Profiling of microRNAs in extracellular vesicles derived from human hepatoma cell line HepG2

Previously we have shown that HepG2 cell-derived extracellular vesicles (EVs) exhibit anti-fibrotic effects in human hepatic stellate cells in vitro and during toxin-induced liver injury in mice. The mechanisms by which HepG2 EVs ameliorate liver fibrosis have not been fully investigated but microRNAs are among the EV payload components that are delivered to recepient cells to regulate their functionsl properties. In this study, EVs were isolated by differential centrifugation of culture supernatants from HepG2 cells that ahd been maintained in serum-free conditions. The EVs were characterized by Nanosight Tracking Analysis and Western blot. Three different batches of HepG2 EVs were used to isolate total RNAs by QIAGEN miRNeasy kit, and approximately 200 ng of RNAs were subjected to small RNA-seq. A total of 205 miRNAs were identified, with miR-191-5p, miR-148a-3p, miR-320a, miR-423-5p, and miR-483-5p ranked as the top five most abundant miRNAs. HepG2 cells were cultured with serum-free medium for 48 hr for supernatant collection and EV isolation. Three batches of HepG2 EVs were used to isolate RNA, which was subjected to small RNA-seq.