Warren G. Foster

Detection of phytoestrogens in samples of second trimester human amniotic fluid

There is widespread concern that fetal exposure to hormonally active chemicals may adversely affect development of the reproductive tract. Therefore, the present study was performed to develop the necessary analytical methods and test the hypothesis that dietary phytoestrogens can be quantified in second trimester human amniotic fluid. Amniotic fluid samples (n=59) from women (n=53) undergoing routine amniocentesis between 15 and 23 weeks of gestation were analyzed by gas chromatography/mass spectrometric (GC/MS). Analytes included the phytoestrogens daidzein, genistein, formononetin, biochanin A, and coumestrol. Dietary phytoestrogens were quantified in 96.2% of second trimester amniotic fluid samples tested. The mean (+/- standard deviation (S.D.)) concentration of daidzein and genistein in amniotic fluid was 1.44 +/- 1.34 and 1.69 +/- 1.48 ng/ml with maximum levels of 5.52 and 6.54 ng/ml, respectively. Second trimester amniotic fluid contains quantifiable levels of dietary phytoestrogens and thus is a marker of mid pregnancy fetal exposure.

Appendiceal Disease in Women with Endometriosis and Right Lower Quadrant Pain

STUDY OBJECTIVEnTo evaluate the frequency and range of appendiceal disease in women with endometriosis and right lower quadrant (RLQ) pain, and to estimate the value of preoperative gastrograffin enema (GGE) as a screen for the disease.nnnDESIGNnNonrandomized clinical trial (Canadian Task Force classification II-2).nnnSETTINGnUniversity-affiliated hospital with a private practice setting.nnnPATIENTSnA subpopulation of 65 women from a group of 337 patients undergoing laparoscopy for symptomatic endometriosis.nnnINTERVENTIONnPreoperative GGE was performed whenever possible in these women. At laparoscopy, the appendix was removed if it appeared abnormal or if the preoperative GGE was positive.nnnMEASUREMENTS AND MAIN RESULTSnOf 65 women (19%) with symptomatic endometriosis and preoperative RLQ pain, 52 (80%) underwent appendectomy as part of surgery. Of these 52 excised appendixes, 39 (75%) had histologically confirmed pathology including appendicitis or periappendicitis, endometriosis, fibrous obliteration, lymphoid hyperplasia, and carcinoid tumor. Preoperative GGE had sensitivity of 74% and specificity of 83% for appendiceal disease. Its positive predictive value was 95% and negative predictive value was 42%. No complications from laparoscopic appendectomy occurred.nnnCONCLUSIONnDisease of the appendix is common in women with endometriosis and RLQ pain. Appendectomy is particularly likely if preoperative GGE is positive.

Environmental Contaminants and Dietary Factors in Endometriosis

Abstract: Endometriosis is an estrogen‐dependent disease characterized by the presence of endometrial glands and stroma outside the uterine cavity. The etiology of this disease remains elusive, but is clearly influenced by genetic, immune, and endocrine factors. Exposure to environmental contaminants has recently been added to the list of potential factors that contribute to the pathogenesis of endometriosis. The objective of this paper is to review the weight of the evidence from hospital‐based case‐control studies and animal experiments for an association between exposure to environmental contaminants and endometriosis.

Human developmental exposure to endocrine active compounds

Quantification of exposure to environmental contaminants such as endocrine active chemicals (EACs) during critical periods of development, particularly in utero, remains largely unexplored. Therefore, we tested the hypothesis that EACs can be detected and quantified in second trimester human amniotic fluid. Amniotic fluid was obtained from women (n=175) undergoing routine amniocentesis between 14 and 21 weeks gestation. Samples were assayed by gas chromatography/mass spectrometry (GC/MS) for common organochlorine contaminants and dietary phytoestrogens. The DDT metabolite p,p-DDE was found in approximately 25% of amniotic fluid samples (mean±S.D., 0.15±0.06 ng/ml) whereas the dietary phytoestrogens, genistein and or daidzein were found in 96.2% of samples tested (0.94±0.91 and 1.08±0.91 ng/ml, respectively). Our results demonstrate that: (1) human amniotic fluid is a suitable biological medium to evaluate developmental exposure to EACs, and (2) fetuses are exposed to biologically active levels of EACs in mid pregnancy.

Extrapolation of Rodent Studies on Amniotic Fluid Contaminatnts to Human Populations

If endocrine active chemicals (EACs) adversely affect human development, then there must be evidence of effects in animal models at properly scaled levels of exposure during pertinent sensitive periods as derived from quantified exposures of the human fetus. Our recent studies attempt to address both effects and exposures. First Study: Dams were gavaged from Gestation Day (GD) 14 through weaning on Post-Natal Day (PND) 21 with either corn oil alone (unexposed controls) or Low DES (0.5u2009mg/kg BW); High DES (5.0u2009mg/kg BW); GEN (15u2009mg/kg BW); GEN + DES (GEN at 15u2009mg/kg BW and DES at 0.5u2009mg/kg BW). No treatments affected duration of gestation, litter size or birth anogenital distance / birth body weights ((bAGD/bBW) or ratios of bAGD/cube root of bBW of pups of either sex. The ratio of weaning AGD to weaning body weight (wAGD/wBW) differed significantly between the control group and each of the estrogenic treatments in both sexes with larger wAGD/wBW values associated with each of the estrogenic treatments. Males exposed to High DES and GEN alone exhibited earlier onset of puberty. Only females in the low DES group showed an earlier onset of puberty. At 50 to 70 days of age, the ratios of male reproductive organ weights/body weight were unaffected by estrogen treatment in all groups except high DES which increased testicle weight and decreased epididymis, seminal vesicle, and prostate weights. Initial vaginal cycle lengths were affected only in the high DES group. Thus low doses of DES and GEN at levels comparable to the upper range of human exposure affect some but not all markers of sexual development. Second Study: Amniotic fluid samples obtained at routine amniocentesis between 15 and 23 weeks of gestation were assayed by gas chromatographic/mass spectrometric (GC/MS) analysis. The first group of amniotic fluid samples (n = 53) from 51 women were analysed for several xenobiotic EACs. Alpha-hexachlorocyclohexane, with a mean (± SD) concentration of 0.15 ± 0.06 (ng/ml), and p,p′-DDE, with a mean (± SD) concentration of 0.21 ± 0.18u2009ng/ ml, were detected in several specimens. Overall one in three amniotic fluid samples tested positive for at least one xenobiotic EAC. Another group of amniotic fluid samples (n = 62) from 56 women were analysed for phytoestrogenic EACs. The mean (± SD) concentration of daidzein and genistein in amniotic fluid were 1.14 ± 1.04 and 1.37 ± 1.00u2009ng/ml with maximum levels of 5.52 and 4.86u2009ng/ml, respectively. Overall, 26 and 34 of the samples had quantifiable levels of daidzein and genistein, respectively. Conclusions: One in three human fetuses were exposed to xenobiotic EACs and two of three human fetuses were exposed to phytoestrogenic EACs in utero. Our demonstrations that EACs have developmental effects in an animal model at levels of exposure that mimic those found in humans in North America during sensitive time-frames sustains concerns about potential adverse health effects of developmental exposures to EACs for the human fetus/neonate.

Hypoxia-induced autophagy, epithelial to mesenchymal transition and invasion in the pathophysiology of endometriosis: A perspective.

Endometriosis is a common estrogen-dependent disease characterized by growth of endometrial glands and stroma outside the uterine cavity. Associated with pelvic pain and infertility, endometriosis is estimated to affect up to 10% of women or 176 million women worldwide. Despite its common occurrence and debilitating impact on the lives of affected women, the cause of endometriosis remains ill defined. While retrograde menstruation is the most widely accepted theory of causation, regurgitation of menstrual effluent through the fallopian tubes occurs in approximately 90% of women. Since only a fraction of women develop endometriosis, factors other than access of menstrual effluent to the pelvic cavity alone must contribute to development of endometriosis. Therefore, unique features of menstruated cells destined to become endometriotic lesions; defects in immune surveillance or some combination of the above are thought to contribute to formation of endometriotic lesions. However, critical steps in the pathogenesis of endometriosis have yet to be elucidated. Emerging evidence over the last several years has revealed signaling pathways central to the pathophysiology of endometriosis. Dysregulation of aromatase expression leading to de novo production of estrogens [1] together with downregulation of 17β-hydroxysteroid dehydrogenase type-II result in high local tissue estrogen concentrations of endometriotic implants. Furthermore, recent studies revealed that endometriotic implants over express estrogen receptor β (EsR2) [1,2] leading to transcriptional repression of estrogen receptor α (EsR1) and progesterone receptor expression. Overexpression of EsR2 modulates tumor necrosis factor-α (TNF-α) mediated apoptosis [2] and enhanced recruitment of inflammatory mediators. In animal models, endometriotic implants formed in wild-type mice form larger implants than implants generated with loss of EsR2 function [2]. Taken together, these studies demonstrate that endometrial implant survival, growth, evasion from apoptosis, and immune dysregulation are estrogen-dependent processes driven by dysregulation of local tissue estrogen levels and preferential upregulation of EsR2. However, although the central role of estrogen signaling in established endometrial lesions is clear, it remains uncertain if these are adaptive responses limited to established lesions or key events leading to the development of disease. Several lines of evidence suggest that lesion initiation and progression are pathophysiologically distinct phases. Results of a recent study [3], using estrogen receptor and interleukin 6 receptor knockout mice and tissue recombination experiments, showed that, unlike established endometriotic implants, newly forming implants are estrogen independent. Rather the initial steps in endometriotic implant development are dependent on immune system modulation. Further evidence supporting the proposal of functionally different phases in endometriotic lesion lifespan is derived from changes in markers of endometriosis. Specifically, higher serum microRNA-451 (miR-451) has recently been demonstrated in women with endometriosis compared to a reference population [4], an observation replicated in a baboon model of endometriosis [5]. Dysregulation of miR-451 expression was detectable in eutopic endometrium as early as 3 months postinduction of endometriosis in the baboon model [5]. However, also in the baboon model, differences in circulating miR-451 peaked and only achieved significance at 6 months after endometriotic lesion induction [6]. Taken together, these data suggest that established disease is functionally distinct from implant development. Endometriosis is a benign disease that, unlike cancer, lacks the capacity for metastasis but, similar to cancer, shed endometrial cells survive under hypoxic conditions, attach to distant anatomical sites, undergo epithelial to mesenchymal transition (EMT), and develops an invasive phenotype. A recent study [7] has shown that under hypoxic conditions, endometrial stromal cells upregulate hypoxiainducible factor-1α (HIF-1α) expression, activate the self-digestion pathway (autophagy), and promotes cell migration and invasion; all key steps in the development of endometriosis. Expanding upon their prior findings, Liu and colleagues [8] have shown increased expression of HIF-1α and microtubule light chain protein (LC3), a marker of autophagy, in ectopic endometrial lesions compared to eutopic endometrium of women with and without endometriosis.

MicroRNA expression pattern differs depending on endometriosis lesion type

Abstract MicroRNA (miRNA), noncoding segments of RNA involved in post-transcriptional regulation of protein expression are differentially expressed in eutopic endometrium of women with and without endometriosis compared to endometriotic lesions. However, endometriotic lesion types are known to be biochemically distinct and therefore hypothesized that miRNAs are differentially expressed in endometriomas compared to peritoneal and deep-infiltrating lesions. Therefore, endometrial biopsies and ectopic implants from women (n = 38) undergoing laparoscopic surgery for chronic pelvic pain were collected. Samples of endometriomas, peritoneal or deep-infiltrating lesions were selected from our tissue bank for study participants who exclusively had only one lesion type noted on their surgical report. Quantitative real-time polymerase chain reaction for miR-9, miR-21, miR-424, miR-10a, miR-10b, and miR-204 was performed. miR-204 expression was significantly lower (P = 0.0016) in the eutopic endometrium of women with endometriosis compared to controls. Relative expression of miR-21, miR-424, and miR-10b differed significantly (P < 0.05) across endometriotic lesion types. Finally, all miRNAs isolated from endometriomas, peritoneal and deep-infiltrating lesions studied were differentially expressed compared to matched eutopic endometrium samples. We therefore conclude thatmiRNA expression in the eutopic endometrium from women with endometriosis differs from symptomatic controls. Moreover, miRNA expression pattern is dependent on the endometriotic lesion type studied. We suggest that identification of different miRNA expression patterns for endometriomas, peritoneal and deep-infiltrating lesions could contribute to individualized patient care for women with endometriosis. Summary Sentence MicroRNAs are differentially expressed in endometriomas, peritoneal and deep infiltration endometriotic lesions, suggesting that endometriotic lesions from different anatomical sites are biochemically distinct.

What motivates women to take part in clinical and basic science endometriosis research

BACKGROUNDnThe objective of this study was to identify factors motivating women to take part in endometriosis research and to determine if these factors differ for women participating in clinical versus basic science studies.nnnMETHODSnA consecutive series of 24 women volunteering for participation in endometriosis-related research were asked to indicate, in their own words, why they chose to volunteer. In addition, the women were asked to rate, on a scale of 0 to 10, sixteen potentially motivating factors. The information was gathered in the form of an anonymous self-administered questionnaire.nnnRESULTSnStrong motivating factors (mean score > 8) included potential benefit to other womens health, improvement to ones own condition, and participation in scientific advancement. Weak motivating factors (mean score < 3) included financial compensation, making ones doctor happy, and use of natural products. No difference was detected between clinical and basic science study participants.nnnCONCLUSIONnThis study is the first study to specifically investigate the factors that motivate women to take part in endometriosis research. Understanding why women choose to take part in such research is important to the integrity of the informed consent process. The factors most strongly motivating women to participate in endometriosis research related to improving personal or public health; the weakest, to financial compensation and pleasing the doctor.