Leishmania donovani mitochondrial iron superoxide dismutase A is released into the cytosol during miltefosine induced programmed cell death

Trypanosoma cruzi undergo PCD (programmed cell death) under appropriate stimuli, the mechanisms of which remain to be established. In the present study, we show that stimulation of PCD in T. cruzi epimastigotes by FHS (fresh human serum) results in rapid (<1 h) externalization of phosphatidylserine and depletion of the low molecular mass thiols dihydrotrypanothione and glutathione. Concomitantly, enhanced generation of oxidants was established by EPR and immuno-spin trapping of radicals using DMPO (5,5-dimethylpyrroline-N-oxide) and augmentation of the glucose flux through the pentose phosphate pathway. In the early period (<20 min), changes in mitochondrial membrane potential and inhibition of respiration, probably due to the impairment of ADP/ATP exchange with the cytosol, were observed, conditions that favour the generation of O2•−. Accelerated rates of mitochondrial O2•− production were detected by the inactivation of the redox-sensitive mitochondrial aconitase and by oxidation of a mitochondrial-targeted probe (MitoSOX). Importantly, parasites overexpressing mitochondrial FeSOD (iron superoxide dismutase) were more resistant to the PCD stimulus, unambiguously indicating the participation of mitochondrial O2•− in the signalling process. In summary, FHS-induced PCD in T. cruzi involves mitochondrial dysfunction that causes enhanced O2•− formation, which leads to cellular oxidative stress conditions that trigger the initiation of PCD cascades; moreover, overexpression of mitochondrial FeSOD, which is also observed during metacyclogenesis, resulted in cytoprotective effects.

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Expression of a Mitochondrial Peroxiredoxin Prevents Programmed Cell Death in Leishmania donovani

Eukaryotic Cell ◽

10.1128/ec.5.5.861-870.2006 ◽

2006 ◽

Vol 5 (5) ◽

pp. 861-870 ◽

Cited By ~ 34

Author(s):

Simone Harder ◽

Meike Bente ◽

Kerstin Isermann ◽

Iris Bruchhaus

Keyword(s):

Cell Death ◽

Programmed Cell Death ◽

Mitochondrial Membrane ◽

Leishmania Donovani ◽

Physiological Role ◽

Logarithmic Phase ◽

Insect Vector ◽

Oxygen Species ◽

Late Logarithmic Phase

ABSTRACT Leishmania promastigote cells transmitted by the insect vector get phagocytosed by macrophages and convert into the amastigote form. During development and transformation, the parasites are exposed to various concentrations of reactive oxygen species, which can induce programmed cell death (PCD). We show that a mitochondrial peroxiredoxin (LdmPrx) protects Leishmania donovani from PCD. Whereas this peroxiredoxin is restricted to the kinetoplast area in promastigotes, it covers the entire mitochondrion in amastigotes, accompanied by dramatically increased expression. A similar change in the expression pattern was observed during the growth of Leishmania from the early to the late logarithmic phase. Recombinant LdmPrx shows typical peroxiredoxin-like enzyme activity. It is able to detoxify organic and inorganic peroxides and prevents DNA from hydroxyl radical-induced damage. Most notably, Leishmania parasites overexpressing this peroxiredoxin are protected from hydrogen peroxide-induced PCD. This protection is also seen in promastigotes grown to the late logarithmic phase, also characterized by high expression of this peroxiredoxin. Apparently, the physiological role of this peroxiredoxin is stabilization of the mitochondrial membrane potential and, as a consequence, inhibition of PCD through removal of peroxides.

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Characterization of Metacaspases with Trypsin-Like Activity and Their Putative Role in Programmed Cell Death in the Protozoan Parasite Leishmania

Eukaryotic Cell ◽

10.1128/ec.00123-07 ◽

2007 ◽

Vol 6 (10) ◽

pp. 1745-1757 ◽

Cited By ~ 91

Author(s):

Nancy Lee ◽

Sreenivas Gannavaram ◽

Angamuthu Selvapandiyan ◽

Alain Debrabant

Keyword(s):

Hydrogen Peroxide ◽

Cell Death ◽

Programmed Cell Death ◽

Leishmania Donovani ◽

Protozoan Parasite ◽

Protein Band ◽

Mrna Levels ◽

Enzymatic Assays ◽

Rich Domain ◽

Axenic Amastigotes

ABSTRACT In this report, we have characterized two metacaspases of Leishmania donovani, L. donovani metacaspase-1 (LdMC1) and LdMC2. These two proteins show 98% homology with each other, and both contain a characteristic C-terminal proline-rich domain. Both genes are transcribed in promastigotes and axenic amastigotes of L. donovani; however, LdMC1 shows increased mRNA levels in axenic amastigotes. An anti-LdMC antibody was obtained and showed reactivity with a single ∼42-kDa protein band in both promastigote and axenic amastigote parasite whole-cell lysates by Western blotting. Pulse-chase experiments suggest that LdMCs are not synthesized as proenzymes, and immunofluorescence studies show that LdMCs are associated with the acidocalcisome compartments of L. donovani. Enzymatic assays of immunoprecipitated LdMCs show that native LdMCs efficiently cleave trypsin substrates and are unable to cleave caspase-specific substrates. Consistently, LdMC activity is insensitive to caspase inhibitors and is efficiently inhibited by trypsin inhibitors, such as leupeptin, antipain, and N α-tosyl-l-lysine-chloromethyl ketone (TLCK). In addition, our results show that LdMC activity was induced in parasites treated with hydrogen peroxide, a known trigger of programmed cell death (PCD) in Leishmania and that parasites overexpressing metacaspases are more sensitive to hydrogen peroxide-induced PCD. These findings suggest that Leishmania metacaspases are not responsible for the caspase-like activities reported in this organism and suggest a possible role for LdMCs as effector molecules in Leishmania PCD.

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In vitro antileishmanial activity and iron superoxide dismutase inhibition of arylamine Mannich base derivatives

Parasitology ◽

10.1017/s0031182017001123 ◽

2017 ◽

Vol 144 (13) ◽

pp. 1783-1790 ◽

Cited By ~ 5

Author(s):

ALVARO MARTIN-MONTES ◽

MERY SANTIVAÑEZ-VELIZ ◽

ELSA MORENO-VIGURI ◽

RUBÉN MARTÍN-ESCOLANO ◽

CARMEN JIMÉNEZ-MONTES ◽

...

Keyword(s):

Superoxide Dismutase ◽

Leishmania Donovani ◽

Mannich Base ◽

Neglected Diseases ◽

Leishmania Braziliensis ◽

Reference Drug ◽

Iron Superoxide Dismutase ◽

Therapeutic Tools

SUMMARYLeishmaniasis is one of the world's most neglected diseases, and it has a worldwide prevalence of 12 million. There are no effective human vaccines for its prevention, and treatment is hampered by outdated drugs. Therefore, research aiming at the development of new therapeutic tools to fight leishmaniasis remains a crucial goal today. With this purpose in mind, we present 20 arylaminoketone derivatives with a very interesting in vitro and in vivo efficacy against Trypanosoma cruzi that have now been studied against promastigote and amastigote forms of Leishmania infantum, Leishmania donovani and Leishmania braziliensis strains. Six out of the 20 Mannich base-type derivatives showed Selectivity Index between 39 and 2337 times higher in the amastigote form than the reference drug glucantime. These six derivatives affected the parasite infectivity rates; the result was lower parasite infectivity rates than glucantime tested at an IC25 dose. In addition, these derivatives were substantially more active against the three Leishmania species tested than glucantime. The mechanism of action of these compounds has been studied, showing a greater alteration in glucose catabolism and leading to greater levels of iron superoxide dismutase inhibition. These molecules could be potential candidates for leishmaniasis chemotherapy.

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