基于CDKN2A通路的益气活血法治疗心肌梗死恢复期气虚血瘀证表观遗传调控研究

Myocardial infarction is one of the diseases with the highest morbidity and mortality in the world. It is an important clinical research topic to take active intervention measures to prevent myocardial remodeling and improve long-term prognosis in the recovery period of myocardial infarction..Myocardial fibrosis is the key link in this study，selecting the CDKN2A gene methylation on the chromosome 9p21 as the target .A case-control design was used to evaluate the level of methylation of different Cp G loci, and the correlation between CDKN2Asites by using Spearman correlation analysis. To determine the relationship between the level of CDKN2A methylation and the syndrome of qi deficiency and blood stasis in recovering period of myocardial infarction..In order to eliminate the influence of factors such as environment and lifestyle, the epigenetic regulation of Chinese medicine to related genes can be analyzed. The traditional Chinese medicine was used to treat the Qi deficiency and blood stasis syndrome n recovering period of myocardial infarction. by using Yiqi Huoxue Chinese medicine, epigenetic changes of CDKN2A before and after are observed and analysised of its correlation with curative effect..In the animal experiment, the environmental factors were excluded to study the traditional Chinese medicine, and the clinical study and animal experiment were combined to carry out the comprehensive curative effect analysis.In order to eliminate the influence of factors such as environment and lifestyle, the epigenetic regulation of Chinese medicine to related genes can be analyzed.

心肌梗死是世界发病率及致死率最高的疾病之一。心肌梗死恢复期采取积极干预措施,防治心肌重构和改善长期预后,是临床的重要研究课题。. 本研究以心肌纤维化为关键环节，选择染色体9p21上的CDKN2A基因甲基化为研究靶点，采用病例对照设计，运用Spearman相关分析，评估CDKN2A不同Cp G位点的甲基化水平及各位点间的两两相关性，以判断CDKN2A甲基化水平与心肌梗死恢复期气虚血瘀证的关联性。. 中药干预采用益气活血中药治疗心肌梗死恢复期气虚血瘀证，观察治疗前后CDKN2A的表观遗传学改变并分析其与疗效的相关性；并在动物实验中排除环境因素进行中药干预研究，将临床研究与动物实验相结合，进行综合疗效分析，排除环境及生活方式等外在因素的影响，可准确分析中药对相关基因的表观遗传调控。

心肌梗死是全球范围内亟需防控与解决的导致高死亡率的重大公共卫生问题之一。气虚血瘀证是心肌梗死最常见的核心证候，然而其证候本质尚未明确。CDKN2A/2B位于染色体9p，是两个邻近的依赖细胞周期蛋白激酶的抑制基因，对调节细胞增殖、衰老及凋亡都起着重要作用。我们的前期研究证实了CDKN2A/2B基因多态性与冠心病气虚血瘀证潜在关联，然其调控机制尚不明晰。本研究围绕心肌梗死恢复期气虚血瘀证的表观遗传学特征，通过临床和基础实验探究心肌梗死恢复期气虚血瘀证物质基础，阐明益气活血中成药芪参益气（QSYQ）滴丸调控CDKN2A抗心肌梗死后纤维化及延缓心室重构过程中的新机制。研究结果显示：1.QSYQ能够改善心肌梗死恢复期气虚血瘀证患者中医证候表现，其本质与DNA甲基化改变水平相关。QSYQ能够直接调控细胞周期通路中的核心靶标CDKN2A的DNA甲基化水平，这可能是益气活血药发挥改善气虚血瘀证的作用机制之一；2.QSYQ能够通过调控DNMTs活性水平及细胞周期调控核心靶标CDKN2A的蛋白表达和mRNA的转录，有效改善心肌梗死大鼠心肌病理学状态、下调与心肌纤维化相关的蛋白表达、改善心功能和发挥抗心肌梗死作用。因此，充分验证了以CDKN2A为核心的通路是益气活血中药抗心肌梗死纤维化的关键机制之一；3.QSYQ可以调控低氧诱导的CFs细胞中甲基化酶水平，进而促进CDKN2A基因在细胞周期调控过程中发挥作用，抑制细胞纤维化进程，减少CFs细胞过度增殖。最终证实QSYQ通过CDKN2A介导的细胞周期通路发挥对低氧诱导CFs细胞的抑制过度纤维化作用，这也是益气活血中药的抗心肌梗死恢复期气虚血瘀证的核心机制。本研究围绕CDKN2A基因甲基化对心室重构的影响以及对细胞周期的调控作用，进一步阐明心肌梗死气虚血瘀证的科学内涵及益气活血中药的延缓心室重构作用机制。