CD44单克隆抗体A3D8通过抑制ERK1_2上调Bim诱导HL-60细胞早期凋亡.doc

  CD44 单克隆抗体 A3D8 通过抑制 ERK1/2 上调 Bim 诱导 HL-60 细胞早期凋亡# 常国强，王建，高伟，王若君，王立洪，金薇娜，蔺亚妮，李华文，庞天翔 5 10 15 20 25 30 35 40 （中国医学科学院北京协和医学院血液学研究所血液病医院，实验血液学国家重点实验室， 天津 300020） 摘要：CD44 单克隆抗体 A3D8 可引起急性髓系白血病（acute myeloid leukemia，AML）细 胞增殖抑制，并诱导其早期凋亡，本研究主要探讨在此过程中 ERK 以及 Bcl-2 家族成员的 作用机制，为治疗白血病提供新方法和新的药物靶点。用 MTT 法检测 A3D8 对 HL-60 细胞 的增值抑制效应；用流式细胞术检测 A3D8 对 HL-60 细胞线粒体膜电位的变化；用实时定 量 PCR 技术检测 Bim 基因在 mRNA 水平的表达变化；用 western blot 检测 A3D8 处理后磷 酸化 ERK-1/2 的表达变化；结果表明：A3D8 能显著抑制 HL-60 细胞的增殖，抑制效率呈现 剂量和时间的量效关系；并可进一步诱导 HL-60 细胞的早期凋亡，而 Bcl-2 家族成员 Bim 的表达水平也随时间和浓度的变化而改变，磷酸化 ERK-1/2 的表达水平明显降低。结论： CD44 单克隆抗体 A3D8 可通过降低磷酸化 ERK-1/2 的表达来调节 Bim，从而诱导 HL-60 细 胞的增殖抑制和早期凋亡。 关键词：HL-60 细胞；A3D8 抗体；p-ERK；Bim 中图分类号：R733.7 Early Apoptosis of HL-60 Cells Induced by Anti-CD44 McAb A3DR Inhibiting ERK1/2 Up-regulated Bim Expression CHANG Guoqiang, WANG Jian, GAO Wei, WANG Ruojun, WANG Lihong, JIN Weina, LIN Yani, LI Huawen, PANG Tianxiang (State Key Laboratory of Experimental Hematology, Institute of Hematology Blood Disease Hospital, Chinese Academy of Medical Sciences Peking Union Medical College, TianJin 300020) Abstract: In this paper, to investigate the effects of anti-CD44 mAb A3D8 on the cell proliferation and apoptosis and to study the mechanism of regulating these progresses via ERK1/2 and Bim. Effects of the anti-CD44 mAb A3D8 on the HL-60 cells proliferation were assayed with MTT method, and the change of mitochondrial transmembrane potential of HL-60 cells was analyzed by flow cytometry. The mRNA expression of Bim was determined by real-time quantitative RT-PCR. Wersten blot was used to determine the protein expression of p-ERK1/2. mAb A3D8 could remarkably inhibit the proliferation capacity of the HL-60 cells in a manner of dosage- and time-dependent way. The mitochondrial transmembrane potential was significantly decreased than the control group when the cells was treated with A3D8 (3.0μg/ml). Furthermore, the mRNA expression of Bim was much higher than that in normal controls. Expression of t