BCL9基因沉默对乳腺癌细胞株侵袭及迁移能力作用研究

发布时间：2018-05-30 04:13

  本文选题：乳腺癌 + B细胞淋巴因子9　； 参考：《河北医科大学》2017年硕士论文

【摘要】：目的:初步研究BCL9基因在三种乳腺癌细胞株中的表达差异,分析三种不同乳腺癌细胞株细胞生物学特性:增殖、侵袭及迁移能力的差异及其意义,并进一步探讨BCL9基因的沉默对乳腺癌细胞株侵袭及迁移能力的影响及意义。方法:1分析三种不同表型的人乳腺癌细胞株,采用实时荧光定量PCR法(Real-time PCR)和蛋白免疫印迹法(Western blot)检测人乳腺癌细胞株MDA-MB-231、MCF-7和MDA-MB-453中BCL9的mRNA及蛋白表达水平,筛选出高表达BCL9的细胞株。2通过流式细胞术检测细胞周期,计算增殖指数,比较三种细胞株之间增殖能力。采用细胞划痕实验和Transwell细胞侵袭实验比较三者细胞迁移和侵袭能力。3构建BCL9慢病毒质粒shRNA,针对高表达BCL9的细胞株做BCL9基因沉默实验。4采用Transwell细胞小室以及细胞划痕实验进一步研究BCL9基因的敲除对乳腺癌细胞株侵袭及迁移能力的影响。结果:1 RT-PCR检测细胞株BCL9的mRNA表达:MDA-MB-231(0.016±0.004)MCF-7(0.008±0.002)MDA-MB-453(0.004±0.002),(各组间比较,P0.05)差异有统计学意义。BCL9蛋白水平在细胞间表达量为MDA-MB-231(0.629±0.101)MCF-7(0.397±0.196)MDA-MB-453(0.204±0.990)(各组间比较,P0.05),差异有统计学意义;2流式细胞术增殖指数显示MDA-MB-231(0.483±0.010)MDA-MB-453(0.358±0.032),MCF-7(0.397±0.061)(各组间比较,P0.05),差别有统计学意义;Transwell细胞侵袭实验:穿膜细胞数:MDA-MB-231(417.80±6.94)MDA-MB-453(379.60±11.99)MCF-7(9.40±1.14)(各组间比较,P0.05),差异有统计学意义;细胞划痕实验:MDA-MB-231(0.716±0.021)MDA-MB-453(0.331±0.021)MCF-7(0.272±0.017)(各组间比较,P0.05),差异有统计学意义。3针对人乳腺癌细胞株MDA-MB-231运用慢病毒细胞转染技术进行BCL9基因沉默;4 Transwell细胞小室实验细胞数:空白组(420.50±3.704)/阴性对照组(417.00±5.310)实验组(151.50±7.580)(P0.05),差异有统计学意义;细胞划痕实验划痕愈合率:空白组(0.712±0.127)/阴性对照组(0.709±0.015)实验组(0.382±0.012)(P0.05),差异有统计学意义。结论:1通过检测三种人乳腺癌细胞株中BCL9基因及蛋白的表达水平,发现BCL9在三阴性乳腺癌细胞株MDA-MB-231细胞中表达水平较高。2通过分析三种不同表型乳腺癌细胞株,细胞的增殖、侵袭和迁移能力均有差别,三阴性乳腺癌的MDA-MB-231细胞株的增殖、侵袭和迁移能力均较高。3 BCL9基因的沉默可有效抑制人乳腺癌细胞株MDA-MB-231侵袭及迁移能力,提示BCL9可能成为抑制乳腺癌细胞侵袭及转移新的分子靶点。
[Abstract]:Objective: to study the difference of BCL9 gene expression in three breast cancer cell lines, and analyze the biological characteristics of three different breast cancer cell lines: proliferation, invasion and migration. The effect and significance of BCL9 gene silencing on the invasion and migration of breast cancer cell lines were investigated. Methods three human breast cancer cell lines with different phenotypes were analyzed. The mRNA and protein expression of BCL9 in MDA-MB-231 MCF-7 and MDA-MB-453 were detected by real-time fluorescence quantitative PCR and Western blot. The cell lines with high expression of BCL9 were selected. The cell cycle was detected by flow cytometry, the proliferation index was calculated, and the proliferative ability of the three cell lines was compared. Cell scrape assay and Transwell cell invasion assay were used to compare the cell migration and invasion ability of three cells to construct BCL9 lentivirus plasmid shRNAs. The BCL9 gene silencing assay was performed on the cell lines with high expression of BCL9. 4. Transwell cell compartment and cell division were used. The effect of BCL9 knockout on the invasion and migration of breast cancer cell lines was further studied. 缁撴灉:1 RT-PCR妫，

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