isolation-dendritic-cell-uptake-small-extracellular-vesicles-from

Isolation And Dendritic Cell-Uptake of Small Extracellular Vesicles from <em>Echinococcus granulosus</em>

The secretion of extracellular vesicles by cestodes is crucial for enabling cellular communication not only among parasites but also with host tissues. In ...

Universidad Nacional de Mar del Plata (UNMdP)

Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET)

Cystic echinococcosis is a chronic, complex, and neglected disease. Novel therapeutical tools are needed to optimize human treatment. A number of compounds have been investigated, either using in vitro cultured parasites and/or applying in vivo rodent models. Although some of these compounds showed promising activities in vitro, and to some extent also in the rodent models, they have not been translated into clinical applications. Membrane enzyme activities in culture supernatants of treated protoscoleces with calcium modulator drugs and anthelmintic drugs were measured and provided an indication of compound efficacy. This work describes for the first time the detection of alkaline phosphatase, gamma-glutamyl-transpeptidase and acetylcholinesterase activities in supernatants of in vitro treated Echinococcus granulosus protoscoleces. Marked differences on the enzymatic activities in supernatants from drug treated cultures were detected. We demonstrated that those genes that show the highest degree of conservation when compared to orthologs, are constitutively and highly expressed in protoscoleces and metacestodes. Due to high sensibility and the lack of activity in supernatants of intact protoscoleces, gamma-glutamyl-transpeptidase is proposed as the ideal viability marker during in vitro pharmacological studies against E. granulosus protoscoleces.

Echinococcus granulosus tegumental enzymes as in vitro markers of pharmacological damage: a biochemical and molecular approach.

Cystic echinococcosis (CE) is a worldwide zoonosis caused by the Echinococcus granulosus larval stage. The currently available therapy for this disease is based on benzimidazoles, which are rarely curative and cause several adverse effects. Therefore, new treatment options are needed. Octreotide (Oct) is a somatostatin analogue which exhibits anti-proliferative and anti-secretory effects over several cancer cell lines expressing somatostatin receptors. Here, we assessed the in vitro pharmacological effect of Oct against the E. granulosus larval stage. The drug caused a significant dose-dependent decrease in the viability of both protoscoleces and metacestodes. SEM and TEM analysis showed ultrastructural damage in both larval forms under drug treatment. Based on this, we investigated the possible presence of an Oct binding receptor in the parasite. The putative somatostatin/allatostatin-like receptor (Eg-s/ast) conserves the characteristic topology and signature sequences of the prototype somatostatin receptor common to vertebrates and is expressed in both metacestodes and protoscoleces. Moreover, Oct treated-parasites showed the presence of autophagic structures and a significant increase in transcriptional expression of autophagy key genes such as Eg-atg6, Eg-atg8, Eg-atg12 and Eg-atg16. In addition, by in toto immunolocalization assays, an increase in the punctate pattern and Eg-Atg8 protein expression was detected in Oct-treated metacestodes. Subsequently, the combination of Oct and Met had an additive effect on the viability of both larval forms. Our results provide additional evidence for the participation of PI3K/AKT/TOR/autophagy pathway in the Echinococcus survival and suggest the concomitant use of these drugs as potential therapeutic agents in treating of CE.

In vitro anti-echinococcal activity of octreotide: Additive effect of metformin linked to autophagy.

Alveolar echinococcosis (AE) is a severe disease caused by the larval stage of the tapeworm  Current chemotherapeutic treatment options based on benzimidazoles are of limited effectiveness, which underlines the need to find new antiechinococcosis drugs. Metformin is an antihyperglycemic and antiproliferative agent that shows activity against the related parasite  Hence, we assessed the  and  effects of the drug on  Metformin exerted significant dose-dependent killing effects on  cultured parasite stem cells and protoscoleces and significantly reduced the dedifferentiation of protoscoleces into metacestodes. Likewise, oral administration of metformin (50 mg/kg of body weight/day for 8 weeks) was effective in achieving a significant reduction of parasite weight in a secondary murine AE model. Our results revealed mitochondrial membrane depolarization, activation of Em-AMPK, suppression of Em-TOR, and overexpression of Em-Atg8 in the germinal layer of metformin-treated metacestode vesicles. The opposite effects on the level of active Em-TOR in response to exogenous insulin and rapamycin suggest that Em-TOR is part of the parasite's insulin signaling pathway. Finally, the presence of the key lysosomal pathway components, through which metformin reportedly acts, was confirmed in the parasite by  assays. Taken together, these results introduce metformin as a promising candidate for AE treatment. Although our study highlights the importance of those direct mechanisms by which metformin reduces parasite viability, it does not necessarily preclude any additional systemic effects of the drug that might reduce parasite growth .

Metformin Suppresses Development of the Echinococcus multilocularis Larval Stage by Targeting the TOR Pathway.

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.

Article	Year
Echinococcus granulosus tegumental enzymes as in vitro markers of pharmacological damage: a biochemical and molecular approach. Parasitology international\| PubMed ID: 22609954	2012
In vitro anti-echinococcal activity of octreotide: Additive effect of metformin linked to autophagy. Acta tropica\| PubMed ID: 31870710	2020
Metformin Suppresses Development of the Echinococcus multilocularis Larval Stage by Targeting the TOR Pathway. Antimicrobial agents and chemotherapy\| PubMed ID: 32540980	2020
Characterization of protein cargo of Echinococcus granulosus extracellular vesicles in drug response and its influence on immune response. Parasites & vectors\| PubMed ID: 37516852	2023

Andrea C. Cumino

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