BAG-1通过MAPK/ERK通路调控乳腺癌细胞对吉非替尼敏感性的研究  

BAG-1 regulates the sensitivity of breast cancer cells to gefitinib through MAPK/ERK pathway

作　　者：高永昌 Gao Yongchang(Department of General Surgery,General Hospital of Tianjin Medical University,Tianjin 300052,China;Tianjin Institute of General Surgery,Tianjin 300052,China)

机构地区：[1]天津医科大学总医院普通外科,天津300052 [2]天津市普通外科研究所,天津300052

出　　处：《医药前沿》2024年第23期11-15,共5页Journal of Frontiers of Medicine

基　　金：天津市自然科学基金(应用基础研究多元投入基金项目)(21JCQNJC01660);天津市卫生健康科技项目(科技人才培育项目)(KJ20180)。

摘　　要：目的:探讨B淋巴细胞瘤-2(Bcl-2)结合抗凋亡基因1(BAG-1)在乳腺癌细胞中的表达及与吉非替尼作用敏感相关性研究。方法:应用小干扰RNA(siRNA)在MDA-MB-231和T47D细胞系中特异性干扰BAG-1的表达,应用0、0.1、1、5、10、20及50μmol/L不同梯度浓度吉非替尼暴露BAG-1 siRNA转染的细胞并应用MTT实验检查细胞生长增殖情况。以BAG-1 siRNA组为实验组,以未转染细胞和阴性对照control siRNA为对照组,在吉非替尼(10μmol/L)作用下检测不同组别细胞凋亡和周期变化。应用蛋白质印迹检测不同组别丝裂原活化蛋白激酶/细胞外信号调节激酶(MAPK/ERK)相关通路蛋白和磷酸化蛋白的表达。免疫组织化学分析87例三阴性乳腺癌BAG-1与表皮生长因子受体(EGFR)表达相关性。结果:干扰BAG-1表达后,相对于阴性对照,BAG-1siRNA实验组EGFRmRNA及其蛋白表达同样显著增加(P<0.05)。MTT检测MDA-MB-231以及T47D细胞中BAG-1表达减低后对不同浓度梯度吉非替尼介导的生长抑制变化差异有统计学意义(P<0.05),吉非替尼(10μmol/L)作用下MDA-MB-231、T47D的对照组比实验组细胞增殖抑制显著(P<0.05)。干扰BAG-1基因表达增加吉非替尼作用下细胞凋亡并促进细胞G1期阻滞(P<0.05)。BAG-1敲低时下调磷酸化MAPK激酶(p-MEK)和磷酸化ERK(p-ERK)表达(P<0.05),并上调小泛素相关修饰物(SUMO)特异性蛋白酶1(SENP1)和COP9信号小体5(CSN5)蛋白水平(P<0.05)。免疫组织化学结果验证三阴性乳腺癌标本中BAG-1与组织学分级和EGFR表达密切相关(P<0.05)。结论:BAG-1通过EGFR/MAPK/ERK通路抑制信号传导可能增强乳腺癌细胞对吉非替尼的敏感性,BAG-1基因是乳腺癌治疗的前瞻性生物学靶点。Objective The purpose of this study was to investigate the expression of B lymphocytoma-2(Bcl-2)associated athanogene-1(BAG-1)in breast cancer cells and its correlation with gefitinib sensitivity.Methods Small interfering RNA(siRNA)was used to specifically interfere with the expression of BAG-1 in MDA-MB-231 and T47D cell lines.BAG-1 siRNA transfected cells were exposed to gefitinib at different gradient concentrations of 0,0.1,1,5,10,20 and 50μmol/L and the cell viability was examined by MTT assay.Then take BAG-1 siRNA group as the experimental group,and control siRNA of non transfected cells and negative control as the control group.Under the effect of gefitinib(10μmol/L),detect the apoptosis and cycle change of cells in different groups.Western blot was used to detect the expression of mitogen-activated protein kinase/extracellular signal-regulated kinase(MAPK/ERK)related pathway proteins and phosphorylated proteins in different groups.In addition,87 cases of triple negative breast cancer were selected to analyze the correlation between BAG-1 and epidermal growth factor receptor(EGFR)expression in breast cancer patients by immunohistochemistry.Results After interfering with BAG-1 gene expression,the changes of EGFR mRNA and protein were detected.Compared with the negative control,the EGFR mRNA(P<0.05)and protein expression in BAG-1 siRNA experimental group were also significantly increased(P<0.05).MTT assay showed that the decreased expression of BAG-1 in MDAMB-231 and T47D cells had significant difference in the growth inhibition induced by gefitinib at different concentration gradients(P<0.05).When gefitinib(10μmol/L)was used,the cell proliferation of MDA-MB-231 control group was significantly inhibited compared with the experimental group(P<0.05).Similar results were obtained in T47D(P<0.05).Interference with BAG-1 gene expression increased cell apoptosis under the action of gefitinib(P<0.05)and promoted cell G1 phase arrest(P<0.05).At the same time,BAG-1 knockdown showed that p-mitogen extracellular kin

关 键 词：乳腺癌 BAG-1 表皮生长因子受体 吉非替尼 

分 类 号：R737.9[医药卫生—肿瘤]