Exosomes in the ascites of ovarian cancer patients: origin and effects on anti-tumor immunity.

This study was performed to identify the origin of the ascites-derived exosomes from patients with ovarian cancer and to observe the effect of exosomes on anti-tumor immunity. Exosomes were isolated from the ascites of patients with ovarian epithelial cancer by ultracentrifugation plus density gradient centrifugation. The origin of exosomes was identified by immunoelectronmicroscopy (IEM). The growth curve of the tumor cell line SKOV3 cultured with or without exosomes was analyzed. The apoptosis of autogeneic tumor cells (ATCs) and SKOV3 cells affected by exosomes was measured by flow cytometry (FCM) and light phase contrast microscopy. The cytotoxic effect of the peripheral blood mononuclear cells (PBMCs) stimulated by exosomes and/or dendritic cells (DCs) on ovarian cancer cells was measured using a CCK-8 assay. The levels of IFN-γ released by PBMCs stimulated by exosomes and/or DCs were measured by ELISA. The apoptosis of PBMCs and DCs affected by exosomes was measured by FCM and light microscopy. Whether the mature process of DCs was affected by exosomes was studied by FCM. The ratio of CD4+ T cell and CD8+ T cell were measured by FCM. FasL and TRAIL molecules on exosomes were detected by western blot analysis. The human FasL antagonistic antibody was used to block the apoptosis of DCs and PBMCs induced by exosomes. The receptors of TRAIL DR4 and DR5 on PBMCs and DCs were detected by FCM. In 41 patients examined, we isolated exosomes from the ascites of 35 patients. We detected TCR, CD20, HLA-DR, B7-2, HER2/neu, CA125 and Histone H2A on exosomes. We found that exosomes might impair the cytotoxic activity of PBMCs when DCs are present. We found that exosomes had no effect on the growth and apoptosis of SKOV3 cells. However, exosomes may induce apoptosis of precursors, mature DCs and PBMCs. We found that FasL and TRAIL were present in the exosome suspension and addition of an anti-FasL antibody may decrease the percentage of apoptosis of DCs and PBMCs. We conclude that exosomes exist in ascites of 85.4% of patients with ovarian cancer. Moreover, these exosomes may be of multi-origin. Exosomes had no effect on the growth and apoptosis of tumor cells but impaired the cytotoxic activity of PBMCs in the presence of DCs. Exosomes also may induce apoptosis of the precursors of DCs, DCs and PBMCs. FasL and TRAIL on exosomes may partly account for the apoptosis of cells of the immune system.