LncRNA DLEU2对胃癌细胞增殖、凋亡和PI3K/Akt信号通路的影响  

Effect of lncRNA DLEU2 on proliferation, apoptosis and PI3K/Akt signaling pathway of gastric cancer cells

作　　者：秦延[1] 陈晓[1] 赵星[1] 霍志斌[1] 姜春艳 吴殿超[1] 赵笑枫 侯艳须 QIN Yan;CHEN Xiao;ZHAO Xing;HUO Zhibin;JIANG Chunyan;WU Dianchao;ZHAO Xiaofeng;HOU Yanxu(Department of Gastrointestinal Cancer,Xingtai People s Hospital,Xingtai 054001,China)

机构地区：[1]邢台市人民医院胃肠肿瘤外科

出　　处：《临床肿瘤学杂志》2020年第2期115-120,共6页Chinese Clinical Oncology

摘　　要：目的探讨长链非编码RNA淋巴细胞白血病缺失基因2(DLEU2)对胃癌细胞增殖、凋亡和磷脂酰肌醇3激酶(PI3K)/蛋白激酶B(Akt)信号通路的影响。方法通过GEPIA在线分析来自TCGA数据库和GTEx项目的胃癌组织DLEU2水平,采用实时定量PCR(QPCR)检测正常胃黏膜上皮GES-1细胞和胃癌BGC-823细胞的DLEU2水平;脂质体法将3条靶向DLEU2的小干扰RNA(si-DLEU2-1、si-DLEU2-2和si-DLEU2-3)转染至BGC-823细胞中,筛选最佳si-DLEU2干扰序列进行实验。体外常规培养BGC-823细胞并分为转染si-DLEU2的干扰组、转染无义siRNA序列的阴性对照(NC)组和未转染的空白对照(Blank)组,采用MTT比色法和AnnexinⅤ-FITC/PI双染流式细胞术检测细胞的增殖活力和凋亡率,QPCR和Western blotting检测凋亡相关因子Bcl-2、Bax和caspase-3的水平,Western blotting检测磷酸化的PI3K调节亚基p85(p-p85)和Akt(p-Akt)水平。结果GEPIA在线分析显示胃癌组织的DLEU2水平高于正常组织(P<0.05);QPCR结果显示BGC-823细胞的DLEU2水平高于GES-1细胞(P<0.05),且3条si-DLEU2序列转染后,BGC-823细胞的DLEU2水平均降低(P<0.05),其中si-DLEU2-3序列的效果最佳,故选取si-DLEU2-3序列进行后续实验;干扰组的DLEU2水平及转染24、48 h后的增殖活力均低于Blank组和NC组(P<0.05)。干扰组的凋亡率为(21.529±1.320)%,高于Blank组的(9.032±0.536)%和NC组的(8.641±0.365)%(P<0.05)。与Blank组和NC组相比,干扰组的Bcl-2、p-p85和p-Akt水平降低,而Bax和caspase-3水平均升高(P<0.05)。Blank组和NC组上述指标的差异无统计学意义(P>0.05)。结论DLEU2在胃癌组织和细胞中均升高且发挥促癌作用,可能通过激活PI3K/Akt信号通路来增强细胞增殖能力并抑制凋亡,有望成为胃癌治疗的新靶点。Objective To investigate the effect of long non-coding RNA deleted in lymphocytic leukaemia 2(DLEU2)on the proliferation,apoptosis and the phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt)signal transduction pathway of gastric cancer cells.Methods DLEU2 level of gastric cancer tissues from TCGA database and GTEX project was analyzed online by GEPIA.The DLEU2 levels of mormal gastric epithelial GES-1 cells and gastric cancer BGC-823 cells were detected by real-time quantitative PCR(QPCR).Three small interfering RNAs(si-DLEU2-1,si-DLEU2-2 and si-DLEU2-3)targeting DLEU2 were transfected into BGC-823 cells by liposome method,and the best si-DLEU2 interfering sequence was screened for experiment.BGC-823 cells were cultured in vitro and divided into three groups:Interference group(cells transfected with si-DLEU2),negative control(NC)group(cells transfected with nonsense siRNA sequence)and blank control(Blank)group(cells without transfection).MTT assay and flow cytometry via AnnexinⅤ-FITC/PI double staining were used to detect cell proliferation ability and apoptotic rate.QPCR and Western blotting were used to detect levels of Bcl-2,Bax and caspase-3.Phosphorylated levels of PI3K regulatory subunits p85(p-p85)and Akt(p-Akt)were detected by Western blotting.Results Online analysis of GEPIA showed that the DLEU2 level in gastric cancer tissues was higher than that in normal tissues(P<0.05).Results of QPCR showed that the DLEU2 level in BGC-823 cells was higher than that in GES-1 cells(P<0.05),and DLEU2 level in BGC-823 cells decreased after transfection with three si-DLEU2 sequences(P<0.05).Effect of si-DLEU2-3 sequence was the best,so the si-DLEU2-3 sequence was selected for following experiment.Proliferation activity after 24 and 48-h transfection and DLEU2 level of Interference group were lower than those of Blank group and NC group(P<0.05).The apoptotic rate of the Interference group was(21.529±1.320)%,higher than(9.032±0.536)%of Blank group and(8.641±0.365)%of NC group(P<0.05).Compared with Blank group a

关 键 词：胃癌 淋巴细胞白血病缺失基因2 增殖 凋亡 磷脂酰肌醇3激酶/蛋白激酶B通路 

分 类 号：R735.2[医药卫生—肿瘤]