放射线诱导BRCA1出核效应对PARP抑制剂的增敏作用

发布时间：2018-10-23 12:58

【摘要】：目的探讨放射线诱导的乳腺癌1号基因(BRCA1)出核效应对同源重组介导的DNA双链断裂损伤(DSB)修复功能的影响及对聚腺苷二磷酸核糖聚合酶(PARP)抑制剂的增敏作用。方法采用亚细胞纯化及蛋白印迹法检测乳腺癌细胞株MCF7经放射线处理后BRCA1的亚细胞定位,同时采用免疫荧光法检测细胞核磷酸化的H2AX组蛋白(γ-H2AX)、Rad51核焦点形成。流式细胞技术检测细胞凋亡、克隆形成实验检测体外细胞存活情况,并制作裸鼠皮下移植瘤模型,分为4组,分别为:对照组、放射+DMSO组、假照+ABT-888(ABT-888为PARP抑制剂)组、放射+ABT-888组,每5 d为1个治疗周期,周期第1天给予2 Gy照射或假照射。第2~5天给予溶剂(对照组、放射+DMSO组)或20 mg/(kg·d)的ABT-888(假照+ABT-888组),总共给予4个周期的治疗。检测各组的体内抑瘤作用。结果经4 Gy放射处理后,MCF7胞核BRCA1表达量减少,仅为对照组的41%,但胞浆内表达量明显增多,是对照组的2.14倍(P0.01);同时MCF7细胞经4 Gy放射处理后ABT-888诱导的Rad51核焦点形成阳性细胞也较对照组明显减少(7%vs.30%,P=0.01)。放射线和ABT-888联合应用导致MCF7细胞凋亡率增高(19%),与对照组(5%)、放射+DMSO组(9%)或假照+ABT-888组(6.2%)相比较差异有统计学意义(P0.01)。裸鼠移植瘤模型中放射+ABT-888组对肿瘤生长抑制作用明显优于假照+DMSO组或假照+ABT-888组(P0.01)。结论放射线可诱导BRCA1出核并增强PARP抑制剂的抗肿瘤作用。
[Abstract]:Objective to investigate the effect of radiation-induced nucleation of breast cancer gene 1 (BRCA1) on the repair function of (DSB) induced by homologous recombination of DNA double strand break and to enhance the sensitivity of polyadenosine diphosphate ribose polymerase (PARP) inhibitor. Methods the subcellular localization of BRCA1 in breast cancer cell line MCF7 was detected by subcellular purification and Western blotting. Meanwhile, nuclear phosphorylation of H2AX histone (纬-H2AX) and nuclear focus formation of Rad51 were detected by immunofluorescence. Flow cytometry was used to detect cell apoptosis and clone formation assay was used to detect cell survival in vitro. The nude mice were divided into 4 groups: control group, radiation DMSO group, pseudoradiographic ABT-888 (ABT-888 as PARP inhibitor) group. ABT-888 group was given 2 Gy irradiation or false irradiation every 5 days. A total of 4 cycles of treatment were given on day 2 to day 5: solvent (control group, radiation DMSO group) or 20 mg/ (kg d) ABT-888 (pseudoradiographic ABT-888 group). The inhibitory effect in vivo of each group was detected. Results after 4 Gy irradiation, the expression of BRCA1 in the nucleus of MCF7 was decreased to 41% of that in the control group, but the expression of BRCA1 in the cytoplasm was significantly increased. At the same time, the number of Rad51 focus forming positive cells induced by ABT-888 in MCF7 cells after 4 Gy irradiation was significantly lower than that in the control group (7vs.30). The apoptosis rate of MCF7 cells was increased by radiation combined with ABT-888 (19%), which was significantly higher than that of control group (5%), radiation DMSO group (9%) or pseudoradiographic ABT-888 group (6.2%) (P0.01). In nude mice, the inhibitory effect of radiation ABT-888 on tumor growth was significantly better than that of DMSO or ABT-888 (P0.01). Conclusion radiation can induce the nucleation of BRCA1 and enhance the anti-tumor effect of PARP inhibitor.
【作者单位】：广州医科大学附属第一医院呼吸疾病国家重点实验室广州呼吸疾病研究所;广州医科大学附属第一医院病理科;
【基金】：国家自然科学基金资助项目(编号:81272901) 广州市医药卫生科技项目(编号:20151A011067)
【分类号】：R737.9

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