| 摘要: |
| 目的通过RNA干扰技术抑制卵巢癌SKOV3细胞分泌型clusterin(sCLU)基因的表达,研究其对卵巢癌细胞增殖、凋亡及侵袭能力的影响。方法通过脂质体介导靶向sCLU基因的siRNA转染卵巢癌SKOV3细胞,实验分为4组: 实验组(sCLUsiRNA +脂质体)、阴性对照组Ⅰ(negative control siRNA +脂质体)、阴性对照组Ⅱ(sCLUsiRNA)、空白对照组(等体积的完全培养液)。RTPCR和蛋白质印迹分析法检测sCLU表达；利用MTT法、Transwell小室体外侵袭实验及流式细胞仪Annexin Ⅴ/PI法检测评价sCLU基因抑制后对卵巢癌细胞增殖、侵袭及凋亡的影响。结果靶向sCLU基因的siRNA明显、特异性地抑制了sCLU mRNA和蛋白的表达水平；MTT实验结果显示实验组细胞增殖能力较各对照组明显降低,差异有统计学意义(P<0.05)。流式细胞仪Annexin Ⅴ/PI法检测结果显示实验组细胞凋亡率较各对照组明显升高,达到(15.84±1.53)%,较空白对照组升高了约9%,差异有统计学意义(P<0.01)。侵袭实验结果提示实验组细胞侵袭能力受到明显抑制,实验组穿膜细胞数量为(26.52±6.22)个/视野,较各对照组明显降低,差异有统计学意义(P<0.01)。结论sCLU基因沉默明显抑制了卵巢癌SKOV3细胞的增殖、侵袭能力,并增加了其凋亡率,sCLU基因有望成为卵巢癌治疗的新靶点。 |
| 关键词: clusterin 卵巢肿瘤 小分子干扰RNA 细胞增殖 肿瘤侵袭 |
| DOI：10.3724/SP.J.1008.2011.028 |
| 投稿时间：2010-08-19修订日期：2010-12-28 |
| 基金项目: |
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| Influence of clusterin gene silencing on proliferation and invasion of human ovarian carcinoma SKOV3 cell line |
| HUANG Luo1,LUO Qian2,CHEN Cheng1,ZHANG Xing-ping1,WU Yong-zhong2* |
| (1. Department of Oncology, the First Affiliated Hospital, Chongqing Medical University, Chongqing 400016, China;;2. Department of Radiotherapy,Chongqing Tumor Hospital, Clinical College of Chongqing Medical University, Chongqing 400030, China) |
| Abstract: |
| ObjectiveTo investigate the influence of secretory clusterin(sCLU) gene silencing by RNA interference on the proliferation, apoptosis and invasion of human ovarian carcinoma SKOV3 cell line. MethodsThe human ovarian carcinoma SKOV3 cells were transfected with sCLUtargeting siRNA by Lipofectamine 2000. The study included four groups: the experimental group (sCLUsiRNA+Lipofectamine 2000), negative groupⅠ (negative control siRNA+Lipofectamine 2000),negative groupⅡ(sCLUsiRNA) and blank control group. The expressions of mRNA and protein were examined by RTPCR and Western blotting analysis. The proliferation, apoptosis and invasion ability of SKOV3 cells were evaluated by MTT assay, flow cytometry and Transwell assay. ResultsThe sequencespecific siRNA of sCLU effectively and specifically downregulated the expression of sCLU at both mRNA and protein levels. The MTT results showed that the proliferation of cells in the experimental group was significantly inhibited compared with those in the other groups(P<0.05). Flow cytometry demonstrated that the apoptotic rate of cells in the experimental group([15.84±1.53]%) increased by 9% compared with those in the rest groups(P<0.01).We also found that the matrigel penetrating capacity of cells in the experimental group ([26.52±6.22] cells/field)was significantly inhibited compared with those in the rest groups(P<0.01).ConclusionSilencing of sCLU can effectively inhibit the proliferation and invasion of human ovarian carcinoma SKOV3 cells, and it can also greatly promote their apoptosis, making sCLU gene a potential new target for ovarian carcinoma treatment. |
| Key words: clusterin ovarian neoplasms small interfering RNA cell proliferation neoplasm invasiveness |
