Associations between follicular fluid trace elements and ovarian response during in vitro fertilization.

INTRODUCTION: Exposure to trace elements has been associated with ovarian response in experimental studies. We conducted a hypothesis-generating study of associations between ovarian follicular fluid (FF) trace elements and measures of ovarian response among women using in vitro fertilization (IVF). METHODS: We collected ovarian FF specimens from 56 women. We determined concentrations (µg/L) of 11 trace elements using inductively coupled plasma-tandem mass spectrometry. We estimated associations between women's FF trace elements per interquartile range difference, and measures of ovarian response using linear (peak estradiol (E2), baseline anti-mullerian hormone (AMH), and follicle stimulating hormone (FSH)) and negative binomial (baseline antral follicle count (AFC) and oocyte count) regression, adjusting for confounding factors. We used principal component analysis (PCA) to estimate the associations of the FF trace elements mixture. We also explored FF oxidative stress enzymes as causal mediators of the associations. RESULTS: Higher FF cobalt was associated with greater peak E2 (mean difference = 351.48 pg/mL; 95%CI: 21.76, 724.71) and AFC (rate ratio = 1.14; 95%CI: 1.01, 1.28), and higher FF copper was associated with greater peak E2 (mean difference = 335.66 pg/mL; 95%CI: 81.77, 753.08) and oocyte count (rate ratio = 1.19; 95%CI: 1.02, 1.43). Higher FF mercury was also associated with greater peak E2 (mean difference = 410.70 pg/mL; 95%CI: 61.90, 883.39). Higher FF lead was associated with lesser AFC (rate ratio = 0.85; 95%CI: 0.73, 0.98). Using PCA, the mixture of Sr, Hg, and As was associated with higher peak estradiol, AFC, and oocyte count. FF glutathione peroxidase, paraoxonase, and arylesterase activities were inconsistent mediators of the associations, but the effect estimates were imprecise. CONCLUSION: Our results suggest that essential and non-essential trace elements in FF were associated with ovarian response during IVF.

Variability of essential and non-essential trace elements in the follicular fluid of women undergoing in vitro fertilization (IVF).

Both essential and non-essential elements have been associated with female reproductive function in epidemiologic investigations, including among IVF populations. To date, most investigators have used blood or urine to assess biomarkers of exposure, with few employing ovarian follicular fluid (FF). FF may offer a more direct "snapshot" of the oocyte microenvironment than blood or urine, however previous studies report follicle-to-follicle variability in FF constituents that may contribute to exposure misclassification. Our objectives were to investigate sources of trace element variability, to estimate FF biomarker reliability among women undergoing IVF (n = 34), and to determine the minimum number of follicles required to estimate subject-specific mean concentrations. We measured As, Hg, Cd, Pb, Cu, Mn, Se, and Zn in FF samples using inductively coupled plasma tandem mass spectrometry. Inter-subject (between-women) variability contributed most of the variability in FF element concentrations, with ovarian, follicular, and analytical as smaller sources of variability. The proportion of variability attributable to sources between-follicles differed by age, body mass index (BMI), race, and cigarette smoking for Cu, Se, and Zn, by BMI and cigarette smoking for As, by primary infertility diagnosis for Hg, Cu, Se, and Zn, and by ovarian stimulation protocol for Mn and Se. Four to five individual follicles were sufficient to estimate subject-specific mean Cu, Se, and Zn concentrations, while >14 were necessary for As, Hg, Cd, Pb, and Mn. Overall, our results suggest that FF is a suitable source of biomarkers of As and Hg exposure in ovarian follicles. Although limited in size, our study offers the most comprehensive exploration of biological variation in FF trace elements to date and may provide guidance for future studies of ovarian trace element exposures.

Seafood consumption is associated with higher follicular fluid arsenic (As) and mercury (Hg) concentrations in women undergoing in vitro fertilization (IVF).

Human exposure to non-essential toxic metals such as cadmium (Cd), mercury (Hg), and lead (Pb), and metalloids such as arsenic (As) commonly occurs through diet. Toxic trace element exposures have been reported in association with fertility and fecundity in epidemiologic studies even at low to moderate levels. While most previous studies employed blood and urine biomarkers of exposure, few have assessed toxic trace elements in ovarian follicular fluid (FF), which surrounds the developing oocyte and hence may better reflect concentrations potentially affecting reproductive outcomes. Our objective was to identify dietary predictors of FF toxic trace elements in n = 56 women (mean age: 38.3 years) undergoing in vitro fertilization (IVF) at the University of California at San Francisco. We determined As, Hg, Cd, and Pb in 197 FF specimens, collected on the day of oocyte retrieval, using inductively coupled plasma tandem mass spectrometry. A comprehensive food frequency questionnaire was used to assess the weekly and annual dietary "patterns" of participants. Consumption of specific seafood items and turkey were correlated with individual FF toxic trace elements. We also found that each unit higher seafood consumption in the past week dominated by mollusks, shrimp, and bass was associated with 60% higher FF As (95% confidence interval (CI): 25%, 105%) and FF Hg (95%CI: 7%, 136%) concentrations. Higher annual seafood consumption dominated by urchin, crab, and trout was associated with 16% higher FF As (95%CI: -2%, 38%) and 31% higher FF Hg (95%CI: 7%, 60%) concentrations per unit intake. No associations were noted between diet and Cd and Pb levels in FF. Overall, our results suggest that higher seafood consumption contributes to elevated levels of As and Hg in FF. These findings are consistent with previous IVF studies that assessed toxic trace element exposures in blood and urine. To our knowledge, this is the first study to report that diet might be a source of As, Hg, Cd, and Pb in FF.

Preconception serum lipids and lipophilic micronutrient levels are associated with live birth rates after IVF.

RESEARCH QUESTION: Is a mixture of preconception serum lipids and lipophilic micronutrients associated with clinical pregnancy and live births? DESIGN: In this prospective cohort study, blood serum was collected on the day of oocyte retrieval for 180 women undergoing IVF at an academic reproductive health centre. Concentrations of lipids (phospholipids, total cholesterol, high- and low-density lipoproteins, and triglycerides) and lipophilic micronutrients (α-, δ-, and γ-tocopherols, retinol, ß- and α-carotenes, ß-cryptoxanthin, lutein and lycopene) were determined using diagnostic reagent kits and high-performance liquid chromatography. Poisson regression was used with robust variance estimation to evaluate changes in Z-scores for the mixture of serum lipid and lipophilic micronutrient concentrations as predictors of embryo implantation, clinical pregnancy and live birth, adjusted for age, body mass index (BMI), race, smoking status, infertility diagnosis, ovarian stimulation protocol and other measured lipid and lipophilic micronutrient concentrations. RESULTS: Each SD higher serum triglyceride concentration was associated with a lower chance of live birth (RR 0.54; 95% CI 0.33 to 0.90) whereas a 1 SD higher serum α-tocopherol concentration, as part of a mixture of serum lipids and lipophilic micronutrients, was associated with a higher likelihood for a live birth (RR 1.61; 95% CI 1.11 to 2.36). Serum ß-carotene concentrations were associated with live birth in a non-linear fashion; low ß-carotene was associated with a lower chance of live birth and high ß-carotene with a higher chance of live birth. CONCLUSION: Although components of a mixture of lipids and lipophilic micronutrients were associated with live birth outcomes after IVF, a larger investigation is necessary to fully evaluate the potential clinical implications.

Ultra-trace element analysis of human follicular fluid by ICP-MS/MS: pre-analytical challenges, contamination control, and matrix effects.

Follicular fluid (FF), which is the fluid that envelops the developing oocyte (egg cell) in the ovary, can be analyzed to assess trace element content as well as to determine potential exposure to toxic elements in women seeking in vitro fertilization (IVF) treatment. Such measurements may be useful in establishing associations with potential adverse effects on oocyte viability and subsequent pregnancy outcomes. The principal goal of this study was to leverage the next generation of inorganic mass spectrometry based on ICP-MS/MS to address the numerous analytical challenges of (ultra-)trace element analysis of human FF specimens. Ultra-trace element measurements are defined by the Clinical Laboratory Standards Institute as fluid concentrations below 10 µg L-1 or tissue mass fractions below 1 µg g-1. Stringent pre-analytical procedures were developed to minimize exogenous contamination during FF specimen collection and storage in a prospective study of 56 women seeking IVF treatment. ICP-MS/MS instrumental parameters were carefully optimized, and the method validated for 11 biologically important elements that included 4 at trace levels (Cu, Se, Sr, and Zn) and 7 at ultra-trace levels (As, Cd, Co, Mo, Mn, Hg, and Pb). Method limits of detection (LODs) for ultra-trace elements varied from 5.6 ng L-1 for Cd to 0.11 µg L-1 for Mo. A total of 197 human FF specimens were analyzed using the proposed ICP-MS/MS method with 84% of specimens detectable for Pb and 100% detectable for Co, Cu, Mn, Mo, Sr, and Zn. The method based on ICP-MS/MS was compared to a previous method developed for FF using SF-ICP-MS.

Decreased clinical pregnancy and live birth rates after short interval from delivery to subsequent assisted reproductive treatment cycle.

STUDY QUESTION: Does the interval from delivery to initiation of a subsequent ART treatment cycle impact clinical pregnancy or live birth rates? SUMMARY ANSWER: An interval from delivery to treatment start of <6 months or ≥24 months is associated with decreased likelihood of clinical pregnancy and live birth. WHAT IS KNOWN ALREADY: Short interpregnancy intervals are associated with poor obstetric outcomes in the naturally conceiving population prompting birth spacing recommendations of 18-24 months from international organizations. Deferring a subsequent pregnancy attempt means a woman will age in the interval with an attendant decline in her fertility. STUDY DESIGN, SIZE, DURATION: Retrospective analysis of the Society for Assisted Reproductive Technology Clinical Outcome Reporting System (SARTCORS) cohort containing 61 686 ART cycles from 2004 to 2013. PARTICIPANTS/MATERIALS, SETTING, METHODS: The delivery-to-cycle interval (DCI) was calculated for patients from SARTCORS with a history of live birth from ART who returned to the same clinic for a first subsequent treatment cycle. Generalized linear models were fit to determine the risk of clinical pregnancy and live birth by DCI with subsequent adjustment for factors associated with outcomes of interest. Predicted probabilities of clinical pregnancy and live birth were generated from each model. MAIN RESULTS AND THE ROLE OF CHANCE: A DCI of <6 months was associated with a 5.6% reduction in probability of clinical pregnancy (40.1 ± 1.9 versus 45.7 ± 0.6%, P = 0.009) and 6.8% reduction in live birth (31.6 ± 1.7 versus 38.4 ± 0.6%, P = 0.001) per cycle start compared to a DCI of 12 to <18 months. A DCI of ≥24 months was associated with a 5.1% reduction in probability of clinical pregnancy (40.6 ± 0.5 versus 45.7 ± 0.6%, P < 0.001) and 5.7% reduction in live birth (32.7 ± 0.5 versus 38.4 ± 0.6%, P < 0.001) compared to 12 to <18 months. LIMITATIONS, REASONS FOR CAUTION: The SART database is reliant upon self-report of many variables of interest including live birth. It remains unclear whether poorer outcomes are a result of residual confounding from factors inherent to the population with a very short or long DCI or the interval itself. WIDER IMPLICATIONS OF THE FINDINGS: Birth spacing recommendations for naturally conceiving populations may not be generally applicable to patients with a history of infertility. Patients planning ART treatment should wait a minimum of 6 months, but not more than 24 months, from a live birth for optimization of clinical pregnancy and live birth rates. STUDY FUNDING/COMPETING INTEREST(S): National Center for Advancing Translational Sciences, National Institutes of Health, UCSF-CTSI Grant number UL1TR001872. The authors have no competing interests.

A higher prevalence of endometriosis among Asian women does not contribute to poorer IVF outcomes.

PURPOSE: The purpose of the study was to determine whether diagnosis of endometriosis or endometriosis with endometrioma influences in vitro fertilization (IVF) outcomes in an ethnically diverse population. METHODS: Women undergoing a first IVF cycle (n = 717) between January 1, 2008 and December 31, 2009, at a university-affiliated infertility clinic, were retrospectively assessed for an endometriosis diagnosis. Differences in prevalence of endometriosis by ethnicity were determined, as well as differences in IVF success by ethnicity, with a focus on country of origin for Asian women. A multivariate model was generated to assess the relative contributions of country of origin and endometriosis to chance of clinical pregnancy with IVF. RESULTS: Endometriosis was diagnosed in 9.5% of participants; 3.5% also received a diagnosis of endometrioma. Endometriosis prevalence in Asian women was significantly greater than in Caucasians (15.7 vs. 5.8%, p < 0.01). Women of Filipino (p < 0.01), Indian (p < 0.01), Japanese (p < 0.01), and Korean (p < 0.05) origin specifically were more likely to have endometriosis than Caucasian women, although there was no difference in endometrioma presence by race/ethnicity. Oocyte quantity, embryo quality, and fertilization rates did not relate to endometriosis. Clinical pregnancy rates were significantly lower for Asian women, specifically in Indian (p < 0.05), Japanese (p < 0.05), and Korean (p < 0.05) women, compared to Caucasian women, even after controlling for endometriosis status. CONCLUSIONS: The prevalence of endometriosis appears to be higher in Filipino, Indian, Japanese, and Korean women presenting for IVF treatment than for Caucasian women; however, the discrepancy in IVF outcomes was conditionally independent of the presence of endometriosis. Future research should focus on improving pregnancy outcomes for Asian populations whether or not they are affected by endometriosis, specifically in the form of longitudinal studies where exposures can be captured prior to endometriosis diagnoses and infertility treatment.

Associations between IVF outcomes and essential trace elements measured in follicular fluid and urine: a pilot study.

PURPOSE: A hypothesis-generating pilot study exploring associations between essential trace elements measured in follicular fluid (FF) and urine and in vitro fertilization (IVF) endpoints. METHODS: We recruited 58 women undergoing IVF between 2007 and 2008, and measured cobalt, chromium, copper, manganese, molybdenum, and zinc in FF (n = 46) and urine (n = 45) by inductively coupled plasma mass spectrometry (ICP-MS). We used multivariable regression models to assess the impact of FF and urine trace elements on IVF outcomes, adjusted for age, body mass index, race, and cigarette smoking. RESULTS: Trace elements were mostly present at lower concentrations in FF than in urine. The average number of oocytes retrieved was positively associated with higher urine cobalt, chromium, copper, and molybdenum concentrations. FF chromium and manganese were negatively associated with the proportion of mature oocytes, yet urine manganese had a positive association. FF zinc was inversely associated with average oocyte fertilization. Urine trace elements were significant positive predictors for the total number of embryos generated. FF copper predicted lower embryo fragmentation while urine copper was associated with higher embryo cell number and urine manganese with higher embryo fragmentation. No associations were detected for implantation, pregnancy, or live birth. CONCLUSIONS: Our results suggest the importance of trace elements in both FF and urine for intermediate, although not necessarily clinical, IVF endpoints. The results differed using FF or urine biomarkers of exposure, which may have implications for the design of clinical and epidemiologic investigations. These initial findings will form the basis of a more definitive future study.

Variability in follicular fluid high density lipoprotein particle components measured in ipsilateral follicles.

PURPOSE: The purpose of this study was to examine the biological variability of follicular fluid (FF) high density lipoprotein (HDL) particle components measured in ipsilateral ovarian follicles. METHODS: We collected FF from two ipsilateral follicles among six women undergoing in vitro fertilization (IVF). We measured concentrations of 19 FF HDL particle components, including HDL cholesterol, free cholesterol, four cholesteryl esters, phospholipids, triglycerides, paraoxonase and arylesterase activities, apolipoproteins A-1 and A-2 (ApoA-1 and ApoA-2), and seven lipophilic micronutrients, by automated analysis and with high-performance liquid chromatography. We assessed biological variability using two-stage nested analysis of variance and compared values with those previously published for contralateral follicles. RESULTS: For most FF HDL analytes, there was little variability between follicles relative to the variability between women (i.e., %σ(2) F:%σ(2) B <0.5). Intraclass correlation coefficients were >0.80 for HDL cholesterol (0.82), phospholipids (0.89), paraoxonase (0.96), and arylesterase (0.91) activities, ApoA-1 (0.89), and ApoA-2 (0.90), and single specimen collections were required to estimate the subject-specific mean, demonstrating sufficient reliability for use as biomarkers of the follicular microenvironment in epidemiologic and clinical studies. CONCLUSIONS: These preliminary results raise the possibility for tighter regulation of HDL in follicles within the same ovary vs. between ovaries. Thus, collection of a single FF specimen may be sufficient to estimate HDL particle components concentrations within a single ovary. However, our results should be interpreted with caution as the analysis was based on a small sample.

Toxic metals in seminal plasma and in vitro fertilization (IVF) outcomes.

We measured toxic metals in seminal plasma collected from 30 men using vitro fertilization (IVF), to evaluate associations with semen quality and IVF outcomes. A doubling in Hg-adjusted Pb concentration was associated with 47% lower total motile sperm. Positive associations were suggested for Hg with pregnancy and live birth, adjusted for Cd or Pb. A negative association was suggested for Hg-adjusted Cd with pregnancy. These data add to evidence indicating that toxic metals impact IVF.

Urine cortisol concentration as a biomarker of stress is unrelated to IVF outcomes in women and men.

PURPOSE: Our primary objective was to assess associations between urine cortisol as a biomarker of psychological stress and in vitro fertilization (IVF) outcomes. A secondary objective was to assess associations between toxic metals and cortisol. METHODS: Urine and blood specimens were collected from 52 women and 28 male partners completing a first IVF procedure, on the day of oocyte retrieval. Urine cortisol was measured with an enzyme-linked immunosorbent assay. Mercury (Hg), cadmium (Cd), and lead (Pb) were determined in blood and Cd in urine by inductively coupled plasma-mass spectrometry. RESULTS: No associations were indicated for cortisol with IVF outcomes in multivariable regression models adjusted for covariates. However, we detected positive linear associations for cortisol and urine Cd (ß = 9.96, 95%CI 1.52, 21.44) and blood Hg (ß = 1.44, 95%CI 0.31, 3.18). An exploratory stratified analysis suggested a potential inverse association between urine cortisol and oocyte fertilization among women with low, but not high blood Hg. CONCLUSION: While limited, these preliminary data suggest that psychological stress may not play a major role in IVF outcomes, which therefore could be one less concern for couples and their clinicians. Our data also raise the possibility for toxic metals to modify associations between cortisol and IVF outcomes among women. However, these preliminary results require corroboration in an experimental animal model and confirmation in a larger, more definitive observational study.

Risk factors associated with endometriosis: importance of study population for characterizing disease in the ENDO Study.

OBJECTIVE: We sought to identify risk factors for endometriosis and their consistency across study populations in the Endometriosis: Natural History, Diagnosis, and Outcomes (ENDO) Study. STUDY DESIGN: In this prospective matched, exposure cohort design, 495 women aged 18-44 years undergoing pelvic surgery (exposed to surgery, operative cohort) were compared to an age- and residence-matched population cohort of 131 women (unexposed to surgery, population cohort). Endometriosis was diagnosed visually at laparoscopy/laparotomy or by pelvic magnetic resonance imaging in the operative and population cohorts, respectively. Logistic regression estimated the adjusted odds ratios (AORs) and 95% confidence intervals (CIs) for each cohort. RESULTS: The incidence of visualized endometriosis was 40% in the operative cohort (11.8% stage 3-4 by revised criteria from the American Society for Reproductive Medicine), and 11% stage 3-4 in the population cohort by magnetic resonance imaging. An infertility history increased the odds of an endometriosis diagnosis in both the operative (AOR, 2.43; 95% CI, 1.57-3.76) and population (AOR, 7.91; 95% CI, 1.69-37.2) cohorts. In the operative cohort only, dysmenorrhea (AOR, 2.46; 95% CI, 1.28-4.72) and pelvic pain (AOR, 3.67; 95% CI, 2.44-5.50) increased the odds of diagnosis, while gravidity (AOR, 0.49; 95% CI, 0.32-0.75), parity (AOR, 0.42; 95% CI, 0.28-0.64), and body mass index (AOR, 0.95; 95% CI, 0.93-0.98) decreased the odds of diagnosis. In all sensitivity analyses for different diagnostic subgroups, infertility history remained a strong risk factor. CONCLUSION: An infertility history was a consistent risk factor for endometriosis in both the operative and population cohorts of the ENDO Study. Additionally, identified risk factors for endometriosis vary based upon cohort selection and diagnostic accuracy. Finally, endometriosis in the population may be more common than recognized.

Bisphenol-A (BPA), BPA glucuronide, and BPA sulfate in midgestation umbilical cord serum in a northern and central California population.

Bisphenol-A (BPA) is an endocrine disrupting chemical used in numerous consumer products, resulting in universal exposure in the United States. Prenatal exposure to BPA is associated with numerous reproductive and developmental effects in animals. However, little is known about human fetal exposure or metabolism of BPA during midgestation. In the present study, we present a new liquid chromatography-tandem mass spectrometry method to directly measure concentrations of BPA and two predominant metabolic conjugates-BPA glucuronide and BPA sulfate-in umbilical cord serum collected from elective second trimester pregnancy terminations. We detected at least one form of BPA in all umbilical cord serum samples: BPA (GM 0.16, range

Exploratory cohort study into underlying mechanism of differences in estrogen metabolism between Asian and Caucasian women during assisted reproductive technology treatment.

Objective: Explore whether racial differences in prevalence of CYP1A2∗1F polymorphism underlies estrogen metabolism differences among Asians and Caucasians. Design: Prospective cohort study. Setting: University-based fertility practice. Patients: Asian or Caucasian patients who underwent ovarian stimulation (OS) or programmed cycle frozen embryo transfer (FET) between October 2019 and April 2021. Interventions: None. Main Outcome Measures: Trigger-day serum E2 per oocyte retrieved in OS cycles, and E2 on day of lining check in FET cycles. Results: Seventy-one participants were enrolled, 55 in OS group (29 Caucasian and 26 Asian) and 16 in FET group (10 Caucasian and 6 Asian). Peak E2 per oocyte retrieved in the OS group (n = 48) differed by race, with significantly lower levels in Caucasians compared with Asians (177.5 ± 64.2 vs. 261.1 ± 139.5 pg/mL). Prevalence of CYP1A2∗1F polymorphism did not significantly differ by race. Compared using Kruskal-Wallis test, peak E2 per oocyte retrieved did not differ by CYP1A2∗1F genotype. In multivariate linear regression model, adjusting for body mass index, caffeine intake, and self-reported race, there remained no significant correlation. In FET group, serum E2 on day of lining check was also not significantly different by CYP1A2∗1F genotype. Conclusions: Although a consistent difference in serum E2 between Asians and Caucasians undergoing OS was noted, the CYP1A2∗1F polymorphism is unlikely the primary driver of this difference.

Perfluorochemicals and endometriosis: the ENDO study.

BACKGROUND: Environmental chemicals may be associated with endometriosis. No published research has focused on the possible role of perfluorochemicals (PFCs) despite their widespread presence in human tissues. METHODS: We formulated two samples. The first was an operative sample comprising 495 women aged 18-44 years scheduled for laparoscopy/laparotomy at one of 14 participating clinical sites in the Salt Lake City or San Francisco area, 2007-2009. The second was a population-based sample comprising 131 women matched to the operative sample on age and residence within a 50-mile radius of participating clinics. Interviews and anthropometric assessments were conducted at enrollment, along with blood collection for the analysis of nine PFCs, which were quantified using liquid chromatography-tandem mass spectrometry. Endometriosis was defined based on surgical visualization (in the operative sample) or magnetic resonance imaging (in the population sample). Using logistic regression, we estimated odds ratios (ORs) and 95% confidence intervals (CIs) for each PFC (log-transformed), adjusting for age and body mass index, and then parity. RESULTS: Serum perfluorooctanoic acid (PFOA; OR = 1.89 [95% CI = 1.17-3.06]) and perfluorononanoic acid (2.20 [1.02-4.75]) were associated with endometriosis in the operative sample; findings were moderately attenuated with parity adjustment (1.62 [0.99-2.66] and 1.99 [0.91-4.33], respectively). Perfluorooctane sulfonic acid (1.86 [1.05-3.30]) and PFOA (2.58 [1.18-5.64]) increased the odds for moderate/severe endometriosis, although the odds were similarly attenuated with parity adjustment (OR = 1.50 and 1.86, respectively). CONCLUSIONS: Select PFCs were associated with an endometriosis diagnosis. These associations await corroboration.

DNA methylation changes in whole blood is associated with exposure to the environmental contaminants, mercury, lead, cadmium and bisphenol A, in women undergoing ovarian stimulation for IVF.

BACKGROUND: Changes in DNA methylation may play an important role in the deleterious reproductive effects reported in association with exposure to environmental pollutants. In this pilot study, we identify candidate methylation changes associated with exposure to pollutants in women undergoing in vitro fertilization (IVF). METHODS: Blood and urine were collected from women on the day of oocyte retrieval. Whole blood was analyzed for mercury and lead, and urine for cadmium using inductively coupled plasma mass spectrometry. Unconjugated bisphenol A (BPA) was analyzed in serum using high-performance liquid chromatography with Coularray detection. Participants were dichotomized as higher or lower exposure groups by median concentrations. Using the Illumina GoldenGate Methylation Cancer Panel I, DNA methylation in whole blood from 43 women was assessed at 1505 CpG sites for association with exposure levels of each pollutant. Candidate CpG sites were identified using a Diff Score >|13| (P< 0.05) and an absolute difference >10% which were confirmed using bisulfite pyrosequencing. RESULTS: Methylation of the GSTM1/5 promoter was increased for women with higher mercury exposure (P= 0.04); however, no correlation was observed (r= 0.17, P= 0.27). Reduced methylation was detected in the COL1A2 promoter in women with higher exposure to lead (P= 0.004), and an inverse correlation was observed (r = - 0.45, P= 0.03). Lower methylation of a promoter CpG site at the TSP50 gene was detected in women with higher BPA exposure (P= 0.005), and again an inverse correlation was identified (r = - 0.51, P= 0.001). CONCLUSIONS: Altered DNA methylation at various CpG sites was associated with exposure to mercury, lead or BPA, providing candidates to be investigated using a larger study sample, as the results may reflect an independently associated predictor (e.g. socioeconomic status, diet, genetic variants, altered blood cell composition). Further studies accommodating variations in these factors will be needed to confirm these associations and identify their underlying causes.

Meiotic recombination in human oocytes.

Studies of human trisomies indicate a remarkable relationship between abnormal meiotic recombination and subsequent nondisjunction at maternal meiosis I or II. Specifically, failure to recombine or recombination events located either too near to or too far from the centromere have been linked to the origin of human trisomies. It should be possible to identify these abnormal crossover configurations by using immunofluorescence methodology to directly examine the meiotic recombination process in the human female. Accordingly, we initiated studies of crossover-associated proteins (e.g., MLH1) in human fetal oocytes to analyze their number and distribution on nondisjunction-prone human chromosomes and, more generally, to characterize genome-wide levels of recombination in the human female. Our analyses indicate that the number of MLH1 foci is lower than predicted from genetic linkage analysis, but its localization pattern conforms to that expected for a crossover-associated protein. In studies of individual chromosomes, our observations provide evidence for the presence of "vulnerable" crossover configurations in the fetal oocyte, consistent with the idea that these are subsequently translated into nondisjunctional events in the adult oocyte.

Associations between toxic metals in follicular fluid and in vitro fertilization (IVF) outcomes.

PURPOSE: We previously reported associations between trace concentrations of Hg, Cd and Pb in blood and urine and reproductive outcomes for women undergoing in-vitro fertilization (IVF). Here we assess measurements in single follicular fluid (FF) specimens from 46 women as a presumably more relevant marker of dose for reproductive toxicity. METHODS: FF specimens were analyzed for Hg, Cd and Pb using sector field-inductively coupled plasma-mass spectrometry (SF-ICP-MS). Variability sources were assessed by nested ANOVA. Multivariable regression was used to evaluate associations for square root transformed metals with IVF outcomes, adjusting for confounders. RESULTS: An inverse association is detected for FF Pb and fertilization (relative risk (RR) = 0.68, P = 0.026), although positive for Cd (RR = 9.05, P = 0.025). While no other statistically significant associations are detected, odds ratios (OR) are increased for embryo cleavage with Hg (OR = 3.83, P = 0.264) and Cd (OR = 3.18, P = 0.644), and for embryo fragmentation with Cd (OR = 4.08, P = 0.586) and Pb (OR = 2.22, P = 0.220). Positive estimates are observed for Cd with biochemical (RR = 19.02, P = 0.286) and clinical pregnancies (RR = 38.80, P = 0.212), yet with very low precision. CONCLUSIONS: We have identified associations between trace amounts of Pb and Cd in FF from a single follicle, and oocyte fertilization. Yet, the likelihood of biological variation in trace element concentrations within and between follicles, coupled with levels that are near the limits of detection suggest that future work should examine multiple follicles using a 'one follicle-one oocyte/embryo' approach. A larger study is merited to assess more definitively the role that these environmental factors could play with respect to egg quality in IVF programs.

Female obesity adversely affects assisted reproductive technology (ART) pregnancy and live birth rates.

BACKGROUND: Obesity has risen among women in the USA, including those seeking infertility treatments. In 2007, height and weight were added to the Society for Assisted Reproductive Technology Clinic Online Reporting System (SART CORS), permitting calculation of BMI (weight/height(2)) for the first time using this national dataset. METHODS: The SART CORS was used to evaluate the odds of failure to achieve a clinical intrauterine pregnancy and failure to achieve a live birth by the woman's age, BMI and oocyte source (autologous versus donor), controlling for race and ethnicity, day of embryo transfer, number of embryos transferred and infertility diagnoses. The reference population was women with normal BMI. RESULTS: There were 45 163 ART embryo transfers where maternal height and weight were recorded. Increasing obesity was associated with a significant rise in failure to achieve a clinical pregnancy with the use of autologous oocytes (P< 0.0001), but no difference with the use of donor oocytes. Among women using autologous oocytes who did conceive, failure to achieve a live birth increased with increasing obesity, to a greater extent among women <35 years of age. CONCLUSIONS: Higher BMI is associated with an increased failure to achieve a clinical intrauterine gestation; this risk was overcome with the use of donor oocytes. Failure to achieve a live birth increases with higher BMI, significantly with the use of autologous oocytes (P< 0.0001), and to a greater extent among women <35 years of age (P< 0.0001).

Biomonitoring for exposure to multiple trace elements via analysis of urine from participants in the study of metals and assisted reproductive technologies (SMART).

Humans are exposed to concentrations of multiple trace elements through a variety of background sources; many are suspected reproductive toxicants. Prior to investigating associations between trace elements and human reproductive health, potential biomarkers of exposure should be characterized by sources of variability in the population at risk. Factors influencing elemental exposure should also be identified to ensure their consideration as potential confounding variables. The principal aim of this study is to characterize sources of variability for 19 trace elements measured in urine specimens collected from 55 women and 36 male partners completing a 1st cycle of in vitro fertilization (IVF). Urine specimens were analyzed using a biomonitoring method based on inductively coupled plasma-mass spectrometry (ICP-MS). Randomly selected urine specimens (∼6%) were analyzed in duplicate, and these data were used to characterize sources of variability. Nine trace elements including As, Ba, Cd, Cs, Co, Cu, Mn, Mo, and Zn, were quantified in most specimens, indicating their utility in future epidemiologic studies of trace elements exposure and IVF outcomes. With few exceptions, normalizing urine using the traditional creatinine-correction procedure, or an alternative approach based on a linear regression model, increased residual variability only slightly. Sex and race appear to be important factors to consider in epidemiologic studies conducted in this population. Urine concentrations for most elements are similar to those reported in the 2005-2006 National Health and Nutrition Examination Survey (NHANES); however, differences in others may indicate regional trends or a unique exposure history for this infertile study population.