RESUMEN
Dengue virus (DENV) infections may cause life-threatening dengue hemorrhagic fever (DHF). Suppressed protective immunity was shown in these patients. Although several hypotheses have been formulated, the mechanism of DENV-induced immunosuppression remains unclear. Previously, we found that cross-reactive antibodies against tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor 1 (death receptor 4 [DR4]) were elicited in DHF patients, and that anti-DR4 autoantibody fractions were elicited by nonstructural protein 1 (NS1) immunizations in experimental mice. In this study, we found that anti-DR4 antibodies could suppress B lymphocyte function in vitro and in vivo. Treatment with the anti-DR4 immunoglobulin (Ig) induced caspase-dependent cell death in immortalized B lymphocyte Raji cells in vitro. Anti-DR4 Igs elicited by NS1 and DR4 immunizations markedly suppressed mouse spleen transitional T2 B (IgM+IgD+), bone marrow pre-pro-B (B220+CD43+), pre-B (B220+CD43-), and mature B cell (B220+IgD+) subsets in mice. Furthermore, functional analysis revealed that the pre-elicitation of anti-NS1 and anti-DR4 Ig titers suppressed subsequently neutralizing antibody production by immunization with DENV envelop protein. Our data suggest that the elicitation of anti-DR4 titers through DENV NS1 immunization plays a suppressive role in humoral immunity in mice.
Asunto(s)
Anticuerpos Antivirales/inmunología , Inmunidad Humoral , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/inmunología , Dengue Grave/inmunología , Proteínas no Estructurales Virales/inmunología , Animales , Autoanticuerpos/sangre , Células Cultivadas , Virus del Dengue/inmunología , Humanos , Inmunización , Ratones , Ratones Endogámicos C57BLRESUMEN
The accurate examination of paint fragments obtained from an accident, such as those obtained from vehicles involved in a hit-and-run case, is often critical in forensic investigations. However, organic pigments are typically minor components of automotive coatings, which makes discrimination difficult. In this study, direct analysis in real time coupled to Q-orbitrap tandem mass spectrometry (DART-MS) was employed to detect a wide range of common organic pigments in vehicle paints. Twelve common organic pigments used in vehicle paints, such as red, yellow, orange, and purple, were tested, and a database was constructed for future examinations of vehicle paint. Two hit-and-run vehicle accident cases, which occurred in New Taipei City, were investigated by Fourier transform infrared (FTIR) spectroscopy and DART-MS. First, FTIR spectroscopy was employed to study the paint samples as a preliminary screening step. Most of the observed IR peaks were attributed to binder and extenders present in paints. The IR peaks corresponding to the organic pigments were found to be weak and overlapped with those corresponding to resins. On the other hand, DART-MS successfully characterized the organic pigments. DART-MS was found to be excellent for rapidly determining the presence of organic pigments in paint samples without the need for a complicated pre-treatment process or lengthy analysis time.
RESUMEN
Direct analysis in real time coupled to Q-orbitrap tandem mass spectrometry (DART-MS) without requiring preparatory procedures was used to directly detect trace amounts of illegal street drugs, namely p-chloroamphetamine, p-fluoromethamphetamine, γ-hydroxybutyrate, ketamine, methamphetamine, 3,4-methylenedioxypyrovalerone, p-methylethcathinone, methylone, and nimetazepam, in solution and also in real drug samples. Exact mass determination of the drug samples was completed in less than 1min. With the ability to rapidly identify drugs, this technique shows great potential as a useful analytical tool in the analysis of illicit street drugs, and has the significant advantages of simplicity and sensitivity without the sample preparation needed by other methods.
Asunto(s)
Bebidas , Drogas Ilícitas/análisis , Espectrometría de Masas en Tándem/métodos , 4-Butirolactona/análisis , Anfetaminas/análisis , Cromatografía de Gases , Humanos , Ketamina/análisis , Metanfetamina/análogos & derivados , Metanfetamina/análisis , Nitrazepam/análogos & derivados , Nitrazepam/análisis , Oxibato de Sodio/análisisRESUMEN
Plumbagin is found in many herbal plants and inhibits the growth of various bacteria. Escherichia coli strains are relatively resistant to this drug. The mechanism of resistance is not clear. Previous findings showed that plumbagin treatment triggered up-regulation of many genes in E. coli including ahpC, mdaB, nfnB, nfo, sodA, yggX and ygfZ. By analyzing minimal inhibition concentration and inhibition zones of plumbagin in various gene-disruption mutants, ygfZ and sodA were found critical for the bacteria to resist plumbagin toxicity. We also found that the roles of YgfZ and SodA in detoxifying plumbagin are independent of each other. This is because of the fact that ectopically expressed SodA reduced the superoxide stress but not restore the resistance of bacteria when encountering plumbagin at the absence of ygfZ. On the other hand, an ectopically expressed YgfZ was unable to complement and failed to rescue the plumbagin resistance when sodA was perturbed. Furthermore, mutagenesis analysis showed that residue Cys228 within YgfZ fingerprint region was critical for the resistance of E. coli to plumbagin. By solvent extraction and HPLC analysis to follow the fate of the chemical, it was found that plumbagin vanished apparently from the culture of YgfZ-expressing E. coli. A less toxic form, methylated plumbagin, which may represent one of the YgfZ-dependent metabolites, was found in the culture supernatant of the wild type E. coli but not in the ΔygfZ mutant. Our results showed that the presence of ygfZ is not only critical for the E coli resistance to plumbagin but also facilitates the plumbagin degradation.
Asunto(s)
Antineoplásicos Fitogénicos/farmacología , Proteínas Portadoras/metabolismo , Proteínas de Escherichia coli/metabolismo , Escherichia coli/metabolismo , Naftoquinonas/farmacología , Secuencia de Aminoácidos , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Proteínas Portadoras/genética , Escherichia coli/genética , Proteínas de Escherichia coli/genética , Prueba de Complementación Genética , Pruebas de Sensibilidad Microbiana , Datos de Secuencia Molecular , Estructura Molecular , Mutagénesis Sitio-Dirigida , Naftoquinonas/química , Naftoquinonas/metabolismo , Alineación de Secuencia , Superóxido Dismutasa/genética , Superóxido Dismutasa/metabolismo , Superóxidos/metabolismoRESUMEN
The bark and roots of Cinnamomum osmophloeum are widely used in Taiwan as spice substitutes for C. CASSIA. We have isolated three novel lignan esters, one dibenzylbutane-type ligan ester [9,9'-di-O-feruloyl-(+)-5,5'-dimethoxy secoisolariciresinol (3)] and two cyclolignan esters [(7' S,8' R,8 R) -lyoniresinol-9-O-(E)-feruloyl ester ( 5) and (7' S,8' R,8 R)-lyoniresinol-9,9'-di-O-(E)-feruloyl ester (6)], and several known lignans from the heartwood and roots of C. osmophloeum. We identified these compounds using 1D and 2D NMR spectroscopy and mass spectrometry. Cytotoxicity assays of these novel lignan esters revealed that compound 6 has strong activities against human liver cancer (HepG2 and Hep3B) and oral cancer (Ca9-22) cells, with IC(50) values of 7.87, 4.31, and 2.51 microg/mL, respectively.
Asunto(s)
Antineoplásicos Fitogénicos/química , Antineoplásicos Fitogénicos/farmacología , Cinnamomum/química , Lignanos/química , Lignanos/farmacología , Línea Celular Tumoral , Humanos , Corteza de la Planta/química , Raíces de Plantas/químicaRESUMEN
Plant acid invertases, which are either associated with the cell wall or present in vacuoles, belong to family 32 of glycoside hydrolases (GH32). Homology modeling of bamboo vacuolar invertase Bobetafruct3 using Arabidopsis cell-wall invertase AtcwINV1 as a template showed that its overall structure is similar to GH32 enzymes, and that the three highly conserved motifs, NDPNG, RDP and EC, are located in the active site. This study also used site-directed mutagenesis to examine the roles of the conserved amino acid residues in these three motifs, which include Asp135, Arg259, Asp260, Glu316 and Cys317, and a conserved Trp residue (Trp159) that resides between the NDPNG and RDP motifs. The mutants W159F, W159L, E316Q and C317A retained acid invertase activity, but no invertase activity was observed for the mutant E316A or mutants with changes at Asp135, Arg259, or Asp260. The apparent K(m) values of the four mutants with invertase activity were all higher than that of the wild-type enzyme. The mutants W159L and E316Q exhibited lower k(cat) values than the wild-type enzyme, but an increase in the k(cat) value was observed for the mutants W159F and C317A. The results of this study demonstrate that these residues have individual functions in catalyzing sucrose hydrolysis.
Asunto(s)
Poaceae/enzimología , beta-Fructofuranosidasa/genética , beta-Fructofuranosidasa/metabolismo , Secuencia de Aminoácidos , Dominio Catalítico , Regulación de la Expresión Génica de las Plantas , Modelos Moleculares , Datos de Secuencia Molecular , Mutación , Proteínas de Plantas/metabolismo , Conformación ProteicaRESUMEN
In this study, we assessed the antitumor activity of the methanol extract from wood chips of the heartwood of the Taiwan red cypress, Chamaecyparis formosensis Matsumura, which is a precious tree species endemic to Taiwan. A brine shrimp lethality test (BST) indicated that the ethyl acetate (EtOAc)-soluble extract from the MeOH extract was a suitable candidate (LC (50) = 15.36 microg/mL) for further studies of the antitumor activity of its components. From this EtOAc fraction, we isolated six lignans and two norlignans and tested their cytotoxic activities IN VITRO against promyelocytic leukemia (HL-60) and hepatoma (Hepa-G2) cell lines. Among these compounds, 7,7'-( S)-dihydrotaiwanin C, isolated for the first time from nature, with its single crystal structure depicted in this study, exhibited significant cytotoxic activity against HL-60 cell lines IN VITRO (IC (50) = 4.03 microg/mL) after 24 hours. BST:brine shrimp lethality test Hepa-G2:human hepatoma HL-60:human leukemia IC (50):half-maximal inhibitory concentration SARs:structure-activity relationships AS:adenine sulfate.