RESUMEN
The recently described 'gasomediator' hydrogen sulfide (H2S) has been involved in pain mechanisms, but its effect on pruritus, a sensory modality that similarly to pain acts as a protective mechanism, is poorly known and controversial. The effects of the slow-releasing (GYY4137) and spontaneous H2S donors (Na2S and Lawesson's reagent, LR) were evaluated in histamine and compound 48/80 (C48/80)-dependent dorsal skin pruritus and inflammation in male BALB/c mice. Animals were intradermally (i.d.) injected with C48/80 (3µg/site) or histamine (1µmol/site) alone or co-injected with Na2S, LR or GYY4137 (within the 0.3-100nmol range). The involvement of endogenous H2S and KATP channel-dependent mechanism were also evaluated. Pruritus was assessed by the number of scratching bouts, whilst skin inflammation was evaluated by the extravascular accumulation of intravenously injected 125I-albumin (plasma extravasation) and myeloperoxidase (MPO) activity (neutrophil recruitment). Histamine or C48/80 significantly evoked itching behavior paralleled by plasma extravasation and increased MPO activity. Na2S and LR significantly ameliorated histamine or C48/80-induced pruritus and inflammation, although these effects were less pronounced or absent with GYY4137. Inhibition of endogenous H2S synthesis increased both Tyrode and C48/80-induced responses in the skin, whereas the blockade of KATP channels by glibenclamide did not. H2S-releasing donors significantly attenuate C48/80-induced mast cell degranulation either in vivo or in vitro. We provide first evidences that H2S donors confer protective effect against histamine-mediated acute pruritus and cutaneous inflammation. These effects can be mediated, at least in part, by stabilizing mast cells, known to contain multiple mediators and to be primary initiators of allergic processes, thus making of H2S donors a potential alternative/complementary therapy for treating inflammatory allergic skin diseases and related pruritus.
Asunto(s)
Sulfuro de Hidrógeno/metabolismo , Sulfuro de Hidrógeno/farmacología , Inflamación/tratamiento farmacológico , Mastocitos/efectos de los fármacos , Sustancias Protectoras/farmacología , Prurito/tratamiento farmacológico , Piel/efectos de los fármacos , Animales , Gliburida/farmacología , Histamina/metabolismo , Inflamación/metabolismo , Canales KATP/metabolismo , Masculino , Mastocitos/metabolismo , Ratones , Ratones Endogámicos BALB C , Morfolinas/farmacología , Compuestos Organotiofosforados/farmacología , Prurito/metabolismo , Piel/metabolismoRESUMEN
Hydrogen sulfide (H2S) has been highlighted as an endogenous signaling molecule and we have previously found that it can inhibit histamine-mediated itching. Pruritus is the most common symptom of cutaneous diseases and anti-histamines are the usual treatment; however, anti-histamine-resistant pruritus is common in some clinical settings. In this way, the involvement of mediators other than histamine in the context of pruritus requires new therapeutic targets. Considering that the activation of proteinase-activated receptor 2 (PAR-2) is involved in pruritus both in rodents and humans, in this study we investigated the effect of H2S donors on the acute scratching behavior mediated by PAR-2 activation in mice, as well as some of the possible pharmacological mechanisms involved. The intradermal injection of the PAR-2 peptide agonist SLIGRL-NH2 (8-80nmol) caused a dose-dependent scratching that was unaffected by intraperitoneal pre-treatment with the histamine H1 antagonist pyrilamine (30mg/kg). Co-injection of SLIGRL-NH2 (40nmol) with either the slow-release H2S donor GYY4137 (1 and 3nmol) or the spontaneous donor NaHS (1 and 0.3nmol) significantly reduced pruritus. Co-treatment with the KATP channel blocker glibenclamide (200nmol) or the nitric oxide (NO) donor sodium nitroprusside (10nmol) abolished the antipruritic effects of NaHS; however, the specific soluble guanylyl cyclase inhibitor ODQ (30µg) had no significant effects. The transient receptor potential ankyrin type 1 (TRPA1) antagonist HC-030031 (20µg) significantly reduced SLIGRL-NH2-induced pruritus; however pruritus induced by the TRPA1 agonist AITC (1000nmol) was unaffected by NaHS. Based on these data, we conclude that pruritus secondary to PAR-2 activation can be reduced by H2S, which acts through KATP channel opening and involves NO in a cyclic guanosine monophosphate (cGMP)-independent manner. Furthermore, TRPA1 receptors mediate the pruritus induced by activation of PAR-2, but H2S does not interfere with this pathway. These results provide additional support for the development of new therapeutical alternatives, mainly intended for treatment of pruritus in patients unresponsive to anti-histamines.
Asunto(s)
Sulfuro de Hidrógeno/farmacología , Oligopéptidos/farmacología , Prurito/tratamiento farmacológico , Receptor PAR-2/metabolismo , Animales , Modelos Animales de Enfermedad , Gliburida/farmacocinética , Histamina/metabolismo , Antagonistas de los Receptores Histamínicos/farmacología , Isotiocianatos/farmacología , Masculino , Ratones , Ratones Endogámicos C57BL , Morfolinas/farmacología , Nitroprusiato/farmacología , Compuestos Organotiofosforados/farmacología , Prurito/metabolismo , Canales de Potencial de Receptor Transitorio/metabolismoRESUMEN
Leishmania spp are the causative agents of a spectrum of diseases termed leishmaniasis that affect mammals, including humans and dogs. Although reactive nitrogen species are employed in the control of parasitism by the immune system, it is known that Leishmania can withstand this oxidative stress. As the mechanism by which these species are resistant to nitric oxide (NO) is poorly understood, the main objective of this study was to evaluate the expression of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) in Leishmania amazonensis and Leishmania chagasi promastigotes showing natural resistance to NO. GAPDH transcript levels were quantified by real-time polymerase chain reaction amplification, and GAPDH activity (assessed by levels of NADH oxidation) was measured by spectrophotometry. The level of nitration in total protein was assessed by immunoblotting. The results demonstrated an increase in GAPDH expression in resistant isolates of both species compared to susceptible isolates. The increase in GAPDH expression led to an increase in the activity of GAPDH in L. amazonensis human isolates resistant to NO. The pattern of protein nitration did not differ between sensitive and resistant isolates. Our results suggest that changes in expression of GAPDH may be responsible, at least in part, to natural resistance to NO found in human and canine Leishmania spp.
Asunto(s)
Expresión Génica/efectos de los fármacos , Gliceraldehído 3-Fosfato Deshidrogenasa (NADP+)/genética , Leishmania infantum/genética , Leishmania/genética , Estadios del Ciclo de Vida/efectos de los fármacos , Óxido Nítrico/farmacología , Proteínas Protozoarias/genética , Medios de Cultivo , Resistencia a Medicamentos , Gliceraldehído 3-Fosfato Deshidrogenasa (NADP+)/metabolismo , Leishmania/efectos de los fármacos , Leishmania/enzimología , Leishmania/crecimiento & desarrollo , Leishmania infantum/efectos de los fármacos , Leishmania infantum/enzimología , Leishmania infantum/crecimiento & desarrollo , NAD/metabolismo , Óxido Nítrico/metabolismo , Oxidación-Reducción , Proteínas Protozoarias/metabolismo , Nitrito de Sodio/química , Nitrito de Sodio/farmacologíaRESUMEN
BACKGROUND AND OBJECTIVE: After activation, platelets express mediators that modulate inflammation. We hypothesized that drug-induced platelet inactivation may interfere in the inflammatory process in experimental periodontal disease by suppressing the release of biological mediators to the injury site. MATERIAL AND METHODS: To evaluate the effects of antiplatelet drugs on experimental periodontal disease, 60 rats were randomly assigned to six groups (n = 10) and ligatures were placed around lower first molars in three groups. The other three groups were not subjected to the induction of periodontal disease and were used as negative controls. During the experimental period, animals were given aspirin (30 mg/kg) or clopidogrel (75 mg/kg) intragastrically once daily for 3 d. On day 3, they were killed and gingival tissue were used to evaluate myeloperoxidase activity and the expression of the chemokine CXCL4. Hemi-mandibles were used for microscopic evaluation. RESULTS: Clopidogrel significantly reduced the inflammatory infiltrate and increased the amount of collagen fibers. Histometric analysis showed that clopidogrel impaired alveolar bone loss. Expression of CXCL4 was significantly increased (p < 0.001) in rats subjected to periodontal disease. Systemic administration of aspirin and clopidogrel induced a significant decrease ( p < 0.05) in the expression of CXCL4. Treatment with antiplatelet drugs resulted in a significant reduction of myeloperoxidase activity when compared to saline-treated animals with periodontal disease. CONCLUSION: Clopidogrel but not aspirin showed the ability of preventing bone loss in experimental periodontitis.
Asunto(s)
Antiinflamatorios/uso terapéutico , Periodontitis/tratamiento farmacológico , Inhibidores de Agregación Plaquetaria/uso terapéutico , Pérdida de Hueso Alveolar/prevención & control , Animales , Aspirina/uso terapéutico , Clopidogrel , Colágeno/efectos de los fármacos , Tejido Conectivo/efectos de los fármacos , Tejido Conectivo/patología , Modelos Animales de Enfermedad , Encía/efectos de los fármacos , Encía/patología , Mediadores de Inflamación/antagonistas & inhibidores , Masculino , Enfermedades Mandibulares/prevención & control , Periodontitis/inmunología , Periodontitis/patología , Peroxidasa/análisis , Factor Plaquetario 4/análisis , Distribución Aleatoria , Ratas , Ratas Sprague-Dawley , Ticlopidina/análogos & derivados , Ticlopidina/uso terapéuticoRESUMEN
BACKGROUND: Nitric oxide is a key signalling molecule in the pathogenesis of inflammation, but its role in acute pancreatitis and related abdominal pain induced by secretory phospholipase A2 (sPLA2 ) from Crotalus durissus terrificus (Cdt) venom has not been investigated. METHODS: Male Wistar rats were i.v. injected with L-NAME (20 mg/kg), aminoguanidine (AG, 50 mg/kg), 7-nitroindazole (7-NI, 10 mg/kg) or vehicle 10 min before or 60 min after the injection of sPLA2 (300 µg/kg) into the common bile duct. After 4 h of sPLA2 injection, abdominal hyperalgesia and inflammation were assessed in addition to serum amylase, nitrite/nitrate (NOx), pancreas lipoperoxidation and 3-nitrotyrosine (3-NT) contents. RESULTS: sPLA2 -induced acute pancreatitis, related abdominal hyperalgesia, hyperamylasemia and increased concentration of NOx were not correlated with lipoperoxidation or increased 3-NT in the pancreas. Pretreatment with all the nitric oxide synthase (NOS) inhibitors significantly reduced abdominal mechanical hyperalgesia, but only iNOS blockade by AG suppressed pancreas oedema and serum NOx increase. The therapeutic approach with all the NOS inhibitors produced a similar reduction pattern of the abdominal hyperalgesia, but AG treatment also inhibited serum hyperamylasemia and NOx concentrations and pancreatic myeloperoxidase. The nNOS blockade by 7-NI treatment also inhibited myeloperoxidase activity in both pancreas and lung. CONCLUSIONS: Therapeutic blockade of iNOS or nNOS provides benefits in terms of inhibition of the acute pancreatitis-related abdominal hyperalgesia, while iNOS inhibition also ameliorates the inflammatory cell influx to the pancreas and reduces the resultant hyperamylasemia and NOx levels, thus representing alternative pharmacological strategies for treatment of clinical pancreatitis associated with increased PLA2 .
Asunto(s)
Inhibidores Enzimáticos/uso terapéutico , NG-Nitroarginina Metil Éster/uso terapéutico , Óxido Nítrico Sintasa de Tipo II/antagonistas & inhibidores , Dolor/tratamiento farmacológico , Dolor/etiología , Pancreatitis/complicaciones , Pancreatitis/tratamiento farmacológico , Fosfolipasas A2 Secretoras , Animales , Hiperalgesia/inducido químicamente , Hiperalgesia/tratamiento farmacológico , Masculino , Páncreas/enzimología , Páncreas/patología , Pancreatitis/enzimología , Peroxidasa/metabolismo , Ratas , Ratas Wistar , Sustancias Reactivas al Ácido Tiobarbitúrico/metabolismo , Tirosina/análogos & derivados , Tirosina/metabolismoRESUMEN
We have shown previously that nitric oxide (NO) controls platelet endothelial cell adhesion molecule (PECAM-1) expression on both neutrophils and endothelial cells under physiological conditions. Here, the molecular mechanism by which NO regulates lipopolysaccharide (LPS)-induced endothelial PECAM-1 expression and the role of interleukin (IL)-10 on this control was investigated. For this purpose, N-(G)-nitro-L-arginine methyl ester (L-NAME; 20 mg/kg/day for 14 days dissolved in drinking water) was used to inhibit both constitutive (cNOS) and inducible nitric oxide (iNOS) synthase activities in LPS-stimulated Wistar rats (5 mg/kg, intraperitoneally). This treatment resulted in reduced levels of serum NO. Under this condition, circulating levels of IL-10 was enhanced, secreted mainly by circulating lymphocytes, dependent on transcriptional activation, and endothelial PECAM-1 expression was reduced independently on reduced gene synthesis. The connection between NO, IL-10 and PECAM-1 expression was examined by incubating LPS-stimulated (1 µg/ml) cultured endothelial cells obtained from naive rats with supernatant of LPS-stimulated lymphocytes, which were obtained from blood of control or L-NAME-treated rats. Supernatant of LPS-stimulated lymphocytes obtained from L-NAME-treated rats, which contained higher levels of IL-10, reduced LPS-induced PECAM-1 expression by endothelial cells, and this reduction was reversed by adding the anti-IL-10 monoclonal antibody. Therefore, an association between NO, IL-10 and PECAM-1 was found and may represent a novel mechanism by which NO controls endothelial cell functions.
Asunto(s)
Inflamación/metabolismo , Interleucina-10/metabolismo , Lipopolisacáridos/inmunología , Linfocitos/metabolismo , Óxido Nítrico/metabolismo , Molécula-1 de Adhesión Celular Endotelial de Plaqueta/genética , Molécula-1 de Adhesión Celular Endotelial de Plaqueta/metabolismo , Animales , Anticuerpos Monoclonales , Células Cultivadas , Células Endoteliales/metabolismo , Inhibidores Enzimáticos/administración & dosificación , Inhibidores Enzimáticos/farmacología , Inflamación/inmunología , Interleucina-10/sangre , Interleucina-10/inmunología , NG-Nitroarginina Metil Éster/administración & dosificación , NG-Nitroarginina Metil Éster/farmacología , Óxido Nítrico/sangre , Óxido Nítrico Sintasa de Tipo II/antagonistas & inhibidores , Óxido Nítrico Sintasa de Tipo II/metabolismo , Óxido Nítrico Sintasa de Tipo III/antagonistas & inhibidores , Óxido Nítrico Sintasa de Tipo III/metabolismo , Distribución Aleatoria , Ratas , Ratas Wistar , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
The proteinase-activated receptor 2 (PAR(2)) is a putative therapeutic target for arthritis. We hypothesized that the early pro-inflammatory effects secondary to its activation in the temporomandibular joint (TMJ) are mediated by neurogenic mechanisms. Immunofluorescence analysis revealed a high degree of neurons expressing PAR(2) in retrogradely labeled trigeminal ganglion neurons. Furthermore, PAR(2) immunoreactivity was observed in the lining layer of the TMJ, co-localizing with the neuronal marker PGP9.5 and substance-P-containing peripheral sensory nerve fibers. The intra-articular injection of PAR(2) agonists into the TMJ triggered a dose-dependent increase in plasma extravasation, neutrophil influx, and induction of mechanical allodynia. The pharmacological blockade of natural killer 1 (NK(1)) receptors abolished PAR(2)-induced plasma extravasation and inhibited neutrophil influx and mechanical allodynia. We conclude that PAR(2) activation is pro-inflammatory in the TMJ, through a neurogenic mechanism involving NK(1) receptors. This suggests that PAR(2) is an important component of innate neuro-immune response in the rat TMJ.
Asunto(s)
Artritis/patología , Receptor PAR-2/análisis , Trastornos de la Articulación Temporomandibular/patología , Animales , Artropatía Neurógena/patología , Inmunidad Innata/inmunología , Inyecciones Intraarticulares , Masculino , Fibras Nerviosas/patología , Neuroinmunomodulación/inmunología , Antagonistas del Receptor de Neuroquinina-1 , Neuronas/patología , Infiltración Neutrófila/efectos de los fármacos , Neutrófilos/patología , Oligopéptidos/administración & dosificación , Oligopéptidos/farmacología , Dimensión del Dolor , Piperidinas/farmacología , Plasma , Quinuclidinas/farmacología , Ratas , Ratas Wistar , Receptor PAR-2/agonistas , Células Receptoras Sensoriales/patología , Sustancia P/análisis , Articulación Temporomandibular/inervación , Ganglio del Trigémino/patología , Tripsina/administración & dosificación , Tripsina/farmacología , Ubiquitina Tiolesterasa/análisisRESUMEN
BACKGROUND AND PURPOSE: Recent findings suggest that the noxious gas H(2)S is produced endogenously, and that physiological concentrations of H(2)S are able to modulate pain and inflammation in rodents. This study was undertaken to evaluate the ability of endogenous and exogenous H(2)S to modulate carrageenan-induced synovitis in the rat knee. EXPERIMENTAL APPROACH: Synovitis was induced in Wistar rats by intra-articular injection of carrageenan into the knee joint. Sixty minutes prior to carrageenan injection, the rats were pretreated with indomethacin, an inhibitor of H(2)S formation (DL-propargylglycine) or an H(2)S donor [Lawesson's reagent (LR)]. KEY RESULTS: Injection of carrageenan evoked knee inflammation, pain as characterized by impaired gait, secondary tactile allodynia of the ipsilateral hindpaw, joint swelling, histological changes, inflammatory cell infiltration, increased synovial myeloperoxidase, protein nitrotyrosine residues, inducible NOS (iNOS) activity and NO production. Pretreatment with LR or indomethacin significantly attenuated the pain responses, and all the inflammatory and biochemical changes, except for the increased iNOS activity, NO production and 3-NT. Propargylglycine pretreatment potentiated synovial iNOS activity (and NO production), and enhanced macrophage infiltration, but had no effect on other inflammatory parameters. CONCLUSIONS AND IMPLICATIONS: Whereas exogenous H(2)S delivered to the knee joint can produce a significant anti-inflammatory and anti-nociceptive effect, locally produced H(2)S exerts little immunomodulatory effect. These data further support the development and use of H(2)S donors as potential alternatives (or complementary therapies) to the available anti-inflammatory compounds used for treatment of joint inflammation or relief of its symptoms.
Asunto(s)
Carragenina/efectos adversos , Sulfuro de Hidrógeno/farmacología , Articulación de la Rodilla/patología , Sinovitis/inducido químicamente , Animales , Articulación de la Rodilla/enzimología , Articulación de la Rodilla/metabolismo , Masculino , Óxido Nítrico/biosíntesis , Óxido Nítrico Sintasa de Tipo II/metabolismo , Peroxidasa/metabolismo , Ratas , Ratas Wistar , Sinovitis/enzimología , Sinovitis/metabolismo , Tirosina/análogos & derivados , Tirosina/metabolismoRESUMEN
BACKGROUND AND OBJECTIVE: Platelets contain factors, including VEGF and endostatin, that can modulate the healing process. We evaluated the effects of severe thrombocytopenia on periodontal healing in rats and determined the contribution of VEGF and endostatin to the healing process. MATERIAL AND METHODS: Rats were distributed into three test groups and two control groups. Cotton ligatures were placed at the gingival margin level of the lower first molar in the test groups. Sham-operated rats and rats in one of the periodontitis groups were killed 15 days later. Rats in the remaining two periodontitis groups had the ligatures removed in order to study the spontaneous recovery from the periodontal disease 15 days later, and these rats were treated with rabbit antiplatelet serum, in order to induce thrombocytopenia, or normal rabbit serum. An additional group without ligatures received antiplatet serum in the same period. RESULTS: After ligature removal, rats treated with normal rabbit serum showed reduced myeloperoxidase activity, decreased alveolar bone loss and increased numbers of blood vessels. Thrombocytopenia caused a delay in alveolar bone regeneration, a decrease in the number of vessels and a modest decrease in myeloperoxidase activity. In the rats with periodontitis, serum endostatin concentrations were slightly decreased and serum VEGF remained unchanged compared with sham-operated animals. After ligature removal, a significant VEGF increase and endostatin decrease were observed in the rats treated with normal rabbit serum. Thrombocytopenia led to a dramatic fall in both VEGF and endostatin concentrations. CONCLUSION: Thrombocytopenia leads to a delay of periodontal healing in the situation of experimental periodontitis, which might be mediated in part by a decrease in the serum concentration of VEGF and endostatin derived from the platelets. However, other factors derived from the platelets may also have contributed to a delay of periodontal healing in the rats with thrombocytopenia.
Asunto(s)
Inhibidores de la Angiogénesis/fisiología , Proteínas Angiogénicas/fisiología , Endostatinas/fisiología , Periodontitis/fisiopatología , Trombocitopenia/fisiopatología , Factor A de Crecimiento Endotelial Vascular/fisiología , Pérdida de Hueso Alveolar/patología , Pérdida de Hueso Alveolar/fisiopatología , Inhibidores de la Angiogénesis/sangre , Proteínas Angiogénicas/sangre , Animales , Plaquetas/inmunología , Plaquetas/fisiología , Regeneración Ósea/fisiología , Remodelación Ósea/fisiología , Endostatinas/sangre , Sueros Inmunes , Masculino , Neovascularización Fisiológica/fisiología , Periodontitis/sangre , Periodontitis/patología , Peroxidasa/análisis , Recuento de Plaquetas , Conejos , Ratas , Ratas Sprague-Dawley , Trombocitopenia/sangre , Factores de Tiempo , Factor A de Crecimiento Endotelial Vascular/sangre , Cicatrización de Heridas/fisiologíaRESUMEN
BACKGROUND AND OBJECTIVE: Cyclosporine A treatment is important in the therapy of a number of medical conditions; however, alveolar bone loss is an important negative side-effect of this drug. As such, we evaluated whether concomitant administration of simvastatin would minimize cyclosporine A-associated alveolar bone loss in rats subjected, or not, to experimental periodontal disease. MATERIAL AND METHODS: Groups of 10 rats each were treated with cyclosporine A (10 mg/kg/day), simvastatin (20 mg/kg/day), cyclosporine A and simvastatin concurrently (cyclosporine A/simvastatin) or vehicle for 30 days. Four other groups of 10 rats each received a cotton ligature around the lower first molar and were treated similarly with cyclosporine A, simvastatin, cyclosporine A/simvastatin or vehicle. Calcium (Ca(2+)), phosphorus and alkaline phosphatase levels were evaluated in serum. Expression levels of interleukin-1beta, prostaglandin E(2) and inducible nitric oxide synthase were evaluated in the gingivomucosal tissues. Bone volume and numbers of osteoblasts and osteoclasts were also analyzed. RESULTS: Treatment with cyclosporine A in rats, with or without ligature, was associated with bone loss, represented by a lower bone volume and an increase in the number of osteoclasts. Treatment with cyclosporine A was associated with bone resorption, whereas simvastatin treatment improved cyclosporine A-associated alveolar bone loss in all parameters studied. In addition, simvastatin, in the presence of inflammation, can act as an anti-inflammatory agent. CONCLUSION: This study shows that simvastatin therapy leads to a reversal of the cyclosporine A-induced bone loss, which may be mediated by downregulation of interleukin-1beta and prostaglandin E(2) production.
Asunto(s)
Pérdida de Hueso Alveolar/inducido químicamente , Ciclosporina/efectos adversos , Inhibidores de Hidroximetilglutaril-CoA Reductasas/farmacología , Inmunosupresores/efectos adversos , Simvastatina/farmacología , Fosfatasa Alcalina/sangre , Pérdida de Hueso Alveolar/fisiopatología , Proceso Alveolar/efectos de los fármacos , Proceso Alveolar/patología , Animales , Antiinflamatorios/farmacología , Densidad Ósea/efectos de los fármacos , Calcio/sangre , Recuento de Células , Dinoprostona/análisis , Regulación hacia Abajo , Encía/efectos de los fármacos , Encía/patología , Interleucina-1beta/análisis , Masculino , Mucosa Bucal/efectos de los fármacos , Mucosa Bucal/patología , Óxido Nítrico Sintasa de Tipo II/análisis , Osteoblastos/efectos de los fármacos , Osteoblastos/patología , Osteoclastos/efectos de los fármacos , Osteoclastos/patología , Fósforo/sangre , Distribución Aleatoria , Ratas , Ratas Sprague-DawleyRESUMEN
BACKGROUND AND PURPOSE: The inflammation-resolving lipid mediator resolvin E1 (RvE1) effectively stops inflammation-induced bone loss in vivo in experimental periodontitis. It was of interest to determine whether RvE1 has direct actions on osteoclast (OC) development and bone resorption. EXPERIMENTAL APPROACH: Primary OC cultures derived from mouse bone marrow were treated with RvE1 and analysed for OC differentiation, cell survival and bone substrate resorption. Receptor binding was measured using radiolabelled RvE1. Nuclear factor (NF)-kappaB activation and Akt phosphorylation were determined with western blotting. Lipid mediator production was assessed with liquid chromatography tandem mass spectrometry. KEY RESULTS: OC growth and resorption pit formation were markedly decreased in the presence of RvE1. OC differentiation was inhibited by RvE1 as demonstrated by decreased number of multinuclear OC, a delay in the time course of OC development and attenuation of receptor activator of NF-kappaB ligand-induced nuclear translocation of the p50 subunit of NF-kappaB. OC survival and apoptosis were not altered by RvE1. Messenger RNA for both receptors of RvE1, ChemR23 and BLT(1) is expressed in OC cultures. Leukotriene B(4) (LTB(4)) competed with [(3)H]RvE1 binding on OC cell membrane preparations, and the LTB(4) antagonist U75302 prevented RvE1 inhibition of OC growth, indicating that BLT(1) mediates RvE1 actions on OC. Primary OC synthesized the RvE1 precursor 18R-hydroxy-eicosapentaenoic acid and LTB(4). Co-incubation of OC with peripheral blood neutrophils resulted in transcellular RvE1 biosynthesis. CONCLUSIONS AND IMPLICATIONS: These results indicate that RvE1 inhibits OC growth and bone resorption by interfering with OC differentiation. The bone-sparing actions of RvE1 are in addition to inflammation resolution, a direct action in bone remodelling.
Asunto(s)
Resorción Ósea , Diferenciación Celular/fisiología , Ácido Eicosapentaenoico/análogos & derivados , Inflamación/prevención & control , Osteoclastos/citología , Animales , Apoptosis , Células Cultivadas , Dentina/metabolismo , Ácido Eicosapentaenoico/biosíntesis , Ácido Eicosapentaenoico/fisiología , Leucotrieno B4/biosíntesis , Ratones , Osteoclastos/metabolismo , Ensayo de Unión RadioliganteRESUMEN
BACKGROUND AND PURPOSE: Overactive bladder is a complex and widely prevalent condition, but little is known about its physiopathology. We have carried out morphological, biochemical and functional assays to investigate the effects of long-term nitric oxide (NO) deficiency on muscarinic receptor and beta-adrenoceptor modulation leading to overactivity of rat detrusor muscle. EXPERIMENTAL APPROACH: Male Wistar rats received N(omega)-nitro-L-arginine methyl ester (L-NAME) in drinking water for 7-30 days. Functional responses to muscarinic and beta-adrenoceptor agonists were measured in detrusor smooth muscle (DSM) strips in Krebs-Henseleit solution. Measurements of [(3)H]inositol phosphate, NO synthase (NOS) activity, [(3)H]quinuclidinyl benzilate ([(3)H]QNB) binding and bladder morphology were also performed. KEY RESULTS: Long-term L-NAME treatment significantly increased carbachol-induced DSM contractile responses after 15 and 30 days; relaxing responses to the beta(3)-adrenoceptor agonist BRL 37-344 were significantly reduced at 30 days. Constitutive NOS activity in bladder was reduced by 86% after 7 days and maintained up to 30 days of L-NAME treatment. Carbachol increased sixfold the [(3)H]inositol phosphate in bladder tissue from rats treated with L-NAME. [(3)H]QNB was bound with an apparent K(D) twofold higher in bladder membranes after L-NAME treatment compared with that in control. No morphological alterations in DSM were found. CONCLUSIONS AND IMPLICATIONS: Long-term NO deficiency increased rat DSM contractile responses to a muscarinic agonist, accompanied by significantly enhanced K(D) values for muscarinic receptors and [(3)H]inositol phosphate accumulation in bladder. This supersensitivity for muscarinic agonists along with reductions of beta(3)-adrenoceptor-mediated relaxations indicated that overactive DSM resulted from chronic NO deficiency.
Asunto(s)
Contracción Muscular/efectos de los fármacos , Óxido Nítrico/deficiencia , Receptores Adrenérgicos beta 3/metabolismo , Receptores Muscarínicos/metabolismo , Vejiga Urinaria Hiperactiva/fisiopatología , Agonistas de Receptores Adrenérgicos beta 3 , Agonistas Adrenérgicos beta/farmacología , Animales , Carbacol/farmacología , Fosfatos de Inositol/metabolismo , Masculino , Agonistas Muscarínicos/farmacología , Músculo Liso/metabolismo , NG-Nitroarginina Metil Éster , Óxido Nítrico Sintasa/metabolismo , Ratas , Ratas Wistar , Receptores Muscarínicos/efectos de los fármacos , Vejiga Urinaria/metabolismo , Vejiga Urinaria/fisiopatologíaRESUMEN
Numerous vasoactive agents play an important physiological role in regulating vascular tone, reactivity and structure. In pathological conditions, alterations in the regulation of vasoactive peptides result in endothelial dysfunction, vascular remodeling and vascular inflammation, which are important processes underlying vascular damage in cardiovascular disease. Among the many vasoactive agents implicated in vascular (patho)biology, angiotensin II (Ang II), endothelin (ET), serotonin and natriuretic peptides appear to be particularly important because of their many pleiotropic actions and because they have been identified as potential therapeutic targets in cardiovascular disease. Ang II, ET-1, serotonin and natriuretic peptides mediate effects via specific receptors, which belong to the group of G-protein-coupled receptors. ET, serotonin and Ang II are primarily vasoconstrictors with growth-promoting actions, whereas natriuretic peptides, specifically atrial, brain and C-type natriuretic peptides, are vasodilators with natriuretic effects. Inhibition of vasoconstrictor actions with drugs that block peptide receptors, compounds that inhibit enzymes that generate vasoactive peptides or agents that increase levels of natriuretic peptides are potentially valuable therapeutic tools in the management of cardiovascular diseases. This review focuses on ET, natriuretic peptides and serotonin. The properties and distribution of these vasoactive agents and their receptors, mechanisms of action and implications in cardiovascular (patho)physiology will be discussed.
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Enfermedades Cardiovasculares/fisiopatología , Endotelina-1/fisiología , Corazón/fisiología , Péptidos Natriuréticos/fisiología , Serotonina/fisiología , HumanosRESUMEN
We have previously reported that in comparison with normal rats, the presence of experimental allergic encephalomyelitis (EAE) leads to decreased endogenous inhibitory activity (EIA) of Ca2+-dependent nitric oxide synthase (NOS) in both brain and serum, and increased expression of protein 3-nitrotyrosine (NT) in brain. In this work we show that animals recovered from the clinical signs of EAE are not different from controls in terms of either brain NOS activity, EIA of NOS, or NT expression. These results suggest that parallel to the reversal of the disease symptoms, a normalization of the production of nitric oxide and related species occurs.
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Encéfalo/enzimología , Encefalomielitis Autoinmune Experimental/enzimología , Óxido Nítrico Sintasa de Tipo III/metabolismo , Óxido Nítrico/metabolismo , Animales , Dineínas Citoplasmáticas , Dineínas/metabolismo , Encefalomielitis Autoinmune Experimental/sangre , Masculino , Óxido Nítrico Sintasa de Tipo III/antagonistas & inhibidores , Ratas , Ratas Endogámicas LewRESUMEN
We have previously reported that in comparison with normal rats, the presence of experimental allergic encephalomyelitis (EAE) leads to decreased endogenous inhibitory activity (EIA) of Ca2+-dependent nitric oxide synthase (NOS) in both brain and serum, and increased expression of protein 3-nitrotyrosine (NT) in brain. In this work we show that animals recovered from the clinical signs of EAE are not different from controls in terms of either brain NOS activity, EIA of NOS, or NT expression. These results suggest that parallel to the reversal of the disease symptoms, a normalization of the production of nitric oxide and related species occurs.
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Animales , Masculino , Ratas , Encéfalo/enzimología , Encefalomielitis Autoinmune Experimental/enzimología , Óxido Nítrico Sintasa de Tipo III/metabolismo , Óxido Nítrico/metabolismo , Dineínas/metabolismo , Encefalomielitis Autoinmune Experimental/sangre , Óxido Nítrico Sintasa de Tipo III/antagonistas & inhibidores , Ratas Endogámicas LewRESUMEN
OBJECTIVE: The present study analyzed the effects of acute amphetamine (AMPH) treatment on immune-mediated lung inflammatory response in rats. METHODS: There were four experiments. In the first and second experiments, rats were treated with AMPH (1 mg/kg) or 0.9% NaCl, and locomotor activity (experiment 1) and serum AMPH concentrations (experiment 2) were measured 1 or 12 h after treatment. In the third experiment, rats which were immunized with ovalbumin (OVA) were treated 14 days later with 0.9% NaCl or AMPH (1 mg/kg). Twelve hours after these treatments, all animals were submitted to challenge by 1% OVA inhalation being analyzed afterwards for bronchoalveolar lavage fluid (BAL), peripheral blood and bone marrow cellularity. In the fourth and final experiment, rats were treated and studied as for experiment 3, except that half of the animals within each group were previously treated with metyrapone prior to the OVA challenge. RESULTS: In the non-immunized rats, AMPH treatment induced an increase in locomotor activity synchronized to high serum AMPH concentrations 1 h after, but not 12 h after treatment. In OVA-challenged rats, AMPH treatment decreased the total number of inflammatory cells, recovered in both BAL and peripheral blood and increased the total number of bone marrow cells. These effects, observed 1 day after OVA challenge, were abrogated by previous metyrapone treatment. CONCLUSION: AMPH treatment changed HPA-axis responsiveness to the stress condition imposed by the OVA challenge decreasing lung and blood leukocytes cellularity most probably via corticosterone actions on bone marrow activity.
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Anfetamina/farmacología , Quimiotaxis de Leucocito/efectos de los fármacos , Sistema Hipotálamo-Hipofisario/efectos de los fármacos , Mielopoyesis/efectos de los fármacos , Neuroinmunomodulación/efectos de los fármacos , Neumonía/inmunología , Anfetamina/sangre , Animales , Médula Ósea/efectos de los fármacos , Médula Ósea/inmunología , Líquido del Lavado Bronquioalveolar/citología , Líquido del Lavado Bronquioalveolar/inmunología , Quimiotaxis de Leucocito/inmunología , Corticosterona/metabolismo , Regulación hacia Abajo/efectos de los fármacos , Regulación hacia Abajo/inmunología , Inhibidores Enzimáticos/farmacología , Sistema Hipotálamo-Hipofisario/inmunología , Sistema Hipotálamo-Hipofisario/metabolismo , Mediadores de Inflamación/farmacología , Recuento de Leucocitos , Leucocitos/citología , Leucocitos/efectos de los fármacos , Leucocitos/inmunología , Masculino , Metirapona/farmacología , Actividad Motora/efectos de los fármacos , Actividad Motora/fisiología , Mielopoyesis/inmunología , Neuroinmunomodulación/inmunología , Ovalbúmina/farmacología , Neumonía/inducido químicamente , Neumonía/fisiopatología , Ratas , Ratas WistarRESUMEN
A method based on liquid chromatography with negative ion electrospray ionization and tandem mass spectrometry is described for the determination of nimesulide in human plasma. Liquid-liquid extraction using a mixture of diethyl ether and dichloromethane was employed and celecoxib was used as an internal standard. The chromatographic run time was 4.5 min and the weighted (1/x) calibration curve was linear in the range 10.0-2000 ng x ml(-1). The limit of quantification was 10 ng x ml(-1), the intra-batch precision was 6.3, 2.1 and 2.1% and the intra-batch accuracy was 3.2, 0.3 and 0.1% for 30, 300 and 1200 ng x ml(-1) respectively. The inter-batch precision was 2.3, 2.8 and 2.7% and the accuracy was 3.3, 0.3 and 0.1% for 30, 300 and 1200 ng x ml(-1) respectively. This method was employed in a bioequivalence study of one nimesulide drop formulation (nimesulide 50 mg x ml(-1) drop, Medley S/A Indústria Farmacêutica, Brazil) against one standard nimesulide drop formulation (Nisulid, 50 mg x ml(-1) drop, Astra Médica, Brazil). Twenty-four healthy volunteers (both sexes) took part in the study and received a single oral dose of nimesulide (100 mg, equivalent to 2 ml of either formulation) in an open, randomized, two-period crossover way, with a 2-week washout interval between periods. The 90% confidence interval (CI) for geometric mean ratios between nimesulide and Nisulid were 93.1-109.6% for C(max), 87.7-99.8% for AUC(last) and 88.1-99.7% for AUC(0-infinity). Since the 90% CI for the above-mentioned parameters were included in the 80-125% interval proposed by the US Food and Drug Administration, the two formulations were considered bioequivalent in terms of both rate and extent of absorption.
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Cromatografía Líquida de Alta Presión/métodos , Espectrometría de Masas/métodos , Sulfonamidas/sangre , Equivalencia Terapéutica , Adolescente , Adulto , Estabilidad de Medicamentos , Éter , Femenino , Humanos , Masculino , Cloruro de Metileno , Persona de Mediana Edad , Control de Calidad , Sensibilidad y Especificidad , Sulfonamidas/administración & dosificación , Sulfonamidas/farmacocinéticaRESUMEN
1. Nonsteroidal anti-inflammatory drugs have been reported to exacerbate hypertension and to interfere with the effectiveness of some anti-hypertensive therapies. In this study, we tested the effects of a gastric-sparing, nitric oxide-releasing derivative of aspirin (NCX-4016) on hypertension in rats. 2. Hypertension was induced by administering L-NAME in the drinking water (400 mg l(-1)). Groups of rats were treated daily with aspirin, NCX-4016 or vehicle. 3. NCX-4016 significantly reduced blood pressure relative to the aspirin-treated group over the 2-week period of treatment. Aspirin and, to a lesser extent, NCX-4016 suppressed whole blood thromboxane synthesis. 4. In anaesthetized rats, acute intravenous administration of NCX-4016 caused a significant fall in mean arterial pressure in hypertensive rats, but was devoid of such effects in normotensive controls. 5. In vitro, NCX-4016 relaxed phenylephrine-pre-contracted aortic rings obtained from both normotensive and hypertensive rats, and significantly reduced their responsiveness to the contractile effects of phenylephrine. 6. These results suggest that NCX-4016 reduces blood pressure in hypertensive rats, not simply through the direct vasodilatory actions of the nitric oxide released by this compound, but also through possible interference with the effects of endogenous pressor agents. These properties, added to its anti-thrombotic effects, suggest that NCX-4016 may be a safer alternative to aspirin for use by hypertensive patients.
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Antiinflamatorios no Esteroideos/farmacología , Aspirina/farmacología , Presión Sanguínea/efectos de los fármacos , Hipertensión/fisiopatología , Óxido Nítrico/metabolismo , Vasodilatación/efectos de los fármacos , Anestesia , Animales , Antiinflamatorios no Esteroideos/farmacocinética , Aorta/efectos de los fármacos , Aorta/metabolismo , Aorta/fisiología , Aorta/fisiopatología , Aspirina/análogos & derivados , Aspirina/farmacocinética , Estado de Conciencia , Relación Dosis-Respuesta a Droga , Técnicas In Vitro , Masculino , NG-Nitroarginina Metil Éster/farmacología , Nitratos/metabolismo , Nitritos/metabolismo , Nitroprusiato/farmacología , Fenilefrina/farmacología , Ratas , Ratas Wistar , Renina/sangre , Tromboxanos/metabolismoRESUMEN
Venoms from snakes of the genus Bothrops cause pronounced local effects in the victims. These alterations result not only from the direct toxic action of venom components, but also from the prominent inflammatory reaction associated with these envenomations. In this study we investigated the ability of Bothrops asper (BaV) and Bothrops jararaca (BjV) venoms to induce cellular influx and microbicidal functions in leukocytes. BaV and BjV (5 microg/animal) caused a long lasting infiltration of leukocytes (3-48 h) when injected into mouse peritoneal cavity. Both venoms increased phagocytosis and production of hydrogen peroxide (H2O2) by polymorphonuclear (PMN) and mononuclear (MN) peritoneal leukocytes. In addition, nitric oxide (NO) production by macrophages was also enhanced after the venom injections. This effect was inhibited by treating animals with L-NAME and aminoguanidine, thus suggesting the induction of iNOS synthesis by the venoms. Western blot analysis confirmed the expression of iNOS in macrophages. BaV and BjV injection led to increased levels of IFN-gamma at the site of inflammation. Since IFN-gamma is an effective inducer of iNOS expression, an indirect action of the venoms on iNOS expression can be proposed. A marked formation of nitrotyrosine-containing proteins was also observed in macrophage homogenates. Based on these results, we suggest that reactive oxygen and nitrogen-derived species are involved in the pathogenesis of the local tissue damage characteristic of Bothrops sp envenomations.
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Bothrops/fisiología , Quimiotaxis de Leucocito/efectos de los fármacos , Venenos de Crotálidos/toxicidad , Guanidinas/farmacología , Interferón gamma/farmacología , Macrófagos Peritoneales/efectos de los fármacos , NG-Nitroarginina Metil Éster/farmacología , Óxido Nítrico/biosíntesis , Fagocitosis/efectos de los fármacos , Animales , Anticoagulantes/farmacología , Antineoplásicos/farmacología , Western Blotting , Quimiotaxis de Leucocito/inmunología , Venenos de Crotálidos/administración & dosificación , Venenos de Crotálidos/inmunología , Inhibidores Enzimáticos/farmacología , Heparina/farmacología , Peróxido de Hidrógeno/análisis , Técnicas para Inmunoenzimas , Inyecciones Intraperitoneales , Masculino , Ratones , Óxido Nítrico/sangre , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa/antagonistas & inhibidores , Oxidantes/farmacología , Fagocitosis/inmunología , Serpientes , Factores de TiempoRESUMEN
The introduction of selective inhibitors of cyclo-oxygenase-2 to the marketplace has been much anticipated for several years. It would appear that these compounds have lived up to the expectations of having reduced gastrointestinal toxicity and, at least for some indications, of efficacy similar to that of conventional non-steroidal anti-inflammatory drugs. However, there is a growing body of evidence suggesting that cyclo-oxygenase-2 plays a very important role in gastrointestinal mucosal defence, particularly in situations in which the mucosa is damaged or inflamed. Moreover, physiological roles for cyclo-oxygenase-2 both in the renal and cardiovascular systems are becoming better recognized. Inhibition of cyclo-oxygenase-2 can lead to peripheral oedema and hypertension, and may promote thrombosis. Indeed, there is recent evidence of increased rates of myocardial infarction in arthritis patients taking a selective cyclo-oxygenase-2 inhibitor. Use of low-dose aspirin concurrently with use of a selective cyclo-oxygenase-2 inhibitor may provide some degree of protection against the potential cardiovascular toxicity of the latter but both laboratory and clinical studies suggest that the concomitant use of these two types of drugs results in gastrointestinal ulceration comparable to what is seen with conventional non-steroidal anti-inflammatory drugs. These recent results suggest that care must be exercised in the use of selective cyclo-oxygenase-2 inhibitors by individuals who are at increased risk of myocardial infarction and stroke, and the use of low-dose aspirin by these patients may place them at increased risk of gastrointestinal complications.