Dynamics of circulating liver-derived and CD36+ extracellular vesicles in metabolic dysfunction-associated steatotic liver disease during weight loss intervention

INTRODUCTION Metabolic dysfunction-associated steatotic liver disease (MASLD) is an increasing health challenge, with early diagnosis hindered by the lack of reliable and non-invasive biomarkers. The fatty acid transporter, CD36, plays a significant role in ectopic liver fat accumulation during MASLD development. Extracellular vesicle (EV) secretion is modulated by lipotoxicity and likely represent the phenotype of the cell of origin. MATERIALS AND METHODS Using high-resolution flow cytometry (hFCM) and our in-house-developed EV Array technology for high-throughput multiplexed phenotyping of EVs, our study is the first to characterize and report levels of circulating liver-derived and CD36+ EVs directly in plasma of obese individuals with MASLD (n=36) compared with obese (n=25) or lean (n=27) control individuals without MASLD. Moreover, these EV populations were assessed in the MASLD group before (baseline) and during weight-loss intervention (1 month and 5 months). RESULTS Using a panel of antibodies and labels to CD9, lactadherin (EV markers), asialoglycoprotein receptor 1 (ASGR1, liver marker), and CD36, hFCM analysis revealed that at baseline, liver-EVs were significantly increased in the MASLD group compared with the lean control group, while CD36+-EVs were significantly increased in the MASLD group compared with either control group. During the weight loss intervention, levels of liver-derived EVs, CD36⁺ EVs, and liver-EVs expressing CD36 were significantly reduced in the MASLD group as early as 1 month, with further reductions observed after 5 months. Preliminary results from EV Array analysis confirm these findings and reveal a significant correlation between CD36⁺ EVs and ectopic liver fat accumulation. CONCLUSION Our study advances the understanding of EVs in the development of MASLD and introduces a novel approach for characterizing organ-specific circulating EV phenotypes. Profiling the composition of these EV populations offers valuable insights into organ physiology and opens new avenues for biopsy-free diagnosis and monitoring of complex diseases such as MASLD.