BTBD7基因通过作用于Wnt/β-catenin信号通路促进人乳腺癌MCF-7细胞的侵袭转移  

BTBD7 promotes the invasion and metastasis of breast cancer MCF-7 cells by targeting Wnt / β-catenin signaling

作　　者：李峻[1] 李刚[1] 谢玉莲[1] 王敏洪[1] 

机构地区：[1]解放军第458医院乳腺外科,广州510602

出　　处：《中华乳腺病杂志（电子版）》2015年第6期357-362,共6页Chinese Journal of Breast Disease(Electronic Edition)

摘　　要：目的观察敲减BTBD7基因后人乳腺癌MCF-7细胞的运动侵袭能力的变化,并探讨其作用机制。方法将经慢病毒转染shRNA,敲减BTBD7基因表达后的细胞作为实验组（MCF-7-sh BTBD7）,未进行慢病毒转染的细胞为对照组（MCF-7-vec）。应用划痕愈合实验、Transwell侵袭实验、MTT法绘制细胞生长曲线等方法来检测实验组和对照组内MCF-7细胞的侵袭转移以及增殖能力之间的区别,两组试验结果比较采取独立样本t检验。在此基础上通过肿瘤信号通路筛选芯片筛选出BTBD7作用的下游信号通路,并以Western blot实验验证这条信号通路是否确实参与其中发挥作用。结果划痕愈合实验结果显示,与对照组相比,实验组MCF-7细胞的迁移能力显著降低[24 h愈合率：（85.95±5.30）%比（43.38±4.01）%,t=18.108,P〈0.001];Transwell侵袭实验显示实验组和对照组细胞侵袭能力出现明显的差别,与对照组相比,实验组细胞的侵袭能力显著降低（24 h细胞计数：152±7比50±8,t=27.378,P〈0.001）;MTT实验显示从第4天开始两组细胞增殖能力出现差异,对照组细胞数为（3.17±0.23）×10^4/ml,明显高于实验组的细胞数为（2.58±0.21）×10^4/ml,差异具有统计学意义（t=3.189,P〈0.050）;第7天对照组的细胞数为（9.57±0.36）×10~4/ml,仍然显著高于实验组的细胞数（7.29±0.37）×10^4/ml,差异具有统计学意义（t=7.654,P〈0.050）。通过双荧光素酶检测系统检测肿瘤信号通路筛选芯片的结果表明Wnt信号通路的相对荧光强度在两组间差异有统计学意义（t=12.509,P〈0.001）,Western blot实验结果同样表明,与对照组相比,实验组的c-Myc、MMP-7以及β-catenin的蛋白水平均发生变化。结论 BTBD7基因可能通过作用于Wnt/β-catenin信号通路促进人乳腺癌MCF-7细胞的侵袭转移。Objective To explore whether knock-down of BTBD7 inhibits the invasion and metastasis of breast cancer cells MCF-7 and the underlying mechanism. Methods The expression of BTBD7 was knocked down in MCF-7 cells by lentivirus transfection. The transfected MCF-7 cells served as the experimental group（ MCF-7-sh BTBD7） and the untransfected cells served as the control group（ MCF-7-vec）. Then wound healing assay,Transwell assay and MTT method（ to draw cell growth curve） were performed to detect the proliferation,invasion and metastasis abilities of MCF-7 cells of two groups. The results were compared between two groups using independent sample t test. Moreover,Cignal finder cancer 10-pathway reporter array was also performed to identify the exact pathway participating in BTBD7 functioning. Western blot assay was performed to confirm the related pathways. Results After knocking down BTBD7,wound healing assay showed the migration ability in the experimental group were significantly inhibited compared with the control group[24 h healing rate：（ 43. 38 ± 4. 01） % vs（ 85. 93 ± 5. 30） %,t = 18. 108,P〈0. 001]. Transwell assay demonstrated that the invasion of MCF-7 cells were also significantly suppressed in the experimental group compared with the control group（ 24 h cell count： 50 ± 8 vs 152 ± 7,t = 27. 378,P〈0. 001）. MTT assay showed that there was a significant difference between the experimental group and the control group in cell count on day 4 [（ 3. 17 ± 0. 23） × 10^4/ ml vs（ 2. 58 ± 0. 21） × 10^4/ ml,t = 3. 189,P〈0. 050],and on day 7 [（ 7. 29 ± 0. 37） × 10^4/ ml,as compared to the control group（ 9. 57 ± 0. 36） × 10^4/ ml,t = 7. 654,P〈0. 050]. Result of Cignal finder cancer 10-pathway reporter array by dual luciferase reporter system showed that the relative fluorescence intensity of Wnt signaling pathways was significantly different between two groups（ t = 12. 509,P〈0. 001）. Western Blot assay confirmed that the protein levels of c-Myc,MMP-7 and β-ca

关 键 词：乳腺肿瘤 同源盒结构域蛋白质类 WNT蛋白质类 肿瘤侵润 肿瘤转移 

分 类 号：R737.9[医药卫生—肿瘤]