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1.
Phys Rev Lett ; 131(9): 092502, 2023 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-37721815

RESUMO

We successfully performed electron scattering off unstable nuclei which were produced online from the photofission of uranium. The target ^{137}Cs ions were trapped with a new target-forming technique that makes a high-density stationary target from a small number of ions by confining them in an electron storage ring. After developments of target generation and transportation systems and the beam stacking method to increase the ion beam intensity up to approximately 2×10^{7} ions per pulse beam, an average luminosity of 0.9×10^{26} cm^{-2} s^{-1} was achieved for ^{137}Cs. The obtained angular distribution of elastically scattered electrons is consistent with a calculation. This success marks the realization of the anticipated femtoscope which clarifies the structures of exotic and short-lived unstable nuclei.

2.
Clin Exp Allergy ; 48(3): 278-287, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29315896

RESUMO

BACKGROUND: Respiratory impedance comprises the resistance and reactance of the respiratory system and can provide detailed information on respiratory function. However, details of the relationship between impedance and morphological airway changes in asthma are unknown. OBJECTIVE: We aimed to evaluate the correlation between imaging-based airway changes and respiratory impedance in patients with asthma. METHODS: Respiratory impedance and spirometric data were evaluated in 72 patients with asthma and 29 reference subjects. We measured the intraluminal area (Ai) and wall thickness (WT) of third- to sixth-generation bronchi using three-dimensional computed tomographic analyses, and values were adjusted by body surface area (BSA, Ai/BSA, and WT/the square root (√) of BSA). RESULTS: Asthma patients had significantly increased respiratory impedance, decreased Ai/BSA, and increased WT/√BSA, as was the case in those without airflow limitation as assessed by spirometry. Ai/BSA was inversely correlated with respiratory resistance at 5 Hz (R5) and 20 Hz (R20). R20 had a stronger correlation with Ai/BSA than did R5. Ai/BSA was positively correlated with forced expiratory volume in 1 second/forced vital capacity ratio, percentage predicted forced expiratory volume in 1 second, and percentage predicted mid-expiratory flow. WT/√BSA had no significant correlation with spirometry or respiratory impedance. CONCLUSIONS & CLINICAL RELEVANCE: Respiratory resistance is associated with airway narrowing.


Assuntos
Asma/diagnóstico por imagem , Asma/fisiopatologia , Imageamento Tridimensional/métodos , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Resistência das Vias Respiratórias/fisiologia , Impedância Elétrica , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Testes de Função Respiratória , Tomografia Computadorizada por Raios X/métodos , Adulto Jovem
4.
Phys Rev Lett ; 118(26): 262501, 2017 Jun 30.
Artigo em Inglês | MEDLINE | ID: mdl-28707914

RESUMO

The first elastic electron scattering has been successfully performed at the self-confining radioactive-isotope ion target (SCRIT) facility, the world's first electron scattering facility for SCRIT technique achieved high luminosity (over 10^{27} cm^{-2} s^{-1}, sufficient for determining the nuclear shape) with only 10^{8} target ions. While ^{132}Xe used in this time as a target is a stable isotope, the charge density distribution was first extracted from the momentum transfer distributions of the scattered electrons by comparing the results with those calculated by a phase shift calculation.

6.
Clin Exp Immunol ; 177(1): 110-20, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24580543

RESUMO

Dermatomyositis (DM) and polymyositis (PM) are collectively termed autoimmune myopathy. To investigate the difference between muscle- and skin-infiltrating T cells and to address their role for myopathy, we characterized T cells that were directly expanded from the tissues. Enrolled into this study were 25 patients with DM and three patients with PM. Muscle and skin biopsied specimens were immersed in cRPMI medium supplemented with interleukin (IL)-2 and anti-CD3/CD28 antibody-conjugated microbeads. The expanded cells were subjected to flow cytometry to examine their phenotypes. We analysed the cytokine concentration in the culture supernatants from the expanded T cells and the frequencies of cytokine-bearing cells by intracellular staining. There was non-biased in-vitro expansion of tissue-infiltrating CD4(+) and CD8(+) T cells from the muscle and skin specimens. The majority of expanded T cells were chemokine receptor (CCR) type 7(-) CD45RO(+) effecter memory cells with various T cell receptor (TCR) Vßs. The skin-derived but not muscle-derived T cells expressed cutaneous lymphocyte antigen (CLA) and CCR10 and secreted large amounts of IL-17A, suggesting that T helper type 17 (Th17) cells may have a crucial role in the development of skin lesions. Notably, the frequency of IL-4-producing chemokine (C-X-C motif) receptor (CXCR)4(+) Th2 cells was significantly higher in the muscle-derived cells and correlated inversely with the serum creatine phosphokinase (CPK) and lactate dehydrogenase (LDH) levels. stromal-derived factor (SDF)-1/CXCL12, a ligand for CXCR4, was expressed at a high level in the vascular endothelial cells between muscular fasciculi. Our study suggests that T cell populations in the muscle and skin are different, and the Th2 cell infiltrate in the muscle is associated with the low severity of myositis in DM.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Dermatomiosite/imunologia , Músculos/imunologia , Pele/imunologia , Células Th2/imunologia , Adulto , Idoso , Antígenos de Diferenciação de Linfócitos T/metabolismo , Movimento Celular , Separação Celular , Células Cultivadas , Quimiocina CXCL12/metabolismo , Feminino , Citometria de Fluxo , Humanos , Memória Imunológica , Imunofenotipagem , Interleucina-17/metabolismo , Interleucina-4/metabolismo , Masculino , Glicoproteínas de Membrana/metabolismo , Pessoa de Meia-Idade , Receptores CCR10/metabolismo , Receptores CXCR4/metabolismo
7.
Dev Dyn ; 242(2): 179-88, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-23192966

RESUMO

BACKGROUND: In the cochlea, patterning of the organ of Corti is tightly regulated to produce a single row of sound-detecting inner hair cells and three rows of outer hair cells, which amplify and refine the signal. The recently identified R-Spondin family of signaling molecules usually act as co-activators of Wnt signaling; it is thought that they regulate turnover of Wnt receptors at the membrane. We sought to test whether R-Spondins function in the developing cochlea. RESULTS: Expression analysis of all four members of the R-Spondin family showed that only R-Spondin2 (Rspo2) is expressed in the cochlea during development of the sensory epithelium. Examination of an Rspo2(-/-) mouse showed that loss of Rspo2 results in an additional single row of outer hair cells and disruption of peripheral innervation pattern. Addition of Rspo2 recombinant protein to organotypic cochlear cultures resulted in a small but significant decrease in the number of outer hair cells. CONCLUSIONS: Rspo2 is required to limit the number of outer hair cells to three rows and for optimal arrangement of peripheral nerve fibers. The Rspo2 gain- and loss-of-function studies show that in the ear, Rspo2 function is not consistent with its assigned role as a Wnt potentiator.


Assuntos
Cóclea/embriologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Morfogênese/fisiologia , Trombospondinas/metabolismo , Animais , Cóclea/metabolismo , Primers do DNA/genética , Imunofluorescência , Regulação da Expressão Gênica no Desenvolvimento/genética , Células Ciliadas Auditivas/fisiologia , Processamento de Imagem Assistida por Computador , Hibridização In Situ , Camundongos , Camundongos Knockout , Microscopia Confocal , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Trombospondinas/genética
8.
Gene Ther ; 20(8): 816-23, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23344066

RESUMO

The development of a safe and reproducible gene delivery system is an essential step toward the clinical application of the hydrodynamic gene delivery (HGD) method. For this purpose, we have developed a novel electric power-driven injection system called the HydroJector-EM, which can replicate various time-pressure curves preloaded into the computer program before injection. The assessment of the reproducibility and safety of gene delivery system in vitro and in vivo demonstrated the precise replication of intravascular time-pressure curves and the reproducibility of gene delivery efficiency. The highest level of luciferase expression (272 pg luciferase per mg of proteins) was achieved safely using the time-pressure curve, which reaches 30 mm Hg in 10 s among various curves tested. Using this curve, the sustained expression of a therapeutic level of human factor IX protein (>500 ng ml(-1)) was maintained for 2 months after the HGD of the pBS-HCRHP-FIXIA plasmid. Other than a transient increase in liver enzymes that recovered in a few days, no adverse events were seen in rats. These results confirm the effectiveness of the HydroJector-EM for reproducible gene delivery and demonstrate that long-term therapeutic gene expression can be achieved by automatic computer-controlled hydrodynamic injection that can be performed by anyone.


Assuntos
Fator IX/genética , Técnicas de Transferência de Genes , Vetores Genéticos , Animais , Expressão Gênica , Humanos , Hidrodinâmica , Injeções , Fígado/metabolismo , Luciferases/metabolismo , Ratos
9.
Clin Exp Allergy ; 43(5): 521-6, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23600542

RESUMO

BACKGROUND: Clinical application of the forced oscillation technique (FOT) has progressed with the spread of commercially available FOT devices. The correlation between respiratory impedance and spirometry has been reported; however, the association with airway inflammation and pulmonary function, in the lung periphery in particular, is unclear. OBJECTIVE: To assess whether respiratory impedance is associated with peripheral airway inflammation and dysfunction in asthma. METHODS: Subjects included 78 patients with overall controlled asthma. We measured whole-breath or within-breath respiratory system resistance (Rrs) and reactance (Xrs) using a commercially available multi-frequency FOT device (MostGraph-01), and assessed the correlation with the fraction of exhaled nitric oxide (FeNO), alveolar nitric oxide concentration (CANO), maximal NO flux in the conductive airways (J'awNO), and the N2 phase III slope of single breath N2 washout (delta N2 ). RESULTS: The differences between inspiratory and expiratory phases of Xrs at 5 Hz (X5), resonant frequency (Fres), and a low-frequency reactance area (ALX) were significantly correlated with CANO; however, there was no correlation between respiratory impedance and FeNO or J'awNO. The delta N2 values were significantly correlated with whole-breath, inspiratory, and expiratory Rrs and Xrs, except for R20. CONCLUSIONS AND CLINICAL RELEVANCE: We conclude that respiratory impedance reflects peripheral airway inflammation and ventilation inhomogeneity.


Assuntos
Asma/fisiopatologia , Mecânica Respiratória , Adulto , Idoso , Estudos de Casos e Controles , Expiração , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Óxido Nítrico , Testes de Função Respiratória , Fatores de Risco
10.
J Viral Hepat ; 19(2): e212-9, 2012 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-22239521

RESUMO

Acoustic Radiation Force Impulse (ARFI) imaging is a novel ultrasound-based elastography method that is integrated in a conventional ultrasound machine enabling the exact localization of measurement site. It might present an alternative method to transient elastography for the noninvasive assessment of liver fibrosis. At present, studies with small patient population have shown promising results. A systematic review and meta-analysis of pooled patient data were performed to evaluate the overall performance of ARFI for the staging of liver fibrosis. Literature databases were searched up to 10/2010. The authors of the original publication were contacted, and the original patient data were requested. A meta-analysis was performed using a random effect meta-analytic method for diagnostic tests. In addition, available data comparing ARFI with FibroScan with the DeLong test were evaluated. Literature search yielded nine full-paper publications evaluating ARFI while using liver biopsy as reference method. Original patient data were available from eight studies including 518 patients. The mean diagnostic accuracy of ARFI expressed as areas under ROC curves (AUROC) was 0.87 for the diagnosis of significant fibrosis (F ≥ 2), 0.91 for the diagnosis of severe fibrosis (F ≥ 3), and 0.93 for the diagnosis of cirrhosis. ARFI can be performed with good diagnostic accuracy for the noninvasive staging of liver fibrosis.


Assuntos
Técnicas de Imagem por Elasticidade , Cirrose Hepática/diagnóstico , Cirrose Hepática/patologia , Fígado/diagnóstico por imagem , Fígado/patologia , Índice de Gravidade de Doença , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Adulto Jovem
11.
Sarcoidosis Vasc Diffuse Lung Dis ; 29(1): 69-73, 2012 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23311128

RESUMO

BACKGROUND: Pulmonary dendritic cells (DCs) are key regulators of immune responses. An increased accumulation of DCs was reported in the lungs of patients with idiopathic interstitial pneumonia (IIP). OBJECTIVE: This study aimed to investigate the number of pulmonary DCs in patients with collagen vascular disease associated interstitial lung diseases (CVD-ILDs). DESIGN: Lung tissue samples obtained from 27 patients with IIP and 39 patients with CVD-ILD were detected using monoclonal antibodies against CD1a, CD1c, CD83, Langerin and DC-specific intercellular adhesion molecule-3-grabbing nonintegrin (DC-SIGN). RESULTS: No significant differences in the number or distribution of DCs were observed between patients with IIP and CVD-ILDs. When DC marker expression was analyzed according to pathological subgroup, patients with idiopathic usual interstitial pneumonia (UIP) showed increased DC-SIGN staining when compared with CVD-UIP (p < 0.05). CONCLUSION: Both mature and immature DCs accumulate in CVD-ILDs. The number of DCs expressing DC-SIGN in CVD-UIP was decreased compared with that in idiopathic UIP. The variation in accumulated DC-SIGN-positive cells might help to explain the differences in the development and maintenance of lung inflammation between idiopathic UIP and CVD-UIP.


Assuntos
Células Dendríticas/imunologia , Fibrose Pulmonar Idiopática/imunologia , Doenças Pulmonares Intersticiais/imunologia , Pulmão/imunologia , Idoso , Idoso de 80 Anos ou mais , Antígenos CD/análise , Antígenos CD1/análise , Biomarcadores/análise , Biópsia , Moléculas de Adesão Celular/análise , Feminino , Glicoproteínas/análise , Humanos , Fibrose Pulmonar Idiopática/classificação , Fibrose Pulmonar Idiopática/patologia , Imunoglobulinas/análise , Lectinas Tipo C/análise , Pulmão/patologia , Doenças Pulmonares Intersticiais/classificação , Doenças Pulmonares Intersticiais/patologia , Masculino , Lectinas de Ligação a Manose/análise , Glicoproteínas de Membrana/análise , Pessoa de Meia-Idade , Receptores de Superfície Celular/análise , Antígeno CD83
12.
Eur Respir J ; 37(2): 371-5, 2011 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-20530033

RESUMO

There are very few data on serum procalcitonin (PCT) levels in pulmonary tuberculosis (PTB) patients who are negative for HIV. We assessed serum PCT in consecutive patients diagnosed with pulmonary tuberculosis or community-acquired pneumonia (CAP) on admission to discriminate between PTB and CAP, and examined the value of prognostic factors in PTB. 102 PTB patients, 62 CAP patients, and 34 healthy volunteers were enrolled. Serum PCT in PTB patients was significantly lower than in CAP patients (mean ± sd 0.21 ± 0.49 versus 4.10 ± 8.68 ng·mL⁻¹; p < 0.0001). By receiver-operating characteristic curve analysis, serum PCT was an appropriate discrimination marker for PTB and CAP (area under the curve 0.866). PTB patients with ≥ 0.5 ng·mL⁻¹ (normal cut-off) had significantly shorter survival than those with < 0.5 ng·mL⁻¹ (p < 0.0001). Serum PCT is not habitually elevated in HIV-negative PTB patients and is a useful biomarker for discriminating between PTB and CAP; however, when serum PCT is outside the normal range, it is a poor prognostic marker.


Assuntos
Calcitonina/sangue , Pneumonia Bacteriana/diagnóstico , Precursores de Proteínas/sangue , Tuberculose Pulmonar/sangue , Tuberculose Pulmonar/diagnóstico , Idoso , Idoso de 80 Anos ou mais , Biomarcadores/sangue , Peptídeo Relacionado com Gene de Calcitonina , Infecções Comunitárias Adquiridas/sangue , Infecções Comunitárias Adquiridas/diagnóstico , Diagnóstico Diferencial , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Pneumonia Bacteriana/sangue , Prognóstico , Curva ROC , Índice de Gravidade de Doença
13.
Nat Med ; 3(4): 409-13, 1997 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-9095174

RESUMO

The Fas ligand (FasL) is expressed in activated T cells and induces apoptosis in Fas-bearing cells. A cytotoxic T lymphocyte (CTL) clone specific for hepatitis B surface antigen (HBsAg) causes an acute liver disease in HBsAg transgenic mice. Here we observed that the CTL clone killed hepatocytes expressing HBsAg in a Fas-dependent manner. Administration of the soluble form of Fas into HBsAg transgenic mice prevented the CTL-induced liver disease. In the second model, mice were primed with Propionibacterium acnes. A subsequent challenge with lipopolysaccharide (LPS) killed the mice by inducing liver injury. Neutralization of FasL rescued the mice from LPS-induced mortality, and Fas-null mice were resistant to LPS-induced mortality. These results suggest that FasL has an essential role in the development of hepatitis.


Assuntos
Encefalopatia Hepática/etiologia , Glicoproteínas de Membrana/metabolismo , Receptor fas/metabolismo , Animais , Citotoxicidade Imunológica , Endotoxinas/farmacologia , Proteína Ligante Fas , Encefalopatia Hepática/imunologia , Encefalopatia Hepática/mortalidade , Encefalopatia Hepática/prevenção & controle , Antígenos de Superfície da Hepatite B/genética , Antígenos de Superfície da Hepatite B/imunologia , Camundongos , Camundongos Transgênicos , Solubilidade , Análise de Sobrevida , Linfócitos T Citotóxicos , Receptor fas/farmacologia
14.
Nat Med ; 4(11): 1287-92, 1998 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-9809553

RESUMO

Fas ligand is a well-characterized apoptosis inducer. Here we demonstrate that Fas ligand induces the processing and secretion of interleukin-1beta (IL-1beta) in peritoneal exudate cells. This IL-1beta secretion is independent of IL-1beta converting enzyme (caspase 1), yet it is inhibited by caspase inhibitors, indicating that a caspase(s) in addition to IL-1beta converting enzyme can process IL-1beta. Inoculation of tumor cells expressing Fas ligand into wild-type mice induces a massive neutrophil infiltration that is, in contrast, suppressed in IL-1alpha/beta knockout mice. These results demonstrate a newly discovered role for Fas ligand in inflammation, and challenge the dogma that apoptosis does not induce inflammation.


Assuntos
Apoptose , Caspase 1/metabolismo , Inflamação/fisiopatologia , Interleucina-1/biossíntese , Glicoproteínas de Membrana/fisiologia , Receptor fas/fisiologia , Animais , Caspase 1/deficiência , Caspase 1/genética , Células Cultivadas , Cruzamentos Genéticos , Proteína Ligante Fas , Feminino , Fibrossarcoma/imunologia , Fibrossarcoma/patologia , Homozigoto , Inflamação/imunologia , Interleucina-1/deficiência , Interleucina-1/genética , Linfócitos/imunologia , Masculino , Glicoproteínas de Membrana/biossíntese , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos , Camundongos Knockout , Neutrófilos/fisiologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Baço/imunologia
15.
Nat Med ; 2(3): 317-22, 1996 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-8612231

RESUMO

The Fas ligand (FasL), a member of the tumor necrosis factor family, induces apoptosis in Fas-bearing cells. The membrane-bound human FasL was found to be converted to a soluble form (sFasL) by the action of a matrix metalloproteinase-like enzyme. Two neutralizing monoclonal anti-human FasL antibodies were identified, and an enzyme-linked immunosorbent assay (ELISA) for sFasL in human sera was established. Sera from healthy persons did not contain a detectable level of sFasL, whereas those from patients with large granular lymphocytic (LGL) leukemia and natural killer (NK) cell lymphoma did. These malignant cells constitutively expressed FasL, whereas peripheral NK cells from healthy persons expressed FasL only on activation. These results suggested that the systemic tissue damage seen in most patients with LGL leukemia and NK-type lymphoma is due to sFasL produced by these malignant cells. Neutralizing anti-FasL antibodies or matrix metalloproteinase inhibitors may be of use in modulating such tissue damage.


Assuntos
Glicoproteínas de Membrana/sangue , Animais , Anticorpos Monoclonais , Sequência de Bases , Primers do DNA , Ensaio de Imunoadsorção Enzimática , Proteína Ligante Fas , Humanos , Técnicas In Vitro , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Leucemia de Células T/sangue , Leucemia de Células T/imunologia , Ligantes , Ativação Linfocitária , Linfoma/sangue , Linfoma/genética , Linfoma/imunologia , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/imunologia , Metaloendopeptidases/metabolismo , Camundongos , Dados de Sequência Molecular , Testes de Neutralização , Solubilidade , Linfócitos T/imunologia , Linfócitos T/metabolismo , Transformação Genética
16.
Clin Transl Oncol ; 23(2): 418-423, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32533317

RESUMO

PURPOSE: Tryptophan metabolites have immunomodulatory functions, suggesting possible roles in cancer immunity. METHODS: Plasma tryptophan metabolites were measured using liquid chromatography/mass spectrometry before immune checkpoint inhibitors (ICIs) in patients with non-small cell lung cancer (NSCLC). RESULTS: The 19 patients with NSCLC had significantly lower levels of tryptophan (p = 0.002) and xanthurenic acid (p = 0.032), and a significantly higher level of 3-hydroxyanthranilic acid (3-HAA) (p = 0.028) compared with the 10 healthy volunteers. The patients achieving objective responses had significantly lower levels of 3-HAA than those who did not (p = 0.045). Receiver operating characteristic analyses determined that the cutoff value of 3-HAA for objective response was 35.4 pmol/mL (sensitivity: 87.5% and specificity: 83.3%). The patients with 3-HAA < 35.4 pmol/mL had significantly longer median progression-free survival (7.0 months) than those without (1.6 months, p = 0.022). CONCLUSIONS: Tryptophan metabolites may have a potential for predicting the efficacy of ICIs. REGISTRATION NUMBER: University Hospital Medical Information Network Clinical Trial Registry 000026140.


Assuntos
Ácido 3-Hidroxiantranílico/análise , Carcinoma Pulmonar de Células não Pequenas/sangue , Inibidores de Checkpoint Imunológico/uso terapêutico , Neoplasias Pulmonares/sangue , Triptofano/sangue , Xanturenatos/sangue , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígeno B7-H1/sangue , Antígeno B7-H1/metabolismo , Biomarcadores/sangue , Biomarcadores/metabolismo , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/mortalidade , Feminino , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/mortalidade , Masculino , Pessoa de Meia-Idade , Intervalo Livre de Progressão , Estudos Prospectivos , Curva ROC , Análise de Regressão , Sensibilidade e Especificidade , Resultado do Tratamento , Triptofano/metabolismo
17.
J Exp Med ; 179(3): 873-9, 1994 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-7509364

RESUMO

Fas is a 45-kD cell surface protein belonging to the tumor necrosis factor/nerve growth factor receptor family, and transduces the signal for apoptosis. The cytotoxic T lymphocyte (CTL) hybridoma, PC60-d10S requires the presence of Fas on target cells to induce cytolysis in target cells. This CTL cell line was weakly but specifically stained by a chimeric protein that consisted of the extracellular domain of mouse Fas and the Fc portion of human immunoglobulin G1 (mFas-Fc). Moreover, mFas-Fc inhibited the cytotoxic activity of PC60-d10S. Sublines of d10S that were stained intensively by mFas-Fc were isolated by repetitive fluorescence-activated cell sorter sorting. A cell-surface protein of about 40 kD was specifically precipitated by mFas-Fc from the lysates of these sublines. This protein was homogeneously purified by sequential affinity chromatographies using mFas-Fc and concanavalin A beads. The purified protein exhibited cytotoxic activity against cells expressing Fas but not to the cells which do not express Fas. These results indicated that the 40-kD membrane glycoprotein expressed on PC60-d10S cells is the Fas-ligand that induces the apoptotic signal by binding to Fas.


Assuntos
Antígenos de Superfície/metabolismo , Apoptose , Glicoproteínas de Membrana/isolamento & purificação , Proteínas de Membrana/isolamento & purificação , Linfócitos T Citotóxicos/imunologia , Animais , Antígenos de Superfície/biossíntese , Sequência de Bases , Cromatografia de Afinidade , Concanavalina A , Primers do DNA , Eletroforese em Gel de Poliacrilamida , Humanos , Hibridomas/imunologia , Fragmentos Fc das Imunoglobulinas/análise , Fragmentos Fc das Imunoglobulinas/metabolismo , Imunoglobulina G/análise , Imunoglobulina G/metabolismo , Glicoproteínas de Membrana/metabolismo , Glicoproteínas de Membrana/fisiologia , Proteínas de Membrana/metabolismo , Camundongos , Dados de Sequência Molecular , Peso Molecular , Reação em Cadeia da Polimerase , Proteínas Recombinantes de Fusão/análise , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/metabolismo , Transdução de Sinais , Receptor fas
18.
J Exp Med ; 181(2): 485-91, 1995 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-7530758

RESUMO

Fas is a cell surface protein that mediates apoptosis. A mouse mutant, lpr (lymphoproliferation), has a mutation in the Fas gene. In this report, we studied the expression and function of Fas in various subpopulations of mouse thymocytes. Abundant expression of Fas was detected on CD4+CD8+ double positive as well as CD4+ or CD8+ single positive thymocytes in wild-type mice. Little or low levels of Fas were expressed in CD4-CD8- double negative thymocytes except for the CD4-CD8-CD3+ phenotype, which expresses Fas as abundantly as double positive or single positive subsets. On the other hand, no Fas expression was detected in any population of thymocytes from lpr mice. When the wild-type thymocytes were treated with the agonistic anti-Fas antibody, double positive cells from the wild-type mice were selectively killed by apoptosis, whereas, the single positive cells were resistant to its cytolytic activity despite their abundant expression of Fas. Unlike the apoptosis of thymocytes induced by glucocorticoid or T cell activator, the Fas-induced apoptosis of thymocytes was enhanced by metabolic inhibitors such as cycloheximide. Furthermore, intraperitoneal administration of the anti-Fas antibody into mice caused rapid apoptosis of thymocytes in vivo.


Assuntos
Antígenos de Superfície/fisiologia , Apoptose , Antígenos CD4 , Antígenos CD8 , Timo/citologia , Animais , Antígenos de Superfície/genética , Antígenos de Superfície/imunologia , Diferenciação Celular , Células Cultivadas , Cricetinae , Citotoxicidade Imunológica , Camundongos , Camundongos Endogâmicos BALB C , Timo/imunologia , Timo/metabolismo , Receptor fas
19.
J Exp Med ; 186(12): 2045-50, 1997 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-9396774

RESUMO

It has been believed that the Fas expressed on human peripheral blood T cells (PBT) is nonfunctional, because these cells are insensitive to agonistic anti-Fas/Apo-1 mAbs that efficiently kill in vitro-activated T cells and many Fas-expressing cell lines. Here, we demonstrate that membrane-bound Fas ligand (FasL) kills both fresh and in vitro-activated PBT, indicating that the Fas expressed on fresh PBT is functional. In contrast, soluble FasL kills only the latter. Naive T cells in umbilical cord blood do not express Fas, but can be induced to express Fas by IFN-gamma or by a combination of IL-2 and anti-CD28 mAb, after which they acquire sensitivity to membrane but not to soluble FasL. Soluble FasL inhibited the killing of fresh PBT by membrane FasL. These results indicate that the shedding of FasL from the membrane is a mechanism for downregulating at least part of its killing activity.


Assuntos
Apoptose , Glicoproteínas de Membrana/farmacologia , Linfócitos T/efeitos dos fármacos , Receptor fas/farmacologia , Adulto , Anticorpos Monoclonais/farmacologia , Apoptose/efeitos dos fármacos , Antígenos CD28/imunologia , Proteína Ligante Fas , Humanos , Interferon gama/farmacologia , Interleucina-2/farmacologia , Ligantes , Solubilidade , Linfócitos T/citologia
20.
J Exp Med ; 176(2): 351-61, 1992 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-1380060

RESUMO

The preadipose cell line, PA6, can support long-term hemopoiesis. Frequency of the hemopoietic stem cells capable of sustaining hemopoiesis in cocultures of bone marrow cells and PA6 cells for 6 wk was 1/5.3 x 10(4) bone marrow cells. In the group of dishes into which bone marrow cells had been inoculated at 2.5 x 10(4) cells/dish, 3 of 19 dishes (16%) contained stem cells capable of reconstituting erythropoiesis of WBB6F1-W/Wv mice, indicating that PA6 cells can support the proliferation of primitive hemopoietic stem cells. When the cocultures were treated with an antagonistic anti-c-kit monoclonal antibody, ACK2, only a small number of day 12 spleen colony-forming units survived; and hemopoiesis was severely reduced. However, when the cocultures were continued with antibody-free medium, hemopoiesis dramatically recovered. To examine the proliferative properties of the ACK2-resistant stem cells, we developed a colony assay system by modifying our coculture system. Sequential observations of the development of individual colonies and their disappearance demonstrated that the stem cells having higher proliferative capacity preferentially survive the ACK2 treatment. Furthermore, cells of subclones of the PA6 clone that were incapable of supporting long-term hemopoiesis expressed mRNA for the c-kit ligand. These results suggest that a mechanism(s) other than that involving c-kit receptor and its ligand plays an important role in the survival and proliferation of primitive hemopoietic stem cells.


Assuntos
Células-Tronco Hematopoéticas/citologia , Proteínas Proto-Oncogênicas/metabolismo , Receptores de Superfície Celular/metabolismo , Animais , Anticorpos Monoclonais/imunologia , Sequência de Bases , Células da Medula Óssea , Divisão Celular , Linhagem Celular , Células Clonais , DNA , Hematopoese , Células-Tronco Hematopoéticas/metabolismo , Cinética , Ligantes , Camundongos , Camundongos Endogâmicos C57BL , Dados de Sequência Molecular , Proteínas Proto-Oncogênicas/imunologia , Proteínas Proto-Oncogênicas c-kit , Receptores de Superfície Celular/imunologia
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