Association Of Fâ…« C46T Polymorphism With Coronary Artery Disease In Chinese

BackgroundCoronary artery disease (CAD) is associated intensely with the coagulation system. The disorder of blood coagulation are often found in CAD patients. Coagulation factors and antifibrinolytic mass are important in atherosclerosis. The rupture of atheromatous plaque and thrombosis is the major cause of acute myocardial infarction and unstable angina.Cogulation factor XII (FXII: Hageman factor ) is an 80-kDa serine ptorease circulating at an average concentration of 30ug/ml. In vitro, FXI1 plays a central role in the initiation of coagulation and fibrinolysis, but its physiological role is still under discuss. Usually, no increased bleeding tendency is observed in patients with a"severe deficiency of FXII. In contrast, severe as well as mild factor XII deficiency have even been associated with an increased risk for venous and arterial thrombolism. But Miller, Kohler found high FXIIa level is a risk factor to coronary artery disease.Recently, a C46T polymorphism in the promoter region of the FXII gene at nt46 was identified. The polymorphism destroys a Kozak consensus sequence, resulting in a lower translation efficiency and a decrease in FXII plasma level. This polymorphism is frequent in Oriental and may explain the lower plasma FXII activity found in this population. The role of this polymorphism for the development and progression of coronary artery disease is still under discussion. We hypothesize the low incidence of coronary artery disease in oriental because of the high incidence of this polymorphism.We investigated the FXII C46T polymorphism using mutagenically separated polymerase chain reaction assay ( MSPCR ) technique. Our purpose is to known the distribution of FXII C46T in Chinese people and the relationship with CAD.Meterial and Methods:1. Subjects:335 Han ethnic persons without blood relationship were recruited to the study. All of the person were performed selected coronary angiography and from the Cardiovascular Department of the Second Affiliated Hospital, Medical College of Zhejiang University. Among them, 150 CAD patients (118male and 32 female, the average age is 63.9±9.4) at least 50% obstruction of one major coronary vessel. The CAD group is divided into acute coronary syndrome subgroup (n=108) and no acute coronary syndrome subgroup(n=42) using criteria defined by the World Health Organisation(WHO);185 control subjects(121 male and 64 female, the average age is 62.3±9.4) were from patients with normal angiography.2. Methods:2.1 Collection of clinical features: All subjects were enquired in detail for history of hypertention diabets hyperlipemia smoking. The stature weight and major biochemical data were also determinated.2.2 Isolation of DNA. Cells were collected from EDTA anticoagulated blood and stored at -72℃, pending extraction by the salting out method.2.3 FXI1 C46T genotype was determined by MSPCR.(l) The primers of Georg Endler are referenced, the forward primer:46C 5'-GGC AGC TGG ACC AAC GGA CCG AC-3\ 46T 5'-TCC TGG ATA AAC AGC TGG ACC AAC GGA CGA AT-3' and the reverse primer: 5' - GAA CAA TCC TGG GAC AAT CCT GGT -3' . (2) PCR was carried out using a 25ul reaction mix containing 4pmol of primer 46C, 6pmol of primer 46T , 5pmol of reverse primer, 100ng of genomic DNA, 200uM of each dNTPs, 2.5ul of 10 X buffer, 1.5mM of MgCI2, 0.75U of Taq DNA polymerase. The PCR cycles were modified as follows: 5 min initial at 95℃,followed by 35 cycles of 30 sec of denaturationat 95℃,60 sec of annealing at 55℃ and 30 sec of extension at 72℃ followed by a final 10 min extension step at 72℃. (4)Electrophoresis: PCR products were loaded onto a 12% polyacrylamide gel(29:l) containing 5% glycerol for electrophoresis. Gels were visualized by silver nitrate staining and analyzed with macrography.3. Statistical analysis:Using the SPSS 10.0 for windows statistical package. Count data were performed by Student's t-test and measure data were performed by chi-squared test. Genotype and allele frequencies betwe...