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1.
J Mech Behav Biomed Mater ; 117: 104392, 2021 Feb 12.
Artigo em Inglês | MEDLINE | ID: mdl-33601300

RESUMO

Stabilization of the resin-dentin interface to increase the durability of adhesive dental restorations is a challenging task. The use of naturally occurring collagen crosslinking agents has been proposed to prevent degradation of the hybrid layer. Myricetin (MYR) is a flavonoid with a wide variety of beneficial effects and it has been used for the treatment of different systemic pathologies. The chemical structure of MYR makes it a powerful antioxidant, an inhibitor of matrix metalloproteinase (MMP) activity, and a collagen cross-linker. This study presents MYR as a novel treatment in operative dentistry to stabilize the resin-dentin interface by inhibiting MMPs and crosslinking the collagen. Viability tests carried out using a resazurin assay showed that MYR had no cytotoxic effects on human odontoblast-like cells and the phenotype was preserved. Fluorometric MMP activity assay and fluorescence microscopy revealed that the MMPs in the demineralized dentin were effectively inhibited by the application of MYR (600 µM for 120 s). A microtensile bond strength test was performed immediately and after six months of storage. The bond strength to dentin was not affected by MYR and was preserved over time. Demineralized dentin beams were evaluated to determine the dentin biomodification using microtensile strength and elastic modulus assays. MYR improved the biomechanical behavior of the demineralized dentin similarly to glutaraldehyde, a recognized crosslinking agent. These findings indicated that MYR acts as an MMP inhibitor, collagen cross-linker, and preserver of the bond strength. Furthermore, MYR is an ethanol-soluble molecule with a lower molecular weight than the other polyphenols; hence, it can be applied for a short time and diffuses deeply through the dentin without any associated cytotoxicity. This molecule has beneficial effects on the biological and mechanical behavior of the resin-dentin interface and may be used to effectively stabilize the hybrid layer in a clinical setting.

2.
Infectio ; 24(3,supl.1): 5-10, oct.-dic. 2020. tab, graf
Artigo em Espanhol | LILACS-Express | LILACS | ID: biblio-1143093

RESUMO

Resumen Se formulan recomendaciones de un grupo de consenso de expertos sobre los criterios para evaluar el desempeño diagnóstico (tamaño y selección de muestras para sensibilidad y especificidad analíticas, criterios para establecer límites de detección, criterios para establecer el estándar de oro para las serologías) que deberían ser tenidos en cuenta al evaluar y validar las pruebas diagnósticas para SARS CoV-2. Con el propósito de asegurar la calidad de las pruebas serológicas a utilizar en el país, se recomienda la participación en un programa de control de calidad externo, que garantice la idoneidad y desempeño en la realización de las pruebas diagnósticas serológicas y moleculares durante esta pandemia, ya que su uso tiene profundas implicaciones para las medidas de intervención clínicas individuales y de seguimiento y control en salud pública.


Abstract We formulate recommendations from a consensus working group on the criteria to evaluate the diagnostic performance (size and criteria of selection of samples to determine sensitivity, analytical specificity, criteria for limit of detection, criteria for gold standard to evaluate serological assays) that should be taken into account during the evaluation and validation/verification of diagnostic tests for SARS CoV-2 infection. A national external quality control program should be established to guarantee the suitability and performance of these diagnostic serological and molecular tests during this pandemic, that will have deep implications for decisions on clinical and public health.

3.
Chem Biol Interact ; 333: 109336, 2020 Nov 25.
Artigo em Inglês | MEDLINE | ID: mdl-33248029

RESUMO

Resin-based dental materials consist of filler particles and different monomers that are light cured in situ to re-establish dental function and aesthetics. Due to the degree of conversion of adhesive polymers, the monomers triethyleneglycol dimethacrylate (TEGDMA) and 2-hydroxyethyl methacrylate (HEMA) are released in relatively high amounts and are susceptible to degradation, acting as bioactive compounds and affecting cell and tissues. This study aimed to assess the effect of HEMA and TEGDMA exposure on metabolic activity, membrane integrity, and cell survival of human odontoblast-like cell (hOLCs). Exposure to resin monomers for 24 h induced major changes in cell membrane integrity, metabolic activity, and survival, which were measured by the calcein method and lactate dehydrogenase release. Increased and early reactive oxygen species (ROS) production was observed leading to degradative oxidation of membrane lipids identified as malondialdehyde production. Severe alteration in mitochondria occurred due to transmembrane mitochondrial potential collapse, possibly inducing activation of apoptotic cell death. hOLCs exposure to resin monomers modified the cell redox potential, with consequences on membrane permeability and integrity, including mitochondrial function. Lipid peroxidation appears to be a key phenomenon for the membrane structures oxidation after HEMA and TEGDMA exposure, leading to cell death and cytotoxicity. hOLCs respond early by differential induction of adaptive mechanisms to maintain cell homeostasis. Modulation of oxidative stress-induced response involves the regulation of genes that encode for antioxidant proteins such as catalase and heme oxygenase-1; regulation that functions as a critical protection mechanism against oxidative cell damage induced by HEMA and TEGDMA. Ascorbic acid as an antioxidant substance mitigates the oxidative damage associated with exposure to monomers.

4.
Int J Infect Dis ; 102: 97-102, 2020 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-33075526

RESUMO

OBJECTIVE: To identify the arbovirus involved in febrile cases identified in a pediatric clinic in Cali, Valle del Cauca province, Colombia, and study the clinical characteristics. METHODS: A descriptive, prospective study enrolled 345 febrile children for 12 months in a pediatric clinic. Serum samples and medical record registers documenting signs and symptoms were analyzed to detect DENV, CHIKV, and ZIKV by reverse transcription-polymerase chain reaction and serology methods. Diagnosis at the time of admission and discharge were compared based on laboratory test results. RESULTS: All patients were diagnosed as severe dengue at admission. Molecular detection and serology tests identified 143 CHIKV-positive (41.4%), 20 DENV-positive (5.8%), and 123 DENV-CHIKV coinfection patients (35.7%). DENV or CHIKV serology test results of these double-infected patients yielded poor performance to confirm patient cases. ZIKV infection was detected in 5 patients (1.4%) as double or triple infections. CONCLUSION: A sustained CHIKV circulation and transmission was confirmed causing febrile illness in children and indicating that this virus spreads even during the regular DENV season, leading to double infections and altering clinical symptoms. Specific clinical tests are necessary to closely identify the arbovirus involved in causing infectious diseases that can help in better treatment and mosquito-transmitted virus surveillance.

7.
Int J Dent ; 2020: 8860840, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32676112

RESUMO

Introduction: The aim of the present study was to determine the effects of vitamins D and E on the proliferation, morphology, and differentiation of human dental pulp stem cells (hDPSCs). Methods: In this in vitro experimental study, hDPSCs were isolated, characterized, and treated with vitamins D and E, individually and in combination, utilizing different doses and treatment periods. Changes in morphology and cell proliferation were evaluated using light microscopy and the resazurin assay, respectively. Osteoblast differentiation was evaluated with alizarin red S staining and expression of RUNX2, Osterix, and Osteocalcin genes using real-time RT-PCR. Results: Compared with untreated cells, the number of cells significantly reduced following treatment with vitamin D (49%), vitamin E (35%), and vitamins D + E (61%) after 144 h. Compared with cell cultures treated with individual vitamins, cells treated with vitamins D + E demonstrated decreased cell confluence, with more extensive and flatter cytoplasm that initiated the formation of a significantly large number of calcified nodules after 7 days of treatment. After 14 days, treatment with vitamins D, E, and D + E increased the transcription of RUNX2, Osterix, and Osteocalcin genes. Conclusions: Vitamins D and E induced osteoblastic differentiation of hDPSCs, as evidenced by the decrease in cell proliferation, morphological changes, and the formation of calcified nodules, increasing the expression of differentiation genes. Concurrent treatment with vitamins D + E induces a synergistic effect in differentiation toward an osteoblastic lineage.

8.
Braz. j. infect. dis ; 24(1): 13-24, Feb. 2020. tab, graf
Artigo em Inglês | LILACS | ID: biblio-1089334

RESUMO

ABSTRACT Dengue has been a significant public health problem in Colombia since the simultaneous circulation of the four dengue virus serotypes. The replicative fitness of dengue is a biological feature important for virus evolution and contributes to elucidating the behavior of virus populations and viral pathogenesis. However, it has not yet been studied in Colombian isolates. This study aimed to compare the replicative fitness of the four dengue virus serotypes and understand the association between the serotypes, their in vitro infection ability, and their replication in target cells. We used three isolates of each DENV serotype to infect Huh-7 cells at an MOI of 0.5. The percentage of infected cells was evaluated by flow cytometry, cell viability was evaluated by MTT assay, and the pathogenicity index was calculated as a ratio of both parameters. The replicative fitness was measured by the number of viral genome copies produced using quantitative PCR and the production of infectious viral progeny was measured by plaque assay. We showed that Huh-7 cells were susceptible to infection with all the different strain isolates. Nevertheless, the biological characteristics, such as infectious ability and cell viability, were strain-dependent. We also found different degrees of pathogenicity between strains of the four serotypes, representative of the heterogeneity displayed in the circulating population. When we analyzed the replicative fitness using the mean values obtained from RT-qPCR and plaque assay for the different strains, we found serotype-dependent behavior. The highest mean values of replicative fitness were obtained for DENV-1 (log 4.9 PFU/ml) and DENV-4 (log 5.28 PFU/ml), followed by DENV-2 (log 3.9 PFU/ml) and DENV-3 (log 4.31 PFU/ml). The internal heterogeneity of the replicative fitness within each serotype could explain the simultaneous circulation of the four DENV serotypes in Colombia.


Assuntos
Humanos , Replicação Viral/genética , Vírus da Dengue/genética , Vírus da Dengue/patogenicidade , Sorogrupo , Ensaio de Placa Viral , Valores de Referência , Sais de Tetrazólio , Fatores de Tempo , RNA Viral/genética , Linhagem Celular , Sobrevivência Celular , Células Cultivadas , Colômbia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Citometria de Fluxo , Formazans , Fígado/citologia
9.
PLoS One ; 15(1): e0227030, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31910224

RESUMO

Endothelial activation and alteration during dengue virus (DENV) infection are multifactorial events; however, the role of extracellular vesicles (EVs) in these phenomena is not known. In the present study, we characterized the EVs released by DENV-2 infected U937 macrophage cell line and evaluated the changes in the physiology and integrity of the EA.hy926 endothelial cells exposed to them. U937 macrophages were infected, supernatants were collected, and EVs were purified and characterized. Then, polarized endothelial EA.hy926 cells were exposed to the EVs for 24 h, and the transendothelial electrical resistance (TEER), monolayer permeability, and the expression of tight junction and adhesion proteins and cytokines were evaluated. The isolated EVs from infected macrophages corresponded to exosomes and apoptotic bodies, which contained the viral NS3 protein and different miRs, among other products. Exposure of EA.hy926 cells to EVs induced an increase in TEER, as well as changes in the expression of VE-cadherin and ICAM in addition leads to an increase in TNF-α, IP-10, IL-10, RANTES, and MCP-1 secretion. These results suggest that the EVs of infected macrophages transport proteins and miR that induce early changes in the physiology of the endothelium, leading to its activation and eliciting a defense program against damage during first stages of the disease, even in the absence of the virus.


Assuntos
Vírus da Dengue , Células Endoteliais/metabolismo , Vesículas Extracelulares/virologia , Macrófagos/ultraestrutura , Antígenos CD/metabolismo , Caderinas/metabolismo , Moléculas de Adesão Celular/metabolismo , Citocinas/metabolismo , Dengue/imunologia , Vírus da Dengue/imunologia , Células Endoteliais/imunologia , Vesículas Extracelulares/fisiologia , Humanos , Macrófagos/virologia , Permeabilidade , Células U937
10.
Vaccines (Basel) ; 8(1)2020 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-31979145

RESUMO

Dengue virus (DENV) is an arbovirus of the Flaviviridae family and is an enveloped virion containing a positive sense single-stranded RNA genome. DENV causes dengue fever (DF) which is characterized by an undifferentiated syndrome accompanied by fever, fatigue, dizziness, muscle aches, and in severe cases, patients can deteriorate and develop life-threatening vascular leakage, bleeding, and multi-organ failure. DF is the most prevalent mosquito-borne disease affecting more than 390 million people per year with a mortality rate close to 1% in the general population but especially high among children. There is no specific treatment and there is only one licensed vaccine with restricted application. Clinical and experimental evidence advocate the role of the humoral and T-cell responses in protection against DF, as well as a role in the disease pathogenesis. A lot of pro-inflammatory factors induced during the infectious process are involved in increased severity in dengue disease. The advances in DF research have been hampered by the lack of an animal model that recreates all the characteristics of this disease. Experiments in nonhuman primates (NHP) had failed to reproduce all clinical signs of DF disease and during the past decade, humanized mouse models have demonstrated several benefits in the study of viral diseases affecting humans. In DENV studies, some of these models recapitulate specific signs of disease that are useful to test drugs or vaccine candidates. However, there is still a need for a more complete model mimicking the full spectrum of DENV. This review focuses on describing the advances in this area of research.

11.
Braz J Infect Dis ; 24(1): 13-24, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31843340

RESUMO

Dengue has been a significant public health problem in Colombia since the simultaneous circulation of the four dengue virus serotypes. The replicative fitness of dengue is a biological feature important for virus evolution and contributes to elucidating the behavior of virus populations and viral pathogenesis. However, it has not yet been studied in Colombian isolates. This study aimed to compare the replicative fitness of the four dengue virus serotypes and understand the association between the serotypes, their in vitro infection ability, and their replication in target cells. We used three isolates of each DENV serotype to infect Huh-7 cells at an MOI of 0.5. The percentage of infected cells was evaluated by flow cytometry, cell viability was evaluated by MTT assay, and the pathogenicity index was calculated as a ratio of both parameters. The replicative fitness was measured by the number of viral genome copies produced using quantitative PCR and the production of infectious viral progeny was measured by plaque assay. We showed that Huh-7 cells were susceptible to infection with all the different strain isolates. Nevertheless, the biological characteristics, such as infectious ability and cell viability, were strain-dependent. We also found different degrees of pathogenicity between strains of the four serotypes, representative of the heterogeneity displayed in the circulating population. When we analyzed the replicative fitness using the mean values obtained from RT-qPCR and plaque assay for the different strains, we found serotype-dependent behavior. The highest mean values of replicative fitness were obtained for DENV-1 (log 4.9 PFU/ml) and DENV-4 (log 5.28 PFU/ml), followed by DENV-2 (log 3.9 PFU/ml) and DENV-3 (log 4.31 PFU/ml). The internal heterogeneity of the replicative fitness within each serotype could explain the simultaneous circulation of the four DENV serotypes in Colombia.


Assuntos
Vírus da Dengue/genética , Vírus da Dengue/patogenicidade , Sorogrupo , Replicação Viral/genética , Linhagem Celular , Sobrevivência Celular , Células Cultivadas , Colômbia , Citometria de Fluxo , Formazans , Humanos , Fígado/citologia , RNA Viral/genética , Valores de Referência , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sais de Tetrazólio , Fatores de Tempo , Ensaio de Placa Viral
12.
Int J Infect Dis ; 91: 9-16, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31733358

RESUMO

OBJECTIVES: Colombia is a dengue hyperendemic country; however, the prevalence of antibodies against dengue in the general population including the inhabitants of rural areas is unknown. This study aimed to determine the prevalence of dengue IgM and IgG antibodies in healthy children and adults in urban and rural areas of seven different endemic regions in Colombia between 2013 and 2015. DESIGN OR METHOD: Blood samples from healthy volunteers (1,318) were processed by serology (by indirect IgG and capture IgM and IgG ELISA) and molecular tests to detect viral RNA and circulating serotypes. RESULTS: The seroprevalence of IgG for dengue were 85% in children and over 90% for adults. In addition to the high IgM positive rate (14.9%) and secondary recent infection marker rate (capture IgG, 16%), 8.4% of the healthy volunteers were positive for dengue virus (DENV) RNA. CONCLUSION: This study confirmed the broad and permanent circulation of DENV in Colombia and the high rates of infection and reinfection suffered by its inhabitants. This information can be used by the health authorities to strengthen vector control and vaccine policies and review the algorithms of diagnosis and disease management in children and adults.


Assuntos
Anticorpos Antivirais/sangue , Vírus da Dengue/imunologia , Dengue/epidemiologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Coinfecção , Colômbia/epidemiologia , Dengue/imunologia , Vírus da Dengue/genética , Doenças Endêmicas , Feminino , Humanos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Masculino , Pessoa de Meia-Idade , RNA Viral/imunologia , Estudos Soroepidemiológicos , Sorogrupo , Adulto Jovem
13.
Front Microbiol ; 10: 1435, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31293558

RESUMO

More than 500 million people worldwide are infected each year by any of the four-dengue virus (DENV) serotypes. The clinical spectrum caused during these infections is wide and some patients may develop neurological alterations during or after the infection, which could be explained by the cryptic neurotropic and neurovirulent features of flaviviruses like DENV. Using in vivo and in vitro models, researchers have demonstrated that DENV can affect the cells from the blood-brain barrier (BBB) in several ways, which could result in brain tissue damage, neuronal loss, glial activation, tissue inflammation and hemorrhages. The latter suggests that BBB may be compromised during infection; however, it is not clear whether the damage is due to the infection per se or to the local and/or systemic inflammatory response established or activated by the BBB cells. Similarly, the kinetics and cascade of events that trigger tissue damage, and the cells that initiate it, are unknown. This review presents evidence of the BBB cell infection with DENV and the response established toward it by these cells; it also describes the consequences of this response on the nervous tissue, compares these evidence with the one reported with neurotropic viruses of the Flaviviridae family, and shows the complexity and unpredictability of dengue and the neurological alterations induced by it. Clinical evidence and in vitro and in vivo models suggest that this virus uses the bloodstream to enter nerve tissue where it infects the different cells of the neurovascular unit. Each of the cell populations respond individually and collectively and control infection and inflammation, in other cases this response exacerbates the damage leaving irreversible sequelae or causing death. This information will allow us to understand more about the complex disease known as dengue, and its impact on a specialized and delicate tissue like is the nervous tissue.

14.
BMC Microbiol ; 19(1): 45, 2019 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-30786858

RESUMO

BACKGROUND: Pseudomonas aeruginosa Sequence Type 235 is a clone that possesses an extraordinary ability to acquire mobile genetic elements and has been associated with the spread of resistance genes, including genes that encode for carbapenemases. Here, we aim to characterize the genetic platforms involved in resistance dissemination in blaKPC-2-positive P. aeruginosa ST235 in Colombia. RESULTS: In a prospective surveillance study of infections in adult patients attended in five ICUs in five distant cities in Colombia, 58 isolates of P. aeruginosa were recovered, of which, 27 (46.6%) were resistant to carbapenems. The molecular analysis showed that 6 (22.2%) and 4 (14.8%) isolates harboured the blaVIM and blaKPC-2 genes, respectively. The four blaKPC-2-positive isolates showed a similar PFGE pulsotype and belonged to ST235. Complete genome sequencing of a representative ST235 isolate shows a unique chromosomal contig of 7097.241 bp with eight different resistance genes identified and five transposons: a Tn6162-like with ant(2″)-Ia, two Tn402-like with ant(3″)-Ia and blaOXA-2 and two Tn4401b with blaKPC-2. All transposons were inserted into the genomic islands. Interestingly, the two Tn4401b copies harbouring blaKPC-2 were adjacently inserted into a new genomic island (PAGI-17) with traces of a replicative transposition process. This double insertion was probably driven by several structural changes within the chromosomal region containing PAGI-17 in the ST235 background. CONCLUSION: This is the first report of a double Tn4401b chromosomal insertion in P. aeruginosa, just within a new genomic island (PAGI-17). This finding indicates once again the great genomic plasticity of this microorganism.


Assuntos
Cromossomos Bacterianos , Elementos de DNA Transponíveis , Farmacorresistência Bacteriana Múltipla/genética , Genoma Bacteriano , Ilhas Genômicas , Pseudomonas aeruginosa/genética , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Técnicas de Tipagem Bacteriana , Carbapenêmicos/farmacologia , Colômbia , DNA Bacteriano/genética , Humanos , Unidades de Terapia Intensiva/estatística & dados numéricos , Testes de Sensibilidade Microbiana , Tipagem de Sequências Multilocus , Estudos Prospectivos , Pseudomonas aeruginosa/enzimologia , Sequenciamento Completo do Genoma , beta-Lactamases/genética
15.
Am J Trop Med Hyg ; 99(6): 1422-1429, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30398143

RESUMO

Dengue is the most important arbovirosis in the world. In this study, we assessed the knowledge, attitudes, and practices (KAP) regarding dengue in parents from two small Colombian municipalities in the Cundinamarca Province. Parents and their healthy children from 4 to 14 years of age were included in some public elementary schools. After a medical examination, blood samples were taken for diagnosis of dengue using enzyme-linked immunosorbent assays (capture immunoglobulin M and capture immunoglobulin G [IgG], indirect IgG and detection non-structural viral protein 1) and detection of viral RNA by reverse transcription polymerase chain reaction. In addition, a KAP survey was applied to the children's parents or tutors. The indirect IgG test determined that of the 347 examined children, 87.9% had a previous infection with the dengue virus (DENV), 12.7% of them were positive for viral RNA (asymptomatic infection), and 32.0% presented reinfections. Risk factors evaluation showed that children aged 8 years and older living in the municipalities for more than 7 years were more likely to be infected or reinfected by DENV. In the same way, poor nutrition, lack of water supply, sewer service, or waste disposal services could raise the likelihood of dengue infections. The surveys indicated that parents have unhealthy practices and a low knowledge about the transmission of the disease, which could result in an increase of mosquito breeding sites, allowing sustained dengue transmission.


Assuntos
Aedes/virologia , Infecções Assintomáticas/epidemiologia , Vírus da Dengue/isolamento & purificação , Dengue/epidemiologia , Conhecimentos, Atitudes e Prática em Saúde , Desnutrição/epidemiologia , Mosquitos Vetores/virologia , Adolescente , Animais , Anticorpos Antivirais/sangue , Criança , Pré-Escolar , Cidades , Colômbia/epidemiologia , Dengue/transmissão , Dengue/virologia , Vírus da Dengue/genética , Vírus da Dengue/imunologia , Água Potável/análise , Feminino , Humanos , Imunoglobulina G/sangue , Masculino , Desnutrição/virologia , Pais/educação , Pais/psicologia , RNA Viral/sangue , Recidiva , Fatores de Risco , Saneamento , Proteínas não Estruturais Virais/sangue , Proteínas não Estruturais Virais/imunologia , Qualidade da Água
16.
Biomedica ; 38(0): 135-143, 2018 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-30184368

RESUMO

INTRODUCTION: Dengue virus replication has been considered mainly cytoplasmic, however, studies indicate that some flaviviruses may use the intranuclear pathway as part of the machinery that the virus uses to increase infection capacity in the host cell. This paper describes alterations at nuclear level in the cell infected with dengue, which are likely involved in the virus replication processes. OBJECTIVE: This paper addresses the ultrastructural observations of C6/36 cells of the Aedes albopictus mosquito infected with dengue virus type 2. MATERIALS AND METHODS: C6/36 cells were infected in culture medium with the serum of a patient positively diagnosed for dengue 2. Subsequently, the cells were incubated for 10 days and the cytopathic effect was assessed. The cells were processed for immunofluorescence assays and transmission electron microscopy. RESULTS: The immunofluorescence assays confirmed the presence of viral protein E associated with cellular syncytia in the culture. In the ultrastructural study, the infected cells showed vesicular-tubular structures and dilated cisterns of the endoplasmic reticulum at the cytoplasmic level. Viral particles were found exclusively in cytoplasm localized within the vacuoles. Nuclei of cellular syncytia showed membrane structures arranged in a circular shape and, in some cases, these syncytia displayed lysis; in no case viral particles were observed at the nuclear level. CONCLUSIONS: The ultrastructural alterations of nuclei in cells infected with the dengue virus using electron microscopy techniques had not been reported before, as far as we know. It is likely that such modifications are associated with replicative processes at an intranuclear level as an alternate replication mechanism.


Assuntos
Núcleo Celular/ultraestrutura , Efeito Citopatogênico Viral , Vírus da Dengue/fisiologia , Aedes/citologia , Animais , Linhagem Celular , Citoplasma/virologia , Dengue/virologia , Vírus da Dengue/isolamento & purificação , Células Gigantes/virologia , Humanos , Microscopia Eletrônica , Microscopia de Fluorescência , Vacúolos/virologia , Proteínas do Envelope Viral/análise , Viremia/virologia , Replicação Viral
17.
Viruses ; 10(9)2018 09 07.
Artigo em Inglês | MEDLINE | ID: mdl-30205518

RESUMO

The high levels of dengue-virus (DENV) seroprevalence in areas where the Zika virus (ZIKV) is circulating and the cross-reactivity between these two viruses have raised concerns on the risk of increased ZIKV disease severity for patients with a history of previous DENV infections. To determine the role of DENV preimmunity in ZIKV infection, we analyzed the T- and B-cell responses against ZIKV in donors with or without previous DENV infection. Using peripheral blood mononuclear cells (PBMCs) from donors living in an endemic area in Colombia, we have identified, by interferon (IFN)-γ enzyme-linked immunospot (ELISPOT) assay, most of the immunodominant ZIKV T-cell epitopes in the nonstructural (NS) proteins NS1, NS3, and NS5. Analyses of the T- and B-cell responses in the same donors revealed a stronger T-cell response against peptides conserved between DENV and ZIKV, with a higher level of ZIKV-neutralizing antibodies in DENV-immune donors in comparison with DENV-naïve donors. Strikingly, the potential for antibody-mediated enhancement of ZIKV infection was reduced in donors with sequential DENV and ZIKV infection in comparison with donors with DENV infection only. Altogether, these data suggest that individuals with DENV immunity present improved immune responses against ZIKV.


Assuntos
Imunidade Adaptativa , Dengue/imunologia , Infecção por Zika virus/imunologia , Zika virus/imunologia , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Linfócitos B/imunologia , Colômbia , Humanos , Interferon gama/metabolismo , Leucócitos Mononucleares/imunologia , Linfócitos T/imunologia
18.
Biomédica (Bogotá) ; 38(supl.2): 135-143, ago. 2018. tab, graf
Artigo em Inglês | LILACS-Express | LILACS | ID: biblio-1038798

RESUMO

ABSTRACT Introduction: Dengue virus replication has been considered mainly cytoplasmic, however, studies indicate that some flaviviruses may use the intranuclear pathway as part of the machinery that the virus uses to increase infection capacity in the host cell. This paper describes alterations at nuclear level in the cell infected with dengue, which are likely involved in the virus replication processes. Objective: This paper addresses the ultrastructural observations of C6/36 cells of the Aedes albopictus mosquito infected with dengue virus type 2. Materials and methods: C6/36 cells were infected in culture medium with the serum of a patient positively diagnosed for dengue 2. Subsequently, the cells were incubated for 10 days and the cytopathic effect was assessed. The cells were processed for immunofluorescence assays and transmission electron microscopy. Results: The immunofluorescence assays confirmed the presence of viral protein E associated with cellular syncytia in the culture. In the ultrastructural study, the infected cells showed vesicular-tubular structures and dilated cisterns of the endoplasmic reticulum at the cytoplasmic level. Viral particles were found exclusively in cytoplasm localized within the vacuoles. Nuclei of cellular syncytia showed membrane structures arranged in a circular shape and, in some cases, these syncytia displayed lysis; in no case viral particles were observed at the nuclear level. Conclusions: The ultrastructural alterations of nuclei in cells infected with the dengue virus using electron microscopy techniques had not been reported before, as far as we know. It is likely that such modifications are associated with replicative processes at an intranuclear level as an alternate replication mechanism.


RESUMEN Introducción. La replicación del virus del dengue se ha considerado principalmente citoplásmica; sin embargo, en diversos estudios se ha informado que algunos flavivirus pueden utilizar factores intranucleares como parte de la maquinaria que utilizan para aumentar la capacidad de infección en la célula huésped. En este trabajo se describen las alteraciones a nivel nuclear en células infectadas con dengue, probablemente involucradas en procesos de replicación viral. Objetivo. Presentar las observaciones ultraestructurales de células C6/36 de Aedes albopictus infectadas con el virus del dengue de tipo 2. Materiales y métodos. Se infectaron células C6/36 con suero de un paciente con diagnóstico de dengue 2; posteriormente, se mantuvieron en medio de cultivo durante 10 días y se evaluó el efecto citopático. Las células se procesaron para los ensayos de inmunofluorescencia y microscopía electrónica de transmisión, con el fin de hacer el estudio ultraestructural. Resultados. Los ensayos de inmunofluorescencia confirmaron la presencia de la proteína E viral asociada con sincitios celulares en el cultivo. En el estudio ultraestructural, las células infectadas tenían estructuras vesiculares y tubulares, y cisternas dilatadas del retículo endoplásmico en el citoplasma. Las partículas virales se encontraron exclusivamente en vacuolas localizadas en el citoplasma. Los núcleos de los sincitios celulares contenían estructuras de membrana dispuestas en forma circular y, en algunos casos, dichos sincitios presentaban lisis. En ningún caso se observaron partículas virales en el núcleo. Conclusiones. No se habían reportado alteraciones ultraestructurales en los núcleos de células infectadas con el virus del dengue detectadas mediante técnicas de microscopia electrónica. Es probable que tales modificaciones estén asociadas con procesos intranucleares de replicación como un mecanismo alternativo.

19.
Diseases ; 6(3)2018 Jul 06.
Artigo em Inglês | MEDLINE | ID: mdl-29986388

RESUMO

Natural infection with dengue virus (DENV) induces an increase in the production of cytokines that play an important role in disease pathogenesis. Despite numerous scientific studies, there are still no commercially available disease-specific therapeutics. Previous evidence shows that inhibiting histone deacetylase enzymes (HDACs) regulates the immune response in several inflammatory disease models. The aim of the current study was to evaluate the effect of HDAC inhibition in the production of inflammatory cytokines in human monocyte-derived macrophages infected with DENV serotype 2 (DENV-2). To this end, human monocyte-derived macrophages (MDMs) were treated with valproic acid (VPA) before or after infection and the inflammatory cytokine concentration was quantified by flow cytometry. We found that infected MDMs secreted IL-8, IL-1b, IL-6, TNF-alpha, and IL-10, but not IL-12. Strikingly, treatment of infected cells with VPA had a differential and concentration-dependent effect on the production of specific cytokines without eliciting significant changes in cell viability. Using the highest concentration of VPA, a significant reduction in the production of all cytokines was observed. These results suggest that HDAC inhibition during DENV-2 infection could exert an important regulatory effect in the production of inflammatory cytokines, representing a significant advance in the design of novel therapeutic dengue treatments.

20.
Plant Pathol J ; 34(3): 171-181, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29887773

RESUMO

Fruit rot is the principal phytopathological problem of pipiana pumpkin (Cucurbita argyrosperma Huber) in the state of Guerrero. The aims of this research were to 1) identify the causal agent of southern blight on pumpkin fruits by morphological, pathogenic, and molecular analysis (ITS1, 5.8S, ITS2); 2) evaluate in vitro Trichoderma spp. strains and chemical fungicides; and 3) evaluate under rainfed field conditions, the strains that obtained the best results in vitro, combined with fungicides during two crop cycles. Number of commercial and non-commercial fruits at harvest, and seed yield (kg ha-1) were registered. Morphological, pathogenic and molecular characterization identified Sclerotium rolfsii as the causal agent of rot in pipiana pumpkin fruits. Now, in vitro conditions, the highest inhibition of S. rolfsii were obtained by Trichoderma virens strain G-41 (70.72%), T. asperellum strain CSAEGro-1 (69%), and the fungicides metalaxyl (100%), pyraclostrobin (100%), quintozene (100%), cyprodinil + fludioxonil (100%), and prochloraz (100%). Thiophanate-methyl only delayed growth (4.17%). In field conditions, during the spring-summer 2015 cycle, T. asperellum strain CSAEGro-1 + metalaxyl, and T. asperellum + cyprodinil + fludioxonil, favored the highest number of fruits and seed yield in the crop.

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