| Part I Study on Biochemical Markers of Thrombosis in Coronary Heart DiseaseObjective There are close relationships between disfunctions of coagulation, anticoagulation, fibrinolysis as well as endothelium and development of coronary heart disease. Some studies found that during pre-infarction phases, a hypercoagulable state may be detectable. Single research referring to multiple thrombosis factors is rare. Thus, the main objective of this study was to evaluate the effects of multiple thrombosis factors as well as some risk factors on coronary heart disease. Methods 56 patients with coronary artery disease (including 26 stable angina pectoris and 30 acute coronary syndromes) admitted to department of cardiology in Wuhan Union Hospital and 54 control patients excluded thromboembolic disease, diabetes mellitus, hypertension, malignant tumor, acute/chronic liver or kidney disease and connective tissue disease were enrolled from January to July 2006. Coronary artery disease group and control group were matched for age, sex and sampling time. Plasma levels of protein C, free protein S, total protein S, thrombomodulin, activated factor VII, factor VII Antigen, P-selectin, tissue-type plasminogen activator, plasminogen activator inhibitor-1 were measured by enzyme linked immunosorbent assay, activity of tissue factor was measured by chromogenic activity assay, and activated protein C ratio, prothrombin time, activated partial thromboplastin time, fibrinogen, D-dimmer and thrombin time were detected by full-automated coagulation analyzer (Sysmex CA-7000, Japan). Levels of blood lipids were assayed by Department of Laboratory of Union Hospital. Results The levels of tissue factor activity in patients with acute coronary syndromes were found to be significantly higher than those in controls (p<0.01) or in stable angina patients (p<0.05); the levels of FVIIag in ACS patients were significantly higher than those of controls; compared with controls, the plasma levels of FVIIa (P<0.01) and TM (P<0.05) as well as prevalences of APC ratio below 2.4 (P<0.01) in ACS and SAP groups were higher, however, APC ratios were lower (P<0.01). The result of binary logistic regression analysis showed that activated factor VII (OR 2.680, 95%CI 1.539-4.665) and tissue factor activity (OR 1.019, 95%CI 1.004-1.035) were risk factors, and high density lipoprotein (OR 0.008, 95%CI 0-0.478) and activated protein C ratio (OR 0.001, 95%CI 0-0.011) were protective factors for coronary heart disease. Conclusion There are activated extrinsic coagulation, lower response to activated protein C and damaged or activated endothelium function in SAP and ACS patients, plasma TFc may be a predictor for onset of ACS, plasma FVIIa and TFc are correlated with CHD. Part II Study on Protein Markers of Thrombosis in Coronary Heart DiseaseObjective To detect the plasma proteomic patterns in acute myocardial infarction (AMI) patients and controls, screen specific biomarkers and build diagnostic models by Surface-enhanced laser desorption/ionization time-of-flight mass spectrometer (SELDI-TOF-MS) and weak cation exchange protein chip (CM10). Methods 77 patients with acute myocardial infarction admitted to department of cardiology in Wuhan Union Hospital, 60 healthy controls and 32 patients with acute cerebral infarction (ACI) were enrolled from January to July 2006. Acute myocardial infarction group and control groups were matched for age, sex and sampling time. Protein chips combined with human plasma were placed in SELDI-TOF-MS for detecting protein profiling, analysis of the total experiment data was implemented by the Zhejiang University Cancer Institute-ProteinChip Data Analysis System (ZUCI-PDAS) for finding out discrepancy protein and building diagnostic models. Results A pattern composed five protein peaks (m/z: 3321.6, 16099.3, 8058.7, 15933.0 and 16286.7, respectively) with a specificity of 95% and a sensitivity of 88.3% was selected based on their collective contribution to the optimal separation between patients with AMI and healthy controls; A pattern composed seventeen protein peaks (m/z: 6682.6, 6485.6, 3321.6, 16682.9, 16297.7, 7977.5, 3424.7, 4821.1, 11748.7, 4775.3, 8977.3, 9094.8, 4361.4, 11479.9, 2941.1, 11399.7 and 2462.3, respectively) with a total accuracy of 100% was selected based on their collective contribution to the optimal separation between patients with AMI and patients with ACI. Conclusion Plasma proteomic profiling with SELDI-TOF-MS and ProteinChip technologies provides high sensitivity and specificity in discriminating patients with AMI and controls, and the discovered protein peaks might show great potential for early diagnosis of AMI. Part III Study on Gene Markers of Thrombosis in Coronary Heart Disease by Multi-Analyte Suspension Array technologyObjective Coronary heart disease is caused by multiple factors, which also include genetic factors. The main objective of this study was to detect 7 gene polymorphisms in acute myocardial infarction (AMI) patients by liquid gene chip technology (MASA technology), and positive findings were verified, so as to find out validity of MASA technology in detecting gene variations. Methods 34 patients with acute myocardial infarction admitted to department of cardiology in Wuhan Union Hospital from January to July 2006 were enrolled in this trail. DNA was amplified by PCR technology after being extracted, nucleic acid probes of 7 gene variations (FV Leiden G/A mutation, Fbg Beta -455 G/A mutation, Fbg Beta -148 C/T mutation, Fbg Beta 448 G/A mutation, FVII -323 10bp deletion/insertion mutation, prothrombin 20210 G/A mutation and MTHFR 677 C/T mutation) were combined with microspheres (beads) by covalent bonds, the latter were coded with specified fluorescence, PCR products was added after various beads were mixed, the mixture was detected by Luminex 100? Multi-Analyte Suspension Array, finally, positive findings were verified by Sanger dideoxynucleotide chain termination. Results There were 19 patients with gene variations, including 4 patients with FV Leiden mutation, 14 patients with MTHFR 677 C/T mutation, 5 patients with fibrinogenβgene 448 G/A mutation, among them there were 1 patient with FV Leiden and Fbgβ448G/A mutation, 1 with FV Leiden and MTHFR677 C/T mutation, 2 with MTHFR677 C/T mutation and Fbgβ448G/A mutation, the total accuracy of MASA in positive findings was 73.9%. Conclusion Liquid gene chips technology is feasible in detecting gene variations in AMI patients, nevertheless, the MASA technology needs to improve.
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