Luiz Gustavo Rodrigues Oliveira

Enhanced protection by melatonin and meloxicam combination in experimental infection by Trypanosoma cruzi

The aim of this study was to evaluate a possible synergism between melatonin and meloxicam in up‐regulating the immune response in male Wistar rats infected with Trypanosoma cruzi during immunosuppression phenomenon, which characterizes the acute phase of the Chagas’ disease. Male Wistar rats were infected with the Y strain of T. cruzi. Experiments were performed on 7, 14 and 21 days post‐infection. Several immunological parameters were evaluated including γ‐interferon (IFN‐γ), interleukin‐2 (IL‐2), nitric oxide (NO) and prostaglandin E2 (PGE2). The combined treatment with melatonin and meloxicam significantly enhanced the release of IL‐2 and INF‐γ into animals’ serum, when compared with the infected control groups during the course of infection. Furthermore, the blockade of PGE2 synthesis and the increased release of NO by macrophage cells from T. cruzi‐infected animals contributed to regulate the production of Th1 subset cytokines significantly reducing the parasitaemia in animals treated with the combination of both substances. Therefore, our results suggest that the association of melatonin and meloxicam was more effective in protecting animals against the harmful actions of T. cruzi infection as compared with the treatments of meloxicam or melatonin alone.

In vitro Leishmanicidal and Cytotoxic Activities of the Glycoalkaloids from Solanum lycocarpum (Solanaceae) Fruits

Leishmaniasis is an infection caused by a protozoan parasite of the genus Leishmania and is the second most prevalent parasitic protozoal disease after malaria in the world. We report the in vitro leishmanicidal activity on promastigote forms of Leishmania amazonensis and cytotoxicity, using LLCMK2 cells, of the glycoalkaloids from the fruits of Solanum lycocarpum, determined by colorimetric methods. The alkaloidic extract was obtained by acid‐base extraction; solamargine and solasonine were isolated by silica‐gel chromatography, followed by reversed‐phase HPLC final purification. The alkaloidic extract, solamargine, solasonine, as well as the equimolar mixture of the glycoalkaloids solamargine and solasonine displayed leishmanicidal activity against promastigote forms of L. amazonensis, whereas the aglycone solasodine was inactive. After 24 and 72 h of incubation, most of the samples showed lower cytotoxicities (IC50 6.5 to 124 μM) as compared to leishmanicidal activity (IC50 1.1 to 23.6 μM). The equimolar mixture solamargine/solasonine was the most active with an IC50 value of 1.1 μM, after 72 h. Likewise, solamargine was the most active after 24 h with an IC50 value of 14.4 μM, both in comparison with the positive control amphotericin B.

Protective actions of melatonin against heart damage during chronic Chagas disease.

Chronic cardiomyopathy is the most important clinical form of Chagas disease, and it is characterised by myocarditis that is associated with fibrosis and organ dysfunction. Alternative treatment options are important tools to modulate host immune responses. The main goal of this work was to evaluate the anti-inflammatory actions of melatonin during the chronic phase of Chagas disease. TNF-α, IL-10 and nitrite concentrations were evaluated as predictive factors of immune modulation. Creatine phosphokinase-MB (CK-MB), cardiac inflammatory foci and heart weight were assessed to evaluate the efficacy of the melatonin treatment. Male Wistar rats were infected with 1×10(5) blood trypomastigotes of the Y strain of Trypanosoma cruzi and kept untreated for 60 days to mimic chronic infection. After this period, the rats were orally treated with melatonin 50mg/kg/day, and the experiments were performed 90, 120, and 180 days post-infection. Melatonin treatment significantly increased the concentration of IL-10 and reduced the concentrations of NO and TNF-α produced by cardiomyocytes. Furthermore, it led to decreased heart weight, serum CK-MB levels and inflammatory foci when compared to the untreated and infected control groups. We conclude that melatonin therapy is effective at protecting animals against the harmful cardiac inflammatory response that is characteristic of chronic T. cruzi infection.

Melatonin and dehydroepiandrosterone combination: does this treatment exert a synergistic effect during experimental Trypanosoma cruzi infection?

Abstract:  Previous studies showed that melatonin or dehydroepiandrosterone (DHEA) enhances the immune response against parasitic pathogens. The present study investigated the in vitro activity of melatonin combined with DHEA in a period of 24 hr during the course of in vivo T. cruzi infection. The in vitro activity of melatonin or DHEA alone, as well as together, were tested for the trypomastigote forms (doses ranging from 0.5 to 128 μm). In vitro, neither melatonin nor DHEA alone had any activity against trypomastigote forms, although when the highest concentration of combined melatonin and DHEA was used, it was active against the trypomastigote forms of the parasite. However, for this concentration, a quite toxicity on peritoneal macrophages was observed. For in vivo evaluation, male Wistar rats were infected with the Y strain of T. cruzi. They were orally treated with 10 mg/kg body weight/day of melatonin and subcutaneously with 40 mg/kg body weight/day of DHEA. Treatment with melatonin, DHEA and the association showed a significant reduction in the number of blood trypomastigotes during the acute phase of infection as compared to untreated animals (P < 0.05). A significant increase in the number of macrophages and nitric oxide (NO) concentrations were observed during the peak of parasitaemia with melatonin alone or combined with DHEA. However, with DHEA alone the highest concentration of NO was observed (P < 0.05). Moreover, DHEA treatment increased TNF‐alpha levels during the infection (P < 0.05). These results show that melatonin, DHEA or the combination of both reduces parasitemia during the acute phase of infection. The combined action of both molecules did not exert a synergic action on the host’s ability to fight infection, and it seems that among all treatments DHEA induces a more efficient immune response.

Immunomodulatory effect of the alkaloidic extract of Solanum lycocarpum fruits in mice infected with Schistosoma mansoni

Schistosomiasis is a chronic disease caused by trematode flatworms of the genus Schistosoma; it accounts for more than 280,000 deaths annually. In this work we investigated the effect of the alkaloidic extract obtained by acid-base extraction of the dried fruits of Solanum lycocarpum on schistosomiasis. We used this extract at concentrations of 10, 20, and 40 mg/kg to treat mice infected with Schistosoma mansoni in different phases of the parasite cycle, and we compared its effect with that of the positive control praziquantel (60 mg/kg). We evaluated the results on the basis of the number of macrophages, eggs, and granulomas; we also assessed nitric oxide (NO) and interferon-gamma (IFN-γ) production. Animals treated with a daily dose of 10 or 20 mg/kg alkaloidic extract between the 37th and 41st day of infection showed increased number of macrophages, elevated NO and IFN-γ concentrations, and reduced number of eggs and granulomas in the liver. The alkaloidic extract of S. lycocarpum fruits displayed an immunomodulatory effect on mice infected with S. mansoni, so its potential to treat schistosomiasis deserves further studies.

Prior and concomitant dehydroepiandrosterone treatment affects immunologic response of cultured macrophages infected with Trypanosoma cruzi in vitro

DHEA, a steroid hormone synthesized from cholesterol by cells of the adrenal cortex, plays an essential role in enhancing the hosts resistance to different experimental infections. Receptors for this hormone can be found in distinct immune cells (especially macrophages) that are known to be the first line defense against Trypanosoma cruzi infection. These cells operate through an indirect pathway releasing nitric oxide (NO) and cytokines such TNF-α and IL-12 which in turn trigger an enhancement of natural killer cells and lymphocytes which finally secrete pro and anti-inflammatory cytokines. The effects of pre- and post-infection DHEA treatment on production of IL-12, TNFα and NO were evaluated. T. cruzi infected macrophages post treated with DHEA displayed enhanced concentrations of TNF-α, IL-12 and NO. Probably, the mechanisms that induced the production of cytokines by infected cells are more efficient when the immune system has been stimulated first by parasite invasion, suggesting that the protective role of DHEA is greater when administered post infection.

Effects of dehydroepiandrosterone-sulfate (DHEA-S) and benznidazole treatments during acute infection of two different Trypanosoma cruzi strains.

A significant role for hormones in regulating the balance of Th1- and Th2-associated cytokines with a role in modulating diseases has been accumulating. Previously, we reported that dehydroepiandrosterone (DHEA), the most abundant steroid hormone synthesized by the adrenal cortex, markedly reduced the blood and tissue parasites in experimentally Trypanosoma cruzi-infected rats. Based on these findings, the main purpose of this study was to investigate the effect of dehydroepiandrosterone-sulfate ester (DHEA-S) therapy alone or in combination with benznidazole (BNZ) (recommended in Brazil for the treatment of T. cruzi infection) will be effective during the acute phase of two different lineages of T. cruzi strains: type I (Y strain) and type II (Bolivia strain) of T. cruzi. Administration of either DHEA-S or BNZ alone or in combination significantly reduced the Y strain parasite load as compared with untreated. Furthermore treatment with DHEA-S resulted in Bolivia strain clearance. This protective effect of DHEA-S was associated with the hosts immune response, as evidence by enhanced levels of interferon-gamma and interleukin-2. DHEA-S treatment also increased peritoneal macrophages levels and nitrite production. DHEA-S treatment was effective in reducing the mortality rate as compared to BNZ alone or to combiner DHEA-S+BNZ treatment of T. cruzi Bolivia strain infected animals. These findings suggest that hormonal therapy may have a protective effect in the treatment of T. cruzi infection.

Distinctive histopathology and modulation of cytokine production during oral and intraperitoneal Trypanosoma cruzi Y strain infection.

Acute Chagas disease outbreaks are related to the consumption of food or drink contaminated by triatomine feces, thus making oral infection an important route of transmission. Both vector-borne and oral infections trigger important cardiac manifestations in the host that are related to a dysregulated immune response. The aims of this work were to evaluate possible alterations of lymphocyte CD4+/CD8+ sub-populations, Th1 and Th2 cytokines, nitrite concentrations and cardiac histopathology. One group of male Wistar rats was intraperitoneally infected (I.P.) with 1×105 metacyclic trypomastigotes of the T. cruzi Y strain, and another group of Wistar rats was orally infected (O.I.) with 8×105 metacyclic trypomastigotes of the same strain. The intraperitoneal infection triggered statistically enhanced parasite and peritoneal macrophage numbers, increased concentrations of NO and IL-12 and elevated cardiac inflammatory foci when compared with the oral infection. However, proliferation of CD4+ and CD8+ T cells were not statistically different for oral and intraperitoneal routes.