RNAs are generated by trans-splicing and polyadenylation of precursor RNAs, and regulatory information in the 3' un-translated region (UTR), rather than promoter activity/sequence-specific transcription factors, controls mRNA levels. Given this peculiar genome structure, special strategies must be utilized to control transcription in T. brucei. TTSs are deposition sites for three non-essential chromatin factors-two of non-canonical histone variants (H3v and H4v) and a DNA modification (base J, which is a hydroxyl-glucosyl dT). This association generated the hypothesis that these three chromatin marks define a transcription termination site in T. brucei. Using a panel of null mutants lacking H3v, H4v, and base J, here I show that H4v is a major sign for transcription termination at TTSs. While having a secondary function at TTSs, H3v is important for monoallelic transcription of telomeric antigen genes. The simultaneous absence of both histone variants leads to proliferation and replication defects, which are exacerbated by the J absence, accompanied by accumulation of sub-G1 population. Thus, I propose that the coordinated actions of H3v, H4v, and J provide compensatory mechanisms for each other in chromatin organization, transcription, replication, and cell-cycle progression.

18.
Artigo em Inglês | MEDLINE | ID: mdl-33494300

RESUMO

We aimed to describe the infectious disease (ID) mortality trends and evaluate age-period-cohort (APC) effects on ID mortality in Korea. Using cause-of-death and census population estimates data from 1983-2017, age-standardized ID mortality trends were investigated by joinpoint regression analysis. The APC effects on ID mortality were estimated using intrinsic estimator models. The age effect showed a J-shaped concave upward curve. Old age, especially ≥70 years, was a critical factor for ID deaths. Similar to the W-shaped period curve, ID mortality rapidly decreased due to economic development and the expansion of health coverage in the 1980s, decelerated with increasing inequality, surged due to the 1997 economic crisis, and has gradually increased since the mid-2000s. The cohort effect showed an inverted U-shape. The increasing cohort effect due to the deterioration of living standards led to a decreasing trend after the independence of Korea. Notwithstanding the slowdown during the 1950-1953 Korean War, educational expansion, economic growth, fertility reduction, and the improvement of ID-related policies might have led to a continued decline among the cohorts born since the 1960s. Diverse socioeconomic events may have influenced ID mortality trends in Korea via period and cohort effects. Policies to reduce the growing burden of ID deaths should be further improved.


Assuntos
Doenças Transmissíveis , Idoso , Efeito de Coortes , Estudos de Coortes , Humanos , Mortalidade , República da Coreia/epidemiologia , Fatores Socioeconômicos
19.
Biotechnol Lett ; 32(11): 1685-91, 2010 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-20574831

RESUMO

A triple-point mutated fish microsomal epoxide hydrolase (mEH) gene from Mugil cephalus was expressed in Escherichia coli in the presence of various chaperones to prevent protein aggregations. The enantioselective hydrolytic activity was more than doubled by co-expressing the EH mutant gene with pGro7 plasmid. The highly active EH mutant with a his-tag was immobilized onto magnetic silica assembled with NiO nanoparticles. The immobilized mEH mutant was re-used more than 10 times with less than 10% activity loss. (S)-Styrene oxide with 98% enantiopurity was repeatedly obtained with over 50% of the theoretical yield by the magnetically separable high-performance mEH mutant.


Assuntos
Epóxido Hidrolases/genética , Epóxido Hidrolases/metabolismo , Compostos de Epóxi/metabolismo , Expressão Gênica , Nanopartículas de Magnetita , Microssomos/enzimologia , Smegmamorpha , Animais , Cromatografia de Afinidade , Enzimas Imobilizadas/metabolismo , Epóxido Hidrolases/isolamento & purificação , Escherichia coli/genética , Hidrólise , Proteínas Mutantes/genética , Proteínas Mutantes/isolamento & purificação , Proteínas Mutantes/metabolismo , Plasmídeos , Mutação Puntual , Ligação Proteica , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/isolamento & purificação , Proteínas Recombinantes de Fusão/metabolismo
20.
J Korean Acad Nurs ; 50(2): 200-209, 2020 Apr.
Artigo em Coreano | MEDLINE | ID: mdl-32376809

RESUMO

PURPOSE: The purpose of this study was to examine the effects of oral gargling with an aromatic solution on xerostomia, objective oral status, and oral health-related quality of life in psychiatric inpatients. METHODS: A nonequivalent control group with a non-synchronized design was used in this study. The experimental group (n=34) received oral gargling with an aroma solution, while the control group (n=33) gargled with 0.9% normal saline. Dependent variables were measured at pre-, post-, and follow-up test. Data were analyzed using an χ²-test, Fisher's exact probability test, t-tests, and repeated measures ANOVA using SPSS/WIN v.21.0. RESULTS: After the intervention, significant differences were revealed in xerostomia (F=15.30, p <.001), objective oral status (F=38.44, p <.001), and oral health-related quality of life (F=62.70, p <.001) with an interaction effect between group and time. CONCLUSION: These findings indicate that gargling with an aroma solution is more effective than 0.9% normal saline for the oral health of psychiatric inpatients. Therefore gargling with an aroma can be safely recommended as a brief, economical, and positive intervention in clinical settings.


Assuntos
Aromaterapia , Transtornos Mentais/psicologia , Antissépticos Bucais , Adulto , Feminino , Humanos , Masculino , Transtornos Mentais/patologia , Pessoa de Meia-Idade , Antissépticos Bucais/química , Antissépticos Bucais/uso terapêutico , Saúde Bucal , Qualidade de Vida , Inquéritos e Questionários , Xerostomia/tratamento farmacológico , Xerostomia/patologia , Adulto Jovem
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA