The relationship between risk factor levels and presence of coronary artery calcification is dependent on apolipoprotein E genotype.

An important research question in the study of the genetics of coronary artery disease (CAD) is whether information about genetic variation will improve our ability to predict CAD beyond established risk factors. This question is especially relevant to the goal of identifying young, asymptomatic adults with coronary atherosclerosis who would benefit most from interventions to reduce risk. Coronary artery calcification (CAC) detected by electron-beam computed tomography is a relatively new method for detecting coronary atherosclerosis in asymptomatic individuals that has been shown to be a more accurate indicator of coronary atherosclerosis in asymptomatic individuals than other noninvasive techniques. In a study of asymptomatic women (n=169) and men (n=160) between the ages of 20 and 59 representative of the Rochester, Minnesota population, we used logistic regression to ask whether the most common Apolipoprotein (Apo) E genotypes (epsilon3/2, epsilon3/3, and epsilon4/3) predict the presence of CAC. The addition of information about ApoE genotypes to logistic models containing each separate risk factor did not improve prediction of CAC (P>0.10 in both women and men). However, there was significant evidence (P<0.10) that associations between variation in the probability of having CAC and variation in body mass index, plasma total cholesterol, and plasma ApoB in men and body mass index, plasma triglycerides, plasma ApoA1, and plasma ApoE in women were dependent on ApoE genotype. Thus, variation in the gene coding for ApoE may play a role in determining the contribution of established risk factors to risk of CAC.

Correlates of family history of coronary artery disease in children.

The atherosclerotic process begins in childhood but, in general, does not reach the clinical horizon until after the fifth decade of life, at which point the best opportunities for prevention and intervention have been lost. In order to identify children with a high risk of developing coronary artery disease (CAD), risk factors measured in children that are the most informative indicators of future risk must be identified. Using a novel analytical strategy that incorporates a continuum of information about context dependency, we investigated whether there were significant differences in intermediate biochemical and physiological traits between children (189 females and 188 males, ages 5-20.5 years) with and without a strong family history of clinically-defined CAD at three levels of context dependency (coarse grain, medium grain, and fine grain). In the coarse-grained analysis we tested for differences in mean levels of nine intermediate traits (lipids, apolipoproteins, blood pressure traits) and indices of external and internal environmental context (age, body mass index, smoking status). Female children with a strong family history had higher average levels for total cholesterol, triglyceride, Apo B, and systolic blood pressure and were on average older and weighed more than female children with a weak family history of CAD. Male children with a strong family history of CAD had higher average levels of triglycerides and were on average older than male children with a weak family history. In the medium-grained analysis we investigated whether the regression relationships between each intermediate trait and each measure of environmental context was significantly different between children with and without a strong family history of CAD. Our results indicate that children with a strong family history of CAD have a significantly different relationship between their intermediate traits and environmental contexts than children with a weak family history. In the fine-grained analysis, we stratified the sample into age, BMI, and smoking subgroups and tested for mean differences in the intermediate traits between children with and without a strong family history. For seven of the nine intermediate traits we found evidence of significant mean differences between children with and without a strong family history of CAD in particular age and BMI subgroups in nonsmokers that were not expected given the results from separate age-dependent or BMI-dependent marginal analyses. From these analyses, we conclude that the inferences about intermediate biochemical and physiological trait associations with family history of CAD depend on where on the coarse-grain to fine-grain continuum of context dependency the analysis is performed. In many cases, inferences at one level of investigation are different than the inferences made at a coarser or finer level. This study documents the complexity of the associations between intermediate traits and risk of CAD and raises the question of how many models are needed to maximize disease prediction and where these models should fall on the coarse- to fine-grain continuum.

Syndrome X: is it for real?

The term syndrome X has been applied to the association of hypertension, non-insulin-dependent diabetes mellitus (NIDDM), android obesity, insulin resistance, and dyslipidemia. In this paper, based on population samples from Tecumseh, Michigan, and Hiroshima, Japan, characterized by persons > or = 40 years of age, we examine the validity of regarding this constellation of traits as a true syndrome, i.e., an array of traits with a single, unifying pathophysiology underlying its components. Data were not available on insulin resistance and dyslipidemia, and obesity was expressed as body mass index (BMI) without the division into android and non-android types. The four ethnic-gender data sets were analyzed on the basis of two age classes, age > or = 40 years and age > or = 50 years, and two obesity classes, BMI > or = 27 and > or = 30. A simple chi 2 test of goodness-of-fit under a model of independence revealed non-random associations between hypertension, NIDDM, and BMI which were in part attributable to an excess of persons with all three traits. However, when the four data sets were subjected to separate log-linear analyses of the three-way association tables, none of the three-factor interaction terms (i.e., syndrome X) was significant. High significance was, however, observed in the two-factor interaction term for BMI*hypertension. It is concluded that the significant association between these three traits is driven by the BMI*hypertension interaction, and there is no evidence in these data sets of a significant role for a syndrome X. Genet.

Association between common alleles of the low-density lipoprotein receptor gene region and interindividual variation in plasma lipid and apolipoprotein levels in a population-based sample from Rochester, Minnesota.

This paper presents an analysis of the relationship between variation in the low-density lipoprotein receptor (LDLR) gene region and interindividual variation in plasma lipid and apolipoprotein levels in a sample representative of the adult population of Rochester, Minn. (217 females and 187 males aged 26 to 63). This relationship was analyzed by estimating the average excesses of alleles of the LDLR gene defined using RFLP markers both singly and simultaneously. We also used a cladistic approach to illustrate the consequences of incorporating evolutionary information into the analysis of genotype-phenotype relationships. Although results from both approaches supported the inference that common variation in the LDLR gene region associates with small effects on plasma lipid and apolipoprotein levels, only the cladistic approach provides direction for further work aimed at identifying the functional DNA sequence variations responsible for the observed associations.

Genetic architecture of common multifactorial diseases.

The purpose of this paper is to foster a dialogue among those interested in the genetic analysis of common chronic diseases. The need for an alternative to the Cartesian-Mendelian research strategy is discussed. A biological model that considers an individual's health to be an emergent property of a complex adaptive system is presented. A review of the contribution of the gene coding for apolipoprotein E to the genetic architecture of measures of lipid metabolism documents that an individual's coronary artery health has all the features expected of a complex adaptive system. Suggestions for enhancing our ability to use genetic information to predict onset, progression and severity of disease are offered.

Impact of apolipoprotein E genotype variation on means, variances, and correlations of plasma lipid, lipoprotein, and apolipoprotein traits in octogenarians.

The impact of apolipoprotein (apo) E genotype variation on means, variances and correlations between plasma lipid traits was studied in male and female octogenarians. Females had significantly higher mean levels of all 10 of the measured plasma lipid traits than males. The subset of concomitants (i.e., age, height, weight, body mass index, glucose and uric acid) that made a statistically significant contribution to interindividual variability was different in males and females for every trait considered. Gender-specific associations between variation in apo E genotype and variation in particular measures of lipid metabolism, adjusted for concomitant variation, were observed: in females there were no statistically significant associations while in males the means of the three common apo E genotypes were significantly different for adjusted measures of total cholesterol, low density lipoprotein cholesterol and low density lipoprotein-apo B. The common apo E genotypes were heterogeneous with respect to intragenotypic variance for adjusted log-transformed triglyceride levels in females only. Finally, the three common apo E genotypes were heterogeneous with respect to the correlation between traits, adjusted for concomitant variation, and gender influenced the manner in which the genotypes differed for specific correlations. This study documents that variation in the apo E gene has a significant impact on means, variances and correlations of plasma lipid traits in octogenarians, but the effects are context-, that is gender- and age-, dependent.

Cladistic analysis of the apolipoprotein AI-CIII-AIV gene cluster using a healthy French Canadian sample. I. Haploid analysis.

A cladistic analysis was carried out to identify haplotypes hypothesized to differ for functional DNA sequence variations within the apolipoprotein (apo) AI-CIII-AIV gene cluster that affect plasma lipid, lipoprotein and apolipoprotein levels. A sample of unrelated healthy French Canadians was studied. First, a cladogram of the observed apo AI-CIII-AIV haplotypes was estimated. Then this cladogram was used to define a statistical analysis of the association between haplotype variation and variation in plasma lipid, lipoprotein and apolipoprotein levels. Three haplotypes were identified which were associated with small (5-12% of the total sum of squares) pleiotropic effects on plasma lipid, lipoprotein and apolipoprotein traits and these effects were context, i.e. gender, dependent.

Biological complexity and strategies for finding DNA variations responsible for inter-individual variation in risk of a common chronic disease, coronary artery disease.

Most common chronic diseases of humans aggregate, but do not segregate, in families. The segregation-linkage research paradigm has not provided great insights into their genetic etiology. In this paper, using coronary artery disease as an example, we discuss hierarchical organization, coherence, emergent properties and dynamism as features that characterize the complexity of genotype-phenotype relationships. We summarize a research strategy for evaluating the contribution of genetic and environmental factors to the prediction of inter-individual variation in risk of disease. We then review a statistical strategy that employs cladistic theory to identify individuals carrying mutant DNA sequences responsible for an observed association between marker variation in a gene and inter-individual variation in biological traits that determine risk of a common multifactorial disease. Finding these DNA sequences is a necessary step in our search for an understanding of the nature of the mapping of genetic variation into variation in risk of a disease like coronary artery disease.

Application of cladistics to the analysis of genotype-phenotype relationships.

We seek to understand the relative contribution of allelic variations of a particular gene to the determination of an individual's risk of atherosclerosis or hypertension. Work in progress is focusing on the identification and characterization of mutations in candidate genes that are known to be involved in determining the phenotypic expression of intermediate biochemical and physiological traits that are in the pathway of causation between genetic variation and variation in risk of disease. The statistical strategy described in this paper is designed to aid geneticists and molecular biologists in their search to find the DNA sequences responsible for the genetic component of variation in these traits. With this information we will have a more complete understanding of the nature of the organization of the genetic variation responsible for quantitative variation in risk of disease. It will then be possible to fully evaluate the utility of measured genetic information in predicting the risk of common diseases having a complex multifactorial etiology, such as atherosclerosis and hypertension.

Estimation of Hardy-Weinberg and pairwise disequilibrium in the apolipoprotein AI-CIII-AIV gene cluster.

Departures from Hardy-Weinberg (HW) equilibria and pairwise disequilibria were estimated in a sample of unrelated healthy individuals typed for six RFLPs in the apo AI-CIII-AIV gene region. The sample was composed of males and females, selected for health, from two populations, those of exclusively French-Canadian (FC) and those of some non-French-Canadian (NFC) ancestry. An approach suggested by Weir and Cockerham, which includes estimates of nonrandom association (disequilibria) between three and four alleles at two loci as well as the traditional associations between two alleles, at two loci was used. The pattern of departures from HW equilibria suggested that the genetic structures of the FC and NFC are different. Departure from HW equilibrium at an RFLP locus could not be predicted from information about other loci in the same gene region. Nonrandom associations were also evident from the pairwise analyses. Two pairs of loci had significant diallelic disequilibria, while two other pairs had significant triallelic disequilibria. All of the RFLP pairs had at least one measure of disequilibrium at its maximum value determined by allele frequencies. Inferences about pairwise disequilibria depended on the statistical approach used. Sizes of the pairwise disequilibria were not correlated with the physical distance between loci. The impact of these disequilibria on RFLP-phenotype association studies is discussed.

Expression of the Axd (axial defects) mutation in the mouse is insensitive to retinoic acid at low dose.

The Axd mutation in the mouse acts by an unknown mechanism to cause lumbosacral open neural tube defects and a variety of tail anomalies. Retinoic acid (RA) plays a number of different physiological and developmental roles and has been shown to affect neurulation in mice and other species. Indeed, reports have shown that this biologically active compound (or its metabolites) at low dose can alter the incidence of neural tube defects (NTD) in curly-tail (ct), splotch (Sp), and delayed splotch (Spd) mice, strains that are genetically predisposed to such abnormalities. The aim of the present study was to determine if RA administered under similar conditions would affect the penetrance or expression of the Axd mutation or survival of Axd homozygotes. Axd/+ and +/+ dams were exposed to RA intraperitoneally (5 mg/kg) on D9 postcoitus. No difference in incidence or extent of neural tube defects or other axial anomalies was detected among embryos of Axd/+ dams given RA compared with those administered vehicle only. This finding is consistent with the diversity of gene-controlled steps required for neurulation and the differing sensitivities of specific mutants to rescue by extrinsic agents.

Expression of a new mutation (Axd) causing axial defects in mice correlates with maternal phenotype and age.

A new autosomal mutation, Axd (axial defects), is described. Axd segregates in a simple Mendelian fashion, and it is dominant with incomplete penetrance and variable expressivity. The phenotype of Axd heterozygotes ranges from a variety of tail anomalies to visibly normal tails. Approximately 12% of neonates from curly-tail (CT) F1 (Axd/+) x F1 (Axd/+) matings exhibit open neural tube defects (NTD) in the lumbosacral region and 16% have curly tails. Mean litter sizes and resorption rates comparable to wild type indicate that homozygosity for Axd is not obligately lethal. Genetic background plays a major role in Axd expression. Strains such as BALB/cByJ allow the highest penetrance of the mutation in single dose (46%), whereas, in CF-1 mice Axd is recessive. The tail phenotype of heterozygous Axd/+ dams, in part reflective of their genetic background, correlates with the incidence of NTD in F2 offspring: CT mothers produce significantly more neonates with frank NTD than normal tail mothers. At the one embryonic period examined for this study (D13/D14 post-coitus), an 85% higher incidence of total axial defects is observed than among the F2 at birth. Unchanging litter size and the relative increase in phenotypically normal offspring by birth suggest that Axd acts by delaying posterior neural tube closure. One of the most significant findings in this study is that maternal age influences the survival of Axd embryos in utero. Axd/+ dams older than 8 months yield fewer mean implants, higher resorption rates, and fewer viable embryos with axial defects than do Axd/+ dams younger than 8 months. Axd is not allelic to nor linked to the Sp (splotch) gene which also affects neurulation.