• Yan-hui Lou, Zhu-mei Cui, Fu-ling Wang, and Xing-sheng Yang.
• Department of Gynecology, Qingdao University, Qingdao, China. lyh7497@163.com
• Zhonghua Fu Chan Ke Za Zhi. 2011 Sep 1; 46 (9): 684-9.

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 ± 0.21 and 1.00 respectively. The level of miR-21 in the cells in transfection group was significantly lower than those in the negative 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 transfection, 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 transfection, 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.

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