三阴乳腺癌患者血液外泌体miRNA-27a/P-gp在新辅助化疗后的变化及其对血管新生的影响  

Changes of miRNA-27a/ P-gp in blood exosomes of TNBC patients after neoadjuvant chemotherapy and its effects on angiogenesis

作　　者：李会欣 刘伟[1] 邵建强 王遵义[1] LI Huixin;LIU Wei;SHAO Jianqiang;WANG Zunyi(The Third Department of Thyroid and Breast Surgery,Cangzhou Central Hospital,Cangzhou,Hebei 061001,China)

机构地区：[1]沧州市中心医院甲状腺乳腺外三科,河北沧州061001

出　　处：《徐州医科大学学报》2021年第5期357-362,共6页Journal of Xuzhou Medical University

基　　金：河北省医学科学项目研究课题(20200343)。

摘　　要：目的研究新辅助化疗(NACT)前后三阴乳腺癌(TNBC)患者血液外泌体中miR-27a/P-gp对血管新生的调节机制。方法分离并鉴定NACT组与对照组(单独表柔比星治疗)TNBC患者血液外泌体。RT-qPCR法检测血液外泌体中miR-27a/P-gp的表达量。荧光素酶报告基因实验研究miR-27a/P-gp的关系。低表达miR-27a的外泌体(EXO-miR-27a-)、高表达miR-27a的外泌体(EXO-miR-27a+)、miR-27a拟似物(miR-27a-mimic)和阴性对照(mimic-NC)、P-gp过表达质粒(pcDNA3.1-P-gp)及相应的对照物(pcDNA3.1-null)分别处理人微血管内皮细胞(HMVEC)。ELISA法检测血管内皮生长因子(VEGF)水平,蛋白免疫印迹法检测VEGF-A的表达,小管形成实验评估血管生成能力。结果与对照组比较,NACT组的血液和外泌体中miR-27a的表达下调,P-gp表达上调(P<0.05);与治疗前比较,NACT组治疗后的血液和外泌体中miR-27a表达下调,而P-gp的表达上调(P<0.05);HMVEC细胞中,EXO-miR-27a+组的VEGF水平和血管生成能力较EXO-miR-27a-组增加(P<0.05)。miR-27a-mimic组的VEGF水平和血管生成能力较mimic-NC组增加(P<0.05)。miR-27a靶向抑制P-gp。pcDNA3.1-P-gp组VEGF水平和血管生成能力较pcDNA3.1-null组降低(P<0.05)。pcDNA3.1-P-gp+miR-27a-mimic组VEGF水平和血管生成能力较pcDNA3.1-null+miR-27a-mimic组降低(P<0.05)。结论NACT治疗TNBC患者后外泌体miR-27a下调,而P-gp上调,miR-27a通过抑制P-gp促进血管新生。Objective To investigate the regulatory mechanism of miR-27a/P-gp in blood exosomes of patients with triple-negative breast cancer(TNBC)before and after neoadjuvant chemotherapy(NACT).Methods Blood exosomes from TNBC patients in the NACT treatment group and the control group(epirubicin treatment alone)were isolated and identified.The expression of miR-27a/P-gp in blood exosomes was detected by RT-qPCR.The relationship between miR-27a and P-gp was investigated by luciferase reporter gene assay.Human microvessels endothelial cells(HMVECs)were treated with exosomes with low expression of miR-27a(the Exo-miR-27a-group),exosomes with high expression of miR-27a(the Exo-miR-27a+group),miR-27a mimics(the miR-27a-mimic group),negative control(the mimic-NC group)and plasmid with overexpression of P-gp(the pcDNA3.1-P-gp group).The level of VEGF was detected by ELSIA.The expression of VEGF-A was detected by Western blot.The angiogenesis was evaluated by tubule formation assay.Results Compared with the control group,the expression of miR-27a in blood and exosomes in the NACT group was down-regulated,while the expression of P-gp was up-regulated(P<0.05).Compared with those before NACT treatment,the expression of miR-27a in blood and exosomes was down-regulated after NACT treatment,while the expression of P-gp was up-regulated(P<0.05).In HMVECs,the level of VEGF and angiogenesis increased in the Exo-miR-27a+group,compared with those in the Exo-miR-27a-group(P<0.05).The level of VEGF and angiogenesis in the miR-27a-mimic group were higher than those in the mimic-NC group(P<0.05).MiR-27a targeted P-gp inhibition.The level of VEGF and angiogenesis in the pcDNA3.1-P-gp group were lower than those in the pcDNA3.1-null group(P<0.05).The level of VEGF and angiogenesis were lower in the pcDNA3.1-P-gp+miR-27a-mimic group than those in the pcDNA3.1-null+miR-27a-mimic group(P<0.05).Conclusions Exosome miR-27a is down-regulated and P-gp up-regulated in TNBC patients after NACT treatment,and miR-27a promotes angiogenesis through inhibiting

关 键 词：微小RNA-27a P-GP 新辅助化疗 三阴乳腺癌 人微血管内皮细胞 血管生成 

分 类 号：R737.9[医药卫生—肿瘤]