摘要
目的探讨解聚素-金属蛋白酶17(ADAM17)-短发夹RNA(shRNA)对人乳腺癌MCF-7细胞增殖的影响及其可能机制。方法将MCF-7细胞分为无义序列组、对照组、转染组。针对ADAM17基因设计合成4个特异性ADAM17-shRNA(ADAM17-shRNA-1219、ADAM17-shRNA-1508、ADAM17-shRNA-1134、ADAM17-shRNA-297),采用电穿孔法转染MCF-7细胞:无义序列组转染无义序列ADAM17-sh NC,转染组分别转染ADAM17-shRNA序列ADAM17-shRNA-1219、ADAM17-shRNA-1508、ADAM17-shRNA-1134、ADAM17-shRNA-297。对照组加入空白PBS,分别采用利用Real-time PCR、Western blotting法、i CELLigence法及流式细胞仪检测各组ADAM17 mRNA表达水平、ADAM17蛋白表达情况、细胞生长曲线和增殖活性及细胞周期变化。结果 4个ADAM17-shRNA序列对MCF-7细胞的ADAM17基因表达均有抑制作用,以shRNA1219载体的抑制效率最高,与对照组和无义序列组比较差异均具有统计学意义(P均<0.05);转染组ADAM17 mRNA及蛋白的表达水平明显低于无义序列组和对照组(P均<0.05);转染组细胞增殖活性和细胞生长速度较无义序列组和对照组均显著降低(P均<0.05);转染组细胞进入S期和G2/M期的比例降低,绝大多数细胞停留在G0/G1期,与对照组、无义序列组比较差异显著(P均<0.05)。结论 ADAM17-shRNA可抑制人乳腺癌MCF-7细胞的增殖活性,机制与其对ADAM17基因具有沉默作用有关。
Objective To investigate the effect of a disintegrin and metalloprotease 17 （ ADAM17 ）-shRNA on the proliferation of human breast cancer MCF-7 cells and to explore its possible mechanism .Methods MCF-7 cells were di-vided into the nonsense group , control group （ PBS） and transfection group .Four specific ADAM17-shRNAs （ ADAM17-shRNA-1219, ADAM17-shRNA-1508, ADAM17-shRNA-1134 and ADAM17-shRNA-297） were designed and synthesized for ADAM17 gene.MCF-7 cells were transfected by electroporation .The nonsense group was transfected by ADAM 17-shNC, the control group was added with PBS , and the transfection group was transfected by ADAM 17-shRNA-1219, AD-AM17-shRNA-1508, ADAM17-shRNA-1134 and ADAM17-shRNA-297, respectively.The expression of ADAM17 mRNA, the expression of ADAM17 protein, cell growth curve, proliferation activity and cell cycle of MCF-7 cells was respectively detected by real-time PCR, Western blotting, iCELLigence and flow cytometry .Results Four ADAM17-shRNA se-quences inhibited the expression of ADAM 17 gene, the shRNA1219 vector had the highest inhibitory efficiency , and com-pared with the nonsense group and control group , there was significant difference （all P〈0.05）.The expression level of ADAM17 mRNA and protein in the transfection group was significantly lower than that in the nonsense group and control group （all P〈0.05）.The cell proliferation activity and cell growth rate of the transfection group were significantly lower than those of the nonsense group and the control group （all P〈0.05）.The percentage of cells in S phase and G 2/M phase decreased in the transfection group , and most of the cells stayed in G0/G1 phase, which were significantly different from those in the control group and nonsense group （all P〈0.05）.Conclusion ADAM17-shRNA can inhibit the proliferation of human breast cancer MCF-7 cells, and the mechanism is related to its silencing effect on ADAM 17 gene.
作者
孟祥潮
蔡准
陈国福
吴丽君
张雪鹏
MENG Xiangchao CAI Zhun CHEN Guofu WU Lijun ZHANG Xuepeng(Affiliated Hospital of North China University of Science and Technology, Tangshan 063000, Chin)
出处
《山东医药》
CAS
北大核心
2017年第4期21-24,I0001,共5页
Shandong Medical Journal
基金
河北省自然科学基金资助项目(C2010001767)
唐山市科学技术研究与发展计划项目[唐科计(2014)16号]
作者简介
孟祥潮（1989-），男，硕士，研究方向为乳腺癌基础与临床。E—mail：13191777313@163．com
通信张雪鹏（1974-），男，博士后，教授，主任医师，研究方向为乳腺癌基础与临床。E-mml：syzxp@sina．COm
