Hepatic Stellate Cells Activated by Cancer Cell-derived AMIGO2-containing Small Extracellular Vesicles Promote Cancer Cell Migration by Producing IL-8

Our previous studies have demonstrated that amphoterin-induced gene and open reading frame 2 (AMIGO2) functions as a driver gene for liver metastasis, regulating adhesion between cancer cells and liver endothelial cells. AMIGO2-containing small extracellular vesicles (sEVs) derived from gastric cancer (GC) cells were shown to enhance adhesion to hepatic endothelial cells, contributing to pre-metastatic niche formation. However, their role in promoting cancer cell migration into the liver parenchyma remained unclear. This study investigated whether AMIGO2-containing sEVs activate hepatic stellate cells (HSCs) and promote cancer cell migration. AMIGO2-over-expressing and control cell lines (MKN28) were established. sEVs isolated from each cell line were added to human HSCs (TWINT-1). The supernatant collected was added to MKN28 to quantitatively evaluate migration ability and nuclear translocation of NF-kB. A chemokine array identified secreted factors affected by sEV treatment. HSCs were activated by AMIGO2-containing EVs, resulting in increased IL-8 secretion through NF-kB nuclear translocation. This IL-8-rich supernatant significantly enhanced GC cell migration. Neutralizing IL-8 with antibodies suppressed this migration, confirming its pivotal role. AMIGO2-containing sEVs derived from GC cells actively modify the hepatic microenvironment by activating HSCs and inducing IL-8 secretion, which promotes GC cell migration into the liver parenchyma. This process contributes to the formation of a pre-metastatic niche, highlighting AMIGO2-containing sEVs as potential therapeutic targets for preventing liver metastasis.