抗CD44 mAb A3D8对卵巢癌球形体形成细胞增殖和凋亡的影响*

摘要 目的：探讨抗CD44单克隆抗体(mAb)A3D8对3种卵巢癌球形体形成细胞增殖和凋亡的影响，并探讨其作用机制。方法：用MTS法检测A3D8对细胞增殖的影响；采用PI染色及流式细胞仪检测A3D8对细胞周期的影响；用Annexin V-FITC/PI凋亡检测试剂盒检测A3D8在细胞凋亡中的作用；用罗丹明123试剂盒检测A3D8对细胞线粒体膜电位的影响；并采用Western Blotting法检测A3D8作用于3种卵巢癌球形体形成细胞后，CDK2、cyclinA、Bcl-2和caspase-3的改变。结果：A3D8可抑制3种卵巢癌球形体形成细胞的增殖，且此抑制作用呈现剂量和时间依赖性；A3D8可在阻滞S期的同时降低G0/G1期比例；A3D8可促进3种细胞凋亡，与DDP联用，细胞凋亡率较单独使用DDP增高；A3D8的处理可导致3种细胞的线粒体膜电位损失，CDK2、cyclinA、Bcl-2蛋白表达下调，caspase-3表达上调。结论：A3D8可能是通过影响p21/CDK2/cyclinA途径阻滞细胞周期达到抑制3种卵巢癌球形体形成细胞增殖的结果，并且可通过线粒体途径促进细胞凋亡。

Influence of Anti-CD44 mAb A3D8 on Proliferation and Apoptosis of three Sphere-forming Cells from Ovarian Cancer

Abstract Objective: To explore the effects and mechanisms of A3D8(mAb) against CD44 on proliferation and apoptosis in three kinds of sphere-forming cells of ovarian cancer. Methods: The MTS assay was used to evaluate the cell proliferation. The PI staining and flow cytometry was used to analyze the cell cycle distribution. Annexin V-FITC/PI apoptosis detection kit was used to analyze cell apoptosis. Rhodamine123 apoptosis detection kit was used to detect the change of mitochondrial transmembrane potential (ΔψM). Western blotting assay was used to detect the CDK2,cyclinA,Bcl-2 and caspase-3 protein levels. Results: A3D8 inhibited the proliferation in three sphere-forming cells of ovarian cancer, the inhibition was in concentration-and time-dependant manner. A3D8 could cause S arrest with G0/G1 decrease via p21/CDK2/cyclinA pathways. A3D8 can increase the apoptosis rates of three kinds of cells, compared with DDP treatment alone, the apoptosis rates caused by A3D8 combined with DDP was increased significantly.Meanwhile A3D8 can lead ΔψM to decrease, CDK2,cyclinA,Bcl-2 protein level decrease and caspase-3 increase in above three kinds of sphere-forming cells. Conclusion: A3D8 inhibit the proliferation in three sphere-forming cells of ovarian cancer probably by the cell cycle arrest via p21/CDK2/cyclinA pathways and increase the apoptosis rates via mitochondrial pathways.