LncRNA PVT1对脑胶质瘤中GLUT3表达及细胞增殖、侵袭的影响  

Influence of long non-coding RNA plasmacytoma variant translocation 1 in cell proliferation,invasion and glucose transporter 3 expression in glioma

作　　者：卓胜华 张金本 羊良旺 陈申波 张召腾 李争争 杨堃[1] Zhuo Shenghua;Zhang Jinben;Yang Liangwang;Chen Shenbo;Zhang Zhaoteng;Li Zhengzheng;Yang Kun(Department of Neurosurgery,First Affiliated Hospital of Hainan Medical College,Haikou 570102,China)

机构地区：[1]海南医学院第一附属医院神经外科,海口570102

出　　处：《中华神经医学杂志》2021年第6期541-549,共9页Chinese Journal of Neuromedicine

基　　金：海南省财政科技计划资助项目(ZDYF2019129);国家自然科学基金(82060456);海南省研究生创新科研课题基金(Hys2019-312)。

摘　　要：目的探讨长链非编码RNA(LncRNA)浆细胞瘤变异易位1(PVT1)对脑胶质瘤中葡萄糖转运体3(GLUT3)表达及细胞增殖、侵袭能力的影响。方法(1)下载中国脑胶质瘤基因组图谱数据库中mRNAseq_325数据集,应用R语言分析222例原发性脑胶质瘤样本中PVT1与GLUT3基因表达的相关性以及二者对患者总生存期的影响。(2)收集海南医学院第一附属医院神经外科自2019年1月至2019年12月手术切除并经病理证实的8例低级别胶质瘤标本(低级别胶质瘤组)及7例胶质母细胞瘤标本(胶质母细胞瘤组),采用荧光原位杂交法检测PVT1的表达,免疫组化染色检测GLUT3蛋白的表达。(3)体外培养人星形胶质细胞NHA及胶质母细胞瘤细胞U87、LN229和U251(NHA组、U87组、LN229组、U251组),采用实时荧光定量PCR(RT-qPCR)检测各组细胞中PVT1、GLUT3 mRNA的表达,采用Western blotting实验检测GLUT3蛋白的表达。将U87、LN229细胞分别分为PVT1过表达质粒组与空白质粒组、PVT1短发夹RNA(shRNA)组与阴性对照shRNA组,分别构建慢病毒稳定转染过表达PVT1及其空白对照细胞系、敲低PVT1及其阴性对照细胞系,应用RT-qPCR和Western blotting实验检测各组细胞系中GLUT3 mRNA和蛋白的表达。(4)取PVT1 shRNA组与阴性对照shRNA组U87、LN229细胞,分别采用细胞计数试剂盒-8(CCK-8)实验、平板克隆形成实验检测细胞的增殖能力,采用Transwell侵袭实验检测细胞的侵袭能力。(5)将10只BALB/C-nu雌性裸鼠按随机数字表法分为实验组与对照组(n=5),分别移植PVT1 shRNA组与阴性对照shRNA组U87细胞以构建皮下移植瘤模型。移植4周后处死取瘤,称重及测量体积,并采用免疫组化染色检测肿瘤中Ki-67、GLUT3蛋白的表达。结果(1)mRNAseq_325数据集中222例原发性脑胶质瘤样本的PVT1与GLUT3基因表达呈正相关关系(r=0.514,P=0.000);与PVT1低表达组相比,PVT1高表达组的总生存期明显缩短,差异有统计学意义(P<0.05);与GLUT3�Objective To study the influence of long non-coding RNA(LncRNA)plasmacytoma variant translocation 1(PVT1)in glucose transporter 3(GLUT3)expression,and cell proliferation and invasion in glioma.Methods(1)The correlation between PVT1 and GLUT3 gene expressions and their influences in overall survival(OS)were analyzed using data from 222 cases of primary gliomas from Chinese Glioma Genome Atlas mRNAseq_325 data set.(2)Fifteen glioma specimens,including 8 from patients with low-grade glioma(LGG group)and 7 from patients with glioblastoma(GBM group),were collected in our hospital from January 2019 to December 2019;the PVT1 expression was detected by fluorescence in situ hybridization(FISH);the GLUT3 protein expression was detected by immunohistochemistry.(3)Normal human astrocyte(NHA)and glioblastoma cell lines U87,LN229 and U251(NHA group,U87 group,ln229 group and U251 group)were cultured in vitro;real-time fluorescent quantitative PCR(RT-qPCR)was used to detect the PVT1 and GLUT3 mRNA expressions;Western blotting was used to detect the GLUT3 protein expression;U87 and LN229 cells were divided into PVT1 overexpression plasmid group and blank plasmid group,PVT1 short hairpin RNA(shRNA)group and negative control shRNA group;the GLUT3 mRNA and protein expressions were detected by RT-qPCR and Western blotting.(4)In U87 and LN229 cells of negative control shRNA group and PVT1 shRNA group,CCK-8 assay and colony formation assay were used to detect the cell proliferation and Transwell assay was used to detect the cell invasion.(5)Ten female BALB/c-nu nude mice were randomly divided into experimental group and control group(n=5);the U87 cells from PVT1 shRNA group and negative control shRNA group were transplanted into the mice to establish subcutaneously transplanted tumor models.The animals were sacrificed and the tumors were weighed and measured 4 weeks after transplantation;the Ki-67 and GLUT3 protein expressions were detected by immunohistochemistry.Results(1)The gene expressions of PVT1 and GLUT3 were positively correla

关 键 词：神经胶质瘤 浆细胞瘤变异易位1 葡萄糖转运蛋白3 

分 类 号：R739.41[医药卫生—肿瘤]