In vitro antileishmanial effects of Physalis angulata root extract on Leishmania infantum.

OBJECTIVE: In the present study, we evaluated the effects of the aqueous extract of Physalis angulata root (AEPa) on Leishmania infantum proliferation, morphology, and the driving mechanism in leishmanicidal activity and modulatory action on macrophages. METHODS: L. infantum promastigotes were treated with 50 and 100 µg/mL AEPa for 72 h and then antipromastigote assay was performed by counts in a Newbauer chamber, morphological changes were analyzed by transmission electron microscopy and the mechanism of the leishmanicidal activity was detected. In addition, macrophages were infected with L. infantum and were used to evaluate anti-amastigote activity of AEPa and effects of AEPa on cytokine secretion after 72-hour treatment. RESULTS: Treatment with AEPa reduced the numbers of L. infantum promastigotes (50% inhibitory concentration (IC50) = 65.9 µg/mL; selectivity index (SI) = 22.1) and amastigotes (IC50 = 37.9 µg/mL; SI = 38.5) compared with the untreated control. Amphotericin B reduced 100% of the promastigote numbers after 72 h of treatment (IC50 = 0.2 µg/mL). AEPa induced several morphological changes and increased the production of reactive oxygen species and apoptotic death in promastigotes after treating for 72 h. AEPa (100 µg/mL) promoted tumor necrosis factor-α secretion in macrophages infected with L. infantum after 72 h of treatment, but did not induce an increase in this cytokine in noninfected macrophages. In addition, AEPa showed no cytotoxic effect on J774-A1 cells (50% cytotoxic concentration >1000 µg/mL). CONCLUSION: AEPa presented antileishmanial activity against the promastigotes and amastigotes of L. infantum without macrophage cytotoxicity; these results show that natural products such as P. angulata have leishmanicidal potential and in the future may be an alternative treatment for leishmaniasis.

Selective effects of Euterpe oleracea (açai) on Leishmania (Leishmania) amazonensis and Leishmania infantum.

Leishmania (Leishmania) amazonensis and Leishmania infantum (=Leishmania chagasi) are protozoa that cause American cutaneous and visceral leishmaniasis, respectively. These diseases show a high incidence in developing countries such as Brazil. The treatments used for leishmaniasis are still limited due to their high cost and toxicity. Currently, some natural products are considered an important alternative source of new leishmanicidal agents. Euterpe oleracea Martius, a palm producing black fruits, is frequently consumed in the Amazon region, as a juice, known as açai, with potent antioxidant, anti-inflammatory and anticonvulsant properties. Interestingly, the biological activity of clarified açai juice (EO) on L. (L.) amazonensis and L. infantum (=L. chagasi) is unknown. Therefore, the mechanism of anti-leishmanial action of EO has been evaluated on L. (L.) amazonensis and L. infantum (=L. chagasi). EO reduced the number of promastigotes and caused morphological alterations, increased the production of reactive oxygen species (ROS) and induced cell death phenotypes probably seems by apoptosis in the promastigotes of L. (L.) amazonensis (IC50 = 1:40) and L. infantum (=L. chagasi) (IC50 = 1:38). EO also presented activity against Leishmania amastigotes. Treatment with EO for 72 h strongly reduced IL-17 cytokine levels at all tested concentrations and decreased the number of intracellular amastigotes in macrophages infected with L. (L.) amazonensis (IC50 = 1:30) and L. infantum (=L. chagasi) (IC50 = 1:38). Additionally, no cytotoxic effect was observed in murine macrophages treated with EO (72 h - CC50 > 1:1). Our results demonstrated that EO has leishmanicidal activity against two different species that cause American visceral and cutaneous leishmaniasis without cytotoxic effects for the host cell.

Medicinal plants from the Brazilian Amazonian region and their antileishmanial activity: a review.

Leishmaniasis, a neglected disease caused by Leishmania protozoans, primarily affects people in tropical and subtropical areas. Chemotherapy based on the use of pentavalent antimonials, amphotericin B, paromomycin, miltefosine and liposomal amphotericin B is currently the only effective treatment. However, adverse effects, long-term treatment and the emergence of parasite resistance have led to the search for alternative treatments. Natural products used in traditional medicine provide an unlimited source of molecules for the identification of new drugs, and the Amazon region has abundant biodiversity that includes several species of plants and animals, providing a rich source of new products and compounds. Although the literature describes numerous promising compounds and extracts for combating Leishmania protozoans, the results of such research have not been embraced by the pharmaceutical industry for the development of new drugs. Therefore, this review focused on the antileishmanial activity of extracts, isolated compounds and essential oils commonly used by the local population in the Brazilian Amazonian region to treat several illnesses and described in the literature as promising compounds for combating leishmaniasis.

Biological effects of kojic acid on human monocytes in vitro.

Monocytes are mononuclear phagocytes in peripheral blood that can differentiate into macrophages and dendritic cells. Macrophages play a specific role in the inflammatory process and are essential for the innate response. Given the important role of monocytes/macrophages in the immune response, this study aimed to evaluate the activity of kojic acid (KA), a natural product of certain fungal species, on human peripheral blood monocytes in vitro. Purified monocytes isolated from human blood were incubated with KA (50 µg/mL for 48 h) and analyzed by light microscopy, scanning electron microscopy, transmission electron microscopy and flow cytometry. Host cell cytotoxicity was measured by the colorimetric MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. KA treatment induced morphological alterations in monocytes, such as increased cell size, as well as numerous cellular projections. Furthermore, flow cytometry revealed increased labeling of cell surface EMR1-F4/80 but decreased labeling of CD11b and CD14. KA also promoted increased IL-6 cytokine production but did not cause cytotoxic effects in monocytes. In conclusion, our results show that KA promotes the differentiation of monocytes into macrophages and can act as an immunomodulatory agent.

Comparative analysis of carbohydrate residues in the midgut of phlebotomines (Diptera: Psychodidae) from colony and field populations from Amazon, Brazil.

Leishmaniasis are worldwide diseases that occur in 98 countries including Brazil, transmitted by the bite of female phlebotomines during blood feeding. In Brazil it is known that some species of sand flies as Lutzomyia longipalpis sensun latum (vector of Leishmania infantum chagasi), Lutzomyia flaviscutellata (vector of Leishmania (Leishmania) amazonensis) and Lutzomyia antunesi [suspected vector of Leishmania (Viannia) lindenbergi] are incriminated of transmitting the parasite Leishmania for the vertebrate host. The phlebotomine-parasite is mediated by the attachment of the promastigote lipophosphoglycan (LPG) to the midgut epithelium. However, another mechanism that is LPG-independent and mediated by N-acetyl-galactosamine (GalNAc) seems to occur in some species of phlebotomines that are classified as permissive. The aim of this study was to characterize the carbohydrate residues that, probably, play a role in parasite attachment to the midgut of phlebotomine from colony and field populations from the Brazilian Amazonian region. We observed the presence of GalNAc, mannose, galactose and GlcNAc in all phlebotomine species. A binding assay between L. (L.) amazonensis and L. i.chagasi to the midguts of different species of phlebotomines was performed. The attachment of both Leishmania and vector species suggests the presence of GalNAc on the midgut surfaces. Thus, these results suggested that GalNAc is a possible binding sites of Leishmania in sand flies from the Brazilian Amazonian region.

Physalis angulata induces in vitro differentiation of murine bone marrow cells into macrophages.

BACKGROUND: The bone marrow is a hematopoietic tissue that, in the presence of cytokines and growth factors, generates all of the circulating blood cells. These cells are important for protecting the organism against pathogens and for establishing an effective immune response. Previous studies have shown immunomodulatory effects of different products isolated from plant extracts. This study aimed to evaluate the immunomodulatory properties of aqueous Physalis angulata (AEPa) extract on the differentiation of bone marrow cells. RESULTS: Increased cellular area, higher spreading ability and several cytoplasmatic projections were observed in the treated cells, using optical microscopy, suggesting cell differentiation. Furthermore, AEPa did not promote the proliferation of lymphocytes and polymorphonuclear leukocytes, however promotes increased the number of macrophages in the culture. The ultrastructural analysis by Transmission Electron Microscopy of treated cells showed spreading ability, high number of cytoplasmatic projections and increase of autophagic vacuoles. Moreover, a high level of LC3b expression by treated cells was detected by flow cytometry, suggesting an autophagic process. Cell surface expression of F4/80 and CD11b also indicated that AEPa may stimulate differentiation of bone marrow cells mainly into macrophages. In addition, AEPa did not differentiate cells into dendritic cells, as assessed by CD11c analysis. Furthermore, no cytotoxic effects were observed in the cells treated with AEPa. CONCLUSION: Results demonstrate that AEPa promotes the differentiation of bone marrow cells, particularly into macrophages and may hold promise as an immunomodulating agent.

Supplementation of bovine embryo culture medium with L-arginine improves embryo quality via nitric oxide production.

Nitric oxide (NO) is a cell-signaling molecule that regulates a variety of molecular pathways. We investigated the role of NO during preimplantation embryonic development by blocking its production with an inhibitor or supplementing in vitro bovine embryo cultures with its natural precursor, L-arginine, over different periods. Endpoints evaluated included blastocyst rates, development kinetics, and embryo quality. Supplementation with the NO synthase inhibitor N-Nitro-L-arginine-methyl ester (L-NAME) from Days 1 to 8 of culture decreased blastocyst (P < 0.05) and hatching (P < 0.05) rates. When added from Days 1 to 8, 50 mM L-arginine decreased blastocyst rates (P < 0.001); in contrast, when added from Days 5 to 8, 1 mM L-arginine improved embryo hatching rates (P < 0.05) and quality (P < 0.05) as well as increased POU5F1 gene expression (P < 0.05) as compared to the untreated control. Moreover, NO levels in the medium during this culture period positively correlated with the increased embryo hatching rates and quality (P < 0.05). These data suggest exerts its positive effects during the transition from morula to blastocyst stage, and that supplementing the embryo culture medium with L-arginine favors preimplantation development of bovine embryos.