Jason G. Bromer

Bisphenol-A exposure in utero leads to epigenetic alterations in the developmental programming of uterine estrogen response

Bisphenol‐A (BPA) is a nonsteroidal estrogen that is ubiquitous in the environment. The homeobox gene Hoxa10 controls uterine organogenesis, and its expression is affected by in utero BPA exposure. We hypothesized that an epigenetic mechanism underlies BPA‐mediated alterations in Hoxa10 expression. We analyzed the expression pattern and methylation profile of Hoxa10 after in utero BPA exposure. Pregnant CD‐1 mice were treated with BPA (5 mg/kg IP) or vehicle control on d 9–16 of pregnancy. Hoxa10 mRNA and protein expression were increased by 25% in the reproductive tract of mice exposed in utero. Bisulfite sequencing revealed that cytosine‐guanine dinucleotide methylation was decreased from 67 to 14% in the promoter and from 71 to 3% in the intron of Hoxa10 after in utero BPA exposure. Decreased DNA methylation led to an increase in binding of ER‐α to the Hoxa10 ERE both in vitro as and in vivo as determined by EMSA and chromatin immunoprecipitation, respectively. Diminished methylation of the ERE‐containing promoter sequence resulted in an increase in ERE‐driven gene expression in reporter assays. We identify altered methylation as a novel mechanism of BPA‐induced altered developmental programming. Permanent epigenetic alteration of ERE sensitivity to estrogen may be a general mechanism through which endocrine disruptors exert their action.—Bromer, J. G., Zhou, Y., Taylor, M. B., Doherty, L., Taylor, H. S.. Bisphenol‐A exposure in utero leads to epigenetic alterations in the developmental programming of uterine estrogen response. FASEBJ. 24, 2273–2280 (2010). www.fasebj.org

In utero exposure to diethylstilbestrol (DES) or bisphenol-A (BPA) increases EZH2 expression in the mammary gland: an epigenetic mechanism linking endocrine disruptors to breast cancer.

Diethylstilbestrol (DES) and bisphenol-A (BPA) are estrogen-like endocrine-disrupting chemicals that induce persistent epigenetic changes in the developing uterus. However, DES exposure in utero is also associated with an increased risk of breast cancer in adult women. Similarly, fetal exposure to BPA induces neoplastic changes in mammary tissue of mice. We hypothesized that epigenetic alterations would precede the increased risk of breast neoplasia after in utero exposure to endocrine disruptors. Enhancer of Zeste Homolog 2 (EZH2) is a histone methyltransferase that has been linked to breast cancer risk and epigenetic regulation of tumorigenesis. We examined the effect of BPA and DES on EZH2 expression and function in MCF-7 cells and in mammary glands of mice exposed in utero. DES and BPA treatment approximated human exposure. EZH2 functional activity was assessed by measuring histone H3 trimethylation. Treatment of MCF-7 cells with DES or BPA led to a 3- and 2-fold increase in EZH2 mRNA expression, respectively (p < 0.05) as well as increased EZH2 protein expression. Mice exposed to DES in utero showed a >2-fold increase in EZH2 expression in adult mammary tissue compared with controls (p < 0.05). EZH2 protein was elevated in mammary tissue of mice exposed to DES or BPA. Histone H3 trimethylation was increased in MCF-7 cells treated with BPA or DES. Similarly, mice exposed to BPA or DES in utero showed increased mammary histone H3 trimethylation. Developmental programming of EZH2 is a novel mechanism by which in utero exposure to endocrine disruptors leads to epigenetic regulation of the mammary gland.

Assessment of embryo viability in assisted reproductive technology: shortcomings of current approaches and the emerging role of metabolomics.

Purpose of review The present article describes the current status of embryo assessment in assisted reproductive technologies and discusses two important issues that derive, at least in part, from our inability to adequately assess the reproductive potential of individual embryos: low implantation rates and high multiple pregnancy rates. After an overview of studies evaluating embryo metabolism as a predictor of embryo viability, use of metabolomics and additional emerging approaches for rapid, noninvasive, embryo assessment are discussed. Recent findings Morphology and cleavage rate remain the mainstay of embryo assessment. A number of technologies are, however, under investigation. These include the assessment of glucose, lactate, pyruvate, and amino acid metabolism, proteomic profiling, evaluation of oxygen consumption, and most recently, examination of the metabolome. Summary As the number of assisted reproductive technology cycles increases worldwide, improvements in the ability to quickly and noninvasively determine the best embryos for transfer remain a critical goal for reproductive medicine. Recent studies suggest that metabolomic profiling of embryo culture media may provide a useful adjunct to the current embryo assessment strategies based on morphology and cleavage rate.

Insurance coverage and in vitro fertilization outcomes: a U.S. perspective

OBJECTIVE To compare the impact of mandated IVF insurance coverage on ET practices and resulting multiple pregnancy rates. DESIGN Retrospective analysis of all fresh, nondonor IVF cycles performed in the United States in 2006. SETTING United States. PATIENT(S) A total of 91,753 fresh, nondonor IVF cycles in the United States. INTERVENTION(S) None. MAIN OUTCOME MEASURE(S) Pregnancy rate, live-birth rate, embryos transferred, multiple pregnancy rate. RESULT(S) Overall, nonmandated states had a significantly higher pregnancy rate (38.8% vs. 35%) and live-birth rate (32.2% vs. 29.1%) than mandated states. Nonmandated states also had a significantly higher twin rate (28.1% vs. 26%) and triplet rate (3.9% vs. 3.4%). The average number of embryos transferred was also significantly higher in nonmandated states (2.6 vs. 2.2). These findings were more pronounced in the <35 and 35-37 age groups. CONCLUSION(S) In the last 8 years, despite a reduction in the average number of embryos transferred and multiple pregnancy rates, there is a continued association between mandated IVF coverage, the transfer of fewer embryos, and lower rates of multiple pregnancies and births, particularly in the younger age groups.

Preterm deliveries that result from multiple pregnancies associated with assisted reproductive technologies in the USA: a cost analysis.

Purpose of review Simultaneous transfer of multiple embryos in an assisted reproductive technology (ART) cycle results in a high rate of multiple pregnancy. Besides the medical complications associated with multiple pregnancy, the financial burden of the resultant preterm infants is also substantial. The current review evaluates the costs associated with the care of preterm infants that are born as a result of ART-associated multiple pregnancies. Recent findings In 2006, 30% of all ART live births were multiple infant deliveries in the USA. This resulted in 48% of all ART neonates being the product of a multiple infant delivery. In the same year, 62% of ART twins and 97% of ART triplets were delivered preterm, corresponding to approximately 17 000 infants. The Board of Health Sciences Policy has estimated the mean cost of each preterm infant to be US

Defining the proliferative phase endometrial defect

51 600. Therefore, the financial burden of ART-associated preterm deliveries is estimated to be approximately US

Fertility Preservation: The Rationale for Cryopreservation of the Whole Ovary

1 billion annually. This figure has remained essentially unchanged between 2001 and 2006, despite decreasing number of embryos transferred, due to increasing total number of ART cycles performed. Summary Preterm deliveries that result from ART-associated multiple pregnancies add a substantial burden to overall US healthcare expenditure annually. Policies limiting the number of embryos transferred should be considered with a perspective to increase elective single embryo transfers.

Live babies born per oocyte retrieved in a subpopulation of oocyte donors with repetitive reproductive success

OBJECTIVE To evaluate proliferative phase endometrial development in a heterogeneous infertility population. DESIGN Retrospective study. SETTING University-based infertility practice. PATIENT(S) Two hundred forty-six treatment cycles. INTERVENTION(S) Clomiphene citrate or FSH ovarian stimulation, followed by IUI or IVF. MAIN OUTCOME MEASURE(S) Endometrial thickness according to transvaginal ultrasonography; clinical pregnancy rate. RESULT(S) Endometrial growth began from a nadir of approximately 4.5 mm on cycle day 4 and increased linearly to a plateau of approximately 10 mm on cycle day 9. This same pattern was observed in all cycles, regardless of pregnancy, drug, or underlying diagnosis. Follicle-stimulating hormone-stimulated cycles showed a significantly increased endometrial thickness compared with clomiphene citrate cycles (10.1 vs. 8.3 mm). Maximum endometrial thickness achieved showed a correlation with age, body mass index, and maximum E(2) level. Subjects who carried a primary diagnosis of polycystic ovary syndrome, endometriosis, or recurrent pregnancy loss all achieved a significantly lower peak endometrial thickness than control subjects. There was a trend toward increased endometrial thickness in cycles resulting in pregnancy compared with those not (10.1 vs. 9.6 mm, respectively). CONCLUSION(S) Endometrial development follows a predictable pattern, with a plateau in growth at cycle day 9. Diseases associated with infertility manifest a proliferative phase defect that can be recognized clinically.

Pretreatments before the Induction of Ovulation in Assisted Reproduction Technologies: Evidence-based Medicine in 2007

Recently, much progress has been made in the area of cryopreservation of ovarian tissue, one of the only options for fertility preservation available to women who require immediate gonadotoxic chemotherapy. Human ovarian cortical tissue strips have been cryopreserved, thawed, and autotransplanted with successful reproductive function. Cryopreservation of ovarian cortical strips, however, is limited by the ischemia that occurs at the time of retransplantation. Thus for patients that desire long-term resumption of endocrine function, cryopreservation of the whole ovary with an intact pedicle and vascular supply may be a better option. This article describes recent advances in whole ovary cryopreservation in both animal and human models, with a focus on surgical technique for removal, choice of cryoprotectants, freezing and thawing protocols, and preliminary results with organ retransplantation. Although no human cases of whole ovary retransplantation after cryopreservation have been performed to date, these preliminary studies have been encouraging, and it is likely that this option for fertility preservation will be a viable treatment option in the future.

Reproductive efficiency of women over the age of 40 and the low risk of multiple pregnancies

OBJECTIVE To investigate the oocyte-to-baby rate when controlled ovarian stimulation was performed on a highly successful donor population and to evaluate whether they produce a higher proportion of reproductively competent oocytes compared with standard donors. DESIGN Retrospective analysis of clinical and embryological database. SETTING University center. PATIENT(S) A total of 191 oocyte donation cycles were analyzed from 53 donors (28 best-prognosis donors and 23 standard donors). INTERVENTION(S) None. MAIN OUTCOME MEASURE(S) Total number of oocytes collected, the number of embryos transferable (fresh and frozen). and corresponding oocyte to live baby born (LBB) rates. In patients with remaining frozen embryos, the final LBB rate was estimated according to our reported rates. RESULT(S) A total of 130 oocyte retrievals from the best-prognosis donors yielded 2,470 oocytes. The total LBB per oocyte retrieved and LBB per embryo transferred was 7.3% and 24.6%, respectively. A total of 61 oocyte retrievals from the standard donors yielded 1,044 oocytes. The total LBB per oocyte and LBB per embryo transferred was 5.0% and 16.6%, respectively. This is significantly different from the best-prognosis donor group. Success rates were also analyzed after grouping donors based on the number of oocytes retrieved per cycle. In the best-prognosis group, the oocyte use rate increased significantly when fewer oocytes were retrieved, whereas the oocyte-to-baby rate was similar regardless of the number of oocytes for the standard donor group. CONCLUSION(S) This retrospective analysis revealed the existence of a subset of good-prognosis donors who produce a higher oocyte-to-baby rate that is indicative of a more biologically efficient reproductive system. Their identification, albeit a posteriori, has clinical implications for safety, by reducing ovarian hyperstimulation syndrome and multiple pregnancies, as well as for assisted reproductive technology success.