RESUMEN

Leishmania infantum infantum (LII) is one of the species that causes visceral leishmaniasis (VL) in the Old World, while L. infantum chagasi (LIC) is present in the New World. Few studies address biological differences or the behavior of these strains during infection. These parasites live inside cells of their hosts, continuously evading microbicidal mechanisms and modulating the immune responses of these cells. One of the mechanisms used by these protozoa involves the L-arginine metabolism. Understanding the differences between Leishmania species and establishing an improved murine model for study of leishmaniasis are matters of extreme importance. Thereby, the objectives of this work were to analyze the biological and molecular differences between two Leishmania infantum strains (LII and LIC) and the degree of susceptibility to infection of mice with different genetic backgrounds. The infectivity in vivo and in vitro of LII and LIC strains was evaluated in BALB/c and Swiss Webster mice, as well the NOS and ARG activities. The LII strain was more infective than the LIC strain both in vivo and in vitro. In animals infected by the LII and LIC strains, differences in NOS and ARG activities occurred. In vitro, promastigotes of LII isolated from BALB/c and Swiss Webster mice showed higher ARG activity than LIC promastigotes during the growth curve. However, no difference was observed in intracellular NO production by promastigotes of these strains. The ARG gene sequences were compared, and those of both strains were identical. However, despite the similarity, the strains showed different expression levels of this gene. It can be concluded that although L. chagasi strains are considered identical to L. infantum strains from a molecular point of view, these strains have different biological behavior.

Asunto(s)

Leishmania infantum/inmunología , Leishmaniasis Cutánea/inmunología , Leishmaniasis Cutánea/parasitología , Leishmaniasis Visceral/inmunología , Leishmaniasis Visceral/parasitología , Animales , Femenino , Ratones , Ratones Endogámicos BALB C

RESUMEN

The leishmaniases represent a public health problem in under-developed countries and are considered a neglected disease by the World Health Organization (WHO). They are cuased by Leishmania  parasites with different clinical manifestations. Currently, there is no vaccine, and treatment is in-efficient and is associated with both serious side effects often leading to resistance to the parasites. Thus, it is essential to search for new treatment strategies, such as drug repurposing, i.e., the use of drugs that are already used for other diseases. The discovery of new clinical applications for approved drugs is strategic for lowering the cost of drug discovery since human toxicity assays are already conducted. Here, we review a broad analysis of the different aspects of this approach for anti-leishmanial treatment.

Asunto(s)

Reposicionamiento de Medicamentos/métodos , Leishmania/efectos de los fármacos , Leishmaniasis/tratamiento farmacológico , Tripanocidas/farmacología , Animales , Interacciones Huésped-Parásitos/efectos de los fármacos , Humanos , Leishmania/fisiología , Leishmaniasis/parasitología , Resultado del Tratamiento

RESUMEN

This study evaluated the in vitro sensitivity of paired Leishmania braziliensis samples isolated from the same patient before pentavalent antimonial treatment (Sample A) and after treatment failure or cutaneous leishmaniasis reactivation (Sample B) in patients undergoing intralesional administration or injections (5 mgSb(V)/kg/d) of meglumine antimoniate. Fourteen samples from 7 patients were studied. After 24 h of drug exposure, 50% lethal dose (LD50) values for promastigotes ranged from 0.37 mg/mL to 5.86 mg/mL for samples obtained before treatment (A) and 0.89 mg/mL to 7.80 mg/mL for samples obtained after treatment (B). After 48 h, LD50 values ranged from 0.37 mg/mL to 5.75 mg/mL and 0.70 mg/mL to 7.68 mg/mL for A and B samples, respectively. After 48 h, LD50 values for amastigotes ranged from 11.7 to 44.3 µg/mL for A samples and 13.7 to 52.7 µg/mL for B samples. Of 7 patients, 1 discontinued treatment and 6 were cured after retreatment with amphotericin B (4 cases) or meglumine antimoniate (2 cases). Overall the B samples had higher LD50 values than A samples; however the difference was not significant. These results do not support the hypothesis that low-dose and intralesional treatments induce selection of resistant parasites in vitro and suggest that other factors may influence therapeutic outcome in patients with poor response to initial treatment.

Asunto(s)

Antiprotozoarios/farmacología , Leishmania braziliensis/efectos de los fármacos , Leishmaniasis/tratamiento farmacológico , Meglumina/farmacología , Compuestos Organometálicos/farmacología , Adolescente , Adulto , Anciano , Antiprotozoarios/uso terapéutico , Femenino , Humanos , Leishmania braziliensis/aislamiento & purificación , Leishmaniasis/parasitología , Dosificación Letal Mediana , Masculino , Meglumina/uso terapéutico , Antimoniato de Meglumina , Persona de Mediana Edad , Compuestos Organometálicos/uso terapéutico , Piel/parasitología

RESUMEN

Leishmaniasis is a spectrum of infectious diseases caused by Leishmania protozoan parasites. The purpose of this study was to perform, in vitro, a comparative analysis of the activity amastigotes. Results showed excellent efficacy of all compounds against axenic amastigotes, compared to pentamidine isethionate, the reference drug used. The cytotoxic effect of these mesoionic compounds of six mesoionic compounds (three 1,3,4-thiadiazolium-2-aminide and three 1,2,3-oxadiazolium-5-olate class compounds) was evaluated in mouse peritoneal macrophages using MTT assay, low toxicity (≈ 10%) for these mammalian cells being observed. In an attempt to define a potential drug target, the activities of nitric oxide synthase (NOS) and arginase of the parasites treated with the mesoionic derivatives were evaluated. NOS was purified from a cell-free extract of infective promastigotes and axenic amastigotes and all derivatives tested were able to inhibit the enzyme as monitored by the decrease of NADPH consumption. Arginase activity from both stages of the parasite was measured using urea production and none of the compounds inhibited the enzyme activity of axenic amastigotes. However, the compounds without substituents (MI-H and SID-H) were able to inhibit arginase activity of these parasites.

Asunto(s)

Arginasa/metabolismo , Leishmania mexicana/efectos de los fármacos , Óxido Nítrico Sintasa/metabolismo , Oxadiazoles/farmacología , Tiadiazoles/farmacología , Animales , Arginasa/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Cinamatos/síntesis química , Cinamatos/química , Cinamatos/farmacología , Concentración 50 Inhibidora , Leishmania mexicana/enzimología , Leishmania mexicana/crecimiento & desarrollo , Macrófagos Peritoneales/efectos de los fármacos , Macrófagos Peritoneales/parasitología , Ratones , Ratones Endogámicos BALB C , Óxido Nítrico Sintasa/efectos de los fármacos , Oxadiazoles/síntesis química , Oxadiazoles/química , Cavidad Peritoneal/citología , Cavidad Peritoneal/parasitología , Tiadiazoles/síntesis química , Tiadiazoles/química

RESUMEN

Drug resistance in protozoan parasites has been associated with the P-glycoprotein (Pgp), an energy-dependent efflux pump that transports substances across the membrane. Interestingly, the genes TcPGP1 and TcPGP2 have been described in Trypanosoma cruzi, although the function of these genes has not been fully elucidated. The main goal of this work was to investigate Pgp efflux pump activity and expression in T. cruzi lines submitted to in vitro induced resistance to the compounds 4-N-(2-methoxy styryl)-thiosemicarbazone (2-Meotio) and benznidazole (Bz) and to verify the stability of the resistant phenotypes during the parasite life cycle. We observed that the EC50 values for the treatment of epimastigotes with 2-Meotio or Bz were increased at least 4.7-fold in resistant lines, and this phenotype was maintained in metacyclic trypomastigotes, cell-derived trypomastigotes, and intracellular amastigotes. However, in epimastigotes, 2-Meotio resistance is reversible, but Bz resistance is irreversible. When compared with the parental line, the resistant lines exhibited higher Pgp efflux activity, reversion of the resistant phenotypes in the presence of Pgp inhibitors, cross-resistance with Pgp modulators, higher basal Pgp ATPase activity, and overexpression of the genes TcPGP1 and TcPGP2. In conclusion, the resistance induced in T. cruzi by the compounds 2-Meotio and Bz is maintained during the entire parasite life cycle. Furthermore, our data suggest the participation of the Pgp efflux pump in T. cruzi drug resistance.

Asunto(s)

Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/metabolismo , Antiprotozoarios/metabolismo , Antiprotozoarios/farmacología , Resistencia a Medicamentos , Proteínas Protozoarias/metabolismo , Trypanosoma cruzi/efectos de los fármacos , Trypanosoma cruzi/metabolismo , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/genética , Adenosina Trifosfatasas/genética , Adenosina Trifosfatasas/metabolismo , Perfilación de la Expresión Génica , Nitroimidazoles/metabolismo , Nitroimidazoles/farmacología , Tiosemicarbazonas/metabolismo , Tiosemicarbazonas/farmacología , Trypanosoma cruzi/genética

RESUMEN

In Leishmania amazonensis, kinetoplastid membrane protein-11 (KMP-11) expression increases during meta-cyclogenesis and is higher in amastigotes than in promastigotes, suggesting a role for this protein in the infection of the mammalian host. We show that the addition of KMP-11 exacerbates L. amazonensis infection in peritoneal macrophages from BALB/c mice by increasing interleukin (IL)-10 secretion and arginase activity while reducing nitric oxide (NO) production. The doses of KMP-11, the IL-10 levels and the intracellular amastigote loads were strongly, positively and significantly correlated. The increase in parasite load induced by KMP-11 was inhibited by anti-KMP-11 or anti-IL-10 neutralising antibodies, but not by isotype controls. The neutralising antibodies, but not the isotype controls, were also able to significantly decrease the parasite load in macrophages cultured without the addition of KMP-11, demonstrating that KMP-11-induced exacerbation of the infection is not dependent on the addition of exogenous KMP-11 and that the protein naturally expressed by the parasite is able to promote it. In this study, the exacerbating effect of KMP-11 on macrophage infection with Leishmania is for the first time demonstrated, implicating it as a virulence factor in L. amazonensis. The stimulation of IL-10 production and arginase activity and the inhibition of NO synthesis are likely involved in this effect.

Asunto(s)

Arginasa/metabolismo , Interleucina-10/inmunología , Leishmania mexicana/efectos de los fármacos , Macrófagos Peritoneales/parasitología , Proteínas de la Membrana/farmacología , Óxido Nítrico/biosíntesis , Proteínas Protozoarias/farmacología , Animales , Células Cultivadas , Femenino , Interleucina-10/metabolismo , Leishmania mexicana/inmunología , Macrófagos Peritoneales/enzimología , Macrófagos Peritoneales/inmunología , Ratones , Ratones Endogámicos BALB C

RESUMEN

In Leishmania amazonensis, kinetoplastid membrane protein-11 (KMP-11) expression increases during metacyclogenesis and is higher in amastigotes than in promastigotes, suggesting a role for this protein in the infection of the mammalian host. We show that the addition of KMP-11 exacerbates L. amazonensis infection in peritoneal macrophages from BALB/c mice by increasing interleukin (IL)-10 secretion and arginase activity while reducing nitric oxide (NO) production. The doses of KMP-11, the IL-10 levels and the intracellular amastigote loads were strongly, positively and significantly correlated. The increase in parasite load induced by KMP-11 was inhibited by anti-KMP-11 or anti-IL-10 neutralising antibodies, but not by isotype controls. The neutralising antibodies, but not the isotype controls, were also able to significantly decrease the parasite load in macrophages cultured without the addition of KMP-11, demonstrating that KMP-11-induced exacerbation of the infection is not dependent on the addition of exogenous KMP-11 and that the protein naturally expressed by the parasite is able to promote it. In this study, the exacerbating effect of KMP-11 on macrophage infection with Leishmania is for the first time demonstrated, implicating it as a virulence factor in L. amazonensis. The stimulation of IL-10 production and arginase activity and the inhibition of NO synthesis are likely involved in this effect.

Asunto(s)

Animales , Femenino , Ratones , Arginasa/metabolismo , /inmunología , Leishmania mexicana/efectos de los fármacos , Macrófagos Peritoneales/parasitología , Proteínas de la Membrana/farmacología , Óxido Nítrico/biosíntesis , Proteínas Protozoarias/farmacología , Células Cultivadas , Leishmania mexicana/inmunología , Ratones Endogámicos BALB C , Macrófagos Peritoneales/enzimología , Macrófagos Peritoneales/inmunología

RESUMEN

The essential oils obtained from leaves of Piper duckei and Piper demeraranum by hydrodistillation were analyzed by gas chromatography-mass spectrometry. The main constituents found in P. demeraranum oil were limonene (19.3%) and ß-elemene (33.1%) and in P. duckei oil the major components found were germacrene D (14.7%) and trans-caryophyllene (27.1%). P. demeraranum and P. duckei oils exhibited biological activity, with IC(50) values between 15 to 76 µg mL(-1) against two Leishmania species, P. duckei oil being the most active. The cytotoxicity of the essential oils on mice peritoneal macrophage cells was insignificant, compared with the toxicity of pentamidine. The main mono- and sesquiterpene, limonene (IC(50) = 278 µM) and caryophyllene (IC(50) = 96 µM), were tested against the strains of Leishmania amazonensis, and the IC(50) values of these compounds were lower than those found for the essential oils of the Piper species. The HET-CAM test was used to evaluate the irritation potential of these oils as topical products, showing that these oils can be used as auxiliary medication in cases of cutaneous leishmaniasis, with less side effects and lower costs.

Asunto(s)

Aceites Volátiles/farmacología , Piper/química , Hojas de la Planta/química , Animales , Macrófagos Peritoneales/efectos de los fármacos , Ratones , Aceites Volátiles/química

RESUMEN

The present study was designated to evaluate semi-quantitative antileishmanial activity of alkaloidal extracts that were obtained from 1g of different parts of Aspidosperma ramiflorum (leaves, roots, seeds, and stem barks). Alkaloidal extracts of barks and leaves presented a good activity against the extracellular form (promastigotes) of Leishmania (L.) amazonensis. It is known that compounds responsible for the antileishmanial activity in the alkaloidal extracts from A. ramiflorum are the monoterpenoid indole alkaloids ramiflorine A and ramiflorine B, therefore extracts obtained from different plant parts were analyzed by electrospray ionization mass spectrometry (ESI-MS) in order to evidence the presence of these bioactive alkaloids. Based on these findings, alkaloidal extract from leaves was fractionated on preparative thin-layer chromatography in a bioassay-guided fractionation affording individual purified ramiflorines A and B. Both ramiflorines A and B showed significant activity against Leishmania (L.) amazonensis (LD(50) values of 18.5±6.5µg/ml and 12.63±5.52µg/ml, respectively). Our results are showing that alkaloidal extract from leaves is a promising alternative to the use of stem barks from A. ramiflorum.

Asunto(s)

Alcaloides/farmacología , Aspidosperma/química , Leishmania/efectos de los fármacos , Extractos Vegetales/farmacología , Hojas de la Planta/química , Alcaloides/química , Alcaloides/aislamiento & purificación , Bioensayo , Cromatografía en Capa Delgada , Corteza de la Planta/química , Extractos Vegetales/química , Extractos Vegetales/aislamiento & purificación , Raíces de Plantas/química , Alcaloides de Triptamina Secologanina/química , Alcaloides de Triptamina Secologanina/aislamiento & purificación , Alcaloides de Triptamina Secologanina/farmacología , Semillas/química , Espectrometría de Masa por Ionización de Electrospray

RESUMEN

Trypanosoma cruzi is the causal agent of Chagas' disease, an infection with a great impact on public health in Latin America. One of the challenges to understand Chagas' disease lies on the complex host-parasite interaction. The understanding of this interaction requires the use of appropriate experimental models that mimic the human disease. Here, we have used two lineages of rats (Wistar and Holtzman) to comparatively evaluate the course of the acute infection (Y strain). Infection was monitored by parasitemia, cardiac and skeletal muscle parasitism and inflammation, heart ultrastructure, recruitment of monocytes/macrophages and nitric oxide, and arginase production by these cells. Although both rats were able to infect, only Holtzman rats developed a marked infection in the cardiac and skeletal muscles, in parallel to a high recruitment of first-line defense cells. A high number of inflammatory macrophages directed parasite clearance. By the end of the acute phase, Holtzman rats showed consistent disease control. Interestingly, parasite killing was not related to nitric oxide production likely inhibited by an arginase-dependent mechanism. Our work demonstrates differential responses of Holtzman and Wistar rats to T. cruzi, and highlights the use of Holtzman rats as useful models for further studies of cardiac/skeletal muscle tropism and innate immune responses that protect the host against parasite replication. This is important for the development of proper therapeutic interventions.

Asunto(s)

Enfermedad de Chagas/inmunología , Modelos Animales de Enfermedad , Corazón/parasitología , Interacciones Huésped-Parásitos , Macrófagos/inmunología , Miocardio/inmunología , Trypanosoma cruzi/patogenicidad , Animales , Enfermedad de Chagas/parasitología , Enfermedad de Chagas/fisiopatología , Femenino , Humanos , Inmunidad Innata , Inflamación/inmunología , Monocitos/inmunología , Miocardio/patología , Óxido Nítrico , Parasitemia , Ratas , Ratas Sprague-Dawley , Ratas Wistar , Trypanosoma cruzi/fisiología

RESUMEN

Extracts of propolis samples collected in Brazil and Bulgaria were assayed against four Leishmania species--Leishmania amazonensis, L. braziliensis, L. chagasi from the New World, and L. major from the Old World--associated to different clinical forms of leishmaniasis. The composition of the extracts has been previously characterized by high temperature high resolution gas chromatography coupled to mass spectrometry. Considering the chemical differences among the extracts and the behavior of the parasites, it was observed significant differences in the leishmanicidal activities with IC50/1 day values in the range of 2.8 to 229.3 microg/ml . An overall analysis showed that for all the species evaluated, Bulgarian extracts were more active than the ethanol Brazilian extract. As the assayed propolis extracts have their chemical composition determined it merits further investigation the effect of individual components or their combinations on each Leishmania species.

Asunto(s)

Antiprotozoarios/farmacología , Leishmania/efectos de los fármacos , Própolis/farmacología , Animales , Cromatografía de Gases , Espectrometría de Masas , Pruebas de Sensibilidad Parasitaria

RESUMEN

Extracts of propolis samples collected in Brazil and Bulgaria were assayed against four Leishmania species - Leishmania amazonensis, L. braziliensis, L. chagasi from the New World, and L. major from the Old World - associated to different clinical forms of leishmaniasis. The composition of the extracts has been previously characterized by high temperature high resolution gas chromatography coupled to mass spectrometry. Considering the chemical differences among the extracts and the behavior of the parasites, it was observed significant differences in the leishmanicidal activities with IC50/1 day values in the range of 2.8 to 229.3 µg/ml . An overall analysis showed that for all the species evaluated, Bulgarian extracts were more active than the ethanol Brazilian extract. As the assayed propolis extracts have their chemical composition determined it merits further investigation the effect of individual components or their combinations on each Leishmania species.

Asunto(s)

Animales , Antiprotozoarios/farmacología , Leishmania/efectos de los fármacos , Própolis/farmacología , Brasil , Bulgaria , Cromatografía de Gases , Espectrometría de Masas , Pruebas de Sensibilidad Parasitaria

RESUMEN

The activity of several diarylheptanoid derivatives (curcuminoids) was previously evaluated against Leishmania amazonensis promastigotes and among them the most active compound was the [1-(4-methoxy-phenyl)-7-(3,4-methoxy-4-hydroxy-phenyl)-1,6-heptadien-3, 5-dione]. This derivative was chosen to be assayed in vivo in a treatment trial. For these experiments, the curcuminoid compound was used in a concentration equivalent to the IC50/24 h, obtained from the previous study. Balb/c mice were inoculated subcutaneously in the footpad with L. amazonensis infective promastigotes and 4 weeks after the inoculation, the animals were treated with different schemes, varying from 1 to 3 doses. In all the experiments, Pentamidine Isethionate was used as reference drug under the same experimental conditions. The results showed that one dose was not enough to heal the lesion, however, with 2 and 3 doses the efficiency of the assayed compound was clear. On the other hand, treatment with Pentamidine Isethionate using the three different schemes was not satisfactory when compared to the curcuminoid derivative.

Asunto(s)

Antiprotozoarios/farmacología , Curcumina/análogos & derivados , Leishmania/efectos de los fármacos , Pentamidina/farmacología , Animales , Dosificación Letal Mediana , Masculino , Ratones , Ratones Endogámicos BALB C , Pruebas de Sensibilidad Parasitaria , Pentamidina/química

RESUMEN

The activity of several diarylheptanoid derivatives (curcuminoids) was previously evaluated against Leishmania amazonensis promastigotes and among them the most active compound was the [1-(4-methoxy-phenyl)-7-(3,4-methoxy-4-hydroxy-phenyl)-1,6-heptadien-3, 5-dione]. This derivative was chosen to be assayed in vivo in a treatment trial. For these experiments, the curcuminoid compound was used in a concentration equivalent to the IC50/24 h, obtained from the previous study. Balb/c mice were inoculated subcutaneously in the footpad with L. amazonensis infective promastigotes and 4 weeks after the inoculation, the animals were treated with different schemes, varying from 1 to 3 doses. In all the experiments, Pentamidine Isethionate was used as reference drug under the same experimental conditions. The results showed that one dose was not enough to heal the lesion, however, with 2 and 3 doses the efficiency of the assayed compound was clear. On the other hand, treatment with Pentamidine Isethionate using the three different schemes was not satisfactory when compared to the curcuminoid derivative

Asunto(s)

Animales , Ratones , Antiprotozoarios , Curcumina , Leishmania , Pentamidina , Curcumina , Ratones Endogámicos BALB C , Pentamidina