Our principal findings are as follows: 1) SEC yielded the most number of proteins, while preserving the top 100 EV proteins and select EV marker proteins in comparison to UTC and DG; 2) lipotoxic EVs are enriched in many distinct DAMPs and ribosomal proteins; 3) ICAM1 and PRKCA were the most significant ceramide trafficking-dependent lipotoxic EV proteins; and 4) F13A1, haptoglobin, VNN1, and IGFALS were detected in lipotoxic hepatocyte-derived EVs and in NASH plasma EVs, providing candidates for further validation studies. Here, HP is linked to metabolic dysfunction-associated steatohepatitis.