Steatotic Hepatocyte-Derived Exosomes Increase Free Fatty Acids Uptake by Transporting miR-122-5p to Target ATP6V1H-Mediated CD36 Translocation in Hepatocytes

Hepatic steatosis is a notable feature of metabolic associated fatty liver disease (MAFLD), which is associated with exosomal miR-122-5p. However, the mechanisms by which exosomal miR-122-5p is involved in transport of free fatty acid (FFA) remain uncharacterized. The aim of this study was to investigate the functional role of exosomal miR-122-5p derived from steatotic hepatocytes in hepatic CD36 translocation. Exosomes from PBS- or FFA-treated AML12 cells were extracted via ultracentrifugation (Ctrl-exo or SH-exo), and recipient AML12 cells were treated with them for 24 h. Additionally, AML12 cells were transfected with miR-122-5p mimic or inhibitor for 24 h. Dual-luciferase reporter assay was applied to identify miR-122-5p targeting 3'UTR of ATP6V1H mRNA. RT-qPCR and western blot assessed the expressions of miR-122-5p, CD36 and ATP6V1H in exosomes and recipient AML12 cells. The miR-122-5p downregulates ATP6V1H expression by directly targeting the 3'UTR of ATP6V1H mRNA. SH-exo were absorbed by recipient AML12 cells, and exhibited elevated levels of miR-122-5p. The SH-exo increased FFA uptake, promoted CD36 translocation and disrupted v-ATPase assembly in recipient AML12 cells by downregulating the ATP6V1H expression, in an insulin-independent manner. MiR-122-5p mimic similarly increased FFA uptake and CD36 translocation by inhibiting ATP6V1H expression in AML12 cells. Furthermore, miR-122-5p inhibitor reduced the FFA content and increased ATP6V1H expression in SH-exo-treated AML12 cells. Our findings indicated that exosomal miR-122-5p, derived from steatotic hepatocytes, can increase FFA uptake and promote CD36 translocation in recipient hepatocytes by directly targeting ATP6V1H expression, suggesting that hepatic exosomal miR-122-5p may serve as a potential therapeutic target for MAFLD.