Regular Article
Leishmaniaspp.: Mechanisms of Toxicity of Nitrogen Oxidation Products

https://doi.org/10.1006/expr.1997.4205Get rights and content

Abstract

Intracellular killing ofLeishmaniaparasites within activated murine macrophages is thought to result from the toxic activities of nitrogen oxidation products (referred to as NO) released by the activated cells. In order to determine possible mechanisms of NO toxicity for these microorganisms, promastigotes ofLeishmania majorandLeishmania enriettiiwere exposed to NO generated chemically from acidified nitrite,S-nitrosocysteine, diethylamine NONOate, or nitroprusside. Treatment with these agents led to loss of viability (as determined from decreased motility and inhibition of [3H]TdR uptake upon reincubation in NO-free medium) with kinetics characteristic for each compound.L. majorwas less sensitive to these effects thanL. enriettii,and amastigotes displayed the same sensitivity as promastigotes of the same species. The early effects of NO toxicity could be detected within minutes of exposure to the NO donors; they included decreased respiration rate and inhibition of glucose, proline, and adenine incorporation. Inhibition of the activities of glyceraldehyde 3-phosphate dehydrogenase and of aconitase were also evidenced. In order to determine whether these phenomena reflected the mechanisms of toxicity ofbona fideNO generated by macrophages, promastigotes were exposed to IFN-γ + LPS-activated macrophages across permeable membranes. This resulted in marked inhibition of proline and adenine uptake in the parasites, which was restored, however, to control levels when macrophages were activated in the presence of the nitric oxide synthase inhibitor NGMMA. These results indicate that several cellular targets may be subject to NO toxicity inLeishmaniaparasites, including enzymes of glycolysis and respiratory metabolism as well as trans-membrane transport systems.

References (58)

  • J.M. May

    Differential labeling of the erythrocyte hexose carrier byN

    Biochimica et Biophysica Acta

    (1989)
  • S. Mohr et al.

    Mechanism of covalent modification of glyceraldehyde-3-phosphate dehydrogenase at its active site thiol by nitric oxide, peroxynitrite and related nitrosating agents

    FEBS Letters

    (1994)
  • S. Mohr et al.

    Posttranslational modification of glyceraldehyde-3-phosphate dehydrogenase by S-nitrosylation and subsequent NADH attachment

    Journal of Biological Chemistry

    (1996)
  • M.A. Munoz-Fernandez et al.

    Activation of human macrophages for the killing of intracellularTrypanosoma cruzi

    Immunology Letters

    (1992)
  • J. Radons et al.

    Nitric oxide toxicity in islet cells involves poly(ADP-ribose) polymerase activation and concomitant NAD+

    Biochemical and Biophysical Research Communications

    (1994)
  • C. Richter et al.

    Nitric oxide kills hepatocytes by mobilizing mitochondrial calcium

    Biochemical and Biophysical Research Communications

    (1994)
  • Y. Sugiura et al.

    Nucleotide-selective cleavage of duplex DNA by nitric oxide

    Biochemical and Biophysical Research Communications

    (1995)
  • C. Szabo et al.

    Endogenous peroxynitrite is involved in the inhibition of mitochondrial respiration in immuno-stimulated J774.2 macrophages

    Biochemical and Biophysical Research Communications

    (1995)
  • D. Zilberstein

    Transport of nutrients and ions across membranes of trypanosomatid parasites

    Advances in Parasitology

    (1993)
  • D. Zilberstein et al.

    Glucose transport inLeishmania donovani

    Molecular and Biochemical Parasitology

    (1984)
  • J.A. Alspaugh et al.

    Inhibition ofCryptococcus neoformans

    Infection and Immunity

    (1991)
  • J.S. Beckman et al.

    Nitric oxide and peroxynitrite

    Methods in Nitric Oxide Research

    (1996)
  • F.T. Bonner et al.

    The chemistry of nitric oxide and redox-related species

  • J.C. Drapier et al.

    Murine cytotoxic activated macrophages inhibit aconitase in tumor cells. Inhibition involves the iron-sulfur prosthetic group and is reversible

    Journal of Clinical Investigation

    (1986)
  • J.C. Drapier et al.

    Differentiation of murine macrophages to express nonspecific cytotoxicity for tumor cells results inl

    Journal of Immunology

    (1988)
  • T.G. Evans et al.

    Effect of in vivo inhibition of nitric oxide production in murine leishmaniasis

    Journal of Immunology

    (1993)
  • M. Feelisch et al.

    Donors of nitrogen oxides

  • B. Gaston et al.

    Endogenous nitrogen oxides and bronchodilatorS

    Proceedings of the National Academy of Sciences of the United States of America

    (1993)
  • Cited by (53)

    • Nitric oxide-loaded chitosan nanoparticles as an innovative antileishmanial platform

      2019, Nitric Oxide - Biology and Chemistry
      Citation Excerpt :

      Thus, we hypothesize that the use of a higher dose (400 μM) could support parasite reduction for longer periods. This hypothesis was confirmed and we observed a noticeable and sustained promastigote killing even after 5 days, which corroborates with other studies, where high levels of NO were necessary to promote an efficient antiparasitic activity [43–45]. Before evaluation of drug efficiency against intracellular amastigotes, we evaluated the toxicity of NONPs on macrophages.

    • Leishmaniasis drug discovery: recent progress and challenges in assay development

      2017, Drug Discovery Today
      Citation Excerpt :

      Disadvantages include the need for specialised equipment and high running costs of flow-cytometry-based assays with limited capability for compound screening. To assess parasite viability, a 3[H] thymidine uptake assay was established for Leishmania enriettii and L. major promastigotes [89]. New strands of chromosomal DNA incorporate 3[H] thymidine during mitotic cell division.

    • Induction, Propagation, and Activity of Host Nitric Oxide: Lessons from Leishmania Infection

      2015, Trends in Parasitology
      Citation Excerpt :

      Moreover, NO cytotoxic effect in trans could explain how parasites are controlled by NO in fibroblasts that do not express iNOS during the latency phase of Leishmania infection [59]. Several studies have investigated the molecular mechanisms underlying NO cytotoxic activity on Leishmania parasites [60,61]. Addition of authentic NO gas or NO donors (such as acidified nitrite or S-nitroso-cysteines) in axenic promastigote or amastigote cultures leads to a rapid loss of parasite viability and to DNA fragmentation.

    • Role of trypanosomatid's arginase in polyamine biosynthesis and pathogenesis

      2012, Molecular and Biochemical Parasitology
      Citation Excerpt :

      A different approach is to use NO or nitrosating compounds or NOS2-inducing agents. There is some evidence that sodium nitroprusside – a relatively strong nitrosating agent – acts as a nitric oxide (NO) donor, reducing L. amazonensis viability in vitro [124,125]. The second approach is more interesting and would prevail macrophages in a manner similar to the way Th1 activation acts to destroy the parasite burden.

    View all citing articles on Scopus

    M. FeelishJ. S. Stamler, Eds.

    1

    To whom correspondence should be addressed. Fax: ++41 21 692 57 05.

    View full text