GM1 oligosaccharide reprograms Neuro2a-derived extracellular vesicles for selective, low-disruption membrane interactions.

The GM1 oligosaccharide β-Gal-(1-3)-β-GalNAc-(1-4)-[α-Neu5Ac-(2-3)]-β-Gal-(1-4)-Glc (OligoGM1) represents the soluble, bioactive portion of GM1 ganglioside and recapitulates many of its trophic, protective, and restorative effects. Our previous proteomic studies on OligoGM1-treated neuroblastoma cells revealed alterations in endocytic trafficking and extracellular vesicles (EVs) biogenesis suggesting a potential role for OligoGM1 in modulating EV biogenesis. Here, we apply a multi-technique biophysical platform to investigate the physico-chemical properties and functionality of EVs released by murine neuroblastoma Neuro2a cells, treated with OligoGM1. We show that OligoGM1 treatment slightly shifts to lower values the EVs' size population distribution and increases their relative lipid content, accompanied by a higher proportion of membrane-associated proteins. Our analysis further indicates that OligoGM1-treated cells release EVs with altered bilayer structural parameters, including increased apparent membrane thickness, consistent with a modified distribution of lipid/protein contributions within the adopted membrane model. Functional characterization using biomimetic model membranes indicates that EVs from treated and untreated cells differ in protein insertion and lipid exchange efficiency, pointing to distinct membrane-interaction modes relevant to downstream cellular interactions. Overall, these findings suggest that OligoGM1 reshapes the molecular architecture and biophysical behaviour of Neuro2a-derived EVs, providing membrane-level fingerprints that may be relevant for future studies of EV-cell interactions in neurodegenerative models.