Role of microRNA-21 in the proliferation and apoptosis of ovarian epithelial carcinoma cells
10.3760/cma.j.issn.0529-567x.2011.09.010
- VernacularTitle:微小RNA-21表达在卵巢癌细胞生长和凋亡过程中的作用
- Author:
Yanhui LOU
;
Zhumei CUI
;
Fuling WANG
;
Xingsheng YANG
- Publication Type:Journal Article
- Keywords:
Ovarian neoplasms;
MicroRNAs;
RNA-binding proteins;
Apoptosis regulatory proteins;
Cell line,tumor;
Apoptosis
- From:
Chinese Journal of Obstetrics and Gynecology
2011;46(9):684-689
- CountryChina
- Language:Chinese
-
Abstract:
ObjectiveTo investigate the role and mechanism of microRNA-21 (miR-21) in the proliferation and apoptosis of ovarian epithelial carcinoma cells. MethodsA short-hairpin RNA specifically targeting miR-21 plasmid was constructed, and the recombinant was identified by restriction endonuclease analysis and DNA sequencing. Three experimental groups were included, transfection group (transfected with pSIREN-miR-21 ), negative control group ( transfected with pSIREN-miR-21-neg) and blank control group (without transfection plasmid ). The expression of miR-21 was detected by stem-loop real-time reverse transcription (RT)-PCR in OVCAR3 cells ,and western blot was used to detect the expression of programmed cell death 4 ( PDCD4 ) protein. Tethyl thiazolyl tetrazolium (MTT) and flow cytometry method were used respectively. ResultsRecombinant plasmid (pSIREN-miR-21) was constructed successfully and identified by restriction endonuclease analysis and DNA sequencing. The relative expression level of miR-21 in cells transfection, negative control and blank control group was 0.26 ± 0.08, 1.26 ± 0.21and 1.00 respectively. The level of miR-21 in the cells in transfection group was significantly lower than those in the negtive control and blank control group(P ＜0. 01 ). The gray scale of PDCD4. protein was 1443 ±33,858 ± 19 and 846 ± 16 in the transfection group, negative control and blank control group respectively. The value of PDCD4 in transfection group was higher than other control groups, and there were significantly difference among them( P ＜0. 01 ). Moreover, the optical density of the cells in transfection group was 0. 661 ±0. 015,significantly lower than those in two control groups (0. 848 ± 0. 150 for negative control, 0. 935 ± 0. 133 for blank control,P ＜ 0. 01 ). Forty-eight hours after tranfection, the rate of viable apoptotic cell was significantly higher than negative control and blank control group [(25.821 ± 0. 763 )％ vs.(0. 010 ± 0. 003 ) ％ vs. (0. 238 ± 0. 023) ％ ; P ＜ 0. 01];72 hours after tranfection, the rates of viable apoptotic cell and necrotic cell were all higher than the two control groups [the rate of viable apoptotic cell was ( 30. 480 ±0. 821 ) ％, ( 7. 792 ± 0. 312 ) ％ and ( 7. 033 ± 0. 257 ) ％ respectively ( P ＜ 0. 01 ) ; the rate of necrotic cell was (3.558 ±0.211)％, (1.557 ±0.067)％ and (1.049 ±0.028)％, respectively (P＜0. 01)].ConclusionmiR-21 might play an important role in the proliferation and apoptosis of ovarian epithelial carcinoma cells through negatively control the expression of PDCD4.
