LncRNA-MALAT1通过调控miR-106b-5p介导结直肠癌细胞对5-氟尿嘧啶耐药机制研究  

LncRNA-MALAT1 mediated colorectal cancer cells to 5-fluorouracil resistance by targeting miR-106b-5p

作　　者：张朴花 徐志广[3] 阳美玲 阳婵娟 ZHANG Pu-hua;XU Zhi-guang;YANG Mei-ling;YANG Chan-juan(AffHated Nanhua Hospital,University of South China,Hengyang 421002,China;Basic Experimental Center,Hengyang Medical College,University of South China,Hengyang 421002,China)

机构地区：[1]南华大学附属南华医院肿瘤内科,湖南衡阳421002 [2]南华大学衡阳医学院基础实验中心,湖南衡阳421002 [3]南华大学附属南华医院肝胆外科,湖南衡阳421002

出　　处：《中华肿瘤防治杂志》2021年第12期914-920,共7页Chinese Journal of Cancer Prevention and Treatment

摘　　要：目的探讨LncRNA-MALAT1(MALAT1)通过调控miR-106b-5p介导结直肠癌细胞5-氟尿嘧啶(5-FU)耐药分子机制。方法收集2016-03-17-2019-04-10就诊于南华大学附属南华医院资料完整的30例5-FU化疗敏感和30例5-FU化疗耐药的结直肠癌患者癌组织。采用5-FU诱导结直肠癌细胞HCT116产生耐药性细胞HCT116/5-FU,将HCT116/5-FU细胞分为空白对照组(Control)、siRNA阴性对照组(siRNA-NC)、MALAT1 siRNA干扰组(si-MALAT1)、si-MALAT1+inhibitor-NC组和si-MALAT1+miR-106b-5p inhibitor组。采用不同浓度(0、5、10、20 mg/L)5-FU处理细胞48 h,四甲基偶氮唑盐法检测细胞增殖活性;实时荧光定量PCR法检测结直肠癌组织或细胞中MALAT1和miR-106b-5p表达水平;双荧光素酶报告基因实验检测MALAT1、miR-106b-5p和缺氧诱导因子-1α(HIF-1α)的靶向关系。采用SPSS 21.0进行数据分析,多组间比较采用单因素方差分析或两因素方差分析,两两多重比较采用LSD-t检验。结果 (1)临床标本测定:5-FU耐药结直肠癌组织中MALAT1表达量为7.57±1.76,高于5-FU敏感结直肠癌组织(3.15±1.13),t=11.541,P<0.001;而miRNA-106b-5p表达量为8.59±2.32,低于5-FU敏感结直肠癌组织(11.57±2.97),t=4.329,P<0.001。(2)体外细胞实验:0、5、10、20 mg/L 5-FU处理48 h后,HCT116/5-FU细胞在450 nm波长处光密度值D(450 nm)高于HCT116细胞,差异有统计学意义,F组别=93.335,F浓度=48.983,F组别×浓度=11.848,均P<0.001。HCT116/5-FU细胞中MALAT1表达量高于HCT116细胞(t=12.883,P<0.001),miRNA-106b-5p表达量低于HCT116细胞,t=8.101,P<0.001;si-MALAT1组HCT116/5-FU细胞中MALAT1表达量为0.04±0.01,低于Control组(1.01±0.07)和si-NC组(0.99±0.02),F=397.841,P<0.001;0、5、10、20 mg/L 5-FU处理48 h后,si-MALAT1组HCT116/5-FU细胞增殖率低于Control组和si-NC组,差异有统计学意义,F组别=263.160,F浓度=180.634,F组别×浓度=33.675,均P<0.001。si-MALAT1组HCT116/5-FU细胞中miRNA-106b-5p表达量为8.33±0.76,高于Control组(1.10±0.30)和si-NC组Objective To explore the molecular mechanism of LncRNA-MALAT1(MALAT1) mediating 5-fluorouracil(5-FU) resistance in colorectal cancer cells by regulating miR-106 b-5 p.Methods 5-FU-sensitive and 5-FU-resistant colorectal cancer tissues(30 samples for each) were collected in Affliated Nanhua Hospital,University of South China from March 17,2016 to April 10,2019.5-FU was employed to construct resistance cells HCT116/5-FU,and HCT116/5-FU cells were divided into blank group(Control),siRNA negative control group(siRNA-NC),MALAT1 siRNA interference group(siRNA-MALAT1),si-MALAT1+inhibitor-NC group and si-MALAT1+miR-106 b-5 p inhibitor group.Cells were treated with different concentrations(0,5,10,20 mg/l)of 5-FU for 48 h,and the proliferation activity of cells were detected by MTT assay.The expression levels of MALAT1 and miR-106 b-5 p in colorectal cancer tissues or cells were detected by qRT-PCR.The targeting relationships of MALAT1,miR-106 b-5 p and hypoxia inducible factor-1α(HIF-1α)were detected by using the dual luciferase reporter gene system.SPSS 21.0 statistical software was applied for statistical analysis of data,one-way ANOVA or two-factor ANOVA were used for comparing the among multiple groups and LSD-t test was used for pairwise multiple comparisons of the between groups.Results The expression levels of MALAT1 in 5-FU-resistant colorectal cancer tissues(7.57±1.76)were higher than that in 5-FU-sensitive colorectal cancer tissues(3.15±1.13,t=11.541,P<0.001),while the expression levels of miRNA-106 b-5 p(8.59±2.32)were lower than that in 5-FU-sensitive colorectal cancer tissues(8.59±2.32,t=4.329,P<0.001).After 5-FU treatment at different concentrations(0,5,10,20 mg/l)for 48 h,the optical density value D(450 nm)of HCT116/5-FU cells was higher than that of HCT116 cells,and the difference was statistically significant(Fgroup=93.335,Fconcentration=48.983,Fgroup×concentration=11.848,all P<0.001).The expression levels of MALAT1 in HCT116/5-FU cells was higher than that in HCT116 cells(t=12.883,P<0.001),and the

关 键 词：结直肠癌 耐药性 HCT116/5-FU细胞 LncRNA-MALAT1 miR-106b-5p 5-氟尿嘧啶 

分 类 号：R735.3[医药卫生—肿瘤]