Exogenous Estrogen Exposure Research Articles

Estrogens and development of the rete testis, efferent ductules, epididymis and vas deferens

Estrogen has always been considered the female hormone and testosterone the male hormone. However, estrogen's presence in the testis and deleterious effects of estrogen treatment during development have been known for nearly 90 years, long before estrogen receptors (ESRs) were discovered. Eventually it was learned that testes actually synthesize high levels of estradiol (E2) and sequester high concentrations in the reproductive tract lumen, which seems contradictory to the overwhelming number of studies showing reproductive pathology following exogenous estrogen exposures. For too long, the developmental pathology of estrogen has dominated our thinking, even resulting in the “estrogen hypothesis” as related to the testicular dysgenesis syndrome. However, these early studies and the development of an Esr1 knockout mouse led to a deluge of research into estrogen's potential role in and disruption of development and function of the male reproductive system. What is new is that estrogen action in the male cannot be divorced from that of androgen. This paper presents what is known about components of the estrogen pathway, including its synthesis and target receptors, and the need to achieve a balance between androgen- and estrogen-action in male reproductive tract differentiation and adult functions. The review focuses on what is known regarding development of the male reproductive tract, from the rete testis to the vas deferens, and examines the expression of estrogen receptors and presence of aromatase in the male reproductive system, traces the evidence provided by estrogen-associated knockout and transgenic animal models and discusses the effects of fetal and postnatal exposures to estrogens. Hopefully, there will be enough here to stimulate discussions and new investigations of the androgen:estrogen balance that seems to be essential for development of the male reproductive tract.

The impact of exogenous estrogen exposure on the characteristics of estrogen receptor (ER) positive, early-stage breast cancer (EBC).

e12606 Background: Oral contraceptives (OC) and hormone replacement therapy (HRT) are well-established risk factors for ER positive breast cancer. Infertility is associated with an increased breast cancer risk and there is conflicting data on the influence of fertility treatments on breast cancer risk. The impact of exogenous estrogen exposure on breast cancer characteristics is not well described. Methods: A single center retrospective cohort study comprising all women with ER positive, human epidermal growth factor receptor 2 (HER2) negative, EBC whose tumors were sent to OncotypeDX analysis treated in our institute between 2005 and 2012. Data on exogenous estrogen exposure were collected including: OC and HRT use and prior fertility treatments. The impact of these exposures was assessed on pre-specified histopathological features including: tumor size, nodal status, ER and progesterone receptor (PR) staining, grade, Oncotype recurrence score (RS), ki67, lymphovascular and perineural invasion. Results: A total of 620 women were included, 79% were postmenopausal. Prior exposure to OC, HRT and fertility treatments was documented in 19% (103), 30% (136) and 11% (62), respectively. OC use was associated with smaller (≤1cm) tumors (30% vs. 20%, p = 0.023) and were less likely to have grade 3 disease (10% vs. 19%, p = 0.049). No other associations were found between exogenous estrogen exposures and tumor characteristics (Table). Conclusions: Use of OC may be associated with breast cancer with a distinct features compared to women with luminal breast cancer without history of OC use. Large scale studies are needed to better characterize these findings. [Table: see text]

Estrogen Receptor 1 (ESR1) Agonist Induces Ovarian Differentiation and Aberrant Müllerian Duct Development in the Chinese Soft-shelled Turtle, Pelodiscus sinensi.

Estrogens play critical roles in ovarian and reproductive organ development, but the molecular signaling pathways in non-mammalian vertebrates are not well understood. Studies of reptiles have indicated that administration of exogenous estrogens during embryonic development causes ovarian differentiation and presumptive male to female sex-reversal. The Chinese soft-shelled turtle, Pelodiscus sinensis, belongs to the family Trionychidae and exhibits genotypic sex determination system with ZZ/ZW sex chromosomes. In order to assess the role of estrogens and their signaling pathway on sex determination and differentiation, P. sinensis eggs were given a single administration of endogenous estrogen,17β-estradiol (E2) or a synthetic estrogen receptor 1 (ESR1) agonist, 4,4',4"-(4-propyl-[1H]-pyrazole-1,3,5-triyl) trisphenol (PPT) in ovo during gonadal differentiation, and the subsequent effects were examined during a final developmental stage prior to hatching. The administration of both E2 and PPT induced ovarian differentiation in genetic male embryos. Intriguingly, PPT but not E2 induced the Müllerian duct enlargement and aberrant glandular development. These data suggest that ovarian differentiation and reproductive tract anomalies induced by the exogenous estrogen exposure act through ESR1 in the Chinese soft-shelled turtles.

Kisspeptin and neurokinin B analogs use in gynecological endocrinology: where do we stand?

Recent studies have found that kisspeptin/neurokinin B/dynorphin neurons (KNDy neurons) in the infundibular nucleus play a crucial role in the reproductive axis. Analogs, both agonists and antagonists, of kisspeptin and neurokinin B (NKB) are particularly important in explaining the physiological role of KNDy in the reproductive axis in animals. The use of kisspeptin and NKB analogs has helped elucidate the regulators of the hypothalamic reproductive axis. This review describes therapeutic uses of Kiss-1 and NKB agonists, most obviously the use of kisspeptin agonists in the treatment for infertility and the induction of ovulation. Kisspeptin antagonists may have potential clinical applications in patients suffering from diseases associated with enhanced LH pulse frequency, such as polycystic ovary syndrome or menopause. The inhibition of pubertal development using Kiss antagonists may be used as a therapeutic option in precocious puberty. Kisspeptin antagonists have been found capable of inhibiting ovulation and have been proposed as novel contraceptives. Hypothalamic amenorrhea and delayed puberty are conditions in which normalization of LH secretion may potentially be achieved by treatment with both kisspeptin and NKB agonists. NKB antagonists are used to treat vasomotor symptoms in postmenopausal women, providing rapid relief of symptoms while supplanting the need for exogenous estrogen exposure. There is a wide spectrum of therapeutic uses of Kiss-1 and NKB agonists, including the management of infertility, treatment for PCOS, functional hypothalamic amenorrhea or postmenopausal vasomotor symptoms, as well as contraceptive issues. Nevertheless, further research is needed before kisspeptin and NKB analogs are fully incorporated in clinical practice.

A critical assessment of case reports describing absent uterus in subjects with oestrogen deficiency

The dual diagnosis of hypoplastic uterus in association with ovarian dysgenesis is regularly reported but the pathogenesis of the association is unclear. The uterus, however, may be invisible to all imaging modalities without at least six months of exogenous oestrogen exposure in complete ovarian failure. We assessed all available case reports in this category to estimate whether the apparent association between primary ovarian insufficiency or Turner syndrome and Mullerian agenesis can be largely accounted for by oestrogen deficiency. A literature review of all cases in which an association between ovarian insufficiency or Turner syndrome and hypoplastic uterus has been reported. PubMed was searched for all case reports associated with relevant key terms. In total, 22 publications with a total of 25 patients were identified and reviewed; 14 subjects had the normal female karyotype (46,XX), and 11 subjects had Turner Syndrome. Proportion of subjects who had been exposed to adequate oestrogen prior to the absent uterine diagnosis. A diagnosis of absent uterus was made prior to exposure to exogenous oestrogen in 22/25 (88%) of subjects with primary hypogonadism including 14/14 females with normal karyotype and 8/11 females with Turner syndrome. Oestrogen deficiency is a possible explanation for most subjects being reported as having Mullerian agenesis in association with Turner syndrome or primary ovarian insufficiency. In the presence of oestrogen deficiency, no conclusion can be made about the status of the uterus until adequate exposure to exogenous oestrogen has been completed and we suggest reassessment of the uterus when full adult dose has been reached towards the end of induction of puberty.

Exogenous estrogen therapy, testicular cancer, and the male to female transgender population: a case report

BackgroundOver the last 40 years, there has been a significant increase in the incidence of testicular cancer. The epidemiologic evidence to understand this phenomenon is unclear, however exogenous estrogen exposure is thought to be a driver in the development of testicular cancer. This is of particular importance in the transgender population because utilization of exogenous estrogen therapy is an essential aspect of the transition process.CaseWe present the case of a 38-year-old Caucasian male to female transgender patient who presented with metastatic testicular cancer 15 months after initiating estrogen therapy. She presented to our emergency department with worsening back pain and fatigue. A clinical examination revealed a right-sided testicular mass. A computed tomography scan of her abdomen/pelvis identified a right groin lesion measuring 6.4 cm, a retroperitoneal mass causing right-sided hydronephrosis, an extensive deep vein thrombosis, and pathologic abdominal lymphadenopathy. Germ cell tumor markers revealed an alpha-fetoprotein of < 2.5 μg/L and a beta-human chorionic gonadotrophin of 2526 IU/L. Her lactate dehydrogenase was 5294 U/L. Medical oncology advised the discontinuation of hormonal therapy at this time. On the basis of elevation in germ cell tumor markers and the burden of disease, she was treated with four cycles of bleomycin, etoposide, and cisplatin chemotherapy. A decision to defer upfront radical inguinal orchiectomy was made due to not wanting to have an early interruption in anticoagulation.Following the completion of the chemotherapy, a 6 cm retroperitoneal mass persisted. Due to the location of the mass and surgical morbidity associated with excision, she was followed with positron emission tomography-computed tomography by Uro-oncology, with no evidence of recurrent disease 2 years since the time of diagnosis.ConclusionsWhile there are recognized risks associated with estrogen therapy less is known about the extent to which exogenous estrogen can serve as a driver of malignancy. With recent experimental evidence revealing a pro-growth impact of estrogen on human testicular cells, continued reporting of similar cases in the literature is imperative to see if a link between exogenous estrogen exposure and testicular cancer exists.

Leptin gene variants and colorectal cancer risk: Sex-specific associations.

BackgroundHigh levels of serum leptin and low levels of serum adiponectin are strongly correlated with obesity, a well-established risk factor for colorectal cancer (CRC). Growing evidence suggests that dysregulation of leptin and adiponectin levels may play an etiological role in colorectal carcinogenesis. We evaluated 20 candidate variants in 4 genes previously shown to alter serum leptin and adiponectin levels for associations with obesity (BMI>30 kg/m2) and CRC risk.MethodsWe analyzed 6,246 CRC cases and 7,714 population-based controls from 11 studies within the Genetics and Epidemiology of Colorectal Cancer Consortium (GECCO). Associations of each variant with obesity or CRC were evaluated using multivariate logistic regression models stratified by sex and adjusted for age, a study variable, and the first three principal components of genetic ancestry. Gene-specific False Discovery Rate (FDR)-adjusted p-values <0.05 denoted statistical significance.ResultsTwo variants in the leptin gene showed statistically significant associations with CRC among women: LEP rs2167270 (OR = 1.13, 95% CI: 1.06–1.21) and LEP rs4731426 (OR = 1.09, 95% CI: 1.02–1.17). These associations remained significant after adjustment for obesity, suggesting that leptin SNPs may influence CRC risk independent of obesity. We observed statistically significant interactions of the leptin variants with hormone replacement therapy (HRT) for CRC risk; these variant associations were strengthened when analyses were restricted to post-menopausal women with low estrogen exposure, as estimated by ‘never use’ of HRT and/or non-obese BMI. No variants were associated with CRC among men.ConclusionsLeptin gene variants may exhibit sex-specific associations with CRC risk. Endogenous and exogenous estrogen exposure may modify the association between these variants, leptin levels, and CRC risk.

Behind the bleed: analysis of the formation of subchorionic hematomas (SCH) in single euploid embryo transfer cycles by protocol

Subchorionic hematomas (SCH) are a known risk factor for poor obstetric outcomes such as spontaneous abortion and preterm delivery. We sought to determine if there is a difference in the prevalence of SCH formation in pregnancies after frozen embryo transfer (FET) between natural and programmed preparation. Retrospective cohort study of single euploid FET cycles resulting in clinical pregnancy performed between 1/2016 and 12/2017 at our center. All single euploid FET cycles utilizing Next Generation Sequencing were reviewed. Exclusion criteria included FET cycles with ploidy determined by aCGH, or cycles in which untested, mosaic, or multiple embryos were transferred. All transfer cycles that resulted in clinical pregnancy (defined as presence of gestational sac) were included. These cycles were analyzed by protocol; natural versus programmed. Patients undergoing programmed cycles took oral estradiol daily followed by either 50-75mg intramuscular progesterone (P4) in oil or vaginal P4 suppository. Patients undergoing natural cycles (or supplemented with letrozole for 5 days) were monitored until a dominant follicle reached >18mm and ovulation was confirmed followed by the initiation of vaginal P4 suppository. SCH were documented at the time of luteal ultrasound as a measurable clot behind the gestational sac. Primary outcome was the rate of SCH noted on luteal ultrasound. Statistical analyses included students T-test, chi-squared and Fischer’s Exact test where appropriate (mean +/- SD, p < 0.05). A total of 719 cycles were included; 86 natural and 633 programmed (Table 1). SCH formation was diagnosed in 12.9% (n = 93) of clinical pregnancies. The formation of SCH was significantly lower in natural cycles compared to programmed cycles (RR 0.4, 95% CI 0.17 - 0.98, p < 0.04). Relevant cycle parameters were similar between groups (Table 1); cycle day 28 estrogen was higher in programmed cycles reflecting exogenous estrogen exposure. When programmed cycles utilizing vaginal P4 only were excluded (n=36), the formation of SCH is still statistically lower in natural compared to programmed cycles (5.8% v 14.2%, RR 0.4 (0.17-0.97, p <0.03). SCH occurred less frequently in single euploid embryo transfers using a “natural” endometrial preparation. Further analyses of long term outcomes could help provide insight into estimates of obstetrical risk.Tabled 1SCH Formation in Clinical Pregnancies by FET Protocol: Patient and Cycle CharacteristicsNatural Cycles (n=86)Programmed Cycles (n=633)p - valueDeveloped SCH5.8% (5/86)14.0% (88/633)p < 0.04 RR 0.4 (0.17-0.98)Average CD SCH diagnosed44.8±3.547.7±7.3p < 0.40Age37.45±4.336.81±4.4p < 0.20CD 28 hCG202.3±147.4189.3±176.5p < 0.52CD 28 E2284.9±234.5477.4±395.2p < 0.0001CD 28 P426.4±11.326.7±17.9p < 0.89Transfer Difficulty Marked “Easy” by Provider91.9% (79/86)95.6% (605/633)p < 0.13 RR 1.84 (0.83-4.1)Endometrial Thickness (mm)8.8±1.510.4±2.4p < 0.54 Open table in a new tab

A Longitudinal Study of Estrogen-Responsive Tissues and Hormone Concentrations in Infants Fed Soy Formula.

Chemicals with hormonelike activity, such as estrogenic isoflavones, may perturb human development. Infants exclusively fed soy-based formula are highly exposed to isoflavones, but their physiologic responses remain uncharacterized. Estrogen-responsive postnatal development was compared in infants exclusively fed soy formula, cow-milk formula, and breast milk. We enrolled 410 infants born in Philadelphia-area hospitals between 2010 and 2014; 283 were exclusively fed soy formula (n = 102), cow-milk formula (n = 111), or breast milk (n = 70) throughout the study (birth to 28 or 36 weeks for boys and girls, respectively). We repeatedly measured maturation index (MI) in vaginal and urethral epithelial cells using standard cytological methods, uterine volume and breast-bud diameter using ultrasound, and serum estradiol and follicle-stimulating hormone levels. We estimated MI, organ-growth, and hormone trajectories by diet using mixed-effects regression splines. Maternal demographics did not differ between cow-milk-fed and soy-fed infants but did differ between formula-fed and breastfed infants. Vaginal-cell MI trended higher (P = 0.01) and uterine volume decreased more slowly (P = 0.01) in soy-fed girls compared with cow-milk-fed girls; however, their trajectories of breast-bud diameter and hormone concentrations did not differ. We observed no significant differences between boys fed cow-milk vs soy formula; estradiol was not detectable. Breastfed infants differed from soy-formula-fed infants in vaginal-cell MI, uterine volume, and girls' estradiol and boys' breast-bud diameter trajectories. Relative to girls fed cow-milk formula, those fed soy formula demonstrated tissue- and organ-level developmental trajectories consistent with response to exogenous estrogen exposure. Studies are needed to further evaluate the effects of soy on child development.

Hormone factors play a favorable role in female head and neck cancer risk.

Due to lower female incidence, estimates of exogenous and endogenous hormonal factors in head and neck cancers (HNCs, comprising cancers of the oral cavity, oropharynx, hypopharynx, and larynx) among women have been inconsistent and unable to account for key HNC risk factors. We pooled data from 11 studies from Europe, North America, and Japan. Analysis included 1572 HNC female cases and 4343 controls. Pooled odds ratios (ORs) estimates and their 95% confidence intervals (CIs) were calculated using multivariate logistic regression models adjusting for tobacco smoking and alcohol drinking. Lower risk was observed in women who used hormone replacement therapy (HRT) (OR = 0.58; 95% CI: 0.34–0.77). Pregnancy (OR = 0.61; 95% CI: 0.42–0.90) and giving birth (OR = 0.59; 95% CI: 0.38–0.90) at <35 years of age were inversely associated with HNCs. An inverse association with HNC was observed with age at start of HRT use (OR = 0.59; 95% CI: 0.39–0.90) for each additional 10 years and with duration of use (OR = 0.87; 95% CI: 0.76–0.99 for every 3 years). Exogenous female hormone use is associated with a nearly twofold risk reduction in female HNCs. The lower female HNC incidence may, in part, be explained by endogenous and exogenous estrogen exposures.

Hormone Replacement Therapy and Oral Contraceptives and Risk of Oesophageal Adenocarcinoma: A Systematic Review and Meta-analysis.

There is an unexplained strong male predominance in the aetiology of oesophageal adenocarcinoma (OAC). The hypothesis that oestrogens are protective deserves attention. A potential protective influence of exogenous oestrogen exposure that is hormone replacement therapy (HRT) and oral contraceptives (OC) has been addressed only in studies of limited statistical power and the individual studies have not provided conclusive results. We conducted a systematic literature search and meta-analysis on HRT and OC and the risk of OAC. We used the databases PubMed and the Web of Science. Odds ratios (ORs) and 95% confidence intervals (CIs) were estimated by the Mantel-Haenszel random-effect method. A total of five studies were included. Compared to never users ever users of HRT had a statistically significantly decreased risk of OAC (pooled OR = 0.75; 95% CI: 0.58-0.98) and ever users of OC had a borderline significantly decreased risk of this cancer (pooled OR = 0.76; 95% CI: 0.57-1.00). In conclusion HRT and OC use seems to be associated with a decreased risk of OAC. However further research is warranted. (c) 2014 UICC.

Pre- and postnatal bisphenol A treatment does not alter the number of tyrosine hydroxylase-positive cells in the anteroventral periventricular nucleus (AVPV) of weanling male and female rats

Exposure to Bisphenol A (BPA) may interfere with brain sexual differentiation. Altered numbers of tyrosine hydroxylase (TH) cells in the rodent anteroventral periventricular nucleus (AVPV) after developmental BPA treatment have been reported; however, definitive conclusions are lacking. The current study incorporated many of the guidelines suggested for endocrine disrupter research. Specifically, ethinyl estradiol (EE2) served as a reference estrogen, exogenous environmental estrogen exposure was controlled, BPA was administered orally, and subjects consumed a low phytoestrogen diet. Here, on gestational days 6–21, Sprague-Dawley rats (10–15/group) were gavaged with 2.5 or 25.0µg BPA/kg/day or 5.0 or 10.0µg EE2/kg/day or the vehicle (5ml of 0.3% aqueous carboxymethylcellulose/kg/day). A naïve control group was weighed and restrained, but not gavaged. Beginning on postnatal day (PND) 1 and continuing until PND 21, the 4 pups/sex/litter were orally treated with the same dose their dam had received. On PND 21, 1/sex/litter was perfused and the brain removed. TH immunoreactivity (TH-ir) was counted in 8 images/pup by a technician blind to treatment status. ANOVA results indicated significantly higher TH-ir cells/mm2 in females (main effect of sex: p<0.01); however, there was no significant effect of treatment or a significant interaction of treatment with sex. In a separate untreated group of PND 21 Sprague-Dawley pups, AVPV volume was quantified and no significant sexual dimorphism was apparent. Similar to our reported results of behavioral assessments, the BPA treatment paradigm used here (2.5 or 25.0µg BPA/kg/day administered orally) does not appear to cause significant alterations in AVPV TH-ir.

The Critical Hormone-Sensitive Window for the Development of Delayed Effects Extends to 10 Days after Birth in Female Rats Postnatally Exposed to 17alpha-Ethynylestradiol.

Neonatal exposure to estrogens is known to cause delayed effects, a late-occurring adverse effect on adult female reproductive functions, such as early onset of age-matched abnormal estrous cycling. However, the critical period in which neonates are sensitive to delayed effects inducible by exogenous estrogen exposure has not been clearly identified. To clarify this window, we examined the intensity and timing of delayed effects using rats exposed to ethynylestradiol (EE) at various postnatal ages. After subcutaneous administration of a single dose of EE (20 μg/kg, which induces delayed effects) on Postnatal Day (PND) 0, 5, 10, or 14 in Wistar rats, hypothalamic and hormonal alterations in young adults and long-term estrous cycling status were investigated as indicators of delayed effects. In young adults, peak luteinizing hormone concentrations at the time of the luteinizing hormone surge showed a decreasing trend, and KiSS1 mRNA expression of the anterior hypothalamus and number of KiSS1-positive cells in the anteroventral periventricular nucleus were significantly decreased in the PND 0, 5, and 10 groups. The reduction in KiSS1 mRNA and KiSS1-postive cells was inversely correlated with age at time of exposure. These groups also exhibited early onset of abnormal estrous cycling, starting from 17 wk of age in the PND0 group and 19 wk of age in the PND5 and 10 groups. These indicators were not apparent in the PND14 group. Our results suggest that PND0-PND10 is the critical window of susceptibility for delayed effects, and PND14 is presumed to be the provisional endpoint of the window.

Reproductive and hormonal factors and mortality among women with colorectal cancer in the NIH-AARP Diet and Health Study.

Background:Although use of menopausal hormone therapy (MHT) and some reproductive factors have been associated with colorectal cancer (CRC) risk, relations between these factors and survival after CRC diagnosis are unclear.Methods:Among 2053 post-menopausal women diagnosed with incident CRC in the NIH-AARP Diet and Health Study, we calculated hazard ratios (HRs) and 95% confidence intervals (CIs) using multivariable Cox proportional hazards regression to test associations between oral contraceptive (OC) use, menarche age, age at first birth, parity, menopausal age, and MHT use with all-cause and CRC-specific mortality.Results:There were 759 deaths (332 CRC-related deaths) over a median follow-up of 7.7 years. We observed no statistically significant associations between OC use, menarche age, age at first birth, parity, menopausal age, and mortality. Compared with never MHT use, former use was not associated with mortality, but we found an inverse association among baseline current users, for both all-cause (HR=0.79, 95% CI 0.66–0.94) and CRC mortality (0.76, 0.59–0.99).Conclusion:Future studies should further focus on the mechanisms by which exogenous oestrogen exposure might affect tumour progression and CRC survival.

Abstract 4213: Evaluation of LMTK3 expression and tumor phenotype in estrogen-dependent colorectal cancer

Abstract Background: Incompatible physiology between genders influences development and progression of colorectal cancer (CRC). It is postulated that improved survival of women with CRC may be due to exogenous estrogen exposure. A recent study used the age of 50 as a surrogate for menopause, evaluating survival in two age groups: women ≥50 and ≤50. It was observed that women ≤50 had a survival advantage that was contributed to protection by estrogen- a mechanism lost in menopause; with no significant difference found in men. Lemur tyrosine kinase 3 (LMTK3) was recently identified, that when in association with an estrogen receptor (ER) α modulator increased growth, progression, and altered resistance to endocrine therapy was observed in breast cancer. The ER family (ERα and ERβ) exerts contrasting actions of estrogen on different target tissues, where ERβ is the dominant receptor expressed in the gastrointestinal tract. The different ER distribution between breast and colon cancer led to the hypothesis that LMTK3 overexpression should stabilize ERβ and decrease tumorigenesis in CRC. Methods: CRC cell lines were characterized for LMTK3 mRNA (qRT-PCR) and protein expression (Western blot) in both transfected and non-transfected cell lines. Overexpression of LMTK3 was achieved utilizing pCMV6-XL4-LMTK3. Overexpressing LMTK3 CRC cells were evaluated for alterations to cell viability by MTT, colony formation (CFAs), invasion assays and chemosensitivity to 5-Fluororuacil, oxaliplatin, and tamoxifen. Results: CRC cell lines demonstrated a range of LMTK3, ERα and ERβ mRNA and protein expression. Caco-2, HCT116 and LoVo cells demonstrated estrogen sensitivity as evidenced by treatment with ER antagonist, tamoxifen. CFA demonstrated that the presence of estrogen plus overexpression of LMTK3 decreased cell viability by 27, 28 and 36% in Caco2, HCT116 and LoVo cells respectively, had limited impact on chemosensitivity, and decreased their ability to invade. Conclusion: This study demonstrates that estrogen mediated cell signaling and LMTK3 play an important role in decreasing CRC tumorigenesis. Further studies are underway to better understand estrogen and LMTK3's role in CRC. Citation Format: Sybil Magturo, Georgios Giamas, Snay Mallick, Austin Layton, Justin Stebbing, Heinz-Josef Lenz, Melissa J. LaBonte. Evaluation of LMTK3 expression and tumor phenotype in estrogen-dependent colorectal cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4213. doi:10.1158/1538-7445.AM2014-4213

Hormone replacement therapy and oral contraceptives and risk of oesophageal adenocarcinoma: A systematic review and meta‐analysis

There is an unexplained strong male predominance in the aetiology of oesophageal adenocarcinoma (OAC). The hypothesis that oestrogens are protective, deserves attention. A potential protective influence of exogenous oestrogen exposure, that is, hormone replacement therapy (HRT) and oral contraceptives (OC) has been addressed only in studies of limited statistical power, and the individual studies have not provided conclusive results. We conducted a systematic literature search and meta-analysis on HRT and OC and the risk of OAC. We used the databases PubMed and the Web of Science. Odds ratios (ORs) and 95% confidence intervals (CIs) were estimated by the Mantel-Haenszel random-effect method. A total of five studies were included. Compared to never users, ever users of HRT had a statistically significantly decreased risk of OAC (pooled OR = 0.75; 95% CI: 0.58-0.98), and ever users of OC had a borderline significantly decreased risk of this cancer (pooled OR = 0.76; 95% CI: 0.57-1.00). In conclusion, HRT and OC use seems to be associated with a decreased risk of OAC. However, further research is warranted.

Abstract PL01-04: Stress and the social environment: Impact on cancer risk

Abstract The breast cancer research community has long considered the impact of exogenous estrogen exposure, nutrition, alcohol consumption, and exercise on breast cancer biology. More recently, experimental studies of exposure to chronic stressors in the social environment and the resulting stress response have suggested that epinephrine- and glucocorticoid-mediated signaling pathways might potentiate breast cancer growth both at primary and metastatic tumor sites. To study this hypothesis in more detail, we initiated a transdisciplinary study of a well-established social stressor for female rodents- chronic social isolation. We found that exposing a young female rodent to chronic socially isolation for 12 weeks (compared to group living/social interaction with three other female mice) was associated with increased mammary tumor growth in genetically predisposed rodents. For example, in the C3(1)/SV40 T-antigen (Tag) FVB/N mouse model of “triple-negative” (ER,PR and ErbB-negative breast cancer) the heightened stress response elicited by chronic social isolation was associated with increased expression of metabolic genes in the mammary gland before invasive tumors develop (i.e. during the in situ carcinoma stage when the animals are 15 weeks of age). This suggested that metabolic changes within the mammary gland could affect breast cancer biology. To further understand how accelerated mammary tumor growth may be associated with the physiological response to social isolation, we separated the mammary gland adipose tissue from its adjacent ductal epithelial cells and then analyzed individual cell types for changes in metabolic gene expression. We found that the major increase in expression of the key metabolic genes Acaca, Hk2 and Acly was primarily in the adipocyte (fat cell) fraction of the mammary gland. In contrast, metabolic gene expression was not significantly increased in visceral or gonadal adipose depots of socially isolated female mice, suggesting that the stress response affects individual fat depots differently in this model. We confirmed that the increased metabolic gene expression in the mammary adipocytes of socially isolated mice coincided with increased glucose metabolism, lipid synthesis, and local leptin production; interestingly, systemic leptin levels were not significantly altered. Furthermore, compared to the group housed mammary fat, application of media that had been cultured with isolated mouse mammary adipose tissue resulted in relatively increased proliferation of mammary cancer cells. These results suggest that the physiological response to a chronic social stressor (isolation) results in metabolic changes in mammary gland adipocytes that in turn contribute to increased proliferation of pre-invasive malignant epithelial cells. Adipocyte-secreted metabolites and/or tumor growth-promoting proteins could identify biomarkers and/or targets for preventive intervention in breast cancer. We are currently examining the mammary fat of the isolated versus group-housed mice to identify differential production of adipokines and lipid species that may influence early breast cancer growth in this model. Citation Format: Suzanne Conzen. Stress and the social environment: Impact on cancer risk. [abstract]. In: Proceedings of the Eleventh Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2012 Oct 16-19; Anaheim, CA. Philadelphia (PA): AACR; Cancer Prev Res 2012;5(11 Suppl):Abstract nr PL01-04.

Reference gene selection for qPCR in mussel, Mytilus edulis, during gametogenesis and exogenous estrogen exposure

The aim of this study is to develop a normalization method for real-time PCR data by analyzing the most stably expressed control genes in mussel (Mytilus edulis) reproductive tissue. To facilitate this, six candidate genes, including several commonly used in the literature, were investigated in mussels at different stages of gametogenesis and following experimental exposure to a model estrogen (17b-estradiol). GeNorm and NormFinder softwares were employed to assess the stability of the reference genes. Our results demonstrate that the most stable reference genes are not the same in mussels at different stages of gametogenesis and in experimentally E2-exposed mussels. Interestingly, HEL (helicase) and ACT (actin) mRNA expression levels were most affected by the stage of gametogenesis and yet, in molluscan studies, ACT is possibly the most frequently used reference gene. We demonstrate that the experimental results are highly dependent on the reference gene chosen and that statistically significant contrasting differences between sample groups are present or absent depending on the reference gene employed.

Exposure of neonatal rats to anti‐androgens induces penile mal‐developments and infertility comparable to those induced by oestrogens

We previously reported that oestrogen exposure in neonatal rats induced permanent infertility and malformed penis characterized by fat accumulation, which replaced most of the smooth muscle cells and cavernous spaces in the body of the penis, structures essential for erection. The objective of this study was to determine if reduced androgen production/action in the neonatal period, in the absence of exogenous oestrogen exposure, induces penile deformities similar to those caused by oestrogen. Male rats were treated from postnatal days 1-6 with GnRH antagonist antide (A, 10 mg/kg) or androgen receptor (AR) antagonist flutamide (F, 50 mg/kg) or F + A, with or without AR agonist dihydrotestosterone (DHT, 20 mg/kg). For comparison, pups received diethylstilbestrol (DES, 0.1 mg/kg), with or without DHT. Tissues were collected at ages 7 and 12 days and at adulthood. Flutamide alone decreased penile length and weight significantly (p < 0.05), but it caused neither fat accumulation, nor affected fertility (80% vs. 87% in controls). Antide alone reduced penile length and weight significantly, and induced fat accumulation in 4/11 rats and infertility in 13/14 rats. Conversely, all 11 F + A-treated rats, similar to all nine DES-treated rats, had fat accumulation and loss of smooth muscle cells and cavernous spaces in the body of the penis and were infertile. In addition, reductions in penile length and weight were higher than in rats treated with F or A alone. DHT co-administration mitigated penile deformities in the DES group, but did not in the F + A group. Testicular testosterone was reduced by 70-95% at 7 or 12 days of age in all treated groups, except in the F group, which had threefold higher testosterone than controls. Collectively, data unequivocally show that reduced androgen production/action in the neonatal period, in the absence of oestrogen exposure, induces permanent infertility and malformed penis similar to that caused by oestrogen.

Identification of Reproduction-Specific Genes Associated with Maturation and Estrogen Exposure in a Marine Bivalve Mytilus edulis

BackgroundWhile it is established that vertebrate-like steroids, particularly estrogens (estradiol, estrone) and androgens (testosterone), are present in various tissues of molluscs, it is still unclear what role these play in reproductive endocrinology in such organisms. This is despite the significant commercial shellfishery interest in several bivalve species and their decline.Methodology/Principal FindingsUsing suppression subtraction hybridisation of mussel gonad samples at two stages (early and mature) of gametogenesis and (in parallel) following controlled laboratory estrogen exposure, we isolate several differentially regulated genes including testis-specific kinases, vitelline lysin and envelope sequences.ConclusionsThe differentially expressed mRNAs isolated provide evidence that mussels may be impacted by exogenous estrogen exposure.