Expression of miRNA-4766-5p in ovarian cancer tissues and its effect on proliferation and invasion of ovarian cancer cells / 肿瘤研究与临床

ABSTRACT

Objective:To investigate the expression of miRNA-4766-5p (miR-4766-5p) in ovarian cancer tissues, the effect of miR-4766-5p on the proliferation and invasion of ovarian cancer cells in vitro and its related mechanism.Methods:The cancerous tissues and adjacent tissues of 32 patients with ovarian cancer who underwent surgical treatment in the Central Hospital of Wuhan, Tongji Medical College, Huazhong University of Science and Technology from February 2019 to December 2020 were selected, as well as the ovarian cancer cell lines (A2780, OC3, SKOV-3. HO-8910, and OVCAR-3) and normal ovarian epithelial cell line IOSE80 were selected for subsequent experiments. Real-time fluorescent quantitative polymerase chain reaction (qRT-PCR) was used to detect the relative expression of miR-4766-5p in each cell line and ovarian cancer tissues. The cell line with the lowest relative expression of miR-4766-5p was taken as the experimental subject, and the negative control plasmid (control group) and the plasmid expressing miR-4766-5p (miR-4766-5p group) were transfected respectively into the ovarian cancer cells. CCK-8 method and Transwell experiment were used to detect the effect of overexpression of miR-4766-5p on the proliferation and invasion ability of the selected cells. PITA and starBase V2.0 softwares were used to predict the target genes of miR-4766-5p, and the dual luciferase reporter gene method was used for verification. qRT-PCR and Western blot were used to detect the effect of overexpression of miR-4766-5p on target gene expression of the selected cell lines.Results:The relative expressions of miR-4766-5p in ovarian cancer tissues and adjacent tissues were 1.06±0.17 and 5.25±0.70, respectively, and the difference was statistically significant ( t = 5.86, P < 0.01). Compared with IOSE80 cells, the relative expression of miR-4766-5p in all ovarian cancer cell lines decreased (all P < 0.01), and the relative expression of miR-4766-5p in OC3 cells was the lowest, so this cell line was used for subsequent experiments. The result of CCK-8 method showed that the absorbances of OC3 cells in the miR-4766-5p group were lower than those in the control group after 1 d, 2 d and 3 d of cell culture (all P < 0.05). The result of Transwell experiment showed that the number of penetrating cells in OC3 cells of the miR-4766-5p group and the control group were 25±6 and 86±11, respectively, and the difference was statistically significant ( t = 4.77, P < 0.01). PITA and starBase V2.0 softwares predicted that miR-4766-5p may have binding sites with microtubule unstable protein 1 (STMN1) mRNA; the result of dual luciferase reporter gene showed that the target gene of miR-4766-5p may be STMN1. The relative expression of STMN1 mRNA in OC3 cells of the miR-4766-5p group and the control group were 0.28±0.05 and 1.00±0.05, respectively, and the difference was statistically significant ( t = 10.47, P < 0.01). Compared the control group, the expression of STMN1 protein in the miR-4766-5p group decreased, the expression of epithelial cell phenotype protein β-catenin increased, the expression of mesenchymal cell phenotype protein Snail decreased, and the expressions of cyclin CDK2 and cyclin E decreased. Conclusion:miR-4766-5p is under-expressed in ovarian cancer tissues and ovarian cancer cell lines, and miR-4766-5p may inhibit the proliferation and invasion of ovarian cancer cells by down-regulating the expression of its target gene STMN1.