凋亡蛋白Bax在肿瘤坏死因子凋亡诱导配体诱导恶性淋巴瘤细胞耐药中的调节作用  

The modulating effect of proapoptotic protein Bax on the resistance of malignant lymphoma cells to tumor necrosis factor related apoptosis inducing ligand.induced apoptosis

作　　者：郝继辉[1] 俞鸣[1] 贾丽[1] 柳凤亭[1] 李强[1] 郝希山[1] 

机构地区：[1]天津医科大学附属肿瘤医院腹部外科,30060

出　　处：《中华医学杂志》2007年第2期134-137,共4页National Medical Journal of China

基　　金：天津市教育委员会自然科学基金(20040217)

摘　　要：目的探讨凋亡蛋白 Bax 在恶性淋巴瘤细胞对肿瘤坏死因子凋亡诱导配体(TRAIL)凋亡抵抗中的调节作用,为临床克服肿瘤耐药提供治疗靶点。方法分别采用500μg/L TRAIL 和/或10 nmol/L 蛋白酶体抑制剂 PS-341处理恶性淋巴瘤细胞(CRL)和正常对照细胞(HRC)。流式细胞仪检测细胞凋亡情况。荧光显色技术检测细胞半胱氨酸蛋白水解酶-8酶活性。Western 印迹检测细胞中 Bax 蛋白表达水平。免疫沉淀技术(IP)检测 Bax 蛋白的活性及构象变化。结果与 HRC 细胞比较,CRL 细胞对 TRAIL 存在显著抵抗,24 h 凋亡指数仅为21%,前者为32%,两者比较差异有统计学意义(P<0.01);同时在 TRAIL 作用过程中 CRL 细胞中 Bax 表达呈下降趋势,24 h 表达量仅为正常量的5/17。当联合使用 PS-341后,可有效抑制 Bax 蛋白降解,6 h 表达量为正常量的1.2倍,显著增加活性;同时增加半胱氨酸蛋白酶-8酶活性,6 h 为26.5 μmol·L^(-1)·h^(-1)·mg^(-1)蛋白,从而增加细胞凋亡,6 h 凋亡指数达54%,与对照组比较差异均有统计学意义(均 P<0.01)。结论促凋亡蛋白 Bax 的下调表达参与恶性淋巴瘤细胞对 TRAIL 耐药性。合理的使用蛋白酶体抑制剂可通过抑制蛋白降解,增加其活性来有效克服肿瘤耐药。Objective To elucidate the modulating effect of proapoptotic protein Bax on the resistance of malignant lympboma cells to tumor necrotic factor-related apoptosis inducing ligand （TRAIL）- induced apoptosis and offer evidence for clinic work. Methods Human malignant lymphoma cells of the line CRL and normal HRC cells were cultured and treated by 500μg/L TRAIL（ Group T）, treated by 10 nmol/L PS-341 （ Group P）, or pre-treated by 10 nmol/L proteasome inhibitor PS-341 for 1 hour and then treated by 500 μg/L TRAIL（ Group P ＋ T）. Flow cytometry was used to detect the cell apoptosis. Western blotting was used to detect the expression of Bax protein. Caspase-8 activity was tested by fluorophotometer. Immunoprecipitation method was used to examine the conformation change of Bax protein. Results The apoptosis index 24 hours after treatment of the CRL cells of Group T was 21%, significantly lower than that of the HRC cells of Group T （32%, P 〈0. 01 ）. The Bax protein expression amount 24 hours after treatment of the HRC cells of Group T was 1.8 times that of the normal cells, and the Bax protein expression amount 24 hours after treatment of the CRL cells of Group T was only 5/17 that of the normal amount. The apoptotic index 6 hours after treatment of the CRL cells of Group P ＋ T was 54%, significantly higher than those of Groups T and P （ both P 〈 0. 01 ）. The caspace-8 activity 6 hours after treatment of the CRL cells of Group P ＋ T was 26. 5 μ mol · L^-1 · h^-1 ·mg^-1 protein, similar to that of the HRC cells of Group P ＋ T （27.2 Pμ mol · L^-1 · h^-1 ·mg^-1 protein）, and significantly higher than those of the other cells （all P 〈0.01 ）. The Bax protein expression 5 hours after treatment of the Group P HRC and CRL cells were 2.5 times and 1.2 times that of the control cells. The Bax protein expression of the HRC cells of Group P ＋ T was 3.3 times that of the normal controls, and the Bax degradation of the CRL cells of Group P ＋ T was significantly reduced. The com

关 键 词：淋巴瘤 脱噬作用 BAX TRAIL 

分 类 号：R733.1[医药卫生—肿瘤]