RESUMO
BACKGROUND: qSOFA has been proposed as a prognostic tool in patients with sepsis. This study set out to assess the sensitivity of several scores, namely: the pre-ICU qSOFA, the qSOFA with lactate (qSOFA L), SIRS score, qSOFA + SIRS score (qSIRS) and qSIRS with lactate (qSIRS L) in predicting in-hospital mortality in patients with surgical sepsis as well as the sensitivity of these scores in predicting high-grade sepsis. The secondary aim was to determine which of these scores is best suited to predict high-grade surgical sepsis. METHODS: This was a retrospective cohort study that was conducted between December 2012 and August 2017 in a public metropolitan surgical service. Data from patients aged > 13 years, who were admitted to the hospital and who had an emergency surgical procedure for source control were retrieved from a prospectively maintained hybrid electronic database. The qSOFA, qSOFA plus lactate (qSOFA L), SIRS and qSOFA + SIRS (qSIRS), as well as the qSIRS plus lactate (qSIRS L), were calculated for each patient. A lactate level that was greater than 2mmol/L was deemed to be a positive finding. Any score ≥2 was deemed to be a positive score. The outcome measure was in-hospital mortality. The prognostic value of qSOFA, qSOFA L, SIRS, qSIRS and qSIRS L was studied. Receiver operating characteristic analyses were performed to determine the area under the curve (AUC), sensitivity, specificity and positive and negative likelihood ratios for positive qSOFA, qSOFA L, SIRS, qSIRS, and qSIRS L. Contingency tables were used to calculate the sensitivity, specificity, PPV and NPV for predicting severe or high-grade surgical sepsis. RESULTS: There were a total number of 1884 patients in the sample group of whom 855 were female (45.4%). The median patient age was 36 years (IQR 23-56). A total of 1489 patients (79%) were deemed to have high-grade sepsis based on an advanced EGS AAST grading, whilst 395 patients (21%) had low-grade sepsis. A total of 71 patients died (3.8%). Of these patients who died, 67 (94.4%) had high-grade sepsis and 4 (5.6%) had low-grade sepsis. The mortality rate in the high-grade sepsis group was 4.5%, whilst the mortality rate in the low-grade sepsis group was 1%. The scores with the greatest accuracy in predicting mortality were qSIRS (AUROC 0.731, 95% CI 0.68-0.78), followed by SIRS (AUROC 0.70, 95% CI 0.65-0.75). The qSOFA and qSOFA L were the least accurate in predicting mortality (AUROC 0.684, 95% CI 0.63-0.74 for both). The addition of lactate had no significant effect on the accuracy of the five scores in predicting mortality. Patients with a qSOFA ≥ 2 have an increased risk of dying (OR 5.8), as do patients with a SIRS score ≥2 (OR 2.7). qSIRS L had the highest sensitivity (69%) in predicting the presence of high-grade surgical sepsis, followed by qSIRS (65.5% sensitivity). qSOFA showed a very low sensitivity of only 4.5% and a high specificity of 99.2%. The addition of lactate to the score marginally improved the sensitivity. Lactate of 2mmol/L or more was also an independent predictor of high-grade sepsis. CONCLUSION: The qSIRS score is most accurate in predicting mortality in surgical sepsis. The qSOFA score is inferior to both the SIRS and the qSIRS scores in predicting mortality. The qSIRS score with the addition of lactate to the qSIRS score made it the most sensitive score in predicting high-grade surgical sepsis.
Assuntos
Países em Desenvolvimento , Serviço Hospitalar de Emergência , Escores de Disfunção Orgânica , Sepse/diagnóstico , Sepse/mortalidade , Síndrome de Resposta Inflamatória Sistêmica/diagnóstico , Síndrome de Resposta Inflamatória Sistêmica/mortalidade , Adulto , Área Sob a Curva , Feminino , Mortalidade Hospitalar , Hospitalização , Humanos , Ácido Láctico/sangue , Masculino , Pessoa de Meia-Idade , Prognóstico , Curva ROC , Estudos Retrospectivos , Sensibilidade e Especificidade , Adulto JovemRESUMO
BACKGROUND: Obstetric spinal hypotension is a common and important problem during caesarean delivery. Identifying patients at risk for hypotension may guide clinical decision-making and allow timeous referral. OBJECTIVE: Using preoperative risk factors, to develop a simple scoring system to predict systolic hypotension. METHODS: This prospective, single-centre, observational study of patients undergoing elective or urgent caesarean delivery assessed body mass index, baseline heart rate, baseline mean arterial pressure (MAP), maternal age, urgency of surgery (elective v. non-elective) and preoperative haemoglobin concentration as predictors of spinal hypotension (systolic blood pressure <90 mmHg). We used empirical cut-point estimations in a logistic regression model to develop a scoring system for prediction of hypotension. RESULTS: From 504 eligible patients, preoperative heart rate (odds ratio (OR) 1.02, 95% confidence interval (CI) 1.00 - 1.03; p=0.012), preoperative MAP (OR 0.97, 95% CI 0.95 - 0.98; p<0.001) and maternal age (OR 1.05, 95% CI 1.02 - 1.08; p=0.002) were found to be predictors of hypotension. We derived a preliminary scoring system (pulse rate >90 bpm, age >25 years, MAP <90 mmHg - the PRAM score) for the prediction of systolic hypotension following obstetric spinal anaesthesia. Patients with three factors had a 53% chance of developing hypotension, compared with the overall incidence of 30%. The PRAM score showed good discrimination, with a c-statistic of 0.626 (95% CI 0.576 - 0.676) and good calibration. CONCLUSIONS: Preoperative heart rate, preoperative MAP and maternal age were predictive of hypotension in elective and emergency caesarean delivery. The PRAM score shows promise as a simple, practical means to identify these patients preoperatively, but requires prospective validation.
RESUMO
As excess mucin expression can contribute to the exacerbation of asthma, the present authors hypothesised that Mycoplasma pneumoniae significantly induces MUC5AC (the major airway mucin) expression in airway epithelial cells isolated directly from asthmatic subjects. A total of 11 subjects with asthma and six normal controls underwent bronchoscopy with airway brushing. Epithelial cells were cultured at an air-liquid interface and incubated with and without M. pneumoniae for 48 h, and in the presence and absence of nuclear factor (NF)-kappaB and a toll-like receptor (TLR)2 inhibitor. Quantitative PCR was performed for MUC5AC and TLR2 mRNA. MUC5AC protein and total protein were determined by ELISA. M. pneumoniae exposure significantly increased MUC5AC mRNA and protein expression after 48 h in epithelial cells isolated from asthmatic, but not from normal control subjects, at all concentrations as compared to unexposed cells. TLR2 mRNA expression was significantly increased in asthmatic epithelial cells at 4 h compared with unexposed cells. NF-kappaB and TLR2 inhibition reduced MUC5AC expression to the level of the unexposed control in both groups. Mycoplasma pneumoniae exposure significantly increased MUC5AC mRNA and protein expression preferentially in airway epithelial cells isolated from asthmatic subjects. The toll-like receptor 2 pathway may be involved in this process.
Assuntos
Asma/metabolismo , Células Epiteliais/microbiologia , Regulação da Expressão Gênica , Mucinas/biossíntese , Mycoplasma pneumoniae/metabolismo , Adulto , Asma/imunologia , Asma/microbiologia , Broncoscopia/métodos , Ensaio de Imunoadsorção Enzimática , Células Epiteliais/citologia , Feminino , Humanos , Masculino , Modelos Biológicos , Mucina-5AC , Mucinas/metabolismo , Mycoplasma pneumoniae/fisiologia , NF-kappa B/metabolismo , Fatores de Tempo , Receptor 2 Toll-Like/metabolismoRESUMO
The 5-Lipoxygenase pathway results in the formation of leukotrienes, including leukotriene B(4) (LTB(4)), 5-oxo-6E,8Z,11Z,14Z-eicosatetranoic acid and the cysteinyl leukotrienes (LTC(4), LTD(4) and LTE(4)) and activates all four leukotriene receptors, BLT1, BLT2, cysLT(1) and cysLT(2). Zileuton is the only commercially available inhibitor of the 5-Lipoxygenase pathway. In a number of clinical trials, zileuton has been shown to improve airway function and inflammation, asthma symptom control and quality of life in asthmatics. Given the important role that leukotrienes play in airway inflammation, zileuton provides an additional therapeutic option in the management of chronic, persistent asthma, particularly those asthmatics with more severe disease. In addition, zileuton has shown promise in a number of other conditions, including upper airway inflammatory conditions, dermatological disease and chronic obstructive pulmonary disease. The development of new formulations, including a controlled release tablet formulation for b.i.d. dosing and an intravenous preparation for acute asthma exacerbations may enhance clinical utility and expand therapeutic indications.
Assuntos
Anti-Inflamatórios não Esteroides/uso terapêutico , Asma/tratamento farmacológico , Hidroxiureia/análogos & derivados , Inibidores de Lipoxigenase/uso terapêutico , Anti-Inflamatórios não Esteroides/farmacocinética , Anti-Inflamatórios não Esteroides/farmacologia , Asma/metabolismo , Dermatite Atópica/tratamento farmacológico , Esquema de Medicação , Humanos , Hidroxiureia/farmacocinética , Hidroxiureia/farmacologia , Hidroxiureia/uso terapêutico , Inibidores de Lipoxigenase/farmacocinética , Inibidores de Lipoxigenase/farmacologia , Doença Pulmonar Obstrutiva Crônica/tratamento farmacológico , Rinite/tratamento farmacológicoRESUMO
OBJECTIVES: To measure the effect of tricyclic antidepressant drugs (TCAs) on human myocardial contractility. METHODS: Human atrial tissue was obtained during cardiac bypass surgery. The tissue was harvested, suspended in a Tyrode buffer at 37 degrees C, and perfused with a 95%/5% oxygen-carbon dioxide mixture. Developed force was continuously measured using a force transducer and recorded by computer. After an equilibration period, escalating doses of amitriptyline or desipramine were added to the bath. All strips were exposed to the following five concentrations of each drug: 0 (control) 0.4, 4, 40, and 400 microM. The results for each experiment were expressed as the difference between the developed force measured prior to the addition of each concentration of drug and the developed force measured after a 30-minute exposure to the drug. RESULTS: Desipramine decreased the developed force by 27%, 49%, and 74% at concentrations of 0.4, 40, and 400 microM, respectively. Amitriptyline decreased the developed force by 38% at the 40-microM concentration and by 89% at the 400-microM concentration. Untreated strips retained 94% of baseline developed force at 150 minutes. CONCLUSIONS: Tricyclic antidepressants depress human myocardial function in a dose-dependent fashion independent of the effects on the cardiac conduction system. While previous work has demonstrated the effect of therapies for the reversal of impaired cardiac conduction following TCA poisoning, to the best of the authors' knowledge, no reports have documented the effects of therapy on direct myocardial depression. Additional therapies targeted at reversing the direct cardiodepressive effects of TCA may improve outcome following TCA poisoning.
Assuntos
Amitriptilina/farmacologia , Antidepressivos Tricíclicos/farmacologia , Desipramina/farmacologia , Átrios do Coração/efeitos dos fármacos , Contração Miocárdica/efeitos dos fármacos , Análise de Variância , Técnicas de Cultura , Relação Dose-Resposta a Droga , Humanos , Condução Nervosa/efeitos dos fármacos , Condução Nervosa/fisiologia , Probabilidade , Valores de Referência , Sensibilidade e EspecificidadeRESUMO
The C chemokine lymphotactin has been characterized as a T cell chemoattractant both in vitro and in vivo. To determine whether lymphotactin expression within tumors could influence tumor growth, we transfected an expression vector for lymphotactin into SP2/0 myeloma cells and tested their ability to form tumors in BALB/c and nude mice. Transfection did not alter cell growth in vitro. Whereas SP2/0 cells gave rise to a 100% tumor incidence, lymphotactin-expressing SP2/0-Lptn tumors invariably regressed in BALB/c mice and became infiltrated with CD4(+) and CD8(+) T cells and neutrophils. Regression of the SP2/0-Lptn tumors was associated with a type 1 cytokine response and dependent on both CD4(+) and CD8(+) T cells, but not NK cells. Both SP2/0 and SP2/0-Lptn tumors grew in nude mice, but growth of the latter tumors was retarded and associated with heavy neutrophil responses; this retardation of SP2/0-Lptn tumor growth was reversed by neutrophil depletion of the mice. Our data also indicate that mouse neutrophils express the lymphotactin receptor XCR1 and that lymphotactin specifically chemoattracts these cells in vitro. Thus, lymphotactin has natural adjuvant activities that may augment antitumor responses via effects on both T cells and neutrophils and thereby could be important in gene transfer immunotherapies for some cancers.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Linfocinas/biossíntese , Proteínas de Membrana , Mieloma Múltiplo/imunologia , Mieloma Múltiplo/prevenção & controle , Neutrófilos/imunologia , Receptores de Superfície Celular/biossíntese , Receptores Acoplados a Proteínas G , Sialoglicoproteínas/biossíntese , Células Tumorais Cultivadas/metabolismo , Animais , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Vacinas Anticâncer/administração & dosagem , Vacinas Anticâncer/uso terapêutico , Quimiocinas C/genética , Quimiocinas C/fisiologia , Quimiotaxia de Leucócito/imunologia , Feminino , Rejeição de Enxerto/genética , Rejeição de Enxerto/imunologia , Injeções Subcutâneas , Linfocinas/genética , Linfocinas/fisiologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Mieloma Múltiplo/genética , Mieloma Múltiplo/patologia , Transplante de Neoplasias , Neutrófilos/metabolismo , Engenharia de Proteínas , Sialoglicoproteínas/genética , Sialoglicoproteínas/fisiologia , Células Tumorais Cultivadas/imunologia , Células Tumorais Cultivadas/transplanteRESUMO
The C chemokine lymphotactin (Lptn) has been reported to act specifically on CD4(+) and CD8(+) T lymphocytes and natural killer (NK) cells, but not monocytes. However, the chemotactic effect of Lptn on other types of hematopoietic cells has not been well studied. In this study we investigated (i) the chemotactic influences of Lptn on T and B lymphocytes, neutrophils, monocytes, and dendritic cells, and (ii) the expression of the Lptn receptor (XCR1) by these cells, using RT-PCR. Our data showed that Lptn is chemotactic for B lymphocytes and neutrophils as well as T lymphocytes, but not for monocytes or dendritic cells, and that XCR1 expression is found only in association with T and B lymphocytes and neutrophils, but not monocytes or dendritic cells. Thus, this study is the first demonstration of a chemotactic effect of Lptn on neutrophils and confirms the association of this effect with expression of the XCR1 receptor on these cells. These data suggest that Lptn could potentially be an important protein in the regulation of T and B lymphocytes and neutrophil trafficking, and thereby also their roles in inflammatory and immunological responses.
Assuntos
Quimiocinas C , Quimiotaxia/efeitos dos fármacos , Linfocinas/farmacologia , Proteínas de Membrana , Neutrófilos/efeitos dos fármacos , Receptores de Superfície Celular/genética , Receptores Acoplados a Proteínas G , Sialoglicoproteínas/farmacologia , Animais , Linfócitos B/citologia , Linfócitos B/efeitos dos fármacos , Linfócitos B/metabolismo , Fatores Quimiotáticos/farmacologia , Relação Dose-Resposta a Droga , Expressão Gênica , Camundongos , Camundongos Endogâmicos BALB C , Neutrófilos/citologia , Neutrófilos/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Linfócitos T/citologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismoRESUMO
Increased rates of RNA polymerase (pol) III transcription constitute a central feature of the mitogenic response, but little is known about the mechanism(s) responsible. We demonstrate that the retinoblastoma protein RB plays a major role in suppressing pol III transcription in growth-arrested fibroblasts. RB knockout cells are compromised in their ability to down-regulate pol III following serum withdrawal. RB binds and represses the pol III-specific transcription factor TFIIIB during G(0) and early G(1), but this interaction decreases as cells approach S phase. Full induction of pol III coincides with mid- to late G(1) phase, when RB becomes phosphorylated by cyclin D- and E-dependent kinases. TFIIIB only associates with the underphosphorylated form of RB, and overexpression of cyclins D and E stimulates pol III transcription in vivo. The RB-related protein p130 also contributes to the repression of TFIIIB in growth-arrested fibroblasts. These observations provide insight into the mechanisms responsible for controlling pol III transcription during the switch between growth and quiescence.
Assuntos
Ciclo Celular/fisiologia , Regulação da Expressão Gênica , RNA Polimerase III/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Gênica , Células 3T3 , Animais , Meios de Cultura Livres de Soro , DNA/biossíntese , Embrião de Mamíferos , Fibroblastos/citologia , Fibroblastos/enzimologia , Fibroblastos/fisiologia , Fase G1 , Fase G2 , Camundongos , Mitose , Fosforilação , Reação em Cadeia da Polimerase , RNA Polimerase III/genética , Fase de Repouso do Ciclo Celular , Fator de Transcrição TFIIIBRESUMO
In 1862, Samuel Gross described shock as the "rude unhinging" of the machinery of life. As noted above, adequate oxygen delivery and metabolism are essential to the maintenance of cellular energy stores. Failure of adequate tissue oxygen delivery and utilization during shock can lead to organ dysfunction and death. Hemorrhagic shock after trauma can result in inherent mitochondrial dysfunction as manifested by decoupling. This pathologic condition has been recently termed cytopathic hypoxia. Since mitochondria are the ultimate consumer of oxygen in cells, mitochondria might indeed be the machinery of life rudely unhinged by shock. Yet, therapeutic strategies have been recently developed to support mitochondrial function in shock and related states. If these therapeutic interventions directed towards organelle and cellular resuscitation are proven to enhance human organ function and improve survival, then these strategies could augment current therapeutic regimens directed exclusively towards hemodynamic and ventilatory homeostasis.
Assuntos
Doenças Mitocondriais/metabolismo , Consumo de Oxigênio/fisiologia , Choque Hemorrágico/metabolismo , Metabolismo Energético , Humanos , Isquemia/metabolismo , Mitocôndrias/enzimologia , Mitocôndrias/metabolismo , Doenças Mitocondriais/etiologia , Insuficiência de Múltiplos Órgãos/etiologia , NAD/metabolismo , Traumatismo por Reperfusão/metabolismo , Choque Hemorrágico/complicações , Choque Hemorrágico/terapia , Síndrome de Resposta Inflamatória Sistêmica/etiologia , Síndrome de Resposta Inflamatória Sistêmica/metabolismo , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Proinflammatory mediators such as tumor necrosis factor-alpha (TNF) have been implicated in the pathophysiology in a number of acute disease states. Tumor necrosis factor-alpha can contribute to cell death, apoptosis, and organ dysfunction. Tumor necrosis factor-alpha can be generated with sepsis or ischemia-reperfusion by activation of cell mitogen-activated protein kinases and nuclear factor kappa B, leading to TNF production. A number of strategies to modulate TNF have been recently explored, including factors directed toward mitogen-activated protein kinases, TNF transcription, anti-inflammatory ligands, heat shock proteins, and TNF-binding proteins. However, TNF may also play an important role in the adaptive response to injury and inflammation. Control of the deleterious effects of TNF and other proinflammatory cytokines represents a realistic goal for clinical emergency medicine. The purpose of this article is to provide a background of relevance to emergency medicine academicians on the production and regulation of TNF, the acute effects of TNF on pathophysiology, and the rationale for therapeutic interventions directed toward TNF and the clinical experience with these strategies.
Assuntos
Medicina de Emergência , Fator de Necrose Tumoral alfa , Apoptose , Humanos , Inflamação/tratamento farmacológico , Ensaios Clínicos Controlados Aleatórios como Assunto , Fator de Necrose Tumoral alfa/efeitos adversos , Fator de Necrose Tumoral alfa/biossíntese , Fator de Necrose Tumoral alfa/fisiologia , Fator de Necrose Tumoral alfa/uso terapêuticoRESUMO
RNA polymerase III (Pol III) transcription is subject to repression by the retinoblastoma protein RB, both in vitro and in vivo (R. J. White, D. Trouche, K. Martin, S. P. Jackson, and T. Kouzarides, Nature 382:88-90, 1996). This is achieved through a direct interaction between RB and TFIIIB, a multisubunit factor that is required for the expression of all Pol III templates (C. G. C. Larminie, C. A. Cairns, R. Mital, K. Martin, T. Kouzarides, S. P. Jackson, and R. J. White, EMBO J. 16:2061-2071, 1997; W.-M. Chu, Z. Wang, R. G. Roeder, and C. W. Schmid, J. Biol. Chem. 272:14755-14761, 1997). p107 and p130 are two closely related proteins that display 30 to 35% identity with the RB polypeptide and share some of its functions. We show that p107 and p130 can both repress Pol III transcription in transient transfection assays or when added to cell extracts. Pull-down assays and immunoprecipitations using recombinant components demonstrate that a subunit of TFIIIB interacts physically with p107 and p130. In addition, endogenous TFIIIB is shown by cofractionation and coimmunoprecipitation to associate stably with both p107 and p130. Disruption of this interaction in vivo by using the E7 oncoprotein of human papillomavirus results in a marked increase in Pol III transcription. Pol III activity is also deregulated in fibroblasts derived from p107 p130 double knockout mice. We conclude that TFIIIB is targeted for repression not only by RB but also by its relatives p107 and p130.
Assuntos
Proteínas Nucleares/genética , Fosfoproteínas/genética , Proteínas , RNA Polimerase III/genética , Fatores de Transcrição/genética , Células 3T3 , Animais , Northern Blotting , Western Blotting , Cloranfenicol O-Acetiltransferase/metabolismo , Fibroblastos/metabolismo , Humanos , Camundongos , Camundongos Knockout , Osteossarcoma/metabolismo , Papillomaviridae/metabolismo , Plasmídeos , Testes de Precipitina , Proteínas Recombinantes de Fusão , Proteína p107 Retinoblastoma-Like , Proteína p130 Retinoblastoma-Like , Fator de Transcrição TFIIIB , Transcrição Gênica , Células Tumorais CultivadasRESUMO
Hemoglobin is perhaps the most intensively studied of the biologically important molecules. Much is known of its structure, its function, and its regulation. In addition to well-characterized processes of oxygen, carbon dioxide, and carbon monoxide transport, new data suggest a key role of hemoglobin as a carrier of nitric oxide. In this review, we describe the basis of this interaction, as well as its clinical relevance to such problems as acute respiratory distress syndrome, percutaneous transluminal coronary angioplasty, and transplant allograft survival.
Assuntos
Proteínas de Transporte/fisiologia , Hemoglobinas/fisiologia , Óxido Nítrico/fisiologia , HumanosRESUMO
Ischemic preconditioning has been shown to ameliorate injury due to subsequent ischemia in several organs. However, relatively little is known about preconditioning and the kidney. To address this, rats were randomized to control (C, N = 14), 2 min of ischemic preconditioning (P2 N = 10), 3 periods of 2 min of ischemia separated by 5 min periods of reflow (P2,3 N = 7), or three 5 min periods of ischemia separated by 5 min of reflow (P5,3 N = 6) prior to 45 min of bilateral renal ischemia followed by 24 hours of reperfusion. We observed a lower serum creatinine after 24 hours of reflow in P2, P2, 3 but not P5, 3 rats compared with C. Histology was examined in the C and P2, 3 groups and demonstrated less severe injury in the P2, 3 group. To gain insight into the mechanism by which preconditioning ameliorated ischemic injury, we performed near IR spectroscopy and 31P NMR spectroscopy. Based on near IR spectroscopy, the P2, 3 group had closer coupling of cytochrome aa3 redox state with that of hemoglobin during reflow. In the 31P NMR studies, the changes in ATP and pHi were similar during ischemia, but the P2, 3 group recovered ATP and pHi faster than C. These data suggest that ischemic preconditioning may ameliorate ischemic renal injury as assessed by functional, metabolic and morphological methods. The mechanism(s) by which this occurs requires additional study.
Assuntos
Injúria Renal Aguda/prevenção & controle , Trifosfato de Adenosina/metabolismo , Creatinina/sangue , Isquemia/metabolismo , Precondicionamento Isquêmico , Rim/irrigação sanguínea , Injúria Renal Aguda/etiologia , Injúria Renal Aguda/metabolismo , Animais , Temperatura Corporal , Modelos Animais de Doenças , Seguimentos , Hemoglobinas/metabolismo , Concentração de Íons de Hidrogênio , Isquemia/complicações , Isquemia/patologia , Rim/metabolismo , Rim/fisiopatologia , Espectroscopia de Ressonância Magnética , Masculino , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Espectroscopia de Luz Próxima ao InfravermelhoRESUMO
p53 is a major tumour suppressor that is inactivated in a large proportion of human cancers. We show that p53 serves as a general repressor of transcription by RNA polymerase (pol) III. It can inhibit the synthesis of a range of essential small cellular RNAs including tRNA, 5S rRNA and U6 snRNA, as well as viral products such as the adenovirus VAI RNA. Fibroblasts derived from p53 knock-out mice display a substantial increase in pol III transcriptional activity. Endogenous cellular p53 is shown to interact with the TATA-binding protein (TBP)-containing general factor TFIIIB, thereby compromising its function severely. However, assembly of TFIIIB into a pre-initiation complex confers substantial protection against the inhibitory effects of p53. Since TFIIIB is an essential determinant of the biosynthetic capacity of cells, its release from repression by p53 may contribute to a loss of growth control during the development of many tumours.
Assuntos
Genes p53/fisiologia , RNA Polimerase III/biossíntese , RNA Polimerase III/genética , Proteínas Repressoras/genética , Transcrição Gênica , Células 3T3 , Animais , Repressão Enzimática/efeitos dos fármacos , Repressão Enzimática/genética , Fibroblastos/enzimologia , Células HeLa , Humanos , Camundongos , Camundongos Knockout , Iniciação Traducional da Cadeia Peptídica/genética , RNA Polimerase III/efeitos dos fármacos , Proteínas Repressoras/fisiologia , Transcrição Gênica/efeitos dos fármacos , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/fisiologiaRESUMO
The rate of protein synthesis is a critical determinant of cellular growth. Abnormal activation of this process is a frequent feature of transformed and tumour cells. Several distinct components of the translation apparatus have been shown to be deregulated in response to malignant transformation. Indeed, overexpression of certain translation factors has been found to predispose cells to transformation or even initiate it. The latest twist to this story comes from the discovery that the retinoblastoma protein RB plays a major role in restricting the production of tRNA and rRNA. RB is an important tumour suppressor. Its ability to limit the synthesis of these principle determinants of biosynthetic capacity could provide a mechanism for restraining cell growth. The loss of this control may constitute a significant step towards tumour progression.
Assuntos
Biossíntese de Proteínas , RNA Polimerase III/metabolismo , RNA Polimerase I/metabolismo , Proteína do Retinoblastoma/metabolismo , Transcrição Gênica , Animais , Divisão Celular , Regulação da Expressão Gênica , Humanos , Neoplasias/genética , Neoplasias/metabolismo , Proteína do Retinoblastoma/genéticaRESUMO
BACKGROUND: Ischemia and reperfusion (IR) can lead to impaired myocardial mechanical function and inhibition of key metabolic enzyme systems after IR. In this study, we sought to identify the postischemic lesion in oxidative phosphorylation and hypothesized that selective substrate repletion would restore mitochondrial metabolic function during reperfusion. METHODS AND RESULTS: Isolated rat hearts were subjected to global ischemia (25 minutes; 37 degrees C) and reperfusion (40 minutes). Left ventricular developed pressure (LVDP) and the cytochrome a,a3 redox state (near infrared spectroscopy) were continuously monitored. Oxygen consumption was measured for the NADH (mitochondrial complex I) and FADH2 (complex II) pathways in both the resting and maximal ADP-stimulated states. Myocellular oxidative phosphorylation capacity was measured using an NADPH-linked assay specific for mitochondrial ATPase. The hearts were randomized to either succinate (200 micromol/L) or control for the first 5 minutes of reperfusion after ischemia. IR in the control group resulted in an impairment of NADH (complex I) oxidative phosphorylation capacity (1.4+/-0.4 versus control 3.9+/-0.6 nmol ATP/min/mg) and depressed LVDP (49+/-3% of baseline; P<.05). The oxidative phosphorylation capacity for the succinate-using FADH2 pathway remained intact (2.6+/-0.3 versus 2.4+/-0.4). Postischemic succinate administration enhanced LVDP recovery after IR (89+/-8% of baseline; P<.05). Diminished electron transport resulted in depletion of electrons from cytochrome a,a3 during ischemia and early reperfusion, which was reversed by providing succinate as substrate. CONCLUSIONS: Cardiac ischemia and reperfusion results in a defect at mitochondrial complex I but not complex II. Cytochrome a,a3 undergoes anomalous oxidation during ischemia. Postischemic administration of succinate infusion restores the cytochrome a,a3 redox state balance and myocardial function after IR.
Assuntos
Isquemia Miocárdica/metabolismo , Traumatismo por Reperfusão Miocárdica/metabolismo , Miocárdio/metabolismo , Ácido Succínico/farmacologia , Trifosfato de Adenosina/análise , Animais , Oxirredução , Fosforilação Oxidativa , Consumo de Oxigênio , Ratos , Ratos Sprague-DawleyRESUMO
One hypothesized mechanism of the cardioprotection provided by preconditioning is decreased utilization of ATP during ischemia. Although ATP levels in preconditioned heart during ischemia have been previously studied, contractile activity during ischemia has not been investigated. Contractile activity accounts for significant ATP consumption during ischemia. We hypothesized that preconditioning stimuli may conserve energy during the ischemic period by decreasing myocardial contractile energy expenditure prior to asystolic cardiac arrest. We studied three preconditioning stimuli: (i) four cycles of 5-min periods of ischemia (4 x 5' CI), (ii) 2 min of alpha 1-adrenergic stimulation (phenylephrine; PE), and (iii) 2 min of P1-purinergic stimulation (adenosine). The effects of these stimuli on myocardial ATP, ventricular contractility, and the time to cessation of electromechanical function (asystole) during the sustained ischemic period were then examined. Preconditioning stimuli (4 x 5' CI, phenylephrine, and adenosine) improved postischemic functional recovery compared with nonpreconditioned controls. Myocardial ATP contents at the end of 20 min of global ischemia were higher for adenosine-treated (9.0 +/- 1.5 mumol/g dry weight; p < 0.05) and PE-treated (9.9 +/- 1.9 mumol/g dryweight; p < 0.05) hearts than for controls (6.6 +/- 1.2 mumol/g dry weight). The CI hearts began with lower myocardial ATP levels (9.9 +/- 1.2 mumol/g dry weight; p < 0.05) than other groups prior to the sustained ischemic period (control 13.4 +/- 1.0 mumol/g dry weight). As a result of a lower rate of ATP depletion, ATP levels in the CI group were similar to the untreated control after 20 min of sustained ischemia (5.5 +/- 0.7 mumol/g dry weight). Preconditioning with 4 x 5' CI or adenosine (but not PE) led to earlier ventricular arrest. Only adenosine-treated hearts demonstrated a more rapid decline in ventricular contractility during sustained ischemia than did nonpreconditioned control hearts. We conclude that while the final recovery of ventricular contractility after asystolic arrest and reperfusion is improved by preconditioning with different stimuli (4 x 5' CI, adenosine, or PE), each stimulus conferred a characteristic electromechanical and energy conservation strategy during sustained ischemia. Adenosine conserved myocardial ATP content and reduced total cardiac work (developed pressure and heart beats). CI conserved myocardial ATP and minimized the number of ischemic cardiac beats. PE preserved myocardial ATP during ischemia without changing contractile behavior. Thus, energy conservation strategies during ischemia could contribute to the protection afforded by preconditioning stimuli, but the mechanisms appear to differ among stimuli.
Assuntos
Coração/fisiopatologia , Precondicionamento Isquêmico Miocárdico , Isquemia Miocárdica/fisiopatologia , Adenosina/farmacologia , Trifosfato de Adenosina/metabolismo , Animais , Pressão Sanguínea/efeitos dos fármacos , Cardiotônicos/farmacologia , Coração/efeitos dos fármacos , Frequência Cardíaca/efeitos dos fármacos , Técnicas In Vitro , Masculino , Contração Miocárdica/efeitos dos fármacos , Fenilefrina/farmacologia , Ratos , Ratos Sprague-DawleyRESUMO
The tumour suppressor protein RB restricts cellular growth. This may involve inhibiting the synthesis of tRNA and 5S rRNA by RNA polymerase (pol) III. We have shown previously that RB can repress pol III transcription when overexpressed either in vitro or in vivo. We also demonstrated that pol III activity is elevated substantially in primary fibroblasts from RB-deficient mice. Here we address the molecular mechanism of this regulation. RB is shown to repress all types of pol III promoter. It can do this even if added after transcription complex assembly. Functional assays demonstrate that RB targets specifically the general pol III factor TFIIIB. A physical interaction between TFIIIB and RB is indicated by fractionation, pull-down and immunoprecipitation data. We show that TFIIIB activity is elevated in primary fibroblasts from RB-deficient mice. TFIIIB is a multisubunit complex that includes the TATA-binding protein (TBP) and a TFIIB-related factor called BRF. We show that RB itself contains regions of homology to both TBP and BRF and propose a model in which RB disrupts TFIIIB by mimicking these two components.
Assuntos
RNA Polimerase III/metabolismo , Proteína do Retinoblastoma/metabolismo , Fatores de Transcrição TFIII , Transcrição Gênica/fisiologia , Sequência de Aminoácidos , Animais , Células Cultivadas , Proteínas de Ligação a DNA/metabolismo , Fibroblastos , Células HeLa , Humanos , Camundongos , Camundongos Knockout , Dados de Sequência Molecular , Testes de Precipitina , Regiões Promotoras Genéticas/genética , Ligação Proteica , Proteínas Recombinantes/metabolismo , Proteína do Retinoblastoma/fisiologia , Proteínas de Saccharomyces cerevisiae , Homologia de Sequência de Aminoácidos , Fatores Associados à Proteína de Ligação a TATA , Proteína de Ligação a TATA-Box , Fator de Transcrição TFIIIB , Fatores de Transcrição/metabolismoRESUMO
In a double-blind study forty-two children scheduled for elective adenotonsillectomy were randomized to receive peritonsillar infiltration, following induction of anaesthesia, with either 0.25% plain bupivacaine or 0.9% saline, 0.5 ml/kg to a maximum of 10 ml. The children were assessed on awakening, and then 10 minutes, 1 hour, 4 hours and 24 hours later. On each occasion the observer gave the child a pain score from 1 (no pain) to 5 (severe pain). The scores on awakening and after 10 minutes were significantly lower in the bupivacaine group (P < 0.05, Mann-Whitney U test). Thereafter there was no difference between the groups. The authors conclude that peritonsillar infiltration with bupivacaine is only moderately useful as analgesia for children having tonsillectomy.
Assuntos
Analgesia , Anestesia Local , Bupivacaína/administração & dosagem , Tonsila Palatina , Tonsilectomia , Período de Recuperação da Anestesia , Criança , Pré-Escolar , Método Duplo-Cego , Feminino , Humanos , Masculino , Medição da Dor , Dor Pós-Operatória/prevenção & controle , PlacebosRESUMO
The purpose of this study was to determine the regional and myofibrillar ATPase (M-ATPase) fibre type glycogen utilization patterns in response to increased ventilation induced by pre-exhaustive (Pre-Exh) and exhaustive (Exh) durations of swimming. Twenty-eight hamsters were studied: six controls (Con), 11 Pre-Exh (swam 82 min), 11 Exh (swam to exhaustion). We examined the optical density of PAS-stained fibres from the different regions of the diaphragm as a measure of glycogen remaining after the exercise or control period. The optical densities of PAS-stained fibres in most M-ATPase fibre types and diaphragmatic regions for the Pre-Exh and Exh groups was less than those in the Con hamsters except for the optical densities of all the M-ATPase fibre types in the sternal region. The optical densities of PAS-stained fibres in different regions and M-ATPase fibre types did not differ in the Exh and Pre-Exh groups. This data indicates that significant glycogen utilization occurred in all three M-ATPase fibre types in the costal, and both the thoracic and abdominal surface of the crural diaphragm in hamsters following pre-exhaustive and exhaustive durations of swimming. Glycogen utilization was greater in type 1 fibres of the thoracic surface of the crural region than in the type 1 fibres of the sternal region of the Pre-Exh group. Further, significant utilization of glycogen did not occur in any of the three M-ATPase fibre types of the sternal region of the diaphragm following prolonged durations of swimming. It would appear that glycogen is an important substrate in the hamster diaphragm during swimming.(ABSTRACT TRUNCATED AT 250 WORDS)