Acute dietary fat formulations shape circulating postprandial extracellular vesicle-associated lipid signatures

Humans spend much of the day in the postprandial state, yet how acute dietary fat formulations shape lipid signatures associated with circulating extracellular vesicle (EV)-enriched fractions remains poorly understood. We conducted a randomized acute crossover pilot trial in apparently healthy adults who consumed fat-free, saturated fatty acid (SFA)-, monounsaturated fatty acid (MUFA)-, or MUFA-based meals supplemented with ω3 long-chain polyunsaturated fatty acids (MUFA + ω3). Plasma SEC-derived EV-enriched fractions were collected at fasting baseline (B), 2-3 h after meal ingestion (postprandial peak, P), and 5-6 h after ingestion (late postprandial phase, LP), and characterized by electron microscopy, nanoparticle tracking analysis, bead-based surface-marker profiling, untargeted UHPLC-MS/MS lipidomics, and targeted integration of proteomic markers. The recovered fractions showed vesicle-like morphology, comparable NTA-derived particle size distributions and concentrations, dynamic surface-marker profiles, and broad lipidomic coverage comprising 373 lipid species. Significant within-diet species-level changes were detected only at LP under SFA and MUFA + ω3, involving 45 and 57 lipid species, respectively. Eighteen altered species were shared between both formulations, all of them triacylglycerols (TG), whereas 27 were specific to SFA and 39 to MUFA + ω3. Between-diet analyses revealed persistent differences dominated by TG but also involving glycerophospholipids and sphingolipids, with the strongest divergence between SFA and MUFA + ω3 and greater concordance between MUFA and MUFA + ω3. Integration with plasma biochemistry identified an exploratory TG-centered systemic pattern, while proteomic analyses revealed associations of lipid remodeling with both EV-associated tetraspanins and ApoA/ApoB-related signals. These findings support plasma SEC-derived EV-enriched fractions as a dynamic postprandial molecular interface in which dietary fat quality is reflected through coordinated lipid remodeling across vesicular and apolipoprotein-related features.