Abstract
BackgroundPathogenic protozoans use extracellular vesicles (EVs) for intercellular communication and host manipulation. Acanthamoeba castellanii is a free-living protozoan that may cause severe keratitis and fatal granulomatous encephalitis. Although several secreted molecules have been shown to play crucial roles in the pathogenesis of Acanthamoeba, the functions and components of parasite-derived EVs are far from understood.MethodsPurified EVs from A. castellanii were confirmed by electron microscopy and nanoparticle tracking analysis. The functional roles of parasite-derived EVs in the cytotoxicity to and immune response of host cells were examined. The protein composition in EVs from A. castellanii was identified and quantified by LC-MS/MS analysis.ResultsEVs from A. castellanii fused with rat glioma C6 cells. The parasite-derived EVs induced an immune response from human THP-1 cells and a cytotoxic effect in C6 cells. Quantitative proteomic analysis identified a total of 130 proteins in EVs. Among the identified proteins, hydrolases (50.2%) and oxidoreductases (31.7%) were the largest protein families in EVs. Furthermore, aminopeptidase activities were confirmed in EVs from A. castellanii.ConclusionsThe proteomic profiling and functional characterization of EVs from A. castellanii provide an in-depth understanding of the molecules packaged into EVs and their potential mechanisms mediating the pathogenesis of this parasite.
Highlights
Pathogenic protozoans use extracellular vesicles (EVs) for intercellular communication and host manipulation
Physical characterization of EVs produced by A. castellanii To test whether A. castellanii produces EVs, we isolated the fraction from cell-free medium that contained secreted proteins, followed by a series of centrifugation and concentration steps
The purified fraction of EVs was confirmed by electron microscopy (EM) (Fig. 1b) and nanoparticle tracking analysis (NTA) (Fig. 1c)
Summary
Pathogenic protozoans use extracellular vesicles (EVs) for intercellular communication and host manipulation. Acanthamoeba castellanii is a free-living protozoan that may cause severe keratitis and fatal granuloma‐ tous encephalitis. Several secreted molecules have been shown to play crucial roles in the pathogenesis of Acanthamoeba, the functions and components of parasite-derived EVs are far from understood. Acanthamoeba spp. are free-living amoeba and distributed in diverse natural environments, including water and soil. Acanthamoeba is capable of causing a rare but fatal encephalitis known as granulomatous amoebic meningoencephalitis (GAE) [1] or a painful and sightthreatening corneal infection termed Acanthamoeba keratitis (AK) [2]. It is essential to further elucidate the mechanism of Acanthamoeba spp. pathogenesis and develop chemotherapeutic agents to combat Acanthamoeba infections
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