ABSTRACT
Ecto-nucleoside triphosphate diphosphohydrolases (NTPDases) are enzymes located on the surface of the T. cruzi plasma membrane, which hydrolyze a wide range of tri-/-diphosphate nucleosides. In this work, we used previously developed genetically modified strains of Trypanosoma cruzi (T. cruzi), hemi-knockout (KO +/−) and overexpressing (OE) the TcNTPDase-1 gene to evaluate the parasite infectivity profile in a mouse model of acute infection (n = 6 mice per group). Our results showed significantly higher parasitemia and mortality, and lower weight in animals infected with parasites OE TcNTPDase-1, as compared to the infection with the wild type (WT) parasites. On the other hand, animals infected with (KO +/−) parasites showed no mortality during the 30-day trial and mouse weight was more similar to the non-infected (NI) animals. In addition, they had low parasitemia (45.7 times lower) when compared with parasites overexpressing TcNTPDase-1 from the hemi-knockout (OE KO +/−) group. The hearts of animals infected with the OE KO +/− and OE parasites showed significantly larger regions of cardiac inflammation than those infected with the WT parasites (p < 0.001). Only animals infected with KO +/− did not show individual electrocardiographic changes during the period of experimentation. Together, our results expand the knowledge on the role of NTPDases in T. cruzi infectivity, reenforcing the potential of this enzyme as a chemotherapy target to treat Chagas disease (CD).
Subject(s)
Chagas Disease , Trypanosoma cruzi , Mice , Animals , Chagas Disease/genetics , Chagas Disease/parasitology , Heart , Disease Models, AnimalABSTRACT
Background: Autologous whole blood (AWB) administration is described as alternative/complementary medical practice widely employed in medical and veterinary therapy against infections, chronic pathologies and neoplasias. Our aim is to investigate in vivo biological effect of AWB using healthy murine models under the course of Trypanosoma cruzi acute infection. Methods: The first set of studies consisted of injecting different volumes of AWB and saline (SAL) into the posterior region of quadriceps muscle of healthy male Swiss mice under distinct therapeutic schemes evaluating: animal behavior, body and organ weight, hemogram, plasmatic biochemical markers for tissue damage and inflammatory cytokine levels and profile. To assess the impact on the experimental T. cruzi infection, different schemes (prior and post infection) and periods of AWB administration (from one up to 10 days) were conducted, also employing heterologous whole blood (HWB) and evaluating plasma cytokine profile. Results: No major adverse events were observed in healthy AWB-treated mice, except gait impairment in animals that received three doses of 20 L AWB in the same hind limb. AWB and SAL triggered an immediate polymorphonuclear response followed by mononuclear infiltrate. Although SAL triggered an inflammatory response, the kinetics and intensity of the histological profile and humoral mediator levels were different from AWB, the latter occurring earlier and more intensely with concomitant elevation of plasma IL-6. Inflammatory peak response of SAL, mainly composed of mononuclear cells with IL-10, was increased at 24 h. According to the mouse model of acute T. cruzi infection, only minor decreases ( 30%) in the parasitemia levels were produced by AWB and HWB given before and after infection, without protecting against mortality. Rises in IFN-gamma, TNF-alpha and...
Subject(s)
Animals , Mice , Autoantigens/therapeutic use , Blood Transfusion, Autologous , Trypanosoma cruziABSTRACT
Drug repurposing has been an interesting and cost-effective approach, especially for neglected diseases, such as Chagas disease. Methods: In this work, we studied the activity of the antidepressant drug sertraline against Trypanosoma cruzi trypomastigotes and intracellular amastigotes of the Y and Tulahuen strains, and investigated its action mode using cell biology and in silico approaches. Results: Sertraline demonstrated in vitro efficacy against intracellular amastigotes of both T. cruzi strains inside different host cells, including cardiomyocytes, with IC50 values between 1 to 10 µM, and activity against bloodstream trypomastigotes, with IC50 of 14 µM. Considering the mammalian cytotoxicity, the drug resulted in a selectivity index of 17.8. Sertraline induced a change in the mitochondrial integrity of T. cruzi, resulting in a decrease in ATP levels, but not affecting reactive oxygen levels or plasma membrane permeability. In silico approaches using chemogenomic target fishing, homology modeling and molecular docking suggested the enzyme isocitrate dehydrogenase 2 of T. cruzi (TcIDH2) as a potential target for sertraline. Conclusions: The present study demonstrated that sertraline had a lethal effect on different forms and strains of T. cruzi, by affecting the bioenergetic metabolism of the parasite. These findings provide a starting point for future experimental assays and may contribute to the development of new compounds.(AU)
Subject(s)
Trypanosoma cruzi , In Vitro Techniques , Sertraline , Drug RepositioningABSTRACT
Autologous whole blood (AWB) administration is described as alternative/complementary medical practice widely employed in medical and veterinary therapy against infections, chronic pathologies and neoplasias. Our aim is to investigate in vivo biological effect of AWB using healthy murine models under the course of Trypanosoma cruzi acute infection. Methods: The first set of studies consisted of injecting different volumes of AWB and saline (SAL) into the posterior region of quadriceps muscle of healthy male Swiss mice under distinct therapeutic schemes evaluating: animal behavior, body and organ weight, hemogram, plasmatic biochemical markers for tissue damage and inflammatory cytokine levels and profile. To assess the impact on the experimental T. cruzi infection, different schemes (prior and post infection) and periods of AWB administration (from one up to 10 days) were conducted, also employing heterologous whole blood (HWB) and evaluating plasma cytokine profile. Results: No major adverse events were observed in healthy AWB-treated mice, except gait impairment in animals that received three doses of 20 µL AWB in the same hind limb. AWB and SAL triggered an immediate polymorphonuclear response followed by mononuclear infiltrate. Although SAL triggered an inflammatory response, the kinetics and intensity of the histological profile and humoral mediator levels were different from AWB, the latter occurring earlier and more intensely with concomitant elevation of plasma IL-6. Inflammatory peak response of SAL, mainly composed of mononuclear cells with IL-10, was increased at 24 h. According to the mouse model of acute T. cruzi infection, only minor decreases (< 30%) in the parasitemia levels were produced by AWB and HWB given before and after infection, without protecting against mortality. Rises in IFN-gamma, TNF-alpha and IL-6 were detected at 9 dpi in all infected animals as compared to uninfected mice but only Bz displayed a statistically significant diminution (p= 0.02) in TNF-alpha levels than infected and untreated mice. Conclusions: This study revealed that the use of autologous whole blood (AWB) in the acute model employed was unable to reduce the parasitic load of infected mice, providing only a minor decrease in parasitemia levels (up to 30%) but without protecting against animal mortality. Further in vivo studies will be necessary to elucidate the effective impact of this procedure.(AU)
Subject(s)
Animals , Male , Rats , Trypanosoma cruzi , Blood Transfusion, Autologous , Chagas Disease/blood , Complementary TherapiesABSTRACT
Chagas disease (CD) is caused by the intracellular protozoan parasite Trypanosoma cruzi and affects more than 10 million people in poor areas of Latin America. There is an urgent need for alternative drugs with better safety, broader efficacy, lower costs and shorter time of administration. Thus the biological activity of viniconazole, a chloroaryl-substituted imidazole was investigated using in vitro and in vivo screening models of T. cruzi infection. Ultrastructural findings demonstrated that the most frequent cellular damage was associated with plasma membrane (blebs and shedding events), Golgi (swelling aspects) and the appearance of large numbers of vacuoles suggesting an autophagic process. Our data demonstrated that although this compound is effective against bloodstream and intracellular forms (16 and 24 µ m, respectively) in vitro, it does not present in vivo efficacy. Due to the urgent need for novel agents against T. cruzi, the screening of natural and synthetic products must be further supported with the aim of finding more selective and affordable drugs for CD.
Subject(s)
Chagas Disease/drug therapy , Imidazoles/pharmacology , Trypanocidal Agents/pharmacology , Trypanosoma cruzi/drug effects , Animals , Chagas Disease/parasitology , Disease Models, Animal , Dose-Response Relationship, Drug , Humans , Imidazoles/chemistry , Imidazoles/therapeutic use , Male , Mice , Parasitemia/drug therapy , Parasitemia/parasitology , Parasitic Sensitivity Tests , Primary Cell Culture , Trypanocidal Agents/chemistry , Trypanocidal Agents/therapeutic use , Trypanosoma cruzi/ultrastructureABSTRACT
Trypanosoma cruzi is a parasite that causes Chagas disease, which affects millions of individuals in endemic areas of Latin America. One hundred years after the discovery of Chagas disease, it is still considered a neglected illness because the available drugs are unsatisfactory. Aromatic compounds represent an important class of DNA minor groove-binding ligands that exhibit potent antimicrobial activity. This study focused on the in vitro activity of 10 aromatic dicationic compounds against bloodstream trypomastigotes and intracellular forms of T. cruzi. Our data demonstrated that these compounds display trypanocidal effects against both forms of the parasite and that seven out of the 10 compounds presented higher anti-parasitic activity against intracellular parasites compared with the bloodstream forms. Additional assays to determine the potential toxicity to mammalian cells showed that the majority of the dicationic compounds did not considerably decrease cellular viability. Fluorescent microscopy analysis demonstrated that although all compounds were localised to a greater extent within the kinetoplast than the nucleus, no correlation could be found between compound activity and kDNA accumulation. The present results stimulate further investigations of this class of compounds for the rational design of new chemotherapeutic agents for Chagas disease.
Subject(s)
Animals , Mice , Trypanocidal Agents/pharmacology , Trypanosoma cruzi/drug effects , Microscopy, Fluorescence , Myocytes, Cardiac/parasitology , Parasitic Sensitivity Tests , Time FactorsABSTRACT
A doença de Chagas, causada pelo protozoário Trypanosoma cruzi, é uma parasitose relevante e negligenciada afetando 16-18 milhões de indivíduos em áreas endêmicas da América Latina. A quimioterapia corrente, baseada em 2 nitroderivados (nifurtimox e benznidazol), é insatisfatória principalmente pela baixa atividade na fase crônica, alta toxicidade e devido ao aumento de isolados resistentes. Diamidinas aromáticas apresentam importante atividade anti-tumoral e antiparasitária, e vêem sendo utilizadas no tratamento de doenças parasitárias humanas e animais. Porém, devido à baixa biodisponibilidade oral e alta toxicidade, estudos têm sido desenvolvidos de modo a sintetizar e avaliar a ação de novos análogos dicatiônicos. Assim, a presente dissertação teve por objetivo caracterizar a atividade de amidinas reversas (ARs) contra Trypanosoma cruzi in vitro, comparando-a com a ação da diguanidina DB711. Nossos dados revelaram que ARs exibem importante efeito antiparasitário sobre amastigotas e tripomastigotas de T. cruzi em doses micro e nanomolares que não afetam a viabilidade de células de mamíferos. As amidinas DB889 e DB702 apresentaram superior atividade na maioria dos sistemas avaliados (diferentes formas evolutivas, diferentes condições de temperatura, e adição de sangue) em relação às duas outras ARs testadas, DB786 e DB811, sugerindo que pequenas varuações na estrutura química resultam em diferenças na potência antiparasitária. A análise das ARs sobre amastigotas não revelou diferenças no perfil de susceptibilidade entre os 2 estoques de T. cruzi testados, cepa Y e clone DM28c. Observamos superior atividade das ARs sobre tripomastigotas durante incubação a 37grausC, e que a adição de sangue reduziu a atividade tripanocida da maioria dos compostos testados à 4grausC, possivelmente devido à sua associação a componentes plasmáticos, como já relatado em outros estudos. Ensaios...A análise por microscopia eletrônica de transmissão (MET) revelou importantes altera...
