Rac1蛋白调控乳腺癌MDA-MB-231细胞分泌血管内皮生长因子的研究  

Regulatory effect of Rac1 protein on vascular endothelial growth factor secreted in breast cancer MDA-MB-231 cells

作　　者：马骥[1] 赵庆丽[1] 赵易[1] 刘颖[1] 

机构地区：[1]解放军兰州军区兰州总医院乳腺科,730050

出　　处：《中华乳腺病杂志（电子版）》2016年第1期10-13,共4页Chinese Journal of Breast Disease(Electronic Edition)

基　　金：国家青年科学基金资助项目(81202085);甘肃省科技计划资助项目(1506RJDA296);兰州市科技计划资助项目(2014-1-39)

摘　　要：目的探讨Rac1蛋白对乳腺癌MDA-MB-231细胞分泌血管内皮生成因子(VEGF)的调控作用。方法在乳腺癌MDA-MB-231细胞中分别转染Rac1正显性真核表达质粒(V12Rac1组)及空白对照质粒(空白对照组)和Rac1小干扰RNA(siRac1组)及对照小干扰RNA(沉默对照组)后,通过ELISA分别检测转染后24、48、72 h由MDA-MB-231细胞分泌的VEGF蛋白水平;通过Western blot检测各组血管生成相关因子p53和VEGF表达的变化。两组Western blot定量分析数据比较采用t检验,VEGF分泌数据采用重复测量的方差分析。结果 ELISA分析结果显示:当MDA-MB-231细胞中高表达Rac1时,细胞分泌的VEGF水平随着时间的延长而逐渐升高,与空白对照组相比组间差异有统计学意义(组间比较:F=837.122,P<0.001;不同时间点比较:F=57 806.374,P<0.001;交互作用:F=7 663.095,P<0.001);当MDA-MB-231细胞中低表达Rac1时,细胞分泌的VEGF水平随着时间的延长逐渐升高,但低于沉默对照组,差异有统计学意义(组间比较:F=511.891,P<0.001;不同时间点比较,F=268.078,P<0.001;交互作用:F=120.708,P=0.001)。Western blot结果显示,当MDA-MB-231细胞高表达Rac1,与空白对照组相比,p53蛋白表达降低(t=-6.392,P=0.003),VEGF蛋白表达升高(t=7.497,P=0.002);当MDA-MB-231细胞中低表达Rac1,与沉默对照组相比,p53蛋白表达增加(t=5.307,P=0.006),VEGF蛋白表达降低(t=-7.395,P=0.002)。结论在乳腺癌MDA-MB-231细胞中,Rac1表达高低可以引起细胞分泌VEGF的水平变化,Rac1可能通过抑制p53表达而增加VEGF表达。Objective To investigate the regulatory effect of Racl, one member of Rho protein family, on vascular endothelial growth factor （VEGF） in breast cancer MDA-MB-231 cells. Methods MDA- MB-231 cells were transfected by Racl constitutively-active mutant plasmid pCEFL-GST-V12Racl （V12Racl group） , blank plasmid pCEFL-GST-neo （blank control group） , small interference Racl RNA （siRacl group） and small interference RNA of control （si-control group） respectively. VEGF expression levels in MDA-MB- 231 cells were determined by ELISA assay at different time points（24, 48, 72 h after transfection）. Western blot assay was performed to detect the relative expressions of angiogenesis-related factors p53 and VEGF. VEGF expression levels were compared by repeated measurement analysis of variance and t test was used to compare the quantitative data of Western blot. Results ELISA assays showed that after up-regulating Racl expression in MDA-MB-231 cells, VEGF level was gradually increased with time, indicating a significant difference compared with blank control group （group comparison： F= 837. 122, P〈0.001; comparison of different time points： F=57 806. 374,P〈0. 001 ; interaction of grouping and time： F=7 663. 095, P〈0. 001 ）.After down-regulating Racl, VEGF level was significantly lower than that in si-control group （group comparison： F=511. 891 ,P〈0. 001 ; comparison of different time points： F= 268.078,P〈0. 001 ; interaction of grouping and time： F= 120.708,P= 0. 001 ）. Western blot assay showed that in MDA-MB-231 cells, the increased Racl expression could inhibit p53 expression （t = -6. 392, P= 0. 003 ） and enhance VEGF expression （ t = 7. 497, P=0. 002） compared with blank control group, while decreased Racl expression could elevate p53 expression （t=-5. 307, P=0. 006） and inhibit VEGF expression （t=-7. 395, P=0. 002） compared with si- control group. Conclusion Racl expression could affect VEGF expression in breast cancer MDA-MB-231 ceils and Racl may promo

关 键 词：乳腺肿瘤 血管内皮生长因子类 肿瘤抑制蛋白质P53 Rac1蛋白质 人 

分 类 号：R737.9[医药卫生—肿瘤]