| BackgroundWith the rise and development of multi-component research technology,genomics,proteomics and metabonomics are more and more used in the molecular feature,classification and standardization of Zhenghou(TCM syndrome).Zhenghou is a certain pathological stage of the body in the development of the disease,including the location,cause and nature of the lesion,as well as the relationship between anti-pathogenic factors and pathogenic factors,which could reflect the essential pathological changes at a certain stage in the development of the disease.Therefore,it reveals the essence of the disease more comprehensively rather than the symptoms,and is the main way of TCM to understand the disease.In the biomedicine,the concept "phenotype" is the summation of the characters and characteristics with the interaction between genotype and environment,which is shown as a system with multi-gene interacting complexity and dynamic evolution of pathogenism.And a group of complex phenotypes is significantly similar to the concept Zhenghou in TCM.At present,the research on the relationship between Zhenghou and its omic profiles are mostly qualitative,or even semi-quantitative.There are few studies on the quantitative relationship between Zhenghou and its corresponding gene coexpressed network.Using the method of genotype-phenotypic quantitative analysis,the molecular feature of Zhenghou could be uncovered.In this study,taking chronic stable angina with Xueyu Zheng treated by Danhong injection as an example,the molecular mechanism of chronic stable angina with Xueyu Zheng were uncovered by the quantitative correlation method between Zhenghou and its corresponding gene expressed levels.ObjectiveQuantitative correlation analysis was used to analyze the correlation between the phenotypic changes of Xueyu Zheng in chronic stable angina treated with Danhong injection and its corresponding mRNA molecular,and to explore the molecular mechanism and potential biomarkers of Danhong injection for the treatment of Xueyu Zheng in chronic stable angina,so as to provide new strategy for the diagnosis and treatment of Zhenghou.MethodsSpearman correlation method was used to calculate the correlation between gene expression and symptoms in patients with chronic stable angina pectoris before and after treatmented by Danhong injection(0 days,14 days,30 days),and the genes related to symptoms of Xueyu Zheng were screened out.The screening condition was that the absolute value of correlation coefficient was more than 0.4 and P<0.05.The functional enrichment of related genes of six symptoms in Xueyu Zheng under different intervention time points of Danhong injection was performed with Metascape(https://metascape.org/).After comparing the pathways and GO functions of related genes,the molecular mechanism of Danhong injection in the treatment of the symptoms of Xueyu Zheng in chronic stable angina would be explored.The gene co-expression networks of Danghong injection at 14 days and 30 days were established by WGCNA,and compared with the baseline of the control group and the treatment group respectively,the targeted modules of Danhong injection for chronic stable angina at 14 days and 30 days were determined,and the correlation coefficients(r value)between the targeted modules and Xueyu Zheng at two time points were calculated by canonical correlation analysis,and the.targeted modules of Danhong injection for Xueyu zheng in chronic stable angina at two time points were found out.The screening condition was that the absolute r value was greater than 0.5.The functional enrichment analysis was carried out on the to explore the molecular mechanism of Danhong injection in the treatment of Xueyu Zheng in chronic stable angina.The genes in the targeted modules of Danhong injection for Xueyu Zheng in chronic stable angina at two time points with high r value,which were greater than 0.8,were screening for the protential biomakers of Danhong injection for the treatment of chronic stable angina pectoris with Xueyu Zheng based on its normalized degree centrality and r value to Xueyu Zheng.Results1.The genes related to the symptoms in chronic stable angina with Xueyu Zheng on the 0th,14th and 30th day were as follows:the number of genes related to chest pain was 77 in 0 days,60 in 14 days and 1726 in 30 days.The number of genes related chest tightness was 34 in 0 days,192 in 14 days and 82 in 30 days.The number of genes related to palpitation restlessness:155 in 0 days,47 in 14 days and 341 in 30 days.The number of genes related to dark purple lip was 1617 in 0 days,1112 in 14 days and 757 in 30 days.The number of genes related to dark tongue was 333,220,30 days in 0 days,14 days and 30 days respectively.The number of genes related to fine and unsmooth pulse was 1007 in 0 days,812 in 14 days and 684 in 30 days.2.Functional enrichment analysis of genes associated with symptoms of blood stasis syndrome showed that:(1)For chest pain,5 KEGG pathways and 97 GO functions were enriched at Day 0,mainly with the molecular functions as transport and metabolism,41 GO functions enriched at Day 14,mainly with the molecular functions as transport and cell organization and biogenesis,as well as 66 KEGG pathways and 1482 GO functions enriched at Day 30,mainly with the molecular functions as signal transduction,metabolism and immune system.(2)For Chest tightness,2 GO functions were enriched at Day 0,mainly with the molecular functions as cell and cell organization and biogenesis,16 KEGG pathways and 230 GO functions enriched at Day 14,mainly with the molecular functions as immune system,metabolism and transport,as well as 2 KEGG pathways and 60 GO functions enriched at Day 30,mainly with the molecular functions as cell cycle and cell organization and biogenesis.(3)For Palpitations,9 KEGG pathways and 156 GO functions were enriched at Day 0,mainly with the molecular functions as metabolism and cell communication,2 GO functions enriched at Day 14,mainly with the molecular functions as signal transduction and cell communication,as well as 19 KEGG pathways and 139 GO functions enriched at Day 30,mainly with the molecular functions as endocrine system,nervous system and metabolism.(4)For dark purple lip,73 KEGG pathways and 909 GO functions were enriched at Day 0,mainly with the molecular functions as signal transduction,immune system and metabolism,18 KEGG pathways and 424 GO functions enriched at Day 14,mainly with the molecular functions as metabolism,as well as 15 KEGG pathways and 265 GO functions enriched at Day 30,mainly with the molecular functions as metabolism.(5)For dark tongue,9 KEGG pathways and 249 GO functions were enriched at Day 0,mainly with the molecular functions as cancer,nervous system and development,5 KEGG pathways and 149 GO functions enriched at Day 14,mainly with the molecular functions as cardiovascular diseases and metabolism,as well as 25 KEGG pathways and 542 GO functions enriched at Day 30,mainly with the molecular functions as genetic information processing and metabolism.(6)For fine and unsmooth pulse,53 KEGG pathways and 471 GO functions were enriched at Day 0,mainly with the molecular functions as signal transduction,cancer,immune system and metabolism,14 KEGG pathways and 230 GO functions enriched at Day 14,mainly with the molecular functions as metabolism,as well as 3 KEGG pathways and 191 GO functions enriched at Day 30,mainly with the molecular functions as metabolism and development.3.Based on the analysis of the relationship between symptoms at the same time by KEGG pathway and GO function analysis,it was found that there was a high correlation among "dark purple lips","dark tongue" and "fine and unsmooth pulse"。"dark purple lips" and" fine and unsmooth pulse" have 299 common genes,52 GO functions and 1 KEGG pathway at Day 14 and 100 common genes,15 GO functions at Day 30,mainly with metabolism,transport and development,especially.The main symptoms "chest pain" and "chest tightness" only have 3 common GO functions,without common genes at Day 14.However,there were 35 common genes and 23 common GO functions at Day 30 of the two main symptoms."Chest tightness"had a high correlation with "dark purple lips","dark tongue" and " fine and unsmooth pulse"at Day 14,mainly with metabolism,cell differentiation,and signal transduction,while "chest pain" has a high correlation with these three signs metioned above at Day 30,mainly with transport,metabolism and signal transduction.4.Compared with Danhong group at Day 0,the gene co-expression network of Danhong group at Day 14 was divided into 99 modules and 75 differential modules were detected.Compared with the control group at Day 14,the gene co-expression network of the Danhong group at Day 14 were divided into 90 modules and 70 differential modules were detected.And then,5 target modules of Danhong for chronic stable angina at Day 14 were determined.Compared with the Danhong group at Day 0,the gene co-expression network of the Danhong group at Day 30 was divided into 122 modules and 101 differential modules were detected.Compared with the control group at Day 30,the gene co-expression network of Danhong group at Day 30 were divided into 126 modules and 87 differential modules were detected.Therefore,5 target modules of Danhong for chronic stable angina at Day 30 were determined.And then,7 target modules of Danhong for Xueyu Zheng in chronic stable angina were identified by CCA analysis.5.The 14-day target module enriched 6 GO functions and 1 KEGG pathway(RAS signaling pathway),and the 30-day target module enriched 12 GO functions.After classifying the function of GO,it was found that the target module at Day 14 was mainly related to binding,enzyme regulator activity and cell proliferation.The target module at Day 30 was mainly related to transporter activity,cell organization and biogenesis,cell differentiation and cell proliferation.6.According to the normalized degree centrality and r value to Xueyu Zheng of the genes in the high relevant target modules of Xueyu Zheng at two time points for network analysis with the absolute r value greater than 0.8,potential target genes,MIR1183,LOC643355,and SKINTL,for Danhong injection for Xueyu Zheng in chronic stable angina were screened out.with both normalized degree centrality and r value to Xueyu Zheng greater than 0.75.Conclusion1.The symptoms of Xueyu Zheng in chronic stable angina are related to cancer,metabolism,signal transduction,and immune function before drug intervention.At Day 14,the main molecular mechasim of Danhong injection on the symptoms in Xueyu Zheng is mainly related to immune system,metabolism,signal transduction,and genetic information processing,and at day 30,the main molecular functions were turned to correlated with metabolism and signal transduction.2.Under the intervention of Danhong injection,the "chest tightness","dark purple lip","dark tongue" and "fine and unsmooth pulse" are closely related at Day 14,mainly related to some metabolism,signal transduction and cell differentiation;by the 30th day,"chest pain" turned to be is closely related to the 3 signs mentioned above,and related functions involved metabolism,transportation,and signal transduction.3.The main mechanism of Danhong injection in treating Xueyu Zheng in chronic stable angina is related to cell proliferation.At Day 14,it was also associated with RAS signaling pathways,binding,calcium ion binding,and enzyme-regulated activity;At Day 30,it was associated with transport activities,cellular tissue and biogenesis,and cell differentiation.4.The potential target genes of Danhong injection for chronic stable angina pectoris are MIR1183,LOC643355 and SKINTL. |
PDF Full Text Request