长链非编码RNA MALAT1影响口腔鳞状细胞癌侵袭的实验研究  

Metastasis-associated lung adenocarcinoma transcript 1 modulates oral squamous cell carcinoma invasion in vitro and in vivo

作　　者：刘速[1] 周旋[1] 王晓非[2] 岳恺[1] 段远胜[1] 何清华[1] 王佳鑫[1] 司海山 王旭东[1] 

机构地区：[1]天津医科大学肿瘤医院颌面耳鼻喉科,国家肿瘤临床医学研究中心,天津市肿瘤防治重点实验室,天津市300060 [2]天津市第一中心医院耳鼻喉科

出　　处：《中国肿瘤临床》2015年第9期460-465,共6页Chinese Journal of Clinical Oncology

基　　金：国家自然科学基金(编号:81172573);天津市应用基础与前沿技术研究计划(编号:12JCYBJC33800)资助~~

摘　　要：目的:探讨肺腺癌转移相关转录因子1(metastasis-associated lungadenocarcinoma transcript 1,MALAT1)对口腔鳞状细胞癌侵袭能力的影响。方法:应用RT-PCR方法检测MALAT1在口腔鳞状细胞癌组织标本、正常口腔黏膜组织标本以及口腔鳞癌细胞系中的表达;利用小干扰RNA(si RNA)敲低MALAT1在人舌鳞状细胞癌细胞Tscca中的表达;MTT法检测细胞的增殖能力变化;划痕实验、Transwell实验检测肿瘤细胞迁移、侵袭能力的变化;蛋白质印迹法检测肿瘤细胞迁移、侵袭及上皮间质转化(epithelial-mesenchymal transition,EMT)等相关蛋白的表达变化;免疫荧光法检测细胞EMT相关蛋白表达变化;建立Tscca裸鼠皮下荷瘤模型,免疫组织化学染色法检测细胞增殖、侵袭相关蛋白表达。结果:MALAT1在口腔鳞癌组织中的表达明显高于正常组织。抑制MALAT1表达后细胞增殖率下降,细胞系划痕闭合减慢,通过Transwell小室聚碳酸酯膜的细胞数减少,与对照组比较,差异有统计学意义(P<0.05);基质金属蛋白酶-2、-9(MMP-2、MMP-9)、神经钙黏素(N-cadherin)蛋白表达水平明显下调,钙黏着蛋白(E-cadherin)表达水平上调。免疫荧光显示,细胞神经钙黏素荧光强度明显减弱,钙黏着蛋白荧光强度显著增强。体内实验结果显示,MALAT1 si RNA治疗组裸鼠皮下荷瘤体积小于空白对照组及无义序列组(F=18.664,P<0.001);免疫组织化学染色结果示,MALAT1 si RNA治疗组中增殖核抗原(PCNA)、MMP-2、MMP-9表达减少。结论:MALAT1在口腔鳞癌组织中过表达,敲低人舌鳞癌细胞中MALAT1的表达可抑制舌鳞癌细胞的迁移、侵袭能力,MALAT1可能通过调控EMT促进口腔鳞癌的增殖和侵袭过程。Objective：To investigate the effect of metastasis-associated lung adenocarcinoma transcript 1 （MALAT1） in modulat-ing the effects of oral squamous cell carcinoma （OSCC） invasion. Methods：Real-time polymerase chain reaction was employed to de-tect the expression of MALAT1 in samples of OSCC post-radical resection, normal oral mucosa samples, and oral squamous cell lines. MALAT1-siRNA was transfected into TSCCa human tongue squamous cell carcinoma cell lines. Cell proliferation was determined by methyl-thiazolyl-tetrazolium reduction assay. Cell migration and invasive ability were evaluated by scratch test and transwell assay. The expression of proteins that regulated invasion and apoptosis were examined using Western blot assay. Immunofluorescence assay was used to detect changes in epithelial-mesenchymal transition （EMT）-associated proteins in the cells. Tumor-bearing nude mouse models were established by subcutaneous implantation of TSCCa cells. Immunohistochemistry was used to detect up-regulation of proliferating cell nuclear antigen （PCNA） and matrix metalloproteinase-2/9 （MMP-2/9）. Results：MALAT1 expression was significantly higher in OSCC than in normal tissues （P〈0.05）. MALAT1 expression was inhibited by transfecting MALAT1-siRNA. After MALAT1 expres-sion was down-regulated in TSCCa cells, proliferation was inhibited and invasion was attenuated, showing significant differences com-pared with the cells transfected with scrambled siRNA and control cells （P〈0.05）. Expression of N-cadherin and MMP-2/9 were down-regulated in the cells after MALAT1 was knocked down. Tumor growth was significantly slower in the MALAT1-siRNA group than in the control groups. IHC indicated that PCNA and MMP-2/9 expression of tumor tissues were significantly inhibited in MALAT1-siR-NA group. Conclusion：MALAT1 is over-expressed in human OSCC. MALAT1 reduction can inhibit the proliferation and invasion of OSCC cells. Furthermore, MALAT1 may promote OSCC invasion and metastasis by modulating EMT.

关 键 词：肺腺癌转移相关转录因子1 口腔鳞状细胞癌 肿瘤侵袭 EMT 

分 类 号：R739.8[医药卫生—肿瘤]