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 α1-3,4,6-galactosidase or β1-3-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 β1-3-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 α1-3,4,6-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.
Keywords: Echinococcus multilocularis; Galactosidase; Glycan; PD1.
© 2020 S. Karger AG, Basel.