Genome-Wide DNA Methylation Profiling in Cultured Eutopic and Ectopic Endometrial Stromal Cells

The objective of this study was to characterize the genome-wide DNA methylation profiles of isolated endometrial stromal cells obtained from eutopic endometria with (euESCa) and without endometriosis (euESCb) and ovarian endometrial cysts (choESC). Three samples were analyzed in each group. The infinium methylation array identified more hypermethylated and hypomethylated CpGs in choESC than in euESCa, and only a few genes were methylated differently in euESCa and euESCb. A functional analysis revealed that signal transduction, developmental processes, immunity, etc. were different in choESC and euESCa. A clustering analysis and a principal component analysis performed based on the methylation levels segregated choESC from euESC, while euESCa and euESCb were identical. A transcriptome analysis was then conducted and the results were compared with those of the DNA methylation analysis. Interestingly, the hierarchical clustering and principal component analyses showed that choESC were segregated from euESCa and euESCb in the DNA methylation analysis, while no segregation was recognized in the transcriptome analysis. The mRNA expression levels of the epigenetic modification enzymes, including DNA methyltransferases, obtained from the specimens were not significantly different between the groups. Some of the differentially methylated and/or expressed genes (NR5A1, STAR, STRA6 and HSD17B2), which are related with steroidogenesis, were validated by independent methods in a larger number of samples. Our findings indicate that different DNA methylation profiles exist in ectopic ESC, highlighting the benefits of genome wide DNA methylation analyses over transcriptome analyses in clarifying the development and characterization of endometriosis.

本研究旨在明确三类子宫内膜间质细胞的全基因组DNA甲基化谱特征：分别从携带子宫内膜异位症的在位子宫内膜分离得到的细胞（euESCa）、未携带子宫内膜异位症的在位子宫内膜分离得到的细胞（euESCb），以及卵巢子宫内膜异位囊肿来源的细胞（choESC）。每组均设置3份样本进行分析。采用因菲尼姆甲基化芯片（Infinium Methylation Array）检测发现，与euESCa相比，choESC中存在更多高甲基化与低甲基化的CpG位点；且euESCa与euESCb之间仅存在少量基因的甲基化差异。功能分析显示，choESC与euESCa在信号转导、发育过程、免疫应答等生物学过程中存在显著差异。基于甲基化水平开展的聚类分析与主成分分析结果显示，choESC可与euESC明显区分，而euESCa与euESCb则无显著差异。随后本研究开展了转录组分析，并将其结果与DNA甲基化分析结果进行对比。值得注意的是，层级聚类与主成分分析结果显示，在DNA甲基化分析中choESC可与euESCa、euESCb明显区分，但转录组分析中未观察到此类分组差异。从样本中提取的表观遗传修饰酶（包括DNA甲基转移酶）的mRNA表达水平在各组间无显著差异。部分与类固醇生成相关的差异甲基化和/或差异表达基因（NR5A1、STAR、STRA6及HSD17B2）已通过独立实验方法在更大样本量中得到验证。本研究结果表明，异位子宫内膜间质细胞存在独特的DNA甲基化谱特征，凸显了全基因组DNA甲基化分析相较于转录组分析，在阐明子宫内膜异位症的发生发展与特征分型方面的优势。