RESUMO
Pulmonary thrombosis is a significant cause of patient mortality; however, there are no effective in vitro models of thrombi formation in human lung microvessels that could also assess therapeutics and toxicology of antithrombotic drugs. Here, we show that a microfluidic lung alveolus-on-a-chip lined by human primary alveolar epithelium interfaced with endothelium and cultured under flowing whole blood can be used to perform quantitative analysis of organ-level contributions to inflammation-induced thrombosis. This microfluidic chip recapitulates in vivo responses, including platelet-endothelial dynamics and revealed that lipopolysaccharide (LPS) endotoxin indirectly stimulates intravascular thrombosis by activating the alveolar epithelium, rather than acting directly on endothelium. This model is also used to analyze inhibition of endothelial activation and thrombosis due to a protease activated receptor-1 (PAR-1) antagonist, demonstrating its ability to dissect complex responses and identify antithrombotic therapeutics. Thus, this methodology offers a new approach to study human pathophysiology of pulmonary thrombosis and advance drug development.
Assuntos
Barreira Alveolocapilar/efeitos dos fármacos , Desenvolvimento de Medicamentos/métodos , Descoberta de Drogas/métodos , Fibrinolíticos/farmacologia , Dispositivos Lab-On-A-Chip , Técnicas Analíticas Microfluídicas/instrumentação , Microvasos/efeitos dos fármacos , Alvéolos Pulmonares/irrigação sanguínea , Trombose/tratamento farmacológico , Barreira Alveolocapilar/metabolismo , Barreira Alveolocapilar/patologia , Células Cultivadas , Técnicas de Cocultura , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Medicina Baseada em Evidências/métodos , Humanos , Microvasos/metabolismo , Microvasos/patologia , Segurança do Paciente , Medição de Risco , Transdução de Sinais/efeitos dos fármacos , Trombose/metabolismo , Trombose/patologia , Pesquisa Translacional Biomédica/métodosRESUMO
The development path described for JNJ-26489112 provides perspectives on interpretation of retinal effects observed in nonclinical studies and their implications for clinical development. JNJ-26489112 is a CNS-active investigational drug that has potential as a novel treatment for treatment-resistant and bipolar depression, epilepsy, and neuropathic/inflammatory pain. In a 6-month toxicity study in albino rats, retinal atrophy was observed at supratherapeutic exposures to JNJ-26489112. The histopathological changes and topography of the lesions were characteristic of light-induced damage specific to albino rats. The species/strain specificity is supported by an absence of any ocular effects in dogs and in pigmented and albino rats, housed under standard and reduced lighting, respectively. To further evaluate its potential to cause ocular effects, in vivo functional and structural ocular analyses were included in a 9-month monkey toxicity study. Reductions in rod- and cone-mediated electroretinograms were observed at supratherapeutic exposures but without any histopathologic changes. These data suggested that the effects of JNJ-26489112 in monkeys were neuromodulatory and not neurotoxic. Taken together, data related to the light-induced atrophy in albino rats and reversible neuromodulatory effects in monkeys, supported the safe evaluation of JNJ-26489112 in a clinical proof-of-concept study that included comprehensive functional and structural ocular monitoring.
Assuntos
Fármacos do Sistema Nervoso Central/toxicidade , Dioxanos/toxicidade , Retina/efeitos dos fármacos , Retina/patologia , Doenças Retinianas/induzido quimicamente , Sulfonamidas/toxicidade , Administração Oral , Animais , Fármacos do Sistema Nervoso Central/administração & dosagem , Fármacos do Sistema Nervoso Central/química , Dioxanos/administração & dosagem , Dioxanos/química , Cães , Eletrorretinografia , Feminino , Luz , Macaca fascicularis , Masculino , Conformação Molecular , Ratos , Ratos Sprague-Dawley , Doenças Retinianas/patologia , Sulfonamidas/administração & dosagem , Sulfonamidas/químicaRESUMO
In pre-clinical safety studies, drug-induced vascular injury is an issue of concern because there are no obvious diagnostic markers for pre-clinical or clinical monitoring and there is an intellectual gap in our understanding of the pathogenesis of this lesion. While vasodilatation and increased shear stress appear to play a role, the exact mechanism(s) of injury to the primary targets, smooth muscle and endothelial cells are unknown. However, evaluation of novel markers for potential clinical monitoring with a mechanistic underpinning would add value in risk assessment and management. This mini review focuses on the progress to identify diagnostic markers of drug-induced vascular injury. Von Willebrand factor (vWF), released upon perturbation of endothelial cells, is transiently increased in plasma prior to morphological evidence of damage in dogs or rats treated with vascular toxicants. Therefore, vWF might be a predictive biomarker of vascular injury. However, vWF is not an appropriate biomarker of lesion progression or severity since levels return to baseline values when there is morphological evidence of injury. A potential mechanistically linked biomarker of vascular injury is caveolin-1. Expression of this protein, localized primarily to smooth muscle and endothelial cells, decreases with the onset of vascular damage. Since vascular injury involves multiple mediators and cell types, evaluation of a panel rather than a single biomarker may be more useful in monitoring early and severe progressive vascular injury.
Assuntos
Biomarcadores/análise , Vasos Sanguíneos/efeitos dos fármacos , Endotélio Vascular/efeitos dos fármacos , Fator de von Willebrand/análise , Animais , Endotélio Vascular/citologia , Hemodinâmica , HumanosRESUMO
Recently, there has been an increased incidence of vascular toxicity in pre-clinical toxicology studies. This is of concern because of the uncertain relevance and extrapolation of this finding to humans. In dogs, profound heart rate (HR) and mean arterial pressure (MAP) changes were considered surrogate markers for drug-induced vascular injury until the early 1990s when endothelin receptor antagonists (ETRA) did not significantly alter HR or MAP but induced identical lesions in the coronary arteries of dogs. Thus significant alterations in HR and MAP were found not to be a prerequisite for this lesion. Clinically, the potential for vascular injury coupled with the lack of an unequivocal non-invasive diagnostic marker is an issue of concern to pharmaceutical companies and the regulatory authorities. Therefore, qualification and validation of biomarkers as diagnostic tools for drug-induced vascular injury would add great value to risk management and expedite the drug development process. This review focuses on the status, progress and future trends in vascular biology aimed at identification and development of diagnostic markers that are specific, sensitive and possess potential utility in both a pre-clinical and clinical setting.
Assuntos
Biomarcadores/sangue , Doenças Vasculares/induzido quimicamente , Doenças Vasculares/diagnóstico , Reação de Fase Aguda/sangue , Animais , Vasos Coronários/efeitos dos fármacos , Cães , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Humanos , Mediadores da Inflamação/análise , Peptídeos e Proteínas de Sinalização Intercelular/análise , Doenças Vasculares/fisiopatologiaRESUMO
In toxicology studies, drug-induced arterial injury in laboratory animals continues to be a pressing issue of concern, particularly to those engaged in the discovery and development of novel therapies intended for human use. The concern is justifiably magnified because, currently, there is no unequivocal biochemical marker of arterial injury and/or toxicity in animals or man. Therefore, in laboratory animals used for toxicology studies a precise description of arterial lesions in terms of location, distribution and morphologic character is necessary so that a correlation can be drawn between structural damage and derangement of specific cardiovascular functions. The critical nature of the latter cannot be over-emphasized because this will provide a basis for understanding the mechanism of toxicity, the pathogenesis of the lesion and assessment of human risk. However, in the decision making process, utilization of pattern recognition must be supported by rigorous scientific investigations aimed at establishing a link, where possible, between the deranged cardiovascular function and alterations in cellular, biochemical and molecular events. Conceivably, engagement of the molecular pharmacology target initiates a series of interactive cascades among cellular and non-cellular arterial components that culminate in organ damage. Therefore, any investigative mechanistic studies aimed at understanding the initiation and development of arterial lesions in laboratory animals must make a conscientious attempt to identify and characterize the molecular target of toxicity.
Assuntos
Animais de Laboratório , Artérias/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Xenobióticos/toxicidade , Animais , Artérias/patologia , Artérias/fisiopatologia , Biomarcadores , Modelos Animais de Doenças , Cães , Humanos , Medição de Risco , Especificidade da EspécieRESUMO
BACKGROUND: Idoxifene (ID) is a tissue-selective estrogen receptor modulator (SERM). The pharmacological profile of ID in animal studies suggests that it behaves like an estrogen receptor (ER) agonist in bone and lipid metabolism while having negligible ER activity on the reproductive system. It is unknown whether ID retains the vascular protective effects of estrogen. METHODS AND RESULTS: In cultured vascular smooth muscle cells (VSMCs), ID inhibited platelet-derived growth factor-induced DNA synthesis and mitogenesis with IC(50) values of 20.4 and 27.5 nmol/L, respectively. Treatment with ID resulted in S-phase cell cycle arrest in serum-stimulated VSMCs. ID 1 to 100 nmol/L significantly protected endothelial cells from tumor necrosis factor-alpha (TNF-alpha)-induced apoptosis in vitro. Virgin Sprague-Dawley rats ovariectomized 1 week before the study were treated with ID (1 mg x kg(-1) x d(-1)) or vehicle by gavage for 3 days before balloon denudation in carotid artery. The SMC proliferation in injured vessels was determined by immunostaining for proliferating cell nuclear antigen (PCNA). The number of PCNA-positive SMCs was reduced by 69%, 82%, and 86% in the media at days 1, 3 and 7, respectively, and by 78% in the neointima at day 7 after injury in ID- versus vehicle-treated group (P:<0.01). ID significantly enhanced reendothelialization in the injured carotid arteries as determined by Evans blue stain and immunohistochemical analysis for von Willebrand factor. In the former assay, the reendothelialized area in injured vessels was 43% in ID-treated group versus 24% in the vehicle group (P:<0.05); in the latter assay, the numbers of von Willebrand factor-positive cells per cross section increased from 24. 8 (vehicle) to 60.5 (ID) (P:<0.01) at day 14 after injury. In addition, the production of nitric oxide from excised carotid arteries was significantly higher in ID-treated than the vehicle group (8.5 versus 2.7 nmol/g, P:<0.01). Finally, ID treatment reduced neointimal area and the ratio of intima to media by 45% and 40%, respectively (P:<0.01), at day 14 after balloon angioplasty. CONCLUSIONS: The results indicate that ID beneficially modulates the balloon denudation-induced vascular injury response. Inhibition of VSMC proliferation and acceleration of endothelial recovery likely mediate this protective effect of ID.
Assuntos
Endotélio Vascular/lesões , Endotélio Vascular/patologia , Moduladores de Receptor Estrogênico/farmacologia , Músculo Liso Vascular/efeitos dos fármacos , Tamoxifeno/análogos & derivados , Tamoxifeno/farmacologia , Ferimentos não Penetrantes/patologia , Adulto , Animais , Artérias Carótidas/efeitos dos fármacos , Artérias Carótidas/metabolismo , Artérias Carótidas/cirurgia , Estenose das Carótidas/metabolismo , Estenose das Carótidas/patologia , Estenose das Carótidas/prevenção & controle , Contagem de Células , Ciclo Celular/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Células Cultivadas , Modelos Animais de Doenças , Endotélio Vascular/efeitos dos fármacos , Feminino , Humanos , Imuno-Histoquímica , Músculo Liso Vascular/citologia , Músculo Liso Vascular/metabolismo , Ovariectomia , Antígeno Nuclear de Célula em Proliferação/metabolismo , Ratos , Ratos Sprague-Dawley , Túnica Íntima/efeitos dos fármacos , Túnica Íntima/patologia , Fator de von Willebrand/metabolismoRESUMO
The cardiopulmonary profile of three rat strains (Sprague-Dawley, Wistar and High altitude-sensitive) was compared upon exposure to hypoxia (9% O2) for 0, 7 or 14 days. No differences were observed among the in vitro contractile (ET-1) and relaxant (carbachol) responses of pulmonary artery isolated from the three strains during normoxia. Chronic hypoxia decreased ET-1 contractile responses and diminished relaxant responses to carbachol similarly in all strains. In Sprague-Dawley, Wistar and High altitude-sensitive rats, pulmonary arterial pressure rose time-dependently and was elevated by 108%, 116% and 167%, respectively, after 14 days of hypoxia compared to normoxic controls. Right ventricular hypertrophy was increased by 51%, 93% and 55%, respectively, at 14 days. Hypoxia-induced hypertrophy and medial thickening in the pulmonary vasculature were more pronounced in High altitude-sensitive rats. Sprague-Dawley exhibited hypoxia-induced airway hyperresponsiveness to intravenous methacholine, but there were no hypoxia- or strain-related differences in in vitro tracheal contractility. Although each strain exhibited greater sensitivity for a particular hypoxia-induced parameter, pulmonary vascular functional and structural changes suggest that High altitude-sensitive rats represent a choice model of hypoxia-induced pulmonary hypertension.
Assuntos
Hipóxia/fisiopatologia , Artéria Pulmonar/fisiopatologia , Altitude , Animais , Carbacol/farmacologia , Modelos Animais de Doenças , Endotelina-1/farmacologia , Endotelinas/sangue , Hipertensão Pulmonar/etiologia , Hipertensão Pulmonar/patologia , Hipertensão Pulmonar/fisiopatologia , Hipertrofia , Hipertrofia Ventricular Direita/etiologia , Hipóxia/patologia , Técnicas In Vitro , Masculino , Cloreto de Metacolina/farmacologia , Artéria Pulmonar/patologia , Ratos , Ratos Sprague-Dawley , Ratos Wistar , Especificidade da Espécie , Traqueia/efeitos dos fármacos , Traqueia/fisiopatologia , Vasoconstrição/efeitos dos fármacos , Vasodilatação/efeitos dos fármacosRESUMO
Several vasoactive drugs that lower blood pressure and increase heart rate induce regional cardiotoxicity in the dog, most frequently of right coronary arteries and right atrium. The basis for this selective damage is thought to result from local changes in vascular tone and blood flow. Administration of an endothelin receptor antagonist (ETRA, SB 209670) to dogs induced damage most frequent and severe in the right coronary artery and right atrium. Because site predisposition may correlate with distribution of vasoactive receptors, the objectives of this study were to map endothelin (ET) receptor distribution and density within regions of dog heart using both gene (mRNA) and protein expression endpoints for dog ET(A) and ET(B) receptors, and, additionally, correlate ET receptor subtype density with regional cardiac blood flow. A 10- to 15-mmHg reduction in mean arterial pressure with a concomitant increase in heart rate (10-20%), a six- and twofold increase in regional blood flow to the right and left atrium, respectively, and acute hemorrhage, medial necrosis, and inflammation were observed in the right coronary arteries and arteries of the right atrium after ETRA infusion for 5 days. Radioligand protein binding to quantify both ET receptors in normal dog heart indicated a twofold greater density of ET receptors in atrial regions versus ventricular regions. Importantly, ET receptor density in coronary arteries was markedly (about five- to sixfold) increased above that in atrial or ventricular tissues. ET receptor subtype characterization indicated ET(B) receptors were three times more prevalent in right coronary arteries compared to left coronary arteries and in situ hybridization confirmed localization of ET(B) in vascular smooth muscle. ET(A) receptor density was comparable in right and left coronary arteries. Quantitative real-time polymerase chain reaction for ET(A) and ET(B) receptor mRNA transcripts supported the site prevalence for message distribution. Consequently, the composite of protein and message expression profiles for ET(A) and ET(B) receptors indicated a disproportionate distribution of ET(B) receptors within right coronary artery of dog and this, along with functional measures of blood flow after ETRA infusion indicated a predisposition for exaggerated pharmacological responses and subsequent damage to right coronary arteries by ET and/or ETRAs.
Assuntos
Doença das Coronárias/metabolismo , Receptores de Endotelina/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Doença das Coronárias/etiologia , Doença das Coronárias/fisiopatologia , Vasos Coronários/efeitos dos fármacos , Vasos Coronários/metabolismo , Vasos Coronários/patologia , Cães , Antagonistas dos Receptores de Endotelina , Feminino , Átrios do Coração/efeitos dos fármacos , Átrios do Coração/fisiopatologia , Ventrículos do Coração/efeitos dos fármacos , Ventrículos do Coração/fisiopatologia , Hemodinâmica/efeitos dos fármacos , Hibridização In Situ , Indanos/sangue , Indanos/farmacocinética , Indanos/farmacologia , Masculino , Dados de Sequência Molecular , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Endotelina/genética , Fluxo Sanguíneo Regional/efeitos dos fármacos , Homologia de Sequência de Aminoácidos , Homologia de Sequência do Ácido Nucleico , Distribuição TecidualRESUMO
Urotensin-II (U-II) is a vasoactive 'somatostatin-like' cyclic peptide which was originally isolated from fish spinal cords, and which has recently been cloned from man. Here we describe the identification of an orphan human G-protein-coupled receptor homologous to rat GPR14 and expressed predominantly in cardiovascular tissue, which functions as a U-II receptor. Goby and human U-II bind to recombinant human GPR14 with high affinity, and the binding is functionally coupled to calcium mobilization. Human U-II is found within both vascular and cardiac tissue (including coronary atheroma) and effectively constricts isolated arteries from non-human primates. The potency of vasoconstriction of U-II is an order of magnitude greater than that of endothelin-1, making human U-II the most potent mammalian vasoconstrictor identified so far. In vivo, human U-II markedly increases total peripheral resistance in anaesthetized non-human primates, a response associated with profound cardiac contractile dysfunction. Furthermore, as U-II immunoreactivity is also found within central nervous system and endocrine tissues, it may have additional activities.
Assuntos
Proteínas de Ligação ao GTP/agonistas , Proteínas de Ligação ao GTP/metabolismo , Receptores de Superfície Celular/agonistas , Receptores Acoplados a Proteínas G , Urotensinas/farmacologia , Vasoconstritores/farmacologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Cálcio/metabolismo , Linhagem Celular , Clonagem Molecular , DNA Complementar , Proteínas de Ligação ao GTP/genética , Humanos , Macaca fascicularis , Masculino , Dados de Sequência Molecular , Ratos , Ratos Sprague-Dawley , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Homologia de Sequência de Aminoácidos , Distribuição Tecidual , Urotensinas/metabolismo , Vasoconstritores/metabolismoRESUMO
OBJECTIVES: a trial of the use of integrated care pathways (ICPs) for elective vascular surgical procedures. DESIGN: a 12-month prospective study, following a multi-disciplinary group construction of current "best practice" ICPs, with changes in practice only occurring following careful audit of results. MATERIALS: patients admitted to a single vascular unit for "open" repair of abdominal aortic aneurysm, carotid endarterectomy or femoropopliteal bypass grafting. METHODS: patients followed ICPs on a daily basis with signatures required to confirm that action had been taken and careful recording of variances from the ICPs. Audit of variance data allowed changes in the ICPs and, hence, provision of the best possible nursing and clinical practice. RESULTS: ICPs were well received by patients and staff. They improved communication, promoted an appreciation of each health group's role in patient care, increased nursing autonomy, reduced calls to junior medical staff, improved patient education and confidence and caused a marked reduction in hospital "length of stay". CONCLUSIONS: ICPs have clear benefits. This study realises that benefits might be maximal for high throughput, high-cost procedures. Successful use of ICPs depends upon "clinical champions" and effective project management. Sufficient resource and training are essential.
Assuntos
Prestação Integrada de Cuidados de Saúde , Procedimentos Cirúrgicos Vasculares , Aneurisma da Aorta Abdominal/cirurgia , Estenose das Carótidas/cirurgia , Procedimentos Clínicos , Endarterectomia das Carótidas , Artéria Femoral/cirurgia , Humanos , Isquemia/cirurgia , Perna (Membro)/irrigação sanguínea , Tempo de Internação , Auditoria Médica , Avaliação de Processos e Resultados em Cuidados de Saúde , Equipe de Assistência ao Paciente , Projetos Piloto , Artéria Poplítea/cirurgia , Estudos ProspectivosRESUMO
BACKGROUND: Integrated care pathways (ICPs) represent a multidisciplinary approach to clinical patient care. METHODS: A 1-year prospective trial of the use of ICPs for elective vascular surgical procedures was undertaken. A multidisciplinary group constructed ICPs for patients admitted for open repair of abdominal aortic aneurysm, carotid endarterectomy or femoropopliteal bypass grafting. Patient management followed ICPs on a daily basis with signatures required to confirm that each action had been taken. Variances from the ICPs were carefully recorded. Audit of variance data allowed subsequent revision of the ICPs and hence provision of the best possible nursing and clinical practice. METHODS: A total of 33 patients were entered into the study; 16 had a femoropopliteal bypass graft, eight carotid endarterectomy and nine open repair of an abdominal aortic aneurysm. ICPs were well received by patients and staff. They improved communication, promoted an appreciation of each health group's role in patient care, increased nursing autonomy, reduced calls to junior medical staff, improved patient education and confidence, and caused a marked reduction in length of hospital stay. Overall, patients were discharged 13 per cent earlier after open abdominal aortic aneurysm repair, 22 per cent earlier after carotid endarterectomy and 38 per cent earlier after femoropopliteal bypass grafting. CONCLUSION: ICPs have clear benefits. They improve overall clinical efficiency and enhance clinical governance. Successful use of ICPs depends upon 'clinical champions' and effective project management. Sufficient resources and training are essential.
RESUMO
BACKGROUND: Activation of p38 mitogen-activated protein kinase (MAPK) plays an important role in apoptotic cell death. The role of p38 MAPK in myocardial injury caused by ischemia/reperfusion, an extreme stress to the heart, is unknown. METHODS AND RESULTS: Studies were performed with isolated, Langendorff-perfused rabbit hearts. Ischemia alone caused a moderate but transient increase in p38 MAPK activity (3.5-fold increase, P<0.05 versus basal). Ischemia followed by reperfusion further activated p38 MAPK, and the maximal level of activation (6.3-fold, P<0.01) was reached 10 minutes after reperfusion. Administration of SB 203580, a p38 MAPK inhibitor, decreased myocardial apoptosis (14.7+/-3.2% versus 30.6+/-3.5% in vehicle, P<0.01) and improved postischemic cardiac function. The cardioprotective effects of SB 203580 were closely related to its inhibition of p38 MAPK. Administering SB 203580 before ischemia and during reperfusion completely inhibited p38 MAPK activation and exerted the most cardioprotective effects. In contrast, administering SB 203580 10 minutes after reperfusion (a time point when maximal MAPK activation had already been achieved) failed to convey significant cardioprotection. Moreover, inhibition of p38 MAPK attenuated myocardial necrosis after a prolonged reperfusion. CONCLUSIONS: These results demonstrate that p38 MAPK plays a pivotal role in the signal transduction pathway mediating postischemic myocardial apoptosis and that inhibiting p38 MAPK may attenuate reperfusion injury.
Assuntos
Apoptose , Proteínas Quinases Dependentes de Cálcio-Calmodulina/antagonistas & inibidores , Proteínas Quinases Ativadas por Mitógeno , Isquemia Miocárdica/fisiopatologia , Traumatismo por Reperfusão Miocárdica/fisiopatologia , Miocárdio/enzimologia , Animais , Apoptose/efeitos dos fármacos , Fragmentação do DNA/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Imidazóis/farmacologia , Marcação In Situ das Extremidades Cortadas , Técnicas In Vitro , Isquemia Miocárdica/enzimologia , Isquemia Miocárdica/patologia , Traumatismo por Reperfusão Miocárdica/enzimologia , Traumatismo por Reperfusão Miocárdica/patologia , Miocárdio/patologia , Necrose , Piridinas/farmacologia , Coelhos , Fatores de Tempo , Proteínas Quinases p38 Ativadas por MitógenoRESUMO
The effects of SB 217242, a non-peptide endothelin (ET) receptor antagonist, were investigated against hypoxia-induced cardiopulmonary changes in high altitude-sensitive rats. In isolated pulmonary artery rings, SB 217242 (30 n m) antagonized ET-1-induced contractions with a p KB of 8.0. There was no difference in the sensitivity to ET-1 or the potency of SB 217242 in pulmonary artery from normoxic rats vs. rats exposed to hypoxia (9% O2) for 14 days. However, there was a marked reduction in the maximum response to ET-1, but not to KCl or phenylephrine, in pulmonary artery from hypoxic rats; this phenomenon was inhibited by treatment of animals with SB 217242 (10.8 mg/day, ip by osmotic pump) for the 14-day hypoxic period. Furthermore, there was a significant reduction in carbachol-induced, endothelium-dependent relaxation of precontracted pulmonary artery from hypoxic animals; SB 217242 treatment during the hypoxic period did not influence this difference. Vehicle-treated rats exposed to 14-day hypoxia had 173% higher pulmonary artery pressures and 75% higher right/left+septum ventricular mass ratios compared to normoxic animals. SB 217242 (3.6 or 10.8 mg/day, ip) markedly reduced (80 and 95%, respectively) hypoxia-induced increases in pulmonary artery pressure. Right ventricular hypertrophy was inhibited by 40% at the 10.8 mg/day dose. Marked medial thickening and luminal stenosis of small and medium-sized pulmonary arteries was observed in hypoxic rats. The SB 217242-treated, hypoxia-exposed rats had comparable small and medium-sized arteries to normoxic rats. Rats treated with SB 217242 (10.8 mg/day) for the last 14 days of a 28-day hypoxic exposure had significantly lower pulmonary artery pressures than those of vehicle-treated rats. In addition, the effects of the selective ETA receptor antagonist, SB 247083, and the selective ETB receptor antagonist, A-192621 (3.6 or 10.8 mg/day, ip), were compared against hypoxia-induced increases in pulmonary artery pressure and plasma ET concentrations. SB 247083, but not A-192621, inhibited hypoxia-induced pulmonary hypertension, whereas A-192621, but not SB 247083, significantly exacerbated hypoxia-induced increases in ET concentrations, suggesting that hypoxia-induced pulmonary pressor responses are mediated via ETA receptor activation, while ETB receptor blockade may alter clearance of hypoxia-induced elevated plasma ET. The inhibitory effects of SB 217242 on the functional and remodeling changes induced by hypoxia provide further evidence that ET may play a central role in pulmonary hypertension and that ET receptor antagonists may have a utility in the treatment of this disease.
Assuntos
Ácidos Carboxílicos/farmacologia , Antagonistas dos Receptores de Endotelina , Endotelina-1/metabolismo , Hipertensão Pulmonar/fisiopatologia , Hipertrofia Ventricular Direita/fisiopatologia , Hipóxia/metabolismo , Indanos/farmacologia , Artéria Pulmonar/efeitos dos fármacos , Altitude , Animais , Benzofuranos/farmacologia , Masculino , Propionatos/farmacologia , Pirrolidinas/farmacologia , Distribuição Aleatória , Ratos , Receptores de Endotelina/efeitos dos fármacosRESUMO
Embryo-fetal development studies with toxicokinetic evaluations were conducted in rats and rabbits after oral or intravenous administration of two endothelin receptor antagonists. In the rat studies, females were administered SB-217242 (0.01-300 mg/kg/day) orally or SB-209670 (0.01-50 mg/kg/day) intravenously from days 6-17 postcoitus (pc). External and visceral fetal examinations were performed at necropsy on day 21 pc. Maternal body weight and food consumption were decreased only at 300 mg/kg/day SB-217242. Embryolethality was seen at 300 mg/kg/day SB-217242. Decreased fetal body weight occurred at 300 mg/kg/day SB-217242 and 50 mg/kg/day SB-209670. Dose-dependent increases in the mean percentage of fetuses per litter with malformations were seen at > or = 50 mg/kg/day SB-217242 and > or = 10 mg/kg/day SB-209670. Craniofacial, great vessel, heart, and thyroid were the predominant malformations. In the rabbit studies, females were administered SB-217242 (0.01-50 mg/kg/day) orally or SB-209670 (0.01-25 mg/kg/day) intravenously from days 6-20 pc. There was no drug-related effect on maternal body weight or food consumption. Embryolethality was observed at 50 mg/kg/day of SB-217242. Dose-related increases in the mean percentage of fetuses per litter with malformations were seen at > or = 10 mg/kg/day SB-217242 and > or = 10 mg/kg/day SB-209670. The malformations were similar to those observed in the rat studies, except that craniofacial development was not altered by SB-209670. The malformations observed are consistent with the pattern of endothelin-1 gene expression described in mouse embryonic pharyngeal arches and heart, and with the craniofacial and cardiovascular malformations observed in endothelin-1-deficient mice. Given the known role for endothelins in development, and concordant malformations in rats and rabbits observed in this study, teratogenicity is likely to be a class effect of endothelin receptor antagonists.
Assuntos
Ácidos Carboxílicos/toxicidade , Desenvolvimento Embrionário e Fetal/efeitos dos fármacos , Antagonistas dos Receptores de Endotelina , Indanos/toxicidade , Anormalidades Induzidas por Medicamentos , Animais , Área Sob a Curva , Ácidos Carboxílicos/sangue , Ácidos Carboxílicos/farmacocinética , Relação Dose-Resposta a Droga , Feminino , Feto/efeitos dos fármacos , Indanos/sangue , Indanos/farmacocinética , Masculino , Gravidez , Coelhos , Ratos , Ratos Sprague-Dawley , Teratogênicos/farmacocinética , Teratogênicos/toxicidadeRESUMO
Structurally and pharmacologically diverse vasodilators are known to lower blood pressure, increase heart rate, and produce acute injury to right coronary arteries in the dog. Administration of low concentrations of endothelin-1 (ET-1) to anesthetized dogs causes coronary vasoconstriction and reductions in coronary blood flow. Therefore, pharmacologic blockade of endothelin receptors (ETA and ETB) with the mixed ET receptor antagonist SB 209670 could lead to coronary vasodilatation. In toxicology studies, continuous administration of SB 209670 to dogs for 5 days at 50 micrograms/kg/min was associated with minor but sustained increases in heart rate (10-30 beats/min), slight decreases in mean arterial pressure (10-15 mm Hg), and medial hemorrhage and necrosis of extramural coronary arteries in the right atria. Doses of 10 micrograms/kg/min had no effect. The lesions in the right atrium were associated with the highest density of ET receptors, approximately 470 fmol/mg compared to 170-200 fmol/mg in the ventricles and septum. Because changes in systemic cardiovascular parameters are minimal, the coronary arterial lesion is most likely due to a local vasodilatory effect in the coronary bed.
Assuntos
Doença das Coronárias/induzido quimicamente , Antagonistas dos Receptores de Endotelina , Indanos/toxicidade , Animais , Doença das Coronárias/fisiopatologia , Cães , Feminino , Indanos/metabolismo , Masculino , Receptores de Endotelina/metabolismo , Vasodilatação/efeitos dos fármacosRESUMO
The cardiopulmonary profile of three different rat strains was compared after exposure to hypoxia (9% O2) for 0, 7, or 14 days. In Sprague-Dawley (SD), Wistar (W), and high altitude-sensitive (HAS) rats, pulmonary arterial pressure (PAP) rose 30, 58, and 85% respectively, after 7 days of hypoxia, and by 108, 116, and 167%, respectively, at 14 days compared to strain- and age-matched normoxic controls. Right ventricular hypertrophy (RVH), expressed as the ratio of right free wall/left wall + septum weight, in SD, W, and HAS was increased by 24, 53, and 48%, respectively, at 7 days, and by 51, 93, and 55% at 14 days compared to normoxic littermates. Histologically, marked medial thickening and luminal stenosis of small and medium-sized arteries were observed in all hypoxic rats, being most pronounced in the HAS rats at 14 days. Treatment of HAS rats with the ET receptor antagonist SB 217242 (3.6 or 10.8 mg/day i.p. by osmotic pump) significantly inhibited the hypoxia-induced increases in PAP (70-75% decrease). RVH was inhibited by 40% at the dose of 10.8 mg/day. Histologically, the SB 217242-treated rats had almost "normal" small and medium-sized arteries, comparable to those of the normoxic HAS controls. This study demonstrates an exaggerated PAP response to chronic hypoxia in HAS compared to SD and W rats. The inhibitory influence of SB 217242 on the functional and morphologic changes induced by hypoxia provides further evidence for a role for ET and the potential utility of ET receptor antagonists in the treatment of pulmonary hypertension.
Assuntos
Ácidos Carboxílicos/uso terapêutico , Antagonistas dos Receptores de Endotelina , Hipóxia/tratamento farmacológico , Hipóxia/patologia , Indanos/uso terapêutico , Pulmão/patologia , Miocárdio/patologia , Animais , Pressão Sanguínea/efeitos dos fármacos , Doença Crônica , Hipóxia/genética , Masculino , Artéria Pulmonar/efeitos dos fármacos , Ratos , Ratos Endogâmicos , Ratos Sprague-Dawley , Fatores de TempoRESUMO
Focal brain ischemia induces inflammation, extracellular matrix remodeling, gliosis, and neovascularization. Osteopontin (OPN) is a secreted glycoprotein that has been implicated in vascular injury by promoting cell adhesion, migration, and chemotaxis. To investigate the possible involvement of OPN in brain matrix remodeling after focal stroke, we examined the expression of OPN in ischemic cortex after permanent or temporary occlusion of the middle cerebral artery (MCAO) of the rat. OPN mRNA and protein levels in nonischemic cortex were not detected consistently, although significant induction of OPN was observed in the ischemic cortex. OPN mRNA increased 3.5-fold at 12 hr and reached peak levels 5 d (49.5-fold; p < 0.001) after permanent MCAO. The profile of OPN mRNA induction after transient MCAO (160 min) with reperfusion was essentially the same as that of permanent MCAO. In situ hybridization and immunohistochemical studies demonstrated strong induction of OPN in the ischemic cortex, which was localized primarily in a subset of ED-1-positive macrophages that accumulated in the ischemic zone. Moreover, OPN immunoreactivity was detected in the matrix of ischemic brain, suggesting a functional role of the newly deposited matrix protein in cell-matrix interactions and remodeling. Indeed, using a modified Boyden chamber, we demonstrated a dose-dependent chemotactic activity of OPN in C6 astroglia cells and normal human astrocytes. Taken together, these data suggest that the upregulation of OPN after focal brain ischemia may play a role in cellular (glia, macrophage) migration/activation and matrix remodeling that provides for new matrix-cell interaction.
Assuntos
Transtornos Cerebrovasculares/metabolismo , Sialoglicoproteínas/metabolismo , Animais , Astrócitos/efeitos dos fármacos , Astrócitos/fisiologia , Isquemia Encefálica/metabolismo , Movimento Celular/efeitos dos fármacos , Movimento Celular/fisiologia , Córtex Cerebral/irrigação sanguínea , Humanos , Imuno-Histoquímica , Masculino , Osteopontina , RNA Mensageiro/metabolismo , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Sialoglicoproteínas/genética , Fatores de TempoRESUMO
Carvedilol, a new vasodilating beta-adrenoceptor antagonist and a potent antioxidant, produces a high degree of cardioprotection in a variety of experimental models of ischemic cardiac injury. Recent clinical studies in patients with heart failure have demonstrated that carvedilol reduces morbidity and mortality and inhibits cardiac remodeling. The present study was designed to explore whether the protective effects of carvedilol on the ischemic myocardium include inhibition of apoptosis of cardiomyocytes and, if so, to determine its mechanism of action. Anesthetized rabbits were subjected to 30 minutes of coronary artery occlusion followed by 4 hours of reperfusion. Detection of apoptosis of cardiomyocytes was based on the presence of nucleosomal DNA fragments on agarose gels (DNA ladder) and in situ nick end labeling. Carvedilol (1 mg/kg IV), administered 5 minutes before reperfusion, reduced the number of apoptotic myocytes in the ischemic area from 14.7 +/- 0.4% to 3.4 +/- 1.8% (77% reduction, P<.001). Propranolol, administered at equipotent beta-blocking dosage, reduced the number of apoptotic myocytes to 8.9 +/- 2.1% (39% reduction, P<.05). DNA ladders were observed in the hearts of all six vehicle-treated rabbits but only one of six carvedilol-treated rabbits (P<.01). Immunocytochemical analysis of rabbit hearts demonstrated an upregulation of Fas protein in ischemic cardiomyocytes, and treatment with carvedilol reduced both the intensity of staining as well as the area stained. Myocardial ischemia/reperfusion led to a rapid activation of stress-activated protein kinase (SAPK) in the ischemic area but not in nonischemic regions. SAPK activity was increased from 2.1 +/- 0.3 mU/mg (basal) to 8.9 +/- 0.8 mU/mg after 30 minutes of ischemia followed by 20 minutes of reperfusion. Carvedilol inhibited the activation of SAPK by 53.4 +/- 6.5% (P<.05). Under the same conditions, propranolol (1 mg/kg) had no effect on SAPK activation. Taken together, these results suggest that carvedilol prevents myocardial ischemia/reperfusion-induced apoptosis in cardiomyocytes possibly by downregulation of the SAPK signaling pathway, by inhibition of Fas receptor expression, and by beta-adrenergic blockade. The former two actions represent novel and important mechanisms that may contribute to the cardioprotective effects of carvedilol.
Assuntos
Antagonistas Adrenérgicos beta/uso terapêutico , Apoptose/efeitos dos fármacos , Carbazóis/uso terapêutico , Glicoproteínas de Membrana/metabolismo , Isquemia Miocárdica/prevenção & controle , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Propanolaminas/uso terapêutico , Proteínas Quinases/metabolismo , Animais , Carvedilol , Fragmentação do DNA/fisiologia , Regulação para Baixo/fisiologia , Ativação Enzimática/fisiologia , Proteína Ligante Fas , Hemodinâmica/efeitos dos fármacos , Masculino , Isquemia Miocárdica/patologia , Isquemia Miocárdica/fisiopatologia , Traumatismo por Reperfusão Miocárdica/patologia , Traumatismo por Reperfusão Miocárdica/fisiopatologia , Miocárdio/metabolismo , Coelhos , Transdução de Sinais/fisiologia , Estresse Fisiológico/metabolismoRESUMO
In the 3-oxo-1,4-benzodiazepine-2-acetic acid series of vitronectin receptor (alpha v beta 3) antagonists, a compound containing an imidazopyridine arginine mimetic was discovered which had sufficient potency and i.v. pharmacokinetics for demonstration of efficacy in a rat restenosis model.
Assuntos
Angioplastia Coronária com Balão/efeitos adversos , Benzodiazepinas/uso terapêutico , Doença das Coronárias/tratamento farmacológico , Imidazóis/uso terapêutico , Piridinas/uso terapêutico , Receptores de Vitronectina/antagonistas & inibidores , Animais , Modelos Animais de Doenças , RatosRESUMO
2-Methoxyestradiol (2-ME) is an endogenous metabolite of estradiol-17beta and the oral contraceptive agent 17-ethylestradiol. 2-ME was recently reported to inhibit endothelial cell proliferation. The current study was undertaken to explore the mechanism of 2-ME effects on endothelial cells, especially whether 2-ME induces apoptosis, a prime mechanism in tissue remodeling and angiogenesis. Cultured bovine pulmonary artery endothelial cells (BPAEC) exposed to 2-ME showed morphological (including ultrastructural) features characteristic of apoptosis: cell shrinkage, cytoplasmic and nuclear condensation, and cell blebbing. 2-ME-induced apoptosis in BPAEC was a time- and concentration-dependent process (EC50 = 0.45 +/- 0.09 microM, n = 8). Nucleosomal DNA fragmentation in BPAEC treated with 2-ME was identified by agarose gel electrophoresis (DNA ladder) as well as in situ nick end labeling. Under the same experimental conditions, estradiol-17beta and two of its other metabolites, estriol and 2-methoxyestriol (< or =10 microM), did not have an apoptotic effect on BPAEC. 2-ME activated stress-activated protein kinase (SAPK)/c-Jun amino-terminal protein kinase in BPAEC in a concentration-dependent manner. The activity of SAPK was increased by 170 +/- 27% and 314 +/- 22% over the basal level in the presence of 0.4 and 2 microM 2-ME (n = 3-6), respectively. The activation of SAPK was detected at 10 min, peaked at 20 min, and returned to basal levels at 60 min after exposure to 2-ME. Inhibition of SAPK/c-Jun amino-terminal protein kinase activation by basic fibroblast growth factor, insulin-like growth factor, or forskolin reduced 2-ME-induced apoptosis. Immunohistochemical analysis of BPAEC indicated that 2-ME up-regulated expression of both Fas and Bcl-2. In addition, 2-ME inhibited BPAEC migration (IC50 = 0.71 +/- 0.11 microM, n = 4) and basic fibroblast growth factor-induced angiogenesis in the chick chorioallantoic membrane model. Taken together, these results suggest that promotion of endothelial cell apoptosis, thereby inhibiting endothelial cell proliferation and migration, may be a major mechanism by which 2-ME inhibits angiogenesis.