Objectives:Exosomal long noncoding RNAs(lncRNAs)might facilitate epithelial–mesenchymal transition(EMT)in liver cancer after transarterial chemoembolization(TACE),thereby enhancing tumor cell invasiveness and migrati...Objectives:Exosomal long noncoding RNAs(lncRNAs)might facilitate epithelial–mesenchymal transition(EMT)in liver cancer after transarterial chemoembolization(TACE),thereby enhancing tumor cell invasiveness and migration.This study investigated the prognostic role of plasma exosomal long noncoding RNA-plasmacytoma variant translocation 1(lncRNA-PVT1)in TACE treated hepatocellular carcinoma(HCC).Methods:Plasma exosomal lncRNA-PVT1 was evaluated via qPCR before and after TACE.Hepatoma cell behavior was investigated in different HCC cell lines.A lncRNA-PVT1 plasmid was synthesized and overexpressed,and si-lncRNA PVT1 was transfected into poorly invasive cells to reveal its influence on cell characteristics.The lncRNA-PVT1–FoxM1 interaction was elucidated using a double-luciferase reporter gene assay.The effect of miRNA-345-5p on minimally invasive hepatoma cells was assessed.Three experimental groups were established:poorly invasive cells,poorly invasive cells co-cultured with exosomes from hypoxia-induced highly invasive cells,and poorly invasive cells co-cultured with normal hepatocyte exosomes.Liver cancer cells were subcutaneously inoculated into nude mice,with subsequent observations of weight,tumor formation,and tumor size.Results:We identified two lncRNAs(lncRNA-PVT1 and GAPLINC)associated with EMT in the hypoxic microenvironment of liver cancer.Cox multivariate regression analysis was used to establish a prognostic model distinguishing high-and low-risk groups.Hypoxia-induced HepG2 exosomes significantly promoted EMT in poorly invasive HCC cells.LncRNA-PVT1 overexpression and silencing altered E-cadherin,vimentin,and FoxM1 expression,cell proliferation,invasion,migration,and apoptosis.miR-345-5p directly targeted lncRNA-PVT1 and FoxM1,affecting downstream targets.In vivo,co-culturing poorly invasive hepatoma cells with exosomes from highly invasive cells increased tumor volumes,upregulated lncRNA-PVT1,FoxM1,Ki67,and MMP9 expression,and downregulated miR-345-5p expression.Conclusions:Plasma exosomal lncRNA-PVT1 expression is upregulated in highly invasive cells post-hypoxia,potentially serving as a biomarker for evaluating liver cancer prognosis after TACE.Through a miRNA-345-5p-mediated competing endogenous RNA mechanism,it promotes EMT in poorly invasive cells,likely contributing to recurrence and metastasis post-HCC interventional embolization.展开更多
[目的]探讨lncRNA-PVT1通过Hp/CagA介导的胃黏膜屏障损伤促进胃癌的发病机制。[方法]检测lncRNA-PVT1在CagA阳性幽门螺杆菌菌株(Hp-CagA~+)和CagA基因敲除突变体(Hp-ΔCagA-)感染的人胃癌上皮细胞系AGS中的表达。过表达PVT1,分为对照组...[目的]探讨lncRNA-PVT1通过Hp/CagA介导的胃黏膜屏障损伤促进胃癌的发病机制。[方法]检测lncRNA-PVT1在CagA阳性幽门螺杆菌菌株(Hp-CagA~+)和CagA基因敲除突变体(Hp-ΔCagA-)感染的人胃癌上皮细胞系AGS中的表达。过表达PVT1,分为对照组和PVT1组。qPCR和荧光原位杂交技术测定lncRNA-PVT1的表达,CCK8和流式细胞术检测细胞活力及凋亡率。检测细胞中活性氧(ROS)水平,透射电子显微镜(TEM)观察细胞中线粒体的超微结构;Western blot检测两组中细胞周期蛋白CDK4和cyclinD1,凋亡蛋白BAX、Bcl-2和裂解的caspase3,线粒体裂变相关蛋白DRP1和MFN2,紧密连接蛋白(occludin,claudin-1和ZO-1)的表达。[结果 ] lncRNA-PVT1在Hp-CagA~+感染AGS细胞中表达增加。过表达PVT1后增加Hp-CagA~+感染AGS细胞中增殖,降低了其凋亡,调节HpCagA~+感染AGS细胞中CDK4(1.22±0.02 vs 1.65±0.03)和cyclinD1(1.16±0.05 vs 1.53±0.01)的表达,增加了ROS生成的增加和线粒体膜电位(MMP)而升高了细胞的比例,增加了线粒体脊的肿胀畸形,降低了DRP1的磷酸化和MFN2(1.16±0.06 vs 0.59±0.03)和BAX(1.05±0.06 vs 1.40±0.03)的表达,升高了Bcl-2(1.66±0.03 vs 1.24±0.02)和裂解的caspase3(1.03±0.06 vs 1.71±0.02)的表达,增加了氧化应激反应诱导的线粒体功能障碍。过表达PVT1通过降低紧密连接蛋白occludin(4.13±0.05 vs 1.31±0.02)、claudin-1 (5.03±0.06 vs 1.33±0.03)和ZO-1 (1.48±0.04 vs 0.60±0.01)表达增加了细胞屏障。[结论]lncRNA-PVT1通过Hp/CagA介导的胃黏膜屏障损伤促进胃癌的发病,促进肿瘤形成和发展。展开更多
基金the Ethics Committee of the West China Hospital,Sichuan University(No.2018256)the Animal Ethical and Welfare Committee of Sichuan University in China(No.20211218A).
文摘Objectives:Exosomal long noncoding RNAs(lncRNAs)might facilitate epithelial–mesenchymal transition(EMT)in liver cancer after transarterial chemoembolization(TACE),thereby enhancing tumor cell invasiveness and migration.This study investigated the prognostic role of plasma exosomal long noncoding RNA-plasmacytoma variant translocation 1(lncRNA-PVT1)in TACE treated hepatocellular carcinoma(HCC).Methods:Plasma exosomal lncRNA-PVT1 was evaluated via qPCR before and after TACE.Hepatoma cell behavior was investigated in different HCC cell lines.A lncRNA-PVT1 plasmid was synthesized and overexpressed,and si-lncRNA PVT1 was transfected into poorly invasive cells to reveal its influence on cell characteristics.The lncRNA-PVT1–FoxM1 interaction was elucidated using a double-luciferase reporter gene assay.The effect of miRNA-345-5p on minimally invasive hepatoma cells was assessed.Three experimental groups were established:poorly invasive cells,poorly invasive cells co-cultured with exosomes from hypoxia-induced highly invasive cells,and poorly invasive cells co-cultured with normal hepatocyte exosomes.Liver cancer cells were subcutaneously inoculated into nude mice,with subsequent observations of weight,tumor formation,and tumor size.Results:We identified two lncRNAs(lncRNA-PVT1 and GAPLINC)associated with EMT in the hypoxic microenvironment of liver cancer.Cox multivariate regression analysis was used to establish a prognostic model distinguishing high-and low-risk groups.Hypoxia-induced HepG2 exosomes significantly promoted EMT in poorly invasive HCC cells.LncRNA-PVT1 overexpression and silencing altered E-cadherin,vimentin,and FoxM1 expression,cell proliferation,invasion,migration,and apoptosis.miR-345-5p directly targeted lncRNA-PVT1 and FoxM1,affecting downstream targets.In vivo,co-culturing poorly invasive hepatoma cells with exosomes from highly invasive cells increased tumor volumes,upregulated lncRNA-PVT1,FoxM1,Ki67,and MMP9 expression,and downregulated miR-345-5p expression.Conclusions:Plasma exosomal lncRNA-PVT1 expression is upregulated in highly invasive cells post-hypoxia,potentially serving as a biomarker for evaluating liver cancer prognosis after TACE.Through a miRNA-345-5p-mediated competing endogenous RNA mechanism,it promotes EMT in poorly invasive cells,likely contributing to recurrence and metastasis post-HCC interventional embolization.
文摘[目的]探讨lncRNA-PVT1通过Hp/CagA介导的胃黏膜屏障损伤促进胃癌的发病机制。[方法]检测lncRNA-PVT1在CagA阳性幽门螺杆菌菌株(Hp-CagA~+)和CagA基因敲除突变体(Hp-ΔCagA-)感染的人胃癌上皮细胞系AGS中的表达。过表达PVT1,分为对照组和PVT1组。qPCR和荧光原位杂交技术测定lncRNA-PVT1的表达,CCK8和流式细胞术检测细胞活力及凋亡率。检测细胞中活性氧(ROS)水平,透射电子显微镜(TEM)观察细胞中线粒体的超微结构;Western blot检测两组中细胞周期蛋白CDK4和cyclinD1,凋亡蛋白BAX、Bcl-2和裂解的caspase3,线粒体裂变相关蛋白DRP1和MFN2,紧密连接蛋白(occludin,claudin-1和ZO-1)的表达。[结果 ] lncRNA-PVT1在Hp-CagA~+感染AGS细胞中表达增加。过表达PVT1后增加Hp-CagA~+感染AGS细胞中增殖,降低了其凋亡,调节HpCagA~+感染AGS细胞中CDK4(1.22±0.02 vs 1.65±0.03)和cyclinD1(1.16±0.05 vs 1.53±0.01)的表达,增加了ROS生成的增加和线粒体膜电位(MMP)而升高了细胞的比例,增加了线粒体脊的肿胀畸形,降低了DRP1的磷酸化和MFN2(1.16±0.06 vs 0.59±0.03)和BAX(1.05±0.06 vs 1.40±0.03)的表达,升高了Bcl-2(1.66±0.03 vs 1.24±0.02)和裂解的caspase3(1.03±0.06 vs 1.71±0.02)的表达,增加了氧化应激反应诱导的线粒体功能障碍。过表达PVT1通过降低紧密连接蛋白occludin(4.13±0.05 vs 1.31±0.02)、claudin-1 (5.03±0.06 vs 1.33±0.03)和ZO-1 (1.48±0.04 vs 0.60±0.01)表达增加了细胞屏障。[结论]lncRNA-PVT1通过Hp/CagA介导的胃黏膜屏障损伤促进胃癌的发病,促进肿瘤形成和发展。
