MALAT-1沉默对口腔鳞癌细胞增殖、凋亡及Caspase-3通路的影响  

Effect of MALAT-1 silencing on proliferation,apoptosis and Caspase-3 pathway of oral squamous cell carcinoma

作　　者：肖宜 韩旭 张建华 徐晓刚 

机构地区：[1]上海市长海医院口腔科,200433 [2]上海市长海医院眼科,200433

出　　处：《临床和实验医学杂志》2018年第3期225-229,共5页Journal of Clinical and Experimental Medicine

基　　金：上海市科协委员会科研项目(编号:08411964600)

摘　　要：目的探究RNA干扰技术(RNAi)沉默肺癌转移相关转录本-1(MALAT-1)基因对口腔鳞状细胞癌(OSCC)细胞增殖、凋亡的影响并探讨其作用机制。方法采用实时荧光定量PCR(qRT-PCR)检测MALAT-1在人正常口腔上皮细胞系HIOE和OSCC细胞系SCC25中的表达情况。实验分为四组:空白对照组不转染序列;阴性对照组采用对照序列进行转染;RNAi组采用MALAT-1SiRNA进行转染;inhibitor组在SiRNA转染SCC25细胞前30 min,向培养基中加入Caspase-3抑制剂Ac-DEVD-CHO进行预处理(终浓度为40μmol/L)。采用RNAi技术敲低SCC25细胞中MALAT-1表达,采用透射电子显微镜(TEM)观察SCC25细胞超微结构;采用CKK-8法检测细胞增殖情况;采用流式膜联蛋白V-异硫氰酸荧光素碘化丙啶(Annexin V-FITC/PI)法检测细胞凋亡情况;采用实时荧光定量PCR(qRTPCR)和蛋白免疫印迹(WB)检测Caspase-3 mRNA和蛋白表达情况。结果与正常细胞比较,MALAT-1在人OSCC细胞系SCC25中表达上调,差异有统计学意义(P<0.05)。转染MALAT-1 SiRNA可明显降低SCC25细胞中MALAT-1表达(P<0.05)。与阴性对照组和空白对照组比较,RNAi组SCC25细胞增殖抑制率明显升高,细胞凋亡明显增加,差异均有统计学意义(P<0.05);细胞呈现出典型的凋亡特征;细胞中Caspase-3 mRNA和蛋白表达水平均升高(P<0.05)。加入Caspase-3抑制剂预处理,细胞凋亡明显减弱(P<0.05)。结论 MALAT-1沉默抑制人OSCC细胞系SCC25细胞增殖,促进凋亡,推测可能通过Caspase-3凋亡通路发挥作用。Objective To investigate the effect of RNA interference(RNAi) silencing the Metastasis associated lung adenocarcinoma transcript 1(MALAT-1) gene on the proliferation and apoptosis of oral squamous cell carcinoma(OSCC) and to explore its mechanism.Methods The expression of MALAT-1 in human normal oral epithelial cell line HIOE and OSCC cell line SCC25 was detected by real-time quantitative PCR(qRT-PCR).The experiment was divided into four groups:blank control group without transfected sequence;negative control group transfected with control sequence.The RNAi group was transfected with MALAT-1 siRNA.The inhibitor group was pretreated with Ac-DEVD-CHO,a Caspase-3 inhibitor(final concentration of 40 μmol/L) for 30 min before siRNA transfection into SCC25 cells.Knockdown the MALAT-1 expression in SCC25 cells by RNAi technique,transmission electron microscopy(TEM) was used to observe the ultrastructure of SCC25 cells.Cell proliferation was detected by CKK-8 assay;the apoptosis was detected by flow Annexin V-FITC/PI.Real time quantitative PCR(qRT-PCR) and Western blot(WB) were used to detect the expression of Caspase-3 mRNA and protein.Results Compared with normal cells,the expression of MALAT-1 was up-regulated in human OSCC cell line SCC25,and the difference was statistically significant(P <0.05).Transfection SiRNA of MALAT-1 could significantly reduce the expression of MALAT-1 in SCC25 cells(P < 0.05).Compared with the negative control group and the blank control group,the proliferation inhibition rate of SCC25 cells in the RNAi group was significantly increased,and the apoptosis rate was significantly increased,the differences were statistically significant(P < 0.05).The cells showed typical apoptotic characteristics,and the expression levels of Caspase-3 mRNA and protein in the cells were increased(P < 0.05).Apoptosis was significantly attenuated by pretreatment with Caspase-3 inhibitor(P < 0.05).Conclusion MALAT-1 silencing inhibits proliferation and promotes apoptosis of human OSCC cell line SCC25,suggesting tha

关 键 词：口腔鳞状细胞癌 肺癌转移相关转录本-1 半胱天冬蛋白酶-3 细胞增殖 细胞凋亡 

分 类 号：R739.8[医药卫生—肿瘤]