The angiotensin-converting enzyme insertion/deletion polymorphism and its associations with carotid artery wall thickness and coronary heart disease: The atherosclerosis risk in communities study

Coronary heart disease (CHD) is the leading cause of death in the United States. Recently, renin-angiotensin system (RAS) was found associated with atherosclerosis formation, with angiotensin II inducing vascular smooth muscle cell growth and migration, platelet activation and aggregation, and stimulation of plasminogen activator inhibitor-1. Angiotensin II is converted from angiotensin I by angiotensin I-converting enzyme (ACE) and this enzyme is mainly genetically determined. The ACE gene has been assigned to chromosome 17q23 and an insertion/deletion (I/D)polymorphism has been characterized by the presence/absence of a 287 bp fragment in intron 16 of the gene. The two alleles form three genotypes, namely, DD, ID and II and the DD genotype has been linked to higher plasma ACE levels and cell ACE activity.^ In this study, the association between the ACE I/D polymorphism and carotid artery wall thickness measured by B-mode ultrasound was investigated in a biracial sample, and the association between the gene and incident CHD was investigated in whites and if the gene-CHD association in whites, if any, was due to the gene effect on atherosclerosis. The study participants are from the prospective Atherosclerosis Risk in Communities (ARIC) Study, including adults aged 45 to 65 years. The present dissertation used a matched case-control design for studying the associations of the ACE gene with carotid artery atherosclerosis and an unmatched case-control design for the association of the gene with CHD. A significant recessive effect of the D allele on carotid artery thickness was found in blacks (OR = 3.06, 95% C.I: 1.11-8.47, DD vs. ID and II) adjusting for age, gender, cigarette smoking, LDL-cholesterol and diabetes. No similar associations were found in whites. The ACE I/D polymorphism is significantly associated with coronary heart disease in whites, and while stratifying data by carotid artery wall thickness, the significant associations were only observed in thin-walled subgroups. Assuming a recessive effect of the D allele, odds ratio was 2.84 (95% C.I:1.17-6.90, DD vs. ID and II) and it was 2.30 (95% C.I:1.22-4.35, DD vs. ID vs. II) assuming a codominant effect of the D allele. No significant associations were observed while comparing thick-walled CHD cases with thin-walled controls. Following conclusions could be drawn: (1) The ACE I/D polymorphism is unlikely to confer appreciable increase in the risk of carotid atherosclerosis in US whites, but may increases the risk of carotid atherosclerosis in blacks. (2) ACE I/D polymorphism is a genetic risk factor for incident CHD in US whites and this effect is separate from the chronic process of atherosclerosis development. Finally, the associations observed here are not causal, since the I/D polymorphism is in an intron, where no ACE proteins are encoded. ^

MATHEMATICAL STUDIES ON THE EVOLUTIONARY CHANGE OF DNA SEQUENCES

With the aim of understanding the mechanism of molecular evolution, mathematical problems on the evolutionary change of DNA sequences are studied. The problems studied and the results obtained are as follows: (1) Estimation of evolutionary distance between nucleotide sequences. Studying the pattern of nucleotide substitution for the case of unequal substitution rates, a new mathematical formula for estimating the average number of nucleotide substitutions per site between two homologous DNA sequences is developed. It is shown that this formula has a wider applicability than currently available formulae. A statistical method for estimating the number of nucleotide changes due to deletion and insertion is also developed. (2) Biases of the estimates of nucleotide substitutions obtained by the restriction enzyme method. The deviation of the estimate of nucleotide substitutions obtained by the restriction enzyme method from the true value is investigated theoretically. It is shown that the amount of the deviation depends on the nucleotides in the recognition sequence of the restriction enzyme used, unequal rates of substitution among different nucleotides, and nucleotide frequences, but the primary factor is the unequal rates of nucleotide substitution. When many different kinds of enzymes are used, however, the amount of average deviation is generally small. (3) Distribution of restriction fragment lengths. To see the effect of undetectable restriction fragments and fragment differences on the estimate of nucleotide differences, the theoretical distribution of fragment lengths is studied. This distribution depends on the type of restriction enzymes used as well as on the relative frequencies of four nucleotides. It is shown that undetectability of small fragments or fragment differences gives a serious underestimate of nucleotide substitutions when the length-difference method of estimation is used, but the extent of underestimation is small when the site-difference method is used. (4) Evolutionary relationships of DNA sequences in finite populations. A mathematical theory on the expected evolutionary relationships among DNA sequences (nucleons) randomly chosen from the same or different populations is developed under the assumption that the evolutionary change of nucleons is determined solely by mutation and random genetic drift. . . . (Author's abstract exceeds stipulated maximum length. Discontinued here with permission of author). UMI ^

Assessing time-by-covariate interactions in Cox proportional hazards regression models using cubic spline functions

This dissertation develops and explores the methodology for the use of cubic spline functions in assessing time-by-covariate interactions in Cox proportional hazards regression models. These interactions indicate violations of the proportional hazards assumption of the Cox model. Use of cubic spline functions allows for the investigation of the shape of a possible covariate time-dependence without having to specify a particular functional form. Cubic spline functions yield both a graphical method and a formal test for the proportional hazards assumption as well as a test of the nonlinearity of the time-by-covariate interaction. Five existing methods for assessing violations of the proportional hazards assumption are reviewed and applied along with cubic splines to three well known two-sample datasets. An additional dataset with three covariates is used to explore the use of cubic spline functions in a more general setting. ^