Objective: Extracellular vesicles derived from oral cancer cells, which include Exosomes\nand Oncosomes, are membranous vesicles secreted into the surrounding extracellular\nenvironment. These extracellular vesicles can regulate and modulate oral squamous cell carcinoma\n(OSCC) progression through the horizontal transfer of bioactive molecules including proteins,\nlipids and microRNA (miRNA). The primary objective of this study was to examine the potential to\nisolate and evaluate extracellular vesicles (including exosomes) from various oral cancer cell lines\nand to explore potential differences in miRNA content. Methods: The OSCC cell lines SCC9, SCC25\nand CAL27 were cultured in DMEM containing 10% exosome-free fetal bovine serum. Cell-culture\nconditioned media was collected for exosome and extracellular vesicle isolation after 72 hours.\nIsolation was completed using the Total Exosome Isolation reagent (Invitrogen) and extracellular\nvesicle RNA was purified using the Total Exosome RNA isolation kit (Invitrogen). Extracellular\nvesicle miRNA content was evaluated using primers specific for miR-16, -21, -133a and -155.\nResults: Extracellular vesicles were successfully isolated from all three OSCC cell lines and total\nextracellular vesicle RNA was isolated. Molecular screening using primers specific for several\nmiRNA revealed differential baseline expression among the different cell lines. The addition of\nmelatonin significantly reduced the expression of miR-155 in all of the OSCC extracellular vesicles.\nHowever, miR-21 was significantly increased in each of the three OSCC isolates. No significant\nchanges in miR-133a expression were observed under melatonin administration. Conclusions:\nAlthough many studies have documented changes in gene expression among various cancers\nunder melatonin administration, few studies have evaluated these effects on microRNAs. These\nresults may be among the first to evaluate the effects of melatonin on microRNA expression in oral\ncancers, which suggests the differential modulation of specific microRNAs, such as miR-21,\nmiR-133a and miR-155, may be of significant importance when evaluating the mechanisms and\npathways involved in melatonin-associated anti-tumor effects.
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