RESUMO
Dengue virus (DENV) and its four serotypes (DENV1-4) belong to the Flavivirus genus of the Flaviviridae family. DENV infection is a life-threatening disease, which results in up to 20,000 deaths each year. Viruses have been shown to encode trans-regulatory small RNAs, or microRNAs (miRNAs), which bind to messenger RNA and negatively regulate host or viral gene expression. During DENV infections, miRNAs interact with proteins in the RNAi pathway, and are processed by ribonucleases such as Dicer and Drosha. This study aims to investigate Drosha, DGCR8, and Dicer expression levels in human A-549 cells following DENV4 infection. DENV4 infected A-549 cells were collected daily for 5 days, and RNA was extracted to quantify viral load. Gene expression of Drosha, Dicer, and DGCR8 was determined using quantitative PCR (RT-qPCR). We found that DENV4 infection exhibited the highest viral load 3 days post-infection. Dicer, Drosha, and DGCR8 showed reduced expression following DENV4 infection as compared with negative controls. In addition, we hypothesize that reduced expression of DGCR8 may not only be related to miRNA biogenesis, but also other small RNAs. This study may change our understanding regarding the relationship between host cells and the dengue virus.
Assuntos
RNA Helicases DEAD-box/biossíntese , Vírus da Dengue/genética , Vírus da Dengue/patogenicidade , Dengue/metabolismo , RNA Mensageiro/biossíntese , Proteínas de Ligação a RNA/biossíntese , Ribonuclease III/biossíntese , Células A549 , RNA Helicases DEAD-box/genética , RNA Helicases DEAD-box/metabolismo , Dengue/genética , Dengue/virologia , Regulação para Baixo , Regulação da Expressão Gênica , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Interferência de RNA , RNA Mensageiro/genética , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Ribonuclease III/genética , Ribonuclease III/metabolismo , Carga ViralRESUMO
Aborda à luz do debate atual sobre o poder local, o processo de descentralizaçäo dos serviços de saúde no Brasil. Trata essa questäo central tendo por eixo a implementaçäo da Lei do SUS em geral, e no particular as questöes de financiamento e de desenvolvimento de recursos humanos. Conclui, apresentando alternativas no sentido de levar a uma maior autonomia local
Assuntos
Cidades , Política , Legislação como Assunto , Política de Saúde , Sistemas de Saúde/organização & administraçãoRESUMO
This article discusses about a recent procedure in health care studies, the comparison as a methodology of analysis. The different analytical currents refer to a particular method of understanding health-disease process. They are: functionalism, the historical-materialism and the new currents. Their phylosophical and sociological basis, concepts, analysis instruments and purposes are showed here by a review of the principal works from representative authors as Navarro, Terris, Roemer, Fry, Illich, Capra and others. The paper suggests that comparative analysis can take two directions: the first is a operational approach for analysing the concrete situations of health's service organization, the second, a more conceptual one, aimed at identifying critical questions and international tendencies in health's systems. The recent discussion search for the overcoming of these dichotomies toward the progress of the production of knowledge and its effects in health's services organization.
RESUMO
A rat brain extract, able to synthesize from UDP-Glc an alpha-1,4-glucan covalently bound to a protein in the absence of added primer is described. The compound formed is precipitable by dilute trichloroacetic acid (TCA). In the presence of glycogen, added as primer, this molecule is enlarged and is not precipitable by TCA. Unprimed and primed activities differ in several aspects, such as the behavior in the presence of some effectors, and the optimum pH. Umprimed and primed activities presented two pHs optima, both sharing only one. The proteoglucans synthesized under the different pHs gave different patterns after analysis under denaturing PAGE and the oligosaccharides synthesized on the protein backbone differ in the glucosyl length. It is concluded that also in rat brain, the initiation process of glycogen biosynthesis is mediated through the formation of a glycoprotein. Our present results showed that the step of the putative "Glycogen Initiator" proposed by use before, requires two enzymes UDPGlc-transglucosylating activities, Glycogen Initiator 1 and Glycogen Initiator 2, before Glycogen Synthase in the alpha-1,4-glucosidic linkages formation.
Assuntos
Encéfalo/enzimologia , Glicogênio Sintase/metabolismo , Glicogênio/biossíntese , Animais , Cromatografia Líquida de Alta Pressão , Glucose/metabolismo , Glicoproteínas/metabolismo , Concentração de Íons de Hidrogênio , Manganês/metabolismo , Proteoglicanas/metabolismo , Ratos , Ratos Endogâmicos , Uridina Trifosfato/metabolismoRESUMO
The effect of ethanol and tunicamycin on synthesis and secretion of galactose oxidase was studied in resting cells of Dactylium dendroides. Ethanol promoted an overall decrease in both intra- and extracellular enzyme levels to the same extent that it inhibited [14C]glucosamine incorporation into total protein. The carbohydrate content of the intracellular enzyme was also depressed (44%) with a simultaneous decrease in O-Ser linked oligosaccharides. The intracellular galactose oxidase obtained after exposure of mycelia to ethanol plus tunicamycin lost 86% of its carbohydrate moieties, whereas the extracellular form lost only 35%. In both cases, residual sugar moieties were not eliminated by mild alkaline treatment. These data suggest that ethanol affects O-glycosylation of galactose oxidase. O-Underglycosylation did not affect the S0.5 values for galactose but diminished the molar catalytic activity. The absence of O-Ser/Thr-linked saccharides turned the intracellular enzyme into a form more susceptible to proteolysis than that devoid of N-linked sugars (tunicamycin-treated). O-Underglycosylation had a significant effect on the renaturation-reactivation of the enzyme after denaturation with 2.4 M Gdn-HCl.
Assuntos
Basidiomycota/enzimologia , Etanol/farmacologia , Galactose Oxidase/biossíntese , Glucosamina/metabolismo , Polyporaceae/enzimologia , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Galactose Oxidase/genética , Glicosilação , Cinética , Leucina/metabolismo , Polyporaceae/efeitos dos fármacos , Polyporaceae/crescimento & desenvolvimento , Tunicamicina/farmacologiaRESUMO
The stability of intracellular, extracellular, and deglycosylated forms of galactose oxidase was compared with respect to the denaturing effects of heat, pH, and guanidine hydrochloride. The highly glycosylated forms were found to be more stable to pH and thermal inactivation. All forms were reversibly denaturated by guanidine hydrochoride, but the extent was dependent on the carbohydrate content. Deglycosylation did not affect the affinity of the enzyme for dihydroxyacetone and galactose. Exposure of different forms of galactose oxidase to proteases like pronase and trypsin resulted in a rapid degradation of the glycoenzymes with the formation of stable products. After pronase digestion of intra- and extracellular forms of galactose oxidase catalytic species were isolated by gel filtration. The species (61 and 42 kDa) isolated from pronase-digested extracellular enzyme lost their ability to oxidize primary alcohols. Species (67 and 46 kDa) obtained from the intracellular enzyme kept the specificity of the original enzyme. Active pronase-derived peptides (42 and 46 kDa, respectively) had a higher carbohydrate content than the inactive ones.