99tcm-mibi显像评价多药耐药及其逆转-第三军医大学学报.docVIP

乳腺癌多药耐药细胞凋亡通路的研究 张雪梅 杨秀蓉 吴华 张振蔚 鲜于志群 目的 探索耐阿霉素的乳腺癌细胞株MCF-7/Adr凋亡的可能途径。方法 采用耐药株MCF-7/Adr和野生株MCF-7/wt两种细胞株进行对比，以噻唑蓝分析法（MTT法）摸索出H2O2导致其线粒体功能下降的浓度，观察细胞形态学和线粒体超微结构的改变。应用DNA凝胶电泳、Annexin V和PI双标记流式细胞术检测细胞凋亡发生，Rh123共聚焦显微镜检测线粒体膜电位和P-gp功能，Western blot 检测细胞色素C（Cyto C）的释放状况。结果　4mmol/L H2O2引起MCF-7/Adr线粒体整体功能不可逆的进行性下降，线粒体空化,嵴肿胀。MCF-7/Adr凋亡发生率显著高于MCF-7/wt，凋亡率分别为96.52％和18.57％（P〈0.001）。凋亡DNA电泳仅见800、1600两个分子片段。MCF-7/Adr线粒体较MCF-7/wt量减少、膜电位下降。4mmol/L H2O2处理使MCF-7/Adr线粒体Cyto C的释放至胞浆，MCF-7/wt仅少量Cyto C释放。结论　P-gp阳性细胞，因其线粒体的结构和功能改变的特点，较其野生株细胞更易经线粒体途径凋亡。提示以线粒体为靶点的药物有助于克服与P-gp表达有关的多药耐药。 [主题词] 肿瘤； 多药耐药； 凋亡； H2O2； 线粒体； 细胞色素C。 _______ 作者单位： 张雪梅，杨秀蓉。430070 广州军区武汉总医院核医学科 吴华，张振蔚，鲜于志群。430030 华中科技大学同济医学院附属同济医院核医学科 通讯作者：张雪梅，E-mail:plumzhang108@ ________ Corresponding author：Zhang Xuemei，E-mail:plumzhang108@ The apoptotic pathway of breast cancer multidrug resistant cell ZHANG Xue-mei, YANG Xiu-rong, WU Hua, Zhang Zhen-wei, XIANYU Zhi-qun. Department of Nuclear Medicine, Wuhan General Hospital, Guangzhou Military Command, Wuhan, 430070, China [Abstract] Objective To investigate the possible apoptotic pathway in breast cancer daunorubicin-resistant subline, MCF-7/Adr cell. Methods MCF-7/Adr,expressing functional P-gp and its wild type MCF-7/wt were used in contrast.The concentration of hydrogen peroxide(H2O2) that may lead to loss of mitochondrial function was determined using a MTT assay. The cell morphology and the mitochondrial ultrasructure were observed with invert and transmission electron microscopy respectively. The apoptosis of the two cells was determined with DNA gel electrophoresis and Annexin V and PI double stained flow cytometry. Mitochondrial membrane potential (MMP) and the function of P-gp was assessed with rhodamine 123 (Rh123) staining intensity on confocal microscopy.The release of cytochrome C from mitochondria was observed with Western blot. Results The mitochondrial function of MCF-7/Adr was irreversible injury when they were exposured to 4mmol/L H2O2. The mitochondria wer