抗人DR5单克隆抗体——mDRA-6与顺铂协同杀伤白血病细胞HL-60作用研究

Synergistic effect of anti-human DR5(death receptor 5)monoclonal antibody(mDRA-6)and Cisplatin on HL-60 cells

目的:观察抗死亡受体5(Death receptor5,DR5)单克隆抗体———mDRA-6与顺铂(DDP)对HL-60细胞的协同杀伤作用。方法:DR5蛋白免疫BALB/c小鼠,融合筛选抗DR5杂交瘤细胞,制备抗DR5单抗———mDRA-6;流式细胞术测定顺铂对HL-60细胞表面DR5表达及细胞凋亡率;荧光显微镜下观察mDRA-6与顺铂协同作用下HL-60细胞形态变化;MTT法测定不同浓度的顺铂与mDRA-6对HL-60细胞存活的影响;琼脂糖凝胶电泳检测mDRA-6与顺铂联合对HL-60细胞DNA片段化的影响。结果:顺铂可诱导HL-60细胞表面DR5表达增加,mDRA-6与顺铂联用致HL-60细胞出现染色质浓缩、断裂,细胞出芽,凋亡小体形成等细胞凋亡形态学变化;250ng/ml的mDRA-6作用于HL-60细胞10小时,细胞凋亡率为16.61%;0.16μg/ml的DDP作用于HL-60细胞10小时,细胞凋亡率为2.35%;二者联合作用后,HL-60细胞凋亡率增至57.10%;mDRA-6与DDP联合作用HL-60细胞,DNA琼脂糖凝胶电泳显示明显“梯形”条带。结论:抗DR5单抗———mDRA-6与DDP对HL-60细胞具有强大的协同杀伤作用。

Objective ：To study the synergistic induction of apoptosis by the combination of anti-human DR5 （ death receptor 5 ） moneclonal antibody（ mDRA-6 ） and cisplatin （DDP） on HL-60 cells. Methods： To produce anti-human DR5 moneclonal antibody （mDRA-6） by immunizing BALB/c mice using DR5 protein. The change of DR5 expression after DDP-treated on HL-60 cells was detected by FACS. Morphologic changes of HL-60 cells were observed under fluorencence microscope. Cytotoxic and apeptotic effects of mDRA-6 and DDP on HL-60 cells were measured by MTT analysis. DNA fragmentation was detected by agarose gel electrophoresis. Results：DDP could induce DR5 expression on HL-60 cells. Chromatin condensation, budding and apeptotic bodies were observed in HL-60 cells treated by the combination of mDRA-6 and DDP; Death and apeptosis of HL-60 cells treated by the combination of mDRA- 6 and DDP were increased obviously. ＂DNA ladder＂ in the combination of mDRA-6 and DDP-treated HL-60 cells was exhibited obviously on agarose gel electrophoresis. Conclusion：Anti-human DR5 monoclonal antibody （mDRA-6） and DDP could synergistic induce apeptosis of HL-60 cells.