抗人DR5抗体mDRA- 6细胞毒作用机制分析

目的:探讨鼠抗人DR5单克隆抗体(mAb)mDRA-6对Jurkat细胞的细胞毒作用及其机制。方法:以流式细胞术测定mAbmDRA-6对Jurkat细胞的细胞毒作用和细胞凋亡作用,以及caspase8、9的抑制剂对mAbmDRA-6诱导的Jurkat细胞凋亡的影响。在荧光显微镜下,观察mAbmDRA-6对Jurkat细胞形态的影响。以琼脂糖凝胶电泳检测Jurkat细胞中的DNA片段化。结果:mAbmDRA-6对Jurkat细胞具有显著的细胞毒作用,并呈剂量和时间依赖性。经mAbmDRA-6处理后,Jurkat细胞可出现典型的细胞凋亡的形态特征:细胞膜皱缩,出泡,染色质浓缩,形成凋亡小体等。经mAbmDRA-6处理后,Jurkat细胞膜表面高表达丝氨酸磷脂,并可导致Jurkat细胞中的DNA片段化。caspase8的抑制剂可明显抑制mAbmDRA-6诱导的Jurkat细胞凋亡,caspase9的抑制剂的影响很小。结论:mAbmDRA-6可通过死亡受体信号传导途径诱导Jurakt细胞凋亡,对Jurkat细胞产生细胞毒作用,其在以TRAIL/DR5系统进行的肿瘤治疗和探讨DR5功能结构域方面具有广阔的应用前景。

Cytotoxic mechanism of anit-human death receptor 5 monoclonal antibody mDRA- 6

s AIM: To investigate the cytotoxic action and its mechanism of a novel anti-human DR5 monoclonal antibody (mAb mDRA-6). METHODS: The cytotoxic action of mAb mDRA-6 on Jurkat cells and the effects of inhibitors of caspase 8 and caspase 9 on apoptosis of Jurkat cells induced with mAb mDRA-6 were detected by flow cytometry. The effects of mAb mDRA-6 on the morpha of Jurkat cells was observed by fluorencence microscope. The apoptosis of Jurkat cells was detected by flow cytometry with Annexin V-FITC/PI staining. The DNA fragmentation in Jurkat cells was analysed by agrose gel electrophoresis. RESULTS: mAb mDRA-6 exerted cytotoxicity on Jurkat cells in dose-dependent and time-dependent manner. Jurkat cells treated with mDRA-6 exhibited typic apoptostic features in morphology, namely, membrane crenation, bubbling, chromatin condensation, and formation of apoptotic bodies. The flow cytometry analysis showed that phosphatidylserine (PS) was highly expressed in Jurkat cells treated with mDRA-6. Agrose gel electrophoresis indicated that DNA fragmentation occurred in Jurkat cells. Inhibitor of caspase 8 inhibited the apoptosis of Jurkat cells induced with mDRA-6 while Inhibitor of caspase 9 showed less effect. CONCLUSION: mDRA-6 may exert cytotoxicity by inducing Jurkat cell apoptosis through signal transduction pathway of death receptors, which may be a useful tool in treating tumors with DR5 as target molecule and exploring the functional domain of DR5.