| The incidence of diabetic coronary heart disease(DCHD)is increasing year by year,and clinical observation shows that patients with DCHD usually have a worse prognosis than those with coronary heart disease(CHD)alone,and are associated with a greater degree of blood stasis.Therefore,it is important to actively explore the plasma metabolic characteristics of patients with diabetic coronary heart disease with blood stasis,in order to understand the intrinsic connection between DCHD and blood stasis and to identify the targets for disease treatment.Metabolomics technology can accurately characterize or quantify small molecule metabolites and analyze the data using statistical method,which has obvious advantages in finding biomarkers,early diagnosis and determining the direction of disease development,and has been widely used in research related to biomarkers of combining with disease and syndrome.In this study,we used a combination of non-targeted and targeted metabolomics techniques to establish a clinical discovery and validation cohort with diabetic coronary artery disease as the entry point,to screen for biomarkers of combining with disease and syndrome,and help to explain the mechanisms of "blood stasis" exacerbated by diabetes mellitus promoting the progression of coronary vascular lesions.Part I Exploration of plasma metabolic characteristics in patients with diabetic coronary heart disease with blood stasis evidenceObjective:To explore plasma differential metabolites in patients with DCHD blood stasis evidence using non-targeted metabolomics techniques.Methods:91 patients with coronary artery disease from Xiyuan Hospital were consecutively included in the plasma non-targeted metabolomics study and grouped in the following three ways:47 patients with DCHD and 44 patients with CHD alone,according to the grouping of simple CHD and CHD combined with diabetes mellitus;grouped according to the different degrees of blood stasis in different diseases:①34 patients with severe blood stasis in the DCHD group and 13 patients with mild blood stasis;②22 patients with severe blood stasis in the CHD group and 22 patients with mild blood stasis.Using liquid chromatography-mass spectrometry,the characteristic plasma differential metabolites of DCHD and severe blood stasis symptoms were screened by principal component analysis,partial least squares analysis,receiver operating characteristic(ROC)curve plotting and metabolic pathway enrichment analysis.Results:①31 different metabolites were found between the DCHD and CHD groups.1,5-anhydro-d-sorbitol,L-fucose,D-lysose,and Beta-hydroxybutyrate were among the more significant differential metabolites in the DCHD group,and the more significantly enriched pathways were fructose and mannose metabolism,amino sugar and nucleotide sugar metabolism and bile secretion.②There were 19 differential metabolites between the CHD-Z and CHD-Q groups,and the metabolites with significant differences were mainly hippuric acid,Pro-Ser,Dl-a-hydroxybutyric acid,Deoxycholic acid,and Propionylglycine,and the metabolic pathways with more significant enrichment were biosynthesis of cofactors,bile secretion,2-oxocarboxylic acid metabolism,biosynthesis of amino acids,and caffeine metabolism.③The DCHD-Z group showed 20 significantly different metabolites compared with the DCHD-Q group,and the metabolites with more significant differences were 1,2dioleoyl-sn-glycero-3-phospho-(1-myo-inositol-3,4-bisphosphate),O-Acetyl-L-serine,4-phosphonobutyric acid,and PC(16:1(9Z)/16:1(9Z)),and the differential metabolites were significantly enriched in the biosynthesis of cofactors,biosynthesis of amino acids,and carbon metabolism pathways.Conclusion:L-fucose,1,5-anhydro-d-sorbitol,L-fucose-1-phosphate and Betahydroxybutyrate were significantly different between plasma metabolism groups of patients with DCHD and patients with CHD;whereas 4-phosphonobutyric acid,OAcetyl-L-serine,1,2-dioleoyl-sn-glycero-3-phospho-(1-myo-inositol-3,4-bisphosphate)and PC(16:1(9Z)/16:1(9Z))differed significantly between different degrees of blood stasis in DCHD.The pathways of amino acid metabolic may play an important role in both CHD progression and changes in the degree of blood stasis.Part II Validation of plasma differential metabolites in patients with diabetic coronary heart disease with blood stasis syndromeObjective:To targeted test the characteristic plasma differential metabolites in patients with blood stasis evidence of DCHD.Methods:Targeted metabolomic validation was performed for the eight plasma significantly different metabolites obtained in the first part of the screening,and the relationship between the validated significantly different metabolites and clinical laboratory testing indicators was analyzed using Pearson correlation analysis.A total of 79 patients with CHD were included and grouped in the same way as in Part I:43 patients with DCHD and 36 patients with CHD alone were grouped differently by disease;26 patients with severe blood stasis and 10 patients with mild blood stasis were grouped differently by degree of blood stasis in DCHD;20 patients with severe blood stasis and 23 patients with mild blood stasis in the CHD group.Results:Compared with the CHD group,the levels of four metabolites,1.5anhydro-d-sorbitol,D-lysose,L-fucose,and Beta-hydroxybutyrate,were significantly higher in the plasma of the DCHD group(P<0.01);compared with the DCHD-Q group,the levels of L-fucose were significantly higher in the DCHD-Z group(P<0.05)and the levels of Lithocholic acid were significantly lower(P<0.05).A highly significant positive correlation was found between 1,5-anhydro-dsorbitol and glycosylated hemoglobin type AlC(HbAlc)(r=0.814,P<0.01),fasting plasma glucose(FPG)(r=0.606,P<0.01),total cholesterol(TC)(r=0.430,P<0.01),and low-density lipoprotein(LDL)(r=0.457,P<0.01)in the DCHD group;D-lyxose had a highly significant positive correlation with HbAlc(r=0.685,P<0.01),FPG(r=0.545,P<0.01)and a significant positive correlation with TC(r=0.355,P<0.05)and LDL(r=0.398,P<0.05);L-fucose had a significant positive correlation with HbAlc(r=0.840,P<0.01),FPG(r=0.695,P<0.01),TC(r=0.497,P<0.01),and LDL-C(r=0.518,P<0.01),and triglycerides(TG)(r=0.371,P<0.05).Beta-hydroxybutyrate showed a highly significant positive correlation with FPG(r=0.487,P<0.01)and HbAlc(r=0.622,P<0.01).In the DCHD severe blood stasis group,L-fucose content showed a significant positive correlation with FPG(r=0.724,P<0.01),HbAlc(r=0.868,P<0.01),TC(r=0.462,P<0.05),and LDL-C(r=0.499.P<0.01):no correlation was found between Lithocholic acid and clinical laboratory testing indicators.Conclusion:1,5-anhydro-d-sorbitol,D-lyxose,L-fucose,and Betahydroxybutyrate are plasma-specific metabolites in patients with DCHD,and L-fucose and Lithocholic acid are closely related to the severity of DCHD with blood stasis syndrome;except for Lithocholic acid,all of the above differential metabolites showed significant positive correlation with HbAlc. |
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