抗人DR5单克隆抗体诱导HL-60细胞凋亡机制  

The mechanism of HL-60 cell apoptosis induced by anti-human DR5 monoclonal antibody

作　　者：李淑莲[1] 王赟[1] 张舒曼[1] 王靖[1] 

机构地区：[1]河南大学细胞与分子免疫学重点实验室、河南大学免疫学研究所,开封475004

出　　处：《现代免疫学》2012年第5期399-403,共5页Current Immunology

基　　金：国家自然科学基金资助项目(30571697)

摘　　要：探讨抗人DR5单克隆抗体(mDRA-6)诱导白血病细胞系HL-60细胞凋亡的可能机制。MTT法检测mDRA-6对HL-60细胞的生长增殖的影响,以及Caspase 8、10、3抑制剂对mDRA-6抑制HL-60细胞生长增殖的影响;琼脂糖凝胶电泳检测HL-60细胞DNA片断化降解;Western blot检测mDRA-6对HL-60细胞Caspase 8、10、3的激活改变。结果发现,mDRA-6呈时间、浓度依赖性地抑制HL-60细胞的生长增殖,10 mg/L的mDRA-6作用HL-60细胞6 h、8 h和10h,细胞增殖抑制率分别为23.96%、44.10%和50.28%;琼脂糖凝胶电泳显示,10 mg/L的mDRA-6作用6 h,HL-60细胞呈现凋亡细胞特有的DNA梯形条带;Western blot检测结果发现,mDRA-6作用HL 60细胞不同时间,Caspase 8均只有酶原条带出现,无激活片段产生,而Caspase 10和Caspase 3显示明显裂解片段产生,并随mDRA-6作用时间延长而增多;预先使用15μmol/L的Caspase 10及Caspase 3抑制剂孵育细胞1 h,能够使mDRA-6对HL-60细胞的生长抑制率分别降低64.15%(t=10.13、P<0.01)和53.69(t=8.93、P<0.01)%,而预先使用15μmol/L的Caspase 8抑制剂孵育细胞1 h,mDRA-6对HL-60细胞的生长抑制率仅降低4.40%(t=0.52、P>0.05)。以上实验结果提示,mDRA-6通过激活Caspase 10启动凋亡信号分子,诱导白血病HL-60细胞凋亡。To study the mechanism of HL60 cell apoptosis induced by antihuman DR5 monoclonal antibody, HL60 cells were treated with mDRA6 or / and Caspase inhibitors, and the inhibition ratio was determined using MTT assay. The DNA ladder of HL60 cells treated with mDRA6（10mg/L） for 6 h was analyzed by 1% agrose gel electrophoresis. The expression of Caspase 8, Caspase 10 and Caspase 3 of HL60 cells after treating with mDRA6 were detected by Western blotting, respec tively. It was demonstrated that the proliferation of HL60 cell after treating with mDRA6 was inhibited in a dosedependent and timedependent manner, the inhibition ratio of HL60 cells treated by 10 /μg/ml mDRA6 for 6 h,8 h and 10 h were 23. 96%, 44.10% and 50.28%, respectively. DNA fragmentation detection indicated that HL60 cell apoptosis was induced by mDRA6. Western blotting revealed that Caspase 10 and Caspase 3 in HL 60 cells treated with mDRA6 were activated in a timedependent manner, but Caspase 8 did not. The HL60 cells were incubated with the inhibitors for Caspase 8, 10 and 3 for 1 hour, respectively, and then the cells were treated with mDRA6, HL60 cell apoptosis induced by mDRA6 were reduced by 4. 40%（t=0. 52, P〉0.05）, 64.15%（t=10.13, P〈0.01） and 53.69（t=8.93, P〈0.0]）%, respectively. Therefore, it is evident that Caspase 10 inhibits apoptosis of HL60 cells pretreated with antihuman DR5 monoclonal antibody mDRA6.

关 键 词：死亡受体5 单克隆抗体 凋亡 半胱天冬酶 

分 类 号：R392[医药卫生—免疫学]