Microgramma vacciniifolia Frond Lectin: In Vitro Anti-leishmanial Activity and Immunomodulatory Effects Against Internalized Amastigote Forms of Leishmania amazonensis.

PURPOSE: The treatment of leishmaniasis, an anthropozoonosis caused by Leishmania protozoa, is limited by factors, such as adverse effects, toxicity, and excessive cost, which has highlighted the importance of novel drugs. In this context, natural products have been considered as sources of antileishmanial agents. This study investigated the leishmanicidal activity of Microgramma vacciniifolia frond lectin (MvFL) on promastigotes and amastigotes of Leishmania amazonensis. METHODS: The effects of MvFL on promastigote proliferation and macrophage infection by amastigotes were evaluated and mean inhibitory concentrations (IC50) were calculated. As a safety assessment, the hemolytic capacity of MvFL (6.25-200 µg/mL) against mouse and human erythrocytes was determined. Additionally, the ability of MvFL (6.25-100 µg/mL) to modulate lysosomal and phagocytic activities and the nitric oxide (NO) production by murine peritoneal macrophages was also investigated. RESULTS: After 24 h, MvFL inhibited the proliferation of L. amazonensis promastigotes, with an IC50 of 88 µg/mL; however, hemolytic activity was not observed. MvFL also reduced macrophage infection by amastigotes with an IC50 of 52 µg/mL. Furthermore, treatment with MvFL reduced the number of amastigotes internalized by infected murine peritoneal macrophages by up to 68.9% within 48 h. At a concentration of 25 µg/mL, MvFL stimulated lysosomal activity of macrophages within 72 h, but did not alter phagocytic activity or induce NO production at any of the tested concentrations. CONCLUSION: MvFL exerts antileishmanial activity and further studies are needed to assess its therapeutic potential in in vivo experimental models of leishmaniasis.

Inhibition of Carrageenan-Induced Acute Inflammation in Mice by the Microgramma vacciniifolia Frond Lectin (MvFL).

Most anti-inflammatory drugs used nowadays have an excessive cost and their prolonged use has been connected with several injurious effects. Thus, the search for new anti-inflammatory agents is increasing. Lectins are carbohydrate-interacting proteins that can modulate immune response and the release of inflammation mediators. The Microgramma vacciniifolia frond lectin (MvFL) was previously reported to be an immunomodulatory agent in vitro. This work aimed to evaluate the effects of MvFL on the in vivo inflammatory status in the carrageenan-induced peritonitis and paw edema, using female Swiss mice. The animals were pretreated intraperitoneally with MvFL (5 and 10 mg/kg). In the peritonitis assay, the total and differential migration of white blood cells was evaluated, as well as the levels of cytokines, nitric oxide (NO), and total proteins in the peritoneal fluid. In the paw edema evaluation, the paw volume was measured in the early (from 30 min-2 h) and late (3-4 h) phases of edema formation. MvFL (5 and 10 mg/kg) was efficient in reducing neutrophil infiltration, pro-inflammatory cytokines (IL-6, IL-17, and TNF-α), NO, and protein content in the peritoneal fluid. It also repressed the edema formation in the late phase of the assay. In conclusion, MvFL showed inhibitory effects in in vivo acute inflammation, which encouraged future studies exploiting its immunomodulatory ability.

Microgramma vacciniifolia (Polypodiaceae) fronds contain a multifunctional lectin with immunomodulatory properties on human cells.

In this study, we report the purification and characterization of a multifunctional lectin (MvFL) from Microgramma vacciniifolia fronds as well as its immunomodulatory properties on human peripheral blood mononuclear cells (PBMCs). MvFL (pI 4.51; 54kDa) is a glycoprotein able to inhibit trypsin activity and that has sequence similarities (32% coverage) with a plant RNA-binding protein. Hemagglutinating activity of MvFL was not altered by heating at 100°C for 30min, but was reduced in alkaline pH (8.0 and 9.0). Fluorimetric analyses showed that this lectin did not undergo marked conformational changes when heated. However, the MvFL conformation changed depending on the pH. MvFL at 6.25-25µg/mL was not cytotoxic to lymphocytes present among PBMCs. The PBMCs incubated for 24h with the lectin (12.5µg/mL) showed increased TNF-α, IFN-γ, IL-6, IL-10, and nitric oxide production. MvFL also stimulated T lymphocytes from PBMCs to differentiate into CD8+ cells. The activation (indicated by CD28 expression) of these cells was also stimulated. In conclusion, MvFL is a heat-stable and multifunctional protein, with both lectin and trypsin inhibitor activities, and capable of inducing predominantly a Th1 response in human PBMCs as well as activation and differentiation of T lymphocytes.

Toxic effects of Microgramma vacciniifolia rhizome lectin on Artemia salina, human cells, and the schistosomiasis vector Biomphalaria glabrata.

The present study evaluated the toxicity of Microgramma vacciniifolia rhizome lectin (MvRL) to Artemia salina, human tumour cell lines (larynx epidermoid carcinoma Hep-2, NCI-H292 lung mucoepidermoid carcinoma, and chronic myelocytic leukaemia K562), and normal peripheral blood mononuclear cells (PBMCs), as well as to Biomphalaria glabrata embryos and adults. MvRL was toxic to A. salina (LC50=159.9 µg/mL), and exerted cytotoxic effects on NCI-H292 cells (IC50=25.23 µg/mL). The lectin (1-100 µg/mL) did not affect the viability of K562 and Hep-2 tumour cells, as well as of PBMCs. MvRL concentration of 1, 10, and 100 µg/mL promoted malformations (mainly exogastrulation) in 7.8%, 22.5%, and 27.7% of embryos, respectively, as well as delayed embryo development in 42.0%, 69.5%, and 54.7% of embryos, respectively. MvRL at a concentration of 100 µg/mL killed B. glabrata embryos (17.7%) and adults (25%). Further, MvRL damaged B. glabrata reproductive processes, which was evidenced by observations that snails exposed to the lectin (100 µg/mL) deposited fewer eggs than those in the control group, and approximately 40% of the deposited eggs exhibited malformations. Comparison of these results with that from A. salina assay indicates that MvRL is adulticidal at the concentration range which is toxic to environment. In conclusion, the cytotoxicity of MvRL on tumour cell and absence of toxicity to normal cell indicate its potential as chemotherapeutic drug. Also, the study revealed that the lectin is able to promote deleterious effects on B. glabrata embryos at environmentally safe concentrations.