Digest the Sugar, Kill the Parasite: A New Experimental Concept in Treating Alveolar Echinococcosis

Introduction: The E. multilocularis laminated layer (LL) is a heavily glycosylated parasitic structure that plays an important role in protecting the larval stage (metacestode) of this parasite from physiological and immunological host reactions. We elaborated an experimental design with the idea to modify the (glycan) surface of the LL by a targeted digestion. This should allow the host defense to more easily recognize and attack (or kill) the parasite by immune-mediated effects. Methods: Experimentally, E. multilocularis (clone H95) metacestodes were cultured in vitro with or without addition of alpha 1-3,4,6-galactosidase or beta 1-3-galactosidase in the medium. Morphological changes were subsequently measured by microscopy at different time points. Parasites were then recovered at day 5 and reinjected into mice for assessing their viability and infectious status. For finally recovered parasites, the respective load was assessed ex vivo by wet weight measurement, and host-related PD1 and IL-10 levels were determined as the key immunoregulators by using flow cytometry. Results: Our experiments demonstrated that the parasite vesicular structure can be directly destroyed by adding galactosidases into the in vitro culture system, resulting in the fact that the parasite metacestode vesicles could not anymore infect and develop in mice after this glycan digestion. Moreover, when compared to the mice inoculated with E. multilocularis metacestode without galactosidases, PD1 expression was upregulated in CD4(+) Teffs from mice inoculated with E. multilocularis metacestode pretreated with beta 1-3-galactosidase, with a lower IL-10 secretion from CD4(+) Teffs; there was no difference of PD1 and IL-10 expression levels regarding CD4(+) Teff from mice inoculated with E. multilocularis metacestode pretreated with alpha 1-3,4,6-galac-tosidase. Discussion: We raised our hypothesis that this aborting effect may be linked to an altered PD1 and IL-10 response fine-tuning between immunopathology and immune protection. These findings justify a continuation of these experiments upon therapeutical in vivo administration of the enzymes.

Automated summary

The study demonstrated that the parasite vesicular structure could be directly destroyed by adding galactosidases into the in vitro culture system, leading to the inability of the parasite metacestode vesicles to infect mice. Pretreatment of E. multilocularis metacestode with beta 1-3-galactosidase resulted in upregulated PD1 expression and lower IL-10 secretion in CD4(+) Teffs from inoculated mice, while there was no significant difference in PD1 and IL-10 expression levels in CD4(+) Teffs from mice inoculated with E. multilocularis metacestode pretreated with alpha 1-3,4,6-galactosidase.