长链非编码RNA转移相关肺腺癌转录本1及微小RNA-874对气道平滑肌细胞功能改善的调控  

Regulation of airway smooth muscle cells function by long non-coding RNA metastasis associated lung adenocarcinoma transcript 1 and microRNA-874

作　　者：陈璐[1] 陈艳萍[1] 段效军[1] 张瑾 李林瑞[1] 郭宽鹏[1] Chen Lu;Chen Yanping;Duan Xiaojun;Zhang Jin;Li Linrui;Guo Kuanpeng(Department of Respiratory Medicine,Hunan Children's Hospital,Changsha 410000,China;不详)

机构地区：[1]湖南省儿童医院呼吸内科,长沙410000

出　　处：《临床内科杂志》2023年第10期700-704,共5页Journal of Clinical Internal Medicine

基　　金：湖南省卫生健康委员会科研计划项目(202206015321)。

摘　　要：目的探讨长链非编码RNA转移相关肺腺癌转录本1(lncRNA MALAT1)与微小RNA-874(miR-874)在血小板衍生因子(PDGF)-BB处理的气道平滑肌细胞(ASMCs)中的表达与功能。方法采用PDGF-BB处理ASMCs建立哮喘细胞模型。将ASMCs分为正常培养的对照组(Control组)、PDGF-BB处理的PDGF-BB组[再根据处理时间分为PDGF-BB(6 h)组、PDGF-BB(12 h)组、PDGF-BB(28 h)组、PDGF-BB(24 h)组)]、PDGF-BB处理的转染sh-MALAT组(PDGF-BB+sh-MALAT组)、PDGF-BB处理的转染sh-NC组(PDGF-BB+sh-NC组)、PDGF-BB处理的转染miR-874 mimics组(PDGF-BB+miR-874 mimics组)、PDGF-BB处理的转染miR-NC组(PDGF-BB+miR-NC组)、PDGF-BB处理的共转染sh-NC与miR-NC组(PDGF-BB+sh-NC+miR-NC组)、PDGF-BB处理的共转染sh-MALAT1与miR-NC组(PDGF-BB+sh-MALAT1+miR-NC组)、PDGF-BB处理的共转染sh-MALAT1与miR-874 inhibitor组(PDGF-BB+sh-MALAT1+miR-874 inhibitor组)。将MALAT-WT或MALAT-MUT序列和miR-874 mimics或miR-NC共同转染入psi-CHECK2荧光素酶报告载体中,并将共转染的细胞分为miR-874 mimics+MALAT-WT组、miR-NC+MALAT-WT组、miR-874 mimics+MALAT-MUT组和miR-NC+MALAT-MUT组。采用RT-PCR检测MALAT1、miR-874表达水平,采用CCK-8法检测吸光度值代表细胞增殖能力,采用Transwell实验计算迁移细胞数代表细胞迁移能力,采用ELISA试验检测IL-1β、IL-6和肿瘤坏死因子(TNF)-α水平。使用在线数据库starbase预测MALAT和miR-874之间的潜在结合位点,采用双荧光素酶报告系统检测各组ASMCs中荧光素酶活性。结果Control组、PDGF-BB(6 h)组、PDGF-BB(12 h)组、PDGF-BB(18 h)组及PDGF-BB(24 h)组细胞MALAT1相对水平依次升高,miR-874相对水平依次降低(P<0.05)。与Control组比较,PDGF-BB组、PDGF-BB+sh-NC组及PDGF-BB+sh-MALAT1组、PDGF-BB+miR-NC组及PDGF-BB+miR-874 mimics组细胞增殖能力、迁移能力及细胞上清液中IL-1β、IL-6和肿瘤坏死因子(TNF)-α水平均明显增强;与PDGF-BB组比较,PDGF-BB+sh-MALAT1组及PDGF-BB+miR-874 mimics组细胞上述能力与Objective To investigate the expression and function of long non-coding RNA metastasis associated lung adenocarcinoma transcript 1(lncRNA MALAT1)and microRNA-874(miR-874)in platelet-derived factor(PDGF)-BB treated airway smooth muscle cells(ASMCs).Methods The asthma cell model was established by treating ASMCs with PDGF-BB.ASMCs were divided into normal culture control group and PDGF-BB treated group[which were divided into PDGF-BB(6 h)group,PDGF-BB(12 h)group,PDGF-BB(28 h)group and PDGF-BB(24 h)group according to the treatment time],transfected sh-MALAT group with PDGF-BB treatment(PDGF-BB+sh-MALAT group),sh-NC group with PDGF-BB treatment(PDGF-BB+sh-NC group),miR-874 mimics group with PDGF-BB treatment(PDGF-BB+miR-874 mimics group),PDGF-BB transfected miR-NC group(PDGF-BB+miR-NC group),the combined transfection of sh-NC and miR-NC group with PDGF-BB treatment(PDGF-BB+sh-NC+miR-NC group),the combined transfection of sh-MALAT1 and miR-NC group with PDGF-BB treatment(PDGF-BB+sh-MALAT1+miR-NC group)the combined tronsfection of sh-MALAT1 and miR-874 inhibitor group with PDGF-BB treatment(PDGF-BB+sh-MALAT1+miR-874 inhibitor group).MALAT-WT or MALAT-MUT sequence and miR-874 mimics or miR-NC were co-transfected into the psi-CHECK2 luciferase reporter vector.The co-transfected cells were divided into miR-874 mimics+MALAT-WT group,miR-NC+MALAT-WT group,miR-874 mimics+MALAT-MUT group and miR-NC+MALAT-MUT group.The expression levels of MALAT1 and miR-874 were detected by RT-PCR.The absorbance value was detected by CCK-8 method,which represented the cell proliferation ability.The number of migrating cells was calculated by transwell experiment to represent the migration ability of cells.The levels of IL-1β,IL-6 and TNF-αwere detected by ELISA.The online database starbase was used to predict potential binding sites between MALAT and miR-874.Double luciferase reporting system was used to detect luciferase activity in ASMCs of each group.Results The relative level of MALAT1 in control group,PDGF-BB(6 h)group,PDGF-BB(12 h)gro

关 键 词：长链非编码RNA转移相关肺腺癌转录本1 微小RNA-874 气道平滑肌细胞 细胞增殖 细胞迁移 

分 类 号：R392.12[医药卫生—免疫学]