Synthesis of Nanoparticles Using Euphorbia prostrata Extract Reveals a Shift from Apoptosis to G0/G1 Arrest in Leishmania donovani

The aim of the present investigation was to synthesize silver (Ag) and titanium dioxide (TiO2) nanoparticles (NPs) using the aqueous leaves extract of Euphorbia prostrata as antileishmanial agents and to explore the mechanism of induced cell death. In vitro antileishmanial activity of synthesized NPs was tested against promastigotes of Leishmania donovani by alamar Blue® cell viability reagent and propidium iodide uptake assay. The effective leishmanicidal activity of synthesized Ag NPs was further confirmed by cell cycle progression, externalized phosphatidylserine, DNA fragmentation assay, reactive oxygen species (ROS) level, intracellular non-protein thiols and transmission electron microscopy (TEM) of the treated parasites. TEM analysis of the synthesized Ag NPs and TiO2 NPs showed spherical shape with an average size of 12.82 ± 2.50 and 83.22 ± 1.50 nm, respectively. Ag NPs was found to be the most active agent against Leishmania parasites after 24 h exposure with IC50 value of 14.94 μg/mL. A significant increase in G0/ G1 phase of the cell cycle with subsequent decrease in S and G2/M phases was observed when compared to control and thus confirming the growth inhibitory effect of synthesized Ag NPs. Decreased ROS level was also observed which could be responsible for caspase independent shift from apoptosis (G0/G1 arrest) to massive necrosis. High molecular weight DNA fragmentation as a positive consequence of necrotic cell death was also visualized. In the present study, the unique trypanothione/trypanothione reductase (TR) system of Leishmania cells was significantly inhibited by synthesized Ag NPs was reported. The green synthesized Ag NPs may provide promising leads for the development of cost effective and safer alternative treatment against visceral leishmaniasis.