摘要
目的探索长链非编码RNA肺腺癌转移相关转录物1(MALAT1)通过调控核富集转录物1(NEAT1)对人肝癌细胞_(外泌体)分泌和肿瘤细胞增殖、侵袭的影响。方法肝癌细胞HuH-7细胞敲低MALAT1表达(si-MALAT1)转染获得si-MALAT1组细胞,转染无意义的小干扰RNA(si-RNA)作为si-NC组,比较si-MALAT1组和si-NC组NEAT1表达、细胞增殖和侵袭能力的变化。过表达NEAT1载体的慢病毒(lv)与HuH-7细胞共培养72 h后获取NEAT1过表达细胞(lv-NEAT1组),感染空转载体的lv作为lv-control组,比较lv-NEAT1组和lv-control组_(外泌体)相关基因表达差异。使用lv-NEAT1组细胞转染si-MALAT1获得si-MALAT1+lv-NEAT1组细胞,与si-NC组、si-MALAT1组细胞比较_(外泌体)分泌能力变化。将si-MALAT1组细胞与lv-NEAT1组细胞的_(外泌体)共培养获得si-MALAT1+lv-NEAT1_(外泌体)组细胞,将si-MALAT1组细胞与lv-control组细胞的_(外泌体)共培养获得si-MALAT1+lv-control_(外泌体)组,考察si-MALAT1+lv-NEAT1_(外泌体)组和si-MALAT1+lv-control_(外泌体)组细胞的增殖和侵袭功能。结果与si-NC组相比,si-MALAT1组中NEAT1的相对表达量[(0.72±0.02)比(0.98±0.01)]、72 h吸光度[(0.66±0.03)比(0.98±0.04)]、下室细胞数量[(88.33±7.26)比(147.70±13.62)]均降低,差异有统计学意义(P<0.05)。与si-NC组相比,si-MALAT1组_(外泌体)CD9、CD63表达减弱;而与si-MALAT1组相比,si-MALAT1+lv-NEAT1组_(外泌体)CD9、CD63表达增加。与si-MALAT1+lv-control_(外泌体)组相比,si-MALAT1+lv-NEAT1_(外泌体)组吸光度[(0.97±0.03)比(0.74±0.05)]和下室细胞数量[(132.70±7.36)比(98.33±6.01)]均增加,差异有统计学意义(P<0.05)。lv-NEAT1组_(外泌体)相关基因HSPA8、SLC3A2、SLC7A5的相对表达量分别为(5.53±0.31)、(0.32±0.07)、(0.77±0.45),与lv-control组表达水平(0.98±0.15)相比,差异均具有统计学意义(P<0.05)。结论MALAT1可通过调控NEAT1改变肝癌细胞_(外泌体)分泌功能,NEAT1可改变_(外泌体)相关基因的表达,进而促进了肝癌细胞的增殖和侵袭。
Objective To investigate the correlations between expression of long noncoding RNA metastasis-associated lung adenocarcinoma transcript 1(MALAT1),nuclear-enriched abundant transcript 1(NEAT1)and their functions on exosome secretion,proliferation and invasion in hepatocellular carcinoma(HCC).Methods We used small interfering RNA of MALAT1(si-MALAT1)to knockdown MALAT1 in HuH-7.At the meanwhile,cells which were transfected with si-NC were used as the negative control group.Expression of NEAT1,cell proliferation and invasion function were detected these two groups.HuH-7 cells were transfected with lentivirus NEAT1 over expressing vector(lv-NEAT1)or negative control(lv-control).Expression of exosomes secretion related genes were analyzed between lv-NEAT1 and lv-control groups.Cells of lv-NEAT1 were knockdown MALAT1 expression using si-MALAT1,which could be si-MALAT1+lv-NEAT1 group.exosomes secretion was detected in si-NC,si-MALAT1 and si-MALAT1+lv-NEAT1 group.We treated cells(si-MALAT1 group)with exosomes from cells with lv-NEAT1 or lv-control to divide cells as si-MALAT1+exosomes of lv-NEAT1 cells and si-MALAT1+exosomes of lv-control groups.Cell proliferation and invasion of cells were detected in two groups.Results Low expression of NEAT1 were found in MALAT1 knockdown cells compared with si-NC group[(0.72±0.02)vs.(0.98±0.01),P<0.05].Cells with MALAT1 knockdown shown diminished proliferation[(0.66±0.03)vs.(0.98±0.04),P<0.05)]and invasion[(88.33±7.26)vs.(147.70±13.62),P<0.05)].Compared with si-NC group,CD9 and CD63 expression were decreased in exosomes of si-MALAT1 group.Compared with si-MALAT1 group,CD9 and CD63 expression was increased in exosomes of si-MALAT1+lv-NEAT1 group.Compared with si-MALAT1+exosomes of lv-control group,proliferation[(0.97±0.03)vs.(0.74±0.05),P<0.05)]and invasion[(132.70±7.36)vs.(98.33±6.01),P<0.05)]were increased in si-MALAT1+exosomes of lv-NEAT1 group.Exosomes related genes expression including HSPA8(5.53±0.31),SLC3A2(0.32±0.07)and SLC7A5(0.77±0.45)were changed in lv-NEAT1 group compared with lv-control group[(0.98±0.15),P<0.05].Conclusion MALAT1 induced exosomes secretion by NEAT1 and exosomes related genes regulation.This regulation might be related with increased proliferation and invasion function in HCC cells with MALAT1 and NEAT1 abnormal expression.
作者
莽源祎
李立
冉江华
张升宁
李来邦
赵英鹏
高杨
赵姣姣
何祥乐
Mang Yuanyi;Li Li;Ran Jianghua;Zhang Shengning;Li Laibang;Zhao Yingpeng;Gao Yang;Zhao Jiaojiao;He Xiangle(Department of Hepato-biliary-pancreatic Surgery,the Calmette Affiliated Hospital of Kunming Medical University(the First People’s Hospital of Kunming),Clinical Medical Center for Organ Transplantation of Yunnan Province,Kunming 650011,China)
出处
《中华肝胆外科杂志》
CAS
CSCD
北大核心
2022年第4期289-294,共6页
Chinese Journal of Hepatobiliary Surgery
基金
昆明市科技计划省级"放管服"科研重点项目(2019-1-C-25318000002252)。
关键词
癌
肝细胞
RNA
外泌体
肺腺癌转移相关转录物1
核富集转录物1
Carcinoma,hepatocellular
RNA
Exosomes
Metastasis-associated lung adenocarcinoma transcript 1
Nuclear-enriched abundant transcript 1
