RESUMO
The aim of the current study was to define how cyclooxygenase (COX)-activity affects airway hyperresponsiveness (AHR) and inflammation using interventions with COX inhibitors at different time points during allergen challenge and/or prior to measurement of AHR in an eosinophil-driven allergic mouse model. Inflammatory cells were assessed in bronchioalveolar lavage (BAL) and AHR was evaluated as the total lung resistance to methacholine (MCh) challenge. Administration of FR122047 (COX-1 inhibitor) during ovalbumin (OVA) challenge and prior to MCh challenge enhanced AHR without affecting the inflammatory cell response. In contrast, administration of lumiracoxib (COX-2 inhibitor) during the same time period had no effect on AHR but reduced the inflammatory cells in BAL. Nonselective COX inhibition with diclofenac both enhanced the AHR and reduced the inflammatory cells. Administration of diclofenac only during OVA challenge reduced the cells in BAL without any changes in AHR, whereas administration of diclofenac only prior to MCh challenge enhanced AHR but did not affect the cells in BAL. The present study implicates distinct roles of prostanoids generated along the COX-1 and COX-2 pathways and, furthermore, that inflammatory cells in BAL do not change in parallel with AHR. These findings support the fact that AHR and the inflammatory response are distinct and, at least in part, uncoupled events.
Assuntos
Hiper-Reatividade Brônquica/imunologia , Inibidores de Ciclo-Oxigenase/farmacologia , Inflamação/imunologia , Alérgenos/química , Animais , Hiper-Reatividade Brônquica/patologia , Lavagem Broncoalveolar , Ciclo-Oxigenase 1/metabolismo , Ciclo-Oxigenase 2/metabolismo , Diclofenaco/farmacologia , Feminino , Imunoglobulina E/metabolismo , Imunoglobulina G/metabolismo , Inflamação/patologia , Cloreto de Metacolina/farmacologia , Camundongos , Camundongos Endogâmicos BALB CRESUMO
BACKGROUND AND PURPOSE: Non-steroidal anti-inflammatory drugs (NSAIDs) are analgesic and anti-inflammatory by virtue of inhibition of the cyclooxygenase (COX) reaction that initiates biosynthesis of prostaglandins. Findings in a pulmonary pharmacology project gave rise to the hypothesis that certain members of the NSAID class might also be antagonists of the thromboxane (TP) receptor. EXPERIMENTAL APPROACH: Functional responses due to activation of the TP receptor were studied in isolated airway and vascular smooth muscle preparations from guinea pigs and rats as well as in human platelets. Receptor binding and activation of the TP receptor was studied in HEK293 cells. KEY RESULTS: Diclofenac concentration-dependently and selectively inhibited the contraction responses to TP receptor agonists such as prostaglandin D2 and U-46619 in the tested smooth muscle preparations and the aggregation of human platelets. The competitive antagonism of the TP receptor was confirmed by binding studies and at the level of signal transduction. The selective COX-2 inhibitor lumiracoxib shared this activity profile, whereas a number of standard NSAIDs and other selective COX-2 inhibitors did not. CONCLUSIONS AND IMPLICATIONS: Diclofenac and lumiracoxib, in addition to being COX unselective and highly COX-2 selective inhibitors, respectively, displayed a previously unknown pharmacological activity, namely TP receptor antagonism. Development of COX-2 selective inhibitors with dual activity as potent TP antagonists may lead to coxibs with improved cardiovascular safety, as the TP receptor mediates cardiovascular effects of thromboxane A2 and isoprostanes.
Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Inibidores de Ciclo-Oxigenase 2/farmacologia , Diclofenaco/análogos & derivados , Diclofenaco/farmacologia , Receptores de Tromboxanos/antagonistas & inibidores , Adulto , Animais , Anti-Inflamatórios não Esteroides/administração & dosagem , Plaquetas/efeitos dos fármacos , Plaquetas/metabolismo , Linhagem Celular , Inibidores de Ciclo-Oxigenase 2/administração & dosagem , Inibidores de Ciclo-Oxigenase/administração & dosagem , Inibidores de Ciclo-Oxigenase/farmacologia , Diclofenaco/administração & dosagem , Relação Dose-Resposta a Droga , Feminino , Cobaias , Humanos , Masculino , Pessoa de Meia-Idade , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/metabolismo , Ligação Proteica , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacosRESUMO
Lungs from skin-sensitised and non-sensitised guinea pigs were exposed via the airways to 3-carene (1900 mg/m3) and perfused with buffer containing either autologous plasma or lymphocytes. The experiments were performed in order to investigate the importance of blood components for the increased lung responsiveness seen in skin-sensitised animals. A reduction in lung function was noted in all lungs during 3-carene exposure. There was no difference in the 3-carene response between lungs from skin-sensitised animals versus lungs from non-sensitised animals when the perfusion buffer contained lymphocytes. However, when plasma diluted with buffer was used as perfusion medium, there was a significant enhancement in the response in lungs from sensitised versus lungs from non-sensitised animals. This implies that skin sensitisation increases lung responses to inhaled 3-carene and those components in plasma, and not the lymphocyte fraction, contributes to the observed increased lung responsiveness.
Assuntos
Hipersensibilidade a Drogas/etiologia , Pulmão/efeitos dos fármacos , Monoterpenos/farmacologia , Animais , Monoterpenos Bicíclicos , Cosméticos/farmacologia , Cosméticos/toxicidade , Hipersensibilidade a Drogas/imunologia , Feminino , Cobaias , Exposição por Inalação , Pulmão/imunologia , Complacência Pulmonar/imunologia , Linfócitos/imunologia , Monoterpenos/imunologia , Monoterpenos/toxicidade , Plasma/imunologia , Pele/imunologiaRESUMO
In this study possible mechanisms underlying the vaso- and bronchoconstriction caused by the tricyclic antidepressant drug amitriptyline in isolated rat lungs were investigated. Some features here are similar to those apparent in adult respiratory distress syndrome and acute lung injury. Amitriptyline exposure (50 and 100 microM) caused a dose-related, pronounced, and rapid vaso- (50 microM, 30 min, p < 0.001 and 100 microM, 30 min, p < 0.001) and bronchoconstriction (50 microM, 30 min, p = 0.01 and 100 microM, 30 min, p < 0.001). The maximal noted decrease in perfusion flow was 28 +/- 2.9% at 25 min and 80 +/- 4.5% at 30 min for 50 and 100 microM amitriptyline, respectively. The maximal noted decrease in airway conductance was 29 +/- 4.7% at 25 min and 68 +/- 5.0% at 30 min. To investigate mechanisms thought to be involved in amitriptyline-induced lung function impairment, lungs were treated with several different substances including antiinflammatory agents, antioxidants, inhibitors of enzymes involved in the arachidonic acid cascade, physiological antagonists, and neurogenic antagonists. A significant reduction of amitriptyline-induced vasoconstriction was observed when lungs were treated with the protein kinase inhibitor staurosporine (3 microM, 30 min, p < 0.001), the NO-donor S-nitrosoglutathione (100 microM, 30 min, p < 0.001) and the combined endothelin A/endothelin B receptor antagonist PD 145065. This latter inhibitor caused a significant attenuation of late vasoconstriction (1 microM, 60 min, p = 0.03). The amitriptyline-induced bronchoconstriction was attenuated by the beta(2)-agonist salbutamol (1 microM, 30 min, p = 0.03) and the platelet-activating factor antagonist WEB2086 (10 microM, 30 min, p = 0.03). Staurosporine had an initial protective effect on bronchoconstriction (3 microM, 5 min, p = 0.003), while PD145065 significantly decreased bronchoconstriction 60 min after start of amitriptyline exposure (1 microM, 30 min, p = 0.003). This indicates that endothelin as well as platelet activating factor and protein kinase activation are important in mediating amitriptyline-induced lung function impairment in our experimental model and perhaps also in acute lung injury.
Assuntos
Amitriptilina/toxicidade , Pulmão/efeitos dos fármacos , Doença Aguda , Agonistas Adrenérgicos beta/farmacologia , Animais , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios não Esteroides/farmacologia , Antioxidantes/farmacologia , Broncoconstrição/efeitos dos fármacos , Broncodilatadores/farmacologia , Bloqueadores dos Canais de Cálcio/farmacologia , Relação Dose-Resposta a Droga , Antagonistas dos Receptores de Endotelina , Inibidores Enzimáticos/farmacologia , Técnicas In Vitro , Pulmão/irrigação sanguínea , Pulmão/patologia , Doadores de Óxido Nítrico/farmacologia , Oligopeptídeos/farmacologia , Inibidores da Agregação Plaquetária/farmacologia , Ratos , Ratos Sprague-Dawley , Síndrome do Desconforto Respiratório/induzido quimicamente , Síndrome do Desconforto Respiratório/patologia , Vasoconstrição/efeitos dos fármacosRESUMO
Inhalation of 3-carene has been shown to induce bronchoconstriction in concentrations not far from the threshold limit value. In this study, one group of guinea-pigs were sensitised by dermal exposure to 3-carene according to the modified Cumulative Contact Enhancement Test protocol and another group of animals was used as controls. Lungs from the skin-sensitised and control guinea-pigs were perfused with diluted autologous blood (13 ml blood/87 ml buffer) and exposed to 3-carene at an air concentration of 3000 mg/m(3). In both groups there was a reduction in compliance and conductance but this reduction was significantly (P<0.05) more pronounced (2.5-3 times) in lungs obtained from sensitised animals than from control animals. In a previous study with similar design, but with plain buffer instead of diluted autologous blood as perfusate, we found no statistically significant difference in lung bronchoconstriction. Thus, it is concluded that skin sensitisation can increase lung reactivity to 3-carene and that important mediators of this effect seem to be present in the blood.
Assuntos
Alérgenos/imunologia , Hiper-Reatividade Brônquica/imunologia , Hipersensibilidade a Drogas/imunologia , Pulmão/imunologia , Monoterpenos , Pele/imunologia , Terpenos/imunologia , Administração por Inalação , Administração Tópica , Animais , Monoterpenos Bicíclicos , Hiper-Reatividade Brônquica/sangue , Hiper-Reatividade Brônquica/induzido quimicamente , Testes de Provocação Brônquica , Hipersensibilidade a Drogas/sangue , Feminino , Cobaias , Imunização , Pulmão/efeitos dos fármacos , Complacência Pulmonar/efeitos dos fármacos , Complacência Pulmonar/imunologia , PerfusãoRESUMO
A number of drugs can induce lung toxicity. The lung manifestations can range from more acute responses such as an acute ARDS-like reaction, seen occasionally at overdosing of drugs to more insidious reactions, which can occur during conventional drug treatment. In many of these cases inflammation is an important component in the pathophysiology of drug induced lung toxicity. Very little is known about the mechanisms and initial events of drug-induced injury. In this review a couple of mechanistic aspects, related to drug induced lung injury, will be discussed such as reactive oxygen species (ROS)-generation, mediator release and disturbances in lung phospholipid turnover.
Assuntos
Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Lipidoses/induzido quimicamente , Pulmão/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Síndrome do Desconforto Respiratório/induzido quimicamente , Animais , Humanos , Inflamação/induzido quimicamente , Lipidoses/patologia , Síndrome do Desconforto Respiratório/metabolismoRESUMO
Tricyclic antidepressants can, when taken in overdose, cause serious pulmonary failure such as the adult respiratory distress syndrome (ARDS). In this study we have examined the effects of some tricyclic antidepressants (amitriptyline, imipramine, nortriptyline and desipramine) on the viability and morphology of human endothelial and smooth muscle cells derived from umbilical cord. Effects of amitriptyline on endothelial cell fluidity, as well as permeability changes to an endothelial-smooth muscle cell bi-layer, were also studied. The tricyclic antidepressants induced acute, sub-lethal toxicity in both cell types above 100 microM as assessed by the MTT reduction assay. Morphological changes were also observed at these concentrations. Such changes were, however, absent at 33 microM and below. Amitriptyline did, however, cause a concentration-dependent fall in the electrical resistance of an endothelial-smooth muscle cell bi-layer, with significant effects already evident at 33 microM. All of these observed effects were fairly rapid and appeared within 5-15 min of exposure. The rapidity of these permeabilisation effects suggests potential membrane perturbations, since tricyclic antidepressants are lipophilic molecules with affinity for cell membranes. However, fluorescence anisotropy measurements showed no significant difference in membrane fluidity between amitriptyline-treated and control endothelial cells. Collectively, these data point to specific mechanisms of action of amitriptyline, and probably also the other tricyclic antidepressants studied, on endothelial permeability, which is a hallmark of ARDS. The data suggest that increased endothelial permeability could be due to impaired tight junction function.
Assuntos
Amitriptilina/toxicidade , Antidepressivos Tricíclicos/toxicidade , Endotélio Vascular/efeitos dos fármacos , Músculo Liso Vascular/efeitos dos fármacos , Junções Íntimas/efeitos dos fármacos , Permeabilidade da Membrana Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Técnicas de Cocultura , Desipramina/toxicidade , Impedância Elétrica , Endotélio Vascular/patologia , Técnica Indireta de Fluorescência para Anticorpo , Formazans/metabolismo , Humanos , Imipramina/toxicidade , Músculo Liso Vascular/patologia , Nortriptilina/toxicidade , Espectrometria de Fluorescência , Sais de Tetrazólio/metabolismo , Junções Íntimas/fisiologia , Cordão Umbilical/citologiaRESUMO
We have previously shown that tricyclic antidepressants can induce vaso- and bronchoconstriction as well as oedema formation in isolated perfused lungs. This is an effect similar to that seen clinically in adult respiratory distress syndrome. In order to investigate whether endothelin can be a mediator of this reaction, isolated perfused rat lungs were exposed to 0.1 mM amitriptyline via the pulmonary circulation, perfusate was collected and endothelin-1 present in the perfusate and lavage fluids was determined by radioimmunoassay. A significant increase in perfusate concentration of endothelin-1 was noted, with the highest release seen within the first 10 min. of exposure. Histamine and thromboxane have also been proposed as mediators in induction of adult respiratory distress syndrome. However, no increased amounts of these mediators were detected in the perfusate. Experiments where lungs were exposed to exogenous endothelin-1(0.1-1 nmol), both via the perfusate and via intratracheal instillation were conducted. Similar effects as observed with amitriptyline (0.1 mM) on lung function and perfusion flow were detected. In conclusion, the detection of endothelin-1 release in our lung model proposes a role for endothelin-1 in amitriptyline-induced vaso- and bronchoconstriction and possibly in adult respiratory distress syndrome type reaction. Further studies with this model are interesting in order to elucidate mechanisms behind the complex issue of adult respiratory distress syndrome-induction.
Assuntos
Amitriptilina/farmacologia , Antidepressivos Tricíclicos/farmacologia , Endotelina-1/metabolismo , Pulmão/efeitos dos fármacos , Animais , Feminino , Liberação de Histamina/efeitos dos fármacos , Técnicas In Vitro , Pulmão/metabolismo , Perfusão , Ratos , Ratos Sprague-Dawley , Tromboxano B2/metabolismoRESUMO
Guinea pigs were sensitised by dermal exposure to 3-carene according to the modified cumulative contact enhancement test (CCET) protocol. Lungs from sensitised and non-sensitised animals were then perfused with buffer and exposed for a period of 10 min to two different air concentrations of 3-carene, 600 and 3000 mg/m3. 3-Carene caused a statistically significant bronchoconstriction even at the relatively low concentration of 600 mg/m3 and the constriction was dose dependent. 600 mg/m3 of 3-carene caused a reduction of 19% in conductance capacity and 16% in compliance capacity. 3000 mg/m3 of 3-carene decreased lung compliance and conductance by 43 and 31%, respectively. The lungs from sensitised animals tended to show a greater response than lungs obtained from control animals. The lower concentration of 3-carene is close to and may even be below, occupational limit values in Sweden, Germany and USA.
Assuntos
Alérgenos/administração & dosagem , Hipersensibilidade a Drogas/imunologia , Pulmão/imunologia , Monoterpenos , Pele/imunologia , Terpenos/administração & dosagem , Alérgenos/farmacologia , Animais , Monoterpenos Bicíclicos , Feminino , Cobaias , Técnicas In Vitro , Pulmão/efeitos dos fármacos , Masculino , Perfusão , Testes de Função Respiratória , Terpenos/farmacologiaRESUMO
Overdosing of several drugs, such as tricyclic antidepressants, salicylates, and opiates, is known to induce effects like those seen in patients with adult respiratory distress syndrome. By exposing isolated perfused and ventilated rat lungs via the perfusate to six different tricyclic antidepressants (amitriptyline, nortriptyline, imipramine, desipramine, mianserine, and maprotiline), we investigated possible effects on ventilation (conductance and dynamic compliance), lung perfusion flow, and edema formation. The effects of these substances were pronounced and appeared within 15 min after exposure. Amitriptyline was studied in greater detail and caused a dose-related (0.01-1.0 mM) reduction in ventilation and perfusion flow. At the highest drug concentration pronounced lung edema was observed. Morphological studies were conducted with a transmission electron microscope. The microscopic preparations showed dose-related edema (amitriptyline 0.1 and 1.0 mM). The effects noted in our experimental studies are similar to those described in patients who have taken an overdose of tricyclic antidepressants. This emphasizes the possibility of a noncardiogenic edema component in these patients.
Assuntos
Amitriptilina/toxicidade , Antidepressivos Tricíclicos/toxicidade , Pulmão/efeitos dos fármacos , Edema Pulmonar/induzido quimicamente , Ventilação Pulmonar/efeitos dos fármacos , Amitriptilina/administração & dosagem , Animais , Antidepressivos Tricíclicos/administração & dosagem , Desipramina/toxicidade , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Overdose de Drogas , Imipramina/toxicidade , Pulmão/fisiopatologia , Pulmão/ultraestrutura , Maprotilina/toxicidade , Mianserina/toxicidade , Microscopia Eletrônica , Nortriptilina/toxicidade , Tamanho do Órgão/efeitos dos fármacos , Perfusão , Edema Pulmonar/fisiopatologia , Ratos , Ratos Sprague-DawleyRESUMO
Isolated, perfused and ventilated guinea pig lungs were exposed to hexamethylene diisocyanate via the air passages. Two air concentrations of hexamethylene diisocyanate were studied (3.5 and 11 mg/m3). There was a statistically significant (P < 0.05-0.001) dose-related reduction in both conductance and compliance but no effects were noted on the pulmonary circulation. With 3.5 mg/m3 hexamethylene diisocyanate the conductance capacity was reduced with 38% and compliance with 30% after 60 min. exposure. Eleven mg/m3 hexamethylene diisocyanate reduced the conductance and compliance capacity with 86 and 69%, respectively, on an average. The reduction in lung function (with 11 mg/m3) was abolished when 100 microM diclofenac, a cyclooxygenase inhibitor, was added to the perfusate (P < 0.01). The thromboxane A2 antagonist L-670, 596 (20 microM) exerted a partial protective effect. The capacity of conductance and compliance decreased with 46 and 32%, respectively, on an average, after preperfusion with L-670, 596 and a following exposure of 11 mg/m3 hexamethylene diisocyanate for 60 min. Statistically significant protection (P < 0.05) was obtained on compliance and the P-value was < 0.1 for conductance. Thus, these data indicate that hexamethylene diisocyanate-induced bronchoconstriction is mediated via arachidonic acid release and thromboxane formation, in isolated, perfused and ventilated guinea pig lungs.
Assuntos
Cianatos/toxicidade , Complacência Pulmonar/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Circulação Pulmonar/efeitos dos fármacos , Animais , Ácido Araquidônico/metabolismo , Broncoconstrição/efeitos dos fármacos , Inibidores de Ciclo-Oxigenase/farmacologia , Diclofenaco/farmacologia , Relação Dose-Resposta a Droga , Cobaias , Técnicas In Vitro , Isocianatos , Pulmão/fisiologia , Masculino , Perfusão , Ventilação Pulmonar , Tromboxanos/metabolismoRESUMO
Inhalation of sodium metabisulfite (MBS; 80 mM; pH 2.9 +/- 0.1) or citric acid (CA; 0.4 M; pH 2.0 +/- 0.1) aerosols induced a reduction in compliance and conductance in the isolated perfused and ventilated guinea pig lung without affecting perfusion flow. The effect was dependent on the pH of the nebulized solution since inhalation of 80 mM MBS aerosols at pH 7.4 did not induce any effect on bronchial tone. Concomitantly to the bronchoconstriction induced by MBS or CA an increased level of calcitonin gene-related peptide (CGRP-LI) in the effluent perfusate was observed, indicating activation of sensory nerves. Sodium sulfite, a dissolution product of MBS, has previously been shown by our studies to reduce bronchoconstriction induced by inhalation of sulfur dioxide, in the isolated perfused and ventilated guinea pig lung. In the present study perfusion of the lung with sodium sulfite (3 mM) before and during exposure to aerosols with either MBS or CA attenuated the bronchoconstriction induced by the acidic solutions. The release of CGRP-LI induced by MBS or CA was not affected by sodium sulfite. Sulfite treatment did not modify perfused guinea pig lung reactivity towards acetylcholine (4 nmol), bradykinin (100 pmol), histamine (10 nmol), serotonin (500 pmol) and substance P fragment 5-11, a substance P analogue resistant to degrading enzyme (500 pmol). However, an inhibitory effect by sodium sulfite was observed on bronchoconstriction induced by the NK-2 agonist neurokinin A fragment 4-10 (NKA 4-10, 25 pmol). These results indicate that MBS- or CA-induced bronchoconstriction was dependent on the low pH of the aerosol solution and coincided with activation of sensory nerves. Sulfite modulation of the bronchoconstricting action of inhaled MBS and CA is suggested to be related to a sulfite-sensitive step in the signal transduction of the neuropeptide NKA.
Assuntos
Brônquios/inervação , Broncoconstrição/efeitos dos fármacos , Broncoconstritores/farmacologia , Ácido Cítrico/farmacologia , Neurônios Aferentes/fisiologia , Sulfitos/farmacologia , Acetilcolina/farmacologia , Administração por Inalação , Resistência das Vias Respiratórias/efeitos dos fármacos , Animais , Autacoides/farmacologia , Bradicinina/farmacologia , Peptídeo Relacionado com Gene de Calcitonina/metabolismo , Ácido Cítrico/administração & dosagem , Cobaias , Histamina/farmacologia , Concentração de Íons de Hidrogênio , Técnicas In Vitro , Pulmão/inervação , Pulmão/metabolismo , Complacência Pulmonar/efeitos dos fármacos , Masculino , Neurocinina A/farmacologia , Neurônios Aferentes/efeitos dos fármacos , Serotonina/farmacologia , Substância P/farmacologia , Sulfitos/administração & dosagemRESUMO
An isolated, perfused, guinea pig lung model was used to investigate the molecular events which occur when a 14C-labeled TDI vapor reaches the airways. Exposure concentrations of 0.2 and 0.7 ppm were tested. Perfusate composition included: Krebs Ringer buffer only, as well as buffer containing either guinea pig serum albumin, human serum albumin, or diluted guinea pig plasma. Radioactivity was detected in the perfusate within minutes of exposure, and following a delay, increased linearly. The rate of uptake was dependent on TDI concentration and the composition of the perfusate. Biochemical characterization of the state of the 14C-labeled material in the perfusate was performed. The distribution between low and high molecular weight reaction products was determined by molecular sieve fractionation and varied as a function of perfusate composition but no variability was observed as a function of time during the 45 min of exposure. An increase in nucleophile concentration in the perfusate was associated with both a higher percentage of conjugated products (from 15% with buffer only to 45% with diluted guinea pig plasma) and an increase in the rate of TDX uptake (from 0.5 microns Eq/min with buffer alone to 0.1 micrograms Eq/min with diluted GPSA as perfusate at 0.7 ppm). GC-MS analysis of the samples for free TDA, before and after acid hydrolysis, showed that the low molecular weight product(s), which represented from 55-85% of the circulating radioactivity, was composed of hydrolyzable and non-hydrolyzable conjugates and metabolites with approximately 4% of the label associated with free TDA. Although the distribution between high and low molecular weight species varies, this result is analogous to the findings from in vivo studies and suggests that the isolated, perfused lung (IVPL) system may be a useful tool in investigating the molecular mechanisms of isocyanate-induced disease and metabolic activity of the lung.
Assuntos
Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Tolueno 2,4-Di-Isocianato/farmacocinética , Tolueno 2,4-Di-Isocianato/toxicidade , Animais , Líquido da Lavagem Broncoalveolar/química , Radioisótopos de Carbono , Cobaias , Técnicas In Vitro , Cinética , Masculino , Perfusão/instrumentação , Tolueno 2,4-Di-Isocianato/administração & dosagemRESUMO
The intracellular events that lead to arachidonic acid release from bovine endothelial cells in culture treated with hydrogen peroxide were characterized. The hydrogen peroxide-stimulated release of arachidonic acid was time- and dose-dependent, with maximal release achieved at 15 minutes after the addition of 100 microM hydrogen peroxide. Hydrogen peroxide-stimulated release of arachidonic acid was blocked with the phospholipase A2 inhibitor quinacrine. Treatment of the cells with hydrogen peroxide did not result in liberation of oleic acid, indicating that hydrogen peroxide exercised its effect on an arachidonate-specific phospholipase. Pretreatment of the cells with antioxidants, transition metal chelators, and hydroxyl radical scavengers did not affect the hydrogen peroxide-stimulated arachidonic acid release, indicating that the response to hydrogen peroxide is not oxygen radical-mediated. The response to hydrogen peroxide does not appear to be calcium-dependent, due to the following two observations: (a) No increase in intracellular calcium was seen upon exposure of the FURA2-loaded cells to hydrogen peroxide at concentrations sufficient to release arachidonic acid, and (b) no change in the release response was detected in cells loaded with the intracellular calcium chelator BAPTA. Significant inhibition of arachidonic acid release was seen when the cells were pretreated with inhibitors of protein kinase C, but not with inhibitors of tyrosine kinase. The results of these studies indicate that hydrogen peroxide-stimulated arachidonic acid release is mediated by a specific signal-responsive phospholipase A2, and that this process is not mediated via the actions of either lipid peroxidation or calcium but, rather, that a stimulation of intracellular kinase activity is necessary for this response.
Assuntos
Peróxido de Hidrogênio/farmacologia , Fosfolipases A/metabolismo , Animais , Ácido Araquidônico/metabolismo , Cálcio/metabolismo , Bovinos , Células Cultivadas , Relação Dose-Resposta a Droga , Endotélio Vascular/enzimologia , Ativação Enzimática/efeitos dos fármacos , Fosfolipases A2 , Proteína Quinase C/antagonistas & inibidores , Proteína Quinase C/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de SinaisRESUMO
In this study the effects of S-nitrosothiols, in particular S-nitrosoglutathione (GSNO), were evaluated with regard to their bronchodilating properties, both after infusion via the pulmonary circulation and after inhalation, in the isolated perfused and ventilated guinea pig lung. Infused GSNO induced bronchorelaxation of lungs that were precontracted with methacholine. During a 15-min period of single-passage perfusion with GSNO (10 microM), maximally 10% was taken up and/or degraded by the lung. A spontaneous breakdown of GSNO in the perfusion buffer was also observed, which was partially accompanied by the formation of nitrite. Low levels of nitric oxide (NO) were detected in the perfusion buffer when GSNO was present. This was due to the presence of contaminating transition metals, because EDTA and 2,2'-dipyridyl largely reduced the formation of NO. The NO-scavenging agents oxyhemoglobin and 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide abolished levels of NO in the buffer but did not abolish GSNO-induced bronchodilation. The effects of infused GSNO are therefore attributed to an action of the intact S-nitrosothiol and not to NO released from GSNO in the perfusion buffer. Similarly, perfusion with S-nitrosated glutathione isopropyl ester, cysteinyl glycine, N-acetyl-L-cysteine or N-acetyl-D,L-penicillamine, but not with nitrosated bovine serum albumin or sodium nitrite, was found to induce bronchodilation. Inhalation of nebulized GSNO induced bronchodilation of methacholine-precontracted lungs with a rapid onset of action, although it was a less potent bronchodilator than salbutamol. The results show that infused or inhaled S-nitrosothiols have bronchodilating properties in the isolated perfused and ventilated guinea pig lung.
Assuntos
Broncodilatadores , Glutationa/análogos & derivados , Compostos Nitrosos/farmacologia , Compostos de Sulfidrila/farmacologia , Aerossóis , Resistência das Vias Respiratórias/efeitos dos fármacos , Animais , Relação Dose-Resposta a Droga , Glutationa/administração & dosagem , Glutationa/metabolismo , Glutationa/farmacologia , Cobaias , Pulmão/metabolismo , Complacência Pulmonar/efeitos dos fármacos , Masculino , Cloreto de Metacolina/antagonistas & inibidores , Óxido Nítrico/biossíntese , Compostos Nitrosos/administração & dosagem , Compostos Nitrosos/sangue , Compostos Nitrosos/metabolismo , S-Nitrosoglutationa , Compostos de Sulfidrila/sangue , Compostos de Sulfidrila/metabolismoRESUMO
Inhaled 3-carene at a concentration of 5,000 mg/m3 caused bronchoconstriction in isolated, ventilated and perfused guinea pig lungs. This effect was inhibited by the cyclooxygenase inhibitor diclofenac (100 microM) and the thromboxane/prostaglandin endoperoxide-receptor antagonist L-670,596 (1 microM). 3-Carene exposure also increased the amount of thromboxane in the perfusate from the lungs. In cultured calf pulmonary arterial endothelial cells 3-carene caused a dose-related release of arachidonic acid. Thus, the results obtained in this experimental model may have implications in the understanding of the pathophysiology of 3-carene-induced obstructive pulmonary disease in humans.
Assuntos
Broncoconstrição/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Monoterpenos , Terpenos/farmacologia , Animais , Ácido Araquidônico/metabolismo , Monoterpenos Bicíclicos , Carbazóis/farmacologia , Bovinos , Células Cultivadas , Diclofenaco/farmacologia , Relação Dose-Resposta a Droga , Endotélio/metabolismo , Cobaias , Técnicas In Vitro , Masculino , Antagonistas de Prostaglandina/farmacologia , Artéria Pulmonar/citologia , Artéria Pulmonar/metabolismo , Mecânica Respiratória/efeitos dos fármacos , Terpenos/química , Terpenos/metabolismo , Tromboxano B2/biossíntese , Fatores de TempoRESUMO
To investigate the mechanisms of nephrotoxicity of the antiviral drug Foscarnet (phosphonoformic acid), an in vitro model was developed based on primary cultures of dog renal proximal tubule epithelial cells. The cells were isolated by the collagenase perfusion technique from kidneys removed post mortem and grown in serum-free conditions. The cell viability was estimated, in confluent cells grown in 'Ca(2+)-supplemented medium' (average free Ca(2+) concentration, 1 mM), by measuring the release of lactate dehydrogenase or by measuring the rate of alpha-methyl glucose uptake. Foscarnet was toxic to the cells at concentrations above 1 mM. The toxicity was characterized by a long exposure time before any loss of viability was detected. The alpha-methyl glucose uptake was not affected until after 2 days of exposure. In actively dividing cells, Foscarnet displayed a concentration-dependent inhibition of thymidine incorporation after only 6 hr of exposure. The toxic effects of Foscarnet may be due to its ability to form complexes with divalent cations. We conclude that we have established an in vitro model system that uses proximal tubular epithelial cells from dogs and in which Foscarnet is toxic. Our model is therefore well suited for mechanistic studies of the nephrotoxicity of Foscarnet.
RESUMO
The effect of the thiol oxidizing agent diamide on airway conductance, dynamic compliance and perfusion flow of isolated perfused and ventilated guinea pig lungs was investigated. When infused in the pulmonary circulation, diamide (100 microM) induced bronchoconstriction, but no effect on perfusion flow was observed. Although diamide exposure induced the formation of thromboxane A2, the thromboxane/prostaglandin endoperoxide receptor antagonist L-670,596 did not affect the decrease in conductance and compliance induced by diamide. Diamide induced the release of the sensory neuropeptide calcitonin gene-related peptide. The bronchoconstriction and the release of calcitonin gene-related peptide induced by diamide were abolished by capsaicin pretreatment of the guinea pigs. Combined pretreatment with the NK1 and NK2 receptor antagonists, CP-96,345 and SR-48968, attenuated the effect of diamide. Hydrogen peroxide-induced vaso- and bronchoconstriction was not affected by capsaicin-pretreatment, nor did hydrogen peroxide induce detectable release of calcitonin gene-related peptide. The results indicate that diamide activates sensory nerves and induces neuropeptide release and neurokinin receptor-mediated bronchoconstriction in the isolated perfused and ventilated guinea pig lung.
Assuntos
Broncoconstrição/efeitos dos fármacos , Diamida/farmacologia , Peróxido de Hidrogênio/farmacologia , Pulmão/efeitos dos fármacos , Neuropeptídeos/fisiologia , Animais , Benzamidas/farmacologia , Compostos de Bifenilo/farmacologia , Peptídeo Relacionado com Gene de Calcitonina/imunologia , Peptídeo Relacionado com Gene de Calcitonina/metabolismo , Capsaicina/farmacologia , Carbazóis/farmacologia , Cobaias , Técnicas In Vitro , Masculino , Neurocinina A/antagonistas & inibidores , Neurônios Aferentes/efeitos dos fármacos , Neurônios Aferentes/fisiologia , Piperidinas/farmacologia , Tromboxanos/antagonistas & inibidores , Tromboxanos/biossínteseRESUMO
In this study the relationship between sulfur dioxide-induced sensory nerve activation and acute bronchoconstriction was assessed. We also studied the effects of sodium metabisulfite, an agent that is suggested to increase airway resistance via activation of sensory nerves. Sulfur dioxide (250 ppm) induced a characteristic biphasic bronchoconstriction. Concomitantly sulfur dioxide induced the release of calcitonin gene-related peptide (CGRP) from capsaicin-sensitive sensory nerves into the pulmonary circulation. In lungs of guinea pigs pretreated with a neurotoxic dose of capsaicin, the first phase of bronchoconstriction was reduced and the overflow of CGRP was not detectable. Tetrodotoxin abolished the initial phase of the bronchoconstriction induced by sulfur dioxide, indicating that a local neural reflex depending on sodium channels was operant. Inhibition of the vanilloid receptor with capsazepine slightly, although not significantly, reduced the contractile responses to sulfur dioxide. Sodium metabisulfite, when infused via the pulmonary circulation (3 mM), induced bronchoconstriction which was abolished by capsaicin pretreatment, but not significantly reduced by capsazepine. The results indicate that in the isolated guinea pig lung inhaled sulfur dioxide induces initial bronchoconstriction in part via sensory nerve activation, while other mechanisms are involved in the late effect. Sensory nerve activation appears to be the only mechanism for bronchoconstriction induced by infused sodium metabisulfite. A role for sensory nerve-mediated bronchoconstriction by sulfur dioxide or sodium metabisulfite via activation of the vanilloid receptor could not be conclusively demonstrated by this study using capsazepine.
Assuntos
Broncoconstrição/efeitos dos fármacos , Capsaicina/farmacologia , Neurônios Aferentes/efeitos dos fármacos , Sulfitos/farmacologia , Dióxido de Enxofre/farmacologia , Animais , Peptídeo Relacionado com Gene de Calcitonina/metabolismo , Cobaias , Técnicas In Vitro , Pulmão/efeitos dos fármacos , Masculino , Neurônios Aferentes/metabolismo , Perfusão , Respiração ArtificialRESUMO
The involvement of reactive oxygen species (ROS) in the pathogenesis of several lung diseases/injuries has been suggested. ROS are believed primarily to be generated by leukocytes (e.g. infiltrating neutrophils) although other ROS generating systems such as the xanthine/xanthine oxidase system may also be of importance. ROS may through oxidative changes exert a number of toxic effects which have been demonstrated in many different biological systems. At limited oxidative stress events such as modification of receptor activity and signalling, as well as release of endogenous mediators of inflammation may occur. One such ROS induced event, probably of importance for several lung diseases, is arachidonic acid (AA) release and metabolism to active product(s). In the lung, the release of AA results in both vaso- and bronchoconstriction, primarily caused by thromboxane A2. The molecular events leading to oxidant induced AA release and thromboxane formation are only partially elucidated.