结直肠腺瘤基因低甲基化谱分析及其标记物的筛选研究

Analysis of gene hypomethylation profiles and screening of biomarkers in colorectal adenoma

目的分析结直肠腺瘤基因组低甲基化谱,并筛选低甲基化基因标记物。方法分别提取9例结直肠腺瘤组织和20例正常结直肠黏膜组织DNA,采用全基因组甲基化芯片(Infinium Human Methylation 450 Bead Chips)进行检测并分析其低甲基化谱,结合GO分析、KEGG_pathyway分析筛选差异低甲基化基因标记物。结果在结直肠腺瘤中共发现40071(61.14%)个差异低甲基化标位点,其中66.3%的低甲基化位点主要分布于Cp G岛以外的区域,在基因结构分析中,40%、37%、33%的低甲基化位点分别分布于基因间隔区域、基因内区域及启动子区。根据Δβ≤-0.4,在所有被研究的Cp G甲基化位点中共发现912个差异低甲基化标记物,包含584个差异低甲基化基因。GO分析、KEGG_pathyway分析表明,差异低甲基化基因在多种功能群落及信号转导通路中出现富集,提示可能通过这些通路参与结直肠腺瘤的发生发展。进一步按腺瘤组中平均甲基化程度β≤0.2以及结合GO、KEEG_pathway分析筛选出15个低甲基化基因标记物。结论在结直肠腺瘤中运用全基因组甲基化芯片筛选出来的低甲基化基因有可能用于结直肠腺瘤的早期诊断,并有可能为进一步探讨其发病机制提供依据。

Objective To analyze gene hypomethylation profiles and to screen biomarkers in colorectal adenoma by using Illumina ＇s Infinium Human Methylation 450 Bead Chip arrays combined with bioinformatics analysis. Methods Illumina＇s Infinium Human Methylation 450 Bead Chip arrays were used to examine DNA methylation profiles in 9 adenomas and 20 normal colorectal mucosa tissue. Hypomethylated gene biomarkers were screened by bioinformatics analysis. Results There were 40071（61.14%） hypomethylated sites in adenoma, of which 66.3% were mainly distributed in the open sea of Cp G island related regions. In the analysis of gene structure related regions, 40%, 37% and 33% of the hypomethylated sites were distributed in the intragenic region, intergenic region（body and 3＇UTR） and promoter region（TSS1500, TSS200, 5 ＇UTR and 1 st Exon）, respectively. According to Δβ ≤-0.4, we found 912 differential Cp G methylated sites, covering 584 genes. Go analysis and KEGG pathway analysis showed that differential hypomethylated genes in adenoma mainly enriched in several functional communities and signal transduction pathways. In this study, 15 hypomethylated gene markers were further screened by average β-value≤0.2 in methylation level of adenoma group and combined by GO, KEEGpathway analysis. Conclusion Genome-scale screening of hypomethylated gene biomarkers in adenoma may provide the possibility of early diagnosis of colorectal adenoma and further our understanding of the pathogenesis.