Inhibition of Exosome-Mediated Crosstalk in HCC

For inhibition of exosome generation, MSCs were treated with a culture medium containing 10 μM of GW4869 for 24 h (Sigma, USA) and marked as MSC-GW. Next, HCC cell co-culturing with MSCs, MSC-GW cells, or MSC exosomes was conducted with a transwell co-culture system (Corning, USA) based on a previous study (29 (link)). In brief, the HCC cells were seeded into the lower chamber. Then the MSCs or MSC-GW cells were seeded into the upper chamber and co-cultured with HCC cells. MSC exosomes were added into the upper chamber with the isovolumetric medium. The cultured Huh7 and PLC cells were divided into four groups: normal group (without any treatment), MSC group (co-cultured with MSCs), MSC-GW group (co-cultured with MSC-GW), and MSC-Exo group (co-cultured with 20 μg/ml of MSC exosomes). PKH-67 (Sigma, USA) staining was carried out to observe exosome uptake of Huh7 and PLC according to the manufacturer’s instructions. Huh7 and PLC cells were collected for proliferation, apoptosis, invasion, sphere formation, and RT-qPCR assays. Huh7 cells were harvested for RNA sequencing (RNA-seq).