Role Of CYP1A1 MspIgene Polymorphism On Cigarette Smoking With Coronary Artery Disease In Chinese

BackgroundCigarette smoking is the most important enviromental contributor of the coronary artery disease.CYPlAl is a member of a large class of cytochrome P-450 enzyme. These enzymes play a significant role in the detoxification of polycycclic aromatic hydrocarbons generated from the combusion of fossil fuels and aromeatic amines present in cigaratte smoke. CYP1A1 encodes a phase I cytochrome P-450 enzyme which is one of the key toxic components produced during cigaratte smoke. A highly inducible form of the enzyme may be associated with an increased risk of arterosclerosis in smokers.However, molecular mechanisms responsible for the high inducibility have not been resolved.Recently a CYPIA1 MspI at the 3'-flanking region (T6235C), closely linked to the IIe→Val mutation in exon 7, and has been reported in many epidemiological studies to be associated with cigarette smoking related coronary artery disease risk but not all studies. Therefore,DNA sequence variant in the CYPIAI contributing tointer-individual susceptibility to cigarette smoking induced arterosclerosis changes. In the present, we explored the hypothesis that the CYP1A1 Msplgene polymorphism may contribute to cigarette smoking related risk of the coronary artery disease.Our purpose is to known the distribution of CYP1A1 Msplgene polymorphism in Chinese and the relationship with CAD.Meterial and MethodsSubjects:453 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, 210 CAD patients (143male and 67 female, the average age is 61.9±9.2) at least 50% obstruction of one major coronary vessel. The lifetime smoking dose was measured smoking index (SI),which calculated by the number of cigarettes that smoked daily,multiplied by the number of years of cintinous smoking ( SI= cigarettes per dailyxyears ). We also classified our patients into no smokers, light smokers(1 SI 400)and heavy smokers ( Sl>400).243 control subjects(165 male and 78 female, the average age is 63.6±6.4) were from patients with normal angiography. 1. 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 CYP1A1 genotype was determined by PCR-RFLP.(1) The primers of Wang XI and BadenhopR are referenced,the forward primer: 5' -CAG TGA AGA GGT GTA GCC GCT-3'; and the reverse primer:5' -TAG GAG TCT TGT CTC ATG CCT-3' .(2)PCR was carried out using a 25ul reaction mix containing 10pmol of each primer ,100ng of genomic DNA,200uM of each dNTP,2.5ul of 10 X buffer,l~3uM of MgCl2,2U of Taq DNA poIymerase.The PCR cycles were modified as follows:5 min initial at 94℃ .followed by 30 cycles of 60 sec of denaturationat 94℃,60 sec of annealing at 53℃ and 60 sec of extension at 72℃ followed by a final 5 min extension step at 72℃.(3) The PCR products were then digested by Mspl restriction enzyme at 37 ℃ for 2 h, separated by electrophoresis in 2% agarose gel for 40 min and stained with ethidium bromide. 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 between groups were analysed by the chi-squared test. Statistical significance was taken as p<0.05.Results1.Prevalence of clinical features in patients and controls:There were no differences in majority clinical features between the groups(P>0.05).Only smoking and hyperlipomia history were significantly more common than in the controls(P<0.05).2.Prevalence of CYP1A1 in all subjects :C,T allele and genotype of CC,CT TT genotype can be found in CAD and control group. The fragment length of all...