Objective To screen the expression differences of miRNAs in supernatant exosomes of cervical cancer cell line(SiHa/HeLa)and normal cervical epithelial cell line(H8)and to identify target genes by bioinformatics analysis.Methods Cervical cancer cell line(SiHa/HeLa)and normal cervical epithelial cell line(H8)were cultured,the supernatant of cultured cells was collected,and exosomes were extracted by ultracentrifugation.The morphology of exosomes was observed by transmission electron microscopy(TEM),and exosome marker proteins were identified by Western blot experiments.The extracted exosomes were subjected to high-throughput sequencing of miRNAs,and the differential miRNAs in SiHa and H8 exosomes,and the differential miRNAs in HeLa and H8 exosomes,respectively,were obtained.The target genes of the obtained intersecting miRNAs were predicted,and the Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway enrichment analysis were performed.Results TEM showed that the exosomes were typical cup-shaped structures with the size of 30～200 nm.Western blot showed that the extracted samples had the expression of exosomes marker proteins.Sequencing results revealed that there were 235 miRNAs with differential expression between HeLa and H8 exosomes,249 miRNAs with differential expression between SiHa and H8 exosomes,and 166 miRNAs with differential expression in exosomes from two cell lines.GO biological function analysis revealed that the differences in cervical cancer exosomal miRNAs were mainly distributed in nucleus,cytoplasm,vesicles and cytoskeleton,and protein binding mainly played a role in metal ion binding,transferase activity,nucleotide binding and other links,and was mainly involved in RNA polymerase Ⅱ's transcription regulation,DNA template,phosphorylation,cell cycle and other biological functions.KEGG pathway enrichment analysis displayed that cervical cancer-related pathways were related to the regulation of metabolic pathways,tumor pathogenesis pathways,axon guidance,PI3K-Akt,calcium ions,Rapl,Ras and other signaling pathways.Conclusion miRNAs such as miR-335-5p,miR-660,miR-155-5p,miR-339-5p,and miR-345-3p are differentially expressed in exosomes derived from cervical cancer,and these differentially expressed miRNAs play a key role in the progression of cervical cancer through metabolic pathways,pathogenesis of cancer,axon guidance,PI3K-Akt,calcium ions,Rap1,Ras and other signaling pathways.
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