RESUMO
Several studies have now clearly established the ability of LPS to induce bradykinin B(1) receptor up-regulation in vivo and the functional relevance of this up-regulation for the pathophysiological effects of LPS. Using an in vivo system in which LPS is injected locally into the rat paw, we have examined the potential contribution of proinflammatory cytokines, NF-kappaB activation, and neutrophil influx for the functional and molecular up-regulation of the bradykinin B(1) receptor. Treatment with LPS resulted in a rapid and sustained functional up-regulation of B(1) receptors in the rat paw that correlated with the increase in B(1) receptor mRNA levels. B(1) receptor up-regulation is preceded by the rapid activation of the transcription factor NF-kappaB and the production of proinflammatory cytokines, including TNF-alpha and IL-1beta. More importantly, blockade of NF-kappaB translocation, TNF-alpha, or IL-1beta prevented the functional and molecular up-regulation of B(1) receptors. Injection of LPS also induced the influx of neutrophils that followed the peak of cytokine production and associated with the persistent activation of NF-kappaB and functional B(1) receptor up-regulation. Blockade of neutrophil influx with platelet-activating factor receptor antagonists or cell adhesion molecule blockers prevented B(1) receptor up-regulation. Thus, by acting in cooperation and in a coordinated, timely manner, TNF-alpha, IL-1beta, neutrophils, and the transcription factor NF-kappaB are major and essential players in the ability of LPS to induce B(1) receptor expression in vivo.
Assuntos
Citocinas/fisiologia , Mediadores da Inflamação/fisiologia , Lipopolissacarídeos/administração & dosagem , Infiltração de Neutrófilos/imunologia , Receptor B1 da Bradicinina/biossíntese , Regulação para Cima/imunologia , Animais , Edema/imunologia , Edema/patologia , Edema/fisiopatologia , Membro Posterior , Injeções Intradérmicas , Interleucina-1/fisiologia , Masculino , NF-kappa B/metabolismo , NF-kappa B/fisiologia , Ratos , Ratos Wistar , Receptor B1 da Bradicinina/metabolismo , Fator de Necrose Tumoral alfa/fisiologiaRESUMO
By the low-stringency single-specific-primer PCR technique, a highly sensitive and rapid method for diagnosis of rifampin resistance in Mycobacterium tuberculosis was established. Seven rifampin-resistant and five rifampin-susceptible specimens were analyzed. Rifampin resistance was determined by MIC measurement. A complex electrophoretic pattern consisting of many bands was obtained for both susceptible and rifampin-resistant isolates. The same pattern was obtained for all of the susceptible specimens, but differences between resistant and susceptible isolates were found. DNA sequencing showed that a particular mutation produces a specific electrophoretic pattern.
Assuntos
Antibióticos Antituberculose/farmacologia , RNA Polimerases Dirigidas por DNA/genética , Farmacorresistência Bacteriana/genética , Mutação , Mycobacterium tuberculosis/efeitos dos fármacos , Reação em Cadeia da Polimerase/métodos , Rifampina/farmacologia , Sequência de Bases , Primers do DNA , Eletroforese em Gel de Poliacrilamida , Humanos , Testes de Sensibilidade Microbiana , Dados de Sequência Molecular , Mycobacterium tuberculosis/classificação , Mycobacterium tuberculosis/genética , Análise de Sequência de DNA , Especificidade da Espécie , Tuberculose Pulmonar/microbiologiaRESUMO
1. The present study evaluated the ability of the administration of platelet activating factor (PAF) to induce the upregulation of B(1) receptors in the rat paw. 2. Local treatment with PAF resulted in a time-dependent increase of oedema formation induced by the B(1) receptor agonist des-Arg(9)-BK (des-Arg(9)-bradykinin), but not by the B(2) receptor agonist tyrosine(8)-bradykinin. Functional upregulation of B(1) receptors was accompanied by a prominent increase of B(1) receptor mRNA expression in the rat paw. 3. In PAF-treated paws, des-Arg(9)-BK-induced oedema formation was significantly inhibited by the B(1) receptor antagonists des-Arg(9)-[Leu(8)]-BK and R-715. The effects of PAF pretreatment were receptor operated, as assessed by the effects of the PAF receptor antagonist WEB2086 or by desensitisation of PAF receptors. 4. The protein synthesis inhibitor cycloheximide, the anti-inflammatory steroid dexamethasone or the nuclear factor (NF-kappaB) blockers pyrrolidine-dithiocarbamate (PDTC) and Nalpha-tosyl-L-chloromethylketone significantly blocked the functional upregulation of B(1) receptors. 5. The selectin inhibitor fucoidin, an anti-CD18 antibody or an anti-rat neutrophil antiserum, also significantly prevented des-Arg(9)-BK-induced paw oedema in rats pretreated with PAF. 6. Intradermal injection of PAF induced a 25-fold increase of myeloperoxidase activity in the rat paw, a response that was significantly inhibited by fucoidin, anti-CD-18, anti-rat neutrophil antiserum or PDTC. 7. Local treatment with PAF also resulted in a marked increase of NF-kappaB activation, an effect largely prevented by PDTC or by the anti-rat neutrophil antiserum. 8. Collectively, the present results indicate that the induction of B(1) receptors following treatment with the chemotatic mediator PAF is dependent on the recruitment of neutrophils, an event that is under the control of adhesion molecules, protein synthesis and NF-kappaB activation. These findings provide new insights into the role played by cell migration and chemotatic factors on B(1) receptor upregulation in vivo.