RESUMEN
BACKGROUND: D-Penicillamine is the most commonly used copper-chelating agent in the treatment of copper-associated hepatitis in dogs. Response to therapy can be variable, and there is a lack of pharmacokinetic information available for dogs. Coadministering the drug with food to alleviate vomiting has been recommended for dogs, which contradicts recommendations for drug administration to humans. HYPOTHESIS: Coadministration of d-penicillamine with food decreases relative bioavailability and maximum plasma drug concentrations (C(max)) in dogs. ANIMALS: Nine purpose-bred dogs with a median body weight of 17.0 kg. METHODS: Dogs received D-penicillamine (12.5 mg/kg PO) fasted and with food in a randomized, crossover design. Blood samples were collected before and 0.25, 0.5, 1, 2, 3, 4, 8, 12, and 24 hours after dosing. Total d-penicillamine concentrations were measured using liquid chromatography coupled with tandem quadrupole mass spectrometry. Pharmacokinetic parameters were calculated for each dog. RESULTS: Two fasted dogs (22%) vomited after receiving d-penicillamine. Mean C(max) ± standard deviation (SD) was 8.7 ± 3.1 µg/mL (fasted) and 1.9 ± 1.6 µg/mL (fed). Mean area under the plasma concentration curve ± SD was 16.9 ± 5.9 µg/mL·h (fasted) and 4.9 ± 3.4 µg/mL·h (fed). There were significant reductions in relative bioavailability and C(max) in fed dogs (P < .001). CONCLUSIONS AND CLINICAL IMPORTANCE: Coadministration of d-penicillamine with food significantly decreases plasma drug concentrations in dogs. Decreased drug exposure could result in decreased copper chelation efficacy, prolonged therapy, additional cost, and greater disease morbidity. Administration of d-penicillamine with food cannot be categorically recommended without additional studies.
Asunto(s)
Quelantes/farmacocinética , Perros/sangre , Privación de Alimentos/fisiología , Penicilamina/farmacocinética , Animales , Área Bajo la Curva , Femenino , Semivida , Masculino , Penicilamina/sangreAsunto(s)
Atrofia Óptica Hereditaria de Leber/tratamiento farmacológico , Ubiquinona/análogos & derivados , Adolescente , Adulto , Anciano , Estudios de Cohortes , Método Doble Ciego , Femenino , Estudios de Seguimiento , Humanos , Masculino , Persona de Mediana Edad , Atrofia Óptica Hereditaria de Leber/epidemiología , Atrofia Óptica Hereditaria de Leber/fisiopatología , Factores de Tiempo , Resultado del Tratamiento , Ubiquinona/farmacología , Ubiquinona/uso terapéutico , Agudeza Visual/efectos de los fármacos , Agudeza Visual/fisiología , Adulto JovenRESUMEN
PURPOSE: The majority of patients with autosomal dominant optic atrophy (DOA) harbour pathogenic OPA1 mutations. Although DOA is characterised by the preferential loss of retinal ganglion cells (RGCs), about 20% of patients with OPA1 mutations will develop a more severe disease variant (DOA+), with additional neuromuscular features. In this prospective, observational case series, optical coherence tomography (OCT) was used to define the pattern of retinal nerve fibre layer (RNFL) loss in patients with both the pure and syndromal forms of DOA. METHODS: Forty patients with a molecular diagnosis of DOA due to OPA1 mutations were prospectively recruited from our neuro-ophthalmology clinic: 26 patients with isolated optic atrophy and 14 patients manifesting DOA+ features. Peripapillary RNFL thickness was measured with the Fast RNFL (3.4) acquisition protocol on a Stratus OCT. RESULTS: There was a statistically significant reduction in average RNFL thickness in the OPA1 group compared with normal controls (P<0.0001). The percentage decrease was greatest in the temporal quadrant (59.0%), followed by the inferior (49.6%), superior (41.8%), and nasal (25.9%) quadrants. Patients with DOA+ features had worse visual outcomes compared with patients with pure DOA. Except in the temporal quadrant, RNFL measurements were significantly thinner for the DOA+ group. There was an inverse correlation between average RNFL thickness and logarithm of the minimum angle of resolution (LogMAR) visual acuity (P<0.0001). CONCLUSIONS: RGC loss in DOA is characterised by severe involvement of the temporal papillomacular bundle, with relative sparing of the nasal fibres. RNFL thinning is more pronounced in patients with DOA+ phenotypes.
Asunto(s)
GTP Fosfohidrolasas/genética , Mutación , Atrofia Óptica Autosómica Dominante/genética , Atrofia Óptica Autosómica Dominante/patología , Células Ganglionares de la Retina/patología , Adolescente , Adulto , Anciano , Niño , Estudios de Cohortes , Femenino , Humanos , Masculino , Persona de Mediana Edad , Disco Óptico/patología , Estudios Prospectivos , Retina/patología , Tomografía de Coherencia Óptica/métodos , Agudeza Visual , Adulto JovenRESUMEN
Evidence derived from human and animal studies strongly supports the notion that dysfunctional alveolar epithelial cells (AECs) play a central role in determining the progression of inflammatory injury to pulmonary fibrosis. We formed the hypothesis that impaired production of the regulatory cytokine granulocyte-macrophage colony-stimulating factor (GM-CSF) by injured AECs plays a role in the development of pulmonary fibrosis. To test this hypothesis, we used the well-characterized model of bleomycin-induced pulmonary fibrosis in rats. GM-CSF mRNA is expressed at a constant high level in the lungs of untreated or saline-challenged animals. In contrast, there is a consistent reduction in expression of GM-CSF mRNA in the lung during the first week after bleomycin injury. Bleomycin-treated rats given neutralizing rabbit anti-rat GM-CSF IgG develop increased fibrosis. Type II AECs isolated from rats after bleomycin injury demonstrate diminished expression of GM-CSF mRNA immediately after isolation and in response to stimulation in vitro with endotoxin compared with that in normal type II cells. These data demonstrate a defect in the ability of type II epithelial cells from bleomycin-treated rats to express GM-CSF mRNA and a protective role for GM-CSF in the pathogenesis of bleomycin-induced pulmonary fibrosis.
Asunto(s)
Bleomicina , Factor Estimulante de Colonias de Granulocitos y Macrófagos/metabolismo , Alveolos Pulmonares/metabolismo , Fibrosis Pulmonar/inducido químicamente , Animales , Anticuerpos/farmacología , Células Cultivadas , Epitelio/efectos de los fármacos , Epitelio/metabolismo , Epitelio/patología , Factor Estimulante de Colonias de Granulocitos y Macrófagos/genética , Factor Estimulante de Colonias de Granulocitos y Macrófagos/inmunología , Hidroxiprolina/metabolismo , Masculino , Alveolos Pulmonares/efectos de los fármacos , Alveolos Pulmonares/patología , Fibrosis Pulmonar/metabolismo , Fibrosis Pulmonar/patología , ARN Mensajero/metabolismo , Ratas , Ratas Endogámicas F344RESUMEN
Intercellular adhesion molecule-1 (ICAM-1) is an adhesion protein involved in immune and inflammatory cell recruitment and activation. In normal, uninflamed adult rat lung, ICAM-1 is expressed at high levels on type I alveolar epithelial cells and is minimally expressed on type II cells. ICAM-1 expression by alveolar epithelial cells in vitro is a function of the state of cellular differentiation, and is regulated by factors influencing cell shape. Based upon this observation, we hypothesized that ICAM-1 expression by fetal lung epithelial cells is developmentally regulated. To investigate this hypothesis, rat and human lung tissues were obtained at time points that represent the canalicular, saccular, and alveolar stages of development. The relative expression of ICAM-1 protein and mRNA were determined in rat lungs from gestational days 18 and 21 (term = 22 days), from day 8 neonatal rats, and from adult rats. ICAM-1 protein was detectable at low level on day 18 and increased progressively during development. Relative expression of ICAM-1 protein was maximal in adult lung. Expression of ICAM-1 mRNA paralleled that of ICAM-1 protein. By immunohistochemical methods in rat and human lung, ICAM-1 was expressed at low level on cuboidal and flattening epithelial cells in the developing alveolar space at the canalicular and saccular stages; however, ICAM-1 expression was increased as epithelial cells spread and flattened during alveolarization. ICAM-1 was predominantly expressed on type I cells rather than type II cells at the alveolar stage in both the rat and human lungs. Thus, relative ICAM-1 expression progressively increased during lung development. ICAM-1 expression is correlated with the increase in surface area as alveolar structures develop and type I cell differentiation takes place. These data indicate that alveolar epithelial cell ICAM-1 expression is developmentally regulated.
Asunto(s)
Células Epiteliales/metabolismo , Regulación del Desarrollo de la Expresión Génica , Molécula 1 de Adhesión Intercelular/genética , Alveolos Pulmonares/metabolismo , Adulto , Animales , Animales Recién Nacidos , Western Blotting , Desarrollo Embrionario y Fetal , Células Epiteliales/citología , Femenino , Edad Gestacional , Humanos , Técnicas para Inmunoenzimas , Molécula 1 de Adhesión Intercelular/biosíntesis , Embarazo , Alveolos Pulmonares/citología , Alveolos Pulmonares/embriología , ARN Mensajero/biosíntesis , Ratas , Ratas Sprague-DawleyRESUMEN
The alveolar macrophage (AM) exhibits a greater capacity to synthesize bioactive leukotrienes from arachidonic acid than does its circulating precursor the peripheral blood monocyte. Macrophage differentiation in the lung entails cellular residence within both the pulmonary interstitial and alveolar compartments. In the present study, we sought to determine 1) whether this enhanced metabolic activity was acquired during maturation within the alveolar space and 2) the underlying mechanisms responsible for this upregulation. Rat AMs were separated by Percoll gradient centrifugation into four density-defined subpopulations thought to reflect their degree of maturation. On stimulation with a calcium ionophore, synthesis of leukotriene B4 increased with the degree of maturation, although it was diminished in the oldest subpopulation. This maturation-dependent upregulation was not explained by increases in arachidonic acid release but was associated with increased expression of 5-lipoxygenase (5-LO) protein as determined by immunoblot analysis. Whereas 5-LO is primarily cytosolic in monocytes, it is known to be primarily intranuclear in unfractionated AMs. Here, the localization of 5-LO was investigated by immunofluorescence microscopy and was found to be predominantly nuclear in all AM subpopulations; by contrast, the protein was cytosolic in interstitial macrophages isolated by mechanical and enzymatic lung digestion. These divergent localization patterns in AMs and interstitial macrophages were verified in situ by immunohistochemical staining of sections of normal rat lung. When unfractionated AMs were isolated and maintained in culture for 3 days, a shift in 5-LO distribution from nucleus to cytosol was observed. We conclude that 1) nuclear import of 5-LO occurs within the alveolar space and is reversible on removal from the alveolar milieu and 2) leukotriene synthetic capacity increases further during AM residence within the alveolar space as a result of a progressive increase in the amount of 5-LO protein.
Asunto(s)
Araquidonato 5-Lipooxigenasa/biosíntesis , Pulmón/citología , Alveolos Pulmonares/citología , Alveolos Pulmonares/enzimología , Animales , Araquidonato 5-Lipooxigenasa/análisis , Diferenciación Celular , Separación Celular , Centrifugación por Gradiente de Densidad , Femenino , Inmunohistoquímica , Pulmón/fisiología , Ratas , Ratas WistarRESUMEN
The intravenous administration of lipopolysaccharide (LPS) to rats results in liver lesions characterized by the infiltration of both platelets and neutrophils into the liver and by midzonal hepatocellular necrosis. The liver injury is dependent on neutrophils, platelets and the coagulation system, as removal or inhibition of any of these components inhibits the development of hepatocellular necrosis. Platelet activating factor (PAF) and the cysteinyl leukotrienes (LTs) are potent inflammatory lipids that are critical for the development of some LPS-mediated alterations. To test the hypothesis that PAF, alone or in combination with LTs, contributes to the development of liver injury during LPS exposure, we conducted studies with the PAF receptor antagonist, WEB 2086, and the 5-lipoxygenase inhibitor, Zileuton. Female, Sprague-Dawley rats were pretreated with WEB 2086 (10 mg/kg, i.p.) alone or with Zileuton (40 mg/kg, p.o.) 1 h before the administration of LPS (4 mg/kg, i.v.) or its saline vehicle. Treatment with WEB 2086, alone or in combination with Zileuton, did not inhibit LPS-mediated hepatic neutrophil infiltration or liver injury, as assessed by histologic evaluation and increases in plasma alanine aminotransferase activity. Pretreatment with these agents also had no effect on the activation of the coagulation system or on the thrombocytopenia induced by LPS. These results suggest that PAF, alone or in combination with 5-lipoxygenase products, is not a critical mediator of LPS-induced hepatocellular injury in this model.
Asunto(s)
Leucotrienos/fisiología , Lipopolisacáridos/toxicidad , Hígado/efectos de los fármacos , Factor de Activación Plaquetaria/fisiología , Administración Oral , Alanina Transaminasa/sangre , Animales , Azepinas/administración & dosificación , Azepinas/farmacología , Azepinas/uso terapéutico , Plaquetas/efectos de los fármacos , Plaquetas/patología , Enfermedad Hepática Inducida por Sustancias y Drogas , Femenino , Fibrinógeno/análisis , Fibrinógeno/efectos de los fármacos , Hidroxiurea/administración & dosificación , Hidroxiurea/análogos & derivados , Hidroxiurea/farmacología , Hidroxiurea/uso terapéutico , Inyecciones Intraperitoneales , Inyecciones Intravenosas , Lipopolisacáridos/administración & dosificación , Inhibidores de la Lipooxigenasa/administración & dosificación , Inhibidores de la Lipooxigenasa/farmacología , Inhibidores de la Lipooxigenasa/uso terapéutico , Hígado/citología , Hígado/patología , Hepatopatías/patología , Hepatopatías/prevención & control , Necrosis , Neutrófilos/efectos de los fármacos , Neutrófilos/patología , Inhibidores de Agregación Plaquetaria/administración & dosificación , Inhibidores de Agregación Plaquetaria/farmacología , Inhibidores de Agregación Plaquetaria/uso terapéutico , Ratas , Ratas Sprague-Dawley , Trombocitopenia/tratamiento farmacológico , Trombocitopenia/prevención & control , Triazoles/administración & dosificación , Triazoles/farmacología , Triazoles/uso terapéuticoRESUMEN
5-Lipoxygenase catalyzes the synthesis of leukotrienes from arachidonic acid. The subcellular distribution of 5-lipoxygenase is known to be cell type-dependent and is cytosolic in blood neutrophils. In this study, we asked whether neutrophil recruitment into sites of inflammation can alter the subcellular compartmentation of 5-lipoxygenase. In peripheral blood neutrophils from rats, 5-lipoxygenase was exclusively cytosolic, as expected. However, in glycogen-elicited peritoneal neutrophils, abundant soluble 5-lipoxygenase was in the nucleus. Upon activation with calcium ionophore A23187, intranuclear 5-lipoxygenase translocated to the nuclear envelope. Elicited neutrophils required a greater concentration of A23187 for activation than did blood neutrophils (half-maximal response, 160 versus 52 nM, respectively) but generated greater amounts of leukotriene B4 upon maximal stimulation (26.6 versus 7.68 ng/10(6) cells, respectively). Intranuclear 5-lipoxygenase was also evident in human blood neutrophils after adherence to a variety of surfaces, suggesting that adherence alone is sufficient to drive 5-lipoxygenase redistribution. These results demonstrate a physiologically relevant circumstance in which the subcellular distribution of 5-lipoxygenase can be rapidly altered in resting cells, independent of 5-lipoxygenase activation. Nuclear import of 5-lipoxygenase may be a universal accompaniment of neutrophil recruitment into sites of inflammation, and this may be associated with alterations in enzymatic function.
Asunto(s)
Araquidonato 5-Lipooxigenasa/metabolismo , Neutrófilos/enzimología , Animales , Calcimicina/farmacología , Adhesión Celular , Núcleo Celular/enzimología , Citosol/enzimología , Activación Enzimática , Glucógeno/metabolismo , Humanos , Leucotrieno B4/metabolismo , RatasRESUMEN
Leukotrienes (LTs) are potent mediators of inflammation derived from the 5-lipoxygenase pathway of arachidonic acid metabolism. Although they are known to enhance leukocyte recruitment and function, their role in antimicrobial host defense has not been established. To determine the role of endogenous LTs in the host response to pulmonary infection, wild-type mice and mice rendered LT-deficient by targeted disruption of the 5-lipoxygenase gene (knockout mice) were studied following intratracheal challenge with Klebsiella pneumoniae. Wild-type mice demonstrated a marked increase in lung LT levels and neutrophil numbers following bacterial challenge. As compared with wild-type animals, knockout animals manifested a greater degree of lethality as well as bacteremia following challenge. Interestingly, they displayed no defect in neutrophil recruitment to the lung. However, alveolar macrophages from knockout animals exhibited impairments in bacterial phagocytosis and killing, and these defects were overcome by in vitro addition of exogenous LTB4. We conclude that endogenous LTs play a critical role in the defense against bacterial pneumonia in this murine model.
Asunto(s)
Araquidonato 5-Lipooxigenasa/deficiencia , Leucotrienos/fisiología , Macrófagos Alveolares/inmunología , Neumonía Bacteriana/inmunología , Animales , Actividad Bactericida de la Sangre , Inmunidad Celular , Klebsiella pneumoniae , Leucotrieno B4/fisiología , Ratones , Ratones Noqueados , FagocitosisRESUMEN
Tumor necrosis factor (TNF) is a proinflammatory cytokine which has recently been shown to have beneficial effects in the setting of acquired host immunity. However, the role of TNF in innate immune responses, as in the setting of bacterial pneumonia, has been incompletely characterized. To determine the role of TNF in gram-negative bacterial pneumonia, CBA/J mice were challenged with 10(2) CFU of Klebsiella pneumoniae intratracheally, resulting in the time-dependent expression of TNF MRNA and protein within the lung. Passive immunization of animals with a soluble TNF receptor-immunoglobulin (Ig) construct (sTNFR:Fc) intraperitoneally 2 h prior to K. pneumoniae inoculation resulted in a significant reduction in bronchoalveolar lavage neutrophils, but not macrophages, at 48 h, as compared with animals receiving control IgG1. Furthermore, treatment with sTNFR:Fc resulted in 19.6- and 13.5-fold increases in K. pneumoniae CFU in lung homogenates and plasma, respectively, as compared with animals receiving control IgG1. Finally, treatment of Klebsiella-infected mice with sTNFR:Fc markedly decreased both short- and long-term survival of these animals. In conclusion, our studies indicate that endogenous TNF is a critical component of antibacterial host defense in murine Klebsiella pneumonia.
Asunto(s)
Infecciones por Klebsiella/inmunología , Klebsiella pneumoniae , Pulmón/microbiología , Neumonía Bacteriana/inmunología , Factor de Necrosis Tumoral alfa/biosíntesis , Animales , Inmunización , Pulmón/inmunología , Ratones , Ratones Endogámicos CBA , ARN Mensajero/biosíntesis , Receptores del Factor de Necrosis Tumoral/inmunología , Factor de Necrosis Tumoral alfa/inmunologíaRESUMEN
Idiopathic pulmonary fibrosis (IPF) is a progressive disorder characterized by inflammation, fibroblast proliferation, and accumulation of extracellular matrix proteins. Leukotrienes (LTs) are pro-inflammatory and pro-fibrogenic mediators derived from the 5-lipoxygenase (5-LO) pathway of arachidonic acid metabolism. They are thought to play a role in a number of disease processes, but have received relatively little attention in investigations into the pathogenesis of IPF. In this study, we measured the levels of immunoreactive LTs B(4) and C(4) in homogenates of lung tissue obtained from patients with newly diagnosed, untreated IPF, as compared to levels measured in homogenates of uninvolved nonfibrotic lung tissue from patients undergoing resectional surgery for bronchogenic carcinoma. Compared to homogenates on nonfibrotic control lung, homogenates from IPF patients contained 15-fold more LTB(4) and 5-fold more LTC(4). IPF homogenate levels of LTB(4) were significantly correlated with histologic indices of both inflammation (r=0.861) and fibrosis (r=0.926). Activation of 5-LO is known from in vitro studies to be associated with localization of the enzyme at the nuclear membrane. Immunohistochemical staining for 5-LO protein in alveolar macrophages (AMs) demonstrated that such an "activated" localization pattern was significantly more frequent in IPF lung (19.2+/-3.3% of cells) than in control lung (9.3+/-0.9%); this localization pattern was rarely seen (3.2%) in sections from a truly normal transplant donor lung. Consistent with these data, AMs obtained from IPF patients by bronchoalveolar lavage, purified by adherence, and cultured in the absence of a stimulus for 16 h elaborated significantly greater amounts of LTB(4) and LTC(4) than did control AMs obtained from normal volunteers. These data indicate that the 5-LO pathway is constitutively activated in the lungs of patients with IPF, and the AM represents at least one cellular source of LT overproduction in this disorder. We speculate that LTs participate in the pathogenesis of IPF, and their overproduction in this disorder may be amenable to specific pharmacotherapy.
Asunto(s)
Araquidonato 5-Lipooxigenasa/metabolismo , Pulmón/enzimología , Fibrosis Pulmonar/enzimología , Adulto , Anciano , Araquidonato 5-Lipooxigenasa/análisis , Células Cultivadas , Activación Enzimática , Femenino , Humanos , Inmunohistoquímica , Inflamación , Leucotrieno B4/análisis , Leucotrieno C4/análisis , Neoplasias Pulmonares/enzimología , Neoplasias Pulmonares/patología , Macrófagos Alveolares/enzimología , Macrófagos Alveolares/patología , Masculino , Persona de Mediana Edad , Fibrosis Pulmonar/patología , FumarRESUMEN
Alpha-naphthylisothiocyanate (ANIT) is a cholangiolitic hepatotoxicant that causes periportal edema, hepatic parenchymal and biliary epithelial cell necrosis, and cholestasis in the rat. A hallmark of ANIT hepatotoxicity is periportal inflammation that includes neutrophil infiltration. Neutrophils are requisite for the expression of ANIT-induced liver injury; however, the mechanism(s) of neutrophil accumulation in the liver and the role of these cells in ANIT hepatotoxicity is incompletely understood. Platelet activating factor (PAF) is a proinflammatory agent that is both a chemoattractant for and an activator of neutrophils. Therefore, we evaluated the role of PAF in ANIT-induced liver injury. Rats were treated with the PAF receptor antagonist, WEB 2086 (WEB), to determine if it afforded protection from ANIT hepatotoxicity. In a separate study, a combination of WEB and the leukotriene synthesis inhibitor, Zileuton (ZIL), was used to address the possible interaction of PAF and leukotrienes in ANIT-induced liver injury. Treatment of rats with WEB, alone or in combination with Zileuton, did not attenuate ANIT-induced liver injury as assessed by increases in plasma alanine aminotransferase or gamma-glutamyl transferase activities. In addition, neither treatment ameliorated ANIT-induced cholestasis assessed as increased plasma bilirubin concentration. These results suggest that PAF, alone or in combination with products of 5-lipoxygenase, does not contribute to ANIT-induced liver injury.
Asunto(s)
1-Naftilisotiocianato/toxicidad , Leucotrienos/fisiología , Hígado/efectos de los fármacos , Factor de Activación Plaquetaria/fisiología , Animales , Azepinas/farmacología , Hidroxiurea/análogos & derivados , Hidroxiurea/farmacología , Inhibidores de la Lipooxigenasa/farmacología , Masculino , Factor de Activación Plaquetaria/antagonistas & inhibidores , Ratas , Ratas Sprague-Dawley , Triazoles/farmacologíaRESUMEN
alpha-Naphthylisothiocyanate (ANIT) is a cholangiolitic hepatotoxicant that causes periportal hepatic injury in the rat that is neutrophil- and platelet-dependent. Since macrophages have recently been implicated as participants in some chemically induced hepatotoxicities, we evaluated the role of these cells in ANIT-induced hepatic injury. Rats were treated with gadolinium chloride (GdCl3), an agent which decreases hepatic macrophage numbers and activity, zymosan, an agent which increases hepatic macrophage numbers, or vitamin A, which increases hepatic macrophage activity. GdCl3 did not ameliorate ANIT-induced hepatotoxicity, as demonstrated by a lack of attenuation of any of the markers of hepatic insult evaluated. In contrast, pretreatment with either zymosan or vitamin A decreased ANIT hepatotoxicity. Zymosan administration reduced blood neutrophil numbers and influx of neutrophils into the peritoneum after intraperitoneal glycogen administration but did not affect hepatic neutrophil accumulation in ANIT-treated rats. To determine if macrophages were important in the protection by vitamin A, rats were cotreated with GdCl3 and vitamin A. GdCl3 did not alter the protection from ANIT hepatotoxicity afforded by vitamin A. Vitamin A treatment decreased ANIT and glutathione concentrations in bile at 1 and 4 hr after ANIT administration but had a minimal effect on plasma ANIT concentration. In summary, pretreatment of rats with zymosan or vitamin A but not GdCl3 attenuated ANIT-induced liver injury. The protection afforded by zymosan may derive from its effects on neutrophils or platelets. The protection by vitamin A appears to result from its effect on the transport of ANIT into bile. The results suggest that hepatic macrophages are not required for the manifestation of ANIT hepatotoxicity.
Asunto(s)
1-Naftilisotiocianato/antagonistas & inhibidores , Hígado/efectos de los fármacos , Vitamina A/farmacología , Zimosan/farmacología , 1-Naftilisotiocianato/toxicidad , Animales , Antiinflamatorios/farmacología , Gadolinio/farmacología , Glucógeno/farmacología , Macrófagos del Hígado/efectos de los fármacos , Macrófagos del Hígado/metabolismo , Recuento de Leucocitos , Hígado/metabolismo , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Masculino , Ratas , Ratas Sprague-Dawley , Vitamina A/administración & dosificación , Zimosan/administración & dosificaciónRESUMEN
alpha-naphthylisothiocyanate (ANIT) administration to rats results in periportal hepatic inflammation and injury. Glutathione (GSH) appears to be necessary for the liver injury to occur. The leukotrienes (LTs) are metabolites of arachidonic acid and potent mediators of inflammation that have been implicated in certain liver injury models. Inasmuch as GSH is a cofactor for the synthesis of cysteinyl-LTs and since inflammation is a prominent component of ANIT injury, we hypothesized that LTs are involved in producing the hepatic insult that results from ANIT administration. To test this hypothesis, rats were treated with one of several inhibitors of LT biosynthesis, A63162, Zileuton or MK-886. Each of these agents prevented the formation of LTB4 in Ca++ ionophore-stimulated whole blood from rats treated with the inhibitors. A63162 attenuated the hepatic parenchymal injury caused by ANIT and resulted in a modest decrease in ANIT-induced cholestasis. In contrast, neither Zileuton nor MK-886 attenuated liver injury. AT-125 (Acivicin) inhibits gamma-glutamyl transferase (GGT), the enzyme that catalyzes the formation of LTD4 from LTC4. AT-125 pretreatment did not prevent ANIT-induced hepatic parenchymal insult. It did, however, ameliorate the cholestasis caused by ANIT. In conclusion, the partial protection afforded by A63162 and AT-125 likely results from effects unrelated to the formation of LTs, since Zileuton and MK-886 inhibited LT synthesis without affording protection. The lack of protection by Zileuton and MK-886 in the face of LT synthesis inhibition suggests that LTs are not necessary for the expression of injury after ANIT administration.
Asunto(s)
1-Naftilisotiocianato/toxicidad , Leucotrienos/fisiología , Hígado/efectos de los fármacos , Acetamidas/administración & dosificación , Acetamidas/farmacología , Acetamidas/uso terapéutico , Animales , Antiinflamatorios no Esteroideos/farmacología , Calcimicina/farmacología , Enfermedad Hepática Inducida por Sustancias y Drogas , Modelos Animales de Enfermedad , Glutatión/metabolismo , Hidroxiurea/administración & dosificación , Hidroxiurea/análogos & derivados , Hidroxiurea/farmacología , Hidroxiurea/uso terapéutico , Indoles/administración & dosificación , Indoles/farmacología , Indoles/uso terapéutico , Inflamación/inducido químicamente , Inflamación/tratamiento farmacológico , Antagonistas de Leucotrieno , Leucotrienos/biosíntesis , Inhibidores de la Lipooxigenasa/administración & dosificación , Inhibidores de la Lipooxigenasa/farmacología , Hígado/patología , Hepatopatías/tratamiento farmacológico , Masculino , Éteres Fenílicos , Ratas , Ratas Sprague-DawleyRESUMEN
1-Naphthylisothiocyanate (ANIT) has been used for many years to study cholangiolitic hepatotoxicity in laboratory animals. Hallmarks of ANIT hepatotoxicity include portal edema and inflammation with bile duct epithelial and hepatic parenchymal cell necrosis. In rats, ANIT hepatotoxicity is dependent upon hepatic glutathione. Studies in vitro have demonstrated that ANIT combines reversibly with glutathione and suggest that intracellular formation and secretion of this glutathione-ANIT conjugate from hepatic parenchymal cells may be responsible for the efflux of glutathione observed upon exposure to ANIT. In vivo, glutathione conjugates produced within hepatic parenchymal cells are typically transported into bile for elimination. Therefore, large concentrations of ANIT in bile may result from hepatic parenchymal cell secretion of a reversible glutathione-ANIT conjugate. To investigate this hypothesis, bile and plasma concentrations of ANIT were determined in rats 1, 4, 8, 12 and 24 hr after administration (100 mg/kg, p.o.). Liver and bile glutathione concentrations were also evaluated. Plasma ANIT concentrations ranged between 2 and 5 microM at 1, 4, 8 and 12 hr and were 0.9 microM at 24 hr after administration. ANIT concentrations in bile at 1, 4, 8 and 12 hr were 60, 28, 21 and 22 microM, respectively. Thus, ANIT was concentrated in bile. Hepatic glutathione was not affected by ANIT during the first 12 hr after administration; however, a moderate elevation occurred by 24 hr. In contrast, a marked elevation in bile glutathione concentration (two times control) occurred 1, 4 and 8 hr after ANIT administration. Thus, the early accumulation of ANIT in bile was coincident with an elevation in bile glutathione. These findings support the hypothesis that glutathione functions to concentrate ANIT in bile. The large concentration of this toxicant in bile may be injurious to bile epithelium, a primary cellular target in ANIT hepatotoxicity.
Asunto(s)
1-Naftilisotiocianato/farmacología , Bilis/metabolismo , Glutatión/metabolismo , 1-Naftilisotiocianato/sangre , 1-Naftilisotiocianato/metabolismo , Animales , Glutatión/química , Hígado/efectos de los fármacos , Hígado/metabolismo , Masculino , Ratas , Ratas Sprague-DawleyRESUMEN
Administration of alpha-naphthylisothiocyanate (ANIT) to rats results in periportal cholangiolitic hepatopathy. Inflammation is a hallmark of the liver injury, and expression of toxicity is dependent on blood neutrophils. The role of other cellular mediators of inflammation in ANIT-induced hepatic insult is unknown. We hypothesized that platelets participate in the expression of ANIT hepatotoxicity. To test this, circulating platelets were decreased by administration of anti-rat platelet serum (PAb) prior to treatment of rats with ANIT. The PAb treatment regimen effectively reduced circulating thrombocytes over the course of the experiment. Twenty-four hours after oral ANIT administration, rats were euthanized and liver injury was estimated by increases in serum alanine aminotransferase (ALT) and gamma-glutamyltransferase (GGT) activities. Cholestasis was assessed by measurement of serum total bilirubin concentration and bile flow. Reduction in platelet numbers was associated with attenuation of the increases in plasma ALT activity and bilirubin concentration seen after ANIT administration. However, PAb treatment did not attenuate the increase in plasma GGT, a marker of biliary epithelial cell injury. ANIT-induced changes in platelet function were assessed by evaluating platelet aggregation responses in platelet-rich plasma from rats treated with ANIT in vivo. ANIT treatment modestly decreased ex vivo platelet aggregation in response to ADP and collagen stimuli. To address further the role of platelet-derived cyclooxygenase products in ANIT hepatotoxicity, rats were treated with aspirin or ibuprofen. Neither pretreatment ameliorated ANIT-induced hepatic insult. These results suggest that platelets contribute to the expression of ANIT-induced liver injury, but they do not appear to act through the production of cyclooxygenase metabolites.
Asunto(s)
1-Naftilisotiocianato/toxicidad , Plaquetas/fisiología , Hígado/efectos de los fármacos , Hígado/patología , Análisis de Varianza , Animales , Aspirina/farmacología , Plaquetas/efectos de los fármacos , Ibuprofeno/farmacología , Sueros Inmunes/farmacología , Hígado/enzimología , Masculino , Agregación Plaquetaria/efectos de los fármacos , Recuento de Plaquetas , Ratas , Ratas Sprague-DawleyRESUMEN
BACKGROUND: Activated neutrophils (PMNs) release cytotoxic agents that can damage surrounding tissue. These studies were performed to determine whether activated PMNs from rat could injure isolated, rat hepatic parenchymal cells (HCs) in vitro. EXPERIMENTAL DESIGN: HCs were cocultured with unstimulated rat PMNs or with PMNs activated with either f-met-leu-phe (FMLP) or phorbol myristate acetate (PMA), that stimulate predominantly degranulation or superoxide production, respectively. Toxicity to HCs was evaluated from release of alanine aminotransferase into the medium. RESULTS: Alanine aminotransferase release was greater in HCs cocultured with FMLP- or PMA-stimulated PMNs compared with unstimulated PMNs. Toxicity was observed by 16 hours after stimulation of PMNs. To test the possible involvement of a soluble mediator released by activated PMNs, HCs were incubated with conditioned medium from PMNs. Compared with unstimulated PMNs, toxicity to HCs was greater in the presence of conditioned medium from FMLP-stimulated PMNs, but not conditioned medium from PMA-activated PMNs. Reactive oxygen species do not appear to be involved in the mechanism by which activated PMNs damage HCs since superoxide dismutase, catalase, superoxide dismutase+catalase, or desferrioxamine failed to prevent the injury. Furthermore, less superoxide anion was detected in PMA-stimulated PMNs when either HCs or HC-conditioned medium was present. Proteolytic enzymes released by stimulated PMNs may play a role in HC damage since an inhibitor of proteases diminished injury due to PMNs activated by either FMLP or PMA. CONCLUSIONS: These results indicate that activated, rat PMNs damage HCs in culture. The data suggest that reactive oxygen species are not involved in the mechanism, but that release of proteolytic enzymes may play a role in the toxic response.
Asunto(s)
Hígado/patología , Hígado/fisiología , Neutrófilos/patología , Neutrófilos/fisiología , Alanina Transaminasa/metabolismo , Animales , Catalasa/farmacología , Células Cultivadas , Medios de Cultivo Condicionados/farmacología , Deferoxamina/farmacología , Hígado/metabolismo , Masculino , N-Formilmetionina Leucil-Fenilalanina/farmacología , Neutrófilos/metabolismo , Ratas , Ratas Sprague-Dawley , Superóxido Dismutasa/farmacología , Superóxidos/metabolismo , Acetato de Tetradecanoilforbol/farmacología , Factores de TiempoRESUMEN
Methylene dianiline (DDM) causes a dose- and time-dependent cholestasis, bile ductular epithelial injury, and hepatic parenchymal insult in rats. The mechanism of toxicity is unknown. Since hepatic leukocyte infiltration is a prominent feature of DDM-induced liver injury, and because leukocytes play a causal role in hepatic and extrahepatic tissue injury, we tested the hypothesis that toxicity caused by DDM is dependent on neutrophils or other circulating inflammatory cells. A polyclonal antibody (NAb) against rat neutrophils was used to address the role of the neutrophil in DDM-induced liver injury. NAb administration caused a significant reduction in circulating neutrophils without altering other leukocyte numbers. Moreover, NAb pretreatment prevented hepatic neutrophil infiltration after administration of DDM. However, neutrophil depletion did not afford protection from DDM-induced liver injury. This result was confirmed and the role of other circulating leukocytes was evaluated by inducing systemic leukopenia using cyclophosphamide (CYCLO). Administration of CYCLO diminished the number of circulating leukocytes within 4 days after treatment. Depletion of leukocytes by CYCLO prevented the hepatic accumulation of leukocytes but did not protect rats from DDM hepatotoxicity. These results suggest that the large numbers of leukocytes that infiltrate the liver after DDM administration do not contribute to hepatic injury.
Asunto(s)
Compuestos de Anilina/toxicidad , Carcinógenos/toxicidad , Enfermedad Hepática Inducida por Sustancias y Drogas , Hepatopatías/inmunología , Neutrófilos/efectos de los fármacos , Animales , Ciclofosfamida/farmacología , Leucopenia , Masculino , Neutropenia , Neutrófilos/fisiología , Ratas , Ratas Sprague-DawleyRESUMEN
Methylene dianiline (DDM) is a chemical intermediate in the production of isocyanates and other industrial chemicals, and it is hepatotoxic in humans. The acute hepatotoxicity of orally administered DDM was characterized in rats. Rats receiving DDM (25-225 mg/kg, per os) demonstrated a dose-dependent elevation in serum alanine aminotransferase activity, g-glutamyltransferase activity, and serum bilirubin concentration. DDM also caused a decrease in bile flow and an elevation in liver weight. Significant changes in these markers of liver injury occurred between 8 and 12 hr after a single, oral administration of DDM. Histologically, DDM caused multifocal, necrotizing hepatitis with neutrophil infiltration. Changes in the portal regions consisted of bile ductular necrosis, portal edema, neutrophil infiltration, mild fibrin exudation, and segmental necrotizing vasculitis. The role of cytochrome P450 monooxygenase (MO)-mediated metabolism in DDM hepatotoxicity was evaluated using the MO inhibitors, aminobenzotriazole and SKF-525A and the MO inducers phenobarbital and beta-naphthoflavone. Aminobenzotriazole provided protection from DDM-induced hepatotoxicity, whereas SKF-525A had no effect. The effect of phenobarbital pretreatment depended on the dose of DDM administered. At a dose of DDM that produced a maximal hepatotoxic response, phenobarbital did not influence hepatotoxicity. However, phenobarbital pretreatment provided protection against the hepatotoxic effects of a lower dose of DDM. beta-naphthoflavone pretreatment had a more modest effect on DDM-induced hepatic insult. These results demonstrate that DDM causes acute hepatotoxicity in the rat that is dose and time dependent. Results using inducers and inhibitors of MO suggest that DDM requires bioactivation to exert toxicity; however, the relationship between metabolism and toxicity may be complex.
Asunto(s)
Compuestos de Anilina/toxicidad , Hígado/efectos de los fármacos , Administración Oral , Animales , Relación Dosis-Respuesta a Droga , Hígado/patología , Masculino , Ratas , Ratas Sprague-DawleyRESUMEN
Diuretics are important drugs in the treatment of fluid overload and other related conditions in infants and children. Their rational use depends on adequate knowledge of how they work and how they interact with other drugs and the changing physiology caused by normal growth and development and by various disease states. It is in the best interest of the patient for the physician prescribing these drugs to have appropriate knowledge of their mode of action. It is clear that with the exception of a few drugs the amount of appropriate data available on the use of these drugs in children is limited at best. As newer drugs become available, the physician caring for such children must be sure that there is some clear indication for its use in preference to those with which we have more experience.