摘要
目的探讨长链非编码核糖核酸淋巴细胞白血病缺失基因2(LncRNA DLEU2)通过靶向微小核糖核酸(miR)-30a-5p/整合素β3(CD61)对人结肠癌SW1116细胞生长和增殖的影响。方法取人结肠癌细胞株SW1116培养并随机分为si-NC组、si-LncRNA DLEU2组、模拟物阴性对照(miR-NC)组、miR-30a-5p mimics组、miR-30a-5p抑制剂阴性对照(NC inhibitor)+si-LncRNA DLEU2组及miR-30a-5p inhibitor+si-LncRNA DLEU2组,同时设置未转染细胞为空白组;转染48 h后,采用荧光显微镜检测细胞转染效率,四甲基噻唑蓝(MTT)法检测细胞增殖活性,实时定量聚合酶链反应(RT-qPCR)和蛋白质免疫印迹法(Western blot)分别检测细胞LncRNA DLEU2和miR-30a-5p表达、CD61、细胞周期蛋白D1(CyclinD1)、c-Myc mRNA和蛋白表达,双荧光素酶报告基因实验和RT-qPCR检测LncRNA DLEU2与miR-30a-5p的靶向关系、miR-30a-5p与CD61的靶向关系;制作雄性胸腺BALB/c裸鼠异种移植瘤模型评估LncRNA DLEU2对肿瘤生长的影响。结果si-NC组、si-LncRNA DLEU2组、miR-NC组、miR-30a-5p mimics组、NC inhibitor+si-LncRNA DLEU2组、miR-30a-5p inhibitor+si-LncRNA DLEU2组转染效率均>85%;与空白组及si-NC组比较,si-LncRNA DLEU2组、NC inhibitor+si-LncRNA DLEU2组增殖抑制率上升(P<0.05),DLEU2表达下降(P<0.05),miR-30a-5p表达增加(P<0.05),CD61、CyclinD1、c-Myc mRNA和蛋白表达减少(P<0.05);与NC inhibitor+si-LncRNA DLEU2组比较,miR-30a-5p inhibitor+si-LncRNA DLEU2组细胞增殖抑制率下降(P<0.05),miR-30a-5p表达下降,CD61、CyclinD1、c-Myc mRNA和蛋白表达上升(P<0.05);与miR-NC组比较,miR-30a-5p mimics组miR-30a-5p表达、增殖抑制率增加(P<0.05),CD61、CyclinD1、c-Myc mRNA和蛋白表达下降(P<0.05);LncRNA DLEU2可直接靶向调控miR-30a-5p、miR-30a-5p可直接靶向调控CD61,抑制LncRNA DLEU2表达可降低BALB/c裸鼠移植瘤生长(P<0.05)。结论沉默LncRNA DLEU2可抑制人结肠癌细胞株SW1116生长和增殖,其机制可能与靶向miR-30a-5p抑制CD61表达,抑制
Objective To investigate the effect of long non-coding ribonucleic deleted in llymphocytic leukemia 2(LncRNA DLEU2)on the growth and proliferation of human colon cancer SW1116 cell by targeting micrornas(miR)-30a-5p/integrinβ3(CD61).Methods Human colon cancer cell strain SW1116 was cultured and randomly divided into:si-NC group,si-LncRNA DLEU2 group,miR-NC negative control group,miR-30a-5p mimics group,miR-30a-5p NC inhibitor+si-LncRNA DLEU2 group,miR-30a-5p Inhibitor+si-LncRNA DLEU2 group;meanwhile,un-transfected cells were set as blank group.After transfection 48h,the transfection efficiency of each group was detected by fluorescence microscope,and MTT assay was used to detect the proliferation activity of cells in each group;and the LncRNA DLEU2 and miR-30a-5p expressions,the mRNA and protein expressions of CD61,CyclinD1,c-Myc in each group were detected by real-time quantitative polymerase chain reaction(RT-qPCR)and western blotting;dual luciferase reporter assay and RT-qPCR were used to detect the targeting relationship between LncRNA DLEU2 and miR-30a-5p,miR-30a-5p,and CD61.The effect of LncRNA DLEU2 on tumor growth in vivo was evaluated in xenograft tumor model of male thymus BALB/C.Results The transfection efficiency of si-NC group,si-LncRNA DLEU2 group,miR-NC group,miR-30a-5p mimics group,NC inhibitor+si-LncRNA DLEU2 group,and miR-30a-5p inhibitor+si-LncRNA DLEU2 group was more than 85%.Compared with blank group,si-NC group,the proliferation inhibition rates of si-LncRNA DLEU2 group,NC inhibitor+si-LncRNA DLEU2 group increased(P<0.05),the expression of DLEU2 decreased(P<0.05),and the expression of miR-30a-5p increased(P<0.05);the mRNA and protein expressions of CD61,CyclinD1,c-Myc decreased(P<0.05).Compared with NC inhibitor+si-LncRNA DLEU2 group,the proliferation inhibition rate of miR-30a-5p inhibitor+si-LncRNA DLEU2 group decreased(P<0.05),and the expression of miR-30a-5p decreased,while the expressions of CD61,CyclinD1,c-Myc mRNA and protein increased(P<0.05).Compared with miR-NC group,the expressio
作者
吕俊
黄可
张恩霖
吴淼
LYU Jun;HUANG Ke;ZHANG Enlin;WU Miao(Department of Gastrointestinal Surgery,Yibin No.2 People's Hospital,Yibing 644000,Sichuan,China;Department of Laboratory Science,Yibin No.2 People's Hospital,Yibing 644000,Sichuan,China)
出处
《贵州医科大学学报》
CAS
2024年第6期836-845,共10页
Journal of Guizhou Medical University
基金
四川省卫生健康委员会科研课题(19PJ301)。
