Background: Exosomes are 30-100nm small membrane vesicles of endocytic origin that are released into the extracellular environment. Studies showed that inside exosomes, cellular gene products including proteins, mRNAs, and microRNAs (miRNAs) are packaged and these molecules might be transferred to recipient cells to deliver their molecular signaling.
Objective: To test whether or not artificial miRNA can be transported into recipient cells and explore the transfer characteristics of exosomes derived from human esophageal cancer cells.
Methods: Cy3labelled-miR-21 mimics were transfected into EC9706 cells which were used as donor cells. Donor cells were washed using PBS to remove remaining mimics and transfection reagent after transfected 12h before replaced with fresh medium. Culture medium from donor cells was collected and centrifuged to remove residual cells after cultured 24h. The culture supernatant was added to the receptor cells cultured for 3h, 6h and 24h, respectively. Transfer efficiency was calculated as the percentage of fluorescent cells by flow cytometry. The expression level of miR-21 in culture supernatant and recipient cells were detected by real time RT-PCR.
Results：The transfer efficiency of cy3 labelled-miR-21 mimics into recipient cells was 59.3%, 83.3% and 85.0% after cultured for 3h, 6h and 24h, respectively using flow cytometry. Real time RT-PCR results showed that the expression level of miR-21 in treatment group was higher than in the control group, with an average fold-change of 1.49, 1.41, 2.08 in recipient cells, while 147.03, 19.29, 15.35 in culture supernatant after cultured for 3h, 6h and 24h, respectively. The results showed that most of exosome-derived miR-21 mimic were transported into recipient cells after cultured for 6h.
Conclusion: It was suggested that artificial miRNA can be transported into recipient cells by exosomes. These specific miRNAs enriched in exosomes may play an important role in intercellular communication mediated.(This study is supported by NSFC (No. 81172747 & 81072259)).