Repositioning of Tamoxifen in Surface-Modified Nanocapsules as a Promising Oral Treatment for Visceral Leishmaniasis.

Standards of care for human visceral leishmaniasis (VL) are based on drugs used parenterally, and oral treatment options are urgently needed. In the present study, a repurposing strategy was used associating tamoxifen (TMX) with polyethylene glycol-block-polylactide nanocapsules (NC) and its anti-leishmanial efficacy was reported in vivo. Stable surface modified-NC (5 mg/mL of TMX) exhibited 200 nm in size, +42 mV of zeta potential, and 98% encapsulation efficiency. Atomic force microscopy evidenced core-shell-NC. Treatment with TMX-NC reduced parasite-DNA quantified in liver and spleen compared to free-TMX; and provided a similar reduction of parasite burden compared with meglumine antimoniate in mice and hamster models. Image-guided biodistribution showed accumulation of NC in liver and spleen after 30 min post-administration. TMX-NC reduced the number of liver granulomas and restored the aspect of capsules and trabeculae in the spleen of infected animals. TMX-NC was tested for the first time against VL models, indicating a promising formulation for oral treatment.

Behavior of two Leishmania infantum strains-evaluation of susceptibility to antimonials and expression of microRNAs in experimentally infected J774 macrophages and in BALB/c mice.

Strains of the same Leishmania parasite species, isolated from different host organisms, may exhibit unique infection profiles and induce a change in the expression of microRNAs among host macrophages and in model host mice. MicroRNAs (MiR) are endogenous molecules of about 22 nucleotides that are involved in many regulatory processes, including the vertebrate host immune response. In this respect, the infectivity and susceptibility to antimonials of two L. infantum strains, BH46, isolated from human, and OP46, isolated from symptomatic dog, were characterized in J774 macrophages and BALB/c mice. Parasite burden was assessed in the liver, spleen, and bone marrow using the serial limiting dilution technique. A higher parasite burden was observed in the spleen and bone marrow of animals infected with OP46 compared to BH46 strain. Our results also showed that OP46 was less susceptible to the antimonials. In addition, miR-122 and miR-155 expression was evaluated in the liver and J774 macrophages, and in spleens from infected animals, respectively. An increase was observed in the expression of miR-155 in J774 macrophages infected with both strains compared to uninfected cells, with a higher expression in cells infected with OP46. However, no difference in the expression of miR-122 and miR-155 was observed in the infected animals. Thus, this study shows that OP46 was more infective for mice, it caused a higher increase in miR-155 expression in infected macrophages and was less susceptible to the antimonials evaluated. These data suggest that alteration in miR-155 level likely plays a role in regulating the response to L. infantum.

Multicomponent LBSap vaccine displays immunological and parasitological profiles similar to those of Leish-Tec® and Leishmune® vaccines against visceral leishmaniasis.

BACKGROUND: In past years, many researchers have sought canine visceral leishmaniasis (CVL) prevention through the characterization of Leishmania antigens as vaccine candidates. Despite these efforts, there is still no efficient vaccine for CVL control. METHODS: In the present study, we performed a pre-clinical vaccine trial using BALB/c mice to compare the effects of the multicomponent LBSap vaccine with those of Leish-Tec® and Leishmune®. Blood was collected to determine the frequency of peripheral blood cells and to evaluate hematologic and immunophenotypic parameters. Liver and spleen samples were collected for parasitological quantification, and spleen samples were used to access the cytokine profile. RESULTS: When measuring total IgG and IgG1 anti-Leishmania levels after the third vaccination and L. infantum challenge, it was evident that all vaccines were able to induce humoral immune response. Regarding the innate immune response, increased levels of NK CD3(-)CD49(+) cells were the hallmark of all vaccinated groups, whereas only the Leish-Tec® group displayed a high frequency of CD14(+) monocytes after L. infantum challenge. Moreover, CD3(+)CD4(+) T cells were the main circulating lymphocytes induced after L. infantum challenge with all evaluated vaccines. Importantly, after L. infantum challenge, splenocytes from the Leishmune® vaccine produced high levels of IL-2, whereas a prominent type 1 immune response was the hallmark of the LBSap vaccine, which presented high levels of IL-2, IL-6, TNF-α, and IFN-γ. The efficacy analysis using real-time polymerase chain reaction demonstrated a reduction in the parasitism in the spleen (Leishmune®: 64 %; LBSap: 42 %; and Leish-Tec®: 36 %) and liver (Leishmune®: 71 %; LBSap: 62 %; and Leish-Tec®: 48 %). CONCLUSIONS: The dataset led to the conclusion that the LBSap vaccination was able to induce immune and efficacy profiles comparable with those of commercial vaccines, thus demonstrating its potential as a promising vaccine candidate for visceral leishmaniasis control.

Leishmania chagasi: effect of the iron deficiency on the infection in BALB/c mice.

Iron deficiency and visceral leishmaniasis are serious problems of public health. The aim of this study was to evaluate the effect of iron deficiency, induced by the iron chelator desferrioxamine, on the course of the infection by Leishmania chagasi in BALB/c mice. Our data show that the iron chelator caused significant reduction in hemoglobin concentration of treated mice and reduction in parasite load in spleen and liver. Significant differences were not observed in the production of IFN-gamma and IL-4 among the experimental groups. In conclusion, the data reported in this paper suggest that iron deficiency may favor the host. If there is not enough iron available to the parasite, its multiplication may be reduced and infection attenuated.

Immune response to Leishmania (Leishmania) chagasi infection is reduced in malnourished BALB/c mice.

Protein-energy malnutrition and micronutrient deficiencies may down-regulate immune response and increase morbidity and mortality due to infection. In this study, a murine model was used to study the effects of protein, iron and zinc deficiencies on the immune response to Leishmania (Leishmania) chagasi infection. Mice were initially fed a standard diet or with a diet containing 3% casein but deficient in zinc and iron. After malnutrition was established, mice were inoculated with L. chagasi and sacrificed four weeks later in order to evaluate liver and spleen parasite loads and serum biochemical parameters. Significant decreases in liver and spleen weight, an increase in the parasite loads in these organs and decreases in serum protein and glucose concentrations in malnourished animals were observed. Furthermore, the production of interferon-gamma by spleen cells from infected malnourished mice stimulated by Leishmania antigen was significantly lower compared with that in control diet mice. These data suggest that malnutrition alters the immune response to L. chagasi infection in the BALB/c model and, in association with the effects on biochemical and anatomical parameters of the host, favored increases in the parasite loads in the spleens and livers of these animals.

Immune response to Leishmania (Leishmania) chagasi infection is reduced in malnourished BALB/c mice

Protein-energy malnutrition and micronutrient deficiencies may down-regulate immune response and increase morbidity and mortality due to infection. In this study, a murine model was used to study the effects of protein, iron and zinc deficiencies on the immune response to Leishmania (Leishmania) chagasi infection. Mice were initially fed a standard diet or with a diet containing 3 percent casein but deficient in zinc and iron. After malnutrition was established, mice were inoculated with L. chagasiand sacrificed four weeks later in order to evaluate liver and spleen parasite loads and serum biochemical parameters. Significant decreases in liver and spleen weight, an increase in the parasite loads in these organs and decreases in serum protein and glucose concentrations in malnourished animals were observed. Furthermore, the production of interferon-gamma by spleen cells from infected malnourished mice stimulated by Leishmaniaantigen was significantly lower compared with that in control diet mice. These data suggest that malnutrition alters the immune response to L. chagasiinfection in the BALB/c model and, in association with the effects on biochemical and anatomical parameters of the host, favored increases in the parasite loads in the spleens and livers of these animals.

Epitope mapping and protective immunity elicited by adenovirus expressing the Leishmania amastigote specific A2 antigen: correlation with IFN-gamma and cytolytic activity by CD8+ T cells.

A2 was identified as an amastigote virulence factor of Leishmania (Leishmania) donovani and as a candidate antigen for vaccine development against visceral leishmaniasis. Here, predicted hydrophilic, class I and II MHC-binding synthetic peptides were used to define epitopes recognized by A2-specific antibodies, CD8+ T and CD4+ T cells, respectively. Immunization of BALB/c mice with adenovirus expressing A2 (AdA2) resulted in low antibody response, contrasting with high levels of IFN-gamma producing CD4+ T and CD8+ T cells specific for A2. Further, A2-specific CD8+ T cells from immunized mice were capable of lysing sensitized target cells in vivo. Finally, we demonstrated an association of A2-specific T cell responses and reduced parasitism in both liver and spleen from mice immunized with AdA2 and challenged with L. (L.) chagasi.

Successful vaccination against Leishmania chagasi infection in BALB/c mice with freeze-thawed Leishmania antigen and Corynebacterium parvum.

This study evaluated the potential of a Leishmania antigen vaccine in protecting BALB/c mice against Leishmania chagasi. Mice received two subcutaneous doses of L. amazonensis vaccine with Corynebacterium parvum and subsequent boost was done without adjuvant. One week later, mice were challenged with L. chagasi. We observed that this vaccine caused a significant reduction in parasite load in liver and spleen and induced a high production of IFN-gamma and IL-4 by spleen cells from vaccinated mice in response to Leishmania antigen. Together, our data show that this vaccine is capable of inducing a Th1/Th2 response that is important to control parasite replication.