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| miR-335 Regulate Cell Proliferation by Targeting Rho Associated Coiled-coil Forming Protein Kinase 1 in Ovarian Cancer Cells SKOV3 |
| QIN Hai-xia1, CUI Hong-kai2, PAN Ying3, HU Rui-li1, ZHU Li-hong1, WANG Shi-jin1 |
1. Department of Obstetrics and Gynecology, The First Affiliated Hospital of Xinxiang Medical University, Weihui 453100, China;
2. Department of Interventional Radiology, The First Affiliated Hospital of Xinxiang Medical University, Weihui 453100, China;
3. Department of Obstetrics and Gynecology, The Third Affiliated Hospital of Xinxiang Medical University, Xinxiang 453000, China |
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Abstract Objective: To evaluate the effect of miR-335 target regulating rho associated coiled-coil forming protein kinase 1(ROCK1)on the cell proliferation of ovarian cancer cells SKOV3. Methods: (1)Chose ovarian cancer cells SKOV3 and human normal ovarian epithelial cells IOSE80 as research object. The miR-335 expression was detected by RT-PCR. The ROCK1 protein expression was detected by Western blot.(2)Chose ovarian cancer cells SKOV3 as research object, transfected miR-335 mimic and mimic control respectively. The miR-335 expression was detected by RT-PCR. (3)Chose ovarian cancer cells SKOV3 as research object, the SKOV3 luciferase report carrier and miR-335 mimic were co-transfection to SKOV3, and the targeting effect of miR-335 to SKOV3 was verified by luciferase activity experiment. (4)Chose ovarian cancer cells SKOV3 as research object, which is divided into three groups: SKOV3 group(transfection mimic control), miR-335 mimic group(transfection miR-335 mimic)and miR-335 mimic+ROCK1 group(co-transfection miR-335 mimic+ROCK1). The cell proliferation activity of each group were detected by MTT method. The ROCK1 protein expression was detected by Western blot. The Cyclin D1 expression was detected by RT-PCR. Results: (1)RT-PCR result showed that, the miR-335 expression of ovarian cancer cells SKOV3 significantly lower than human normal ovarian epithelial cells IOSE80(P < 0.05). Western blot result showed that, the ROCK1 protein expression of ovarian cancer cells SKOV3 significantly higher than human normal ovarian epithelial cells IOSE80(P < 0.05). (2)RT-PCR result showed that, transfection miR-335 mimic made miR-335 expression of ovarian cancer cells SKOV3 significantly increased, and significantly higher than transfection mimic control(P < 0.05).(3)Luciferase activity experiment result showed that, transfection miR-335 mimic made the luciferase activity of ROCK1-Wt significantly decreased, but not inhibited the luciferase activity of ROCK1-Mut.(4)After transfection miR-335 mimic, the proliferative activity of ovarian cancer cells SKOV3 and the expression of Cyclin D1 significantly lower than negative control(P < 0.05). After transfection miR-335 mimic+ROCK1, the proliferative activity of ovarian cancer cells SKOV3 and the expression of Cyclin D1 significantly higher than only transfection miR-335 mimic(P < 0.05), but still significantly lower than negative control(P < 0.05). Western blot result showed that, after transfection miR-335 mimic, the ROCK1 protein expression of ovarian cancer cells SKOV3 significantly lower than negative control(P < 0.05). After transfection miR-335 mimic+ROCK1, the ROCK1 protein expression of ovarian cancer cells SKOV3 significantly higher than only transfection miR-335 mimic(P < 0.05), and still significantly higher than negative control(P < 0.05). Conclusion: miR-335 can target ROCK1 to inhibit the proliferation activity of ovarian cancer cells SKOV3.
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Received: 14 December 2015
Published: 16 March 2016
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