Hypoxia induced changes in miRNAs and their target mRNAs in extracellular vesicles of esophageal squamous cancer cells

BACKGROUND: Extracellular vesicles (EVs) are endogenous membrane vesicles with a diameter of 30-200 nm. It has been reported that hypoxic cancer cells can release numerous EVs to mediate multiple regional and systemic effects in the tumor microenvironment. METHODS: In this study, we used ultracentrifugation to extract EVs secreted by TE-13, an esophageal squamous carcinoma (ESCC) cell line during normoxia and hypoxia and performed high-throughput sequencing to detect exosomal miRNAs. Gene ontology (GO) and KEGG pathway analyses were used to reveal pathways potentially regulated by the miRNAs. RESULTS: A total of 10 810 miRNAs were detected; 50 were significantly upregulated and 34 were significantly downregulated under hypoxic environment. GO analysis identified enrichment of protein binding, regulation of transcription (DNA-templated), and membrane as molecular function, biological process, and cellular component, respectively. KEGG pathway analysis revealed cancer-associated pathways, phospholipase D signaling pathway, autophagy, focal adhesion and AGE-RAGE signaling as the key pathways. Further verification experiment from qRT-PCR indicated that miR-128-3p, miR-140-3p, miR-340-5p, miR-452-5p, miR-769-5p and miR-1304-p5 were significantly upregulated in EVs from hypoxia TE-13 cells while miR-340-5p was significantly upregulated in two other ESCC cells, ECA109 and TE-1. CONCLUSION: This study, for the first time reveals changes in the expression of exosomal miRNAs in hypoxic ESCC cells and these findings will act as a resource to study the hypoxic tumor microenvironment and ESCC EVs.