基于DNA甲基化/羟甲基化修饰探讨早发冠心病血瘀证及家系的表观遗传调控机制及中药干预研究

Geneticist based on DNA methylation/methylolation play an important role in elucidating disease gene and environment interaction.But the mechanism of premature coronary heart disease with blood stasis syndrome has not been given a comprehensive interpretation. In the basis of previous research on genetic genes of blood stasis syndrome in premature coronary heart disease , We 'll do the experiment by detection the whole genic DNA methylation profile through the high- throughput filtration technologies on oligonucleotide microarray , biological information analysis, Methylation-specific polymerase chain reaction, hMeDIP, two-dimension gel electrophoresis and mass spectrum for①To investigate the mechanism of DNA methylation/methylolation and the development of the blood stasis syndrome in patients with premature coronary heart disease ②Yangxin Tongmai Fang`s intervention in the DNA methylation/methylolation in the level of animal and cell and the analysis and discussion of traditional Chinese medicine intervention on the model of DNA methylation/methylolation in epigenetic regulation mechanism.③Though the use of xuefu zhuyu decoction on blood stasis syndrome of premature coronary heart disease patients and pedigree with syndrome differentiation treatment, the connection between research of gene epigenetics and clinical application will be comprehended from the view of systems biology include gene methylation/methylolation - internal phenotype - patterns of disease - Chinese medicine intervention. The above research wil discover the mechanism of Chinese medicine intervention in regulating the methylation of premature coronary heart disease and pedigree with blood stasis , which provide a new idea to promote Chinese medicine prevention and therapy of coronary heart disease with blood stasis syndrome.

基于DNA甲基化/羟甲基化的表观遗传学在阐明疾病基因和环境相互作用方面发挥重要作用，但早发冠心病及血瘀证发生发展的机理及中药干预机理尚未全面阐释。因此，我们在前期早发冠心病血瘀证遗传基因及甲基化研究的基础上,拟通过采用高通量寡核苷酸芯片筛选、甲基化特异PCR、羟甲基化DNA免疫共沉淀法、iTRAQ差异蛋白质组学质谱定量及生物信息分析等技术，开展①探讨DNA甲基化/羟甲基化修饰与早发冠心病血瘀证患者发生发展的相关机制；②养心通脉方干预细胞的DNA甲基化/羟甲基化修饰；③血府逐瘀汤对早发冠心病血瘀证患者和家系进行辨证调理，以方验证和构建早发冠心病血瘀证的甲基化/羟甲基化修饰的调控网络。从基因甲基化/羟甲基化-内部表型-疾病模式-中药干预的系统生物学角度把握早发冠心病血瘀证的基因表达与临床应用的结合点，以期发现中药在冠心病血瘀证表观遗传层面的调控机制,以促进冠心病中医药预防和临床治疗。

基于DNA甲基化/羟甲基化的表观遗传学在阐明疾病基因和环境相互作用方面发挥重要作用，但早发冠心病（PCHD)及血瘀证发生发展的机理及中药干预机理尚未全面阐释。因此，我们在前期PCHD血瘀证遗传基因及甲基化研究的基础上,通过采用高通量寡核苷酸芯片筛选、甲基化特异PCR、羟甲基化DNA免疫共沉淀法、iTRAQ差异蛋白质组学质谱定量及生物信息分析等技术，开展了①探讨DNA羟甲基化对PCHD血瘀证相关易感基因的影响，发现PCHD血瘀证相关的差异羟甲基化基因有128个，相关通路有30条，主要与心肌细胞能量代谢、炎症、斑块形成、内皮损伤、免疫反应等有关; ②根据PCHD血瘀证患者外周血全基因组芯片mRNA表达，发现相关的差异表达基因有405个，相关的信号通路有20余条，主要与血管内皮损伤、炎症、斑块及脂质代谢等有关；③基于iTRAQ技术分析家系PCHD血瘀证的血浆差异蛋白,发现与家系PCHD血瘀证相关的差异蛋白共32个；上调的差异蛋白功能涉及血小板激活、脂质代谢、免疫反应、胰岛素抵抗等;下调的差异蛋白功能涉及补体系统激活、PI3K-Akt通路激活、抗氧化功能等；④基于DNA甲基化/羟甲基化水平探索中药对PCHD血瘀证易感基因的干预前后改变，以方验证探讨中药对血瘀证亚型对应的易感基因甲基化/羟甲基化的表观遗传调控机制。发现养心通脉方加减对PCHD血瘀证三个亚型ZEB2、DES两个易感基因的DNA甲基化有明显的调控作用，进一步证实了ZEB2、DES基因的甲基化修饰是PCHD血瘀证的发病机制之一。初步构建了PCHD血瘀证的甲基化/羟甲基化修饰的调控网络。从基因甲基化/羟甲基化-内部表型-疾病模式-中药干预的系统生物学角度把握了早发冠心病血瘀证的基因表达与临床应用的结合点，探讨了中药在冠心病血瘀证表观遗传层面的调控机制,为促进冠心病中医药预防和临床治疗提供了有益的参考和借鉴。