| Objective:By analyzing about the characterization of coronary heart disease with blood stasis syndrome, which including the symptoms, signs and prethrombotic state moleculars, gene expression profiling and susceptible gene promoter methylation status, to discuss that if coronary heart disease with blood stasis syndrome essence exists heterogeneity?Method:It is the literature research firstly. By searching the CNKI database, the VIP database, the Wanfang database, the Chinese biomedical literature database and the Pubmed database, to query the clinical literature on blood stasis syndrome of coronary heart disease roundly. The clinical literatures mainly include two aspects. One is the studies of the clinical effect of Xuefu Zhuyu decoction in the treatment of coronary heart disease with blood stasis. Other is objective studies of the coronary heart disease with blood stasis syndrome, including using high-throughput techniques (genome, proteome, metabolome, etc) and other microscopic indexes. Then we extract the information about the documents, and carry the system evaluation by utilizing the meta analysis method. Secondly, to analyze the clinical characterization of the coronary heart disease with blood stasis, including symptoms, signs and prethrombotic state molecule, by using the systematic cluster analysis method and according to the principle with large spacing between groups, small spacing within a group. Thirdly, micro array technology was used to analyze gene expression about study groups (family groups:coronary heart disease with blood stasis syndrome, non blood stasis syndrome, non blood stasis coronary heart disease and healthy people; non family group: health people, blood stasis in patients with coronary heart disease). And real time fluorescent quantitative RT-PCR technique was used on numerous differences selected gene expression validation. More, by methylation specific PCR (MS-PCR) was applied to analyze the methylation status of the promoter.Results:The inclusion of the nearly ten years (2003-2012),18articles about Xuefu Zhuyu Decoction in the treatment of angina pectoris of coronary heart disease with blood stasis syndrome were analyzed, and the results showed that Xuefu Zhuyu Decoction could improve the performance of the TCM syndrome curative effect, angina pectoris effects and ECG efficacy, but the independent research within the effect extent existed the heterogeneity of cured, effective, ineffective and worse. Study on coronary heart disease of blood stasis with high-throughput technology access total21literatures study, included the gene expression profile of3articles,11articles on proteomics,6Metabonomics studies. And studies on other microscopic indexes have35articles(13single nucleotide polymorphism studies of peripheral blood white cells,22other articles).some studies, which met the inclusion criteria about ACE, ApoE and F VII gene polymorphism research, other microscopic indicators (nitric oxide, von Willebrand factor, endothelin, soluble intercellular adhesion molecule, tissue plasminogen activator inhibitor-1, tissue plasminogen activator) was analyzed by meta. The results showed that whether the research analysis method or not, the existing research have different degrees of heterogeneity in outcome indicators. In66patients with the coronary heart disease with blood stasis (including39family cases of patients with coronary heart disease with blood stasis,27non family cases of coronary heart disease with blood stasis), system cluster analysis of symptoms and signs in10diagnostic elements of tightness in the chest, the blood stasis prickling, face purple, lip green, nail green, tongue color, tongue petechiae, Knotted and Intermittent Pulse.The results showed that the clinical characterization of blood stasis syndrome could be divided into two subgroups. Cluster1feature:the combinations of symptoms (tightness in the chest, lip green and face purple) as the main performance, other symptoms and signs were rare. Cluster2features:the combinations of symptoms (tightness in the chest, lip green, face purple, blood stasis prickling and the tongue color) were the most common symptoms, tongue petechiaes was extremely atypical symptoms. Through the statistical analysis of the2subgroups respectively with the family blood stasis syndrome of coronary heart disease, which has a genetic background and non-family blood stasis syndrome of coronary heart disease without the chromosomal background. In cluster1, family patients with blood stasis syndrome had12cases, but non family patients with blood stasis syndrome had21cases. However, in cluster2, family patients had27cases, but non family previously had6cases, with statistical significance. In patients with coronary heart disease with blood stasis of the prethrombotic state molecular (PTS) included the8markers: the levels of plasma thrombomodulin (TM), P-selectin, prothrombin fragment1,2(F1+2), soluble fibrin monomer complex(SFMC), tissue type plasminogen activator (tPA), plasminogen activator inhibitor (PAI-1), thrombin antithrombin(TAT) and C protein peptide (PCP) as the object of system clustering analysis and statistics. The results show that the family of blood stasis syndrome of coronary heart disease and family health had statistical differences in TM, P-Selectin,F1+2, t-PA, TAT, PCP six molecules, there was no significant difference in non family blood stasis syndrome and non-family health of human, and the family of blood stasis syndrome of coronary heart disease and non-family of blood stasis syndrome of coronary heart disease, In t-PA, PAI-1, TAT three molecules there was statistical difference.Hierarchical cluster analysis (Ward) showed that PTS molecules could be clustered into two groups. In cluster1,8molecule expressed higher as the characteristic, and in cluster2,8molecules expressed relatively low as the characteristics, and were associated with hereditary factors. In cluster1,14pedigrees of blood stasis syndrome of coronary heart disease compared with4non family cases of coronary heart disease. In cluster2, family blood stasis syndrome of coronary heart disease had10cases, but non family Blood Stasis Syndrome of coronary heart disease had17cases, with statistical difference.6groups had a better distinction in gene expression by microarray analysis.5selected genes were detected by real-time RT-PCR validation and were better consistent with the chip results. So the chip results were reliable. The pedigree of blood stasis syndrome compared with non family in gene expression spectrum, results showed that by screening with FC<2or FC<0.5standards, the two groups had the same78gene probe number, respectively, accounted for only family and non family blood stasis syndrome gene expression ratio was4.12%(78/1893) and3.69%(78/2113). With FC>3or FC<1/3screening, the two groups had the same35gene probe number, respectively, accounted for single family and non family blood stasis syndrome gene expression ratio was5.11%(35/685) and4.62%(35/758).And the two distinct gene expression spectrum exist an obvious difference in the GO distribution and the pathway. Two groups of heterogeneous expression gene probe in173different probe are the same, but there is a difference in the expression of the direction, i.e. a group of "up" but another group of "down". This part of the gene by SAS system on-line comments, results show that at present this part probe had81genes annotated in the database. The gene ontology (GO) analysis and path analysis (pathway), Enrichment test p value<0.05as the selection criteria, by screening for GO analysis of a total of34valuable genes belong to different classes, Pathway analysis can obtain43meaningful pathway.In34GO functional annotations of genes may be associated with blood stasis syndrome gene were CEACAM1, CXCL1and IL8.From the43pathways, Adhesion and Diapedesis of Lymphocytes, Cells and Molecules involved in local acute inflammatory response, Adhesion Molecules on Lymphocyte, Free Radical Induced Apoptosis, Eph Kinases and ephrins support platelet aggregation, Monocyte and its Surface Molecules, B Cell Survival Pathway, CD40L Signaling Pathway, Adhesion and Diapedesis of Granulocytes, IL17Signaling Pathway, Ras-Independent pathway in NK cell-mediated cytotoxicity, PTEN dependent cell cycle arrest and apoptosis, Cytokine Network and Cytokines and Inflammatory Response and other inflammatory immune related pathways may also had potential association with blood stasis syndrome of coronary heart disease. However, in the family blood stasis syndrome and non-family blood stasis syndrome of coronary heart disease in two groups, these genes or pathways of gene expression were the opposite. Expression of susceptibility genes were screened through the identical syndrome in different diseases, the same disease with different combination way.The same syndrome in different diseases screening method of susceptible gene probe number739(FC<2or FC<0.5annotated genes,406) or susceptible gene probe number162(FC<3or FC<1/3,93genes have been annotated); the same disease with different method of screening susceptibility genes probe number1357(FC<2or FC<0.5, has annotated gene830) or susceptible gene probe number516(FC<3or FC <1/3,316genes have been annotated).2genes, namely low density lipoprotein receptor related protein12(LRP12) and Kruppel like factor5(KLF5), were selected from the susceptible genes. In23cases of the crowd (14cases of blood stasis syndrome group, and9cases of healthy adults) were analyzed the methylation status of the selected genes.The results show that between blood stasis syndrome and healthy people, no difference in methylation status of two genes. From the point of view of coronary heart disease with blood stasis group, promoter methylation status between different individuals of these two genes exist difference, there were methylated, not fully methylated and non-methylated three states.Conclusion:1. The difference of curative effect of Xuefu Zhuyu Decoction in treating coronary heart disease with blood stasis and the objective results of blood stasis syndrome in the research, suggested the heterogeneity of syndrome essence.2. There exist differences in characterization of coronary heart disease with blood stasis. The differences may be related to hereditary factors, prompted that coronary heart disease with blood stasis syndrome essence may be genetic heterogeneity.3. Blood stasis syndrome pedigrees patients with non home show differences gene expression spectrum results, coronary heart disease with blood stasis syndrome essence may be genetic heterogeneity.4. Differences in outcome suggest that blood stasis syndrome in coronary heart disease susceptibility gene promoter methylation results, the nature of heterogeneity and scale heterogeneity in genetics might be related to coronary heart disease with blood stasis syndrome. In conclusion, from the results of view, coronary heart disease with blood stasis syndrome essence exists heterogeneity. This heterogeneity generated may be related to genetic and epigenetic and a series of control network system.However, the nature of heterogeneity may also reflect from the characterization of the differences. |
