缺氧相关的长链非编码RNA LINC00970在唾液腺腺样囊性癌中的表达及其作用  

The expression and role of hypoxia-induced long noncoding RNA LINC00970 in salivary adenoid cystic carcinoma cells

作　　者：朱韵莹 高晓琳 戈艳萍 王张嵩 林钊宇[2] 李劲松[2] 武东辉 Yunying Zhu;Xiaolin Gao;Yanping Ge;Zhangsong Wang;Zhaoyu Lin;Jinsong Li;Donghui Wu(Stomatology Hospital of Haizhu District,Guangzhou 510220,China;Sun Yat-sen Memorial Hospital,Sun Yat-sen University,Guangzhou 510120,China)

机构地区：[1]广州市海珠区口腔医院,广州510220 [2]中山大学孙逸仙纪念医院,广州510120

出　　处：《中华口腔医学研究杂志（电子版）》2023年第3期210-217,共8页Chinese Journal of Stomatological Research(Electronic Edition)

基　　金：广东省医学科学技术研究基金(A2021142)。

摘　　要：目的探讨唾液腺腺样囊性癌(SACC)中长链非编码RNA(lncRNA)LINC00970表达与缺氧的关系及其作用。方法低氧(1%O2)诱导48 h后,lncRNA芯片检测SACC-83细胞中缺氧诱导的lncRNA,并用实时荧光定量聚合酶链反应(PCR)验证。使用临床样本分析LINC00970在SACC组织中的表达及其与预后的关系。LINC00970敲低质粒转染SACC-LM和SACC-83细胞后,细胞增殖检测(CCK-8)和平板克隆实验检测细胞增殖能力和克隆形成能力,Transwell实验检测细胞侵袭和迁移能力,Western blot检测E-cadherin、Vimentin、Snail和N-cadherin的表达。荧光定量PCR、Western blot和Transwell等实验均重复3次所得均值为该样本测量值,使用SPSS 20.0软件进行统计学分析。SACC组织细胞与正常唾液腺组织中LINC00970相对表达量差异,采用配对t检验分析;比较沉默LINC00970对SACC-83和SACC-LM细胞增殖、克隆形成、侵袭及迁移能力的影响,以及LINC00970沉默对上皮-间充质转化(EMT)相关蛋白的影响,应用独立t检验进行统计学分析。采用Kaplan-Meier法计算LINC00970的表达与SACC患者总生存时间的相关性,并绘制生存曲线。P<0.05为差异有统计学意义。结果lncRNA芯片及实时荧光定量PCR结果显示,LINC00970是SACC-83细胞中在缺氧反应性表达最高的lncRNA(15.13±0.57 vs 1.08±0.06),差异有统计学意义(t=24.56,P<0.001)。临床样本数据分析显示,LINC00970在SACC组织表达显著高于正常唾液腺组织(0.49±0.41 vs 0.08±0.16,t=2.53,P<0.001),且LINC00970高表达患者生存率明显低于LINC00970低表达患者(HR=0.42,P=0.03)。SACC细胞敲低LINC00970后,缺氧促进的侵袭和迁移能力受抑制,且缺氧升高后的间质标志物N-cadherin表达下调,而缺氧诱导降低的上皮标志物E-cadherin恢复上调。结论缺氧可诱导SACC细胞中LINC00970的表达,其高表达与SACC患者的不良预后密切相关,LINC00970可能通过调控EMT从而促进细胞侵袭和迁移。Objective To investigate the relationship and function between the expression of long noncoding RNA(lncRNA)LINC00970 and hypoxia in salivary adenoid cystic carcinoma(SACC)cells.Methods After 48 hours of hypoxia(1%O2)induction,lncRNA chip was used to detect the lncRNA induced by hypoxia in SACC-83 cells and verified by qRT-PCR.Clinical samples were used to analyze the expression and prognostic impact of LINC00970 in SACC tissues.After transfection of SACC-LM and SACC-83 cells with LINC00970 shRNA plasmids,the cell proliferation,clonal formation ability,cell invasion and migration ability,and the expression of E-cadherin and N-cadherin were detected by CCK-8,plate cloning,Transwell assay and Western blot,respectively.Experiments such as fluorescence quantitative PCR,Western blot and Transwell were repeated three times to get the average value of the sample,and SPSS 20.0 software was used for statistical analysis.The difference between the relative expression of LINC00970 between SACC tissue cells and normal salivary gland tissue was analyzed by paired t-test.The independent t-test was used to compare the influence of silent LINC00970 on the proliferation,cloning,invasion and migration ability of SACC-83 and SACC-LM cells,as well as the effect of LINC00970 silence on EMT-related proteins.The KaplanMeier method was used to calculate the correlation between the expression of LINC00970 and the total survival time of SACC patients,and to draw the survival curve.P<0.05 was taken as statistically significant for the difference.Results The results of lncRNA microarray and qRT-PCR showed that LINC00970 was the most highly expressed lncRNA in SACC-83 cells in response to hypoxia(15.13±0.57 vs 1.08±0.06,t=24.56,P<0.001).Analysis of clinical sample data showed that the expression of LINC00970 in SACC tissues was significantly higher than that in normal salivary gland tissues(0.49±0.41 vs 0.08±0.16,t=2.53,P<0.001),and the survival rate of patients with high expression of LINC00970 was significantly lower than that of patien

关 键 词：癌 腺样囊性 唾液腺 细胞缺氧 长链非编码RNA 上皮-间充质转化 

分 类 号：R782.2[医药卫生—口腔医学]