基于异病同证从分子网络水平探讨缺血性脑卒中与冠心病共享遗传基础

Ischemic stroke and coronary artery disease seriously endanger people's lives. Atherosclerosis is their shared pathological basis, which is similar to the pathogenic mechanism of "phlegm and blood stasis" in traditional Chinese medicin (TCM). We previously had predicted and optimized stroke and coronary artery disease new hub genes based on the network analysis. After pathway enrichment and text mining, we found that these hub genes may be involved in the pathogenic mechanisms of the two diseases by participating in different biological processes. Therefore, we hypothesize that there are risk genetic variations in the new hub genes on the molecular network shared by stroke and coronary artery disease. And these genetic variations may be the underlying mechanism of the two diseases that could be used for explaining the theory of "same TCM syndrome for different diseases". To supported our hypothesis, we firstly detect the risk genetic variations for the susceptibility of stroke, coronary artery disease and their relevant TCM symptom-phlegm and blood stasis based on the association studies. Secondly,the influence of the risk loci on the genetic expression and miRNA combination will be further analyzed by using the luciferase reporter gene system and RT-PCR, thus to investigate their shared genetic basis. It can enrich the scientific implication of "same TCM syndrome for different diseases" and provide molecular genetics evidence for exploring the shared therapeutic targets of the two diseases.

缺血性脑卒中和冠心病严重危害人们的生命健康。现代医学认为动脉粥样硬化是它们共同的病理基础，与中医学痰瘀相关的致病机制的认识有共通之处。我们前期采用网络分析方法预测并优选出缺血性脑卒中和冠心病共享核心基因，并通过通路富集及文本挖掘发现，这些核心基因可能从不同生物过程参与缺血性脑卒中和冠心病的致病机制。基于此，本研究假设这些核心基因可能存在与这两种疾病及其相同中医证候有关联的风险遗传变异，而这些共享风险遗传变异可能是这两种疾病"异病同证"的内在机制。拟通过易感基因关联研究探寻与缺血性脑卒中、冠心病及其中医痰瘀证易感性相关的风险遗传变异，同时利用荧光素酶报告基因系统和RT-PCR深入分析这两种疾病及其痰瘀证相关风险位点对基因表达、miRNA结合的影响，以期探讨这两种疾病及其相同中医证候的共享遗传基础，进一步丰富中医学"异病同证"的科学内涵，为寻找这两种不同疾病共同的治疗靶点提供分子生物学依据。

摘要：缺血性脑卒中和冠心病都是严重危害人们的生命健康的重大疾病，中医学认为两病均与“痰瘀”相关，缺血性脑卒中痰瘀证和冠心病痰瘀证可能有共享分子遗传学基础。本项目主要完成了以下研究：首先，运用MassARRAY基因分型技术，在缺血性脑卒中痰瘀证550例、冠心病痰瘀证550例、健康对照550例样本人群中，完成了60个单核苷酸多态性（SNPs）位点的基因分型，发现CAV1基因rs1049337、SRC基因rs3762203、XRCC6基因rs5751129多态性与缺血性脑卒中痰瘀证、冠心病痰瘀证的发生风险显著关联，是两病的共享风险遗传变异。然后，运用qRT-PCR技术，在缺血性脑卒中痰瘀证患者50例、冠心病痰瘀证患者50例、健康对照50例样本人群中，完成了前期网络分析优选出的23个核心基因的外周血mRNAs表达水平测定，发现CALM1、CALR、DCN、EP300、F10、KDR、MAPK1、PRKCA、SHC1、STAT3、TP53、XRCC6共12个基因外周血mRNA表达与缺血性脑卒中痰瘀证、冠心病痰瘀证的发生均相关。最后，进一步通过细胞实验探索缺血性脑卒中痰瘀证与冠心病痰瘀证共享风险位点的生物学功能。运用双荧光素酶报告基因实验，发现CAV1基因rs1049337野生型通过与miR-1285结合抑制了CAV1基因的表达，而XRCC6基因的转录可被XRCC6基因rs5751129的突变型抑制。本项目鉴别了缺血性脑卒中痰瘀证与冠心病痰瘀证共享的易感遗传变异，并初步阐释了共享风险遗传变异影响两病发生的生物学功能作用。研究结果可为进一步丰富中医学“异病同证”理论提供分子遗传学依据，也为进一步探索“异病同治”的分子治疗靶点打下基础。本项目已发表论文12篇，培养硕士研究生 5 名，其中2名已取得硕士学位、3名在读。