RESUMO
Echocardiography has emerged as an essential tool to guide targeted, transcatheter biopsy of cardiac masses. Options for imaging include transthoracic or transesophageal echocardiography and intracardiac echocardiography, with appropriate use being dictated by specific patient characteristics and institutional experience. The authors present a case of three-dimensional (3-D) transesophageal echocardiography-guided transcatheter biopsy of a right ventricular mass and review the current use of echocardiography to guide these procedures.
Assuntos
Adenocarcinoma de Pulmão/secundário , Cateterismo Cardíaco/métodos , Ecocardiografia Transesofagiana/métodos , Neoplasias Cardíacas/diagnóstico , Biópsia Guiada por Imagem/métodos , Neoplasias Pulmonares/patologia , Adenocarcinoma de Pulmão/diagnóstico , Idoso , Feminino , Neoplasias Cardíacas/secundário , Ventrículos do Coração , Humanos , Metástase NeoplásicaRESUMO
BACKGROUND: Haemolytic disease of the fetus and newborn (HDFN) occurs when maternal IgG alloantibodies to fetal red blood cell antigens cross the placenta, causing haemolysis in the fetus and/or neonate. After delivery, the main concern is hyperbilirubinaemia, which can cause neurological damage. OBJECTIVES: To summarise our current management and outcome data to inform health-care professionals counselling women whose pregnancies are at risk of HDFN and to compare these data with relevant studies. METHODS: This is a retrospective descriptive study of all high-risk pregnancies at risk of HDFN at Guy's and St. Thomas' NHS Foundation Trust (GSTFT) Maternity Unit over a 7-year period. We defined high-risk pregnancies as those in whom anti-D, anti-c, anti-K or high (>32 or doubling strength) titres of all other antibodies were identified. RESULTS: A total of 130 pregnancies in 112 women were followed up. A single alloantibody was found in 93 pregnancies (71.5%) and multiple alloantibodies in 37 pregnancies (28.5%). Anti-D was most commonly encountered (n = 48, 36.9%), followed by anti-c (n = 31, 23.8%) and anti-E (n = 15, 11.5%). In 65 of 130 pregnancies (50%), antibody concentrations triggered scans to screen for fetal anaemia. Of 130 pregnancies, 6 (4.6%) required intrauterine transfusions, and 31 of 130 (26%) neonates required post-natal intervention. Overall, morbidity was 0.1% and mortality 0.002%. CONCLUSIONS: This study demonstrates that morbidity and mortality caused by HDFN is minimal. These results are reassuring for women at risk of HDFN as even severely affected cases are successfully managed in most instances. Further studies are needed to identify predictors of disease severity.
Assuntos
Eritroblastose Fetal/sangue , Eritroblastose Fetal/prevenção & controle , Transfusão Feto-Materna/sangue , Imunoglobulina G/sangue , Isoanticorpos/sangue , Adulto , Eritroblastose Fetal/mortalidade , Feminino , Transfusão Feto-Materna/mortalidade , Transfusão Feto-Materna/prevenção & controle , Seguimentos , Humanos , Recém-Nascido , Masculino , GravidezRESUMO
OBJECTIVES: To demonstrate the feasibility and efficacy of the Google Glass as a tool to improve patient satisfaction and patient-physician communication for otolaryngology residents in the outpatient clinic setting. The primary outcome of the study was to improve patient satisfaction scores based on physician communication-related questions from Consumer Assessment of Healthcare Providers and Systems (CAHPS) surveys. STUDY DESIGN: Prospective randomised trial. SETTING: Tertiary care hospital. SUBJECT AND METHODS: To evaluate the effect on patient satisfaction, five residents were recorded using the Google Glass in an outpatient clinic setting by 50 randomised patients. Modified surveys based on the CG-CAHPS survey were completed by patients at the conclusion of each clinic encounter. The recorded videos were evaluated by two independent faculties. Summarised data and video were distributed to each resident for review as the intervention. The residents were recorded again by 45 additional patients with evaluation by patients and faculties. RESULTS: After intervention, the scores from faculty surveys regarding patient satisfaction including the subject of better explanations (P > 0.001), listening carefully (P > 0.001), addressing patient questions (P > 0.001), displaying respect (P > 0.001) and spending adequate time (P = 0.0005) all significantly improved, as well as overall performance (P = 0.014). The scores from patient surveys did significantly improve. CONCLUSION: This study demonstrates the improvements in patient satisfaction and patient-physician communication can be achieved with the use of Google Glass as a first-person recording device in the outpatient otolaryngology clinic setting.
Assuntos
Óculos , Internato e Residência , Microcomputadores , Otolaringologia/educação , Satisfação do Paciente , Relações Médico-Paciente , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Assistência Ambulatorial , Educação de Pós-Graduação em Medicina , Estudos de Viabilidade , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Projetos Piloto , Estudos Prospectivos , Inquéritos e Questionários , Gravação em VídeoRESUMO
OBJECTIVE: To describe patterns of clinical bleeding in neonates with severe thrombocytopenia (ST and platelet count <60 × 10(9) L(-1)), and to investigate the factors related to bleeding. STUDY DESIGN: Seven tertiary-level neonatal units enrolled neonates (n = 169) with ST. Data were collected prospectively on all clinically apparent haemorrhages. Relationships between bleeding, platelet count and baseline characteristics were explored through regression analysis. RESULTS: Bleeding was recorded in most neonates with ST (138/169; 82%), including 123 neonates with minor bleeding and 15 neonates with major bleeding. The most common sites of minor bleeding were from the renal tract (haematuria 40%), endotracheal tube (21%), nasogastric tube (10%) and skin (15%). Gestational age <34 weeks, development of ST within 10 days of birth and necrotizing enterocolitis were the strongest predictors for an increased number of bleeding events. For neonates with ST, a lower platelet count was not a strong predictor of increased bleeding. CONCLUSIONS: The majority of neonates with ST bleed, although most episodes are minor. These findings establish the importance of clinical factors for bleeding risk, rather than minimum platelet count. Further studies should assess the clinical significance of different types of minor bleed for neonatal outcomes, the predictive value of minor bleeding for major bleeding and the role of platelet transfusions in preventing bleeding.
Assuntos
Hematúria/prevenção & controle , Transfusão de Plaquetas , Trombocitopenia/terapia , Feminino , Idade Gestacional , Hematúria/congênito , Humanos , Lactente , Recém-Nascido , Doenças do Recém-Nascido/terapia , Masculino , Estudos Prospectivos , Trombocitopenia/congênitoRESUMO
This paper calls into question whether and how end-of-life care pathways facilitate the accomplishment of a 'good death'. Achieving a 'good death' is a prominent social and political priority and an ideal which underpins the philosophy of hospice and palliative care. End-of-life care pathways have been devised to enhance the care of imminently dying patients and their families across care settings and thereby facilitate the accomplishment of a 'good death'. These pathways have been enthusiastically adopted and are now recommended by governments in the UK as 'best practice' templates for end-of-life care. However, the literature reveals that the 'good death' is a nebulous, fluid concept. Moreover, concerns have been articulated regarding the efficacy of care pathways in terms of their impact on patient care and close analysis of two prominent end-of-life pathways reveals how biomedical aspects of care are privileged. Nonetheless drawing on a diverse range of evidence the literature indicates that end-of-life care pathways may facilitate a certain type of 'good death' and one which is associated with the dying process and framed within biomedicine.
Assuntos
Atitude Frente a Morte , Procedimentos Clínicos/normas , Cuidados Paliativos/organização & administração , Assistência Terminal/organização & administração , Humanos , Reino UnidoRESUMO
K46J B lymphomas express a T cell costimulatory activity that is not inhibited by CTLA-4Ig, anti-B7-1, anti-B7-2, anti-intercellular adhesion molecule 1 or antibodies to heat stable antigen. In this paper we report that this costimulatory activity is mediated at least in part by 4-1BB ligand, a member of the tumor necrosis factor (TNF) gene family that binds to 4-1BB, a T cell activation antigen with homology to the TNF/nerve growth factor receptor family. A fusion protein between 4-1BB and alkaline phosphatase (4-1BB-AP) blocks T cell activation by K46J lymphomas in both an antigen-specific system and with polyclonally (anti-CD3) activated T cells. 4-1BB-AP also blocks antigen presentation by normal spleen cells. When the antigen-presenting cells express B7 molecules as well as 4-1BB ligand, we find that B7 molecules and 4-1BB-AP both contribute to T cell activation. These data suggest that 4-1BB ligand plays an important role in costimulation of IL-2 production and proliferation by T cells. The B lymphoma M12 expresses low levels of 4-1BB-L but can be induced to express higher levels by treatment of the B cells with cAMP, which also induces B7-1 and B7-2 in these cells. Thus cAMP appears to coordinately induce several costimulatory molecules on B cells.
Assuntos
Antígeno B7-1/fisiologia , AMP Cíclico/farmacologia , Imunoconjugados , Ativação Linfocitária , Receptores de Fator de Crescimento Neural/fisiologia , Linfócitos T/imunologia , Fator de Necrose Tumoral alfa/fisiologia , Ligante 4-1BB , Abatacepte , Fosfatase Alcalina/fisiologia , Animais , Apresentação de Antígeno , Antígenos CD , Antígenos de Diferenciação/fisiologia , Antígeno CTLA-4 , Linhagem Celular , Feminino , Ligantes , Linfoma de Células B/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Regulação para CimaRESUMO
4-1BB ligand (4-1BBL) is a member of the tumor necrosis factor (TNF) family expressed on activated antigen-presenting cells. Its receptor, 4-1BB, is a member of the TNF receptor family expressed on activated CD4 and CD8 T cells. We have produced a soluble form of 4-1BBL using the baculovirus expression system. When coimmobilized on plastic with anti-CD3, soluble 4-1BBL induces interleukin (IL)-2 production by resting CD28+ or CD28- T cells, indicating that 4-1BBL can function independently of other cell surface molecules, including CD28, in costimulation of resting T cell activation. At low concentrations of anti-CD3, 4-1BBL is inferior to anti-CD28 in T cell activation. However, when 4-1BB ligand is provided together with strong TCR signals, then 4-1BBL and anti-CD28 are equally potent in stimulation of IL-2 production by resting T cells. We find that TNF receptor-associated factor (TRAF)1 or TRAF2 associate with a glutathione S-transferase-4-1BB cytoplasmic domain fusion protein in vitro. In T cells, we find that association of TRAF1 and TRAF2 with 4-1BB requires 4-1BB cross-linking. In support of a functional role for TRAF2 in 4-1BB signaling, we find that resting T cells isolated from TRAF2-deficient mice or from mice expressing a dominant negative form of TRAF2 fail to augment IL-2 production in response to soluble 4-1BBL. Thus 4-1BB, via the TRAF2 molecule, can provide CD28-independent costimulatory signals to resting T cells.
Assuntos
Antígenos CD28/imunologia , Proteínas/imunologia , Receptores de Fator de Crescimento Neural/imunologia , Receptores do Fator de Necrose Tumoral/imunologia , Linfócitos T/imunologia , Animais , Anticorpos/imunologia , Antígenos CD , Linhagem Celular , Vetores Genéticos , Interleucina-2/biossíntese , Ligantes , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Nucleopoliedrovírus , Proteínas/genética , Fator 1 Associado a Receptor de TNF , Fator 2 Associado a Receptor de TNF , Membro 9 da Superfamília de Receptores de Fatores de Necrose TumoralRESUMO
The hemolytic effect of diltiazem (a calcium channel blocker) on red blood cells (RBCs) exposed to varying osmolarity was investigated. Previous work on the in vitro hemolytic effect of verapamil shed some light on the potential for some drugs to induce hemolysis at moderate-to-high doses and in pre-existing osmotic stress. The current experimental approach used a modified red cell hemolysis assay with concentrations of diltiazem ranging from 50-1500 µM compared to drug free controls. The time-course of hemolytic effects was also investigated. In conditions representing decreasing osmolarity (dilution from 140 to 0 mM NaCl) there was a significant increase in erythrocyte hemolysis that was also dependent on diltiazem concentration (ANOVA, p<0.05). The red cells also showed a significantly increased rate of hemolysis over 5 h with increasing concentration of diltiazem (ANOVA, p<0.05). Overall the data suggested that diltiazem can cause hemolysis of RBCs in a predictable time- and concentration-dependent manner, and that diltiazem increases the fragility of the erythrocytes further during hypotonic osmotic stress. The mechanism of diltiazem-dependent hemolysis could involve various ion transport pathways (i.e. Ca pump, Ca Channels) and subsequent effects on cell volume control or membrane fragility.
Assuntos
Bloqueadores dos Canais de Cálcio/toxicidade , Diltiazem/toxicidade , Eritrócitos/efeitos dos fármacos , Hemólise/efeitos dos fármacos , Cavalos/fisiologia , Soluções Hipotônicas/efeitos adversos , Animais , Relação Dose-Resposta a Droga , Concentração OsmolarRESUMO
Embryoid bodies formed from teratocarcinoma stem cells differentiate an outer layer consisting of parietal and visceral endoderm or of visceral endoderm exclusively. We have previously shown that when these embryoid bodies are plated on collagen-coated substrates a parietal endoderm-like cell migrates onto the substrate, whereas all of the visceral endoderm remains associated with the stem cell mass, suggesting a role for substrate contact in parietal endoderm differentiation. We now identify fibronectin as the migration-promoting component in these cultures, and note that laminin and collagen type IV are 10-fold less effective at promoting both attachment and endoderm outgrowth. The RGDS tetrapeptide (arg-gly-asp-ser) from the cell attachment domain of fibronectin can specifically block attachment and outgrowth on both fibronectin- and laminin-coated substrates. In addition, the involvement of the 140-kD fibronectin receptor is demonstrated using an antibody directed against this molecule.
Assuntos
Endoderma/patologia , Matriz Extracelular/fisiologia , Teratoma/patologia , Animais , Adesão Celular , Diferenciação Celular , Movimento Celular , Células Cultivadas , Colágeno/fisiologia , Desenvolvimento Embrionário e Fetal , Fibronectinas/fisiologia , Laminina/fisiologia , Camundongos , Oligopeptídeos/farmacologia , Receptores de Fibronectina , Receptores Imunológicos/imunologia , Receptores Imunológicos/fisiologiaRESUMO
As is the case with most proteins of the secretory pathway, the biogenesis of MHC class I and class II molecules occurs in association with molecular chaperones. Considerable progress has been made in identifying the chaperones involved and recent studies on two of these, calnexin and invariant chain, have shown that they influence multiple processes including protein stability, folding, assembly and intercellular retention.
Assuntos
Apresentação de Antígeno/imunologia , Chaperonas Moleculares/imunologia , Animais , HumanosRESUMO
CD28 is the primary co-stimulatory receptor for inducing high-level IL-2 production and survival of naïve CD4(+) T cells. While no other cell surface receptor can be considered fully redundant with CD28, recent developments suggest that additional co-stimulatory pathways have preferential effects at different stages of T cell activation, on different subsets of T cells or contribute to the development of different effector functions.
Assuntos
Antígenos CD28/metabolismo , Glicoproteínas de Membrana , Linfócitos T/imunologia , Animais , Antígenos CD/metabolismo , Antígeno CD24 , Moléculas de Adesão Celular/metabolismo , Humanos , Molécula 1 de Adesão Intercelular/metabolismo , Ativação Linfocitária , Antígeno-1 Associado à Função Linfocitária/metabolismo , Receptores de Fator de Crescimento Neural/metabolismo , Receptores OX40 , Receptores do Fator de Necrose Tumoral/metabolismo , Transdução de Sinais , Membro 7 da Superfamília de Receptores de Fatores de Necrose Tumoral/metabolismo , Membro 9 da Superfamília de Receptores de Fatores de Necrose TumoralRESUMO
We report the identification and control of an outbreak of a ciprofloxacin-susceptible strain of UK epidemic meticillin-resistant Staphylococcus aureus (EMRSA)-15 on a neonatal unit (NNU). All babies were screened for MRSA on admission using ciprofloxacin-containing media which did not detect the outbreak strain. The first identified case was a premature baby who developed MRSA bacteraemia with associated tibial osteomyelitis and multiple subcutaneous abscesses. The outbreak strain was subsequently identified in the nasopharyngeal secretions of a second child who was not clinically infected. Screening of all patients on the NNU using non-ciprofloxacin-media identified two other colonised babies. All four patient isolates were EMRSA-15, spa type t022, SCCmec IV, Panton-Valentine leucocidin (PVL) negative, indistinguishable by pulsed-field gel electrophoresis and susceptible to all non-beta-lactam antimicrobials tested. The outbreak strain was cultured from four of 48 environmental sites in a communal milk-expressing room. Unsupervised movement of mothers to and from the milk-expressing room may have contributed to the outbreak. Control measures included cohort isolation of affected babies, improved environmental cleaning, increased emphasis on hand hygiene and education of mothers. Ciprofloxacin-containing media should be used with caution for MRSA screening in settings where ciprofloxacin-susceptible strains (including community-associated MRSA) are increasing in prevalence.
Assuntos
Antibacterianos/farmacologia , Ciprofloxacina/farmacologia , Infecção Hospitalar/epidemiologia , Surtos de Doenças , Farmacorresistência Bacteriana , Eritromicina/farmacologia , Infecções Estafilocócicas/epidemiologia , Staphylococcus aureus/efeitos dos fármacos , Abscesso/microbiologia , Bacteriemia/microbiologia , Técnicas de Tipagem Bacteriana , Técnicas Bacteriológicas/métodos , Portador Sadio/microbiologia , Infecção Hospitalar/diagnóstico , Infecção Hospitalar/microbiologia , Meios de Cultura/química , Educação , Microbiologia Ambiental , Feminino , Genótipo , Desinfecção das Mãos , Zeladoria Hospitalar , Humanos , Lactente , Recém-Nascido , Londres/epidemiologia , Testes de Sensibilidade Microbiana , Nasofaringe/microbiologia , Osteomielite/microbiologia , Isolamento de Pacientes , Fenótipo , Infecções Estafilocócicas/diagnóstico , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/isolamento & purificaçãoRESUMO
A mutant clone (AU-100) which is 90% deficient in adenylosuccinate synthetase activity was characterized from wild-type murine S49 T-lymphoma cells. This AU-100 cell line and its hypoxanthine-guanine phosphoribosyltransferase-deficient derivative, AUTG-50B, overproduce purines severalfold and excrete massive amounts of inosine into the culture medium (Ullman et al., Proc. Natl. Acad. Sci. U.S.A. 79:5127-5131, 1982). We introduced a mutation into both of these cell lines which make them incapable of taking up nucleosides from the culture medium. The genetic deficiency in nucleoside transport prevents the adenylosuccinate synthetase-deficient AU-100 cells from excreting inosine. Because of an extremely efficient intracellular inosine salvage system, the nucleoside transport-deficient AU-100 cells also no longer overproduce purines. AUTG-50B cells which have been made genetically deficient in nucleoside transport still overproduce purines but excrete hypoxanthine rather than inosine. These studies demonstrate genetically that nucleoside transport and nucleoside efflux share a common component and that nucleoside transport has an important regulatory function which profoundly affects the rates of purine biosynthesis and purine salvage.
Assuntos
Inosina/metabolismo , Nucleosídeos/metabolismo , Purinas/biossíntese , Adenilossuccinato Sintase/deficiência , Animais , Transporte Biológico Ativo , Linhagem Celular , Genes , Hipoxantina Fosforribosiltransferase/deficiência , Camundongos , Mutação , Nucleosídeos/genética , Linfócitos T/metabolismoRESUMO
The haemolytic effect of verapamil on red blood cells (RBCs) exposed to varying osmolarity was investigated. The experimental approach used a modified red cell haemolysis assay with concentrations of verapamil ranging from 50-1500 microM compared to drug free controls. The time-course of haemolytic effects was also investigated. We also briefly determined the haemolytic effects of verapamil in Ca2+-free conditions (with added EGTA). In conditions representing decreasing osmolarity (dilution from 140-0 mM NaCl) there was a significant increase in erythrocyte haemolysis that was also dependent on verapamil concentration (ANOVA, p<0.05). The red cells also showed a significantly increased rate of haemolysis over 5 h with increasing verapamil concentration (ANOVA, p<0.05). The degree of RBC hypotonic haemolysis was significantly increased in a Ca2+-free medium (+EGTA) compared to normal saline and this effect was exacerbated by additions of verapamil (ANOVA, p<0.05). Overall the data suggested that verapamil can cause haemolysis of RBCs in a predictable time- and concentration-dependent manner, and that verapamil increases the fragility of the erythrocytes further during hypotonic osmotic stress and Ca2+-free conditions. The mechanism of verapamil-dependent haemolysis could be directly related to the observed biphasic concentration-effect and could consequently involve several ion transport pathways.
Assuntos
Bloqueadores dos Canais de Cálcio/toxicidade , Eritrócitos/efeitos dos fármacos , Hemólise/efeitos dos fármacos , Verapamil/toxicidade , Animais , Cálcio/metabolismo , Cálcio/fisiologia , Quelantes/farmacologia , Relação Dose-Resposta a Droga , Ácido Egtázico/farmacologia , Cavalos , Soluções Hipotônicas , Técnicas In Vitro , Cinética , Concentração OsmolarRESUMO
PURPOSE: Previous studies have indicated that health beliefs are related to the periodontal disease status and treatment behaviour of patients. However, it is possible that treatment may affect a patient's health beliefs and thus complicate this issue. The present study therefore looked for changes in health control beliefs and oral health impacts in patients undergoing periodontal treatment in a dental school. MATERIALS AND METHODS: Questionnaires assessing dental multidimensional locus of control (LOC) and oral health impact profile (OHIP) were posted to subjects due to attend for initial periodontal consultation and were returned by 127 patients who attended. Repeat questionnaires were sent to all subjects 6 months later when they had received some oral hygiene instruction, scaling and root planing, and 55 were returned. RESULTS: Comparison of data for those subjects who completed both questionnaires showed no difference in LOC but showed a trend (p = 0.065) towards reduced OHIP (i.e. improved oral health-related quality of life). CONCLUSIONS: These subjects apparently did not alter their health control beliefs about periodontal disease as a result of treatment, but there may have been an improvement in their oral health-related quality of life. Further studies are required to confirm these possibilities.
Assuntos
Conhecimentos, Atitudes e Prática em Saúde , Controle Interno-Externo , Doenças Periodontais/terapia , Qualidade de Vida/psicologia , Adulto , Raspagem Dentária , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Aplainamento Radicular , Estatísticas não Paramétricas , Inquéritos e Questionários , Fatores de TempoRESUMO
The induction of long-lived heterotypic T-cell protection against influenza virus remains elusive, despite the conservation of T-cell epitopes. T-cell protection against influenza is critically dependent on lung-resident memory T cells (Trm). Here we show that intranasal administration of 4-1BBL along with influenza nucleoprotein in a replication-defective adenovirus vector to influenza pre-immune mice induces a remarkably stable circulating effector memory CD8 T-cell population characterized by higher IL-7Rα expression than control-boosted T cells, as well as a substantial lung parenchymal CD69+ CD8 Trm population, including both CD103+ and CD103- cells. These T-cell responses persist to greater than 200 days post-boost and protect against lethal influenza challenge in aged (year old) mice. The expansion of the nucleoprotein-specific CD8 Trm population during boosting involves recruitment of circulating antigen-specific cells and is critically dependent on local rather than systemic administration of 4-1BBL as well as on 4-1BB on the CD8 T cells. Moreover, during primary influenza infection of mixed bone marrow chimeras, 4-1BB-deficient T cells fail to contribute to the lung-resident Trm population. These findings establish both endogenous and supraphysiological 4-1BBL as a critical regulator of lung-resident memory CD8 T cells during influenza infection.
Assuntos
Ligante 4-1BB/administração & dosagem , Vírus da Influenza A/imunologia , Vacinas contra Influenza/imunologia , Pulmão/patologia , Infecções por Orthomyxoviridae/imunologia , Subpopulações de Linfócitos T/imunologia , Linfócitos T Reguladores/imunologia , Administração Intranasal , Animais , Antígenos CD/metabolismo , Antígenos CD8/metabolismo , Células Cultivadas , Epitopos de Linfócito T/metabolismo , Memória Imunológica , Cadeias alfa de Integrinas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de Interleucina-7/metabolismo , Subpopulações de Linfócitos T/virologia , Linfócitos T Reguladores/virologiaRESUMO
In this study we analysed the binding of the peptide HEL46-61 to purified Ak molecules which have been altered by site-directed mutagenesis at polymorphic positions to include amino acids from the Ad alpha-chain. We find that changes in the floor of the peptide binding groove, at positions 11, 14 and 28, abolish T cell recognition without changing peptide binding affinity. We further show that amino acid changes at these positions in the Ad molecule result in a conformationally altered molecule as evidenced by loss of binding of the Ad alpha specific monoclonal antibody K24. Thus the T cell receptor is highly sensitive to subtle changes in MHC II structure induced at sites that are unlikely to be involved in direct T cell contact. This has important implications with respect to allorecognition. The binding studies reported here were performed both at pH 7, to reflect binding of peptides at the cell surface, and at pH 5.5, to mimic binding in an intracellular acidic compartment. Binding to wild-type Ak was increased 2-3-fold at pH 5.5, whereas binding to some MHC II mutants was increased by greater than 20-fold at pH 5.5 relative to pH 7. These results show that the apparent peptide binding specificity for the mutants differs at pH 7 and 5.5, and suggest that caution should be used in defining the MHC-restriction of peptide epitopes at neutral pH.
Assuntos
Antígenos/imunologia , Antígenos de Histocompatibilidade Classe II/imunologia , Linfócitos T/imunologia , Sequência de Aminoácidos , Animais , Células Apresentadoras de Antígenos/imunologia , Linhagem Celular , Galinhas , Citometria de Fluxo , Antígenos de Histocompatibilidade Classe II/química , Concentração de Íons de Hidrogênio , Ativação Linfocitária/imunologia , Dados de Sequência Molecular , Muramidase/imunologia , Ligação Proteica/imunologia , Conformação ProteicaRESUMO
The binding of CD40 ligand on activated T cells to CD40 on resting B cells induces the expression of costimulatory molecules B7-1 (CD80) and B7-2 (CD86). The induction of B7 molecules by CD40 ligand-CD40 interaction represents a critical step in rendering B cells competent for antigen presentation. The CBA/N mouse has a defect in CD40 signalling which has been attributed to a mutation in Bruton's tyrosine kinase. We have compared the ability of murine CD40 ligand to induce B7-1 and B7-2 expression on B cells isolated from CBA/N and wild-type CBA/J mice. We find that the CBA/N defect partially impairs both B7-1 and B7-2 induction via CD40. Subsequent experiments investigated the roles of different second messenger systems in B7-1 and B7-2 induction in normal B cells. In M12 B lymphomas either CD40 cross-linking or cAMP treatment can induce B7 molecules. Here we report that treatment with dibutyryl-cAMP also induces B7 molecules in normal B cells provided that they have been preactivated by CD40 cross-linking. We also find that PMA and ionomycin treatment of B cells induces B7-2 but not B7-1 expression. Our data therefore show roles for BTK, cAMP and PMA/ionomycin in B7 induction, as well as providing further evidence for differential regulation of B7-1 and B7-2 induction in B cells.
Assuntos
Antígenos CD/fisiologia , Linfócitos B/fisiologia , Antígeno B7-1/fisiologia , Antígenos CD40/fisiologia , Glicoproteínas de Membrana/fisiologia , Proteínas Tirosina Quinases/metabolismo , Tirosina Quinase da Agamaglobulinemia , Animais , Antígeno B7-2 , Cálcio/fisiologia , AMP Cíclico/fisiologia , Feminino , Ionomicina/farmacologia , Ativação Linfocitária , Masculino , Camundongos , Camundongos Endogâmicos CBA , Proteína Quinase C/fisiologia , Agregação de Receptores , Sistemas do Segundo Mensageiro , Transdução de Sinais , Acetato de Tetradecanoilforbol/farmacologiaRESUMO
Cholinergic toxicity of organophosphate insecticides is regarded as the principal health hazard associated with both human and animal exposures. Recent studies indicate that these pesticides may have important effects on both the immune and hematopoietic systems. In the present study, human bone marrow cells were exposed in vitro to paraoxon and malaoxon (the primary metabolites of parathion and malathion). These compounds produced dose-dependent depression of colony formation by erythrocyte (burst-forming units-erythroid [BFU-E] and colony-forming units-erythroid [CFU-E]) and granulocyte-macrophage progenitors (colony-forming units-granulocyte-macrophage [CFU-GM]). CFU-E colony formation was reduced 15%-57%, by both paraoxon and malaoxon, in the range of 10(-8)-10(-5) M. No effects were seen at 10(-9) and 10(-10) M. Colony formation by BFU-E was reduced 15%-75%, at 10(-9)-10(-5) M organophosphate (OP), then returned to normal at 10(-10) M OP. In comparison to CFU-E, BFU-E appeared to be more sensitive to the suppressive action of OPs. Numbers of CFU-GM colonies were reduced 16%-59% in the range of 10(-9)-10(-5) M OP, then returned to normal at 10(-10) M OP. Choline chloride added to marrow cultures (final concentration, 10 mM) enhanced CFU-GM colony formation at all concentrations of paraoxon and malaoxon. Our results provide a rationale for assessing hematologic parameters in occupationally exposed individuals, and indicate the need to determine both the mechanism and the environmental health consequences of the observed hematopoietic effects.
Assuntos
Células-Tronco Hematopoéticas/citologia , Malation/análogos & derivados , Paraoxon/toxicidade , Medula Óssea/patologia , Células da Medula Óssea , Divisão Celular/efeitos dos fármacos , Ensaio de Unidades Formadoras de Colônias , Relação Dose-Resposta a Droga , Eritrócitos/citologia , Granulócitos/citologia , Células-Tronco Hematopoéticas/efeitos dos fármacos , Humanos , Macrófagos/citologia , Malation/toxicidadeRESUMO
We have investigated the effects of lithium on the proliferative potential of murine megakaryocyte stem cells (CFUMeg) in vitro and on circulating platelet levels in vivo. At optimal levels of megakaryocyte-colony stimulating factor (Meg-CSF) (10%) concentrations of 0.5, 1.0 and 3.0 meq/L augmented CFUMeg numbers. Significant increases of 133% and 125% over control levels were observed at 1.0 and 3.0 meq/L, respectively. At suboptimal concentrations of Meg-CSF (1.0%), Li effected a maximum increase of 200% over control levels at 1.0 meq/L suggesting a greater sensitivity of CFUMeg to stimulatory factors in the presence of Li. To better define the mode of action of Li, heterogenous bone marrow was separated into adherent and non-adherent cell populations and cultured in the presence of Li. Non-adherent cells cultured in the presence of 0.5, 1.0 and 3.0 meq/L Li showed significant increases in CFUMeg over non-adherent cells cultured without Li. These results suggest a direct effect of Li on CFUMeg. Cells cultured from the adherent cell population also responded to Li with enhanced CFUMeg numbers. This response may be a direct stem cell effect or an indirect effect via the production of endogenous Meg-CSF. One meq/L of Li when injected i.p., produced a thrombocytosis from days 4 through 15, with a maximum at 6 days post-Li treatment. These results suggest Li is an effective agent for stimulating megakaryocytopoiesis and has both direct and possibly indirect effects on the megakaryocyte stem cell.