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1.
Phys Rev Lett ; 120(6): 062503, 2018 Feb 09.
Artigo em Inglês | MEDLINE | ID: mdl-29481255

RESUMO

A precision mass investigation of the neutron-rich titanium isotopes ^{51-55}Ti was performed at TRIUMF's Ion Trap for Atomic and Nuclear science (TITAN). The range of the measurements covers the N=32 shell closure, and the overall uncertainties of the ^{52-55}Ti mass values were significantly reduced. Our results conclusively establish the existence of the weak shell effect at N=32, narrowing down the abrupt onset of this shell closure. Our data were compared with state-of-the-art ab initio shell model calculations which, despite very successfully describing where the N=32 shell gap is strong, overpredict its strength and extent in titanium and heavier isotones. These measurements also represent the first scientific results of TITAN using the newly commissioned multiple-reflection time-of-flight mass spectrometer, substantiated by independent measurements from TITAN's Penning trap mass spectrometer.

2.
Pharmacol Res ; 118: 82-92, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-27693910

RESUMO

An interaction of drug with food, herbs, and dietary supplements is usually the consequence of a physical, chemical or physiologic relationship between a drug and a product consumed as food, nutritional supplement or over-the-counter medicinal plant. The current educational review aims at reminding to the prescribing physicians that the most clinically relevant drug-food interactions may not be strictly limited to those with grapefruit juice and with the Saint John's Wort herbal extract and may be responsible for changes in drug plasma concentrations, which in turn decrease efficacy or led to sometimes life-threatening toxicity. Common situations handled in clinical practice such as aging, concomitant medications, transplant recipients, patients with cancer, malnutrition, HIV infection and those receiving enteral or parenteral feeding may be at increased risk of drug-food or drug-herb interactions. Medications with narrow therapeutic index or potential life-threatening toxicity, e.g., the non-steroidal anti-inflammatory drugs, opioid analgesics, cardiovascular medications, warfarin, anticancer drugs and immunosuppressants may be at risk of significant drug-food interactions to occur. Despite the fact that considerable effort has been achieved to increase patient' and doctor's information and ability to anticipate their occurrence and consequences in clinical practice, a thorough and detailed health history and dietary recall are essential for identifying potential problems in order to optimize patient prescriptions and drug dosing on an individual basis as well as to increase the treatment risk/benefit ratio.


Assuntos
Citrus paradisi , Interações Alimento-Droga , Sucos de Frutas e Vegetais , Interações Ervas-Drogas , Hypericum , Inibidores do Citocromo P-450 CYP3A/farmacologia , Suplementos Nutricionais , Humanos , Transportador 1 de Ânion Orgânico Específico do Fígado/antagonistas & inibidores , Micronutrientes/administração & dosagem , Farmacovigilância , Varfarina/farmacologia
3.
Dis Esophagus ; 30(2): 1-5, 2017 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-27766725

RESUMO

The aim of this study was to determine the outcome of salvage definitive chemoradiation (dCRT) for a locoregional recurrence after any prior curative treatment outside previously irradiated areas. Thirty-nine patients treated between January 2005 and December 2014 were reviewed for locoregional recurrent esophageal cancer outside previously irradiated areas. All patients received salvage treatment with external beam radiotherapy (50.4 Gy in 28 fractions) combined with weekly concurrent paclitaxel and carboplatin. The median follow-up period was 15 months (range 1.7-120). The median overall survival (OS) for all patients after salvage dCRT was 22 months (95% CI 6.2-37.6). The 1-, 3-, and 5-year OS was 72%, 31%, and 28%, respectively. Median survival after salvage dCRT for a regional lymph node recurrence was 33 months (95% CI 5.8-60.3) versus 14 months (95% CI 6.8-21.6) for a recurrence at the anastomosis (P = 0.022, logrank). Median OS was 35 months for the squamous cell carcinoma group and 19 months for the adenocarcinoma group (P = 0.67). Sixteen of 39 patients developed a locoregional recurrence after salvaged dCRT. The median locoregional recurrence-free survival (LRFS) was 24 months. The 1-, 3-, and 5-year LRFS was 79%, 36%, and 36%, respectively. Median disease-free survival (DFS) was 15 months. The 1-, 3-, and 5-year DFS was 66%, 27%, and 27%, respectively. Of 16 patients, 8 (50%) with a primary failure at the site of the anastomosis developed a local recurrence after salvaged dCRT compared to 7 of 22 patients (32%) with a primary recurrence in a lymph node. Definitive chemoradiation is a feasible and effective treatment for locoregional recurrent esophageal cancer outside a previously irradiated area, and should be given with a curative intent. This holds true for recurrence of both squamous cell carcinoma and adenocarcinoma. Lymph node recurrences have a markedly better prognosis than recurrences at the site of the anastomosis.


Assuntos
Quimiorradioterapia/métodos , Neoplasias Esofágicas/terapia , Esôfago/cirurgia , Recidiva Local de Neoplasia/terapia , Terapia de Salvação/métodos , Adenocarcinoma/patologia , Adenocarcinoma/terapia , Adulto , Idoso , Idoso de 80 Anos ou mais , Anastomose Cirúrgica/efeitos adversos , Anastomose Cirúrgica/métodos , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Carboplatina/administração & dosagem , Carcinoma de Células Escamosas/patologia , Carcinoma de Células Escamosas/terapia , Intervalo Livre de Doença , Neoplasias Esofágicas/patologia , Esôfago/patologia , Estudos de Viabilidade , Feminino , Seguimentos , Humanos , Linfonodos/patologia , Masculino , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/patologia , Paclitaxel/administração & dosagem , Prognóstico , Estudos Retrospectivos , Resultado do Tratamento , Adulto Jovem
4.
HIV Med ; 17(7): 550-6, 2016 08.
Artigo em Inglês | MEDLINE | ID: mdl-27186847

RESUMO

OBJECTIVES: HIV-infected individuals are at increased risk of incident fractures. Evaluation of trabecular bone micro-architecture is an important tool to assess bone strength, but its use has not yet been reported in middle-aged HIV-infected male individuals. The aim of the study was to compare bone micro-architecture between HIV-infected and HIV-uninfected men. METHODS: In this cross-sectional study, 53 HIV-infected male individuals with a mean (± standard deviation) age of 49 ± 9 years who had been receiving antiretroviral therapy including tenofovir disoproxil fumarate (DF) for at least 60 months were compared with 50 HIV-uninfected male controls, matched for age and ethnic origin. We studied the volumetric bone density and micro-architecture of the radius and tibia using high-resolution peripheral quantitative computed tomography (HR-p QCT). RESULTS: Volumetric trabecular bone density was 17% lower in the tibia (P < 10(-4) ) and 16% lower in the radius (P < 10(-3) ) in HIV-infected patients compared with controls. By contrast, the cortical bone density was normal at both sites. The tibial trabecular micro-architecture differed markedly between patients and controls: bone volume/total volume (BV/TV) and trabecular number were each 13% lower (P < 10(-4) for both). Trabecular separation and inhomogeneity of the network were 18% and 24% higher in HIV-infected patients than in controls, respectively. The radial BV/TV and trabecular thickness were each 13% lower (P < 10(-3) and 10(-2) , respectively). Cortical thickness was not different between the two groups. CONCLUSIONS: The findings of lower volumetric trabecular bone density and disrupted trabecular micro-architectural parameters in middle-aged male HIV-infected treated patients help to explain bone frailty in these patients.


Assuntos
Antirretrovirais/uso terapêutico , Doenças Ósseas/patologia , Osso Esponjoso/patologia , Infecções por HIV/complicações , Infecções por HIV/tratamento farmacológico , Adulto , Densidade Óssea , Doenças Ósseas/diagnóstico por imagem , Estudos Transversais , Humanos , Masculino , Pessoa de Meia-Idade , Rádio (Anatomia)/patologia , Tenofovir/uso terapêutico , Tíbia/patologia , Tomografia Computadorizada por Raios X
5.
Klin Padiatr ; 228(6-07): 325-331, 2016 Nov.
Artigo em Alemão | MEDLINE | ID: mdl-27171924

RESUMO

Background: In Germany some 2 000 children and adolescent are diagnosed with cancer every year. Curing rates are increasing and therewith also the number of survivors is growing. Survivors frequently suffer from long-term effects of the disease and its treatment, but long-term follow-up care shows deficits. Method: The Network for oncological advisory service (NOF) started in 11/2013, researching and building up a network of available support in Lower Saxony. A telephone hotline was installed in 01/2014 in order to advice survivors on their problems. At the same time, an interview study on survivors needs was conducted throughout Germany. Results: In the first 2 years, the NOF gave advice to 79 patients. Whilst enquiries of medical or psychological nature were transferred to the cooperation partner, requests on psychosocial and social legal issues are being deled by the NOF due to lack of appropriate partners. The evaluation of 25 interviews shows key issues in long-term after-care: (1) transition from acute therapy to everyday life, (2) problems due to pediatric cancer and therapy, (3) patients perception of own disposition, (4) social reactions towards survivors, (5) structure of long-term follow-up care, (6) information flow. Conclusion: Many survivors suffer from long-term effects of cancer and treatment. The lack of available contact person and being in limbo between cured and simultaneously affected by the cancer treatment and chronic diseases is perceived as being problematic. This translates to various requirements on a patient-oriented long-term care, mainly in the psychosocial field.


Assuntos
Assistência ao Convalescente/organização & administração , Consultores , Linhas Diretas/organização & administração , Comunicação Interdisciplinar , Colaboração Intersetorial , Assistência de Longa Duração/organização & administração , Equipe de Assistência ao Paciente/organização & administração , Encaminhamento e Consulta/organização & administração , Adolescente , Adulto , Criança , Estudos de Avaliação como Assunto , Seguimentos , Alemanha , Necessidades e Demandas de Serviços de Saúde/organização & administração , Humanos , Entrevista Psicológica , Satisfação do Paciente , Projetos Piloto , Sobreviventes , Cuidado Transicional/organização & administração , Adulto Jovem
6.
Nano Lett ; 14(2): 570-7, 2014 Feb 12.
Artigo em Inglês | MEDLINE | ID: mdl-24417568

RESUMO

Localized surface plasmon resonance (LSPR) as the resonant oscillation of conduction electrons in metal nanostructures upon light irradiation is widely used for sensing as well as nanoscale manipulation. The spectral resonance band position can be controlled mainly by nanoparticle composition, size, and geometry and is slightly influenced by the local refractive index of the near-field environment. Here we introduce another approach for tuning, based on interference modulation of the light scattered by the nanostructure. Thereby, the incoming electric field is wavelength-dependent modulated in strength and direction by interference due to a subwavelength spacer layer between nanoparticle and a gold film. Hence, the wavelength of the scattering maximum is tuned with respect to the original nanoparticle LSPR. The scattering wavelength can be adjusted by a metallic mirror layer located 100-200 nm away from the nanoparticle, in contrast to near-field gap mode techniques that work at distances up to 50 nm in the nanoparticle environment. Thereby we demonstrate, for the first time at the single nanoparticle level, that dependent on the interference spacer layer thickness, different distributions of the scattered signal can be observed, such as bell-shaped or doughnut-shaped point spread functions (PSF). The tuning effect by interference is furthermore applied to anisotropic particles (dimers), which exhibit more than one resonance peak, and to particles which are moved from air into the polymeric spacer layer to study the influence of the distance to the gold film in combination with a change of the surrounding refractive index.

8.
Spinal Cord ; 52(6): 420-7, 2014 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-24752292

RESUMO

STUDY DESIGN: Topical review of the literature. OBJECTIVES: The evaluation of patients with myelopathies requires radiological investigations; however, for the correct interpretation of the neuroimaging findings, the functional assessment of corticospinal conduction is helpful or even mandatory in many conditions. The objective of this review article was to assess the utility of the motor evoked potentials (MEPs) in diagnosis and management of the most frequent spinal cord disorders. SETTING: Salzburg (Austria) and Merano (Italy). METHODS: A MEDLINE search was performed using following terms: 'motor evoked potentials', 'transcranial magnetic stimulation', 'central motor conduction', 'compressive myelopathy', 'spinal cord infarction', 'spinal cord injury', 'syringomyelia', 'myelitis', 'hereditary spastic paraparesis', 'subacute combined degeneration' and 'hepatic myelopathy'. RESULTS: Central motor conduction abnormalities can be detected also in the absence of neuroradiological abnormalities-for example, in patients with subacute combined degeneration or hepatic myelopathy. In the most frequent patients with compressive myelopathies, MEPs were found to be very helpful in determining the functional significance of neuroimaging findings. MEP recording can supplement clinical examination and neuroimaging findings also in the assessment of the spinal cord injury level. In patients with spinal cord infarction, the MEP study can demonstrate spinal involvement even when radiological evidence for spinal cord damage is absent or equivocal, thus allowing an important early diagnosis. CONCLUSION: MEPs represent a highly sensitive and accurate diagnostic tool in many different spinal cord disorders. MEPs can also be useful in follow-up evaluation of motor function during treatment and rehabilitation.


Assuntos
Eletrodiagnóstico/métodos , Potencial Evocado Motor , Doenças da Medula Espinal/diagnóstico , Doenças da Medula Espinal/terapia , Humanos , Medula Espinal/fisiopatologia , Doenças da Medula Espinal/fisiopatologia
9.
J Viral Hepat ; 20(11): 779-89, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-24168257

RESUMO

Narlaprevir, a hepatitis C virus (HCV) NS3/4A serine protease inhibitor, has demonstrated robust antiviral activity in a placebo-controlled phase 1 study. To study evolutionary dynamics of resistant variants, the NS3 protease sequence was clonally analysed in thirty-two HCV genotype 1-infected patients following treatment with narlaprevir. Narlaprevir monotherapy was administered for one week (period 1) followed by narlaprevir/pegylated interferon-alpha-2b combination therapy with or without ritonavir (period 2) during two weeks, interrupted by a washout period of one month. Thereafter, all patients initiated pegylated interferon-alpha-2b/ribavirin combination therapy. Longitudinal clonal analysis was performed in those patients with NS3 mutations. After narlaprevir re-exposure, resistance-associated mutations at position V36, T54, R155 and A156 were detected in five patients in >95% of the clones. Narlaprevir retreatment resulted in a 2.58 and 5.06 log10 IU/mL viral load decline in patients with and without mutations, respectively (P=<0.01). After treatment, resistant variants were replaced with wild-type virus within 2-24 weeks in three patients. However, the R155K mutation was still observed 3.1 years after narlaprevir dosing in two patients in 5% and 45% of the viral population. Resistant variants could be detected early during treatment with narlaprevir. A slower viral load decline was observed in those patients with resistance-associated mutations detectable by direct population sequencing. These mutations disappeared within six months following treatment with the exception of R155K mutation, which persisted in two patients.


Assuntos
Antivirais/uso terapêutico , Dipeptídeos/uso terapêutico , Evolução Molecular , Hepacivirus/efeitos dos fármacos , Hepatite C Crônica/tratamento farmacológico , Inibidores de Proteases/uso terapêutico , Sulfonas/uso terapêutico , Proteínas não Estruturais Virais/genética , Adulto , Ciclopropanos , Quimioterapia Combinada/métodos , Feminino , Hepacivirus/genética , Humanos , Interferon alfa-2 , Interferon-alfa/uso terapêutico , Leucina/análogos & derivados , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Polietilenoglicóis/uso terapêutico , Prolina/análogos & derivados , Proteínas Recombinantes/uso terapêutico , Ribavirina/uso terapêutico , Análise de Sequência de DNA , Ureia , Carga Viral
10.
Int J Oral Maxillofac Surg ; 52(7): 801-805, 2023 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-36443143

RESUMO

This study was performed to present a single operator's experience of in-office (outside of a hospital setting) outpatient orthognathic surgery over a period of 12 years. A total of 254 surgeries were performed during this period. Average procedure times were comparable with published results from studies of similar material. The mean operating time for bimaxillary surgery (n = 21) was 3 hours and 11 minutes. Regarding single-jaw procedures, the mean operating time for Le Fort I osteotomy (n = 115) was 2 hours and 14 minutes and for bilateral sagittal split osteotomy (n = 118) was 2 hours and 1 minute. All patients were discharged from the office the same day, except one patient who was transported to the hospital after surgery due to an anaesthetic complication. This patient was discharged from the hospital later the same day. In this setting, outpatient orthognathic surgery is both safe and practical when careful attention is given to patient preparation and selection. Emergency phone contact with the surgeon in case of complications is important to avoid unnecessary hospitalization.


Assuntos
Cirurgia Ortognática , Procedimentos Cirúrgicos Ortognáticos , Humanos , Procedimentos Cirúrgicos Ortognáticos/métodos , Pacientes Ambulatoriais , Alta do Paciente , Osteotomia de Le Fort/métodos
12.
Rev Med Interne ; 42(2): 79-85, 2021 Feb.
Artigo em Francês | MEDLINE | ID: mdl-33160706

RESUMO

INTRODUCTION: Patients admitted from emergency units represent a large portion of the population in internal medicine departments. The aim of this study is to identify characteristics of patients and organization of these departments. METHODS: Between June 29th and July 26th 2015, voluntary internal medicine departments from the SiFMI group prospectively filled anonymized internet forms to collect data of each patients admitted in their ward from emergency units, during seven consecutive days. RESULTS: Three hundred and sixty-five patients from emergency departments were admitted in 18 internal medicine inpatients departments, totalling 1100 beds and 33,530 annual stays, 56% of them for emergency units inpatients. Mean age was 68 years, 54% were women, mean Charlson score was 2.6 and 44% of the patients took at least three drugs. Main causes of hospitalization were infectious (29%) and neurological (17%) diseases. Mean length of stay was 9.2 days. The medical team was composed by a median value of 4,5 [2,75-6,25] senior full-time equivalents, 86% were internists. Each department except one received residents, two third of them were from general medicine. CONCLUSION: This study highlights a high organizational variability among internal medicine departments and patients, and sets internal medicine as a specialty with a great capacity to achieve an integrative/comprehensive management of patients and to offer a comprehensive basis for physicians in training.


Assuntos
Serviço Hospitalar de Emergência , Medicina Interna , Idoso , Estudos Transversais , Feminino , Hospitalização , Hospitais , Humanos
13.
Med Mal Infect ; 50(1): 36-42, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30982671

RESUMO

OBJECTIVE: The WHO recommends same-day sputum smear microscopy for the diagnosis of smear-positive tuberculosis (TB) in countries with high TB burden for earlier diagnosis and treatment, a cornerstone to prevent air-borne transmission. We aimed to compare the conventional strategy (sputum collection on three consecutive days) and the same-day strategy (hour h, h+1, h+2) in France, a country with low TB burden. PATIENTS AND METHODS: Over a six-month period, all adult individuals presenting with presumptive smear-positive TB were eligible for the study, registered in https://clinicaltrials.gov/ ID (NCT02961569). Sputum specimens were collected three times the first day, then once on the second day and once on the third day. The concordance between the two strategies regarding smears and cultures were assessed. RESULTS: Of the 131 eligible individuals, 34 were given a TB treatment. Smears from hour h, h+1, h+2, day two and three were negative in 19 of these 34 patients. Positive smears were obtained in 15, 14, 15, 14, and 14 patients at hour h, h+1, h+2, on day two and three, respectively. Concordance regarding smear or culture was good, with Kappa 0.69 and 0.64, respectively. CONCLUSION: The same-day strategy seems to be a good alternative to the conventional strategy.


Assuntos
Mycobacterium tuberculosis/isolamento & purificação , Escarro/microbiologia , Tuberculose Pulmonar/diagnóstico , Tuberculose Pulmonar/microbiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Diagnóstico Precoce , Feminino , França , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Fatores de Tempo , Adulto Jovem
14.
Eur J Nucl Med Mol Imaging ; 36(5): 801-10, 2009 May.
Artigo em Inglês | MEDLINE | ID: mdl-19137294

RESUMO

PURPOSE: Comparative evaluation of regional brain perfusion measured by HMPAO-SPECT of patients with mild cognitive impairment (MCI), dementia of Alzheimer's type (DAT) and depression with cognitive impairment (DCI). METHODS: A total of 736 patients were investigated because of suspected cognitive dysfunction. After exclusion of patients with other forms of dementia than DAT or relevant accompanying disorders, SPECT data from 149 MCI, 131 DAT and 127 DCI patients, and 123 controls without any cognitive impairment, were analysed. Relative cerebral blood flow of 34 anatomical regions was assessed with automated analysis software (BRASS). RESULTS: Calculation of global forebrain perfusion discriminated demented from nondemented patients. Compared to controls DCI patients showed hypoperfusion of the thalamus, lentiform nucleus and medial temporal cortex. MCI patients differed significantly from controls concerning perfusion in both hemispheric temporal and parietal areas, and in the (right hemispheric) posterior part of the cingulate gyrus. MCI and DCI patients differed in the parietal, temporal superior and right hemispheric cingulate gyrus posterior cortices. Global forebrain and regional perfusion was more extensively reduced in DAT patients and discriminated them from controls, and MCI and DCI patients. Frontal perfusion disturbance was only present in DAT patients. CONCLUSION: Automated analysis of HMPAO-SPECT data from MCI patients showed significant perfusion deficits in regions also involved in DAT patients, but ROC analysis demonstrated only moderate sensitivity and specificity for differentiating DAT patients from controls and DCI patients. Frontal hypoperfusion seems to correspond with conversion from MCI to DAT. Finally, the results in DCI patients again raise the question of depression as an early symptom of neurodegeneration.


Assuntos
Doença de Alzheimer/complicações , Doença de Alzheimer/diagnóstico , Encéfalo/patologia , Transtornos Cognitivos/complicações , Transtornos Cognitivos/diagnóstico , Depressão/complicações , Depressão/diagnóstico , Tomografia Computadorizada de Emissão de Fóton Único/métodos , Idoso , Doença de Alzheimer/patologia , Automação , Transtornos Cognitivos/patologia , Depressão/patologia , Diagnóstico por Computador , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Doenças Neurodegenerativas/diagnóstico , Doenças Neurodegenerativas/patologia , Perfusão , Curva ROC
15.
J Cell Biol ; 107(5): 1707-15, 1988 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-2846585

RESUMO

Using monoclonal antibodies and indirect immunofluorescence microscopy, we investigated the distribution of the M protein in situ in vesicular stomatitis virus-(VSV) infected MDCK cells. M protein was observed free in the cytoplasm and associated with the plasma membrane. Using the ts045 mutant of VSV to uncouple the synthesis and transport of the VSV G protein we demonstrated that this distribution was not related to the presence of G protein on the cell surface. Sections of epon-embedded infected cells labeled with antibody to the M protein and processed for indirect horseradish peroxidase immunocytochemistry revealed that the M protein was associated specifically with the basolateral plasma membrane. The G and M proteins of VSV have therefore evolved features which bring them independently to the basolateral membrane of polarized epithelial cells and allow virus to bud specifically from that membrane.


Assuntos
Membrana Celular/microbiologia , Vírus da Estomatite Vesicular Indiana/fisiologia , Proteínas da Matriz Viral/metabolismo , Proteínas Virais/metabolismo , Anticorpos Monoclonais/biossíntese , Anticorpos Monoclonais/imunologia , Transporte Biológico , Membrana Celular/metabolismo , Células Cultivadas , Centrifugação com Gradiente de Concentração , Citoplasma/metabolismo , Retículo Endoplasmático/metabolismo , Retículo Endoplasmático/microbiologia , Ensaio de Imunoadsorção Enzimática , Células Epiteliais , Epitélio/microbiologia , Imunofluorescência , Complexo de Golgi/metabolismo , Complexo de Golgi/microbiologia , Imuno-Histoquímica , Metabolismo dos Lipídeos , Testes de Precipitina , Ligação Proteica , Processamento de Proteína Pós-Traducional
16.
J Cell Biol ; 108(3): 811-9, 1989 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-2537836

RESUMO

Cells infected with a temperature-sensitive mutant of vesicular stomatitis virus, ts045, or transfected with the plasmid vector pdTM12 produce mutant forms of the G protein that remain within the ER. The mutant G proteins were isolated by immunoprecipitation from cells metabolically labeled with [2-3H]mannose to facilitate analysis of the protein-linked oligosaccharides. The 3H-labeled glycopeptides recovered from the immunoprecipitated G proteins contained high mannose-type oligosaccharides. Structural analysis, however, indicated that 60-78% of the 3H-mannose-labeled oligosaccharides contained a single glucose residue and no fewer than eight mannose residues. The 3H-labeled ts045 oligosaccharides were deglucosylated and processed to complex-type units after the infected cells were returned to the permissive temperature. When shifted to the permissive temperature in the presence of a proton ionophore, the G protein oligosaccharides were deglucosylated but remained as high mannose-type units. The glucosylated state was observed, therefore, when the G protein existed in an altered conformation. The ts045 G protein oligosaccharides were deglucosylated in vitro by glucosidase II at both the permissive and nonpermissive temperatures. G protein isolated from ts045-infected cells labeled with [6-3H]galactose in the presence of cycloheximide contained 3H-glucose-labeled monoglucosylated oligosaccharides, indicating that the high mannose oligosaccharides were glucosylated in a posttranslational process. These results suggest that aberrant G proteins are selectively modified by resident ER enzymes to retain monoglucosylated oligosaccharides.


Assuntos
Glucose/metabolismo , Glicoproteínas de Membrana , Oligossacarídeos/metabolismo , Processamento de Proteína Pós-Traducional , Proteínas do Envelope Viral/metabolismo , Animais , Linhagem Celular , Manose/análise , Mutação , Oligossacarídeos/análise , Temperatura , Transfecção , Vírus da Estomatite Vesicular Indiana/genética , Proteínas do Envelope Viral/análise , Proteínas do Envelope Viral/genética
17.
J Cell Biol ; 97(6): 1777-87, 1983 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-6315743

RESUMO

An immunoelectron microscopic study was undertaken to survey the intracellular pathway taken by the integral membrane protein (G-protein) of vesicular stomatitis virus from its site of synthesis in the rough endoplasmic reticulum to the plasma membrane of virus-infected Chinese hamster ovary cells. Intracellular transport of the G-protein was synchronized by using a temperature-sensitive mutant of the virus (0-45). At the nonpermissive temperature (39.8 degrees C), the G-protein is synthesized in the cell infected with 0-45, but does not leave the rough endoplasmic reticulum. Upon shifting the temperature to 32 degrees C, the G-protein moves by stages to the plasma membrane. Ultrathin frozen sections of 0-45-infected cells were prepared and indirectly immunolabeled for the G-protein at different times after the temperature shift. By 3 min, the G-protein was seen at high density in saccules at one face of the Golgi apparatus. No large accumulation of G-protein-containing vesicles were observed near this entry face, but a few 50-70-mm electron-dense vesicular structures labeled for G-protein were observed that might be transfer vesicles between the rough endoplasmic reticulum and the Golgi complex. At blebbed sites on the nuclear envelope at these early times there was a suggestion that the G-protein was concentrated, these sites perhaps serving as some of the transitional elements for subsequent transfer of the G-protein from the rough endoplasmic reticulum to the Golgi complex. By 3 min after its initial asymmetric entry into the Golgi complex, the G-protein was uniformly distributed throughout all the saccules of the complex. At later times, after the G-protein left the Golgi complex and was on its way to the plasma membrane, a new class of G-protein-containing vesicles of approximately 200-nm diameter was observed that are probably involved in this stage of the transport process. These data are discussed, and the further prospects of this experimental approach are assessed.


Assuntos
Transformação Celular Viral , Glicoproteínas de Membrana , Vírus da Estomatite Vesicular Indiana/ultraestrutura , Proteínas do Envelope Viral , Proteínas Virais/metabolismo , Animais , Transporte Biológico , Linhagem Celular , Membrana Celular/metabolismo , Membrana Celular/ultraestrutura , Cricetinae , Cricetulus , Feminino , Complexo de Golgi/metabolismo , Complexo de Golgi/ultraestrutura , Microscopia Eletrônica , Membrana Nuclear/metabolismo , Membrana Nuclear/ultraestrutura , Ovário , Vírus da Estomatite Vesicular Indiana/metabolismo
18.
J Cell Biol ; 95(3): 697-703, 1982 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-6296155

RESUMO

We used electron microscope autoradiography (EMAR) to visualize the interaction of 125I-thrombin with its surface receptors on mouse embryo (ME) cells. Autoradiographic grains were spaced over the surface of cells in a periodic nonrandom pattern, indicating 125I-thrombin association with clusters of thrombin receptors. The grain spacing varied slightly from cell to cell, indicating subpopulations of cells with different numbers of thrombin receptors. The average distance between grains on ME cells after binding 125I-thrombin (125 ng/ml) at 37 degrees C was 1.65 +/- 0.49 microns. The average distance between grains on prefixed cells and cells incubated with 125I-thrombin at 4 degrees C was not significantly different from that observed at 37 degrees C. This indicates that thrombin receptors are clustered before thrombin binding and that the thrombin receptor aggregates do not redistribute into large aggregates on the surface of cells subsequent to thrombin binding. The number of grains per cluster also does not change under these three binding conditions. Thus, the number of occupied receptors in each cluster appears to be constant. On the basis of the average grain number and spacing, we estimate that each cluster is approximately 400 nm in diameter containing approximately 550 thrombin-binding sites. These receptor-clusters are not associated with specialized structures or coated regions of the membrane. Additionally, grains observed within cells were not found associated with coated vesicles. Therefore, neither the clustering patterns nor internalization of 125I-thrombin are characteristic of molecules which bind to receptors and are internalized by receptor-mediated endocytosis.


Assuntos
Membrana Celular/metabolismo , Receptores de Superfície Celular/metabolismo , Trombina/metabolismo , Animais , Autorradiografia , Membrana Celular/ultraestrutura , Células Cultivadas , Invaginações Revestidas da Membrana Celular/análise , Embrião de Mamíferos , Radioisótopos do Iodo , Camundongos , Microscopia Eletrônica , Receptores de Superfície Celular/análise , Receptores de Trombina
19.
J Cell Biol ; 110(3): 625-35, 1990 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-2155242

RESUMO

We have investigated the role of the smooth endoplasmic reticulum (SER) of UT-1 cells in the biogenesis of the glycoprotein (G) of vesicular stomatitis virus (VSV). Using immunofluorescence microscopy, we observed the wild type G protein in the SER of infected cells. When these cells were infected with the mutant VSV strain ts045, the G protein was unable to reach the Golgi apparatus at 40 degrees C, but was able to exit the rough endoplasmic reticulum (RER) and accumulate in the SER. Ribophorin II, a RER marker, remained excluded from the SER during the viral infection, ruling out the possibility that the infection had destroyed the separate identities of these two organelles. Thus, the mechanism that results in the retention of this mutant glycoprotein in the ER at 39.9 degrees C does not limit its lateral mobility within the ER system. We have also localized GRP78/BiP to the SER of UT-1 cells indicating that other mutant proteins may also have access to this organelle. Upon incubation at 32 degrees C, the mutant G protein was able to leave the SER and move to the Golgi apparatus. To measure how rapidly this transfer occurs, we assayed the conversion of the G protein's N-linked oligosaccharides from endoglycosidase H-sensitive to endoglycosidase H-resistant forms. After a 5-min lag, transport of the G protein followed first order kinetics (t1/2 = 15 min). In contrast, no lag was seen in the transport of G protein that had accumulated in the RER of control UT-1 cells lacking extensive SER. In these cells, the transport of G protein also exhibited first order kinetics (t1/2 = 17 min). Possible implications of this lag are discussed.


Assuntos
Retículo Endoplasmático/metabolismo , Glicoproteínas/metabolismo , Vírus da Estomatite Vesicular Indiana/metabolismo , Proteínas Virais/metabolismo , Animais , Linhagem Celular , Glicoproteínas/genética , Complexo de Golgi/metabolismo , Hidroximetilglutaril-CoA Redutases/biossíntese , Hidroximetilglutaril-CoA Redutases/genética , Cinética , Mutação , Processamento de Proteína Pós-Traducional , Proteínas Virais/genética
20.
J Cell Biol ; 101(2): 460-9, 1985 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-2991299

RESUMO

The structures of the asparagine-linked oligosaccharides of several variant forms of the vesicular stomatitis virus glycoprotein transiently expressed from cloned cDNAs have been determined. Glycopeptides isolated from forms of the G protein that reach the cell surface or that are secreted into the medium are virtually identical; they contain complex-type oligosaccharides whose nonreducing ends terminate in galactose and sialic acid residues. In contrast, forms of the G protein that remain intracellular possess oligosaccharides at intermediate stages in the processing pathway. One deletion mutant, delta 1473, codes for a protein that remains in the rough endoplasmic reticulum (Rose, J. K., and J. E. Bergmann, 1982, Cell, 30:753-762) and contains only high mannose-type oligosaccharides. Another mutant, delta 1554, codes for a glycoprotein that contains oligosaccharides of primarily two classes. One class is of the high mannose type and is similar to those found on the protein coded for by delta 1473. However, the major class contains biantennary and more highly branched complex-type oligosaccharides that terminate in N-acetylglucosamine rather than galactose or sialic acid residues. These data suggest that the protein coded for by delta 1554 migrates to the Golgi apparatus, but does not enter the more distal compartment(s) of the organelle which contains galactosyl- and sialyltransferases.


Assuntos
Asparagina/metabolismo , Complexo de Golgi/metabolismo , Glicoproteínas de Membrana , Oligossacarídeos/metabolismo , Vírus da Estomatite Vesicular Indiana/metabolismo , Proteínas do Envelope Viral , Proteínas Virais/metabolismo , Animais , Configuração de Carboidratos , Compartimento Celular , Linhagem Celular , Glicopeptídeos/isolamento & purificação , Haplorrinos , Manose/metabolismo , Proteínas de Membrana/isolamento & purificação , Oligossacarídeos/isolamento & purificação , Processamento Pós-Transcricional do RNA , Proteínas Virais/isolamento & purificação
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