Extracellular vesicles from Echinococcus granulosus larval stage: Isolation, characterization and uptake by dendritic cells

Abstract

The secretion of extracellular vesicles (EVs) in helminth parasites is a constitutive mechanism that promotes survival by improving their colonization and adaptation in the host tissue. In the present study, we analyzed the production of EVs from supernatants of cultures of Echinococcus granulosus protoscoleces and metacestodes and their interaction with dendritic cells, which have the ability to efficiently uptake and process microbial antigens, activating T lymphocytes. To experimentally increase the release of EVs, we used loperamide, a calcium channel blocker that increases the cytosolic calcium level in protoscoleces and EV secretion. An exosome-like enriched EV fraction isolated from the parasite culture medium was characterized by dynamic light scattering, transmission electron microscopy, proteomic analysis and immunoblot. This allowed identifying many proteins including: small EV markers such as TSG101, SDCBP, ALIX, tetraspanins and 14-3-3 proteins; proteins involved in vesicle-related transport; orthologs of mammalian proteins involved in the immune response, such as basigin, Bp29 and maspardin; and parasite antigens such as antigen 5, P29 and endophilin-1, which are of special interest due to their role in the parasite-host relationship. Finally, studies on the EVs-host cell interaction demonstrated that E. granulosus exosome-like vesicles were internalized by murine dendritic cells, inducing their maturation with increase of CD86 and with a slight down-regulation in the expression of MHCII molecules. These data suggest that E. granulosus EVs could interfere with the antigen presentation pathway of murine dendritic cells inducing immunoregulation in the host. Further studies are needed to better understand the role of these vesicles in parasite survival and as diagnostic markers and new vaccines.

Characterization of exosome-like vesicles derived from Cysticercus pisiformis and their immunoregulatory role in macrophages

“…The particle size distribution of the vesicles was distributed around 50-150 nm and peaked at a mean diameter of 98.47 nm (Fig. 1b), which had the prototypical size characteristic of exosome-like vesicles and was consistent with exosomes from other parasites [11,24,28,[36][37][38][39]. This data indicated that we successfully isolated and purified exosomelike vesicles from C. pisiformis.…”

“…Recently, helminthes-derived exosomes or extracellular vesicles (EVs) have been proved a new paradigm in the study of parasite-host communication [7,8]. Exosomes are nano-scale (30-150 nm) membrane-bound vesicles that are released into the extracellular environment via the fusion of the plasma membrane with the membrane of multi-vesicular bodies (MVBs) [9][10][11]. These vesicles are produced by normal [12] and pathological cells [13] and can be obtained from extracellular fluids, including urine [14], blood plasma [15], breast milk [16], saliva [17], and cerebrospinal fluid [18].…”

“…Exosome-like vesicles from helminthes may play a pivotal role in parasitic infection [23][24][25][26]. Recent studies of several cestodes have revealed that exosome-like vesicles from parasites can deliver miRNA or protein cargo into host cells to modulate the host immune system [11,27]. Echinococcus granulosus cyst fluids were enriched in some exosome-enriched EVs and specific proteins with host or parasite origins, some of which were associated with cyst survival [28].…”

Characterization of exosome-like vesicles derived from Taenia pisiformis cysticercus and their immunoregulatory role on macrophages

“…The particle size distribution of the vesicles was distributed around 50-150 nm and peaked at a mean diameter of 98.47 nm (Fig. 1b), which had the prototypical size characteristic of exosome-like vesicles and was consistent with exosomes from other parasites [10,23,27,[38][39][40]. This data indicated that we successfully isolated and puri ed exosome-like vesicles from T. pisiformis cysticercus.…”

Biological and morphological consequences of dsRNA-induced suppression of tetraspanin mRNA in developmental stages of Echinococcus granulosus

“…Scale-bars: 2 μm in platyhelminths [47][48][49]. Some membrane-bound proteins have been demonstrated, including important proteins such as binding proteins, heat-shock proteins, enzymatic and transmembrane proteins [50]. Sotillo et al [51] showed the expression profiles of tegument proteins upregulated over 3 h to 5 days during transformation of cercariae to schistosomula in S. mansoni.…”