RESUMO
BACKGROUND: Data about the effects of smoking on thrombo-hemostatic factors (tissue factor [TF] and tissue factor pathway inhibitor [TFPI-1]) are limited and on fibrinolytic factors (tissue plasminogen activator [t-PA] and plasminogen activator inhibitor-1 [PAI-1]) are debatable. The present study investigated the smoking-related, endothelial cell (EC)-specific responses for these factors and their relation to nitric oxide (NO) production in vitro. METHODS AND RESULTS: Serum from 8 nonsmokers and 15 smokers were incubated with confluent (approximately 85%) human umbilical vein endothelial cells (HUVECs) in 24-well tissue-culture plates for 12 hours. After the incubation, basal NO, t-PA, PAI-1, TF, TFPI-1 production, and substance P (SP)-stimulated NO, t-PA, and PAI-1 production were determined. HUVECs treated with smokers' serum showed lower basal (P<0.02) and SP-stimulated (P=0.059) t-PA production but similar basal and stimulated PAI-1 production (P=0.9 and P=0.6) compared with nonsmokers. Basal t-PA/PAI-1 molar ratio was significantly reduced in smokers (P<0.005). TFPI-1 level in the cell culture supernatant was also significantly lower in smokers compared with the nonsmoker group (P<0.05) with no difference in TF level between both groups (P=0.5). As previously reported, both basal (P<0.001) and SP-stimulated (P<0.05) NO production were significantly reduced in smokers. Basal TFPI-1 in culture correlated positively with basal NO production (r=0.42, P=0.04) and negatively with serum cotinine level (r=-0.6, P=0.01). CONCLUSIONS: These results indicate that cigarette smoking is associated with alterations in EC-derived fibrinolytic (t-PA) and antithrombotic (TFPI-1) factors. To our knowledge, this is the first demonstration that EC-derived TFPI is affected by smoking and endogenous NO or that the degree of smoke exposure may influence TFPI levels in an EC milieu.
Assuntos
Endotélio Vascular/metabolismo , Fibrinolíticos/metabolismo , Fumar , Adulto , Células Cultivadas , Cotinina/sangue , Morte Súbita Cardíaca/etiologia , Humanos , Lipoproteínas/biossíntese , Masculino , Infarto do Miocárdio/etiologia , Óxido Nítrico/biossíntese , Inibidor 1 de Ativador de Plasminogênio/biossíntese , Fumar/efeitos adversos , Tromboplastina/biossíntese , Ativador de Plasminogênio Tecidual/biossínteseRESUMO
OBJECTIVES: The goal of this study was to investigate the dose-dependent effects of active cigarette smoking on endothelial nitric oxide (NO) and endothelin-1 (ET-1) biosynthesis. BACKGROUND: Limited studies have suggested that active cigarette smoking may be associated with a dose-dependent reduction of endothelium-dependent vasodilation (EDV). The underlying biochemical changes that cause this dose-specific effect, such as changes in the endothelial NO biosynthetic pathway and ET-1 production, have not been examined. METHODS: Flow- and nitroglycerin-mediated reactivity of the brachial artery were measured in eight nonsmokers, seven light smokers (< or =1 pack/week) and eight heavy smokers (> or =1 pack/day), and their sera were added to confluent ( approximately 85%) monolayers of human umbilical endothelial cells (HUVECs) for 12 h. Basal and substance P-stimulated NO and basal ET-1 production were measured. The HUVECs used for measuring basal NO production were lysed, and both endothelial NO synthase (eNOS) protein expression and eNOS activity were determined. RESULTS: Serum cotinine level and pack-years of smoking were significantly lower in light smokers compared with heavy smokers (p < 0.006 and p < 0.004, respectively). There were no significant differences between heavy smokers and light smokers in EDV (p = 0.52), basal- (p = 0.70) and stimulated-NO production (p = 0.95), eNOS protein (p = 0.40) and eNOS activity (p = 0.63). Compared with nonsmokers, all the parameters were significantly altered in both of the smokers' groups. No differences were found in nitroglycerin-mediated vasodilation and in vitro ET-1 production among the three groups. CONCLUSIONS: These results indicate light smoking may have similar detrimental effects on EDV and NO biosynthetic pathway as does heavy smoking. These data may have important implications concerning the amount of active cigarette exposure that imparts cardiovascular risk.
Assuntos
Endotelina-1/biossíntese , Óxido Nítrico/biossíntese , Fumar/metabolismo , Vasodilatação/efeitos dos fármacos , Artéria Braquial/efeitos dos fármacos , Artéria Braquial/metabolismo , Relação Dose-Resposta a Droga , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Humanos , MasculinoRESUMO
ABSTRACT-We examined the mechanisms and the adhesive molecules mediating platelet-neutrophil adhesion in patients with septic shock. Neutrophils, platelets, and platelet poor plasma (NPPP) were isolated from 12 normal volunteers. Platelets and neutrophils were stimulated with platelet poor plasma (SPPP) removed from 12 patients in septic shock. Cell adhesion was assessed by filtration through 5-microm pore filters and by flow cytometry. Blocking monoclonal antibodies were used against the platelet and neutrophil surface receptors glycoprotein complex IIb/IIla, P-selectin, ICAM-2, CD11a, CD11b, and CD18. The filtration pressure (Pi) of cells suspended in SPPP was significantly greater than that of cells suspended in NPPP (24 +/- 1.0 mmHg vs. 14 +/- 1.0 mmHg; P< 0.05). The difference between the Pi of cells suspended in SPPP or NPPP (deltaPi SPPP-NPPP) in the presence of monoclonal antibodies anti-CD41, anti-CD62P, abciximab, anti-CD11a, anti-CD11b, and anti-CD18 was significantly less than the APi SPPP-NPPP of cell suspensions without the addition of these monoclonal antibodies (P < 0.01). The greatest reduction in Pi occurred when platelet receptor P-selectin was blocked simultaneously with the CD11b receptor on the neutrophil as compared to all other single blocking monoclonal antibodies or combinations of monoclonal antibodies. The mean fluorescence of activated platelet CD63-PE binding to neutrophils suspended in SPPP was significantly greater than that of cells suspended in NPPP (780 +/- 130 Ifu vs. 295 +/- 35 Ifu; P < 0.05). The greatest attenuation in mean fluorescence occurred by blocking the P-selectin receptor on the platelet simultaneously with CD11b receptor on the neutrophil. We conclude that platelet-neutrophil aggregation is increased in septic shock. This aggregation is mediated by the interaction of multiple platelet and neutrophil surface receptors. The platelet receptor P-selectin and the neutrophil receptor CD11b/CD18 appear to play the most important role in these interactions.
Assuntos
Plaquetas/fisiologia , Neutrófilos/fisiologia , Choque Séptico/sangue , Adulto , Idoso , Anticorpos Monoclonais/farmacologia , Antígenos CD/sangue , Antígeno CD11b/sangue , Antígenos CD18/sangue , Estudos de Casos e Controles , Adesão Celular/fisiologia , Comunicação Celular/fisiologia , Feminino , Filtração , Citometria de Fluxo , Humanos , Técnicas In Vitro , Masculino , Pessoa de Meia-Idade , Selectina-P/sangue , Glicoproteínas da Membrana de Plaquetas , Tetraspanina 30RESUMO
UNLABELLED: Drug-induced liver injury (DILI) is the most common cause of acute liver failure in the United-States. The aim of the study was to describe serum immune profiles associated with acute DILI, to investigate whether there are profiles associated with clinical features or types of DILI and/or with prognosis, and to assess temporal changes in levels. Twenty-seven immune analytes were measured in the sera of 78 DILI subjects in the Drug-Induced Liver Injury Network (DILIN) and compared with 40 healthy controls. Immune analytes (14 cytokines, 7 chemokines and 6 growth factors) were measured by BioPlex multiplex ELISA at DILI onset and after 6 months. A modeling process utilizing immune principles was used to select a final set of variables among 27 immune analytes and several additional clinical lab values for prediction of early death (within 6 months of DILI onset). Nineteen of the 27 immune analytes were differentially expressed among healthy control, DILI onset and 6-month cohorts. Disparate patterns of immune responses, especially innate and adaptive cellular (mostly TH17) immunity were evident. Low values of four immune analytes (IL-9, IL-17, PDGF-bb and RANTES) and serum albumin are predictive of early death [PPV = 88% (95% CI, 65%-100%), NPV = 97% (95% CI, 93%-100%), accuracy = 96% (95% CI, 92%-100%)]. CONCLUSIONS: Acute DILI is associated with robust and varying immune responses. High levels of expression of cytokines associated with innate immunity are associated with a poor prognosis, whereas high levels of expression of adaptive cytokines are associated with good long-term prognosis and eventual recovery. Serum immune analyte profiles at DILI onset appear to be of prognostic, and perhaps, diagnostic significance.
Assuntos
Doença Hepática Induzida por Substâncias e Drogas/sangue , Citocinas/sangue , Doença Aguda , Doença Hepática Induzida por Substâncias e Drogas/diagnóstico , Doença Hepática Induzida por Substâncias e Drogas/imunologia , Estudos de Coortes , Ensaio de Imunoadsorção Enzimática , Humanos , Imunidade Inata , Modelos Imunológicos , PrognósticoRESUMO
The effects of acute and prior exposure to lipopolysaccharide (LPS) and staphylococcal enterotoxin B (SEB) on superoxide release by monocytes were examined in control subjects and in patients with sepsis and septic shock during the acute stage and recovery. High doses of LPS, PMA (phorbol 12-myristate 13-acetate), and SEB stimulated monocyte superoxide release in control subjects (P < 0.05). Pretreatment of normal monocytes with these doses of LPS, PMA, and SEB induced significant hyporesponsiveness to subsequent challenge (P < 0.01), and evidence of cross-tolerance was observed. Monocytes isolated from patients with sepsis and septic shock demonstrated high spontaneous superoxide release compared with those of control subjects (P < 0.05). Stimulation of patient monocytes with LPS or SEB resulted in less superoxide production than that spontaneously released by controls (P < 0.01). In patients recovering from their initial infection, spontaneous superoxide release was less than that released during acute stage. In addition, the superoxide release in response to the same stimuli was significantly increased when compared with release during the acute stage (P < 0.05). These data demonstrate that both LPS and SEB induce hyporesponsiveness to LPS- or SEB-stimulated superoxide release. A similar pattern of hyporesponsiveness was observed during sepsis that may represent a mechanism for modulating the inflammatory response during severe infections.
Assuntos
Lipopolissacarídeos/imunologia , Monócitos/imunologia , Sepse/imunologia , Superantígenos/imunologia , Superóxidos/sangue , Adulto , Idoso , Células Cultivadas , Enterotoxinas/imunologia , Feminino , Humanos , Tolerância Imunológica/imunologia , Masculino , Pessoa de Meia-Idade , Choque Séptico/imunologia , Acetato de TetradecanoilforbolRESUMO
We allocated 35 male Sprague-Dawley rats into 7 groups and anesthetized each by using one of the following regimens: ketamine 50 mg+xylaxine 5 mg; ketamine 75 mg+xylazine 5 mg; pentobarbital 45 mg; and Telazol 30, 40, 50, and 60 mg/kg; supplemental doses were used as required. Respiratory rate, heart rate, mean arterial pressure, cardiac index, and stroke index were measured every 30 min for 4 h. The Telazol groups showed a dose-dependent increase in duration of anesthesia. Duration of anesthesia was significantly shorter for the ketamine and pentobarbital groups than for any of the Telazol doses. Heart rate showed a dose-dependent decrease among the Telazol groups, but overall heart rate in these groups was higher than in the ketamine and pentobarbital groups. Mean arterial pressure in the Telazol 40 and 50 groups was significantly higher than the pentobarbital and higher ketamine groups yet lower than that of the Telazol 60 group. Overall animals anesthetized with Telazol showed the highest cardiac index, ketamine intermediate, and pentobarbital the lowest; cardiac index was higher in the Telazol 50 group than in either the Telazol 30 or pentobarbital groups. The pentobarbital group exhibited the lowest stroke index, whereas ketamine-treated animals had an intermediate stroke index. These differing effects of anesthetics on cardiovascular parameters must be considered when choosing an anesthesia regimen or comparing data from different studies. In our model, the Telazol 40 and 50 groups appeared to exhibit the fewest adverse cardiovascular effects.