Characterization of macrophage polarization in lesions of dogs inoculated with Leishmania (Leishmania) infantum (BH401) strain

The aim of this work was to describe the anatomical pathology of dogs experimentally infected with Leishmania (Leishmania) infantum strain MCAN / BR / 2002 / BH401, a Brazilian form of L. infantum isolated from a symptomatic dog from an endemic area. For this, five beagles (three months old and both sexes) composed the experimental group. Markers of macrophage subpopulations M1 and M2 (related to resistance and susceptibility to visceral leishmaniasis) and the tissue cytokine transforming growth factor beta 1 (TGF-ß1) (one of the main cytokines related to the fibrosis process and anti-inflammatory action) were evaluated in livers, lungs and kidneys. The BH 401 L. infantum strain induced classical lesions of the visceral disease where all evaluated organs showed a chronic inflammatory reaction and tissue parasitism associated with a higher expression of CD163 and TGF-ß1 markers, might be related to the progression of the disease. In this work it was possible to conclude that the BH 401 strain reproduces canine visceral leishmaniasis that occurs naturally.(AU)

In vitro and in vivo activity of a hypotoxic copper(I) complex against dermotropic Leishmania species / Atividade in vitro e in vivo de um complexo de cobre(I) hipotóxico contra espécies dermotrópicas de Leishmania

Cutaneous leishmaniasis is a disease caused by protozoa of the genus Leishmania and, currently, the treatment of first choice is meglumine antimoniate. However, due to its limited effectiveness and high toxicity, it is necessary to seek new active principles for leishmaniasis treatment. Metal complexes are gaining importance due to their effectiveness and low toxicity. In this context, the present study aimed to evaluate the in vitro and in vivo antileishmanial activity of the hypotoxic copper(I) complex [HB(pz)3]Cu(PCN). Four dermotropic species of Leishmania were tested with the metal complex and its effectiveness was determined through parasitic viability and infectivity rate, and cytotoxicity was determined using a redox dye (resazurin). For the in vivo tests, hamsters were infected and the lesions treated with a formulated ointment containing the complex, the effectiveness of which was assessed by measuring the diameter of the inoculum/snout location and determining the parasitic load. The results demonstrated moderate toxicity in murine macrophages and human monocytes and better efficacy in Leishmania (V.) braziliensis when compared to the other species tested, with a 50% reduction in the viability of promastigote and amastigote forms (in vitro). General data from daily topical treatment for up to 30 days showed low efficacy for reducing lesions, and no clinical and parasitological cure was observed in the experimental animals. Thus, the [HB(pz)3]Cu(PCN) complex proved to be promising in in vitro studies against L. (V.) braziliensis, and should be further tested in new formulations and new experimental treatment schemes.

A leishmaniose cutânea é uma doença causada por protozoários do gênero Leishmania e, atualmente, o tratamento de primeira escolha é o antimoniato de meglumina. Porém, devido à sua eficácia limitada e alta toxicidade, é necessário buscar novos princípios ativos para o tratamento da leishmaniose. Os complexos metálicos vêm ganhando importância devido à sua eficácia e baixa toxicidade. Nesse contexto, o presente estudo teve como objetivo avaliar a atividade leishmanicida in vitro e in vivo do complexo hipotóxico de cobre(I) [HB(pz)3]Cu(PCN). Quatro espécies dermotrópicas de Leishmania foram testadas com o complexo metálico e sua eficácia foi determinada através da viabilidade parasitária e taxa de infectividade, e a citotoxicidade foi determinada com um corante redox (resazurina). Para os testes in vivo, hamsters foram infectados e as lesões foram tratadas com uma pomada formulada contendo o complexo. A eficácia foi avaliada medindo o diâmetro do inóculo/focinho e determinando a carga parasitária. Os resultados demonstraram toxicidade moderada em macrófagos murinos e monócitos humanos e melhor eficácia em Leishmania (V.) braziliensis quando comparada às demais espécies testadas, com redução de 50% na viabilidade das formas promastigotas e amastigotas (in vitro). Os dados gerais do tratamento tópico diário por até 30 dias mostraram baixa eficácia na redução das lesões, e nenhuma cura clínica e parasitológica foi observada nos animais experimentais. Portanto, o complexo [HB(pz)3]Cu(PCN) mostrou-se promissor em estudos in vitro contra L. (V.) braziliensis, devendo ser empregado em novas formulações e novos esquemas de tratamento experimental.

In vitro and in vivo activity of a hypotoxic copper(I) complex against dermotropic Leishmania species / Atividade in vitro e in vivo de um complexo de cobre(I) hipotóxico contra espécies dermotrópicas de Leishmania

Cutaneous leishmaniasis is a disease caused by protozoa of the genus Leishmania and, currently, the treatment of first choice is meglumine antimoniate. However, due to its limited effectiveness and high toxicity, it is necessary to seek new active principles for leishmaniasis treatment. Metal complexes are gaining importance due to their effectiveness and low toxicity. In this context, the present study aimed to evaluate the in vitro and in vivo antileishmanial activity of the hypotoxic copper(I) complex [HB(pz)3]Cu(PCN). Four dermotropic species of Leishmania were tested with the metal complex and its effectiveness was determined through parasitic viability and infectivity rate, and cytotoxicity was determined using a redox dye (resazurin). For the in vivo tests, hamsters were infected and the lesions treated with a formulated ointment containing the complex, the effectiveness of which was assessed by measuring the diameter of the inoculum/snout location and determining the parasitic load. The results demonstrated moderate toxicity in murine macrophages and human monocytes and better efficacy in Leishmania (V.) braziliensis when compared to the other species tested, with a 50% reduction in the viability of promastigote and amastigote forms (in vitro). General data from daily topical treatment for up to 30 days showed low efficacy for reducing lesions, and no clinical and parasitological cure was observed in the experimental animals. Thus, the [HB(pz)3]Cu(PCN) complex proved to be promising in in vitro studies against L. (V.) braziliensis, and should be further tested in new formulations and new experimental treatment schemes.(AU)

A leishmaniose cutânea é uma doença causada por protozoários do gênero Leishmania e, atualmente, o tratamento de primeira escolha é o antimoniato de meglumina. Porém, devido à sua eficácia limitada e alta toxicidade, é necessário buscar novos princípios ativos para o tratamento da leishmaniose. Os complexos metálicos vêm ganhando importância devido à sua eficácia e baixa toxicidade. Nesse contexto, o presente estudo teve como objetivo avaliar a atividade leishmanicida in vitro e in vivo do complexo hipotóxico de cobre(I) [HB(pz)3]Cu(PCN). Quatro espécies dermotrópicas de Leishmania foram testadas com o complexo metálico e sua eficácia foi determinada através da viabilidade parasitária e taxa de infectividade, e a citotoxicidade foi determinada com um corante redox (resazurina). Para os testes in vivo, hamsters foram infectados e as lesões foram tratadas com uma pomada formulada contendo o complexo. A eficácia foi avaliada medindo o diâmetro do inóculo/focinho e determinando a carga parasitária. Os resultados demonstraram toxicidade moderada em macrófagos murinos e monócitos humanos e melhor eficácia em Leishmania (V.) braziliensis quando comparada às demais espécies testadas, com redução de 50% na viabilidade das formas promastigotas e amastigotas (in vitro). Os dados gerais do tratamento tópico diário por até 30 dias mostraram baixa eficácia na redução das lesões, e nenhuma cura clínica e parasitológica foi observada nos animais experimentais. Portanto, o complexo [HB(pz)3]Cu(PCN) mostrou-se promissor em estudos in vitro contra L. (V.) braziliensis, devendo ser empregado em novas formulações e novos esquemas de tratamento experimental.(AU)

Transmission of Leishmania infantum from cats to dogs / Transmissão de Leishmania infantum de gatos para cães

Leishmania infantum infection in cats has been reported in several countries, including Brazil. However, the transmission of visceral leishmaniasis (VL) from cats to another host has not been proven yet. Therefore, the aim of this study was to verify the possibility of L. infantum transmission from cats to dogs. In order to verify the possibility of VL transmission from the cat to the dog, xenodiagnosis was carried out in a VL-positive cat, using 55 female Lutzomyia longipalpis. Five days later, 40 insects were dissected to verify Leishmania infection. The remaining 15 females were fed in a healthy dog. The potential infection of the dog was verified through clinical, serological, parasitological examinations, and PCR, at three, six, and twelve months post-infection. All 55 L. longipalpis females became visibly engorged. Leishmania promastigotes were detected in 27.5% of the dissected insects. Leishmania infection in the dog was confirmed upon first evaluation. DNA sequencing of the parasite isolated from the cat confirmed L. infantum infection and showed 99% similarity with the L. infantum DNA sequences from the dogs. Through this study, it was possible to confirm the L. infantum experimental transmission from a domestic cat to a domestic dog through its biological vector L. longipalpis.(AU)

A infecção por Leishmania infantum em gatos tem sido relatada em vários países, incluindo o Brasil. No entanto, a transmissão da leishmaniose visceral (LV) de gatos para outro hospedeiro ainda não foi comprovada. Portanto, o objetivo deste estudo foi verificar a possibilidade de transmissão de L. infantum de gatos para cães. Para verificar a possibilidade de transmissão da LV do gato para o cão, foi realizado xenodiagnóstico em um gato com LV, utilizando-se 55 fêmeas de Lutzomyia longipalpis. Cinco dias depois, 40 insetos foram dissecados para verificar a infecção por Leishmania. As 15 fêmeas restantes foram alimentadas em um cão saudável. A possível infecção no cão foi verificada por meio de exames clínicos, sorológicos, parasitológicos e PCR, três, seis e doze meses após a infecção. Todas as 55 fêmeas de L. longipalpis ficaram visivelmente ingurgitadas. Promastigotas de Leishmania foram detectadas em 27,5% dos insetos dissecados. A infecção por Leishmania no cão foi confirmada na primeira avaliação. O sequenciamento do DNA do parasito isolado do gato confirmou a infecção por L. infantum e apresentou 99% de similaridade com sequências de DNA de L. infantum de cães. Através deste estudo, foi possível confirmar a transmissão experimental de L. infantum de um gato doméstico para um cão doméstico através do seu vetor biológico L. longipalpis.(AU)

Testicular and seminal evaluation of dogs naturally infected with Leishmania sp / Avaliação testicular e seminal de cães naturalmente infectados por Leishmania sp

The objective of this study was to evaluate the testicular changes and detect the presence of Leishmania sp. in the testicles and semen of dogs with Visceral Leishmaniasis (VL). The animals were obtained from the Zoonoses Control Department of Teresina, PI, and taken to the kennel of the Agricultural Sciences Center of the Federal University of Piauí, where they remained were maintained for two months and subsequently euthanized for testicles removal. Semen samples were collected from 12 dogs, 06 positive and 06 negative for VL. The following diagnostic techniques readouts were assessed for the sampled animals: testosterone dosage, immunohistochemistry (IMH), histopathology of the slides containing the testicular material, and seminal evaluation by polymerase chain reaction (PCR). Testosterone values remained within the normal range for the canine specie and did not differ statistically among the experimental groups but displayed lower serum concentrations than those of the control group. All the testicular and semen samples from the dogs were negative for VL as determined by techniques IMH and PCR, respectively. The results of testicle histopathology revealed the presence of several lesions with statistical difference among the experimental groups. Parasitized dogs with VL have testicular lesions that may compromise the reproductive efficiency of these animals.(AU)

O objetivo deste estudo foi avaliar as alterações testiculares e detectar a presença de Leishmania sp. nos testículos e sêmen de cães com leishmaniose visceral (LV). Os animais foram obtidos da Gerência de Controle de Zoonoses de Teresina, PI, e levados ao canil do Centro de Ciências Agrárias da Universidade Federal do Piauí, onde permaneceram por dois meses e posteriormente foram submetidos à eutanásia para a remoção dos testículos. As amostras de sêmen foram coletadas de 12 cães, sendo 06 positivos e 06 negativos para LV. Nas amostras dos animais, foram realizadas as seguintes técnicas de diagnóstico: dosagem de testosterona, imunohistoquímica (IMH), histopatologia das lâminas contendo o material testicular e avaliação seminal por reação em cadeia pela polimerase (PCR). Valores de testosterona permaneceram dentro do intervalo normal para a espécie canina, não diferindo estatisticamente entre os grupos experimentais, porém apresentando concentrações séricas mais baixas do que o grupo controle. Todas as amostras testiculares e de sêmen dos cães foram negativas para LV, conforme determinado pelas técnicas IMH e PCR, respectivamente. Os resultados da histopatologia dos testículos revelaram a presença de várias lesões com diferença estatística entre os grupos experimentais. Os cães parasitados com LV apresentam lesões testiculares que podem comprometer a eficiência reprodutiva desses animais.(AU)

Histopathological alterations in the reproductive tract of male dogs with Canine Visceral Leishmaniasis / Alterações histopatológicas no trato reprodutivo de cães machos com Leishmaniose Visceral Canina

Dogs are considered the main domestic host of Leishmania infantum. The transmission between vertebrate hosts normally occurs during blood feeding of Lutzomyia sp., a genus of phlebotomine sand flies. However, other forms of transmission without a vector have been reported. Presence of amastigotes of Leishmania sp. in different organs can cause several pathologies depending on the immune status of the host and the parasite load. The aim of this study was to evaluate histopathological changes in the reproductive tract of male dogs. Samples of testes, epididymis, and prostate gland were collected from 70 animals that were grouped according to serology test results for visceral leishmaniasis and clinical symptoms. Three experimental groups were formed based on serological results and clinical presentation: asymptomatic, symptomatic and control. Prostatic changes were seen regardless of the serological results; however, the changes were more severe in symptomatic animals. In the testes and epididymis, lesions such as interstitial infiltrate, degeneration, and fibrosis were seen more frequently and were more severe in positive animals when compared to the control group. These results demonstrate that dogs infected with Leishmania sp. can develop nonspecific changes in the reproductive tract, which are more severe in symptomatic animals.(AU)

Os cães são considerados os principais hospedeiros domésticos de Leishmania infantum. A transmissão entre hospedeiros vertebrados normalmente ocorre durante o repasto sanguíneo de Lutzomyia sp., gênero de flebotomíneo. Entretanto, outras formas de transmissão sem a presença do vetor têm sido relatadas. A presença de amastigotas de Leishmania sp. em diferentes órgãos pode causar diversas patologias de acordo com o status imune do hospedeiro e a carga parasitária. O objetivo deste trabalho foi avaliar as alterações histopatológicas no trato reprodutivo de cães machos. Amostras de testículo, epidídimo e próstata foram coletadas de 70 animais, os quais foram agrupados conforme o resultado da sorologia para leishmaniose visceral e os sinais clínicos. Três grupos experimentais foram formados baseados nos resultados sorológicos e manifestações clínicas: assintomático, sintomático e controle. Alterações prostáticas foram observadas independentemente dos resultados sorológicos; entretanto, foram mais acentuadas nos animais sintomáticos. Nos testículos e epidídimos, lesões como infiltrado intersticial, degeneração e fibrose foram observadas mais frequentemente e de forma mais severa nos animais positivos quando comparados ao grupo controle. Estes resultados demonstram que cães infectados com Leishmania sp. podem desenvolver alterações não específicas no trato reprodutivo, as quais são...(AU)

O probiótico Saccharomyces boulardii modula a resposta de IgG2a em camundongos expostos aos antígenos de Leishmania infantum / Probiotic Saccharomyces boulardii modulates IgG2a response in mice exposed to Leishmania infantum antigens

O objetivo do presente estudo foiavaliar os efeitos do probiótico Saccharomyces boulardii na modulação da resposta imune humoral de animais expostos a antígenos de Leishmania infantum. Para isso, 16 camundongos BALB/c foram imunizados com antígeno particulado de Leishmania infantum e divididos em dois grupos experimentais, um composto por animais suplementados e outro por animais não suplementados com o probiótico. Amostras de sangue dos animais foram colhidas semanalmente durante o período experimental e submetidas ao Ensaio da Imunoabsorbância Ligado à Enzima indireto para avaliação dos títulos de IgG totais e o perfil dos isotipos de IgG produzidos (IgG1 e IgG2a). A suplementação com o probiótico não exacerbou a produção de IgG total em comparação ao grupo controle, não havendo diferenças significativas entre os dois grupos. Porém, as soroconversões de IgG2a foram mais elevadas no grupo suplementado, no qual registrou-se um aumento de 1,46 vezes no final do experimento. Assim,a suplementação com S. boulardii foi capaz de modular a resposta de IgG2a/IgG1 nos animais expostos aos antígenos de Leishmania infantum.

The aim of the present study was to evaluate the effects of the Saccharomyces boulardii probiotic on the modulation of humoral immune response in animals exposed to Leishmania infantum antigens. For this, 16 BALB/c mice were immunized with Leishmania infantum particulate antigen and divided into two experimental groups, one consisting of supplemented animals and the other not probiotic supplemented animals. Blood samples from the animals were taken weekly during the experimental period and subjected to the Indirect Enzyme-Linked Immunosorbance Assay for evaluation of total IgG titers and the profile of the produced IgG isotypes (IgG1 and IgG2a). Probiotic supplementation did not exacerbate total IgG production compared to the control group, with no significant differences between the two groups. However, IgG2a seroconversions were higher in the supplemented group, which showed a 1.46-fold increase at the end of the experiment. Thus, supplementation with S. boulardii was able to modulate the IgG2a/IgG1 response in animals exposed to Leishmania infantum antigens.

O probiótico Saccharomyces boulardii modula a resposta de IgG2a em camundongos expostos aos antígenos de Leishmania infantum / Probiotic Saccharomyces boulardii modulates IgG2a response in miceexposed to Leishmania infantum antigens

O objetivo do presente estudo foi avaliar os efeitos do probiótico Saccharomyces boulardii na modulação da resposta imune humoral de animais expostos a antígenos de Leishmania infantum. Para isso, 16 camundongos BALB/c foram imunizados com antígeno particulado de Leishmania infantum e divididos em dois grupos experimentais, um composto por animais suplementados e outro por animais não suplementados com o probiótico. Amostras de sangue dos animais foram colhidas semanalmente durante o período experimental e submetidas ao Ensaio da Imunoabsorbância Ligado à Enzima indireto para avaliação dos títulos de IgG totais e o perfil dos isotipos de IgG produzidos (IgG1 e IgG2a). A suplementação com o probiótico não exacerbou a produção de IgG total em comparação ao grupo controle, não havendo diferenças significativas entre os dois grupos. Porém, as soroconversões de IgG2a foram mais elevadas no grupo suplementado, no qual registrou-se um aumento de 1,46 vezes no final do experimento. Assim, a suplementação com S. boulardii foi capaz de modular a resposta de IgG2a/IgG1 nos animais expostos aos antígenos de Leishmania infantum.(AU)

The aim of the present study was to evaluate the effects of the Saccharomyces boulardii probiotic on the modulation of humoral immune response in animals exposed to Leishmania infantum antigens. For this, 16 BALB/c mice were immunized with Leishmania infantum particulate antigen and divided into two experimental groups, one consisting of supplemented animals and the other not probiotic supplemented animals. Blood samples from the animals were taken weekly during the experimental period and subjected to the Indirect Enzyme-Linked Immunosorbance Assay for evaluation of total IgG titers and the profile of the produced IgG isotypes (IgG1 and IgG2a). Probiotic supplementation did not exacerbate total IgG production compared to the control group, with no significant differences between the two groups. However, IgG2a seroconversions were higher in the supplemented group, which showed a 1.46-fold increase at the end of the experiment. Thus, supplementation with S. boulardii was able to modulate the IgG2a/IgG1 response in animals exposed to Leishmania infantum antigens.(AU)

O probiótico Saccharomyces boulardii modula a resposta de IgG2a em camundongos expostos aos antígenos de Leishmania infantum / Probiotic Saccharomyces boulardii modulates IgG2a response in miceexposed to Leishmania infantum antigens

O objetivo do presente estudo foi avaliar os efeitos do probiótico Saccharomyces boulardii na modulação da resposta imune humoral de animais expostos a antígenos de Leishmania infantum. Para isso, 16 camundongos BALB/c foram imunizados com antígeno particulado de Leishmania infantum e divididos em dois grupos experimentais, um composto por animais suplementados e outro por animais não suplementados com o probiótico. Amostras de sangue dos animais foram colhidas semanalmente durante o período experimental e submetidas ao Ensaio da Imunoabsorbância Ligado à Enzima indireto para avaliação dos títulos de IgG totais e o perfil dos isotipos de IgG produzidos (IgG1 e IgG2a). A suplementação com o probiótico não exacerbou a produção de IgG total em comparação ao grupo controle, não havendo diferenças significativas entre os dois grupos. Porém, as soroconversões de IgG2a foram mais elevadas no grupo suplementado, no qual registrou-se um aumento de 1,46 vezes no final do experimento. Assim, a suplementação com S. boulardii foi capaz de modular a resposta de IgG2a/IgG1 nos animais expostos aos antígenos de Leishmania infantum.

The aim of the present study was to evaluate the effects of the Saccharomyces boulardii probiotic on the modulation of humoral immune response in animals exposed to Leishmania infantum antigens. For this, 16 BALB/c mice were immunized with Leishmania infantum particulate antigen and divided into two experimental groups, one consisting of supplemented animals and the other not probiotic supplemented animals. Blood samples from the animals were taken weekly during the experimental period and subjected to the Indirect Enzyme-Linked Immunosorbance Assay for evaluation of total IgG titers and the profile of the produced IgG isotypes (IgG1 and IgG2a). Probiotic supplementation did not exacerbate total IgG production compared to the control group, with no significant differences between the two groups. However, IgG2a seroconversions were higher in the supplemented group, which showed a 1.46-fold increase at the end of the experiment. Thus, supplementation with S. boulardii was able to modulate the IgG2a/IgG1 response in animals exposed to Leishmania infantum antigens.

Second-generation pterocarpanquinones: synthesis and antileishmanial activity

Background: Despite the development of new therapies for leishmaniasis, among the 200 countries or territories reporting to the WHO, 87 were identified as endemic for Tegumentary Leishmaniasis and 75 as endemic for Visceral Leishmaniasis. The identification of antileishmanial drug candidates is essential to fill the drug discovery pipeline for leishmaniasis. In the hit molecule LQB-118 selected, the first generation of pterocarpanquinones was effective and safe against experimental visceral and cutaneous leishmaniasis via oral delivery. In this paper, we report the synthesis and antileishmanial activity of the second generation of pterocarpanoquinones. Methods: The second generation of pterocarpanquinones 2a-f was prepared through a palladium-catalyzed oxyarylation of dihydronaphtalen and chromens with iodolawsone, easily prepared by iodination of lawsone. The spectrum of antileishmanial activity was evaluated in promastigotes and intracellular amastigotes of L. amazonensis, L. braziliensis, and L. infantum. Toxicity was assessed in peritoneal macrophages and selective index calculated by CC50/IC50. Oxidative stress was measured by intracellular ROS levels and mitochondrial membrane potential in treated cells. Results: In this work, we answered two pertinent questions about the structure of the first-generation pterocarpanquinones: the configuration and positions of rings B (pyran) and C (furan) and the presence of oxygen in the B ring. When rings B and C are exchanged, we noted an improvement of the activity against promastigotes and amastigotes of L. amazonensis and promastigotes of L. infantum. As to the oxygen in ring B of the new generation, we observed that the oxygenated compound 2b is approximately twice as active against L. braziliensis promastigotes than its deoxy derivative 2a. Another modification that improved the activity was the addition of the methylenedioxy group... (AU)

4-Phenyl-1, 3-thiazole-2-amines as scaffolds for new antileishmanial agents

Background:There is still a need for new alternatives in pharmacological therapy for neglected diseases, as the drugs available show high toxicity and parenteral administration. That is the case for the treatment of leishmaniasis, particularly to the cutaneous clinical form of the disease. In this study, we present the synthesis and biological screening of eight 4-phenyl-1,3-thiazol-2-amines assayed against Leishmania amazonensis. Herein we propose that these compounds are good starting points for the search of new antileishmanial drugs by demonstrating some of the structural aspects which could interfere with the observed activity, as well as suggesting potential macromolecular targets.Methods:The compounds were easily synthesized by the methodology of Hantzsch and Weber, had their purities determined by Gas Chromatography-Mass spectrometry and assayed against the promastigote forms of Leishmania amazonensis as well as against two white cell lines (L929 and THP-1) and the monkey's kidney Vero cells. PrestoBlue® and MTT viability assays were the methodologies applied to measure the antileishmanial and cytotoxic activities, respectively. A molecular modeling target fishing study was performed aiming to propose potential macromolecular targets which could explain the observed biological behavior.Results:Four out of the eight compounds tested exhibited important anti-promastigote activity associated with good selectivity indexes when considering Vero cells. For the most promising compound, compound 6, IC50 against promastigotes was 20.78 while SI was 5.69. Compounds 3 (IC50: 46.63 μM; SI: 26.11) and 4 (IC50: 53.12 μM; SI: 4.80) also presented important biological behavior. A target fishing study suggested that S-methyl-5-thioadenosine phosphorylase is a potential target to these compounds, which could be explored to enhance activity and decrease the potential toxic side effects....(AU)

Second-generation pterocarpanquinones: synthesis and antileishmanial activity

Despite the development of new therapies for leishmaniasis, among the 200 countries or territories reporting to the WHO, 87 were identified as endemic for Tegumentary Leishmaniasis and 75 as endemic for Visceral Leishmaniasis. The identification of antileishmanial drug candidates is essential to fill the drug discovery pipeline for leishmaniasis. In the hit molecule LQB-118 selected, the first generation of pterocarpanquinones was effective and safe against experimental visceral and cutaneous leishmaniasis via oral delivery. In this paper, we report the synthesis and antileishmanial activity of the second generation of pterocarpanoquinones. Methods: The second generation of pterocarpanquinones 2a-f was prepared through a palladium-catalyzed oxyarylation of dihydronaphtalen and chromens with iodolawsone, easily prepared by iodination of lawsone. The spectrum of antileishmanial activity was evaluated in promastigotes and intracellular amastigotes of L. amazonensis, L. braziliensis, and L. infantum. Toxicity was assessed in peritoneal macrophages and selective index calculated by CC50/IC50. Oxidative stress was measured by intracellular ROS levels and mitochondrial membrane potential in treated cells. Results: In this work, we answered two pertinent questions about the structure of the first-generation pterocarpanquinones: the configuration and positions of rings B (pyran) and C (furan) and the presence of oxygen in the B ring. When rings B and C are exchanged, we noted an improvement of the activity against promastigotes and amastigotes of L. amazonensis and promastigotes of L. infantum. As to the oxygen in ring B of the new generation, we observed that the oxygenated compound 2b is approximately twice as active against L. braziliensis promastigotes than its deoxy derivative 2a. Another modification that improved the activity was the addition of the methylenedioxy group. A variation in the susceptibility among species was evident in the clinically relevant form of the parasite, the intracellular amastigote. L. amazonensis was the species most susceptible to novel derivatives, whilst L. infantum was resistant to most of them. The pterocarpanoquinones (2b and 2c) that possess the oxygen atom in ring B showed induction of increased ROS production. Conclusions: The data presented indicate that the pterocarpanoquinones are promising compounds for the development of new leishmanicidal agents.(AU)

Activity of the antiarrhythmic drug amiodarone against Leishmania (L) infantum: an in vitro and in vivo approach

Background:Considering the high toxicity and limited therapies available for treating visceral leishmaniasis (VL), the drug repositioning approach represents a faster way to deliver new therapies to the market.Methods:In this study, we described for the first time the activity of a potent antiarrhythmic, amiodarone (AMD), against L. (L.)infantum and its in vitro and in vivo activity.Results:The evaluation against promastigotes has shown that amiodarone presents leishmanicidal effect against the extracellular form, with an IC50 value of 10 μM. The activity was even greater against amastigotes in comparison with promastigotes with an IC50 value of 0.5 μM. The selectivity index in relation to the intracellular form demonstrated that the antiparasitic activity was approximately 56 times higher than its toxicity to mammalian cells. Investigation of the in vivo AMD activity in the L. infantum-infected hamster model showed that 51 days after the initial infection, amiodarone was unable to reduce the parasite burden in the spleen and liver when treated for 10 consecutive days, intraperitoneally, at 50 mg/kg/day, as determined by qPCR. Although not statistically significant, AMD was able to reduce the parasite burden by 20% in the liver when treated for 10 consecutive days, orally, at 100 mg/kg/day; no reduction in the spleen was found by qPCR.Conclusions:Our findings may help further drug design studies seeking new AMD derivatives that may provide new candidates with an in vitro selectivity close to or even greater than that observed in the prototype delivering effectiveness in the experimental model of VL.(AU)

Activity of the antiarrhythmic drug amiodarone against Leishmania (L) infantum: an in vitro and in vivo approach

Considering the high toxicity and limited therapies available for treating visceral leishmaniasis (VL), the drug repositioning approach represents a faster way to deliver new therapies to the market. Methods: In this study, we described for the first time the activity of a potent antiarrhythmic, amiodarone (AMD), against L. (L.)infantum and its in vitro and in vivo activity. Results: The evaluation against promastigotes has shown that amiodarone presents leishmanicidal effect against the extracellular form, with an IC50 value of 10 μM. The activity was even greater against amastigotes in comparison with promastigotes with an IC50 value of 0.5 μM. The selectivity index in relation to the intracellular form demonstrated that the antiparasitic activity was approximately 56 times higher than its toxicity to mammalian cells. Investigation of the in vivo AMD activity in the L. infantum-infected hamster model showed that 51 days after the initial infection, amiodarone was unable to reduce the parasite burden in the spleen and liver when treated for 10 consecutive days, intraperitoneally, at 50 mg/kg/day, as determined by qPCR. Although not statistically significant, AMD was able to reduce the parasite burden by 20% in the liver when treated for 10 consecutive days, orally, at 100 mg/kg/day; no reduction in the spleen was found by qPCR. Conclusions: Our findings may help further drug design studies seeking new AMD derivatives that may provide new candidates with an in vitro selectivity close to or even greater than that observed in the prototype delivering effectiveness in the experimental model of VL.(AU)

Topical treatment of experimental cutaneous leishmaniasis in golden hamster (Mesocricetus auratus) with formulations containing pentamidine / Tratamento tópico da leishmaniose cutânea experimental em golden hamsters (Mesocricetus auratus) com fórmulas contendo pentamidina

Current treatment of cutaneous leishmaniasis (CL) relies mainly on pentavalent antimonials salts and second-line drugs include pentamidine and amphotericin B, but these therapies have side effects and require parenteral administration. The aim of this work was to evaluate the topical formulations containing pentamidine isethionate (PI) in the experimental treatment of cutaneous leishmaniasis (CL). Golden hamsters (Mesocricetus auratus) were infected in the nose with Leishmania (Leishmania) amazonensis. Six treatment groups received different topical treatments of anhydrous or hydrating emulsions, for a maximum of 10 days, with an application of 50 mg day-1. After treatment tissue samples of lesions were evaluated by histology, transmission electron microscopy and biopsy cultivation. Compared with untreated group, topical treatment with hydrating emulsion with 10% PI and usnic acid (ACE5AU) showed significantly decrease in volume lesion (P= 0.028) on 20th day after the end of the treatment with reduction of 27.37%. Topical treatment with anhydrous emulsion with 10% PI and usnic acid (ACPU) reduces parasite burden in Golden hamsters. This study demonstrated the potential of topical treatment to reduce the number of parasites that could be combined with others drugs and to have a faster and more effective treatment of cutaneous leishmaniasis.(AU)

O tratamento atual para Leishmaniose Tegumentar Americana é realizado com antimoniais pentavalentes como tratamento padrão, e as drogas de segunda escolha incluem pentamidina e anfotericina B, mas essas terapias apresentam efeitos colaterais e requerem administração parenteral. O objetivo deste trabalho foi avaliar formulações tópicas contendo isetionato de pentamidina comercial (IP) no tratamento da leishmaniose cutânea experimental (LC). Hamsters dourados foram infectados no focinho com Leishmania (Leishmania) amazonensis. Grupos de seis animais receberam tratamento tópico com cremes anidro ou emulsões hidratantes, num máximo de 10 dias, com aplicação de 50 mg dia-1. Amostras de tecido de lesões tratadas foram avaliadas por histologia, microscopia eletrônica de transmissão (MET) e cultivo de biópsia. Comparado ao grupo sem tratamento, o tratamento tópico com emulsão hidratante com 10% de IP e ácido úsnico (ACE5AU) mostrou diminuição significativa (P=0,028) nas medidas de lesões, 20 dias após o final do tratamento e 27,37% de redução. O tratamento tópico com emulsão anidra com 10% de IP e ácido úsnico (ACPU) mostrou redução da carga parasitária em Golden hamsters. Este estudo demonstrou a possibilidade de utilizar o tratamento tópico para reduzir o número de parasitas e que este poderia ser associado a outras drogas para tratamento mais rápido e eficaz da leishmaniose cutânea.(AU)

Antileishmanial activity of extracts from Libidibia ferrea: development of in vitro and in vivo tests / Atividade anti-leishmania de extratos de Libidibia ferrea: desenvolvimento de testes in vitro e in vivo

ABSTRACT Treatment of cutaneous leishmaniasis (CL) is difficult due to the scarce number of drugs able to eliminate completely the intracellular form of the parasite. In the present study, the aim was to evaluate: i) phytochemical properties of extracts from Libidibia ferrea; ii) antileishmanial activity of extracts from L. ferrea against promastigotes and amastigotes of Leishmania (Leishmania) amazonensis and Leishmania (Viannia) guyanensis; iii) the effects of topical treatment using hydrogel containing active extract of L. ferrea on golden hamsters infected with L. (L.) amazonensis. Extracts from leaves, branches and fruits of L. ferrea were obtained with hexane and methanol and were tested by in vitro assays in promastigotes and murine macrophages J774 experimentally infected with amastigotes of Leishmania. Groups of hamsters with CL received topical treatment with a formulation of extract (10%) hydrogels, 50 mg.day-1 for 40 days. In vitro activity of FrMeOH (methanolic extract from fruits without seeds) resulted in significant reduction of viable promastigotes of L. (L.) amazonensis (IC50 of 15.4 µg.mL-1) and demonstrated inhibition potential of amastigote forms of L. (L.) amazonensis and L. (V.) guyanensis and low cytotoxicity in macrophages. The overall data of topical treatment with extract hydrogels (GelFrMeOH) showed that lesion sizes were significantly reduced (42.78%), with low parasite burden by RT-qPCR and culture analysis by microscopy examination, and with histopathological findings such as lower inflammatory cell infiltration 40 days after treatment. Chemical analysis demonstrated FrMeOH contains high levels of phenolic compounds. The results indicate a possible alternative therapy for CL using phytotherapics.(AU)

RESUMO O tratamento da leishmaniose cutânea (LC) é de difícil resultado, devido ao escasso número de fármacos capazes de eliminar completamente a forma intracelular do parasita. No presente estudo, objetivou-se avaliar: i) propriedades fitoquímicas dos extratos de Libidibia ferrea; ii) a atividade antileishmania de extratos de L. ferrea contra promastigotas e amastigotas de Leishmania (Leishmania) amazonensis e Leishmania (Viannia) guyanensis; iii) os efeitos do tratamento tópico utilizando hidrogel contendo extrato ativo de L. ferrea em hamsters dourados (Mesocricetus auratus) infectados com L. (L.) amazonensis. Extratos de folhas, galhos e frutos de L. ferrea foram obtidos com hexano e metanol e foram testados por ensaios in vitro contra promastigotas e macrófagos J774 infectados com amastigotas de Leishmania. Grupos de hamsters infectados receberam hidrogel tópico com extrato (10%), 50 mg.dia-1 durante 40 dias. A atividade in vitro de FrMeOH (extrato metanólico dos frutos) mostrou redução significativa de promastigotas de L. (L.) amazonensis (IC50 de 15,4 g.mL-1), potencial de inibição de formas amastigotas de L. (L.) amazonensis e L. (V.) guyanensis (IC50 303,36 g.mL-1) e baixa citotoxicidade em células de macrófagos J774. Os resultados destacaram que as lesões cutâneas de animais que receberam tratamento com hidrogel + extrato (GelFrMeOH) apresentaram redução significativa (42,78%), menos úlceração e redução da carga parasitária detectada por RT-qPCR, microscopia e análise em cultura e alterações histopatológicas, como menor infiltrado inflamatório após 40 dias de tratamento. A análise química demonstrou que FrMeOH contém altos níveis de compostos fenólicos. Os resultados apontam para uma possível terapia alternativa para CL utilizando fitoterápicos.(AU)

New human case reports of cutaneous leishmaniasis by Leishmania (Viannia) naiffi in the Amazon region, Brazil / Novos relatos de casos de leishmaniose cutânea por Leishmania (Viannia) naiffi na região amazônica, Brasil

Few cases of human cutaneous leishmaniasis (CL) caused by Leishmania naiffi were described in the medical literature. The aim of this study was to report and analyze new cases of L. naiffi in the period between the years 1992 to 2011. The strains were characterized by isoenzyme analysis. All patients assisted had small lesions; ranging from 1.0x1.0 mm and 13.5x11.5 mm. The lesions observed were widely distributed: 55.5% on the lower limb, 5.5% in the abdominal area, 16.6% on upper limb and 22.2% in upper limb and back. Seventy-two percent of patients had ulcerated lesions. Clinical course of the disease varied from 1 to 10 months. According to gender, most infected individuals were men (83.3%). The patients came from Amazonas (10), Pará (01) and Rondônia (01), north States of Brazil. Five patients were infected in experimental stations of the National Institute of Amazonian Research (INPA). Although the results of this study were similar to other reports in the literature, some of the patients had more of one skin lesion. It is also reported the first case of CL caused by L. naiffi in the State of Rondônia and identified an area of disease transmission in the experimental station of INPA.(AU)

Poucos casos de leishmaniose cutânea humana (LC) causada por Leishmania naiffi foram descritos na literatura médica. Assim, o objetivo deste estudo foi relatar e analisar novos casos de infecção por L. naiffi no período compreendido entre os anos de 1992 a 2011. As cepas foram caracterizadas por análise de isoenzimas. Todos os pacientes atendidos apresentavam lesões pequenas; variando entre 1,0x1,0 mm e 13,5x11,5 mm. As lesões observadas foram amplamente distribuídas: 55,5% no membro inferior, 5,5% na área abdominal, 16,6% no membro superior e 22,2% no membro superior e para trás. Setenta e dois por cento dos pacientes tiveram lesões ulceradas. O curso clínico da doença variou de 1 a 10 meses. De acordo com o sexo, a maioria dos indivíduos infectados eram homens (83,3%). Os pacientes vieram de Amazonas (10), Pará (01) e Rondônia (01), estados do norte do Brasil. Cinco pacientes foram infectados em estações experimentais do Instituto Nacional de Pesquisas da Amazônia (INPA). Embora os resultados encontrados neste estudo fossem semelhantes às outras descrições na literatura, alguns dos pacientes apresentavam mais de uma lesão cutânea. Também é relatado o primeiro caso de LC causada por L. naiffi no Estado de Rondônia e identificada uma área de transmissão da doença na estação experimental do INPA.(AU)

New human case reports of cutaneous leishmaniasis by Leishmania (Viannia) naiffi in the Amazon region, Brazil / Novos relatos de casos de leishmaniose cutânea por Leishmania (Viannia) naiffi na região amazônica, Brasil

Few cases of human cutaneous leishmaniasis (CL) caused by Leishmania naiffi were described in the medical literature. The aim of this study was to report and analyze new cases of L. naiffi in the period between the years 1992 to 2011. The strains were characterized by isoenzyme analysis. All patients assisted had small lesions; ranging from 1.0x1.0 mm and 13.5x11.5 mm. The lesions observed were widely distributed: 55.5% on the lower limb, 5.5% in the abdominal area, 16.6% on upper limb and 22.2% in upper limb and back. Seventy-two percent of patients had ulcerated lesions. Clinical course of the disease varied from 1 to 10 months. According to gender, most infected individuals were men (83.3%). The patients came from Amazonas (10), Pará (01) and Rondônia (01), north States of Brazil. Five patients were infected in experimental stations of the National Institute of Amazonian Research (INPA). Although the results of this study were similar to other reports in the literature, some of the patients had more of one skin lesion. It is also reported the first case of CL caused by L. naiffi in the State of Rondônia and identified an area of disease transmission in the experimental station of INPA.

Poucos casos de leishmaniose cutânea humana (LC) causada por Leishmania naiffi foram descritos na literatura médica. Assim, o objetivo deste estudo foi relatar e analisar novos casos de infecção por L. naiffi no período compreendido entre os anos de 1992 a 2011. As cepas foram caracterizadas por análise de isoenzimas. Todos os pacientes atendidos apresentavam lesões pequenas; variando entre 1,0x1,0 mm e 13,5x11,5 mm. As lesões observadas foram amplamente distribuídas: 55,5% no membro inferior, 5,5% na área abdominal, 16,6% no membro superior e 22,2% no membro superior e para trás. Setenta e dois por cento dos pacientes tiveram lesões ulceradas. O curso clínico da doença variou de 1 a 10 meses. De acordo com o sexo, a maioria dos indivíduos infectados eram homens (83,3%). Os pacientes vieram de Amazonas (10), Pará (01) e Rondônia (01), estados do norte do Brasil. Cinco pacientes foram infectados em estações experimentais do Instituto Nacional de Pesquisas da Amazônia (INPA). Embora os resultados encontrados neste estudo fossem semelhantes às outras descrições na literatura, alguns dos pacientes apresentavam mais de uma lesão cutânea. Também é relatado o primeiro caso de LC causada por L. naiffi no Estado de Rondônia e identificada uma área de transmissão da doença na estação experimental do INPA.

Histopathological assessment of the liver during experimental infection with Leishmania (Leishmania) chagasi in immunosuppressed BALB/c mice / Avaliação histopatológica do fígado durante a infecção experimental por Leishmania chagasi (Leishmania) em camundongos BALB / c imunossuprimidos / Evaluación histopatológico del hígado durante la infección experimental por Leishmania chagasi (Leishmania) en ratones BALB / c con inmunosupresión

BALB/c mice were experimentally infected with L. (L.) chagasi and immunosuppressed for the study of the histopathological changes present in their liver. The 96 studied mice were subdivided into four groups: control (I) no treatment; immunosuppressed (II) treatment with dexamethasone (DXM) and pentoxifylline (PTX); infected (III) infection with L. (L.) chagasi; and infected and immunosuppressed (IV) - infection with L. (L.) chagasi and treatment with DXM and PTX. The day of infection was considered the day zero and immunosuppression occurred at 60 days P.I. Samples were obtained at distinct moments, 15, 30, 60, 75 and 90 days P.I., by collecting liver fragments for histopathological assessment. There was progressive and constant inflammatory reaction of inflammatory infiltrate and complete granulomas, which was renewed in all observation phases. For inoculated groups, the presence of amastigotes was confirmed by means of immunohistochemistry. The immunosuppressed group showed weak inflammatory reaction and rare incomplete granulomas. Immunosuppression influenced the immune response of the liver, which is capable of controlling murine visceral leishmaniasis.(AU)

Camundongos BALB / c foram infectados experimentalmente com L. (L.) chagasi e imunossuprimidos para estudar as alterações histopatológicas no fígado presente. Os 96 ratos estudados foram divididos em quatro grupos: controle (I) - sem tratamento; imunossuprimidos (II) - O tratamento com dexametasona (DXM) e pentoxifilina (PTX); infectado (III) - infecção por L. chagasi (L.); e infectados e imunodeprimidos (IV) - infecção por L. chagasi (L.) e tratamento com DXM e PTX. O dia da infecção foi considerado o dia zero e imunossupressão ocorreu 60 dias P.I. As amostras foram obtidas em vários momentos, 15, 30, 60, 75 e 90 dias P.I., por coleta de fragmentos de fígado para avaliação histopatológica. Houve constante reação inflamatória e infiltrado inflamatório progressivo e granulomas completos, que foi renovado em todas as fases de observação. Para os grupos inoculados, a presença de amastigotas foi confirmada por imuno-histoquímica. O grupo de imunodeprimidos apresentou uma resposta inflamatória fraca e granulomas raros e incompletos. Imunossupressão influenciou a resposta imune do fígado, que é capaz de controlar a leishmaniose visceral murina.(AU)

Ratones BALB / c se infectaron experimentalmente con L. chagasi (L.) y inmunosuprimidos para el estudio de los cambios histopatológicos presentes en su hígado. Los 96 ratones estudiados se dividieron en cuatro grupos: control (I) - sin tratamiento; inmunosuprimidos (II) - El tratamiento con dexametasona (DXM) y pentoxifilina (PTX); infectada (III) - la infección con L. chagasi (L.); y infectada y inmunosuprimidos (IV) - infección con L. chagasi (L.) y el tratamiento con DXM y PTX. El día de la infección fue considerado como el día cero y la inmunosupresión se produjo a 60 días P.I. Las muestras se obtuvieron en momentos distintos, 15, 30, 60, 75 y 90 días P.I., mediante la recopilación de fragmentos de hígado para la evaluación histopatológica. Hubo reacción inflamatoria progresiva y constante de infiltrado inflamatorio y granulomas completos, que fue renovado en todas las fases de observación. Para los grupos inoculados, la presencia de amastigotes se confirmó por medio de inmunohistoquímica. El grupo inmunosuprimidos mostró una reacción inflamatoria débil y granulomas incompletas raras. Inmunosupresión influyó en la respuesta inmune del hígado, que es capaz de controlar la leishmaniosis visceral murino.(AU)

Histopathological assessment of the liver during experimental infection with Leishmania (Leishmania) chagasi in immunosuppressed BALB/c mice / Avaliação histopatológica do fígado durante a infecção experimental por Leishmania chagasi (Leishmania) em camundongos BALB / c imunossuprimidos / Evaluación histopatológico del hígado durante la infección experimental por Leishmania chagasi (Leishmania) en ratones BALB / c con inmunosupresión

BALB/c mice were experimentally infected with L. (L.) chagasi and immunosuppressed for the study of the histopathological changes present in their liver. The 96 studied mice were subdivided into four groups: control (I) – no treatment; immunosuppressed (II) – treatment with dexamethasone (DXM) and pentoxifylline (PTX); infected (III) – infection with L. (L.) chagasi; and infected and immunosuppressed (IV) - infection with L. (L.) chagasi and treatment with DXM and PTX. The day of infection was considered the day zero and immunosuppression occurred at 60 days P.I. Samples were obtained at distinct moments, 15, 30, 60, 75 and 90 days P.I., by collecting liver fragments for histopathological assessment. There was progressive and constant inflammatory reaction of inflammatory infiltrate and complete granulomas, which was renewed in all observation phases. For inoculated groups, the presence of amastigotes was confirmed by means of immunohistochemistry. The immunosuppressed group showed weak inflammatory reaction and rare incomplete granulomas. Immunosuppression influenced the immune response of the liver, which is capable of controlling murine visceral leishmaniasis.

Camundongos BALB / c foram infectados experimentalmente com L. (L.) chagasi e imunossuprimidos para estudar as alterações histopatológicas no fígado presente. Os 96 ratos estudados foram divididos em quatro grupos: controle (I) - sem tratamento; imunossuprimidos (II) - O tratamento com dexametasona (DXM) e pentoxifilina (PTX); infectado (III) - infecção por L. chagasi (L.); e infectados e imunodeprimidos (IV) - infecção por L. chagasi (L.) e tratamento com DXM e PTX. O dia da infecção foi considerado o dia zero e imunossupressão ocorreu 60 dias P.I. As amostras foram obtidas em vários momentos, 15, 30, 60, 75 e 90 dias P.I., por coleta de fragmentos de fígado para avaliação histopatológica. Houve constante reação inflamatória e infiltrado inflamatório progressivo e granulomas completos, que foi renovado em todas as fases de observação. Para os grupos inoculados, a presença de amastigotas foi confirmada por imuno-histoquímica. O grupo de imunodeprimidos apresentou uma resposta inflamatória fraca e granulomas raros e incompletos. Imunossupressão influenciou a resposta imune do fígado, que é capaz de controlar a leishmaniose visceral murina.

Ratones BALB / c se infectaron experimentalmente con L. chagasi (L.) y inmunosuprimidos para el estudio de los cambios histopatológicos presentes en su hígado. Los 96 ratones estudiados se dividieron en cuatro grupos: control (I) - sin tratamiento; inmunosuprimidos (II) - El tratamiento con dexametasona (DXM) y pentoxifilina (PTX); infectada (III) - la infección con L. chagasi (L.); y infectada y inmunosuprimidos (IV) - infección con L. chagasi (L.) y el tratamiento con DXM y PTX. El día de la infección fue considerado como el día cero y la inmunosupresión se produjo a 60 días P.I. Las muestras se obtuvieron en momentos distintos, 15, 30, 60, 75 y 90 días P.I., mediante la recopilación de fragmentos de hígado para la evaluación histopatológica. Hubo reacción inflamatoria progresiva y constante de infiltrado inflamatorio y granulomas completos, que fue renovado en todas las fases de observación. Para los grupos inoculados, la presencia de amastigotes se confirmó por medio de inmunohistoquímica. El grupo inmunosuprimidos mostró una reacción inflamatoria débil y granulomas incompletas raras. Inmunosupresión influyó en la respuesta inmune del hígado, que es capaz de controlar la leishmaniosis visceral murino.