A Study Of The Correlation Between The CYP2C9 Gene Polymorphism And Coronary Heart Disease As Well As Plasma Lipid Levels

The coronary heart disease(CHD) is an epidemic disease in people which is caused by many factors such as some incorrect living habits,environment and heredity...etc.And it is also a kind of many gene hereditary disease.Studying the hereditary gene of CHD has important academic meaning for developing the purposeful crowd prevention and deferring the occurrence of CHD.With the quick development of molecular biology and carrying on of the Mankind Genome Plan, looking for CHD correlative genes and illustrating CHD outbreak mechanism on the gene level have becomed a medical science research focus.These genes include the fat-metabolized genes such as apolipoprotein E gene,apolipoprotein A gene,and apolipoprotein B gene...etc,and include angiotensinâ… -coverting enzyme(ACE) gene and angiotensinâ…¡gene which both belong to renin-angiontensin-aldosterone system(RASS) genes.The susceptible genes include also the plas minogen activator inhibitor-1(PAI-1) gene,tissue inhibitors metalloproteinases(TIMP) gene,endothelial nitric oxide synthase(Enos) gene and E-selectin gene...etc.The relationship between CHD and gene is often reserched on polymorphism of CHD correlative genes.Human cytochrome P450 enzymes are an important enzyme family.This family plays a significant role in metabolizing a wide range of drugs in human liver,and also plays an important role in the metabolism of endogenous and extraneous substance. The CYP protein is coded by CYP super-gene family.The CYP enzyme family includes a lot of family and sub-family.CYP2C9 is one of the CYP2C sub-family. The MW of the CYP2C9 protein is 55 kD,consist of about amino acid residues.The protein is mainly expressed in liver,and in small in testine,respiratory organs,brain, artery vessel and kidney.The CYP2C9 protein is coded by CYP2C9 gene.The human CYP2C9 gene is located at chromosomal 24 region 10q,spanning approximately 55kb with nine exons and eight introns.There has been found polymorphism called single nucleotide polymorphism(SNP) in molecular biology in the open reading region of the gene. The allele variants include CYP2C9^*1(wilde type),CYP2C9^*2,and from CYP2C9^*3 to CYP2C9^*30.The distribution of allele variants is different in different populations. The CYP2C9^*3 mutant genotype comes into being by the mutation of A to C at 1075 base spot.This mutation induces the changement from ATT codon which codes isoleucine to CTT codon which codes leucine.The CYP2C9 enzyme plays important roles in metabolism of many drugs such as phenytoin,warfarin,tolbutamide etc.The drugs which the CYP2C9 enzyme is involved to metabolize account for about 12%of those drugs that are commonly used in clinic.The enzyme is also involved in the metabolism of a variety of external carcinogenic and mutagenic substances,such as quinoline,butadiene.In addition to exogenous substances,the CYP2C9 also plays an important role in the metabolism that arachidonic acid is epoxyoxidated into epoxyeicosatrienoic acid(EET) in artery vessel cells.EET are known as a kind of endothelium-derived hyperpolarizing factors (EDHFs).In vascular smooth muscle cells,EET can promote the inflow of Ca²⁺,and beable to activate Ca²⁺ ion-dependent K⁺-Ca²⁺ ion channels to cause hyperpolarization that causes blood vessels to expand in order to lower blood pressure. And it also has an impact on the function of vascular endothelial cells and smooth muscle cells.Lipids is an important endogenous substance which is synthesized and metabolized by the liver,and its metabolity has possibly contacts with the CYP2C9 enzyme. CYP2C9 genetic polymorphism can lead to the individual changes in the structure of CYP2C9 enzyme,and the structure changes can cause changes in protein activity, Researches have shown that the mutant protein activity is lower than that of wild-type protein.Changes in protein activity results in polymorphism of the ability of some drugs metabolism,and the drug metabolism capacity in the individual carrying the mutant gene is often lower than that of the individual carrying wild-type gene.At the same time,this polymorphism of the body not only causes difference in metabolic capacity on external carcinogens and mutagens metabolism,but also causes difference of the body's ability to generate EET.The mutant individual of CYP2C9^*3 genotype has lower EET-synthesized capacity than the wild-type individual of CYP2C9^*1. Through the occurrence and development of some diseases will differently be influenced by the genetic polymorphism of the CYP2C9 gene.This genetic polymorphism often has relationship with the occurrence of some diseases,such as lung cancer and acute myocardial infarction.Objective1.Explore if there is relati6nship between the CYP2C9 gene polymorphism and coronary heart disease.2.Explore if there is relationship between the CYP2C9 gene polymorphism and the plasma lipid level and what the relationship is.Subjects and methods1.SubjectsAll subjects were inpatients in Guangzhou General Hospital of Guangzhou Military Area Command who were given coronary angiography(CAG) to.Those subjects in who the coronary artery stenosis rate is more than 50 percent or who have a clear history of myocatrdial infarction were classified into coronary heart disease group,but those subjects in who coronary angiography displayed no coronary artery stenosis into a control group.All objects are individuals of the Han nationality,and there is not kinship between all individuals.And those who sufferred from liver, kidney,tumor,thyroid and diabetes diseases were excluded.There were 218 cases in the CHD group and 200 cases in the control group.By statistical analysis,the age,sex rate,smoking rate,hypertension morbidity and other conditions in two groups were no significant difference.2.methodsBefore taking lipid-lowering drugs,Subjects after fasting more than eight hours were taken venous blood from.Total cholesterol(TC),triglyceride(TG),low-density lipoprotein cholesterol(LDL-C) and high-density lipoprotein cholesterol(HDL-C) were detected by standard blood enzyme colorimetric,and apolipoprotein A1(apoA1) and apolipoprotein B(apoB) and lipoprotein(a) were measured by immune turbidimetry.Another 5ml venous blood were taken to extract leukocyte genomic DNA from.by ways provided by the blood DNA extraction kit.The polymerase chain reaction(PCR) method was taken to amplify the CYP2C9 gene fragments.PCR products were sent to Shanghai Yingjun Biotechnology Co.Ltd to carry out direct sequencing to difine genotypes.The plasma lipid in all the subjects with the CYP2C9^*3 genotype in all 418 subjects was compared with that in the same number of subjects with the CYP2C9^*1 genotype selected from all 418 subjects.Two Independent-Samples T Test was used to analyze the distinction of age between CHD group and control group,and test was employed to compare the baseline information such as the percentage of men,smoking rates and the incidence of hypertension in two groups.Chi-Square test was employed to compare CYP2C9^*3 genotype frequencies and allele frequencies in two groups,and Two Independent-Samples T Test was used to analyze the distinction of plasma lipid levels between these subjects with CYP2C9^*3 genotype and these subjects with CYP2C9^*1 genotype.A Probability value less than 0.05 was considered statistically significant.Results1.All CYP2C9^*3 mutant genotypes detected in all the subjects were all CYP2C9^*1/ CYP2C9^*3 heterozygote,and CYP2C9^*3/CYP2C9^*3 mutant homozygote wasn't found.2.The percentage of CYP2C9^*3 genotype in CHD group was 8.26%,higher than the percentage of 3.50%in control group,and the difference was statistically significant (P=0.040),allele frequency of CYP2C9^*3(4.13%) in CHD group was also significantly higher than that(1.75%) in control group(P =0.044).3.Plasma lipid and lipoprotein levels between these subjects with CYP2C9^*3 genotype(mutant type) and these subjects with CYP2C9^*1 genotype(wild type) were no statistically significant difference.Conclusion1.In Hart Chinese,CYP2C9^*3 mutant genotype is mainly CYP2C9^*1/CYP2C9^*3 heterozygote.2.The mutant frequency of CYP2C9^*3 genotype in CHD group was higher than that in control group,and the difference between them was statistically significant (P=0.040).Allele frequency of CYP2C9^*3 in CHD group was also significantly higher than that in control group.So these results suggest that CYP2C9 gene polymorphism is possibly in association with the occurrence and development of CHD and CYP2C9^*3 mutation increases the risk of CHD.3.This study did not find CYP2C9 gene polymorphism was significantly correlative with plasma lipid.And though the trend that LDL-C and apoB levels in these subjects with CYP2C9^*3 genotype are higher than that in these subjects with CYP2C9^*1 genotype was seen,it is necessary to increase subjects in order to observe the trend.