Mode of Action of the Sesquiterpene Lactones Psilostachyin and Psilostachyin C on Trypanosoma cruzi

Abstract

Trypanosoma cruzi is the causative agent of Chagas’ disease, which is a major endemic disease in Latin America and is recognized by the WHO as one of the 17 neglected tropical diseases in the world. Psilostachyin and psilostachyin C, two sesquiterpene lactones isolated from Ambrosia spp., have been demonstrated to have trypanocidal activity. Considering both the potential therapeutic targets present in the parasite, and the several mechanisms of action proposed for sesquiterpene lactones, the aim of this work was to characterize the mode of action of psilostachyin and psilostachyin C on Trypanosoma cruzi and to identify the possible targets for these molecules. Psilostachyin and psilostachyin C were isolated from Ambrosia tenuifolia and Ambrosia scabra, respectively. Interaction of sesquiterpene lactones with hemin, the induction of oxidative stress, the inhibition of cruzipain and trypanothione reductase and their ability to inhibit sterol biosynthesis were evaluated. The induction of cell death by apoptosis was also evaluated by analyzing phosphatidylserine exposure detected using annexin-V/propidium iodide, decreased mitochondrial membrane potential, assessed with Rhodamine 123 and nuclear DNA fragmentation evaluated by the TUNEL assay. Both STLs were capable of interacting with hemin. Psilostachyin increased about 5 times the generation of reactive oxygen species in Trypanosoma cruzi after a 4h treatment, unlike psilostachyin C which induced an increase in reactive oxygen species levels of only 1.5 times. Only psilostachyin C was able to inhibit the biosynthesis of ergosterol, causing an accumulation of squalene. Both sesquiterpene lactones induced parasite death by apoptosis. Upon evaluating the combination of both compounds, and additive trypanocidal effect was observed. Despite their structural similarity, both sesquiterpene lactones exerted their anti-T. cruzi activity through interaction with different targets. Psilostachyin accomplished its antiparasitic effect by interacting with hemin, while psilostachyin C interfered with sterol synthesis.

Primary targets of the sesquiterpene lactone deoxymikanolide on Trypanosoma cruzi

“…Based on a previous report about the in vitro and in vivo trypanocidal activity of deoxymikanolide, this compound was selected to carry out a biochemical characterization of its mechanism of action. Even though the presence of an α-β unsaturated lactone moiety is a common structural feature among STLs to which most of their biological activities are attributed, different antiprotozoal mechanisms of action have been described for them (Sülsen et al, 2010;Sülsen et al, 2016;Wulsten et al, 2017).…”

“…The interaction of deoxymikanolide with hemin was examined spectrophotometrically under reducing and non-reducing conditions, following the methodology previously reported (Sülsen et al, 2016). The assay system under non-reducing conditions consisted of 0.23 M sodium phosphate buffer of pH 7.4, 1% SDS, 7.5 µM hemin and different concentrations of deoxymikanolide (7.5 to 45 µM).…”

“…After 24 h of treatment, control parasites (untreated and terbinafine-treated) and deoxymikanolide-treated parasites were harvested and the total lipids were extracted and analyzed as previously described (Sülsen et al, 2016). Qualitative and quantitative analyses were carried out using TLC.…”

In Vitro Anticoccidial Activity of Olive Pulp (Olea europaea L. var. Chemlal) Extract Against Eimeria Oocysts in Broiler Chickens

“…These compounds or secondary metabolites particularly possess also anti-protozoan activities, especially against Plasmodium, Leishmania and Trypanosoma spp. [46][47][48][49][50]. Then, the phenolic compounds are good candidate molecules due to a relatively low or no toxicity and because the presence of some flavonoids limits the resistance of protozoan to other drugs [51].…”

A review of the traditional use of southern African medicinal plants for the treatment of selected parasite infections affecting humans

“…The most frequently encountered mode of action studies have been towards plasmodium parasite studies, but some investigative work has been undertaken on the antitrypanosomal activities where the major target is the inhibition of enzymes responsible for the conversion of glucose to 3-phoshoglycerate and glycerol. Other natural product mode of action studies on trypansomes and schistosomiasis have also been conducted (Morello et al, 1995;Doenhoff et al, 2008;Sülsen et al, 2016), which provide a good reference point with which to perform further studies on Southern African plants.…”