Effects of miR-20a-5p targeting KDM6B on the proliferation, migration and invasion of osteosarcoma cells / 国际肿瘤学杂志

ABSTRACT

Objective:To investigate the expressions of miR-20a-5p and lysine (K) demethylase 6B (KDM6B) in osteosarcoma tissues and the effects of miR-20a-5p targeting KDM6B on the proliferation, migration and invasion of osteosarcoma cells and tumor growth.Methods:The clinicopathological and paracancerous tissues of 20 patients with osteosarcoma admitted to the First Affiliated Hospital of Chinese Medical University from January 2017 to March 2019 were collected. Quantitative real-time PCR (qRT-PCR) was used to detect the expression levels of miR-20a-5p and KDM6B mRNA in tissues. The osteosarcoma MG63 cells were divided into control group, mimic NC group, miR-20a-5p mimic group, and NC+ empty vector group, miR-20a-5p+ empty vector group, miR-20a-5p+ KDM6B group. The expression levels of miR-20a-5p and KDM6B mRNA of all groups were detected by qRT-PCR. Western blotting was used to detect the expression level of KDM6B. CCK-8 assay, cell scratch test and Transwell test were used to detect cell proliferation, migration and invasion ability. According to the random number table method, nude mice were divided into NC+ empty vector group, miR-20a-5p+ empty vector group and miR-20a-5p+ KDM6B group, with 5 mice in each group. Tumor growth ability was detected by tumor xenograft nude mouse models.Results:The relative expression level of miR-20a-5p mRNA in osteosarcoma tissues was 0.55±0.27, and that in paracancerous tissues was 1.22±0.28, with a statistically significant difference ( t=7.701, P<0.001). The relative expression level of KDM6B mRNA in osteosarcoma tissues was 1.66±0.19, and that in paracancerous tissues was 1.00±0.15, with a statistically significant difference ( t=12.219, P<0.001). After transfection of miR-20a-5p, KDM6B mRNA and protein expression levels decreased with the increase of miR-20a-5p expression level. After miR-20a-5p transfection for 48 h, the cell proliferation abilities of the blank control group, mimic NC group and miR-20a-5p mimic group were 0.83±0.04, 0.81±0.03 and 0.52±0.01 ( F=89.655, P<0.001), compared with the blank control group and mimic NC group, the cell proliferation ability was significantly inhibited in the miR-20a-5p mimic group (both P<0.001). The cell proliferation abilities of NC+ empty vector group, miR-20a-5p+ empty vector group and miR-20a-5p+ KDM6B group were 0.83±0.05, 0.52±0.01 and 0.67±0.05 ( F=43.919, P<0.001), compared with the NC+ empty vector group, the cell proliferation ability was significantly inhibited in the miR-20a-5p+ empty vector group ( P<0.001); compared with the miR-20a-5p+ empty vector group, the cell proliferation ability of miR-20a-5p+ KDM6B group increased significantly ( P<0.001). The scratch healing rates of the blank control group, mimic NC group and miR-20a-5p mimic group were (32.51±2.73)%, (30.26±3.22)% and (13.52±1.77)% ( F=46.314, P<0.001), compared with the control group and the mimic NC group, the scratch healing rate of the miR-20a-5p mimic group was significantly decreased (both P<0.001). The scratch healing rates of NC+ empty vector group, miR-20a-5p+ empty vector group and miR-20a-5p+ KDM6B group were (31.34±3.11)%, (12.15±1.64)% and (28.93±2.89)% ( F=47.511, P<0.001), compared with the NC+ empty vector group, the scratch healing rate of the miR-20a-5p+ empty vector group was significantly decreased ( P<0.001); compared with the miR-20a-5p+ empty vector group, the scratch healing rate of miR-20a-5p+ KDM6B group was significantly increased ( P=0.001). The numbers of transmembrane cells in the blank control group, mimic NC group and miR-20a-5p mimic group were 114±16, 108±11 and 42±6 ( F=36.282, P<0.001), compared with the control group and mimic NC group, the number of transmembrane cells of the miR-20a-5p mimic group was significantly decreased (both P<0.001). The numbers of transmembrane cells in the NC+ empty vector group, miR-20a-5p+ empty vector group and miR-20a-5p+ KDM6B group was 143±11, 39±4 and 139±12 ( F=112.120, P<0.001), compared with the NC+ empty vector group, the number of transmembrane cells of the miR-20a-5p+ empty vector group was significantly decreased ( P<0.001); compared with the miR-20a-5p+ empty vector group, the number of transmembrane cells of the miR-20a-5p+ KDM6B group was increased significantly ( P<0.001). The tumor volumes of mice for 21 d in the NC+ empty vector group, miR-20a-5p+ empty vector group and miR-20a-5p+ KDM6B group were (1 667.50±250.40) mm 3, (129.20±21.00) mm 3 and (775.41±77.51) mm 3 respectively, with a statistically significant difference ( F=77.651, P<0.001). The tumor weights of the 3 groups were (1.35±0.18) g, (0.12±0.01) g and (0.61±0.03) g respectively, with a statistically significant difference ( F=104.191, P<0.001). Conclusion:The expression of miR-20a-5p is significantly decreased in osteosarcoma tissues, and the expression of KDM6B is significantly increased in osteosarcoma tissues. Overexpression of miR-20a-5p may inhibit the proliferation, migration and invasion of osteosarcoma cells and tumor growth by targeting to reduce the expression of KDM6B.