Small extracellular vesicles but not microvesicles from<i>Opisthorchis viverrini</i>promote cell proliferation in human cholangiocytes

Chaiyadet, Sujittra; Sotillo, Javier; Smout, Michael; Cooper, Martha; Doolan, Denise L.; Waardenberg, Ashley; Eichenberger, Ramon M.; Field, Matt; Brindley, Paul J.; Laha, Thewarach; Loukas, Alex

doi:10.1101/2023.05.22.540805

Cited by 2 publications

(1 citation statement)

References 74 publications

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“…So far, two different EV subpopulations (exosomes and MVs) have been identi ed in adult trematode specimens in Fasciola spp., Schistosoma mansoni (Digenea, Schistosomatidae) and O. viverrini [41,[50][51][52]. Similarly, the current EV protein composition of S. chrysophrii indicated that S. chrysophrii has two distinctive EV subpopulations.…”

Section: Extracellular Vesicles Isolation and Transmission Electron M...mentioning

confidence: 94%

An inside out journey: Biogenesis, ultrastructure and proteomic characterisation of the monogenean Sparicotyle chrysophrii extracellular vesicles

Riera-Ferrer,

Mazanec,

Mladineo

et al. 2023

Preprint

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Background: Helminth extracellular vesicles (EVs) are known to have a three-way communication role among parasitic helminths, their host and host-associated microbiota. They are considered biological containers carrying virulence factors, being therefore appealing as therapeutic and prophylactic target candidates. This study aims to describe and characterise EVs secreted by the monogenean Sparicotyle chrysophrii (Microcotylidae), a blood-feeding gill parasite of the gilthead seabream (Sparus aurata) inflicting significant economic losses in Mediterranean aquaculture. Methods: To identify proteins involved in extracellular vesicle biogenesis, genomic datasets from S. chrysophrii were mined in silico using known protein sequences from Clonorchis spp., Echinococcus spp., Fasciola spp., Fasciolopsis spp., Opisthorchis spp., Paragonimus spp., and Schistosoma spp. The EV location and ultrastructure was studied using transmission electron microscopy in high-pressure frozen adult S. chrysophrii specimens. Moreover, EVs were isolated and purified from adult S. chrysophrii (N=200) using a newly developed ultracentrifugation-size-exclusion chromatography protocol for Monogenea, and EVs were characterised via nanoparticle tracking analysis and tandem mass spectrometry. Results: Fifty-nine proteins involved in EV biogenesis were identified in S. chrysophrii and EVs compatible with microvesicles were observed in the parasite’s haptoral region syncytial layer lining the clamps. Moreover, isolated and purified nanoparticles presented a mean size of 251.8 ± 2.2 nm (± SD), and yielded 1.71×108 ± 4.55×107 particles·mL-1 (mean ± SD). The protein composition analysis identified proteins related to peptide hydrolases, GTPases, EF-hand domain proteins, aerobic energy metabolism, anticoagulant/ lipid-binding, haem detoxification, iron transport, EV biogenesis-related, vesicle-trafficking and other cytoskeletal-related proteins. From the identified proteins, leucyl and alanyl aminopeptidases, calpain, ferritin, dynein light chain, 14-3-3, heat shock protein 70, annexin, tubulin, glutathione S-transferase, superoxide dismutase, enolase and fructose-bisphosphate aldolase have already been proposed as target candidates for therapeutic or prophylactic purposes in helminths. Conclusions: We have unambiguously demonstrated the secretion of EVs by S. chrysophrii inferring their biogenesis machinery at a genomic and transcriptomic level, ultrastructural location in the parasite, and analysing their protein composition. The identification of several candidate therapeutic targets among EV’s protein repertoire, offers opportunities for target-based drug discovery and vaccine development approaches, for the first time in Monogenea and a fish-ectoparasite model.

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Section: Extracellular Vesicles Isolation and Transmission Electron M...mentioning

confidence: 94%

An inside out journey: Biogenesis, ultrastructure and proteomic characterisation of the monogenean Sparicotyle chrysophrii extracellular vesicles

Riera-Ferrer,

Mazanec,

Mladineo

et al. 2023

Preprint

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An inside out journey: biogenesis, ultrastructure and proteomic characterisation of the ectoparasitic flatworm Sparicotyle chrysophrii extracellular vesicles

Riera-Ferrer,

Mazanec,

Mladineo

et al. 2024

Parasites Vectors

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Background Helminth extracellular vesicles (EVs) are known to have a three-way communication function among parasitic helminths, their host and the host-associated microbiota. They are considered biological containers that may carry virulence factors, being therefore appealing as therapeutic and prophylactic target candidates. This study aims to describe and characterise EVs secreted by Sparicotyle chrysophrii (Polyopisthocotyla: Microcotylidae), a blood-feeding gill parasite of gilthead seabream (Sparus aurata), causing significant economic losses in Mediterranean aquaculture. Methods To identify proteins involved in extracellular vesicle biogenesis, genomic datasets from S. chrysophrii were mined in silico using known protein sequences from Clonorchis spp., Echinococcus spp., Fasciola spp., Fasciolopsis spp., Opisthorchis spp., Paragonimus spp. and Schistosoma spp. The location and ultrastructure of EVs were visualised by transmission electron microscopy after fixing adult S. chrysophrii specimens by high-pressure freezing and freeze substitution. EVs were isolated and purified from adult S. chrysophrii (n = 200) using a newly developed ultracentrifugation-size-exclusion chromatography protocol for Polyopisthocotyla, and EVs were characterised via nanoparticle tracking analysis and tandem mass spectrometry. Results Fifty-nine proteins involved in EV biogenesis were identified in S. chrysophrii, and EVs compatible with ectosomes were observed in the syncytial layer of the haptoral region lining the clamps. The isolated and purified nanoparticles had a mean size of 251.8 nm and yielded 1.71 × 108 particles · mL−1. The protein composition analysis identified proteins related to peptide hydrolases, GTPases, EF-hand domain proteins, aerobic energy metabolism, anticoagulant/lipid-binding, haem detoxification, iron transport, EV biogenesis-related, vesicle-trafficking and other cytoskeletal-related proteins. Several identified proteins, such as leucyl and alanyl aminopeptidases, calpain, ferritin, dynein light chain, 14–3–3, heat shock protein 70, annexin, tubulin, glutathione S-transferase, superoxide dismutase, enolase and fructose-bisphosphate aldolase, have already been proposed as target candidates for therapeutic or prophylactic purposes. Conclusions We have unambiguously demonstrated for the first time to our knowledge the secretion of EVs by an ectoparasitic flatworm, inferring their biogenesis machinery at a genomic and transcriptomic level, and by identifying their location and protein composition. The identification of multiple therapeutic targets among EVs' protein repertoire provides opportunities for target-based drug discovery and vaccine development for the first time in Polyopisthocotyla (sensu Monogenea), and in a fish-ectoparasite model. Graphical Abstract

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Small extracellular vesicles but not microvesicles fromOpisthorchis viverrinipromote cell proliferation in human cholangiocytes

Cited by 2 publications

References 74 publications

An inside out journey: Biogenesis, ultrastructure and proteomic characterisation of the monogenean Sparicotyle chrysophrii extracellular vesicles

An inside out journey: Biogenesis, ultrastructure and proteomic characterisation of the monogenean Sparicotyle chrysophrii extracellular vesicles

An inside out journey: biogenesis, ultrastructure and proteomic characterisation of the ectoparasitic flatworm Sparicotyle chrysophrii extracellular vesicles

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