miR-489协同卡铂通过抑制XIAP的表达发挥抗乳腺癌作用  

MiR-489 Targets X-linked Inhibitor of Apoptosis Protein to Enhance the Anti-tumor Effect of Carboplatin on Breast Cancer

作　　者：郑璐[1] 胡静[1] 陈雪 许权[1] ZHENG Lu, HU Jing, CHEN Xue, XU Quan(Department of Radiotherapy, Li Huili Hospital of Ningbo Medical Center, Ningbo 315000, Chin)

机构地区：[1]宁波市医疗中心李惠利医院放疗科,浙江宁波315000

出　　处：《中国现代应用药学》2018年第3期319-324,共6页Chinese Journal of Modern Applied Pharmacy

摘　　要：目的研究miR-489对卡铂的协同抗乳腺癌效应及机制。方法 MTT细胞活力试验检测卡铂和miR-489对T-47D细胞的杀伤活性。生物信息学、Western blot试验及荧光素酶报告基因试验验证X连锁凋亡抑制蛋白(X-linked inhibitor of apoptosis protein,XIAP)是否为miR-489的靶点。分离去除T-47D细胞中的线粒体,Western blot试验检测miR-489、卡铂及XIAP质粒处理后T-47D细胞Smac/DIABLO的释放和caspase-9及caspase-3的活化。免疫共沉淀试验检测XIAP与Smac/DIABLO的相互作用。流式细胞术检测T-47D细胞的凋亡情况。结果 miR-489处理能显著增强卡铂对T-47D细胞的杀伤活性。XIAP是miR-489的靶点。miR-489不影响卡铂依赖的线粒体中Smac/DIABLO的释放,但能通过下调XIAP的表达减弱XIAP与Smac/DIABLO的相互作用。另外,转染XIAP质粒能显著抑制miR-489对卡铂的协同抗乳腺癌效应,XIAP质粒能显著抑制miR-489联合卡铂对T-47D细胞凋亡的诱导。XIAP质粒能显著抑制miR-489联合卡铂对T-47D细胞caspase-9及caspase-3的活化。结论 miR-489抑制XIAP的表达发挥对卡铂的协同抗乳腺癌作用。OBJECTIVE To investigate the effect and mechanism of miR-489 on enhancing the anti-tumor effect of carboplatin on breast cancer. METHODS Cell viability of T-47 D which were treated with miR-489 and carboplatin was measured by using MTT assay. Bioinformatics, Western blot analysis and luciferase reporter assay were performed to confirm whether the XIAP was the target of miR-489. After removal of mitochondria, Western blot analysis was conducted to detect the release of Smac/DIABLO and activation of caspase-9 and caspase-3 in T-47 D cells treated with carboplatin and miR-489. Co-immunoprecipitation assay was performed to evaluate the interaction with XIAP and Smac/DIABLO. Flow cytometry analysis was used to measure the cell apoptosis of T-47 D. RESULTS Results of MTT assays showed that miR-489 significantly enhanced the cytotoxicity of carboplatin to T-47 D. Results of bioinformatics, Western blot analysis and luciferase reporter assay showed that XIAP was the target of miR-489 in T-47 D. Overexpression of miR-489 couldn＇t influence the carboplatin-induced release of Smac/DIABLO. However, miR-489 was able to inhibit the interaction of Smac/DIABLO and XIAP through down-regulating the expression of XIAP in T-47 D. In addition, transfection with XIAP plasmid significantly abolished the synergistic effect of miR-489 on carboplatin-induced cytotoxicity to breast cancer. Results of flow cytometry showed that transfection with XIAP plasmid significantly abolished the miR-489-promoed apoptosis induced by carboplatin. Results of Western blot analysis showed that transfection with XIAP plasmid significantly abolished the miR-489-promoed activation of caspase-9 and caspase-3 induced by carboplatin. CONCLUSION Mi R-489 targets XIAP to enhance the anti-tumor effect of carboplatin on breast cancer.

关 键 词：乳腺癌 miR-489 X连锁凋亡抑制蛋白 卡铂 凋亡 

分 类 号：R965.2[医药卫生—药理学]