Germaine M. Buck Louis

Lipid Adjustment in the Analysis of Environmental Contaminants and Human Health Risks

The literature on exposure to lipophilic agents such as polychlorinated biphenyls (PCBs) is conflicting, posing challenges for the interpretation of potential human health risks. Laboratory variation in quantifying PCBs may account for some of the conflicting study results. For example, for quantification purposes, blood is often used as a proxy for adipose tissue, which makes it necessary to model serum lipids when assessing health risks of PCBs. Using a simulation study, we evaluated four statistical models (unadjusted, standardized, adjusted, and two-stage) for the analysis of PCB exposure, serum lipids, and health outcome risk (breast cancer). We applied eight candidate true causal scenarios, depicted by directed acyclic graphs, to illustrate the ramifications of misspecification of underlying assumptions when interpreting results. Statistical models that deviated from underlying causal assumptions generated biased results. Lipid standardization, or the division of serum concentrations by serum lipids, was observed to be highly prone to bias. We conclude that investigators must consider biology, biologic medium (e.g., nonfasting blood samples), laboratory measurement, and other underlying modeling assumptions when devising a statistical plan for assessing health outcomes in relation to environmental exposures.

Environmental Factors and Puberty Timing: Expert Panel Research Needs

Serono Symposia International convened an expert panel to review the impact of environmental influences on the regulation of pubertal onset and progression while identifying critical data gaps and future research priorities. An expert panel reviewed the literature on endocrine-disrupting chemicals, body size, and puberty. The panel concluded that available experimental animal and human data support a possible role of endocrine-disrupting chemicals and body size in relation to alterations in pubertal onset and progression in boys and girls. Critical data gaps prioritized for future research initiatives include (1) etiologic research that focus on environmentally relevant levels of endocrine-disrupting chemicals and body size in relation to normal puberty as well as its variants, (2) exposure assessment of relevant endocrine-disrupting chemicals during critical windows of human development, and (3) basic research to identify the primary signal(s) for the onset of gonadotropin-releasing hormone–dependent/central puberty and gonadotropin-releasing hormone–independent/peripheral puberty. Prospective studies of couples who are planning pregnancies or pregnant women are needed to capture the continuum of exposures at critical windows while assessing a spectrum of pubertal markers as outcomes. Coupled with comparative species studies, such research may provide insight regarding the causal ordering of events that underlie pubertal onset and progression and their role in the pathway of adult-onset disease.

Male Reproductive Disorders and Fertility Trends: Influences of Environment and Genetic Susceptibility.

It is predicted that Japan and European Union will soon experience appreciable decreases in their populations due to persistently low total fertility rates (TFR) below replacement level (2.1 child per woman). In the United States, where TFR has also declined, there are ethnic differences. Caucasians have rates below replacement, while TFRs among African-Americans and Hispanics are higher. We review possible links between TFR and trends in a range of male reproductive problems, including testicular cancer, disorders of sex development, cryptorchidism, hypospadias, low testosterone levels, poor semen quality, childlessness, changed sex ratio, and increasing demand for assisted reproductive techniques. We present evidence that several adult male reproductive problems arise in utero and are signs of testicular dysgenesis syndrome (TDS). Although TDS might result from genetic mutations, recent evidence suggests that it most often is related to environmental exposures of the fetal testis. However, environmental factors can also affect the adult endocrine system. Based on our review of genetic and environmental factors, we conclude that environmental exposures arising from modern lifestyle, rather than genetics, are the most important factors in the observed trends. These environmental factors might act either directly or via epigenetic mechanisms. In the latter case, the effects of exposures might have an impact for several generations post-exposure. In conclusion, there is an urgent need to prioritize research in reproductive physiology and pathophysiology, particularly in highly industrialized countries facing decreasing populations. We highlight a number of topics that need attention by researchers in human physiology, pathophysiology, environmental health sciences, and demography.

Designing prospective cohort studies for assessing reproductive and developmental toxicity during sensitive windows of human reproduction and development--the LIFE Study.

The relationship between the environment and human fecundity and fertility remains virtually unstudied from a couple-based perspective in which longitudinal exposure data and biospecimens are captured across sensitive windows. In response, we completed the LIFE Study with methodology that intended to empirically evaluate a priori purported methodological challenges: implementation of population-based sampling frameworks suitable for recruiting couples planning pregnancy; obtaining environmental data across sensitive windows of reproduction and development; home-based biospecimen collection; and development of a data management system for hierarchical exposome data. We used two sampling frameworks (i.e., fish/wildlife licence registry and a direct marketing database) for 16 targeted counties with presumed environmental exposures to persistent organochlorine chemicals to recruit 501 couples planning pregnancies for prospective longitudinal follow-up while trying to conceive and throughout pregnancy. Enrolment rates varied from <1% of the targeted population (n = 424,423) to 42% of eligible couples who were successfully screened; 84% of the targeted population could not be reached, while 36% refused screening. Among enrolled couples, ∼ 85% completed daily journals while trying; 82% of pregnant women completed daily early pregnancy journals, and 80% completed monthly pregnancy journals. All couples provided baseline blood/urine samples; 94% of men provided one or more semen samples and 98% of women provided one or more saliva samples. Women successfully used urinary fertility monitors for identifying ovulation and home pregnancy test kits. Couples can be recruited for preconception cohorts and will comply with intensive data collection across sensitive windows. However, appropriately sized sampling frameworks are critical, given the small percentage of couples contacted found eligible and reportedly planning pregnancy at any point in time.

The relationship between male BMI and waist circumference on semen quality: data from the LIFE study

Michael L. Eisenberg1,*, Sungduk Kim2, Zhen Chen2, Rajeshwari Sundaram2, Enrique F. Schisterman2, and Germaine M. Buck Louis2 Departments of Urology, Obstetrics and Gynecology, Stanford University School of Medicine, 300 Pasteur Drive, Stanford, CA 94305-5118, USA Division of Intramural Population Health Research, Eunice Kennedy Shriver National Institute of Child Health and Human Development, 6100 Executive Blvd., Room 7B03, Rockville, MD 20852, USA

Racial/ethnic standards for fetal growth: The NICHD Fetal Growth Studies

OBJECTIVE Fetal growth is associated with long-term health yet no appropriate standards exist for the early identification of undergrown or overgrown fetuses. We sought to develop contemporary fetal growth standards for 4 self-identified US racial/ethnic groups. STUDY DESIGN We recruited for prospective follow-up 2334 healthy women with low-risk, singleton pregnancies from 12 community and perinatal centers from July 2009 through January 2013. The cohort comprised: 614 (26%) non-Hispanic whites, 611 (26%) non-Hispanic blacks, 649 (28%) Hispanics, and 460 (20%) Asians. Women were screened at 8w0d to 13w6d for maternal health status associated with presumably normal fetal growth (aged 18-40 years; body mass index 19.0-29.9 kg/m(2); healthy lifestyles and living conditions; low-risk medical and obstetrical history); 92% of recruited women completed the protocol. Women were randomized among 4 ultrasonography schedules for longitudinal fetal measurement using the Voluson E8 (GE Healthcare, Milwaukee, WI). In-person interviews and anthropometric assessments were conducted at each visit; medical records were abstracted. The fetuses of 1737 (74%) women continued to be low risk (uncomplicated pregnancy, absent anomalies) at birth, and their measurements were included in the standards. Racial/ethnic-specific fetal growth curves were estimated using linear mixed models with cubic splines. Estimated fetal weight (EFW) and biometric parameter percentiles (5th, 50th, 95th) were determined for each gestational week and comparisons made by race/ethnicity, with and without adjustment for maternal and sociodemographic factors. RESULTS EFW differed significantly by race/ethnicity >20 weeks. Specifically at 39 weeks, the 5th, 50th, and 95th percentiles were 2790, 3505, and 4402 g for white; 2633, 3336, and 4226 g for Hispanic; 2621, 3270, and 4078 g for Asian; and 2622, 3260, and 4053 g for black women (adjusted global P < .001). For individual parameters, racial/ethnic differences by order of detection were: humerus and femur lengths (10 weeks), abdominal circumference (16 weeks), head circumference (21 weeks), and biparietal diameter (27 weeks). The study-derived standard based solely on the white group erroneously classifies as much as 15% of non-white fetuses as growth restricted (EFW <5th percentile). CONCLUSION Significant differences in fetal growth were found among the 4 groups. Racial/ethnic-specific standards improve the precision in evaluating fetal growth.

Persistent Environmental Pollutants and Couple Fecundity: The LIFE Study

Background: Evidence suggesting that persistent environmental pollutants may be reproductive toxicants underscores the need for prospective studies of couples for whom exposures are measured. Objectives: We examined the relationship between selected persistent pollutants and couple fecundity as measured by time to pregnancy. Methods: A cohort of 501 couples who discontinued contraception to become pregnant was prospectively followed for 12 months of trying to conceive or until a human chorionic gonadotrophin (hCG) test confirmed pregnancy. Couples completed daily journals on lifestyle and provided biospecimens for the quantification of 9 organochlorine pesticides, 1 polybrominated biphenyl, 10 polybrominated diphenyl ethers, 36 polychlorinated biphenyls (PCBs), and 7 perfluorochemicals (PFCs) in serum. Using Cox models for discrete time, we estimated fecundability odds ratios (FORs) and 95% CIs separately for each partner’s concentrations adjusting for age, body mass index, serum cotinine, serum lipids (except for PFCs), and study site (Michigan or Texas); sensitivity models were further adjusted for left truncation or time off of contraception (≤ 2 months) before enrollment. Results: The adjusted reduction in fecundability associated with standard deviation increases in log-transformed serum concentrations ranged between 18% and 21% for PCB congeners 118, 167, 209, and perfluorooctane sulfonamide in females; and between 17% and 29% for p,p´-DDE and PCB congeners 138, 156, 157, 167, 170, 172, and 209 in males. The strongest associations were observed for PCB 167 (FOR 0.79; 95% CI: 0.64, 0.97) in females and PCB 138 (FOR = 0.71; 95% CI: 0.52, 0.98) in males. Conclusions: In this couple-based prospective cohort study with preconception enrollment and quantification of exposures in both female and male partners, we observed that a subset of persistent environmental chemicals were associated with reduced fecundity.

Validity of self-reported time to pregnancy.

Background: The reliability of retrospective time to pregnancy (TTP) has been established, but its validity has been assessed in only 1 study, which had a short follow-up. Methods: Ninety-nine women enrolled a decade earlier in a prospective TTP study were queried by means of mailed questionnaires about the duration of time they had required to become pregnant. Their responses were compared with their earlier data from daily diaries (gold standard). Results: One-third of women could not recall their earlier TTP either in menstrual cycles or calendar months. Only 17%–19% of women recalled their TTP exactly. Agreement increased to 41%–51%, 65%–72%, and 72%–77% when defined as ±1, ±2, and ±3 months, respectively. Women with longer observed TTPs or previous pregnancies were more likely to under-report their TTP. Conclusions: The findings raise questions about the commonly assumed validity of self-reported TTP. Recalled TTP may introduce error when estimating fecundability or classifying couples’ fecundity status.

Association between lead and cadmium and reproductive hormones in peripubertal U.S. girls.

Background Lead (Pb) and cadmium (Cd) are known reproductive toxicants thought to disrupt hormone production throughout sensitive developmental windows, although this has not been previously examined in nationally representative peripubertal children. Objectives We examined the association between blood Pb and urinary Cd concentrations and the reproductive hormones inhibin B and luteinizing hormone (LH) in girls 6–11 years of age who participated in the cross-sectional Third National Health and Nutrition Examination Survey (NHANES III) (1988–1994). Methods Pb (micrograms per deciliter) was measured in whole blood, and Cd was measured in urine (nanograms per milliliter). Inhibin B (picograms per milliliter) and LH (milli–International units per milliliter) were measured in residual sera for 705 girls. Survey logistic regression was used to estimate associations with pubertal onset based on inhibin B concentration > 35 pg/mL or LH concentration > 0.4 mIU/mL, and multinomial logistic regression was used to estimate the association between Pb and increasing categories of hormone concentrations. Results High Pb (≥ 5 μg/dL) was inversely associated with inhibin B > 35 pg/mL [odds ratio (OR) = 0.26; 95% confidence interval (CI), 0.11–0.60; compared with Pb < 1 μg/dL]. At 10 and 11 years of age, girls with low Pb (< 1 μg/dL) had significantly higher inhibin B than did girls with moderate (1–4.99 μg/dL) or high Pb (≥ 5 μg/dL). In the subsample of 260 girls with levels of inhibin B above the level of detection and using survey regression modeling, inhibin B levels were lower among girls with both high Pb and high Cd (β = −0.52; 95% CI, −0.09 to −1.04) than among girls with high Pb alone (β = −0.35; 95% CI, −0.13 to −0.57), relative to girls with low Pb and low Cd. Conclusions Higher Pb was inversely associated with inhibin B, a marker of follicular development, and estimated effects suggestive of pubertal delays appeared to be stronger in the context of higher Cd concentrations. These data underscore the importance of Pb and Cd as reproductive toxicants for young girls.

Analysis of repeated pregnancy outcomes

Women tend to repeat reproductive outcomes, with past history of an adverse outcome being associated with an approximate two-fold increase in subsequent risk. These observations support the need for statistical designs and analyses that address this clustering. Failure to do so may mask effects, result in inaccurate variance estimators, produce biased or inefficient estimates of exposure effects. We review and evaluate basic analytic approaches for analysing reproductive outcomes, including ignoring reproductive history, treating it as a covariate or avoiding the clustering problem by analysing only one pregnancy per woman, and contrast these to more modern approaches such as generalized estimating equations with robust standard errors and mixed models with various correlation structures. We illustrate the issues by analysing a sample from the Collaborative Perinatal Project dataset, demonstrating how the statistical model impacts summary statistics and inferences when assessing etiologic determinants of birth weight.