concanavalin-a and Leishmaniasis

Traditional medicine and scientific studies have shown that the raw extract of Evanta [Galipea longiflora, Angostura longiflora (Krause) Kallunki] exhibits anti-leishmanial activity. We hypothesized that the healing observed when using this plant might not only be due to the direct action on the parasite, but possibly to a parallel effect on the host immune response to the parasite involved in the healing process. We show here that an alkaloid extract of Evanta (AEE) directly killed the parasite already at a dose of 10 microg/ml, but at this low concentration, AEE did not have a major effect on viability and proliferation of eukaryotic cells. The whole extract was also found to be stronger than 2-phenylquinoline, the most prominent alkaloid in AEE. AEE was not directly stimulating B or T cells or J774 macrophages. However, it interfered with the activation of both mouse and human T cells, as revealed by a reduction of in vitro cellular proliferation and interferon-gamma (IFN-gamma) production. The effect was more evident when the cells were pretreated with AEE and subsequently stimulated with the polyclonal T-cell activators Concanavalin A and anti-CD3. Taken together, our results suggest that Evanta have a direct leishmanicidal effect and due to the effect on IFN-gamma production it might contribute to control the chronic inflammatory reaction that characterize Leishmania infection pathology, but in vivo studies are necessary to corroborate this finding.

Topics: Animals; Antiprotozoal Agents; Cell Line; Cell Survival; Concanavalin A; Female; Humans; Immunoglobulins; Interferon-gamma; Leishmania braziliensis; Leishmaniasis; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Nitric Oxide; Plant Extracts; Quinolines; Rutaceae; T-Lymphocytes; Tumor Necrosis Factor-alpha

The responsiveness of peripheral blood lymphocytes from 15 cutaneous leishmaniasis patients and five healthy volunteers to purified surface antigens of Leishmania was studied. The following purified antigens were independently used as stimuli for lymphocyte proliferation in vitro: lipophosphoglycan of L. braziliensis, (Lb-LPG), lipophosphoglycan of L. mexicana (Lm-LPG), glycoprotein 63 of L. braziliensis (Lb-gp 63) and glycoprotein 63 of L. mexicana (Lm-gp 63). Among six patients tested with Lb-gp 63 and Lm-gp 63, two borderline responses and two positive responses were observed, respectively, all the others being negative. Lb-LPG and Lm-LPG induced positive responses in all patients tested (11 with Lb-LPG and five with Lm-LPG). A positive dose-response correlation was observed in the responses of the patients to both Lb-LPG and Lm-LPG. Significant differences (P less than 0.01) were found between the two groups (patients and healthy subjects) with regard to the responses to Lb-LPG. The proliferating cells after stimulation with Lb-LPG were shown to belong to the CD4+ T cell subset by flow cytometry. However, the following evidence suggests that the T cell responses were not induced by LPG itself but rather by protein contaminants in the LPG preparations: a highly purified Lb-LPG batch that had undergone additional purification induced positive responses in cells from only three out of six patients and a proteinase K-treated Lb-LPG batch did not stimulate any positive response among the same six patients.

Topics: Adult; Animals; Antigens, Protozoan; Concanavalin A; Female; Glycosphingolipids; Humans; Immunophenotyping; Leishmania; Leishmania braziliensis; Leishmania mexicana; Leishmaniasis; Lymphocyte Activation; Male; Metalloendopeptidases; Protozoan Proteins; T-Lymphocytes

Leishmania major infection in genetically susceptible BALB/c mice is associated with the development of chronic primary lesions as well as multiple metastatic lesions. Spleen cells from these mice were shown to have depressed in vitro responses to concanavalin A (Con A) that coincided with the development of indomethacin-sensitive suppressor cells. Depressed responses to Con A were noted as early as 1 wk after parasite inoculation and correlated with the increased production of prostaglandin E2 (PGE2) by spleen cells from infected mice. Mice induced by prior irradiation (550 rad) to heal infection did not develop indomethacin-reversible depression in responsiveness to Con A. Although macrophages appear to be the major source of PGE2 production, in vitro studies indicate that infection per se is not a sufficient stimulus to initiate prostaglandin (PG) synthesis, suggesting the involvement of other cell types. Mice treated in vivo with indomethacin exhibited significantly fewer metastatic lesions than control mice, suggesting that PG may play a role in the exacerbation of cutaneous disease in these animals.

Topics: Animals; Arachidonic Acid; Arachidonic Acids; Concanavalin A; Dinoprostone; Female; Immune Tolerance; Indomethacin; Leishmaniasis; Lymphocyte Activation; Macrophages; Mice; Mice, Inbred BALB C; Prostaglandins; Prostaglandins E; Spleen; T-Lymphocytes

The specific immune response of patients with cutaneous leishmaniasis including the ability of their lymphokines to enhance the monocytes' leishmaniacidal activity was studied. In 16 patients with cutaneous leishmaniasis, their concanavalin A-induced lymphocyte proliferative responses, interferon-gamma and interleukin 2 activities and the ability of their concanavalin A-induced lymphokines to kill monocyte intracellular amastigotes were not different from normal controls. Antigen-stimulated lymphocyte cultures showed that 13 of 13 patients had an increased lymphocyte proliferative response; 11 of 16 produced interleukin 2 and 12 of 13 produced interferon-gamma; in addition, 10 of 11 of these antigen-induced supernatants increased the monocytes' killing of Leishmania major amastigotes. Antibody levels to parasite membrane antigens determined by radioimmunoassay showed that 8 of 13 titers were greater than 10 and 4 of 13 titers were 2 to 10 times higher than control. Our findings demonstrate that patients with cutaneous leishmaniasis elicit a specific immune response to L. major antigens and part of this response is the production of lymphokine capable of promoting monocyte killing of intracellular amastigotes.

Topics: Animals; Antibodies, Protozoan; Antigens, Protozoan; Concanavalin A; Humans; Interferon-gamma; Interleukin-2; Leishmania tropica; Leishmaniasis; Leukocytes, Mononuclear; Lymphocyte Activation

Differentiation of Leishmania major promastigotes from a noninfective to an infective stage has been demonstrated for promastigotes growing within axenic culture and within the sandfly vector. We have been attempting to identify specific biochemical or antigenic changes that are associated with the development of infective-stage promastigotes. In this report we demonstrate that during growth, cultured L. major promastigotes undergo selective changes in surface carbohydrates, determined by their agglutination by plant lectins. Thus, although all promastigotes from logarithmic (log)-phase cultures were agglutinated by the two-D-galactose-binding lectins, peanut agglutinin (PNA) and Ricinus communis, identical concentrations of these lectins failed to agglutinate approximately 50% of L. major promastigotes from the stationary-phase cultures. These changes in lectin-agglutinating properties are consistent with the fact that log-phase promastigotes represent a homogeneous population of noninfective parasites, whereas up to 50% of the stationary-phase organisms appear to be transformed into infective-stage promastigotes, as determined by their ability to survive within normal resident mouse peritoneal macrophages in vitro. The identities of the populations defined by infectivity and PNA agglutination were confirmed by the purification of PNA-unagglutinated promastigotes from stationary-phase cultures, which demonstrated that 100% of these promastigotes were able to establish intracellular infections. Lectin-purified, infective-stage promastigotes from the stationary phase were compared with noninfective promastigotes from the log phase for the purpose of identifying stage-specific antigens. On the basis of Western blot analysis and the immunoprecipitation of surface-labeled organisms, we have identified an antigen of roughly 116,000 Mr that is expressed on the surface of infective but not noninfective promastigotes.

Topics: Agglutination Tests; Animals; Antigens, Protozoan; Antigens, Surface; Autoradiography; Carbohydrate Metabolism; Concanavalin A; Epitopes; Humans; Lectins; Leishmania; Leishmaniasis; Mice; Mice, Inbred C57BL; Molecular Weight; Peanut Agglutinin; Plant Lectins; Rabbits; Soybean Proteins

Topics: Animals; Concanavalin A; Female; Immunoglobulin M; Leishmania; Leishmaniasis; Lymphocyte Activation; Lymphocyte Culture Test, Mixed; Macrophage Activation; Mice; Phytohemagglutinins

Mechanisms possibly involved in the regulation of the immune response were evaluated in 49 patients with American cutaneous leishmaniasis (ACL). The patients were classified on the basis of clinical and histopathological criteria as suffering localized (LCL), mucocutaneous (MCL) or diffuse (DCL) forms of the disease. A significant leishmanial antigen-induced suppression of in vitro mitogen responsiveness was demonstrated in the DCL group, but not in the other two diseases states. Lack of suppressive activity was particularly evident in MCL, this being the group that presented the highest in vivo and in vitro reactivity to the parasite antigens. In fact, a significant inverse correlation was found between the degree of suppression and the antigen-induced lymphocyte proliferative response. In contrast, a mixture of mononuclear cells from MCL patients and normal subjects showed higher that expected responses to mitogen, while this increase was not observed in co-cultures of DCL and normal mononuclear cells. Due to their possible modulatory influences, circulating immune complexes were also evaluated in these patient groups, higher levels being found in MCL and DCL patients than in either LCL or controls. The possible mechanisms involved in the regulation of the immune response to the protozoan in the complex disease spectrum of ACL are discussed in relation to anergy in DCL and hyperresponsiveness in MCL.

Topics: Antigen-Antibody Complex; Antigens; Cell Division; Concanavalin A; Female; Humans; Leishmania; Leishmaniasis; Leishmaniasis, Mucocutaneous; Lymphocyte Activation; Male; Phytohemagglutinins

Leishmania organisms are obligate intracellular parasites of mammalian mononuclear phagocytes in vivo. In order to study the interactions of these parasites and mononuclear phagocytes, we have used a model of infection of Leishmania major in human monocytes in vitro. The presence of intracellular parasites did not alter the normal secretion of lysozyme or result in increased secretion of prostaglandin E2 (PGE2) or superoxide anion by the monocytes. Addition of concanavalin A (Con A), which binds to a specific membrane receptor, zymosan particles or endotoxin to infected monocyte monolayers, resulted in the expected increase in PGE2 secretion. In addition, the production of superoxide by infected monocytes treated with phorbol myristate acetate was not different from control uninfected cultures. Despite this evidence of biochemical activation, neither endotoxin, zymosan nor Con A had any parasiticidal effect on the intracellular parasites. In contrast, Con A-induced lymphokines from human mononuclear cells resulted in an increased killing of the intracellular amastigotes. These studies have shown that the induction of leishmaniacidal capacity of human monocytes is dependent on the type of stimulus used to induce activation.

Topics: Cells, Cultured; Concanavalin A; Dinoprostone; Endotoxins; Humans; Leishmania; Leishmaniasis; Lymphokines; Macrophage Activation; Monocytes; Muramidase; Prostaglandins E; Zymosan

Topics: Animals; Concanavalin A; Immunosuppression Therapy; Indomethacin; Leishmania; Leishmaniasis; Lymphocyte Activation; Lymphocytes; Mice; Mice, Inbred BALB C

Leishmania tropica in BALB/c mice causes a fatal infection accompanied by the development of multiple metastatic lesions. Spleen cells from these mice were shown to have depressed proliferative responses to concanavalin A (Con A), phytohemagglutinin (PHA), and lipopolysaccharide (LPS). Coinciding with this immunodepression was the development of a cell population capable of suppressing normal spleen cell responses to Con A. This suppressor cell activity was first observed at 6 wk and was present throughout the remainder of the infection. At 12 wk the suppressor cells could be removed by Sephadex G-10 passage or carbonyl iron treatment; however, Sephadex G-10 passage could not reverse the suppression at 18 wk. Indomethacin, a prostaglandin synthetase inhibitor, was found to abrogate the activity of the adherent suppressor cell, suggesting that prostaglandin production may be involved in the immunosuppression seen in these mice. In addition, Sephadex G-10 passage and indomethacin were found to markedly augment spleen cell responses to leishmanial antigen, indicating that the adherent suppressor cell is capable of regulating specific immunologic responses.

Topics: Animals; Antigens; Chromatography, Gel; Concanavalin A; Female; Immunosuppression Therapy; Indomethacin; Leishmaniasis; Lipopolysaccharides; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Phytohemagglutinins; Spleen; T-Lymphocytes, Regulatory

Amastigotes from Leishmania braziliensis and L. mexicana were isolated from cutaneous lesions in infected animals using the plant lectin Concanavalin A as a specific agglutination agent. Amastigotes were collected in preparations of up to 95% purity as determined by cell count. The parasites obtained by this method showed no apparent loss of viability as measured by growth rated and DNA replication, or pathogenicity as measured by routine passages in hamsters or mice.

Topics: Agglutination; Animals; Concanavalin A; Cricetinae; DNA; Humans; Leishmania; Leishmaniasis; Mice; Microbiological Techniques; Skin

Topics: Animals; Concanavalin A; Leishmania; Leishmaniasis; Lymphocytes; Lymphokines; Macrophages; Mice; Mice, Inbred Strains; Species Specificity