Identifying modifier loci in existing genome scan data.

In many genetic disorders in which a primary disease-causing locus has been identified, evidence exists for additional trait variation due to genetic factors. These findings have led to studies seeking secondary 'modifier' loci. Identification of modifier loci provides insight into disease mechanisms and may provide additional screening and treatment targets. We believe that modifier loci can be identified by re-analysis of genome screen data while controlling for primary locus effects. To test this hypothesis, we simulated multiple replicates of typical genome screening data on to two real family structures from a study of hypertrophic cardiomyopathy. With this marker data, we simulated two trait models with characteristics similar to one measure of hypertrophic cardiomyopathy. Both trait models included 3 genes. In the first, the trait was influenced by a primary gene, a secondary 'modifier' gene, and a third very small effect gene. In the second, we modeled an interaction between the first two genes. We examined power and false positive rates to map the secondary locus while controlling for the effect of the primary locus with two types of analyses. First, we examined Monte Carlo Markov chain (MCMC) simultaneous segregation and linkage analysis as implemented in Loki, for which we calculated two scoring statistics. Second, we calculated LOD scores using an individual-specific liability class based on the quantitative trait value. We found that both methods produced scores that are significant on a genome-wide level in some replicates. We conclude that mapping of modifier loci in existing samples is possible with these methods.

A bootstrapped commingling analysis of platelet monoamine oxidase activity levels corrected for cigarette smoking.

Monoamine oxidase (MAO) activity levels have been suggested as a possible biological marker for alcohol dependence and abuse, as well as for schizophrenia and other psychiatric conditions. Using platelet MAO activities in the Collaborative Study on the Genetics of Alcoholism data set, we applied bootstrapping methods as a novel way to test for admixture in families. This bootstrapping involved resampling in family units and hypothesis testing of the resampled datasets for commingling in the distribution of MAO activity levels. Prior to commingling analysis, we used linear models to find covariates of greatest effect on MAO activity levels. While an alcoholism diagnosis was significant in men (n = 1151, P < 0.0001) and women (n = 1254, P = 0.0003), the effect lost significance after controlling for cigarette smoking, indicating alcoholism and smoking behavior to be highly confounded. When smoking histories were compared, former smokers had levels (mean = 7.1) closer to those who never smoked (mean = 7.0) than to current smokers (mean = 5.4). Furthermore, current daily smoking and time since smoking cessation were significantly related to MAO levels, indicating smoking probably has a direct effect on MAO levels, rather than the reverse. These results suggest that studies using MAO levels as a biological marker should consider smoking as an important covariate. Finally, admixture was found in MAO levels controlled for smoking and sex, possibly indicating a major genetic locus; this confirms previous evidence for admixture.

The importance of connections: joining components of the Hutterite pedigree.

We analyzed a quantitative trait (serum IgE levels), and a binary trait (asthma), in four Hutterite sub-pedigrees. A genome screen for asthma was performed using GENEHUNTER, and interesting regions were followed up using extended pedigrees and the FASTLINK package. Markov chain Monte Carlo (MCMC) methods were used to assess haplotype sharing among affected individuals (MORGAN/AUTOZYG), and to perform a combined oligogenic segregation and linkage analysis (LOKI) for log10(IgE). We found evidence for at least two susceptibility loci for asthma on chromosome 5, and a QTL for log10(IgE) on chromosome 1. Our analyses demonstrate that using the most complete pedigree structure possible is advisable, with attention to the possibility of heterogeneity among subunits of a very large pedigree.

Bias in multipoint linkage analysis arising from map misspecification.

Multipoint linkage analysis methods are often used in human genetic studies. Although multipoint methods increase power for a linkage analysis and will become essential if use of diallelic markers becomes widespread, the methods in use assume an accurate meiotic marker map. Unfortunately, uncertainties in estimates of between-marker meiotic distances are large. Also, sex-averaged maps are generally used, but recombination rates differ in males and females. Both these types of map misspecification can lead to lod score bias, but such bias has not previously been systematically quantified. We examine multipoint lod score bias arising from these map misspecifications, in both the presence and absence of actual linkage. We define bias as the expected difference between the lod score computed under the misspecified map and that computed under the true map. With actual linkage, any map misspecification causes negative bias in lod scores, resulting in loss of power to detect linkage. In most cases, bias is modest, only reaching clearly detectable levels when both types of misspecification are substantial. In the absence of linkage, map misspecification can cause positive or negative bias: falsely assuming a 1:1 female:male ratio always causes positive bias; using too large a distance gives a positive bias; using too small a distance gives a negative bias. This bias can inflate the false-positive rate, especially when the sample size is modest. We conclude that although current sex-averaged maps are suitable for a first-pass multipoint screen, the potential for bias from map misspecification should be evaluated in following up results from such an analysis.

The number of trait loci in late-onset Alzheimer disease.

Although it is clear that apoE plays an important role in the genetics of late-onset Alzheimer disease (AD), evidence exists that additional genes may play a role in AD, and estimates of the total contribution of apoE to the variance in onset of AD vary widely. Unfortunately, little information is available on the number and contribution of additional genes. We estimated the number of additional quantitative-trait loci and their contribution to the variance in age at onset of AD, as well as the contribution of apoE and sex, in an oligogenic segregation analysis of 75 families (742 individuals) ascertained for members with late-onset AD. We found evidence that four additional loci make a contribution to the variance in age at onset of late-onset AD that is similar to or greater in magnitude than that made by apoE, with one locus making a contribution several times greater than that of apoE. Additionally, we confirmed previous findings of a dose effect for the apoE varepsilon4 allele, a protective effect for the varepsilon2 allele, evidence for allelic interactions at the apoE locus, and a small protective effect for males. Furthermore, although we estimate that the apoE genotype can make a difference of

Reproducibility of maximal exercise test data in the HERITAGE family study.

PURPOSE: The reproducibility of responses to maximal cycle ergometer testing was determined using data from the HERITAGE Family study at four Clinical Centers in the United States and Canada. METHODS: Reproducibility was determined from maximal exercise test data obtained a) on 2 d in a sample of 390 subjects (198 men and 192 women), b) across 4 d in an Intracenter Quality Control (ICQC) substudy with 55 subjects who were not part of the main study, and c) across 2 wk in a Traveling Crew Quality Control (TCQC) substudy with the same eight subjects who were tested at each of the four centers. Reproducibility was evaluated using technical errors, coefficients of variation (CV) for repeated measures, and intraclass correlation coefficients (ICC) for selected variables obtained on the main cohort, as well as on the ICQC and TCQC substudies. RESULTS: With the exception of systolic and diastolic blood pressures and respiratory exchange ratio, all the other variables (heart rate, ventilation, VO2, and VCO2) were highly reproducible, with CV below 10% and ICC over 0.86. These results were similar to those previously reported on the same subjects at a submaximal power output associated with 60% VO2max. Results were consistent for the main cohort, the ICQC sample, the TCQC sample, and across all four Clinical Centers. CONCLUSIONS: Day-to-day variations are small and reproducibility is high for maximal values of heart rate, ventilation, VO2 and VCO2 at each of the four Clinical Centers of the HERITAGE Family Study.

Monte Carlo Markov chain methods for genome screening.

We used Monte Carlo Markov chain (MCMC) methods to analyze a quantitative trait, MAO level, and a discrete trait, Collaborative Study on the Genetics of Alcoholism (COGA) alcoholism. Segregation, linkage, and haplotype sharing were analyzed and effects of marker map features were examined. For MAO, modest signals were found on chromosomes 1 and 17 for raw data, and 15 for covariate-adjusted data. For alcoholism, a strong signal was found on chromosome 1 with modest signals on chromosomes 4 and 10.

Multipoint oligogenic analysis of age-at-onset data with applications to Alzheimer disease pedigrees.

It is usually difficult to localize genes that cause diseases with late ages at onset. These diseases frequently exhibit complex modes of inheritance, and only recent generations are available to be genotyped and phenotyped. In this situation, multipoint analysis using traditional exact linkage analysis methods, with many markers and full pedigree information, is a computationally intractable problem. Fortunately, Monte Carlo Markov chain sampling provides a tool to address this issue. By treating age at onset as a right-censored quantitative trait, we expand the methods used by Heath (1997) and illustrate them using an Alzheimer disease (AD) data set. This approach estimates the number, sizes, allele frequencies, and positions of quantitative trait loci (QTLs). In this simultaneous multipoint linkage and segregation analysis method, the QTLs are assumed to be diallelic and to interact additively. In the AD data set, we were able to localize correctly, quickly, and accurately two known genes, despite the existence of substantial genetic heterogeneity, thus demonstrating the great promise of these methods for the dissection of late-onset oligogenic diseases.

Alterations in resting metabolic rate as a consequence of 20 wk of endurance training: the HERITAGE Family Study.

This study determined the effects of endurance exercise training on the resting metabolic rate (RMR). It was hypothesized that the RMR would be increased posttraining, but that this increase would reflect the influence of the last exercise bout, not a chronic adaptation to exercise training. Seventy-four subjects (40 men and 37 women) aged 17-63 y participated in a 20-wk endurance training program. RMR and maximal oxygen uptake (VO2max) were each measured on 2 separate days both pre- and posttraining; the posttraining RMR measurements were taken 24 and 72 h after the last exercise bout. There were small but significant changes posttraining in relative body fat (-1.0%), fat mass (-0.6 kg), and fat-free mass (0.7 kg) and a 17.9% increase in VO2max. The RMR remained unchanged posttraining, both 24 and 72 h after the last exercise bout, even when the data were adjusted to account for the potential confounding effects of age, sex, body composition, and VO2max. In conclusion, 20 wk of endurance exercise training had no effect on the RMR even in the presence of small changes in body composition and a large increase in VO2max.

Familial resemblance for VO2max in the sedentary state: the HERITAGE family study.

This study investigates the familial resemblance of maximal oxygen uptake (VO2max) based on data from 86 nuclear families of Caucasian descent participating in the HERITAGE Family Study. In the current study, VO2max was measured twice on a cycle ergometer in 429 sedentary individuals (170 parents and 259 of their offspring), aged between 16 and 65 yr. The VO2max was adjusted by regression procedures for the effects of 1) age and sex; 2) age, sex, and body mass; and 3) age, sex, body mass, fat mass, and fat-free mass, as determined by underwater weighing. Evidence for significant familial resemblance was observed for each of the three VO2max phenotypes. Spouse, sibling, and parent-offspring correlations were significant, suggesting that both genetic and environmental factors contribute to the familial resemblance for VO2max. Maximal heritability estimates were at least 50%, a value inflated to an undetermined degree by nongenetic factors. The hypothesis of maternal inheritance, with the father's contribution being environmental, was also found to fit the data with estimates of maternal heritability, potentially associated in part with mitochondrial inheritance, reaching about 30%. These results suggest that genetic and nongenetic factors as well as maternal influences contribute to the familial aggregation of VO2max in sedentary individuals.

Reproducibility of cardiovascular, respiratory, and metabolic responses to submaximal exercise: the HERITAGE Family Study.

This study determined the reproducibility of cardiovascular, respiratory, and metabolic responses to submaximal cycle ergometer exercise at two power outputs (50 W and 60% VO2max) on each of two separate days in a sample of 390 subjects (198 men and 192 women) participating in the HERITAGE Family Study. The same protocol was conducted across 3 d in an intracenter quality control substudy which included an additional 55 subjects. Reproducibility estimates included technical error, coefficient of variation, and intraclass correlation for each of the selected variables for both subject populations. Further, since the data were collected across four clinical centers, intraclass correlations were also computed separately by clinical center. Most variables were highly reproducible, with coefficients of variation below 9% and intraclass correlations over 0.80. These results were consistent for both subject populations and across all four clinical centers. Reproducibility indicators were generally better at the higher power output. It is concluded that within-subject day-to-day variation and measurement unreliabilities are generally small compared with the between-subject variance in the response to submaximal exercise at each of the clinical centers of the HERITAGE Family Study.

Temporal stability of antisocial personality disorder: blind follow-up study at 8 years.

The study objective was to examine the temporal stability of the antisocial personality disorder (ASPD) diagnosis based on whether specific antisocial symptoms were considered to be related to substance abuse. A total of 407 adults who were initially part of a family study of alcoholism and sociopathy were blindly reassessed an average of 8 years later, using the Home Environment and Lifetime Psychiatric Evaluation Record (HELPER) and basing diagnoses on the clinician's best final estimate using all sources of data. "Narrow" and "broad" ASPD diagnoses were made at both times based on whether individual symptoms were counted toward diagnosis if they occurred in the setting of significant substance abuse. kappa values varied from 0.31 to 0.68, with more restrictive methods of diagnosis being less stable. After deriving estimates of sensitivity and specificity of diagnosis, the probability of being a "case" could be assigned based on the reported number of conduct problems occurring before age 15 as a clinical covariate for diagnosis. We conclude that diagnosing ASPD without attempting to attribute the cause of individual symptoms to substance abuse results in substantially greater temporal stability. Using a broader definition, the diagnosis of ASPD is highly sensitive (P = .97) and specific (q = 0.93). These findings may allow more accurate diagnosis of ASPD in drug-abusing individuals.

Reproducibility of the HERITAGE Family Study intervention protocol: drift over time.

PURPOSE: The primary goal of the HERITAGE Family Study was to document the role of the genotype in the response to aerobic exercise training. Toward this end, nuclear families were enrolled in a 20-week exercise training program, with a large variety of tests performed before and after the training. Since study drift has the potential to adversely affect the results, reproducibility and potential bias over six consecutive 4-month periods were examined for selected test. METHODS: Intraclass correlations (ICC), technical errors (TE), coefficients of variation within subject (CV), and means were calculated with use of the pretraining test results for each of the six time periods. To check for homogeneity, hypothesis tests were performed on the intraclass correlations and means. If homogeneity was not found across all six periods, further tests were performed to assess differences between pairs of time periods. RESULTS: There was little evidence for real drifts in reproducibility, with most tests having ICCs of 0.8 or better. Only a few tests showed any change over time, and in no case was there evidence of a systematic drift in mean values. CONCLUSIONS: Overall, the reproducibility of the HERITAGE Family Study tests and assays considered in this paper was found to be very good, with no evidence of any systematic drift over time.

Familial resemblance for abdominal visceral fat: the HERITAGE family study.

OBJECTIVES: Abdominal visceral fat (AVF) is considered a risk factor for diabetes, atherogenic lipid profiles and hypertension. However, little is known about the genetic contribution to AVF as compared to total body fat. DESIGN: AVF was assessed by computerized tomography, and total body fat (fat mass) was assessed by underwater weighing in 86 families participating in the Heritage Family Study. All family members were sedentary at baseline examination. The familial factors underlying the variability in age-adjusted AVF, age-fat mass-adjusted AVF and age-adjusted fat mass, were assessed using a familial correlation model. RESULTS: The maximal heritability (including genetic and familial environmental effects) for AVF was comparable before (47%) and after (48%) adjusting for fat mass, and was 55% for fat mass itself in these sedentary families. Spouse correlations were significant for fat mass and for AVF prior to, but not after, adjustment for fat mass. CONCLUSIONS: These results confirm the only previous study which investigated the familial aggregation of AVF (both in pattern and magnitude), suggesting that the factors underlying AVF in these sedentary families may be similar to those in the population at large. Although both genetic and familial environmental factors probably influence each of fat mass and AVF, there appears to be a predominantly genetic etiology for the visceral component which is independent of total body fat. These findings imply that some individuals are more at risk then others because of an inherited tendency to store abdominal fat viscerally rather than subcutaneously.

Reproducibility of anthropometric and body composition measurements: the HERITAGE Family Study.

OBJECTIVE: To determine the reproducibility of anthropometric and body composition measures using the HERITAGE Family Study protocol. DESIGN: Anthropometric and body composition measures were obtained on three separate days within a 3-wk period at each of the four HERITAGE Clinical Centers. SUBJECTS: Sixty men and women representative of the HERITAGE subject population, 15 from each of four Clinical Centers. MEASUREMENTS: Anthropometric measures included eight skinfolds, three girths and one length; and body composition measures included stature, mass, hydrostatic weight, residual volume, and body density, from which relative fat, fat mass and fat-free mass were estimated. RESULTS: Reproducibility as determined by technical error, coefficient of variation, and intraclass correlations was very high for the total sample. For example, intraclass correlations for the total sample generally ranged from 0.95-0.99 for the anthropometric measures, and from 0.97-1.00 for the body composition measures. The results across Clinical Centers were in close agreement with each other and with the pooled data. CONCLUSIONS: The reproducibility of anthropometric and body composition measures using the HERITAGE Family Study protocol is sufficiently high that it should be possible to detect small changes in any of these measures and to determine the genetic basis of these changes consequent to a 20 wk endurance training program.

Familial resemblance for body composition measures: the HERITAGE Family Study.

A sex-specific familial correlation model was used to assess the heritable contributions to several measures of body composition in 86 sedentary white families participating in the HERITAGE Family Study. For this study, sedentary families were recruited, tested for a battery of measures, endurance exercise trained for 20 weeks, and remeasured. This sample is unique in that activity level was controlled for in these families at baseline measurement. In this report, three body composition variables measured at baseline were analyzed, two indexing adiposity (total subcutaneous fat based on eight skinfold measurements [SF8] and percent body fat measured by underwater weighing techniques [%BF]) and one assessing fat free mass ([FFM] derived from underwater weighing). The maximal heritabilities for SF8 (34%) and %BF (62%) were consistent with those reported in previous studies. There were no sex nor generation differences in the familial correlations, and the spouse correlation was significant, consistent with the hypothesis that the familial aggregation reflects genetic and familial environmental factors. However, the results for FFM were very different. The most parsimonious pattern of familial resemblance was consistent with mitochondrial inheritance (i.e., mother-offspring and sibling correlations were equal and were larger than those for spouse and father-offspring pairs). Under the mitochondrial hypothesis, 39% of the variance was accounted for by familial/genetic effects. However, under a nonmitochondrial hypothesis, which could not be ruled out, 65% of the FFM phenotypic variance was accounted for by familial/genetic factors. This high heritability level, as compared with results from previous studies, is consistent with the hypothesis that activity may constitute an important environmental determinant of FFM. These alternative hypotheses for FFM warrant further investigation using complex multilocus-multitrait segregation models, which allow for major genetic, polygenic, and environmental sources of variance, as well as interactions among them.

TDT with covariates and genomic screens with mod scores: their behavior on simulated data.

We describe an extension to the TDT (transmission/disequilibrium test) which allows for more than two marker alleles and for covariates measured on the parent or offspring. We also describe a systematic genomic search where the mod score (maximized lod score) is computed for each marker under constraints on the population prevalence or penetrances of a single locus.