Chalcone-rich extracts from Lonchocarpus cultratus roots present in vitro leishmanicidal and immunomodulatory activity.

OBJECTIVES: This study aimed to evaluate the in vitro anti-Leishmania activity of chalcone-rich three extracts (LDR, LHR and LMR) from Lonchocarpus cultratus (Vell.) A.M.G. Azevedo & H.C. Lima against L. amazonensis. Also, the immunomodulatory and antioxidant capacity was assessed. METHODS: Successive extraction with hexane, dichloromethane and methanol were performed to obtain LHR, LDR and LMR extracts from L. cultratus roots, which were characterized by 1H NMR. Promastigotes, amastigotes and peritoneal macrophages were exposed to crescent concentrations of the three extracts, and after incubation, the inhibition rates were determined to both types of cells, and morphological analyses were performed on the parasite. The immunomodulatory activity was determined against stimulated macrophages. KEY FINDINGS: LDR, LHR and LMR inhibited promastigote cell growth (IC50 0.62 ± 0.3, 0.94 ± 0.5 and 1.28 ± 0.73 µg/ml, respectively) and reduced the number of amastigotes inside macrophages (IC50 1.36 ± 0.14, 1.54 ± 0.26 and 4.09 ± 0.88 µg/ml, respectively). The cytotoxicity against murine macrophages resulted in a CC50 of 13.12 ± 1.92, 92.93 ± 9.1 and >300 µg/ml, resulting in high selectivity index to promastigotes and amastigotes. The extracts also inhibited the nitric oxide secretion in RAW 264.7 macrophages. The antioxidant capacity resulted in a higher scavenger LMR ability. CONCLUSIONS: These results suggest that L. cultratus extracts have anti-Leishmania potential, are non-toxic, and immunosuppress macrophages in vitro.

Anti-Trypanosoma cruzi activity, cytotoxicity, and chemical characterization of extracts from seeds of Lonchocarpus cultratus.

INTRODUCTION: Trypanosoma cruzi is the agent of Chagas' disease and affects approximately 6-8 million people worldwide. The search for new anti-T. cruzi drugs are relevant because only two drugs exist actually. The objective of this study was to investigate the effect of the extracts from the seeds of Lonchocarpus cultratus on T. cruzi, its cytotoxicity as well as to elucidate its chemical profile. METHODOLOGY: The characterization of the extracts was done using 1H-RMN. T. cruzi forms were treated with increasing concentrations of the extracts and after, the percentage of inhibition and IC50 or LC50 were calculated. Murine peritoneal macrophages were treated with different concentrations of the extracts to evaluate the cellular viability. The hemotoxicity was accessed by verifying the levels of hemolysis caused by the extracts on human red blood cells. RESULTS: Chalcones isocordoin and lonchocarpin were detected in the dichloromethane extract, and chalcone lonchocarpin was detected in the hexane extract. The dichloromethane extract showed higher activity against all the forms of T. cruzi compared to the other two extracts, but the hexane showed the best selectivity index. The cytotoxicity observed in murine macrophages was confirmed in human erythrocytes, with dichloromethane extract having the highest toxicity. The methanolic extract showed the lowest anti-T. cruzi activity but was nontoxic to peritoneal murine macrophages and red blood cells. CONCLUSIONS: L. cultratus extracts have the potential to be explored for the development of new anti-trypanosomal drugs. This study was the first to demonstrate the action of extracts from the genus Lonchocarpus on infecting forms of T. cruzi.

Determination of chemical structure and anti-Trypanosoma cruzi activity of extracts from the roots of Lonchocarpus cultratus (Vell.) A.M.G. Azevedo & H.C. Lima.

Trypanosoma cruzi is the agent of Chagas disease, an infection that affects around 8 million people worldwide. The search for new anti-T. cruzi drugs are relevant, mainly because the treatment of this disease is limited to two drugs. The objective of this study was to investigate the trypanocidal and cytotoxic activity and elucidate the chemical profile of extracts from the roots of the Lonchocarpus cultratus. Roots from L. cultratus were submitted to successive extractions with hexane, dichloromethane, and methanol, resulting in LCH, LCD, and LCM extracts, respectively. Characterization of extracts was done using 1H-RMN, 13C-RMN, CC and TLC. Treatment of T. cruzi forms (epimastigotes, trypomastigotes, and amastigotes) with crescent concentrations of LCH, LCD, and LCM was done for 72, 48, and 48 h, respectively. After this, the percentage of inhibition and IC50/LC50 were calculated. Benznidazole was used as a positive control. Murine macrophages were treated with different concentrations of both extracts for 48 h, and after, the cellular viability was determined by the MTT method and CC50 was calculated. The chalcones derricin and lonchocarpine were identified in the hexane extract, and for the first time in the genus Lonchocarpus, the presence of a dihydrolonchocarpine derivative was observed. Other chalcones such as isocordoin and erioschalcone B were detected in the dichloromethane extract. The dichloromethane extract showed higher activity against all tested forms of T. cruzi than the other two extracts, with IC50 values of 10.98, 2.42, and 0.83 µg/mL, respectively; these values are very close to those of benznidazole. Although the dichloromethane extract presented a cytotoxic effect against mammalian cells, it showed selectivity against amastigotes. The methanolic extract showed the lowest anti-T. cruzi activity but was non-toxic to peritoneal murine macrophages. Thus, the genus Lonchocarpus had demonstrated in the past action against epimastigotes forms of T. cruzi but is the first time that the activity against infective forms is showed, which leading to further studies with in vivo tests.

Aqueous ozone therapy improves the standard treatment of leishmaniasis lesions in animals leading to local and systemic alterations.

The current treatment of leishmaniasis presents some problems, such as cell toxicity, parenteral route, and time of treatment. Ozone emerges as an option to accelerate the standard treatment due to the immunomodulatory, antioxidant, and wound healing activity reported in the literature. This work aimed to evaluate the efficacy of aqueous ozone as an adjuvant to the standard treatment of cutaneous lesions caused by Leishmania amazonensis in an experimental model. For in vivo experiments, mice were randomly distributed in 6 groups, which were infected with L. amazonensis and treated in five different schedules using the standard treatment with Glucantime® with or without aqueous ozone. After the last day of treatment, the animals were euthanized and were analyzed: the thickness of lesions; collagen deposition, the parasitic burden of the lesions; blood leukocyte number; NO; and cytokine dosages and arginase activity from peritoneal macrophages. All treated groups showed a decrease in the lesion, but with a significative deposition of collagen in lesions with local ozone treatment. The parasite burden showed that ozone enhanced the leishmanicidal activity of the reference drug. The reduction of NO production and blood leukocyte count and increases in the arginase activity showed an immunomodulatory activity of ozone in the treated animals. Thus, ozone therapy has been shown to work as an adjuvant in the treatment of Leishmania lesions, enhancing leishmanicidal and wound healing activity of standard treatment.