Bioprospecção e seleção de ativos de Amburana cearensis com efeito leishmanicida (Leishmania braziliensis) por imunomodulação

Abstract

The actual pharmacotherapy for the treatment of leishmaniasis has serious side effects, as well as cases of drug resistance. Species such as Amburana cearensis A.C. Smith (cumaru), a caatinga tree, whose extract and / or molecules have shown antiinflammatory, antioxidant, antiparasitic properties can be a potential source for new treatments. The present study evaluated the leishmanicidal effect in vitro and in vivo of secondary metabolites isolated from A. cearensis in infection by L. braziliensis. The dry extract (spray dryer) standard HPLC-PDA (markers / CM: 48.573 ± 0.2674 mg / g and AMB: 15.118 ± 1.1181 mg / g), phenolic fraction of A. cearensis (ESAC and FFAC, respectively) and chemical constituents - coumarin (CUM), amburoside A (AMB), vanillic acid (AV) were added to the suspension of promastigotes for up to 48 hours, and the antipromastigote effect was measured by microscopic analysis with Trypan Blue. To evaluate the effect against amastigote forms Raw 264.7 macrophage infected with L. braziliensis was treated or not with CUM (10, 25,50,10 μg / mL). The cytotoxicity of the drugs was evaluated by the MTT assay, parasitic load and cytokine dosage after 24 and 48 hours of infection. For in vivo tests, lesion development and parasitic load (lesion and lymph node-LN) were evaluated, for which Hamsters (Mesocricetus auratus; n=24) were infected with 107 promastigotes of L. braziliensis in the ear. After the appearance of the lesions, animals were treated Antimony or CUM. All estudied metabolites showed anti-promastigote effect after 48h of incubation, however the CUM, showed better inhibition after 24h of incubation with reduction (100ug / mL: 57.3% ± 3.54%). The effect of CUM was not related to cytotoxicity. CUM (100ug / mL) reduced the intracellular parasitic load in 24 and 48h of incubation (maximum reduction: 49% ± 2.517%) accompanied by an increase in NO production (100ug / mL / 48h: 114, 2 µmol / L). There was an increase in TNFα and IL-10, levels and a decrease in the production of IL-4. Regarding the vivo experiments, CUM administered orally reduced the thickness of the lesion, the parasitic load at the lesion site and prevented the spread of the disease to the lymph node, compared to the untreated control. In view of the results obtained, CUM showed a promising leishmanicidal effect in animals infected with L. braziliensis, being capable to induce the control of the lesion, suggesting that this may occur due to the decrease in the parasitic load as well as a modulation of the immune response with induction of a Th1 response profile (increased TNF-α and IL-12) and increased antiinflammatory cytokines (IL-10), thus providing a favorable microenvironment for the control of intracellular parasitemia in macrophages infected with L. braziliensis.