Atividade terapêutica do 5-nitrofuraldeído-semicarbazona (nitrofurazona) em esquemas de duração prologada na infeccção experimental do camundongo pelo Trypanosoma cruzi / Therapeutic activity of 5-nitrofuraldeído-semicarbazone (nitrofurazone) in schemes prolong the duration of infectious disease in experimental mice by Trypanosoma cruzi

Revisões históricas aos avanços científicos para o controle da doença, o Simpósio Internacional Comemorativo do Centenário da Descoberta da Doença de Chagas (1909-2009).

Parasitological cure of acute and chronic experimental Chagas disease using the long-acting experimental triazole TAK-187. Activity against drug-resistant Trypanosoma cruzi strains.

We investigated the activity of TAK-187, an experimental antifungal triazole with a long terminal half-life in several experimental animals, against Trypanosoma cruzi. In vitro studies showed that the minimal inhibitory concentration (MIC) against the (extracellular) epimastigote form was 0.3-1 microM, while the corresponding concentration against clinically relevant intracellular amastigotes was 1 nM. At the MIC the endogenous epimastigote C4,14-desmethyl sterols were replaced by di- and tri-methylated sterols, supporting the notion that the primary target of TAK-187 is the parasite's sterol C14alpha demethylase. We investigated the in vivo activity of the compound in a murine model of acute Chagas disease, using T. cruzi strains with different susceptibilities to the drugs currently used clinically (nitrofurans and nitroimidazoles). It was found that TAK-187 given orally at 20 mg/kg induced complete protection against death and high levels (60-100%) of parasitological cures, independently of the infecting strain and even when administered every other day (e.o.d.), consistent with its long terminal half-life in mice. Other experiments, using longer treatment periods were carried out in both acute and chronic models of the disease and showed that TAK-187 given at 10-20 mg/kg e.o.d. induced 80-100% survival with 80-100% of parasitological cures of survivors in both models. No toxic side effects were observed in any of the experimental protocols. TAK-187 is a potent anti-T. cruzi compound with trypanocidal activity in vivo and should be considered for further studies as a potential specific treatment of human Chagas disease.

Zigman Brener: depoimento / Zigman Brener: a statement

Fala sobre seu ingresso na carreira científica, as principais contribuições para a pesquisa em doença de Chagas, os grupos de pesquisa em doença de Chagas no Brasil, as pesquisas em doenças endêmicas e saúde pública e as fontes de financiamento à pesquisa em doenças endêmicas.(MAM)

Double-blind study to evaluate flow cytometry analysis of anti-live trypomastigote antibodies for monitoring treatment efficacy in cases of human Chagas' disease.

The validation of flow cytometry analysis of anti-live trypomastigote antibodies (FC-ALTA) to monitor cure after treatment of Chagas' disease was evaluated with serum samples from treated and nontreated chagasic patients. After optimization of the original technique, toward better sensitivity and applicability to field surveys, we design a double blind study of 94 coded samples classified into the following categories: patients not treated (NT) and patients treated but not cured (TNC), both presenting positive conventional serology and xenodiagnosis; patients treated and cured (TC), showing negative serology and xenodiagnosis; and patients treated under evaluation (TUE), who presented positive or oscillating conventional serology (CSA) but negative xenodiagnosis. Coded samples, diluted 1:256, were assayed by incubation with live cell culture trypomastigotes, which were subsequently stained with fluorescein isothiocyanate-conjugated anti-human immunoglobulin G, with prior fixation and analysis by flow cytometry. The results were expressed as the percentages of positive fluorescent parasites (PPFP) for each individual sample, establishing 20% PPFP as the cutoff between negative and positive results. Our data demonstrated that all NT and TNC presented positive results while all but one TC had a PPFP lower than 20%. Analysis of TUE demonstrated a wide degree of reactivity, with PPFP values that were negative (PPFP 50%). As TUE with negative PPFP presented negative xenodiagnosis and positive or oscillating CSA, they were classified as dissociated according to the criteria of Krettli and Brener (J. Immunol. 128:2009-2012, 1982) and could indeed be considered cured after chemotherapy. This study demonstrates and validates the use of FC-ALTA to easily identify anti-live trypomastigote membrane-bound antibodies, offering another approach for investigating and monitoring the efficacy of specific chemotherapy in cases of human Chagas' disease.

Ultrastructural observations on trypanosoma herpetosoma rangeli in the salivary glands of rhodnius ecuadoriensis, hemiptera, reduvidae

Fue realizado un estudio ultraestructural de las glándulas salivares del triatomino rhodnius ecuadoriensis experimentalmente infectado por el tripanosomatídeo, trypanosoma (herpetosoma) rangeli, con la finalidad de documentar los aspectos básicos de la infección glandular por este flagelado. Gran número de formas epimastigotes fueron encontradas en la hemolinfa que baña la parte externa de las glándulas en el hemoceloma. Algunos flagelados parecían intimamente asociados a la membrana basal del epitelio glandular y varios de ellos parecían estar iniciando el proceso de su invasión, rompiendo la lámina basal con el flagelo. En el interior de las células glandulares observamos que, T. rangeli se presentaba en la forma de esteromastigotes, células redondeadas rodeadas externamente por un flagelo, que envolvía apretadamente el parásito. En la luz glandular también observamos gran número de epimastigotes mezclados con los productos de la secreción glandular, el material presentó apenas pocas formas paramastigotes y trypomastigotes identificadas de manera inquívoca. El significado de los paramastigotes en el ciclo evolutivo del T. rangeli aguarda futuros estudios

Evaluation of the rabbit as a model for Chagas disease. II. Histopathologic studies of the heart, digestive tract and skeletal muscle

In order to investigate the value of the rabbit as an experimental model for Chagas'disease, seventy one animals were inoculated with different Trypanosoma cruzi strains and routes. The rabbits were submitted to necropsy in acute (earlier than three months of infection), recent chronic (three to six months) and late chronic (later than six months) phases. Myocarditis, generally focal and endomysial, ocurred in 94.1 per cent, 66.7 per cent and 70.8 per cent of the infected rabbits respectively in the acute, recent chronic and late chronic phases. The myocardial inflammatory exudate was composed by mononuclear cells, and also polymorphonuclear cells in the acute phase. In most cases of the late chronic phase, the myocarditis was similar to that described in the indeterminate form of human chagasic patients. Initial fibrosis occurred in the three phases but was more severe and frequent in the early chronic. Advanced fibrosis occurred only in the late chronic phase. Tissue parasites occurred only in the acute phase. The digestive tract and skeletal muscles showed mild and occasional lesions. Our data indicate that experimentally infected chagasic rabbits repeat some lesions similar to that of humans chagasic patients, specially that of the indetermined form. So, it may be a useful, however not an ideal, model.

Recrudescence induced by cyclophosphamide of chronic Trypanosoma cruzi infection in mice is influenced by the parasite strain

Reactivation of chronic chagasic patients may occur upon of use of immunosupressive drugs related to kidney or heart transplantation or when they are affected by concomitant HIV infection. This recrudescence, however, does not occur in all chagasic patients exposed to immunosuppressive agents. We therefore investigated the influence of Trypanosoma cruzi strains in the recrudescence of the parasitism in mice at the chronic phase treated with cyclophosphamide, an immunosuppressor that blocks lymphocytes DNA synthesis and therefore controls B cells response. A large variation was detected in the percentages of newly established acute phases in the groups of mice inoculated with the different strains. We suggest that reactivation of chronic T. cruzi infections is influenced by the parasite intrinsic characteristics, a phenomenon that might occur in the human disease.

The impact of immunology research in Chagas disease in Brazil: A personal overview

This personal overview intends to demonstrate that the Trypanosoma cruzi model of research has been extensively used by groups of Brazilian scientists who with or without formal training in immunology contributed with basic and applied data related to this discipline for a better knowledge of experimental and human Chagas disease. Since the bulk of information in the immunology of this disease is rather numerous the author selected for this review more familiar topics or those in which he directly participated.

An experimental and clinical assay with ketoconazole in the treatment of Chagas disease

Ketoconazole an azole antifungic drug which is already in the market has also been demonstrated to be active against Trypanossoma cruzi experimental infections. In this paper we confirmed the drug effect and investigated its range of activity against different T. cruzi strains naturally resistant or susceptible to both standard drugs Nifurtimox and Benznidazole used clinically in Chagas disease. Moreover, we have shown that the association of Ketoconazole plus Lovastatin (an inhibitor of sterol synthesis), which has an antiproliferative effect against T. cruzi in vitro, failed to enhance the supressive effect of Ketoconazole displayed when administered alone to infected mice. Finally, administration in chronic chagasic patients of Ketoconazole at doses used in the treatment of deep mycosis also failed to induce cure as demonstrated by parasitological and serological tests. The strategy of identify and test drugs which are already in the market and fortuitously are active against T. cruzi has been discussed

Lectin receptors distribution in the surface membrane of Trypanosoma cruzi blood forms collected from mice submitted to specific treatment

The author investigated the distribution of lectin receptors on Trypanosoma cruzi blood forms collected from mice inoculated with, respectively, the drug-resistant and drug-sensitive strains VL-10 and CL, and treated with the two standard active nitroheterocyclic compounds nifurtimox and benznidazole used for treatment of human Chagas' disease. Blood trypomastigotes purified in Fycoll-Hypaque were incubated with fluorescein-labelled lectins Con A, WGA, EE, WFA, TPA and PNA and then microscopically examined. Neither qualitative or quantitative differences in the fluorescence intensity could be detected between parasites from VL-10 and CL strains submitted or not to treatment. The results suggest that both strains do not differ in their surface membrane carbohydrate moieties. Moreover, the rapid clearance of blood forms the drug-sensitive strain in animals treated with singlo doses of both compounds is not likely to depend on membrane alterations expressed by changes in the carbohydrate components. furthermore, resistance or sensitivity to drugs is not apparently related to carbohydrate distribution on T. cruzi blood forms

Effect of the host specific treatment in the phagocytosis of Trypanosoma cruzi blood forms by mouse peritoneal macrophages

Single doses of drugs active aginst Trypanosoma cruzi (megazol, nifurtimox and benznidazole) induce a rapid clearence of the blood parasites in experimentally infected mice. Furthermore, the in vitro phagocytosis and intracellular destruction by mouse peritoneal macrophage of blood forms collected from the treatment animals is strongly enhanced as compared with parasites from untreated controls. The uptake of the blood forms by macrophages is significantly higher with megazol than with benznidazole and nifurtimox, a finding that concurs with data showing that megazol is also the most active compound in the living host. The possibility that macrophages participate in a synergic effect between the host immune response and chemotherapeutic effect is discussed

Evaluation of the rabbit as a model for chagas' disease: I. Parasitological studies

Com a finalidade de investigar o uso do coelho como modelo experimental da doença de Chagas, 72 desses animais foram inoculados por via intraperitoneal e conjuntival com formas sanguíneas, tripomastigotas metacíclicos obtidos de triatomíneos e tripomastigotas obtidos de cultura de tecido, usando-se as cepas Y, CL e Ernane do T. cruzi. E, 95,6% dos coelhos o exame de sangue a fresco foi positivo, o que evidencia uma alta suscetibilidade desses animais à injecçäo pelo T. cruzi. O curso da parasistemia na fase aguda da infecçäo é intensamente influenciado pela cepa do parasita e via de inoculaçäo usada. Na fase crônica parasitas foram recuperados através de xenodiagnóstico e/ou hemocultura em 40% dos coelhos excaminados. A análise dos resultados dos xenodiagnósticos revelou a existência de interaçöes seletivas entre as cepas do T. cruzi e as quatro espécies de vetores empregados, evidenciadas por significativa variabilidade dos resultados. De um modo geral os resultados obtidos no presente trabalho atendem ao pre-requisito de ordem parasitológica sugerido para um modelo para a doença de Chagas na fase crônica

Xenodiagnóstico artificial "post-mortem" em chagásicos crônicos / Post-mortem artificial xenodiagnosis in chronic chagasic patients

A fim de obter metodologias que permitam estabelecer, com segurança, o diagnóstico "post-mortem" da infecçäo chagásica, adaptou-se o xenodiagnóstico artifical a necropsiados com diferentes tempos de óbito. O teste foi positivo em três (30%) de dez chagásicos autopsiados. O tempo decorrido entre o êxito letal e o início do repasto pelos triatomíneos destes chagásicos foi de duas horas, duas horas e quinze minutos e sete horas, respectivamente. Discutem-se os fatores que podem explicar a sobrevivência do Trypanosoma cruzi no hospedeiro morto bem como as aplicaçöes práticas do achado

Schistosoma mansoni: immunodepresion of hepatic schistosome granuloma formation in mice infected by Trypanosoma cruzi

A infecçäo de camundongos pelo Trypanosoma cruzi inibe a formaçäo do granuloma hepático esquistossomótico. Este efeito imunodepressor ocorreu em camundongos com a fase aguda ou crônica da doença de Chagas, sendo os granulomas significativamente menores que nos animais controles. Os dados sugerem que a doença de Chagas deprime diretamente a imunidade celular

Estudo parasitológico e anátomo-patológico da fase aguda da doença de Chagas em cäes inoculados com duas diferentes cepas do Trypanosoma cruzi / A parasitological and pathological study of the acute phase of Chagas' disease in dogs inoculated with 2 different strains of Trypanosoma cruzi

Cäes jovens foram infectados com as cepas Y e CL do T. cruzi usando-se como inóculos 10**7 formas sangüíneas inoculadas por via intraperitoneal e 2x10**3 tripomastigotas metacíclios obtidos do inseto vetor e inoculados por via conjuntival. As cepas Y e CL induziram nos cäes curvas de parasitemia totalmente distintas, confirmando dados parasitológicos obtidos em camundongos e coelhos. Com a cepa CL a parasitemia, com ambos os inóculos, foi gradualmente ascencional ao passo que com Y a parasitemia foi extremamente baixa, irregular e, com freqüência, subpatente. Com ambas as cepas o parasitismo e as lesöes predominaram no miocárdio. Entretanto, com a cepa Y a miocardite foi sempre intensa desde as fases mais precoces da infecçäo, ao passo que com a cepa CL o processo inflamatório tornou-se acentuado somente a partir do 20§ dia. Freqüentemente a intensidade da miocardite observada em alguns animais näo guardava relaçäo com a parasitemia; em alguns cäes com parasitemia subpatente, nos quais a infecçäo só foi diagnosticada pelo xenodiagnóstico, a intensidade da miocardite foi comparável àquela observada nos animais com parasitemia patente. Idêntica correlaçäo também näo foi assinalada em relaçäo ao parasitismo tissular. Esses achados sugerem a participaçäo de mecanismo imunológicos na gênese das lesöes, ainda na fase aguda da infecçäo