口腔鳞状细胞癌组织特征长链非编码RNA(lncRNA)中CCAT2的表达及作用分析

Expression and Role of lncRNA CCAT2 in Oral Squamous Cell Carcinoma

目的分析口腔鳞癌组织特征长链非编码RNA(lncRNA)表达谱,探讨结肠癌相关转录子2(CCAT2)的表达及作用.方法使用高通量lncRNA芯片检测5例口腔鳞癌患者的癌组织及癌旁正常口腔黏膜组织lncRNA表达水平,分析特征性lncRNA表达谱;应用RT-q PCR检测74例口腔鳞癌组织及癌旁组织中CCAT2表达水平,分析其与临床病理特征的关系;CCK8法检测CCAT2干预细胞增殖的能力,检测CCAT2-siRNA感染序列靶向转染人舌鳞状细胞癌cal27细胞株.结果 5例癌组织和对应的癌旁组织的差异表达谱共检出1 572个差异具有统计学意义的lncRNA,其中882个lncRNA表达上调,690个表达下调;19个lncRNA差异表达倍数≥10,其中11个表达上调,8个表达下调;CCAT2上调最明显.进一步对74例口腔鳞状细胞癌组织行RT-qPCR检测,癌组织中CCAT2的相对表达量高于正常组织,差异具有显著统计学意义(P<0.01).Ⅲ+Ⅳ期患者CCAT2相对表达量高于Ⅰ+Ⅱ期患者,肿瘤低分化组CCAT2相对表达量高于中高分化组,差异均具有统计学意义(P<0.05).cal27细胞转染CCAT2 siRNA1,siRNA2,siRNA3后,CCAT2的相对表达量均低于空白对照组,差异具有统计学意义(P<0.05).其中以siRNA2下降幅度最明显,因此,选用CCAT2-siRNA2进行细胞增殖实验.CCK8检测显示:转染48 h后细胞增殖水平低于阴性对照组,差异具有统计学意义(P<0.05).结论 lncRNA在口腔鳞状细胞癌组织中存在异常表达,其中CCAT2表达上调与口腔鳞状细胞癌的发生、发展有关.

Objective To analyze the characteristics of long chain non coding RNA（ lncRNA） expression in human oral squamous cell carcinoma（ SCC）,and to investigate the expression and role of transcription factor 2（ CCAT2）. Method High-throughput lncRNA microarray was used to detect the lncRNA expression in the oral mucosa carcinoma and the normal oral mucosa tissues of 5 patients with oral cancer squamous cell carcinoma,the characteristic lncRNA expression profile was analyzed,and then RT-q PCR was conducted to detect the expression level of CCAT2 in the carcinoma tissue and the adjacent tissue in 74 cases of oral squamous cell carcinoma and the relationship of CCAT2 expression with the clinical pathological characteristics was analyzed; CCAT2-siRNA-infected sequence was transfected to the target human tongue squamous cell carcinoma cell line cal27,and the proliferation of CCAT2 interference cells was detected by CCK8 method. Results 1572 lncRNA with statistically significant differences were found in the differential expression profile of the cancer tissues of 5 cases and the corresponding paracancerous tissues,of which 882 IncRNA expressions were up-regulated and 690 were down-regulated; the differential expression multiple of 19 lncRNA was more than 10,of which 11 were upregulated and 8 down-regulated,and the expression up-regulation of CCAT2 was the most significant; furthermore,oral squamous cell carcinoma tissues of 74 cases were detected by RT-q PCR,showing that the relative expression of CCAT2 in the cancer tissues was significantly higher than that in the normal tissues（ P〈0. 01）. The relative expression level of CCAT2 in patients at stage III and IV was significantly higher than that in patients at stage I and II,and the relative expression of CCAT2 in the poorly differentiated group was significantly higher than that in the middle and high differentiation group（ P〈0. 05）. The relative expression of CCAT2 in cal27 cells after transfection with CCAT2,siRNA2 and siRNA3 was lower than that in the control group,and the difference was statistically significant（ siRNA1）（ P〈0. 05）. The siRNA2 decreased most significantly,so CCAT2 siRNA2 was selected for the cell proliferation experiment,and CCK8 assay showed that cell proliferation level after the transfection for 48 h was significantly lower than that in the negative control group（ P〈0. 05）. Conclusion There is an abnormal expression of lncRNA in human oral squamous cell carcinoma,and the expression of CCAT2 is related to the occurrence and development of squamous cell carcinoma of the oral cavity.