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Induction of apoptosis by the triterpene saponin PX-6518 in promastigotes and amastigotes of Leishmania infantum
Aim: Current first-line medication of visceral leishmaniasis is increasingly faced with levels of resistance and therefore new drugs are still needed . The 13,28 epoxy-oleanane saponin PX-6518 showed strong antileishmania properties in vivo . Because little is known about its mode-of-action, the induction of apoptosis was studied on promastigotes and extracellular and intracellular amastigotes of L. infantum. Miltefosin, for which apoptosis-inducing action was already demonstrated , was used as reference.
Methods: Early apoptosis, characterized by phosphatidylserine (PS) exposure from the inner to the outer side of the cell membrane, was evaluated by Annexin V-FITC/PI labeling (Ann/PI) in treated promastigotes and extracellular amastigotes. Differential sensitivities of axenically grown and fresh ex vivo amastigotes were compared. Microscopic and flow cytometric follow-up studies included DNA fragmentation in oligonucleosomal fragments using a Tunel assay. The effects on intracellular amastigotes were studied in J774 and primary peritoneal macrophages.
Results: Ann/PI evaluation of PX-6518 treated host cells demonstrated signs of apoptosis induction, whereas treatment of promastigotes and axenic amastigotes did not induce PS exposure. Evaluation of ex vivo amastigotes was hampered by the fact that a large proportion of untreated control cells were Ann-positive and PI-negative. This result confirms earlier reports of apoptotic mimicry . No additional effects of PX-6518 could be observed in the small fraction of Ann+, PI- amastigotes. These initial observations are now studied in greater detail using the Tunel assay.
Conclusion: Features of early apoptosis were observed after treatment of macrophages with PX-6518. The effects on promastigotes and extracellular amastigotes were marginal.
References: 1. Croft S. et al. (2003) Trends Parasitol 19: 502–508 2. Maes L. et al. (2004) Antimicrob Agents Chemother 48: 2056–2060 3. Sudhandiran G. et al. (2003)J Biol Chem 278: 25120–25132 4. Wanderley J. et al. (2006)J Immunol 176: 1834–1839.