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Vivian T Martins,
Miguel A Chávez-Fumagalli,
Lourena E Costa,
Adriana M C Canavaci,
Paula S Lage,
Daniela P Lage,
Mariana C Duarte, Diogo G Valadares,
Rubens D M Magalhães,
Tatiana G Ribeiro,
Ronaldo A P Nagem,
Wanderson D Darocha,
Wiliam C B Régis,
Manuel Soto,
Eduardo A F Coelho,
Ana Paula Fernandes,
Carlos A P Tavares
PLoS Neglected Tropical Diseases 04/2013; 7(4). · 4.69 Impact Factor
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Vivian T Martins,
Miguel A Chávez-Fumagalli,
Lourena E Costa,
Adriana M C C Martins,
Paula S Lage,
Daniela P Lage,
Mariana C Duarte, Diogo G Valadares,
Rubens D M Magalhães,
Tatiana G Ribeiro,
Ronaldo A P Nagem,
Wanderson D Darocha,
Wiliam C B Régis,
Manuel Soto,
Eduardo A F Coelho,
Ana Paula Fernandes,
Carlos A P Tavares
[show abstract]
[hide abstract]
ABSTRACT: BACKGROUND: The present study aimed to evaluate a hypothetical Leishmania amastigote-specific protein (LiHyp1), previously identified by an immunoproteomic approach performed in Leishmania infantum, which showed homology to the super-oxygenase gene family, attempting to select a new candidate antigen for specific serodiagnosis, as well as to compose a vaccine against VL. METHODOLOGYPRINCIPAL FINDINGS: The LiHyp1 DNA sequence was cloned; the recombinant protein (rLiHyp1) was purified and evaluated for its antigenicity and immunogenicity. The rLiHyp1 protein was recognized by antibodies from sera of asymptomatic and symptomatic animals with canine visceral leishmaniasis (CVL), but presented no cross-reactivity with sera of dogs vaccinated with Leish-Tec, a Brazilian commercial vaccine; with Chagas' disease or healthy animals. In addition, the immunogenicity and protective efficacy of rLiHyp1 plus saponin was evaluated in BALB/c mice challenged subcutaneously with virulent L. infantum promastigotes. rLiHyp1 plus saponin vaccinated mice showed a high and specific production of IFN-γ, IL-12, and GM-CSF after in vitro stimulation with the recombinant protein. Immunized and infected mice, as compared to the control groups (saline and saponin), showed significant reductions in the number of parasites found in the liver, spleen, bone marrow, and in the paws' draining lymph nodes. Protection was associated with an IL-12-dependent production of IFN-γ, produced mainly by CD4 T cells. In these mice, a decrease in the parasite-mediated IL-4 and IL-10 response could also be observed. CONCLUSIONSSIGNIFICANCE: The present study showed that this Leishmania oxygenase amastigote-specific protein can be used for a more sensitive and specific serodiagnosis of asymptomatic and symptomatic CVL and, when combined with a Th1-type adjuvant, can also be employ as a candidate antigen to develop vaccines against VL.
PLoS Neglected Tropical Diseases 03/2013; 7(3):e2148. · 4.69 Impact Factor
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Diogo G Valadares,
Mariana C Duarte,
Laura Ramírez,
Miguel A Chávez-Fumagalli,
Vivian T Martins,
Lourena E Costa,
Paula S Lage,
Tatiana G Ribeiro,
Rachel O Castilho,
Ana Paula Fernandes,
Wiliam C B Régis,
Manuel Soto,
Carlos A P Tavares,
Eduardo A F Coelho
[show abstract]
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ABSTRACT: The present study aimed to investigate the in vitro antileishmanial activity of five fractions obtained from Agaricus blazei water extract (AbM), namely, Fab1, Fab2, Fab3, Fab4, and Fab5; and use the selected leishmanicidal fraction to treat BALB/c mice infected with Leishmania chagasi. A curve dose-titration was performed to obtain the concentration to be test in infected animals. In this context, Fab5 fraction and AbM were used in the doses of 20 and 100mg/kg/day, respectively, with the product been administered once a day. The effect induced by a chemo-prophylactic regimen, based on the administration Fab5 fraction and AbM 5days before infection, and maintained for an additional 20days post-infection was compared to a therapeutic regimen, in which the compounds were administered from 0 to 20days of infection. Control animals were either treated with amphotericin B deoxycholate (AmpB) or received distilled water. All groups were followed up for 10weeks post-infection, when parasitological and immunological parameters were analyzed. The Fab5 presented the best results of in vitro leishmanicidal activity. In the in vivo experiments, the use of Fab5 or AbM, as compared to control groups, resulted in significant reduced parasite burdens in the liver, spleen, and draining lymph nodes of the infected animals, as compared to control groups. A Type 1 immune response was observed in the Fab5 or AbM treated animals. No significant toxicity was observed. The chemo-prophylactic regimen proved to be more effective to induce theses responses. In this context, the data presented in this study showed the potential of the purified Fab5 fraction of AbM as a therapeutic alternative to treat visceral leishmaniasis. In addition, it can be postulated that this fraction can be also employed in a chemo-prophylactic regimen associated or not with other therapeutic products.
Experimental Parasitology 07/2012; 132(2):228-36. · 2.12 Impact Factor
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Diogo G Valadares,
Mariana C Duarte,
Laura Ramírez,
Miguel A Chávez-Fumagalli,
Paula S Lage,
Vivian T Martins,
Lourena E Costa,
Tatiana G Ribeiro,
Wiliam C B Régis,
Manuel Soto,
Ana Paula Fernandes,
Carlos A P Tavares,
Eduardo A F Coelho
[show abstract]
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ABSTRACT: The development of therapeutic alternatives to treat leishmaniasis has received considerable attention. The present study aimed to investigate the efficacy of the Agaricus blazei Murill water extract (AbM) to treat BALB/c mice infected with Leishmania amazonensis. First, a dose-titration curve was performed. The most well-defined concentration able to induce the most effective results in the infected animals, considering a daily administration of the product, was that of 100 mg kg(-1) day(-1). In this context, the AbM was administered orally, beginning on day 0 up to 20 days postinfection. Additional animals were treated with amphotericin B (AmpB, 5 mg kg(-1) day(-1)) by peritoneal route for the same period of time, while the control group received distilled water. The animals were evaluated at 14 weeks post-infection, at which time the parasitological and immunological parameters were analyzed. Mice treated with the AbM presented a 60 % reduction in the inflammation of infected footpads as compared to untreated control-infected mice. Moreover, in the treated mice, as compared to the untreated controls, approximately 60 and 66 % reductions could be observed in the parasite burdens of the footpad and draining lymph nodes, respectively. In addition, no parasites could be detected in the spleen of treated mice at week 14 postinfection. These treated animals produced significantly higher levels of interferon gamma (IFN-γ) and nitric oxide (NO), higher levels of parasite-specific IgG2a isotype antibodies, and lower levels of interleukin (IL)-4, and IL-10 in the spleen and lymph node cell cultures than did the controls. Differences could be observed by comparing animals treated with AbM to those treated with AmpB, as indicated by a significant reduction in tissue parasitism, higher levels of IFN-γ and NO, and lower levels of IL-4 and IL-10, as well as by a decreased hepatic toxicity. In conclusion, the present study's data show that the A. blazei Murill water extract presents a high potential for the treatment of leishmaniasis, although additional studies on mice, as well as on other mammal hosts, are warranted in an attempt to determine this extract's true efficacy as compared to other known therapeutic products.
Parasitology Research 07/2012; 111(4):1807-16. · 2.15 Impact Factor
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Vinicio T S Coelho,
Jamil S Oliveira, Diogo G Valadares,
Miguel A Chávez-Fumagalli,
Mariana C Duarte,
Paula S Lage,
Manuel Soto,
Marcelo M Santoro,
Carlos A P Tavares,
Ana Paula Fernandes,
Eduardo A F Coelho
[show abstract]
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ABSTRACT: The present study aims to identify antigens in protein extracts of promastigote and amastigote-like Leishmania (Leishmania) chagasi syn. L. (L.) infantum recognized by antibodies present in the sera of dogs with asymptomatic and symptomatic visceral leishmaniasis (VL).
Proteins recognized by sera samples were separated by two-dimensional electrophoresis (2DE) and identified by mass spectrometry. A total of 550 spots were observed in the 2DE gels, and approximately 104 proteins were identified. Several stage-specific proteins could be identified by either or both classes of sera, including, as expected, previously known proteins identified as diagnosis, virulence factors, drug targets, or vaccine candidates. Three, seven, and five hypothetical proteins could be identified in promastigote antigenic extracts; while two, eleven, and three hypothetical proteins could be identified in amastigote-like antigenic extracts by asymptomatic and symptomatic sera, as well as a combination of both, respectively.
The present study represents a significant contribution not only in identifying stage-specific L. infantum molecules, but also in revealing the expression of a large number of hypothetical proteins. Moreover, when combined, the identified proteins constitute a significant source of information for the improvement of diagnostic tools and/or vaccine development to VL.
PLoS Neglected Tropical Diseases 01/2012; 6(1):e1430. · 4.69 Impact Factor
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Dulcilene M Oliveira,
Mariana Amália F Costa,
Miguel A Chavez-Fumagalli, Diogo G Valadares,
Mariana C Duarte,
Lourena E Costa,
Vivian T Martins,
Rosângela F Gomes,
Maria N Melo,
Manuel Soto,
Carlos Alberto P Tavares,
Eduardo Antonio F Coelho
[show abstract]
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ABSTRACT: Experimental vaccines to protect against visceral leishmaniasis (VL) have been developed by using BALB/c mice infected with a large (10⁷ to 10⁸) inoculum of parasites. Remarkably, prior literature has reported that the poor protection observed is mainly due to the high susceptibility of this strain. To determine factors inherent to mice that might abrogate vaccine-induced efficacy, the present research sought to investigate the impact of the administration of different infective inoculums of Leishmania chagasi (syn. L. infantum) in BALB/c mice, evaluating subcutaneous and intravenous routes of administration as well as parasitological and immunological parameters over different periods of time. This study shows that the injection of a highly infective inoculum in mice, through both subcutaneous and intravenous routes, results in a sustained infection. The mice developed a high parasite load in the liver; however, these values diminished over time. This result did not corroborate with the parasite load in the bone marrow and brain and proved to be expressively different in the spleen and draining lymph nodes, where the values increased over time. Mice infected with a low dose of parasites (10³) showed a certain resistance against infection, based mainly on the IFN-γ and oxide nitric production. Considering all the elements, it could be concluded that the models employing high doses (10⁷) of L. chagasi in BALB/c mice can bring about an imbalance in the animals' immune response, thus allowing for the development of the disease at the expense of efficacy within the vaccine candidates.
Parasitology Research 09/2011; 110(3):1277-85. · 2.15 Impact Factor
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Diogo G Valadares,
Mariana C Duarte,
Jamil S Oliveira,
Miguel A Chávez-Fumagalli,
Vivian T Martins,
Lourena E Costa,
João Paulo V Leite,
Marcelo M Santoro,
Wiliam C B Régis,
Carlos A P Tavares,
Eduardo A F Coelho
[show abstract]
[hide abstract]
ABSTRACT: Leishmaniasis is a major public health problem, and the alarming spread of parasite resistance underlines the importance of discovering new therapeutic products. The present study aims to investigate the in vitro leishmanicidal activity of an Agaricus blazei Murill mushroom extract as compared to different Leishmania species and stages. The water extract proved to be effective against promastigote and amastigote-like stages of Leishmania amazonensis, L. chagasi, and L. major, with IC(50) (50% inhibitory concentration) values of 67.5, 65.8, and 56.8 μg/mL for promastigotes, and 115.4, 112.3, and 108.4 μg/mL for amastigotes-like respectively. The infectivity of the three Leishmania species before and after treatment with the water extract was analyzed, and it could be observed that 82%, 57%, and 73% of the macrophages were infected with L. amazonensis, L. major, and L. chagasi, respectively. However, when parasites were pre-incubated with the water extract, and later used to infect macrophages, they were able to infect only 12.7%, 24.5%, and 19.7% of the phagocytic cells for L. amazonensis, L. chagasi, and L. major, respectively. In other experiments, macrophages were infected with L. amazonensis, L. chagasi, or L. major, and later treated with the aforementioned extract, presented reductions of 84.4%, 79.6%, and 85.3% in the parasite burden after treatment. A confocal microscopy revealed the loss of the viability of the parasites within the infected macrophages after treatment with the water extract. The applied extract presented a low cytotoxicity in murine macrophages and a null hemolytic activity in type O(+) human red blood cells. No nitric oxide (NO) production, nor inducible nitric oxide syntase expression, could be observed in macrophages after stimulation with the water extract, suggesting that biological activity may be due to direct mechanisms other than macrophage activation by means of NO production. In conclusion, the results demonstrate that the A. blazei Murill water extract can potentially be used as a therapeutic alternative on its own, or in association with other drugs, to treat Visceral and Cutaneous Leishmaniasis.
Parasitology International 06/2011; 60(4):357-63. · 2.13 Impact Factor
