miR-143 inhibits cell proliferation through targeted regulating the expression of K-ras gene in HeLa cells
10.3760/cma.j.issn.0253-3766.2016.12.003
- VernacularTitle:miR-143通过靶向调控K-ras基因的表达抑制宫颈癌HeLa细胞的增殖
- Author:
Haixia QIN
1
;
Hongkai CUI
;
Ying PAN
;
Ruili HU
;
Lihong ZHU
;
Shijin WANG
Author Information
1. 新乡医学院第一附属医院妇产科
- Keywords:
Subject words] Cervical neoplasms;
miR-143;
K-ras;
HeLa cells;
Cell proliferation
- From:
Chinese Journal of Oncology
2016;38(12):893-897
- CountryChina
- Language:Chinese
-
Abstract:
Objective To explore the effect of microRNA miR?143 on the proliferation of cervical cancer HeLa cells through targeted regulating the expression of K?ras gene. Methods The luciferase report carrier containing wild type 3′?UTR of K?ras gene ( K?ras?wt) or mutated 3′?UTR of the K?ras ( K?ras?mut) were co?transfected with iR?143 mimic into the HeLa cells respectively, and the targeting effect of miR?143 in the transfectants was verified by the dual luciferase report system. HeLa cells were also transfected with miR?143 mimic ( miR?143 mimic group) , mimic control ( negative control group) , and miR?143 mimic plus K?ras gene ( miR?143 mimic+K?ras group) , respectively. The expression of miR?143 in the transfected HeLa cells was detected by real?time PCR ( RT?PCR ) , and the expression of K?ras protein was detected by Western blot. The cell proliferation activity of each group was examined by MTT assay. In addition, human cervical cancer tissue samples ( n=5) and cervical intraepithelial neoplasia tissue samples ( n=5) were also examined for the expression of miR?143 and K?ras protein by RT?PCR and Western blot, respectively. Results The luciferase report assay showed that co?transfection with miR?143 mimic decreased the luciferase activity of the K?ras?wt significantly, but did not inhibit the luciferase activity of the K?ras?mut. The expression of miR?143 in the HeLa cells transfected with miR?143 mimic was significantly higher than that in the HeLa cells transfected with the mimic control (3.31±0.45 vs 0.97±0.22, P<0.05). The MTT assay revealed that the cell proliferative activity of the miR?143 mimic group was significantly lower than that of the negative control group (P<0.05), and the cell proliferative activity of the miR?143 mimic+K?ras group was also significantly lower than the control group ( P<0.05) but higher than the miR?143 mimic group significantly (P<0.05). The expression levels of K?ras protein in the miR?143 mimic group, the negative control group and the miR?143 mimic+K?ras group were lowest, moderate, and highest, respectively (115.27±34.08, 521.36±41.89, and 706.52±89.44, all P<0.05). In the tissue samples, the miR?143 expression in the cervical cancer group was significantly lower than that of the cervical intraepithelial neoplasia group (0.32±0.06 vs. 0.93±0.17, P<0.05);whereas the K?ras protein expression in the cervical cancer group was significantly higher than that in the cervical intraepithelial neoplasia group ( 584. 39 ± 72.34 vs. 114.23±25.82, P<0.05). Conclusions In vitro, miR?143 can inhibit the proliferative activity of HeLa cells through targeted regulating the expression of K?ras gene. In human cervical cancer tissues of a small sample set, the expression of miR?143 is downregulated, and the expression of K?ras is upregulated.
