miR-186-5p通过下调MYCN轴增强小儿神经母细胞瘤NB1/DDP细胞顺铂敏感性的机制研究  

Mechanism of miR-186-5p enhanced cisplatin sensitivity of pediatric neuroblastoma NB1/DDP cells by MYCN

作　　者：王荣 张旭铭[2] 敬晓莉 成彩虹 WANG Rong;ZHANG Xuming;JING Xiaoli;CHENG Caihong(Academic Affairs Office,Dazhou Vocational College of Chinese Medicine,Dazhou 635000,China;Department of Paediatrics,Dazhou Integrated TCM&Western Medicine Hospital Pediatrics,Dazhou 635000,China;Department of Medical Imaging,Dazhou Vocational and Techinal College,Dazhou 635000,China;Department of Nursing,Dazhou Vocational College of Chinese Medicine,Dazhou 635000,China)

机构地区：[1]达州中医药职业学院教务处,四川达州635000 [2]达州市中西医结合医院儿科,四川达州635000 [3]达州职业技术学院影像检验系,四川达州635000 [4]达州中医药职业学院护理系,四川达州635000

出　　处：《沈阳药科大学学报》2022年第12期1478-1485,共8页Journal of Shenyang Pharmaceutical University

摘　　要：目的 探讨miR-186-5p对小儿神经母细胞瘤NB1/DDP细胞顺铂敏感性的影响。方法 构建小儿神经母细胞瘤细胞顺铂耐药株NB1/DDP后,实验分组为:NC组(未加入DDP的NB1顺铂敏感细胞)、Resistance组(加入0.5 mol·L^(-1)DDP处理的NB1/DDP耐药细胞)、miR-186-5p mimic组(在0.5 mol·L^(-1)DDP处理的NB1/DDP耐药细胞中转染miR-186-5p mimic)、miR-186-5p inhibitor组(在0.5 mol·L^(-1)DDP处理的NB1/DDP耐药细胞中转染miR-186-5p inhibitor)、pcDNA3.1-MYCN组(在0.5 mol·L^(-1)DDP处理的NB1/DDP耐药细胞中转染pcDNA3.1-MYCN)、sh-MYCN组(在0.5 mol·L^(-1)DDP处理的NB1/DDP耐药细胞中转染sh-MYCN)和miR-186-5p mimic+pcDNA3.1-MYCN组(在0.5 mol·L^(-1)DDP处理的NB1/DDP耐药细胞中转染miR-186-5p mimic和pcDNA3.1-MYCN);应用RT-qPCR检测miR-186-5p的表达水平;双荧光素酶报告基因验证miR-186-5p和MYCN的靶向关系;Western blot检测MYCN的表达水平;采用MTT试剂盒和Transwell法分别检测NB1/DDP细胞增殖活力和侵袭,流式细胞术检测细胞凋亡。结果 过表达miR-186-5p显著抑制NB1/DDP细胞增殖和侵袭(P<0.05)并增强NB1/DDP细胞对顺铂的敏感性,而抑制miR-186-5p则相反;miR-186-5p靶向负调控MYCN的表达水平;过表达MYCN逆转了miR-186-5p对NB1/DDP细胞增殖和侵袭的抑制作用(P<0.05)。结论 miR-186-5p通过下调MYCN的表达水平增强NB1/DDP细胞对顺铂的敏感性。Objective To investigate the effect of miR-186-5 p on cisplatin sensitivity of pediatric neuroblastoma NB1/DDP cells.Methods After successfully constructing the cisplatin-resistant strain NB1/DDP of pediatric neuroblastoma cells, the experimental groups were as follows: NC group(NB1 cisplatin-sensitive cells without DDP added),resistance group(Add 0.5 mol·L^(-1)DDP-treated NB1/DDP resistant cells),miR-186-5 p mimic group(Transfection of miR-186-5 p mimic in NB1/DDP resistant cells treated with 0.5 mol·L^(-1)DDP),miR-186-5 p inhibitor group(Transfection of miR-186-5 p inhibitor in NB1/DDP resistant cells treated with 0.5 mol·L^(-1)DDP),pcDNA3.1-MYCN group(Transfection of pcDNA3.1-MYCN in NB1/DDP resistant cells treated with 0.5 mol·L^(-1)DDP),sh-MYCN group(Transfection of sh-MYCN in NB1/DDP resistant cells treated with 0.5 mol·L^(-1)DDP),miR-186-5 p mimic+pcDNA3.1-MYCN group(Transfection of miR-186-5 p mimic and pcDNA3.1-MYCN in NB1/DDP resistant cells treated with 0.5 mol·L^(-1)DDP).The expression of miR-186-5 p was detected by RT-qPCR.The targeted relationship between miR-186-5 p and MYCN was verified by dual luciferase reporter gene.The expression of MYCN was detected by Western blot.The proliferation and invasion of NB1/DDP cells were detected by MTT and Transwell, respectively.Flow cytometry was used to detect NB1/DDP cells apoptosis.Results Overexpression of miR-186-5 p significantly inhibited proliferation and invasion of NB1/DDP cells(P<0.05)and sensitized NB1/DDP cells to cisplatin, whereas inhibition of miR-186-5 p did the opposite.miR-186-5 p targets and negatively regulates the expression level of MYCN(P<0.01).Overexpression of MYCN reversed the inhibitory effect of miR-186-5 p on proliferation and invasion of NB1/DDP cells(P<0.05).Conclusion miR-186-5 p inhibited the proliferation and invasion of NB1/DDP cells by downregulating the expression of MYCN,thereby enhancing the cisplatin sensitivity of NB1/DDP cells.

关 键 词：miR-186-5p MYCN 小儿神经母细胞瘤 顺铂 NB1/DDP 

分 类 号：R96[医药卫生—药理学] R739.41[医药卫生—药学]