RESUMO
Pyrogallol is a polyphenol that generates the superoxide anion. In this study, we investigated the influence of pyrogallol on human platelets. Our data showed that exposure of platelets to pyrogallol induced numerous manifestations of apoptosis including depolarization of mitochondrial inner membrane and release of cytochrome c from the mitochondria. Pyrogallol also induced downstream extra-mitochondrial apoptotic responses, including activation of caspase-3 and phosphatidylserine exposure on the outer leaflet of the plasma membrane. Addition of glutathione significantly rescued cells from pyrogallol- induced apoptosis, as evidenced by a decrease of all markers of apoptosis. Thus, pyrogallol appears to produce depletion of intracellular glutathione content in platelets, the main non-protein antioxidant in the cells. Furthermore, inhibition of γ-glutamyl transpeptidase, an enzyme that plays the main role in the cellular supply of glutathione, reverted the glutathione (GSH) protection over platelet apoptosis. Our results indicate that pyrogallol induces apoptosis by suppressing the natural anti-oxidation in human platelets.
Assuntos
Apoptose/efeitos dos fármacos , Plaquetas/citologia , Pirogalol/farmacologia , Biomarcadores/metabolismo , Plaquetas/efeitos dos fármacos , Plaquetas/metabolismo , Boratos/farmacologia , Citocromos c/metabolismo , Glutationa/farmacologia , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Serina/farmacologiaRESUMO
OBJECTIVE AND DESIGN: Carrageenan-induced paw edema has been described as a local and acute inflammatory process. In fact, little is known about the time course and systemic changes following a carrageenan injection. In this study, we examine the systemic changes that follow carrageenan injection in the paw. METHODS: Acute inflammation was produced by subplantar injection of carrageenan in a hind paw of Sprague-Dawley rats. Saline was used in control rats. Paw volume was measured with a plethysmometer. The hot plate latency test was used to quantify antinociception. C-reactive protein (CRP) levels were measured with a sandwich enzyme immunoassay. Fibrinogen concentration was measured using the gravimetric method. Lung morphometric analysis was performed using an image processing package. Lungs and paws were also examined for tissue factor (TF) and proinflammatory cytokines expression by immunohistochemistry. RESULTS: We found diverse systemic changes including increased levels of acute phase proteins, such as CRP and fibrinogen, and a lung inflammatory process characterized by lung edema, fibrin deposition, and leukocyte infiltration. An elevated expression of TF, IL-6, IL-1ß, and TNFα, was observed in paw and lung tissue sections by immunohistochemical methods. CONCLUSION: This study provides new evidence that a local carrageenan injection induces a systemic response.
Assuntos
Carragenina , Inflamação/patologia , Animais , Proteína C-Reativa/metabolismo , Citocinas/metabolismo , Edema/induzido quimicamente , Edema/patologia , Fibrinogênio/metabolismo , Pé/patologia , Inflamação/induzido quimicamente , Pulmão/patologia , Medição da Dor/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Tromboplastina/metabolismoRESUMO
The role of calcium and its relevance have been deeply revised with respect to trypanosomatids, as the mechanism by which calcium enters trypanosomes was, until now, not well understood. There is evidence supporting the presence of a nAChR in another member of the trypanosomatidae family, Trypanosoma cruzi, these receptors being one entry path to calcium ions. The aims of this work were to determine if there was a nicotinic acetylcholine receptor (nAChR) in Trypanosoma evansi, and to subsequently perform a partial pharmacological characterization of this receptor. After being loaded with FURA-2AM, individual cells of T. evansi, were exposed to cholinergic compounds, and the cells displayed a dose-dependent response to carbachol. This observation indicated that a cholinergic receptor may be present in T. evansi. Although a dose-dependent response to muscarine could not be demonstrated, nicotine could promote an incremental dose-dependent response. The relative potency of this specific agonist of nAChR is in agreement with previous reports. The estimated affinity values were a Kd1 value of 29.6+/-5.72 nM and a Kd2 value of 315.9+/-26.6 nM, which is similar to the Kd value reported for the alpha4 nicotinic receptor. The Hill coefficients were determined to be an n1 of 1.2+/-0.3 and an n2 of 4.2+/-1.3. Finally, our calculations indicated that there are about 1020 receptors in each T. evansi parasite, which is approximately 15-fold lower than the number reported in Torpedo californica electric cells. These results suggest the presence of a nAChR in T. evansi, which is able to bind nicotinic ligands and induce calcium signals.
Assuntos
Cálcio/metabolismo , Nicotina/farmacologia , Agonistas Nicotínicos/farmacologia , Receptores Nicotínicos/metabolismo , Trypanosoma/metabolismo , Animais , Bungarotoxinas/metabolismo , Relação Dose-Resposta a Droga , Masculino , Muscarina/farmacologia , Agonistas Muscarínicos/farmacologia , Antagonistas Nicotínicos/farmacologia , Distribuição Normal , Ratos , Ratos Sprague-Dawley , Receptores Nicotínicos/efeitos dos fármacos , Trypanosoma/efeitos dos fármacosRESUMO
En el músculo traqueal de bovino (MLTB) se ha propuesto una vía de señalización celular asociada con la actividad de una guanililciclasa de membrana plasmática (GCm) la cual se encuentra regulada por dos subtipos de receptores muscarínicos (mAChRs) m2 y m3 acoplados a proteínas G de manera opuesta. El objetivo del presemte estudio fue la identificación de las secuencias primarias de las regiones intracelulares de los receptores muscarínicos implicadas en la interacción con las proteínas G que regulan a esta GCm. El cDNA del MLTB fue usado para la amplificación de las regiones intracelulares de los subtipos m2 y m3 utilizando oligonucleótidos cebadores específicos. Los productos de amplificación fueron identificados en geles de agarosa y posteriormente secuenciados, encontrándose un 99 por ciento de similitud con las secuencias obtenidas del bGen Data Bank de los mACHRs en cerebro de bovino. Este es el primer clonamiento molecular para los mACHRs presentes en MLTB y sugiere posibles similitudes con el cerebro de bovino en relación a la interacción receptor/proteína G
Assuntos
Animais , Bovinos , Clonagem Molecular , Músculo Liso , Receptores Muscarínicos , Patologia , Farmacologia Clínica , VenezuelaRESUMO
Los subtipos M3 y M2 del mAchR fueron localizados en fracciones de membranas plasmáticas de músculos liso traqueal de bovino (MLTB). Los mAchRs nativos de MLTB fueron estudiados usando enlazamientos de [3H]QNB. La hetereogeneidad del receptor fue expresada para agonista y antagonista muscarínicos siendo los estados de alta afinidad sensibles a GTPyS. Mostramos que los efectos de GTPgS parecen ser mediados por proteínas Gi/o sensibles a PTX. Se detectó la hetereogeneidad del mAchR hacia 4-DAMP (M3) y pirenzepina (M1). Sin embargo, los antagonistas subtipo M2 como methoctramina y AF-DX 116 no mostraron tales respuestas en la presencia o ausencia de GTPyS. A través de la alquilación con 4-DAMP del subtipo M3 del mAchR, prevaleció el subtipo M2 el cual exhibió un enlazamiento dependiente de GTPyS parecido a un agonista mientras el enlazamiento de los antagonistas fue insensibles al GTPgS. Por lo anterior, la heterogeneidad a antagonistas del receptor descrita aquí es debida al subtipo M3 del mAchR, el cual muestra dos estados de afinidad siendo uno regulado por proteínas Gi/o