RESUMO
Three IgG1 monoclonal antibodies derived from BALB/c mice immunized with the Caki-1 human renal cell carcinoma (RCC) line react with antigens present in most human RCCs but restricted in their expression in normal adult tissues. Antibody DAL-K20 reacted with five of six RCCs and the lining epithelium of normal proximal and distal convoluted tubules. Antibody DAL-K29 reacted with eight of nine RCCs, with glomeruli, where it outlined the capillaries, and more weakly with prostatic glandular epithelium and the basal layer of the epidermis. K29 precipitated molecules with molecular weights of 118,000 and 150,000 from extracts of surface-labeled Caki-1 cells. Antibody DAL-K45 reacted with four of six RCCs but not with any normal adult tissue including kidney. It precipitated Mr 177,000 and 150,000 antigens. The three antibodies showed distinct patterns of reactivity with human tumor cell lines.
Assuntos
Anticorpos Monoclonais/imunologia , Antígenos de Neoplasias/imunologia , Antígenos de Superfície/imunologia , Carcinoma de Células Renais/imunologia , Neoplasias Renais/imunologia , Ensaio de Imunoadsorção Enzimática , Imunofluorescência , Humanos , Técnicas ImunoenzimáticasRESUMO
BN 50730 [tetrahydro-4,7,8,10 methyl-1(chloro-2 phenyl)-6 (methoxy-4 phenyl-carbamoyl)-9 pyrido [4',3'-4,5] thieno [3,2-f] triazolo-1,2,4 [4,3-alpha] diazepine-1,4], a novel platelet-activating factor (PAF) receptor antagonist with a hetrazepine structure, decreased the maximal number of binding sites (Bmax) of [3H]PAF in rabbit platelet membranes without altering its dissociation constant. Platelet aggregation induced by 1 microM PAF was prevented by preincubation with 1 microM BN 50730. The washing of the platelets preincubated with BN 50730 failed to revert its inhibitory effects. We conclude that BN 50730 acts as a non-competitive antagonist of the PAF receptor, due to the formation of a highly stable drug-receptor complex.
Assuntos
Azepinas/farmacologia , Plaquetas/metabolismo , Inibidores da Agregação Plaquetária/farmacologia , Glicoproteínas da Membrana de Plaquetas/antagonistas & inibidores , Receptores de Superfície Celular , Receptores Acoplados a Proteínas G , Triazóis/farmacologia , Animais , Azepinas/química , Sítios de Ligação , Membrana Celular/metabolismo , Fator de Ativação de Plaquetas/antagonistas & inibidores , Inibidores da Agregação Plaquetária/química , Glicoproteínas da Membrana de Plaquetas/química , Coelhos , Tienopiridinas , Triazóis/químicaRESUMO
Previous studies demonstrated that the selective inhibition of phosphodiesterase type IV suppresses antigen-induced eosinophil infiltration and also downregulates certain eosinophil functions assessed in vitro. In the current study, we compared the effect of selective inhibitors of phosphodiesterase IV with the effect of phosphodiesterase III and V inhibitors, focusing on eosinophil chemotaxis stimulated by platelet-activating factor (PAF) and leukotriene B4 in a modified Boyden chamber. The effect of beta 2-adrenoceptor agonists and forskolin as well as the analogue N6-2'-O-dibutyryladenosine 3':5'-cyclic monophosphate (Bt2 cyclic AMP) was also determined. For this purpose eosinophils were obtained by lavage of the peritoneal cavity of normal Wistar rats and purified on Percoll gradients to 85-95% purity. Our results showed that PAF and leukotriene B4 (0.001-10 microM) elicited a concentration-dependent increase in eosinophil migration with maximal responses observed at 1 microM and 0.1 microM respectively. Pre-incubation with the type IV phosphodiesterase inhibitor, rolipram (1-100 microM), suppressed the chemotactic response triggered by PAF and leukotriene B4, in association with elevation of eosinophil cyclic AMP, whereas the compounds milrinone and SK&F 94836 (type III selective) as well as zaprinast (type V selective) were ineffective. The beta 2-adrenoceptor agonists salbutamol and salmeterol (1-100 microM) did not alter the intracellular levels of cyclic AMP and also failed to inhibit the eosinophil response. Moreover, incubation of eosinophils with the adenylate cyclase activator forskolin (1-100 microM), while inducing a discrete increase in cyclic AMP, markedly inhibited PAF- and leukotriene B4-induced eosinophil chemotaxis. Eosinophils treated with a combination of individually inactive amounts of forskolin plus rolipram significantly inhibited the eosinophil migration elicited by PAF and leukotriene B4, but did not change cyclic AMP baseline levels. Though only at the highest concentration tested (100 microM), the analogue Bt2 cyclic AMP abolished the eosinophil chemotaxis. Thus we conclude that the direct inhibitory effect of phosphodiesterase IV inhibitors on eosinophil chemotaxis may account for their suppressive activity on tissue eosinophil accumulation following antigen challenge.
Assuntos
Quimiotaxia de Leucócito/efeitos dos fármacos , Eosinófilos/imunologia , Inibidores de Fosfodiesterase/farmacologia , Agonistas Adrenérgicos beta/farmacologia , Animais , Bucladesina/farmacologia , Colforsina/farmacologia , AMP Cíclico/análise , Isoenzimas/antagonistas & inibidores , Leucotrieno B4/farmacologia , Fator de Ativação de Plaquetas/farmacologia , Agregação Plaquetária/efeitos dos fármacos , Pirrolidinonas/farmacologia , Ratos , Ratos Wistar , RolipramRESUMO
The pro-inflammatory activity of enterolobin, a haemolytic protein from Enterolobium contortisiliquum seeds, was investigated. In doses ranging from 1 to 20 micrograms/site, enterolobin induced a dose-dependent paw oedema and pleurisy in rats. The effect was apparent after 15 min, peaked at 6 hr and decreased 24 hr after enterolobin was administered. One hour after the intrathoracic injection of enterolobin, the total leukocyte content of the pleural cavity increased significantly, mainly due to mononuclear and neutrophil accumulation. At 24 hr, although the number of mononuclear and neutrophil cells tended to decrease, a great rise in eosinophil counts was noted. Intraperitoneal treatment with the dual lipoxygenase and cyclooxygenase blockers, BW 755c (25 mg/kg) and NDGA (50 mg/kg) or the corticosteroid dexamethasone (0.1 mg/kg) inhibited enterolobin-induced paw oedema by 35, 38 and 47% respectively, whereas indomethacin (2 mg/kg) was inactive. The H1 antagonist, meclizine (25 mg/kg), was also effective against enterolobin oedema while the PAF-antagonists WEB 2086 and PCA 4248 (20 mg/kg) did not modify the reaction. It was concluded that enterolobin is a potent inducer of pleural exudation, cellular infiltration and paw oedema. Furthermore, enterolobin-induced oedema is partially dependent on lipoxygenase metabolites and histamine, while PAF and prostaglandins did not seem to be important in this reaction.
Assuntos
Edema/induzido quimicamente , Leucócitos/efeitos dos fármacos , Proteínas de Plantas/toxicidade , Pleurisia/induzido quimicamente , Árvores , Animais , Anti-Inflamatórios não Esteroides/farmacologia , Inibidores de Ciclo-Oxigenase/farmacologia , Relação Dose-Resposta a Droga , Edema/tratamento farmacológico , Edema/fisiopatologia , Feminino , Contagem de Leucócitos , Leucócitos/química , Masculino , Meclizina/farmacologia , Proteínas de Plantas/administração & dosagem , Fator de Ativação de Plaquetas/antagonistas & inibidores , Pleurisia/tratamento farmacológico , Pleurisia/fisiopatologia , Ratos , Ratos Endogâmicos , Sementes/químicaRESUMO
This study investigated the effect of successive daily intrathoracic (it) injections of PAF-acether upon its demonstrated ability to generate eosinochemotaxin(s). Repeated administration of PAF-acether led to a selective state of desensitization, characterized by a gradual reduction of its ability to induce exudation. Concomitantly, however, there was a progressive pleural accumulation of eosinophils leading to a 7-fold increase in the eosinophil counts after the 4th restimulation. The generation of eosinochemotaxin(s) elicited by PAF-acether was not modified by desensitization, as detected by transferring the cell-free pleural fluid from donor to recipient animals. We conclude that, in contrast to exudation, eosinophil tissue infiltration induced by PAF-acether is not a desensitizable phenomenon.
Assuntos
Eosinofilia/induzido quimicamente , Eosinófilos/fisiologia , Fator de Ativação de Plaquetas/farmacologia , Derrame Pleural/induzido quimicamente , Animais , Agregação Celular/efeitos dos fármacos , Feminino , Masculino , Fator de Ativação de Plaquetas/administração & dosagem , Ratos , Ratos EndogâmicosAssuntos
Doença de Chagas , Insetos Vetores , Análise para Determinação do Sexo , Animais , Larva , Metamorfose Biológica , TriatominaeRESUMO
The ability of a furofuran lignan, epiyangambin, to inhibit PAF-induced rabbit platelet aggregation in vitro and thrombocytopenia in rats was investigated. Epiyangambin dose-dependently inhibited PAF-induced platelet aggregation without modifying the amplitude of the maximal response, indicating a competitive antagonism. The IC50 value of epiyangambin for 10(-9) M PAF-induced aggregation was 6.1 x 10(-7) M and the Schild analysis provided a pA2 of 6.91 +/- 0.2 with a slope of 0.98 +/- 0.25 (n = 4) and a pKb of 6.94 +/- 0.19. Epiyangambin had no effect upon the platelet aggregation induced by collagen, thrombin or ADP. The in vivo administration of the lignan at 20 mg/kg significantly inhibited PAF-induced thrombocytopenia in rats. These data indicate that epiyangambin is a potent and selective antagonist of PAF both in vitro and in vivo.
Assuntos
Furanos/farmacologia , Lignanas , Lignina/farmacologia , Fator de Ativação de Plaquetas/antagonistas & inibidores , Inibidores da Agregação Plaquetária/farmacologia , Trombocitopenia/tratamento farmacológico , Animais , Masculino , Estrutura Molecular , Coelhos , Ratos , Ratos Wistar , Trombocitopenia/induzido quimicamenteRESUMO
The effects of the furofuran lignan yangambin on rabbit platelet aggregation and binding of [3H]-PAF to rabbit platelet plasma membranes were studied. Log concentration-response curves to PAF were obtained in the presence or absence of increasing concentrations of yangambin. This lignan dose-dependently inhibited PAF-induced platelet aggregation in platelet-rich plasma (PRP) and shifted PAF curves to the right without decreasing the maximal response. The Schild plot constructed from these data showed a slope of 1.17 and a pA2 of 6.45. Moreover, yangambin at 10(-5) M did not inhibit the platelet aggregation induced by ADP (5 x 10(-7) M), collagen (0.1 microgram ml-1), or thrombin (0.05 U ml-1). Biochemical studies showed that [3H]-PAF labelled in a saturable manner a single class of binding sites on platelet membranes with a Kd of 1.25 +/- 0.24 nM and a maximal binding capacity (Bmax) of 14.9 +/- 2.4 pmol mg protein-1. Both unlabelled PAF and yangambin competitively displaced [3H]-PAF binding with an IC50 of 1.54 +/- 0.37 nM and 1.93 +/- 0.53 microM, respectively. The incubation of rabbit blood neutrophils with yangambin at 10(-5) M did not prevent PAF-induced in vitro chemotaxis in conditions where the PAF antagonist SR 27417 at 10(-5) M abolished the phenomenon. These results indicate that yangambin is an antagonist that selectively blocks PAF receptors on platelets.
Assuntos
Furanos/farmacologia , Lignanas/farmacologia , Plantas/química , Glicoproteínas da Membrana de Plaquetas/antagonistas & inibidores , Receptores de Superfície Celular , Receptores Acoplados a Proteínas G , Animais , Quimiotaxia de Leucócito/efeitos dos fármacos , Furanos/metabolismo , Lignanas/metabolismo , Neutrófilos/citologia , Neutrófilos/efeitos dos fármacos , Fator de Ativação de Plaquetas/farmacologia , Agregação Plaquetária/efeitos dos fármacos , Inibidores da Agregação Plaquetária/metabolismo , Inibidores da Agregação Plaquetária/farmacologia , CoelhosRESUMO
This study investigated the effect of successive daily intrathoracic (it) injections of PAF-acether upon its demonstrated ability to generate eosinochemotaxin(s). Repeated administration of PAF-acether led to a selective state of desensitization, characterized by a gradual reduction of its ability to induce exudation. Concomitantly, however, there was a progressive pleural accumulation of eosinophilis leading to a 7-fold increase in the eosinophil counts after the 4th restimulation. The generation of eosinochemotaxin(s) elicited by PAF-acether was not modified by desensitization, as detected by transferring the cell-free pleural fluid from donor to recipient animals. We conclude that, in contrast to exudation, eosinophil tissue infiltration induced by PAF-acether is not a desensitizable phenomenon