宫颈和食管鳞状细胞癌组织中DNA甲基化谱分析及肿瘤标志物初步筛选

DNA methylation profiling and preliminary screening of tumor markers in cervical and esophageal squamous cell carcinoma

目的初步筛选与宫颈鳞状细胞癌和食管鳞状细胞癌发生相关的差异甲基化标志物。方法采用Illumina Infinium HumanMethylation450 Bead Chips(甲基化450k芯片)技术分别检测并分析8例宫颈鳞状细胞癌和8例食管鳞状细胞癌及其相应癌旁组织的DNA甲基化谱,筛选差异甲基化基因;结合基因本体(Gene Ontology,GO)分析、京都基因与基因组百科全书(Kyoto Encyclopedia of Genes and Genomes,KEGG)Pathway分析差异甲基化基因生物学功能,并通过UALCAN数据库和GEPIA数据库对差异基因进行验证。结果与癌旁组织相比,宫颈鳞状细胞癌和食管鳞状细胞癌中分别检测出29505个和54009个差异甲基化位点,两种肿瘤组织的共有差异位点5421个。这些共有甲基化差异位点主要位于启动子、gene body和3′UTR等基因功能区,基于这些共有甲基化位点,鉴定出相关基因2041个。结合GO功能注释分析显示在生物进展过程注释类别中,这些基因主要富集于信号传导、细胞黏附、胚胎骨骼系统形态发生;在分子功能注释类别中,主要集中于基因序列特异性结合、基因序列特异性结合转录因子活性、钙通道活性的调节;在细胞组分注释类别中,主要以质膜、质膜的整体组成、基底膜为中心。KEGG通路分析发现差异甲基化基因与黏着斑、癌症信号通路、细胞外基质受体相互作用和肌动蛋白细胞骨架调节有关,并筛选得到ITGA6基因可以作为以上2种肿瘤发生相关的候选基因;利用UALCAN数据库和GEPIA数据库分析显示ITGA6基因在2种肿瘤中高表达。结论ITGA6基因在鳞状细胞癌的形成中发挥重要作用,可作为诊断鳞状细胞癌的重要标志物。

Objective To preliminarily screen the differential methylation genes related to cervical and esophageal squamous cell carcinoma.Methods Illumina Infinium HumanMethylation450 Bead Chips(450k methylation chip)technology was used to detect and analyze DNA methylation profiles in 8 cases of cervical cancer,8 cases of esophageal cancer and their corresponding adjacent tissues.GO analysis and KEGG pathway were used to explore the biological functions of common differential methylated genes,and the common differentially methylated genes were verified by UALCAN database and GEPIA database.Results There were 29505 and 54009 differentially methylated sites in cervical cancer and esophageal cancer tissues,respectively.After further analysis,5421 common differential methylation sites were screened out,and they were mainly distributed in the gene function related regions,including promoter region,gene body region and 3′UTR region,of which,a total of 2041 related genes were identified.The GO function annotation showed these differential methylated genes are enriched in biological process,cell component and molecular function.In terms of biological progress,the differential methylated genes mainly enriched in signal transduction,cell adhesion and embryonic skeletal system morphogenesis;in terms of molecular function,the differential methylated genes mainly concentrated on sequence-specific DNA binding,sequence-specific DNA binding transcription factor activity and calcium ion binding;in terms of cell components,the diffe-rentially methylated genes mainly centered on plasma membrane,integral to plasma membrane and basement membrane.KEGG function annotation indicated that these genes are involved in the focal adhesion,pathways in cancer,ECM-receptor interaction and regulation of actin cytoskeleton.Furthermore,ITGA6 was identified as a key candidate gene in the tumor progression of cervical cancer and esophageal cancer.Analysis of data from UALCAN database and GEPIA database showed that the expression of ITGA6 mRNA in cervical cancer and esophageal cancer was higher than that in normal tissues.Conclusion ITGA6 gene plays an important role in the formation of squamous cell carcinoma and may be an important target in the diagnosis of squamous cell carcinoma.