MicroRNA expression profiling of hepatocellular carcinoma tissues derived exosomes

In this study, we isolated exosomes from liver cancer tissues and exosomal miRNA expression was profiled by high-throughput sequencing to investigate the potential of exosomal contents as biomarkers for HCC. Fresh tumor and adjacent normal tissue samples were gently cut into small pieces and incubated at 37 ◦C for 30–60min in DMEM/F12 media supplemented with collagenase I, collagenase Ⅳ, hyaluronidase and DNase I under continuous rotation. The cell suspension containing the diffusing tissue exosomes was filtered through a 70-μm filter and centrifuged at 300×g for 10 min, followed by centrifugation at 2000×g for 20 min to eliminate cells and tissue debris. The exosomes were extracted by ultra-high speed centrifugation. RNA was isolated from exosome using Trizol reagent. small RNA library was constructed and then sequenced using the Illumina NextSeq 500 platform.