RESUMO
The intracellular metabolites of long-term in vitro cultured axenic amastigotes of Leishmania donovani (strain Dd8) were determined and compared with those of promastigotes and intracellular amastigotes, employing proton NMR spectroscopy. The presence of two new metabolites, i.e. betaine and beta-hydroxybutyrate were reported. Betaine was detected in all the three stages being highest in the promastigotes while beta-hydroxybutyrate could be detected only in promastigotes and axenic amastigotes. Among other metabolites, succinate and valine were found in higher quantities in intracellular amastigotes and axenic amastigotes than in promastigotes. Acetoacetate was present only in axenic and intracellular amastigotes. The comparative metabolite profile of different parasite forms reveals that axenic amastigotes seem to represent an intermediate stage between promastigotes and intracellular amastigotes in spite of their strong resemblance to intracellular amastigotes in morphology, infectivity, biochemical studies and even in the manifestation of amastigote specific A2 protein.
Assuntos
Leishmania donovani/química , Ácido 3-Hidroxibutírico/análise , Animais , Antígenos de Protozoários/análise , Betaína/análise , Hidrogênio , Leishmania donovani/fisiologia , Ressonância Magnética Nuclear Biomolecular , Succinatos/análise , Valina/análiseRESUMO
Proton nuclear magnetic resonance spectroscopy was used for studying the intracellular metabolite profile of promastigotes of Leishmania donovani. The major intracellular metabolites observed in the promastigotes were acetate, alanine, succinate, glycine, alpha-glycerophosphorylcholine, acetoacetate, arginine and ethanol. A comparative study of the intracellular metabolite profile of promastigotes of different strains of L. donovani showed that, all the major intracellular metabolites were present in promastigotes of different strains. A quantitative estimation of metabolites showed a strain specific (Finger print) metabolite profile which can be used for strain/species identification/differentiation.
Assuntos
Leishmania donovani/metabolismo , Acetatos/metabolismo , Acetoacetatos/metabolismo , Alanina/metabolismo , Animais , Arginina/metabolismo , Etanol/metabolismo , Glucose/metabolismo , Glicerilfosforilcolina/metabolismo , Glicina/metabolismo , Ácido Láctico/metabolismo , Leishmania donovani/crescimento & desenvolvimento , Espectroscopia de Ressonância Magnética , Especificidade da Espécie , Succinatos/metabolismo , Ácido SuccínicoRESUMO
In vitro as well as in vivo studies suggest that cytokine-induced synthesis of nitric oxide (NO) from L-arginine is a major effector mechanism against intracellular pathogens. In this study, we demonstrate that golden hamsters infected with Leishmania donovani amastigotes upon treatment with polyinosinic-polycytidylic acid stabilized with polylysine and carboxymethylcellulose (poly ICLC), a potent interferon inducer and immune enhancer, in combination with L-arginine, develop the capacity to eliminate intracellular pathogens. This antileishmanial activity of poly ICLC was suppressed by N w nitro-L-arginine (N w NLA), an inhibitor of inducible NO synthase. Furthermore, prolonged treatment of infected animals with L-arginine alone for 5 days more after 5 day treatment with poly ICLC plus L-arginine increased the antileishmanial activity compared with 5 day treatment with poly ICLC plus L-arginine, suggesting that inducible NO synthase, once activated, produces NO for 5 days more. Our results suggest that an L-arginine-dependent, NO-mediated mechanism is probably responsible for the antileishmanial action of poly ICLC.
Assuntos
Arginina/uso terapêutico , Carboximetilcelulose Sódica/análogos & derivados , Indutores de Interferon/uso terapêutico , Leishmania donovani/efeitos dos fármacos , Leishmaniose Visceral/tratamento farmacológico , Poli I-C/uso terapêutico , Polilisina/análogos & derivados , Animais , Arginina/antagonistas & inibidores , Carboximetilcelulose Sódica/uso terapêutico , Cricetinae , Avaliação Pré-Clínica de Medicamentos , Quimioterapia Combinada , Indução Enzimática , Indutores de Interferon/antagonistas & inibidores , Leishmania donovani/crescimento & desenvolvimento , Masculino , Mesocricetus , Óxido Nítrico/biossíntese , Óxido Nítrico Sintase/biossíntese , Nitroarginina/uso terapêutico , Poli I-C/farmacologia , Polilisina/farmacologia , Polilisina/uso terapêuticoRESUMO
After presenting processed glycoprotein of Leishmania donovani to T-cell, macrophage seeks the help of a panel of T-cells lymphokines to transform from a state that sustains intra cellular replication of parasite to an effector state for destructing parasites. But esterase and trypsin of macrophage membrane prevent T-cells to release MIF. Role of soya-bean trypsin inhibitor (STI) has been exposed in the present study with a view to alter esterase functional behaviour of macrophage for control of T-cell activation and also, if T-cells once made responsive to antigen by STI do alter macrophage response to T-cells or not. Results establish STI as potent effector molecule, which can serve as an adjuvant to candidate T-cell epitope and synthetic peptide for development of anti-Kala-azar vaccine protocol in future.
Assuntos
Imunidade Celular , Leishmaniose Visceral/imunologia , Ativação Linfocitária/efeitos dos fármacos , Linfócitos T/imunologia , Inibidores da Tripsina/farmacologia , Inibição de Migração Celular , Citotoxicidade Imunológica , Relação Dose-Resposta a Droga , Técnicas In Vitro , Ativação de Macrófagos , Macrófagos/imunologiaRESUMO
The efficacy of sodium stibogluconate against Leishmania donovani infections was markedly enhanced by encapsulating this drug in tuftsin-bearing liposomes. Also, pretreatment of the animals with these liposomes (free of drug) rendered them resistant to this infection, possibly by activating the host's macrophages. These results demonstrate that tuftsin-bearing liposomes besides delivering the drug to the target cells could also enhance the nonspecific resistance against infections, thus offering an additional advantage over the use of tuftsin-free liposomes as drug carriers in leishmania therapy.