He Lijie, Wang Jing, Chang Dandan, Lv Dandan, Li Haina, Feng Heqiang
Departments of T Lymphocyte Subpopulation, Tianjin Fifth Central Hospital, Tianjin, 300450, People's Republic of China.
Departments of Immunologic Fuction, Tianjin Fifth Central Hospital, Tianjin, 300450, People's Republic of China.
Open Med (Wars). 2020 Oct 8;15(1):1019-1027. doi: 10.1515/med-2020-0147. eCollection 2020.
This article aims to investigate the effect of miRNA-200b on the proliferation and apoptosis of cervical cancer cells by targeting RhoA.
HeLa cells of cervical cancer were divided into five groups: blank control group, negative control group (miRNA-200b mimic NC), miRNA-200b mimic group, RhoA-negative control group, and RhoA overexpression group. Cells were collected 48 h after transfection. The expression levels of miRNA-200b were detected by RT-PCR. Target relationship between miRNA-200b and RhoA was verified by the dual-luciferase reporter assay. RhoA mRNA and protein expression were detected by western blot and RT-PCR methods. Flow cytometry was used to detect the apoptosis of cells in each group, and the CCK8 method was used to detect the proliferation of cells in each group. The mRNA and protein expression of Bax and cyclin D1 were detected by RT-PCR and western blot.
The results of the dual luciferase reporter assay showed that RhoA was the target gene of microRNA 200b. Compared with the blank control group and the miRNA-200b mimic-NC group, the proportion of apoptotic cells increased significantly in the miRNA-200b mimic group, and the proliferation of cells was inhibited ( < 0.05). After overexpression of RhoA, the percentage of apoptotic cells decreased and the ability of cell proliferation increased significantly ( < 0.05).
miRNA-200b can inhibit the proliferation and promote the apoptosis of cervical cancer cells by targeting the RhoA gene.
本文旨在研究微小RNA-200b通过靶向RhoA对宫颈癌细胞增殖和凋亡的影响。
将宫颈癌HeLa细胞分为五组：空白对照组、阴性对照组（微小RNA-200b模拟物阴性对照）、微小RNA-200b模拟物组、RhoA阴性对照组和RhoA过表达组。转染48小时后收集细胞。采用RT-PCR检测微小RNA-200b的表达水平。通过双荧光素酶报告基因检测验证微小RNA-200b与RhoA之间的靶向关系。采用蛋白质免疫印迹法和RT-PCR法检测RhoA的mRNA和蛋白表达。采用流式细胞术检测各组细胞凋亡情况，采用CCK8法检测各组细胞增殖情况。采用RT-PCR和蛋白质免疫印迹法检测Bax和细胞周期蛋白D1的mRNA和蛋白表达。
双荧光素酶报告基因检测结果显示，RhoA是微小RNA 200b的靶基因。与空白对照组和微小RNA-200b模拟物阴性对照组相比，微小RNA-200b模拟物组凋亡细胞比例显著增加，细胞增殖受到抑制（P<0.05）。RhoA过表达后，凋亡细胞百分比降低，细胞增殖能力显著增强（P<0.05）。
微小RNA-200b可通过靶向RhoA基因抑制宫颈癌细胞的增殖并促进其凋亡。
