For a better understanding of epigenetic regulation of cell differentiation, it is important to analyze DNA methylation at a specific site. In this study, we examined changes in the methylation level of CCGG and GATCG sites during mouse folliculogenesis in paraffin-embedded sections of mouse ovaries. For the purpose, we used a new method, histo endonuclease-linked detection of methylation sites of DNA (HELMET), designed to detect methylation sites of DNA with a specific sequence in a tissue section. Unlike the global level of DNA methylation, which was no change in immunohistochemical staining of 5-methylcytosine throughout folliculogenesis, we found that there were hypermethylation of CCGG and GATCG sites in most of the granulosa cells of tertiary follicles compared to that of primary and secondary follicles. Interestingly, TUNEL-positive granulosa cells, which were frequent in mammalian folliculogenesis, became markedly Hpa II-reactive and Sau3A I-reactive, indicating that the CCGG and GATCG sites may be preferentially demethylated during apoptosis.

中文翻译：

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HELMET评估小鼠卵巢卵泡形成过程中卵母细胞和颗粒细胞DNA甲基化的变化。

为了更好地了解细胞分化的表观遗传学调控，重要的是分析特定位点的DNA甲基化。在这项研究中，我们检查了小鼠卵巢中石蜡包埋的部分在卵泡形成过程中CCGG和GATCG位点甲基化水平的变化。为此，我们使用了一种新方法，即组织内切核酸酶联结的DNA甲基化位点检测（HELMET），该方法旨在检测组织切片中具有特定序列的DNA甲基化位点。与全球甲基化水平不同，在整个卵泡形成过程中5-甲基胞嘧啶的免疫组织化学染色没有变化，我们发现，与初级和次级卵泡相比，第三级卵泡的大多数颗粒细胞中CCGG和GATCG位点存在甲基化。有趣的是，