Characterization of protein cargo of Echinococcus granulosus extracellular vesicles in drug response and its influence on immune response

Abstract

Background: The Echinococcus granulosus sensu lato species complex causes cystic echinococcosis, a zoonotic disease of medical importance. Parasite-derived small extracellular vesicles (sEVs) are involved in the interaction with hosts intervening in signal transduction related to parasite proliferation and disease pathogenesis. Although the characteristics of sEVs from E. granulosus protoscoleces and their interaction with host dendritic cells (DCs) have been described, the effect of sEVs recovered during parasite pharmacological treatment on the immune response remains unexplored. Methods: Here, we isolated and characterized sEVs from control and drug-treated protoscoleces by ultracentrifugation, transmission electron microscopy, dynamic light scattering, and proteomic analysis. In addition, we evaluated the cytokine response profile induced in murine bone marrow-derived dendritic cells (BMDCs) by qPCR. Results: The isolated sEVs, with conventional size between 50 and 200 nm, regardless of drug treatment, showed more than 500 cargo proteins and, importantly, 20 known antigens and 70 potential antigenic proteins, and several integral-transmembrane and soluble proteins mainly associated with signal transduction, immunomodulation, scaffolding factors, extracellular matrix-anchoring, and lipid transport. The identity and abundance of proteins in the sEV-cargo from metformin- and albendazole sulfoxide (ABZSO)-treated parasites were determined by proteomic analysis, detecting 107 and eight exclusive proteins, respectively, which include proteins related to the mechanisms of drug action. We also determined that the interaction of murine BMDCs with sEVs derived from control parasites and those treated with ABZSO and metformin increased the expression of pro-inflammatory cytokines such as IL-12 compared to control cells. Additionally, protoscolex-derived vesicles from metformin treatments induced the production of IL-6, TNF-α, and IL-10. However, the expression of IL-23 and TGF-β was downregulated. Conclusions: We demonstrated that sEV-cargo derived from drug-treated E. granulosus protoscoleces have immunomodulatory functions, as they enhance DC activation towards a type 1 pro-inflammatory profile against the parasite, and therefore support the proposal of a new approach for the prevention and treatment of secondary echinococcosis.