Beclin 1蛋白增强三阴性乳腺癌细胞对5-Fu敏感度的研究  

Beclin 1 Protein Enhances Sensitivity of Triple-negative Breast Cancer Cell Line BT-549 to 5-Fu by Promoting Cell Apoptosis

作　　者：范旭龙[1] 吴爱国 朱大江[1] 焦庆丽 郑临海[2] 邵国利[2] 王梦川[2] 纪术峰[2] 

机构地区：[1]南方医科大学附属佛山妇幼保健院乳腺疾病防治中心,佛山528000 [2]南方医科大学珠江医院普外科,广州510280

出　　处：《肿瘤防治研究》2015年第5期436-441,共6页Cancer Research on Prevention and Treatment

基　　金：广东省自然科学基金(S2012010009276)

摘　　要：目的探讨Beclin 1蛋白与三阴性乳腺癌细胞对5-Fu敏感度的关系。方法应用慢病毒介导p Len O-GTP-BECN1表达载体稳定转染至BT-549细胞,嘌呤霉素筛选阳性细胞,设立对照组。使用不同浓度5-Fu处理各组细胞,MTT法比较细胞对5-Fu的敏感度;流式细胞术和Hoechst33342实验检测细胞凋亡情况;Western blot检测细胞凋亡相关蛋白Caspase-3和Caspase-8表达情况。结果成功构建稳定表达Beclin 1蛋白的BT-549细胞株。p Len O-GTP-BECN1组对5-Fu敏感度显著高于其他两组;5-Fu作用48 h后,p Len O-GTP-BECN1组的IC50为(3.54±0.20)μg/ml,显著低于空白载体p Len O-GTP组(14.45±1.81)μg/ml和空白组(79.40±8.34)μg/ml(F=207.902;P<0.00)。5-Fu作用前,p Len O-GTP-BECN1组凋亡率为(12.14±0.76)%,同p Len O-GTP组(13.57±1.04)%和空白组(13.75±1.29)%(F=2.096;P=0.204)之间差异并无统计学意义;5-Fu作用48 h后,p Len O-GTP-BECN1组凋亡率为(51.01±3.26)%,显著高于p Len O-GTP组(42.17±4.6)%和空白组(38.42±2.93)%(F=9.32;P<0.01);接受5-Fu处理后,p Len O-GTP-BECN1组细胞核凋亡特征改变较其他两组更明显。5-Fu作用后,p Len O-GTP-BECN1组细胞Caspase-3相对表达量为(0.57±0.0028),显著高于p Len O-GTP组(0.42±0.010)和空白组(0.30±0.046)(F=46.134;P<0.01);p Len O-GTP-BECN1组细胞Caspase-8相对表达量为(0.44±0.0038),显著高于p Len O-GTP组(0.18±0.0095)和空白组(0.27±0.00060)(F=1006.757;P<0.00)。结论Beclin 1显著提高三阴性乳腺癌细胞对5-Fu的敏感度,其中促进凋亡发生可能是重要机制之一。Objective To investigate the effect of Beclin 1 protein on the sensitivity of triple-negative breast cancer cell line BT-549 to 5-Fu. Methods p Len O-GTP-BECN1 vectors were transfected into BT-549 cells respectively via lentiviral method. Puromycin selection was used to select positive cells. The negative and the blank control groups were established. BT-549 cells of each group were treated with different concentrations of 5-Fu. MTT assay was performed to compare the sensitivity of BT-549 cells to 5-Fu. FCM and Hoechst33342 dying method were employed to investigate cell apoptosis. Western blot was performed to detect the expression of apoptosis-related protein Caspase-3 and Caspase-8. Results BT-549 cells with stable expression of Beclin 1 were successfully established. The p Len O-GTP-BECN1 group was more sensitive to 5-Fu than other two groups. After treated by 5-Fu for 48 h, IC50 of 5-Fu in p Len O-GTP-BECN1 group was(3.54±0.20) μg/ml, which was significantly lower than those in p Len O-GTP group(14.45±1.81) μg/ml and blank group(79.40±8.34) μg/ml(F=207.902; P<0.00). Before 5-Fu treatment, the apoptosis rate of p Len O-GTP-BECN1 group was(12.14±0.76)%, which had no significant difference with those of p Len OGTP group(13.57±1.04)% and blank group(13.75±1.29)%(F=2.096; P=0.204); After treated by 5-Fu for 48 h, the apoptosis rate of p Len O-GTP-BECN1 group was(51.01±3.26)%, which was significantly higher than those of p Len O-GTP group(42.17±4.6)% and blank group(38.42±2.93)%(F=9.32; P<0.01). The characteristic changes of nucleus apoptosis in p Len O-GTP-BECN1 group were more apparent than those in the other two groups. After 5-Fu treatment, the relative expression of Caspase-3 in p Len O-GTP-BECN1 group was(0.57±0.0028), which was significantly higher than those in p Len O-GTP group(0.42±0.010) and blank group(0.30±0.046)(F=46.134; P<0.01);and the relative expression of Caspase-8 in p Len O-GTP-BECN1 group was(0.44±0.0038),which was significantly higher than those in p Len O-GTP group(0.18±0.00

关 键 词：BECLIN 1 三阴性乳腺癌 5-氟尿嘧啶 凋亡 

分 类 号：R737.9[医药卫生—肿瘤]