异紫堇碱调控LncRNA HCG18/miR-380-5p通路抑制乳腺癌生长及转移  

Inhibitory effects of isocorydine on breast cancer growth and metastasis by regulating the LncRNA HCG18/miR-380-5p pathway

作　　者：丁萌[1] 曲杰[2] 陈向华[1] 张秀琴[1] 张青云[3] DING Meng;QU Jie;CHEN Xianghua;ZHANG Xiuqin;ZHANG Qingyun(Laboratory,Affiliated Hospital of Chengde Medical College,Chengde 067000,China;Oncology Department,Affiliated Hospital of Chengde Medical College,Chengde 067000,China;Vascular Surgery,Affiliated Hospital of Chengde Medical College,Chengde 067000,China)

机构地区：[1]承德医学院附属医院检验科,河北承德067000 [2]承德医学院附属医院肿瘤内科,河北承德067000 [3]承德医学院附属医院血管外科,河北承德067000

出　　处：《沈阳药科大学学报》2022年第12期1486-1493,1520,共9页Journal of Shenyang Pharmaceutical University

基　　金：2020河北省中医药管理局科研计划项目(2020249)。

摘　　要：目的探讨异紫堇碱(ICDE)对乳腺癌生长及转移的影响及其对LncRNA HCG18/miR-380-5p通路的调控作用。方法采用不同剂量的异紫堇碱处理人乳腺癌细胞MDA-MB-231(ICDE 10.8μmol·L^(-1)组、ICDE 32.5μmol·L^(-1)组、ICDE 97.4μmol·L^(-1)组),正常培养的MDA-MB-231细胞记为Con组。MDA-MB-231细胞分别转染si-NC、si-HCG18(si-NC组、si-HCG18组),pcDNA、pcDNA-HCG18分别转染入MDA-MB-231细胞后加入异紫堇碱处理细胞(ICDE 97.4μmol·L^(-1)+pcDNA组、ICDE 97.4μmol·L^(-1)+pcDNA-HCG18组);采用MTT实验、平板克隆形成实验、Transwell小室实验分别检测细胞增殖、克隆形成、迁移及侵袭;采用qRT-PCR法检测乳腺癌组织与癌旁组织以及各组MDA-MB-231细胞中HCG18、miR-380-5p的表达量;双荧光素酶报告基因实验检测HCG18与miR-380-5p的靶向调控关系;采用Western blot法检测MMP-2、MMP-9蛋白表达量。结果与Con组比较,ICDE 10.8μmol·L^(-1)组、ICDE 32.5μmol·L^(-1)组、ICDE 97.4μmol·L^(-1)组细胞增殖抑制率降低,克隆形成数、迁移及侵袭细胞数减少,MMP-2、MMP-9蛋白水平降低;与癌旁组织比较,乳腺癌组织中HCG18的表达水平升高,miR-380-5p的表达水平降低,而ICDE可抑制HCG18的表达及促进miR-380-5p的表达;HCG18可靶向结合miR-380-5p;与si-NC组比较,si-HCG18组细胞增殖抑制率降低(P<0.05),克隆形成数、迁移及侵袭细胞数减少,MMP-2、MMP-9蛋白水平降低;与ICDE 97.4μmol·L^(-1)+pcDNA组比较,ICDE 97.4μmol·L^(-1)+pcDNA-HCG18组miR-380-5p的表达水平降低,细胞增殖抑制率升高,克隆形成数、迁移及侵袭细胞数均明显增多,MMP-2、MMP-9蛋白水平升高。结论异紫堇碱可通过调控HCG18/miR-380-5p通路从而抑制乳腺癌细胞增殖、克隆形成、迁移及侵袭。Objective To explore the effect of isocorydine(ICDE)on the growth and metastasis of breast cancer and its regulation of LncRNA HCG18/miR-380-5 p pathway.Methods Different doses of isocorydine were used to treat human breast cancer cells MDA-MB-231(ICDE 10.8μmol·L^(-1)group,ICDE 32.5μmol·L^(-1)group,ICDE 97.4μmol·L^(-1)group),and normal cultured MDA-MB-231 cells were recorded as Con group.MDA-MB-231 cells were transfected with si-NC and si-HCG18(si-NC group,si-HCG18 group).pcDNA and pcDNA-HCG18 were respectively transfected into MDA-MB-231 cells and then added to isocorydine-treated cells(ICDE 97.4μmol·L^(-1)+pcDNA group,ICDE 97.4μmol·L^(-1)+pcDNA-HCG18 group).MTT experiment,plate clone formation experiment,and Transwell chamber experiment were used todetect cell proliferation,clone formation,migration and invasion,respectively.The qRT-PCR method was used to detect the expression of HCG18 and miR-380-5 p in breast cancer tissues and adjacent tissues and MDA-MB-231 cells in each group.The dual luciferase reporter gene experiment was used to detect the targeted regulation relationship between HCG18 and miR-380-5 p.Western blot was used to detect the protein expression of MMP-2 and MMP-9.Results Compared with the Con group,the cell viability of the ICDE 10.8μmol·L^(-1),ICDE 32.5μmol·L^(-1),and ICDE 97.4μmol·L^(-1)groups was decreased,the number of clones,migration and invasion cells was decreased,the protein levels of MMP-2,MMP-9 were reduced,and was dose-dependent.Compared with adjacent tissues,the expression level of HCG18 in breast cancer tissues was increased,and the expression level of miR-380-5 p was decreased.ICDE could significantly inhibit the expression of HCG18 and promote the expression of miR-380-5 p.HCG18 could target miR-380-5 p.Compared with the si-NC group,the cell viability of the si-HCG18 group was reduced,the number of clone formation,migration and invasion of cells was decreased,and the protein levels of MMP-2 and MMP-9 were decreased.Compared with the ICDE 97.4μmol·L^(-1)+pcDNA

关 键 词：异紫堇碱 LncRNA HCG18 miR-380-5p 乳腺癌 增殖 迁移 侵袭 

分 类 号：R96[医药卫生—药理学]