RESUMO
OBJECTIVES: To explore the value of mast cell tryptase and brain natriuretic peptideï¼BNPï¼ in the differential diagnostic of sudden death due to hypersensitivity and coronary atherosclerotic heart disease. METHODS: Totally 30 myocardial samples were collected from the autopsy cases in the Department of Forensic Pathology, Shanxi Medical University during 2010-2015. All samples were divided into three groupsï¼ death of craniocerebral injury group, sudden death of hypersensitivity group and sudden death of coronary atherosclerotic heart disease group, 10 cases in each group. Mast cell tryptase and BNP in myocardium were detected by immunofluorescence staining and Western Blotting. RESULTS: Immunofluorescence staining showed that the positive staining mast cell tryptase appeared in myocardium of sudden death of hypersensitivity group and coronary atherosclerotic heart disease group. Among the three groups, the expression of mast cell tryptase showed significantly differences through pairwise comparison ï¼P<0.05ï¼; The expression level of BNP in sudden death of coronary atherosclerotic heart disease group were significantly higher than the sudden death of hypersensitivity group and death of craniocerebral injury group ï¼P<0.05ï¼. The difference of the expression level of BNP between the sudden death of hypersensitivity group and the death of craniocerebral injury group had no statistical significance ï¼P>0.05ï¼. CONCLUSIONS: The combined detection of the mast cell tryptase and BNP in myocardium is expected to provide help for the forensic differential diagnosis of sudden death due to hypersensitivity and coronary atherosclerotic heart disease.
Assuntos
Doença da Artéria Coronariana/complicações , Morte Súbita Cardíaca/etiologia , Miocárdio/metabolismo , Peptídeo Natriurético Encefálico/metabolismo , Triptases/metabolismo , Anafilaxia , Autopsia , Western Blotting , Estudos de Casos e Controles , Diagnóstico Diferencial , Imunofluorescência , Patologia Legal , Humanos , Masculino , Infarto do MiocárdioRESUMO
Although radical nephrectomy alone is widely accepted as the standard of care in localized treatment for renal cell carcinoma (RCC), it is not sufficient for the treatment of metastatic RCC (mRCC), which invariably leads to an unfavorable outcome despite the use of multiple therapies. Currently, sequential targeted agents are recommended for the management of mRCC, but the optimal drug sequence is still debated. This case was a 57-year-old man with clear-cell mRCC who received multiple therapies following his first operation in 2003 and has survived for over 10 years with a satisfactory quality of life. The treatments given included several surgeries, immunotherapy, and sequentially administered sorafenib, sunitinib, and everolimus regimens. In the course of mRCC treatment, well-planned surgeries, effective sequential targeted therapies and close follow-up are all of great importance for optimal management and a satisfactory outcome.
RESUMO
Although radical nephrectomy alone is widely accepted as the standard of care in localized treatment for renal cell carcinoma (RCC), it is not sufficient for the treatment of metastatic RCC (mRCC), which invariably leads to an unfavorable outcome despite the use of multiple therapies. Currently, sequential targeted agents are recommended for the management of mRCC, but the optimal drug sequence is still debated. This case was a 57-year-old man with clear-cell mRCC who received multiple therapies following his first operation in 2003 and has survived for over 10 years with a satisfactory quality of life. The treatments given included several surgeries, immunotherapy, and sequentially administered sorafenib, sunitinib, and everolimus regimens. In the course of mRCC treatment, well-planned surgeries, effective sequential targeted therapies and close follow-up are all of great importance for optimal management and a satisfactory outcome.
RESUMO
Cisplatin (cis-diammine-dichloroplatinum; CDDP) is an anticancer drug that induces significant hearing loss and balance dysfunction as side effects. Cilostazol (CS, 6-[4-(1-cyclohexyl-1H-tetrazol-5-yl) butoxy]-3, 4-dihydro-2-(1H)-quinolinone) has neuroprotective and antioxidant effects, whereas Ginkgo biloba extract (GbE) has preventive effects on CDDP-induced hearing loss in rats, and GbE enhances the antiatherogenic effect of CS by inhibiting the generation of reactive oxygen species (ROS). The purpose of this study was to investigate the effects of renexin (RXN), which contains GbE and CS, against CDDP-induced cochleo-vestibular dysfunction in rats and to elucidate the mechanism underlying the protective effects of RXN on auditory cells. Rats intraperitoneally injected with CDDP exhibited an increase in hearing threshold and vestibular dysfunction, which agreed with hair cell damage in the Organ of Corti and otoliths. However, these impairments were significantly prevented in a dose-dependent manner by pre- and co-treatment with RXN, and these preventive effects in RXN-treated rats were more prominent than those in GbE-treated rats. In a CDDP pharmacokinetic study, platinum concentration was very similar between CDDP-only treated and RXN+CDDP cotreated rats. RXN markedly attenuated CDDP-induced intracellular ROS and significantly reduced CDDP-activated expression of p-extracellular regulated kinase (ERK), BAX, cytochrome c, cleaved caspase-3 and cleaved poly (ADP-ribose) polymerase, but increased BCL-XL expression. These results show that RXN may have a synergistic effect by strongly protecting hearing and vestibular dysfunction induced by CDDP by inhibiting ROS production, mitochondrial pathways and the ERK pathway, without interfering with CDDP pharmacokinetics. Therefore, RXN could potentially be used to reduce CDDP-related hearing loss and dizziness.
Assuntos
Antineoplásicos/toxicidade , Apoptose/efeitos dos fármacos , Cisplatino/toxicidade , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Ginkgo biloba/química , Mitocôndrias/metabolismo , Fármacos Neuroprotetores/farmacologia , Extratos Vegetais/farmacologia , Tetrazóis/farmacologia , Animais , Comportamento Animal/efeitos dos fármacos , Linhagem Celular , Cilostazol , Sinergismo Farmacológico , Células Ciliadas Auditivas/citologia , Células Ciliadas Auditivas/efeitos dos fármacos , Células Ciliadas Auditivas/patologia , Perda Auditiva/metabolismo , Perda Auditiva/fisiopatologia , Perda Auditiva/prevenção & controle , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Mitocôndrias/efeitos dos fármacos , Fármacos Neuroprotetores/uso terapêutico , Extratos Vegetais/química , Extratos Vegetais/uso terapêutico , Ratos , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio/metabolismo , Tetrazóis/uso terapêuticoRESUMO
The standardization and validation of a one-step, single-tube, accelerated fluorescent-intercalating-dye-based reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay targeting the NS3 gene of Japanese B encephalitis virus (JEV) is described for rapid, simple, and high-throughput detection of JEV. The amplification can be completed in 35 min under isothermal conditions at 63°C by employing a set of six primers targeting the NS3 gene of JEV. The RT-LAMP assay described demonstrated high sensitivity for detecting JEV, with a detection limit in swine samples of 8.13 PFU/ml. The specificity of the selected primer sets was established by cross-reactivity studies with pathogens that exhibit similar clinical signs and testing of samples from healthy animals. The clinical applicability of the RT-LAMP assay was validated using either spiked samples or samples from seasonal outbreaks. The comparative evaluation of the RT-LAMP assay revealed 79.59 % concordance with conventional RT-PCR targeting the E gene of JEV. The RT-LAMP assay reported here is a valuable tool for rapid real-time and high-throughput seasonal infection surveillance and quarantine after outbreak through blood sampling by using ordinary real-time PCR thermocyclers without purchasing an expensive Loopamp real-time turbidimeter.
Assuntos
Surtos de Doenças/veterinária , Vírus da Encefalite Japonesa (Espécie)/genética , Vírus da Encefalite Japonesa (Espécie)/isolamento & purificação , Encefalite Japonesa/veterinária , Técnicas de Amplificação de Ácido Nucleico/métodos , Doenças dos Suínos/diagnóstico , Proteínas não Estruturais Virais/genética , Animais , China/epidemiologia , Primers do DNA , Encefalite Japonesa/diagnóstico , Encefalite Japonesa/epidemiologia , Encefalite Japonesa/virologia , Corantes Fluorescentes , Técnicas de Diagnóstico Molecular/métodos , RNA Helicases/genética , Sensibilidade e Especificidade , Serina Endopeptidases/genética , Suínos , Doenças dos Suínos/epidemiologia , Doenças dos Suínos/virologiaRESUMO
An orally delivered foot-and-mouth disease (FMD) vaccine has not previously been reported. By using a T4 bacteriophage nanoparticle surface gene-protein display system (T4-S-GPDS), we created a foot-and-mouth disease virus (FMDV) entire capsid protein vaccine candidate. On the T4 phage surface SOC site, a full length FMDV capsid precursor polyprotein (P1, 755 aa) and proteinase 3C (213 aa) derived from an infected pig of serotype O strain GD-10 (1999), were separately displayed on different T4 phage particle surfaces through inserting their coding region DNAs into the T4 phage genome, yielding phage strains T4-P1 and T4-3C. We also constructed a series of FMDV sub-full length capsid structural protein (subunit) containing T4 phage recombinant vaccines. Both sucking and young BALB/c mice were used as two kinds of FMDV vaccine potency evaluation models. Many groups of both model mice were vaccinated orally or by subcutaneous injection with varying FMDV-T4 phage recombinant vaccines, with and without addition of adjuvant, then challenged with a lethal dose of cattle source virulent FMDV. In the case of immunization with a mixture of phage T4-P1 and phage T4-3C particles without any adjuvant added, all mice were 100% protected following either oral or injection immunization, whereas 100% of the control, non-immunized mice and mice immunized with only T4 phage vector Z1/Zh(-) or wild-type T4(+)D phage died; in contrast, with FMDV subunit vaccine, less than 75% protection followed the same potency challenge in both mice model groups. In addition, two pigs immunized with a phage T4-P1 and phage T4-3C mix were protected upon housing together with infected pigs. This study represents a clear example of how FMD and other pathogenic disease vaccines can be prepared by a simple and efficient bacteriophage route.
Assuntos
Bacteriófago T4/imunologia , Capsídeo/imunologia , Vírus da Febre Aftosa/imunologia , Febre Aftosa/prevenção & controle , Vacinas Virais/imunologia , Animais , Animais Recém-Nascidos , DNA Viral/biossíntese , DNA Viral/genética , DNA Viral/imunologia , Eletroforese em Gel de Poliacrilamida , Ensaio de Imunoadsorção Enzimática , Escherichia coli/ultraestrutura , Escherichia coli/virologia , Febre Aftosa/imunologia , Febre Aftosa/virologia , Vírus da Febre Aftosa/genética , Vírus da Febre Aftosa/patogenicidade , Injeções Subcutâneas , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Microscopia Eletrônica , Biblioteca de Peptídeos , Regiões Promotoras Genéticas/genética , Engenharia de Proteínas , Sorotipagem , Suínos , Vacinas Sintéticas/uso terapêuticoRESUMO
Human immunodeficiency virus type 1 (HIV-1) produces two polyproteins, Pr55(Gag) and Pr160(Gag-Pol), that are cleaved into mature functional subunits by the virally encoded protease. Drugs that inhibit this protease are an important part of anti-HIV therapy. We studied the ordered accumulation of Gag and Gag-Pol processing intermediates by variably blocking the protease with HIV-1 protease inhibitors (PIs). Variable protease inhibition caused accumulation of a complex pattern of processing intermediates, which was the same after incubating HIV-1-infected cells with increasing concentrations of either one of the peptidomimetic inhibitors indinavir, saquinavir (SQV), ritonavir (RTV), nelfinavir, and SC-52151 or one of the nonpeptidomimetic inhibitors DMP450, DMP323, PNU-140135, and PNU-109112 for 3 days. The patterns of Gag and Gag-Pol processing intermediate accumulation were nearly identical when the following were compared: cell- versus virion-associated proteins, HIV-1-infected transformed cell lines versus primary human peripheral blood mononuclear cells (PBMCs) and HIV-1(MN) versus HIV-1(IIIB) virus strains. RTV was a more potent inhibitor of p24 production in PBMCs than SQV by approximately 7-fold, whereas SQV was a more potent inhibitor in transformed cells than RTV by approximately 30-fold. Although the antiretroviral potency of HIV-1 PIs may change as a function of cell type, the polyprotein intermediates that accumulate with increasing drug concentrations are the same. These results support sequential processing of Gag and Gag-Pol polyproteins by the HIV-1 protease and may have important implications for understanding common cross-resistance pathways.
Assuntos
Produtos do Gene gag/metabolismo , Inibidores da Protease de HIV/farmacologia , HIV-1/efeitos dos fármacos , Precursores de Proteínas/metabolismo , Transformação Celular Viral , Células Cultivadas , HIV-1/metabolismo , Humanos , Leucócitos Mononucleares/metabolismo , Processamento de Proteína Pós-Traducional , Produtos do Gene gag do Vírus da Imunodeficiência Humana , Produtos do Gene pol do Vírus da Imunodeficiência HumanaRESUMO
The nucleocapsid (NC) domain of the retrovirus Gag protein plays several important roles in the viral life cycle, including virus assembly, viral genomic RNA encapsidation, primer tRNA placement, and enhancement of viral reverse transcription. In this study, deletion of NC domain of human immunodeficiency virus type 1 (HIV-1) Gag was found to drastically reduce virus particle production in CD4(+) T cells. Cellular fractionation experiments showed that although most of the uncleaved wild-type HIV-1 Gag, unmyristylated Gag, and p6(Gag) domain-truncated Gag molecules copurified with the host cell cytoskeleton, most of the mutant Gag molecules lacking both the NC and p6(Gag) domains failed to cofractionate with cytoskeleton. In wild-type virus-infected cells, in which the viral protease was active, the cleaved NCp7 copurified with the cytoskeleton, whereas most of the MAp17 and CAp24 did not. Monoclonal antibody against actin coimmunoprecipitated full-length Gag and p6(Gag) domain-truncated Gag molecules from cell lysates but failed to precipitate the truncated mutant Gag molecules lacking NC plus p6(Gag). Purified recombinant NCp7, but not CAp24, was able to bind F-actin in cosedimentation experiments. Furthermore, wild-type NCp7 and a zinc finger mutant NCp7(F16A), like a cellular actin-binding protein (the villin headpiece), bound F-actin in a dose-dependent fashion in vitro. Taken together, these results suggest that HIV-1 NCp7 can bind F-actin directly and that interaction between HIV-1 Gag and the actin cytoskeleton through the NC domain may play an important role in HIV-1 assembly and/or other steps of the viral life cycle.
Assuntos
Actinas/metabolismo , Produtos do Gene gag/metabolismo , HIV-1/fisiologia , Sequência de Bases , Produtos do Gene gag/genética , Humanos , Dados de Sequência Molecular , Ligação Proteica/fisiologia , Deleção de Sequência , Montagem de Vírus , Replicação ViralRESUMO
It is unclear whether proteolytic processing of the human immunodeficiency virus type 1 (HIV-1) Gag protein is dependent on virus assembly at the plasma membrane. Mutations that prevent myristylation of HIV-1 Gag proteins have been shown to block virus assembly and release from the plasma membrane of COS cells but do not prevent processing of Gag proteins. In contrast, in HeLa cells similar mutations abolished processing of Gag proteins as well as virus production. We have now addressed this issue with CD4(+) T cells, which are natural target cells of HIV-1. In these cells, myristylation of Gag proteins was required for proteolytic processing of Gag proteins and production of extracellular viral particles. This result was not due to a lack of expression of the viral protease in the form of a Gag-Pol precursor or a lack of interaction between unmyristylated Gag and Gag-Pol precursors. The processing defect of unmyristylated Gag was partially rescued ex vivo by coexpression with wild-type myristylated Gag proteins in HeLa cells. The cell type-dependent processing of HIV-1 Gag precursors was also observed when another part of the plasma membrane binding signal, a polybasic region in the matrix protein, was mutated. The processing of unmyristylated Gag precursors was inhibited in COS cells by HIV-1 protease inhibitors. Altogether, our findings demonstrate that the processing of HIV-1 Gag precursors in CD4(+) T cells occurs normally at the plasma membrane during viral morphogenesis. The intracellular environment of COS cells presumably allows activation of the viral protease and proteolytic processing of HIV-1 Gag proteins in the absence of plasma membrane binding.
Assuntos
Produtos do Gene gag/metabolismo , HIV-1/metabolismo , Precursores de Proteínas/metabolismo , Proteínas da Matriz Viral/metabolismo , Animais , Sequência de Bases , Sítios de Ligação , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/virologia , Células COS , Linhagem Celular , Membrana Celular/metabolismo , Membrana Celular/virologia , DNA Viral/genética , Mutação da Fase de Leitura , Inibidores da Protease de HIV/farmacologia , HIV-1/efeitos dos fármacos , HIV-1/genética , Células HeLa , Humanos , Ácido Mirístico/metabolismo , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Transfecção , Proteínas da Matriz Viral/química , Proteínas da Matriz Viral/genéticaRESUMO
One of the crucial steps in the assembly of the human immunodeficiency virus type 1 (HIV-1) and other retroviruses is the incorporation and retention of all the key viral enzymes in released virions. The viral enzymes protease, reverse transcriptase, and integrase of HIV-1 are initially synthesized as Gag-Pol fusion polyproteins. It has been shown that the incorporation of Gag-Pol polyproteins during virus assembly requires the Gag domains that are shared by the Gag and Gag-Pol precursors. We now report that truncation of the C-terminal p6 domain of HIV-1 Gag, which is present in the Gag precursor but not in the Gag-Pol precursor, drastically reduced the amount of Pol proteins in the mutant virions. Mutations in the lentivirus conserved motif P(T/S)APP in p6 also drastically reduced the amount of Pol proteins in mutant virions. The steady-state levels of Gag-Pol precursors and cleaved Pol proteins in the transfected cells were not affected by mutations in p6. The incorporation of unprocessed Gag-Pol precursors into p6 mutant virions was detected when the viral protease was mutated, suggesting that the interactions among mutant Gag molecules and Gag-Pol precursors were not significantly affected. These results suggest that the p6 domain of HIV-1 Gag may play an important role in recruiting or retaining cleaved Pol proteins during virus assembly.