X射线辐照对CAL-27细胞株microRNA表达谱影响的生物信息学分析

摘要

CAL-27, a human tongue squamous carcinoma cell line was selected for the experiment. The cell clone formation rate was detected by a cell clone forming experiment and flow cytometry was used to detect apoptosis. High throughput sequencing technique (HiSeq) was applied for detecting the differentially expressed miRNAs (DEGs) after irradiating CAL-27 cells with X-rays. Using bioinformatics analysis, the distribution of candidate target genes and significantly enriched GO-term genes were obtained. The main signaling pathways involving the DEGs were obtained using KEGG pathway enrichment analysis. The CAL-27 cell clone survival rate of the 2 Gy irradiated group was 59% and the apoptosis rate was 47.5% (p<0.05). After irradiation, 21 miRNAs were altered, of which 18 miRNAs were upregulated (hsa-miR-21-3p, hsa-miR-101-3p, hsa-miR-100-3p, etc.) and 3 miRNAs were downregulated (hsa-miR-204-5p, hsa-miR-10a-5p, hsa-miR-146b-5p). When analyzed by GO function, there were 23 GO-terms enriched in biological process, 20 GO-terms enriched in molecular function, and 18 GO-terms enriched in cellular component. Furthermore, 18 KEGG pathways in which the DEGs participated, were screened out including olfactory transduction, cytokine-cytokine receptor interaction, MAPK signaling pathway-yeast (p<0.05), etc.These results indicate that X-ray irradiation with a dose of 2 Gy altered the miRNA expression profile of CAL-27 cells, and the underlying mechanism may involve MAPK signaling pathways. This study lays a foundation for further study of miRNAs in the molecular mechanism of tongue squamous carcinoma radiotherapy.%选用生长状态良好的人舌鳞癌CAL-27细胞用于实验,X射线辐照至吸收剂量为2 Gy后,通过细胞克隆成形实验检测CAL-27细胞株的存活率,流式细胞技术检测细胞凋亡率;为研究其损伤机理进一步运用高通量测序技术(High throughput sequencing technique,HiSeq)进行测序,分析吸收剂量为2 Gy的受照组CAL-27细胞miRNA的表达变化,并对其靶基因进行基因分类(Gene ontology,GO)和通路分析.结果表明:吸收剂量为2 Gy的受照组CAL-27细胞克隆存活率为59%(p<0.05),细胞凋亡率为47.5%(p<0.05);吸收剂量为2Gy的受照组CAL-27细胞中共有21个已知miRNAs表达差异,其中有18个miRNAs表达上调,3个miRNAs表达下调;在靶基因GO生物学功能分析中,靶基因在生物学过程、细胞组分和分子功能中各有23个条目、18个条目和20个条目;通过KEGG数据库分析筛选出了18条信号通路,如嗅觉传导通路、细胞因子受体交互通路、丝裂原活化蛋白激酶(Mitogen-activated protein kinases,MAPK)信号通路等(p<0.05).结果提示,X射线辐照至吸收剂量为2 Gy时影响了人舌鳞癌CAL-27细胞miRNA表达谱,其机理可能与MAPK信号传导通路有关,这将为研究相关miRNA在舌鳞癌放射治疗中的分子生物学机理并寻找其标志基因打下一定基础.