Novel strategy in Trypanosoma cruzi cell invasion: Implication of cholesterol and host cell microdomains

Trypanosoma cruzi, the etiological agent of Chagas' disease, is an obligatory intracellular parasite in the mammalian host. in order to invade a wide variety of mammalian cells, T cruzi engages parasite components that are differentially expressed among strains and infective forms. Because the identification of putative protein receptors has been particularly challenging, we investigated whether cholesterol and membrane rafts, sterol- and sphingolipid-enriched membrane domains, could be general host surface components involved in invasion of metacyclic trypomastigotes and extracellular amastigotes of two parasite strains with distinct infectivities. HeLa or Vero cells treated with methyl-beta-cyclodextrin (M beta CD) are less susceptible to invasion by both infective forms, and the effect was dose-dependent for trypomastigote but not amastigote invasion. Moreover, treatment of parasites with MPCD only inhibited trypomastigote invasion. Filipin labeling confirmed that host cell cholesterol concentrated at the invasion sites. Binding of a cholera toxin B subunit (CTX-B) to ganglioside GM1, a marker of membrane rafts, inhibited parasite infection. Cell labeling with CTX-B conjugated to fluorescein isothiocyanate revealed that not only cholesterol but also GM1 is implicated in parasite entry. These findings thus indicate that microdomains present in mammalian cell membranes, that are enriched in cholesterol and GM1, are involved in invasion by T cruzi infective forms. (c) 2007 Australian Society for Parasitology Inc. Published by Elsevier B.V. All rights reserved.Universidade Federal de São Paulo, Escola Paulista Med, Dept Microbiol Immunol & Parasitol, BR-04023062 São Paulo, BrazilUniversidade Federal de São Paulo, Escola Paulista Med, Dept Bioquim, BR-04023062 São Paulo, BrazilUniversidade Federal de São Paulo, Escola Paulista Med, Dept Microbiol Immunol & Parasitol, BR-04023062 São Paulo, BrazilUniversidade Federal de São Paulo, Escola Paulista Med, Dept Bioquim, BR-04023062 São Paulo, BrazilWeb of Scienc

Diagnosis of American cutaneous leishmaniasis by enzyme immunoassay in patients from Northern Paraná State, Brazil

American cutaneous leishmaniasis (ACL) is an endemic disease caused by Leishmania parasites. The ACL diagnosis is commonly accomplished by parasitological and immunological methods. The objective of this work was to evaluate the enzyme immunoassay (EIA) for ACL laboratorial diagnosis. IgG antibodies against Leishmania (Viannia) braziliensis promastigotes were researched. For the method standardization 240 sera were used: 72 from patients with positive parasitological diagnosis, 38 from normal individuals and 113 from individuals with other pathologies. The sensibility was 93% and the specificity was 70%. Regarding diagnosis, EIA showed 97% positivity in patients with ACL and 97% negativity in patients with other cutaneous lesions. The EIA presented a good performance when used for diagnosis, thus, it may become an important tool for it and for further ACL epidemiological studies in endemic areas

APPROACHING CARBOHYDRATES AND ITS METABOLISM: AN EXPERIENCE FOR EDUCATIONAL GAMES

INTRODUCTION: The study of carbohydrates, its metabolism and many other fields of biochemistry are often understood by students as a junction of chemical structures and reactions of difficult compression. However, Biochemistry should no longer be seen as an abstruse field, but a way to know the human body and its components, including molecular, structural and functional aspects. Therefore, some alternatives are being evaluated in order to assist and improve the dissemination of knowledge among them highlights are the educational games. OBJECTIVES: The aim of this work is the production of two educational games able to include the contents of carbohydrates and its metabolism in higher education. MATERIALS AND METHODS: The educational games produced were made from available materials and low cost. The games were tested in courses of Biological Sciences and Biotechnology of the Federal University of Uberlândia and the response of the students towards the activities was analyzed. The application, had the presence of trained students to instruct on the activity and correcting. DISCUSSION AND RESULTS: "What is the Carbohydrate?" and "Mastering the metabolism" are two educational games covering the content of structure and function carbohydrates and basal metabolism, respectively. "What is the Carbohydrate?" consists in unravel amid several options the carbohydrate in the hands of the others players. For this, several questions with two possible answers, “yes” or “not”, are accepted each round, and if the player find difficulty in formulating questions, there are cards tips. “Mastering the metabolism” consists in a combination of cards that simulate pieces of a domino that must be mounted following the metabolic pathway of carbohydrates, and as the game progresses, the main points of regulation of the pathway will be accompanied by surprise questions. The games showed great acceptance by students. CONCLUSION: “What is the Carbohydrate?” and “Mastering the Metabolism” are productive methodological alternatives for teaching Biochemistry

In vitro additive interaction between ketoconazole and antimony against intramacrophage Leishmania (Leishmania) amazonensis amastigotes.

Leishmaniasis is a group of diseases caused by protozoa of Leishmania genus. The currently available treatments for this disease are expensive, present high toxicity and are associated to difficulties of healing and parasite resistance. Therefore, the development of strategies for leishmaniasis treatment is indispensable and includes reposition of existing drugs, as well as drug combination therapy. The aim of this study was to assess the nature of ketoconazole and antimony association on the cytotoxic effect against Leishmania (Leishmania) amazonensis amastigotes. The calculated mean sum of fractional 50% inhibitory concentration ([Formula: see text]ΣFIC50) was 2.54 and 1.43 for free and intracellular amastigotes, respectively, values that suggest an additive interaction between ketoconazole and antimony concerning to Leishmania toxicity only in the intramacrophage parasite form. Despite the clinical efficacy of ketoconazole-antimony combination has been shown in the literature, our study is the first to describe the nature of ketoconazole-antimony interaction against L. (L.) amazonensis amastigotes. Moreover, our results point out the need for future in vivo studies to confirm the nature of ketoconazole-antimony interaction and also to determine possible effective dosage regimens related to ketoconazole administration in association with the optimal lower dose of antimony

FROM AMINO ACIDS TO ENZYMES: A PROPOSAL USING DIDACTIC GAMES TO REVIEW THE CONTENT OF PROTEINS

INTRODUCTION: Biochemistry is a discipline offered in graduation courses. In general, the students define Biochemistry as “complex and extensive”, since, in most cases, a broad content is ministered in a short period of time. The consequences of this context may be reflected in the lack of interest by the students. Thus, the use of alternative methodologies presents potential to improve this situation. OBJECTIVES: Develop and implement educational games about protein content to review and arouse students' interest in the discipline. MATERIALS AND METHODS: The educational games were made from available materials and low cost. The games were applied in extracurricular schedules in Biology, Biotechnology, Biomedicine and Agronomy courses of the Federal University of Uberlândia in the second half of 2015. All of this games count with the presence of a mediator to solve questions and correct mistakes that can appear. DISCUSSION AND RESULTS: Three educational games were created: "Amino Game 2.0", "Memo Protein" and "Race of Enzymes" which addressing the content of amino acid ionization, structure and function of proteins and enzymes, respectively. "AminoGame 2.0" is based on the correct assembly of structures of amino acids, dipeptides or tripeptides in according to conditions described on card game randomly selected by the player. "Memoprotein" is a memory game in which the player must correctly answer a question about protein to have the right to turn the cards of the game. Finally, "Race of Enzymes" is a board game in which the player must correctly answer questions about enzymes to have the right to advancing squares on the board. The application of three games showed good acceptance among students. CONCLUSION: “Amino Game 2.0", "Memo Protein" and "Race of Enzymes" were considered to be effective tools in the teaching of Biochemistry

A comparative study on the leishmanicidal activity of the L-amino acid oxidases BjussuLAAO-II and BmooLAAO-II isolated from Brazilian Bothrops snake venoms

This study aims to examine whether two L-amino acid oxidases isolated from Bothrops snake venom (SV-LAAOs) were cytotoxic to Leishmania (Leishmania) amazonensis and Leishmania (Viannia) braziliensis, two causative agents of leishmaniasis, which is an endemic disease in tropical and subtropical countries. The SV-LAAOs BjussuLAAO-II and BmooLAAO-II were isolated from Bothrops jararacussu and Bothrops moojeni venom, respectively, through a three-step chromatography process that used molecular exclusion, hydrophobic interaction, and affinity columns. BmooLAAO-II is a new SV-LAAO isoform that we isolated in this study. The purified BjussuLAAO-II and BmooLAAO-II had high L-amino acid oxidase-specific activity: 3481.17 and 4924.77 U/mg/min, respectively. Both SV-LAAOs were strongly cytotoxic to the two Leishmania species, even at low concentrations. At the same concentration, BjussuLAAO-II and BmooLAAO-II exerted different cytotoxic effects on the parasites. We reported for the first time that the SV-LAAOs suppressed cell proliferation and altered the mitochondrial membrane potential of the two Leishmania species. Surprisingly, BjussuLAAO-II increased the intracellular reactive oxygen species production only in L. (L.) amazonensis, while BmooLAAO-II increased the intracellular reactive oxygen species production only in L. (V.) braziliensis, indicating that these SV-LAAOs had a certain specificity of action