hTERT基因沉默对乳腺癌细胞生物学特性及COX-2表达的影响  

Effect of hTERT gene silencing on biological characteristics and COX-2 expression in breast cancer cells

作　　者：于宏建[1] 赵杨[1] 赵世铭 马洪奎[1] 李光[2] YU Hong-jian ZHAO Yang ZHAO Shi-ming et al(Xianshuigu Hospital of Jinnan District, Tianjin, Laboratory, Tianjin 300350 ,Chin)

机构地区：[1]天津市津南区咸水沽医院检验科,天津300350 [2]天津医科大学基础医学院

出　　处：《中国实验诊断学》2016年第11期1817-1821,共5页Chinese Journal of Laboratory Diagnosis

摘　　要：目的本研究探讨hTERT基因沉默后对乳腺癌细胞生物学特性及环氧化酶-2(COX-2)的影响。方法设计合成特异性针对hTERT mRNA的siRNA,电转染入乳腺癌细胞,以致目的基因沉默。应用RT-PCR及Western blot法检测沉默hTERT基因后乳腺癌MCF-7细胞的mRNA及蛋白的表达状况;应用流式细胞仪检测细胞周期;并通过CCK-8增殖实验检测沉默hTERT基因后乳腺癌MCF-7细胞的增殖能力;经小室侵袭实验检测沉默hTERT基因后乳腺癌MCF-7细胞的侵袭能力。利用免疫组织化学法检测COX-2蛋白的表达。结果 hTERT基因沉默48h后,hTERT转染组在沉默hTERT基因后乳腺癌MCF-7细胞的mRNA及蛋白表达呈现明显降低(P<0.05);细胞周期检测结果示乳腺癌MCF-7肿瘤干细胞停滞在G0/G1周期,S期的细胞数目减低,三组比较统计学差异明显(P<0.05);CCK-8增殖实验结果表明乳腺癌MCF-7肿瘤干细胞在hTERT转染组的增殖得到显著抑制(P<0.05);小室侵袭实验结果表明,hTERT转染组穿过滤膜的细胞数量明显减少(P<0.05);免疫组织化学法检测结果显示,COX-2蛋白在hTERT转染组的表达明显降低,并与hTERT基因的表达相关联。结论 hTERT基因沉默后对乳腺癌细胞的增值和迁移能力均有影响,并能降低COX-2蛋白的表达,从而改善乳腺癌患者预后。Objective This study was to investigate the effect of hTERT gene silencing on the biological characteristics of breast cancer cells and to investigate the effect of hTERT gene silencing on the expression of cyclooxygenase-2（COX-2）in breast cancer cells.Methods The siRNA targeting hTERT mRNA was synthesized and cloned into pGenesil-1.1plasmid.The hTERT mRNA-siRNA expression vector was transfected into breast cancer cells by electroporation to produce RNA interference effect,so that the target gene was silenced.The mRNA and protein expression of breast cancer MCF-7cells were detected by RT-PCR and Western blot.The cell cycle was detected by flow cytometry.MCF-7cells,and the invasive ability of breast cancer MCF-7cells after silencing hTERT gene was detected by cell invasion assay.The expression of COX-2protein was detected by immunohisto chemistry.Results After hTERT gene silencing,the mRNA and protein expression of hTERT gene in breast cancer MCF-7cells after hTERT transfection were significantly lower than those in untransfected plasmid group and untransfected group（P（P〈0.05）.The results of CCK-8proliferation test indicated that the breast cancer MCF-7tumor stem cells stagnated in G0/G1 cycle and the number of cells in S phase was decreased,the difference was statistically significant（P〈0.05）The proliferation of MCF-7tumor stem cells in hTERT-transfected group was significantly inhibited（P〈0.05）.The invasion of MCF-7cells in hTERT-transfected group was significantly lower than that in control group（P〈0.05）,and the difference was statistically significant（P〈0.05）.The expression of COX-2protein in hTERT-transfected group was significantly lower than that in control group（P〈0.05）.The expression of COX-2protein was correlated with the expression of hTERT gene.Conclusion The silencing of hTERT gene may affect the proliferation and migration of breast cancer cells and decrease the expression of COX-2protein,so as to improve the prognosis of patients with breast cancer.

关 键 词：乳腺癌 HTERT基因 基因沉默 COX-2 

分 类 号：R737.9[医药卫生—肿瘤]