Objective: To investigate the whole-genome differential methylation profile of patients with high-altitude polycythemia (HAPC).
Methods: In this study, a total of 20 adult male patients with HAPC were included, including 10 Tibetan and 10 Han patients. The control group consisted of 20 healthy adult males, including 10 Tibetan and 10 Han patients. Peripheral blood was collected from each group for DNA extraction and quality inspection, and DNA libraries were constructed. The differential methylation regions (DMRs) between groups were detected using reduced representation bisulfite sequencing, with enriched regions compared to those of the control group. The differential enrichment regions were selected, and the intersection of the enriched regions was associated with genes. The methylation enrichment regions that differed significantly between groups were filtered based on the number of enriched samples in the enriched regions between the groups. GO, KEGG functional, and pathway analysis were performed on the differentially associated gene sets to reveal significant differences between the patients and control groups at the functional and pathway levels.
Results: In comparison with the control group, 17 152 sites with more than 25% difference and 15 558 sites with less than -25% difference were identified in Tibetan patients. The top 5 genes with the largest methylation differences between the two groups were MCCC2, RP3-399L15.3, ZNF621, RP11-394A14.2 and SLC39A10. The top significantly different pathways annotated in the differentially expressed genes pathway was serotonergic synapse. In comparison with the control group, 2 687 CpG sites with a greater than 25% difference and 2 602 CpG sites with a less than -25% difference were identified in Han patients. The top 5 genes with the largest methylation differences between the two groups were NAA25, CORO2B, PDC, ZNF853, and MLLT10. The top significantly different pathways annotated in the differentially expressed genes pathway were glutamatergic synapse, retrograde endocannabinoid signaling, Rap1 signaling pathway and cholinergic synapse. In comparison with the control group, 3 895 CpG sites with a greater than 25% difference and 3 969 CpG sites with a less than -25% difference were identified in HAPC patients. The maximum methylation difference between the two groups could reach 78.1%, while the minimum was -42.6%. The top 5 genes with the largest methylation differences between the two groups were MCCC2, ARSJ, CTNNA3, SLC39A10, and SWAP70. The top significantly different pathways annotated in the differentially expressed genes pathway was signaling pathways regulating pluripotency of stem cells.
Conclusion: The occurrence of HAPC may be related to abnormal changes in DNA methylation, and methylation sites may be helpful for the early diagnosis of HAPC.
题目: 高原红细胞增多症差异DNA甲基化谱研究.
目的: 探讨高原红细胞增多症（HAPC）患者全基因组差异甲基化谱。.
方法: 研究共纳入HAPC成年男性患者20例，藏、汉族患者各10例。对照组健康成年男性20例，藏、汉族各10例。取各组外周血进行DNA抽取与质检，构建DNA文库，组间的差异甲基化区域（DMR）使用简化代表性亚硫酸氢盐测序的方法进行检测，比对参考基因，将富集区域与对照组比较，取差异富集区域，差异富集区域取交集，将富集区域关联到基因，并根据组间富集区域富集样本个数差异筛选组间差异的甲基化富集区域，针对差异关联基因集进行GO、KEGG功能和通路富集分析。.
结果: 藏族患者与对照组相比单个CpG甲基化差异< 25%的位点共17 152个，< -25%的位点共15 558个。两组间甲基化差值最大的5个基因分别为MCCC2、RP3-399L15.3、ZNF621、RP11-394A14.2和SLC39A10。两组差异基因的信号通路注释中差异最显著的通路为血清素能突触。汉族患者与对照组相比单个CpG甲基化差异>25%的位点共2 687个，< -25%的位点共2 602个。两组间甲基化差值最大的5个基因分别为NAA25、CORO2B、PDC、ZNF853和MLLT10。差异最显著的基因信号通路为谷氨酸能突触、Rap1信号通路、逆行内源性大麻素信号传导和胆碱能突触。HAPC患者与对照组相比单个CpG甲基化差异位点< 25%的位点共3 895个，< -25%的位点共3 969个。两组甲基化差值最大的能达到78.1%，而最小为-42.6%，两组间甲基化差值最大的5个基因分别为MCCC2、ARSJ、CTNNA3、SLC39A10和SWAP70。差异基因最为显著的通路为调节干细胞多能性的信号通路。.
结论: HAPC的发生可能与DNA甲基化异常变化有关，甲基化位点可能对HAPC的早期诊断具有一定的帮助。.
Keywords: high-altitude polycythemia; epigenetics; DNA methylation; functional and pathway enrichment analysis.