Functional characterisation of polymorphisms in candidate genes for coronary heart disease

• Coronary heart disease (CHD) is a multifactorial disorder. Several important risk factors, in particular cigarette smoking, hypertension, hypercholesterolaemia, reduced high density lipoprotein (HDL) cholesterol concentration and hyperglycaemia, have been defined. It appears that most risk factors are influenced by genetic components, suggesting that genetic variants in candidate genes play an important role in the development of CHD. In view of the importance of transcriptional regulation in protein synthesis, it seems appropriate to analyse the promoter region of candidate genes for mutations affecting the basal rate of transcription. The objective of this thesis was therefore to analyse the physiological significance of common polymorphisms in the promoters of candidate genes for CHD. The proximal promoters of the genes for ß-fibrinogen, coagulation factor VII (FVII), apolipoprotein (apo) B, hepatic lipase (HL) and angiotensinogen (AGT) were sequenced in both directions, and in each of these genes one or more common polymorphisms were discovered. Electromobility shift assays were used to determine whether the mutations introduced a change in the binding pattern of nuclear factors. Significant changes were observed for the -455G/A and -854G/A polymorphisms of the ß-fibrinogen gene, the -401G/T and -402G/A polymorphisms of the FVII gene, the -516C/T polymorphism of the apo B gene, the -71OT/C polymorphism of the HL gene, and the -829T/A polymorphism of the AGT gene. The nature of these polymorphisms were subsequently analysed in transient transfection studies in HepG2 cells. It was found that all polymorphisms, with the exception of the -71OT/C polymorphism in the HL gene, influence the basal rate of transcription of the respective genes in vitro. The physiological significance of the promoter polymorphisms in the genes for ß-fibrinogen, FVII, apo B, and AGT was further evaluated in association studies in apparently healthy, middle-aged men. Both the -455G/A and the -854G/A polymorphisms of the ß-fibrinogen gene were associated with an increased plasma fibrinogen concentration. The -40IG/T polymorphism of the FVII gene was related to reduced plasma concentrations of total FVII and fully activated FVII molecules, while the -402G/A polymorphism was associated with increased plasma FVII levels. The -516C/T polymorphism in the apo B gene was associated with an increased level of low density lipoprotein (LDL) cholesterol in plasma. The rare allele of the -829T/A polymorphism of the AGT gene was related to increases in both systolic and diastolic blood pressure. It is concluded that the -455G/A and -854G/A polymorphisms of the p41brinogen gene, the -401G/T and -402G/A polymorphisms of the FVII gene, the -516C/T polymorphism of the apo B gene, and the -829T/A polymorphism of the AGT gene are physiologically relevant mutations with a significant impact on the plasma concentrations of fibrinogen, FVII and LDL cholesterol and blood pressure, respectively, in healthy middle-aged men.

Keyword

fibrinogen, coagulation factor VII, apolipoprotein B, hepatic lipase, angiotensinogen, atherosclerosis, low density lipoprotein, high density lipoprotein, blood pressure

Publication and Content Type

vet (subject category)

dok (subject category)