Docosahexaenoic acid reverses the promoting effects of breast tumor cell-derived exosomes on endothelial cell migration and angiogenesis

Aim: As a natural compound, docosahexaenoic acid (DHA) exerts anti-cancer and anti-angiogenesis functions through exosomes; however, little is known about the molecular mechanisms. Main methods: Breast cancer (BC) cells were treated with DHA (50 mu M) and then tumor cell-derived exosomes (TDEs) were collected and characterized by electron microscopy, dynamic light scattering, and western blot analyses. By the time the cells were treated with DHA, RT-qPCR was used to investigate the expression of vascular endothelial growth factor (VEGF) and the selected pro- and anti-angiogenic microRNAs (miRNAs). The quantification of secreted VEGF protein was measured by enzyme-linked immunosorbent assay (ELISA). The effects of TDEs on endothelial cell angiogenesis were explored by transwell cell migration and in vitro vascular tube formation assays. Key findings: DHA treatment caused a significant and time-dependent decrease in the expression and secretion of VEGF in/from BC cells. This also increased expression of anti-angiogenic miRNAs (i.e. miR-34a, miR-125b, miR-221, and miR-222) while decreased levels of pro-angiogenic miRNAs (i.e. miR-9, miR-17-5p, miR-19a, miR-126, miR-130a, miR-132, miR-296, and miR-378) in exosomes derived from DHA-treated BC cells, TDE (DHA+). While treatment with exosomes (100 mu g/ml) obtained from untreated BC cells, TDE (DHA-), enhanced the expression of VEGF-A in human umbilical vein endothelial cells (HUVECs), incubation with DHA or TDE (DHA+) led to the significant decrease of VEGF-A transcript level in these cells. We indicated that the incubation with TDE (DHA+) could significantly decrease endothelial cell proliferation and migration and also the length and number of tubes made by HUVECs in comparison with endothelial cells incubated with exosomes obtained from untreated BC cells. Significance: DHA alters angiogenesis by shifting the up-regulation of exosomal miRNA contents from pro-angiogenic to anti-angiogenic, resulting in the inhibition of endothelial cell angiogenesis. These data can help to figure out DHA's anti-cancer function, maybe its use in cancer therapy.