Prevalence of C282Y and H63D mutations in the hemochromatosis (HFE) gene in the United States.

CONTEXT: Population-based estimates of the prevalence of disease-associated mutations, such as hemochromatosis (HFE) gene mutations, are needed to determine the usefulness of genetic screening. OBJECTIVE: To estimate the prevalence of the HFE mutations C282Y and H63D in the US population. DESIGN: Cross-sectional population-based study of samples in the DNA bank from phase 2 of the Third National Health and Nutrition Examination Survey conducted from 1992 to 1994. SETTING AND PARTICIPANTS: Genotyped samples of cells from a total of 5171 participants, cross-classified by sex, age, and race/ethnicity in the analysis. MAIN OUTCOME MEASURES: Estimates of the prevalence of C282Y and H63D mutations. RESULTS: The prevalence of C282Y homozygosity is estimated to be 0.26% (95% confidence interval [CI], 0.12%-0.49%); 1.89% (95% CI, 1.48%-2.43%) for H63D homozygosity; and 1.97% (95% CI, 1.54%-2.49%) for compound heterozygosity. The prevalence estimates for C282Y heterozygosity (C282Y/wild type) are 9.54% among non-Hispanic whites, 2.33% among non-Hispanic blacks, and 2.75% among Mexican-Americans. The prevalence estimates of the C282Y mutation in the US population are 5.4% (95% CI, 4.7%-6.2%) and 13.5% (95% CI, 12.5%-14.8%) for the H63D mutation. CONCLUSIONS: Estimates of prevalence of HFE mutations are within the expected range for non-Hispanic whites and blacks but the estimated prevalence of the C282Y mutation among Mexican-Americans is less than expected. Mutation data now need to be linked to clinically relevant indices, such as transferrin saturation level.

Levels of seven urinary phthalate metabolites in a human reference population.

Using a novel and highly selective technique, we measured monoester metabolites of seven commonly used phthalates in urine samples from a reference population of 289 adult humans. This analytical approach allowed us to directly measure the individual phthalate metabolites responsible for the animal reproductive and developmental toxicity while avoiding contamination from the ubiquitous parent compounds. The monoesters with the highest urinary levels found were monoethyl phthalate (95th percentile, 3,750 ppb, 2,610 microg/g creatinine), monobutyl phthalate (95th percentile, 294 ppb, 162 microg/g creatinine), and monobenzyl phthalate (95th percentile, 137 ppb, 92 microg/g creatinine), reflecting exposure to diethyl phthalate, dibutyl phthalate, and benzyl butyl phthalate. Women of reproductive age (20-40 years) were found to have significantly higher levels of monobutyl phthalate, a reproductive and developmental toxicant in rodents, than other age/gender groups (p < 0.005). Current scientific and regulatory attention on phthalates has focused almost exclusively on health risks from exposure to only two phthalates, di-(2-ethylhexyl) phthalate and di-isononyl phthalate. Our findings strongly suggest that health-risk assessments for phthalate exposure in humans should include diethyl, dibutyl, and benzyl butyl phthalates.

Analysis of factors influencing the comparison of homocysteine values between the Third National Health and Nutrition Examination Survey (NHANES) and NHANES 1999+.

Two important changes occurred in the time between the Third National Health and Nutrition Examination Survey (NHANES III) (1991-1994) and the later survey (NHANES 1999+) regarding total homocysteine (tHcy), i.e., a change in matrix from serum to plasma and a change in analytical methods. The goals of this study were to determine the magnitude of potential differences between plasma and serum with regard to tHcy concentrations, and between the two analytical methods used in these surveys. Optimally prepared plasma, serum allowed to clot for 30 and 60 min at room temperature and serum allowed to clot for 30 and 60 min and subjected to four freeze-thaw cycles, prepared from blood samples collected from 30 healthy people, were analyzed by both methods. Serum samples had significantly higher tHcy concentrations than plasma samples, and the difference increased with longer clotting time. Freeze-thaw cycles had little or no effect on the variability or bias in the serum sample results. The tHcy results produced by the two analytical methods were significantly different, but consistent across sample types. On average, the results of the method used in NHANES III were lower by 0.64 micromol/L; however, the relative bias varied with tHcy concentration. The tHcy results determined in surplus serum from NHANES III overestimated tHcy concentrations by approximately 10% compared with optimally prepared plasma. The average method bias was 6% between the two analytical methods. On the basis of changes in matrix and methodology, direct comparison of tHcy results between the two surveys is inappropriate.

Levels of methyleugenol in a subset of adults in the general U.S. population as determined by high resolution mass spectrometry.

We developed a sensitive and accurate analytical method for quantifying methyleugenol (ME) in human serum. Our method uses a simple solid-phase extraction followed by a highly specific analysis using isotope dilution gas chromatography-high resolution mass spectrometry. Our method is very accurate; its limit of detection is 3.1 pg/g and its average coefficient of variation is 14% over a 200-pg/g range. We applied this method to measure serum ME concentrations in adults in the general U.S. population. ME was detected in 98% of our samples, with a mean ME concentration of 24 pg/g (range < 3.1-390 pg/g). Lipid adjustment of the data did not alter the distribution. Bivariate and multivariate analyses using selected demographic variables showed only marginal relationships between race/ethnicity and sex/fasting status with serum ME concentrations. Although no demographic variable was a good predictor of ME exposure or dose, our data indicate prevalent exposure of U.S. adults to ME. Detailed pharmacokinetic studies are required to determine the relationship between ME intake and human serum ME concentrations.

Exposure of the U.S. population aged 6 years and older to cadmium: 1988-1994.

Cadmium was measured in urine specimens from 22,162 participants in the Third National Health and Nutrition Examination Survey (NHANES III 1988-1994). Urine cadmium, expressed either as uncorrected (microg/L) or creatinine corrected (microg/g creatinine) increased with age and with smoking. The arithmetic mean value for urine cadmium in the U.S. population was 0.57 microg/L or 0.48 microg/g creatinine. Based on our estimates, about 2.3% of the U.S. population have urine cadmium concentrations greater than 2 microg/g creatinine, and 0.2% have concentrations greater than 5 microg/g creatinine, the current World Health Organization health-based exposure limit.

Urinary albumin excretion in children: factors related to elevated excretion in the United States population.

Past population studies have indicated a higher prevalence of high albumin excretion in children than in adults. In this study, NHANES III United States population data was analyzed to study factors associated with elevated albumin excretion in children 8 to 18 years of age. The analysis confirmed a higher prevalence of albumin values > 30 mg/g creatinine and > 200 mg/g creatinine in children than in adults, and indicated that girls are two to three times more likely to have albumin excretion above these levels than boys. Neither hypertension nor reported diabetes--major factors influencing albumin excretion in adults--accounted for the higher excretion levels in children. The higher excretion levels were not associated with prescription medications or a poor rating of the child's overall health status by a physician. The higher prevalences is influenced by puberty stage and is more likely to occur in children with lower than average body mass index, independent of the relationship with urine creatinine excretion. The increased prevalence of high albumin excretion is probably associated with normal development in children, but an increased susceptibility to chronic diseases in the future among the children with high excretion cannot be ruled out.

Uranium and thorium in urine of United States residents: reference range concentrations.

We measured uranium and thorium in urine of 500 U. S. residents to establish reference range concentrations using a magnetic-sector inductively coupled argon plasma mass spectrometer (ICP-MS). We found uranium at detectable concentrations in 96.6% of the urine specimens and thorium in 39.6% of the specimens. The 95th percentile concenetration for uranium was 34.5 ng/L (parts per trillion); concentrations ranged up to 4080 ng/L. Thorium had a 95th percentile concentration of 3.09 ng/L; concentrations ranged up to 7.7 ng/L.

Detection of mosaicism in amniotic fluid cultures: a CYTO2000 collaborative study.

PURPOSE: To evaluate the assumptions on which the American College of Medical Genetics (ACMG) Standards and Guidelines for detecting mosaicism in amniotic fluid cultures are based. METHODS: Data from 653 cases of amniotic fluid mosaicism were collected from 26 laboratories. A chi-square goodness-of-fit test was used to compare the observed number of mosaic cases with the expected number based on binomial distribution theory. RESULTS: Comparison of observed data from the in situ colony cases with the expected distribution of cases detected based on the binomial distribution did not reveal a significant difference (P = 0.525). CONCLUSIONS: The empirical data fit the binomial distribution. Therefore, binomial theory can be used as an initial discussion point for determining whether ACMG Standards and Guidelines are adequate for detecting mosaicism.

Assessment of current National Cholesterol Education Program guidelines for total cholesterol triglyceride, HDL-cholesterol, and LDL-cholesterol measurements.

We examine the effect of systematic bias and random error, quality control, and intraperson biological variation on the National Cholesterol Education Program (NCEP) clinical classifications for reported lipid measurements. We consider misclassification to occur if a true lipid homeostatic set point is within a desirable range but the reported lipid value is in a high-risk range, or if a true lipid homeostatic set point is in a high-risk range but the reported lipid value is in a desirable range. To evaluate the overall adequacy of the NCEP guidelines to ensure correct patient classification, we construct operating characteristic curves for total cholesterol, triglycerides, high-density lipoprotein cholesterol, and low-density lipoprotein cholesterol. We demonstrate that if laboratories are meeting the NCEP guidelines for inherent bias and analytic precision and are using standard quality-control (QC) procedures incorporating at least two QC samples per analytical run from each of two QC pools (for a total of 4 QC samples), the current NCEP guidelines are adequate to ensure (probability >0.90) correct patient classifications regardless of the size of the systematic bias of the laboratory or increased random analytic error. Thus we suggest that at least two concentrations of QC material be included in the QC scheme to ensure that the measurement system is operating within desired specifications across the entire range of desirable and high-risk lipid concentrations and to ensure with high probability that patients are correctly classified.

Serum levels of several organochlorine pesticides in farmers correspond with dietary exposure and local use history.

In response to reported increased cancer risks among farmers, the Agricultural Health Study (AHS) was designed to examine health outcomes and environmental exposures among farm families in the United States. In the pilot phase of the AHS, food, beverage, air, dermal, dust, surface wipe, and biological specimens (blood and urine) were collected and analyzed for six farm families in two states (IA and NC). In addition, questionnaires were administered to examine previous pesticide use. This paper reports the organochlorine pesticide results of the serum and dietary analyses as well as questionnaire results from the pilot exposure study of farmers and their families. Note, no organochlorine pesticides were reported as currently being applied to the study farms. In all human serum samples examined, typical U.S. population levels were found for the majority of the pesticides. In addition, human serum levels of organochlorine pesticides showed no significant daily or seasonal variation. However, serum trans-nonachlor levels were found to be higher in people living on the two farms in North Carolina than in people living on the four farms in Iowa (p < 0.05). Further, unusually high dieldrin levels were found in serum samples from a farmer and spouse living on an Iowa farm, and these levels were significantly higher than those of people living on the other farms (p < 0.05). Dieldrin was persistent in the foods consumed on the same Iowa farm where family members showed elevated serum levels. In addition, dietary samples from the North Carolina farms exhibited high levels of chlordane. No organochlorine pesticides were found in any of the drinking water samples. Dietary dieldrin levels on the same Iowa farm exceeded the oral reference dose (RfD) eight- to eleven-fold (50 ng/kg-day). No other pesticide exceeded the RfD. However, dietary chlordane levels at a North Carolina farm reached 17% of the RfD. Previous use of aldrin on an Iowa farm corresponded to dieldrin found in the diet and in the serum of the farmer and spouse. Previous reported use of chlordane on the North Carolina farms corresponded with measurable dietary levels of chlordance and higher serum trans-nonachlor levels than the levels in Iowa farm families.

Results of an international round robin for serum and whole-blood folate.

Because of the increasing significance of folate nutriture to public health, a "round robin" interlaboratory comparison study was conducted to assess differences among methods. Twenty research laboratories participated in a 3-day analysis of six serum and six whole-blood pools. Overall means, SDs, and CVs derived from these results were compared within and across method types. Results reported for serum and whole-blood folate demonstrated overall CVs of 27.6% and 35.7%, respectively, across pools and two- to ninefold differences in concentrations between methods, with the greatest variation occurring at critical low folate concentrations. Although results for serum pools were less variable than those for whole-blood pools, substantial intermethod variation still occurred. The overall results underscore the urgent need for developing and validating reference methods for serum and whole-blood folate and for properly characterized reference materials. For evaluating study or clinical data, method-specific reference ranges (established with clinical confirmation of values for truly folate-deficient individuals) must be used.

Pharmacokinetics of TCDD in veterans of Operation Ranch Hand: 10-year follow-up.

Using multiple measurements from serum collected over 10 yr (1982, 1987, and 1992), we estimated the half-life of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in 213 veterans of Operation Ranch Hand, the Air Force unit responsible for the aerial spraying of Agent Orange in Vietnam. The potential influences of age, percent body fat, and changes in percent body fat on the half-life estimate were also examined. The mean decay rate of TCDD for these veterans is 0.0797 per year with 95% confidence interval 0.0727 to 0.0868 per year; the corresponding half-life estimate is 8.7 yr with 95% confidence interval 8.0-9.5 yr. Half-life increased significantly with increasing body fat, but not with age or relative changes in percent body fat.

An in-depth examination of the excretion of albumin and other sensitive markers of renal damage in mild hypertension.

In an in-depth examination to better define the renal effects of mild hypertension, we used urinary proteins to indicate damage to the glomerulus (albumin), tubular reabsorption capability (retinol-binding protein), and turnover of tubular tissue (alanine aminopeptidase and N-acetyl-beta-D-glucosaminidase) in a group of 18 people with mild hypertension not associated with diabetes and a control group (n = 12). The participants' activity was controlled on a high normal salt diet for 3 days followed by a low salt diet for 4 days. Two distinct patterns of albumin excretion were evident in the hypertensive group: 22% had elevated, highly variable excretion patterns, and the rest had tightly grouped values below 16 mg/g creatinine, 16 micrograms/min, or 16 mg/L, with the lowest within-person biological variability given by albumin calculated as a ratio to creatinine. Albumin and NAG excretion primarily correlated with systolic blood pressure and the best correlations were given by ratios to creatinine. A marked decrease in salt excretion of 71% (to 50.8 mEq/day) resulted in significant (P < .0005) decreases in systolic (13.9 mm Hg), diastolic (6.4 mm Hg), and mean arterial pressures (8.9 mm Hg) only in the group with mild hypertension. However, albumin excretion did not decrease when dietary salt content was lowered. The group with hypertension also had higher urinary excretion of lysosomal N-acetyl-beta-D-glucosaminidase (P < .01), and whites in the group had a higher excretion of retinol-binding protein than did whites in the control group (P < .02). Retinol-binding protein values, however, were within the normal range, indicating that the elevated albumin values were the result of changes in selectivity of the glomerulus.

On combining dose-response data from epidemiological studies by meta-analysis.

Using data from a meta-analysis of the effects of oestrogen replacement therapy on the development of breast cancer, we compared alternative methods for combining dose-response slopes from epidemiological studies. We evaluated issues related both to summarizing data from single studies and to combining results from multiple studies. Findings related to the analysis of individual dose-response studies include: (1) a method of weighing studies that gives greater influence to dose-response slopes that conform to the linear relation of relative risk to duration can lead to large differences in calculated weights as a function of non-linearity; (2) a regression model using a variable-intercept resulted in a mean dose-response slope that increased as much as threefold when compared with the values obtained with a zero-intercept model. When combining results from multiple studies, we found: (1) calculating standard errors of mean dose-response slopes by methods that allow for both among-study and within-study variability (a random-effects type model) gave values different from a method that assumes homogeneity and equal within-study precision (a fixed-effects model); (2) the random-effects model gives mean and standard error results most similar to a bootstrap resampling method as increasing heterogeneity is observed (however, this model could give biased mean estimates compared with the bootstrap method); (3) a components-of-variance model compares favourably with the bootstrap and is easier to apply than the random-effects model. Based on these findings, we recommend the use of methods which incorporate heterogeneity to guard against underestimating the standard error. However, caution is urged because bias in point estimates can occur if extreme heterogeneity is present. Two other observations affect the interpretation of data combined from multiple studies. First, inclusion into a model of quality scores assigned by blinded reviewers had little effect on the mean dose-response slope and its standard error. Second, the number of studies required to achieve desired statistical power, varies with effect size.

Multiple hypothesis tests in multiple investigations.

Inferential statistical methods have traditionally been based on the assumption that one experiment is performed and that interest centres on one or more predetermined hypothesis tests. Exploratory research, on the other hand, often involves multiple hypotheses or repeated investigations under similar or different conditions or both. Several techniques have been proposed to deal with multiple or simultaneous hypothesis testing in single investigations, and procedures to combine observed significance levels for an individual hypothesis test from two or more investigations have been suggested. In this paper we propose a method for identifying important results from multiple statistical tests in multiple investigations. The method is illustrated by using high performance liquid chromatography to identify potential aetiologic contaminants in L-tryptophan samples.

Determinants of TCDD half-life in veterans of operation ranch hand.

The half-life of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) changed significantly with body fat and age in 337 members of Operation Ranch Hand, the Air Force unit responsible for the aerial spraying of Agent Orange in Vietnam. Using paired TCDD measurements derived from serum collected in 1982 and in 1987, we investigated how TCDD half-life varied with percent body fat (PBF), relative changes in PBF, and age. We found that half-life increased significantly with increasing PBF and decreased significantly with increasing relative change in PBF and with age. The median observed half-life of TCDD for these 337 veterans is 11.3 yr with a nonparametric 95% confidence interval of 10.0-14.1 yr.