The putrescine analogue 1,4-diamino-2-butanone affects polyamine synthesis, transport, ultrastructure and intracellular survival in Leishmania amazonensis

Abstract

Polyamines are important regulators of growth and differentiation in a variety of cells, including parasitic protozoa. Promastigotes of Leishmania species have high levels of putrescine and spermidine and their growth can be inhibited by polyamine biosynthesis antagonists. The putrescine analogue 1,4-diamino-2-butanone (DAB) is microbicidal against Tritrichomonas foetus and Trypanosoma cruzi, so we tested its effects on Leishmania amazonensis proliferation, viability, organization, putrescine transport and synthesis as well as in vitro infectivity. DAB impaired promastigote proliferation dose-dependently (IC 50 144 mM) and the parasite putrescine concentration was reduced by nearly 50 %. This analogue markedly inhibited both ornithine decarboxylase activity and [H 3 ]putrescine uptake by promastigotes. Pre-treatment with DAB for 24 h led to compensatory enhancement of putrescine uptake, indicating an adaptive mechanism in DAB-treated parasites. Remarkably, DAB caused mitochondrial damage, assessed by transmission electron microscopy, and 3 h treatment with 1 mM DAB enhanced lipid peroxidation, whereas incubation with 10 mM DAB or for 24 h resulted in decreased peroxidation levels in the parasites. This effect was probably due to the loss of mitochondrial function, demonstrated by the diminished reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), not observed in macrophages. Murine macrophages infected with L. amazonensis amastigotes treated with DAB had parasite loads significantly (P,0.05) lower than controls, presumably due to interference with putrescine uptake and/or synthesis. These results suggest that putrescine may be involved in leishmanial survival, possibly by maintaining the parasite's mitochondrial function. The use of analogues to interfere with polyamine/diamine synthesis and transport may shed light on its function in intracellular parasite survival and lead to identification of new targets for leishmaniasis chemotherapy.

Regulation of Gene Expression by PrrA in Rhodobacter sphaeroides 2.4.1: Role of Polyamines and DNA Topology

“…Likely, the extracellular spermidine concentration used in our experiments might have resulted in an at least transiently higher intracellular polyamine concentration, as previously reported for other organisms (74,82; also as reviewed in references 2, 41, and 78). The fact that the spermidine effects were only observed when the cells grew aerobically might be a reflection of either differential polyamine biosynthesis and transport depending on growth conditions, as inferred from microarray expression data (29) and/or the fact that slight effects on gene regulation and lightharvesting spectral complex formation might not be observed against the background of high photosynthesis gene expression under anaerobic conditions.…”

Induction of Mitochondrial Dysfunction and Oxidative Stress in Leishmania donovani by Orally Active Clerodane Diterpene

“…Subcellular alterations included the rounding up of cells, intense cytoplasmic vacuolation, pyknosis of the nucleus, structural disruption of the mitochondrion, and decondensation of the kinetoplast. Previous studies have described similar observations of mitochondrion-kinetoplast damage upon treatment in Leishmania amazonensis promastigotes with a putrescine analogue (25), the antiarrhythmic drug amiodarone (20), and a squalene synthase inhibitor, BPQ-OH (26).…”

Drug Resistance in Leishmaniasis: Current Drug-Delivery Systems and Future Perspectives

“…They also play a vital role in the regulation of lipid peroxidation caused by the immune system and drugs [41,42]. The polyamine transporters are responsible for transporting both putrescine and spermidine (polyamine) and regulating the intracellular polyamine level.…”