肿瘤坏死因子相关凋亡诱导配体受体单克隆抗体对人肾癌细胞的毒性作用

目的　探讨肿瘤坏死因子相关凋亡诱导配体受体 (TRAIL R)的单克隆抗体对肾癌细胞的毒性作用和机制。方法　在人肾癌细胞系ACHN和Caki 1中 ,应用四甲基偶氮唑盐 (MTT)方法分析细胞毒效应 ,应用流式细胞术检测相关凋亡诱导配体受体 (TRAIL R)的表达 ,应用荧光染色和AnnexinV标记检测细胞凋亡的发生 ,应用比色法分析多种Caspase活性在凋亡过程中的变化。结果　抗TRAIL R2单克隆抗体 (ETR2 )对人肾癌细胞系ACHN和Caki 1都具有明显的细胞毒性 (P <0 .0 1) ,而抗TRAIL R1单克隆抗体 (ETR1)的细胞毒性不显著 (P >0 .0 5 )。TRAIL R2在两种细胞表面均高表达 (ACHN :97.9% ,Caki 1:98.7% ) ,TRAIL R1表达微弱 (ACHN :7.6% ,Caki 1:7.5 4% )。ETR2通过诱导细胞凋亡发挥细胞毒作用 ,Caspase 8, 9, 6和 3活性在凋亡过程中明显增高 (P <0 .0 1)。结论　TRAIL R2的单克隆抗体在体外可诱导肾癌细胞发生凋亡 ,其细胞毒性与TRAIL R2的表达有关 ,在凋亡过程中内源性与外源性凋亡通路均被激活。

The cytotoxicity of monoclonal antibodies specific for TRAIL receptors on human renal cell carcinoma

Objective To study the cytotoxicity of monoclonal antibodies specific for tumor necrosis factor (TNF) related apoptosis inducing ligand (TRAIL) receptors on human renal cell carcinoma cell lines.Methods MTT assay was used to analyze the cytotoxicity.Flow cytometry was used to detect the expression of TRAIL R1,R2.Fluorescence staining and Annexin V were used to detect apoptosis,and colorimetric analysis was used to study Caspase activities.Results The monoclonal antibody specific for TRAIL R2 (ETR2) ( P < 0.01) was much more effective than the antibody for TRAIL R1 (ETR1) ( P > 0.05) to two kinds of renal cell carcinoma cell lines.TRAIL R2 was strongly expressed on both cell lines (ACHN: 97.9%, Caki 1: 98.7%), while TRAIL R1 was not (ACHN: 7.6%, Caki 1: 7.54%). ETR2 could induce apoptosis and the activities of Caspase 8, 9, 6,and 3 was increased during induction of apoptosis ( P < 0.01). Conclusion The monoclonal antibodies specific for TRAIL R2 can induce apoptosis of renal cell carcinoma cell lines in vitro.The cytotoxicity is related with the expression of TRAIL R2.During the induction of apoptosis,both extrinsic and intrinsic pathway is activated.