Bioactivity of Exosomes Derived from Trained Natural Killer Cells versus Non-Trained One: More Functional and Antitumor Activity

Abstract

Background. Natural killer (NK) cells are cytotoxic lymphocytes of the innate immune system, capable of killing viral-infected and cancerous cells. NK cell-mediated immunotherapy has remarkably changed the current paradigm of cancer treatment in recent years. It emerged as a safe and effective therapeutic approach for patients with advanced-stage leukemia. Several immune-escape mechanisms can be enacted by cancer cells to avoid NK-mediated killing. Exosomes released by NK cells that carry proteins and miRNAs can exert an antitumor effect. In the present study, we hypothesized that maybe exosomes derived from trained natural killer cells show more antitumor effect in comparison to non-trained one. Methods. PBMC was separated by the Ficoll method and cultured with IL-2 for 21 days to expand NK cells. The NK cells were co-cultured with K562 for 72 hours and exosome-derived co-cultured (as trained) and natural killer cell-derived exosomes (as non-trained) were extracted by Exo kit. The exosomes were confirmed by dynamic light scattering (DLS), transmission electron microscopy (TEM), flow cytometry, and western blotting. The K562 cells were separately treated by trained and non-trained exosomes and MTT assay, apoptosis, and real-time PCR were performed. Results. Based on flow cytometry, CD56 marker was 89.7% and 40.1% for NK cells and NK-derived exosomes, respectively. CD63 and CD9 were positive for exosomes by western blotting. The morphology of exosome was confirmed by TEM. Treated K562 cells by trained exosomes indicated the diminished cell viability and higher apoptosis. Furthermore, the trained exosomes showed up-regulation in both P53 and caspase3 genes as compared with non-trained sample. Discussion. Trained Exos showed a potent inhibitory effect on proliferation and induced apoptosis on K562 cell lines compared to the same dose of non-trained Exos. According to the results of qRT-PCR, trained Exos exerted an antitumor activity through up-regulation of caspase 3 and P53 in the apoptotic signaling pathway in tumor cells. Our findings indicate an effective action of trained Exos against cancer cells.