Animal models of prenatal exposure to diethylstilboestrol

The multigeneration effect of DES provides a model to test the mechanism of transmission of cancer risk from one generation to the next

1989 Study Abstract

Animals of several species exposed perinatally to diethylstilboestrol (DES) have been evaluated for anomalies and tumours.

In male offspring, anomalies of the testis and epididymis have been reported, but evidence for tumours has been very limited.

Many anomalies and tumours have been recorded in female offspring, and some of these duplicate the anomalies and tumours reported in DES-exposed women, whereas others either have not yet been discovered or else do not occur in the human species..

Animal models of prenatal exposure to diethylstilboestrol, US National Library of Medicine National Institutes of Health, IARC scientific publications, NCBI PubMed PMID: 2680952, 1989.

A variety of abnormal physiological responses has been identified in animals exposed perinatally to DES.

There were altered levels of hormones and receptors; responses to postnatal injection of hormones were often modified; and an increased susceptibility to other carcinogens has been established.

Several mechanisms have been postulated to explain tumour production later in life after perinatal exposure to DES:

  1. Deficiencies in immune function indicate a mechanism of impaired immune surveillance.
  2. The presence of DES and its metabolites in the fetus and neonate raise the issue of somatic mutation. Evidence for sister chromatid exchange, cell transformation in tissue culture and other toxic effects on chromosomes support the somatic mutation hypothesis.
  3. A third hypothesis is involvement of abnormal differentiation of the hypothalamus. Structural, hormonal and behavioural changes support this idea.

Possible additional problems in humans after exposure to DES, on the basis of animal model studies, are increased tumour frequency with ageing and transmission of cancer risk to the third generation.

The multigeneration effect of DES provides a model to test the mechanism of transmission of cancer risk from one generation to the next.

The outcome of such experiments could have considerable impact on the understanding of the association between DES and cancer specifically and transplacental cancer generally.

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Environmental factors, epigenetics, and developmental origin of reproductive disorders

US National Library of Medicine National Institutes of Health, Reproductive toxicology, 2016

Highlights

  • Epidemiological and model system studies support an early origin of reproductive dysfunction.
  • Estrogenic/anti-androgenic chemicals as endocrine disrupting chemicals (EDCs) have vast developmental influences on adult reproductive outcomes.
  • Gestational, perinatal, neonatal, and pubertal periods are “windows of susceptibility” for epigenetic programming.
  • EDCs induce exposure-specific epigenetic modifications in regulatory genes in organs of the reproductive system.
  • Germline epigenetic disruption is a mechanism underlying transgenerational inheritance of reproductive disorders.

2017 Study Abstract

Environmental factors, epigenetics, and developmental origin of reproductive disorders, US National Library of Medicine National Institutes of Health, Reproductive toxicology (Elmsford, N.Y.), NCBI PubMed PMID: 27421580, 2016 Jul.

Image credit Daniel Friedman.

Sex-specific differentiation, development, and function of the reproductive system are largely dependent on steroid hormones.

For this reason, developmental exposure to estrogenic and anti-androgenic endocrine disrupting chemicals (EDCs) is associated with reproductive dysfunction in adulthood.

Human data in support of “Developmental Origins of Health and Disease” (DOHaD) comes from multigenerational studies on offspring of diethylstilbestrol-exposed mothers/grandmothers.

Animal data indicate that ovarian reserve, female cycling, adult uterine abnormalities, sperm quality, prostate disease, and mating behavior are susceptible to DOHaD effects induced by EDCs such as bisphenol A, genistein, diethylstilbestrol, p,p’-dichlorodiphenyl-dichloroethylene, phthalates, and polyaromatic hydrocarbons.

Mechanisms underlying these EDC effects include direct mimicry of sex steroids or morphogens and interference with epigenomic sculpting during cell and tissue differentiation.

Exposure to EDCs is associated with abnormal DNA methylation and other epigenetic modifications, as well as altered expression of genes important for development and function of reproductive tissues.

Here we review the literature exploring the connections between developmental exposure to EDCs and adult reproductive dysfunction, and the mechanisms underlying these effects.

DES DiEthylStilbestrol Resources

Cancer Risk in Women Exposed to Diethylstilbestrol in Utero

Significant increase of breast cancer in DES Daughters

2015 Study Abstract

OBJECTIVE
To evaluate the overall cancer risk, primarily breast cancer, for women exposed to diethylstilbestrol (DES) in utero in France.

METHODS
A cohort of 3 436 prenatally DES exposed women and a comparable cohort of 3256 unexposed women were recruited retrospectively from voluntary responses to questionnaires, and cases were ascertained by medical history at the time of recruitment.

Cancer Risk in Women Exposed to Diethylstilbestrol in Utero, US National Library of Medicine National Institutes of Health, Therapie, NCBI PubMed PMID: 26071143, 2015 Sep-Oct.

Image credit Amy the Nurse.

RESULTS
One hundred ninety-five cancers were observed in exposed women (136 breast cancers, and 59 in other sites) and 141 cancers in unexposed women (90 breast cancers, and 51 others). A significant increase of breast cancers was found in exposed women, with a multivariate incidence rate ratio of 2.10 (95% CI 1.60-2.76) when compared with unexposed women. When exposed women were compared with the general population in France, the standardized incidence ratio was 2.33 (95% CI 1.93-2.72).

CONCLUSION
Our results suggest a significant increase of breast cancer in prenatally DES exposed women when compared with unexposed women and with the general population. For other cancers, except clear cell carcinoma of the cervix or vagina, there was a global non-significant increase.

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Impact of prenatal exposure to diethylstilbestrol (DES) on psychological outcome

A national survey of DES daughters and unexposed controls

2017 Study Abstract

To explore whether prenatal exposure to diethylstilbestrol (DES) is associated with increased risk of poor psychological outcome independently of the occurrence of major somatic complications related to DES exposure.

Data on health outcome were collected in women prenatally exposed to DES (n = 2566) and unexposed women (n = 2967) recruited in a French national survey.

Women prenatally exposed to DES were 1.7 times more likely to have consulted a mental health specialist compared to unexposed women (adjusted odds ratio = 1.69, 95% confidence interval 1.47-1.96), independently of demographic characteristics, poor gynecological or obstetrical outcome, or history of cancer.

Impact of prenatal exposure to diethylstilbestrol (DES) on psychological outcome: a national survey of DES daughters and unexposed controls, US National Library of Medicine National Institutes of Health, Archives of women’s mental health, NCBI PubMed PMID: 28064340, 2017 Jan 7.

Image credit Alessandra.

Frequency of consultation with a mental health specialist in persons with a history of gynecological complications or cancer was comparable in women prenatally exposed to DES and unexposed women.

Findings regarding psychological outcome obtained in the high-risk group of women prenatally exposed to DES may contribute to improving identification of psychological needs of all women presenting with gynecological abnormalities.

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Simultaneous determination of environmental estrogens

DES and estradiol monitoring using Cu-BTC frameworks-sensitized electrode

Abstract

It is quite important to monitor environmental estrogens in a rapid, sensitive, simple and cost-effective manner due to their wide existence and high toxicity.

Schematic representation of the different processes and compartments that need to be monitored to characterize the fate and transport of e-EDCs in the environment via ScienceDirect.

Using 1,3,5-Benzenetricarboxylic acid (H3BTC) as the ligand and copper ions as the center, Cu-BTC frameworks with surface area of 654.6m(2)/g were prepared, and then used to construct a novel electrochemical sensing platform for diethylstilbestrol (DES) and estradiol (E2).

On the surface of Cu-BTC frameworks, two oxidation waves at 0.26V and 0.45V are observed for DES and E2, and the oxidation signals are improved greatly. The prepared Cu-BTC frameworks not only enhance the accumulation efficiency of DES and E2, but also improve their electron transfer ability. The influences of pH value, modification amount of Cu-BTC and accumulation time were examined.

Simultaneous determination of environmental estrogens: Diethylstilbestrol and estradiol using Cu-BTC frameworks-sensitized electrode, NCBI PubMed PMID: 27474301, 2016 Jun 16.

As a result, a highly-sensitive, rapid and convenient electrochemical method was developed for the simultaneous determination of DES and E2, with detection limit of 2.7nM and 1.1nM. The practical applications manifest this new sensing system is accurate and feasible.

DES DiEthylStilbestrol Resources

X chromosome dosage compensation and sex-biased gene expression relashionship

Sex-Biased Expression of the Caenorhabditis elegans X Chromosome is a result of Both X Chromosome Copy Number and Sex-Specific Gene Regulation

Abstract

Dosage compensation mechanisms equalize the level of X chromosome expression between sexes. Yet the X chromosome is often enriched for genes exhibiting sex-biased, i.e. imbalanced expression. The relationship between X chromosome dosage compensation and sex-biased gene expression remains largely unexplored. Most studies determine sex biased gene expression without distinguishing between contributions from X chromosome copy number (dose) and the animal’s sex.

Sex-Biased Expression of the Caenorhabditis elegans X Chromosome is a result of Both X Chromosome Copy Number and Sex-Specific Gene Regulation, Genetics, NCBI pubmed/27356611, 2016 Jun 29.

Here, we uncoupled X chromosome dose from sex-specific gene regulation in C. elegans to determine the effect of each on X expression. In early embryogenesis, when dosage compensation is not yet fully active, X chromosome dose drives the hermaphrodite-biased expression of many X-linked genes, including several genes that were shown to be responsible for hermaphrodite fate. A similar effect is seen in the C. elegans germline, where X chromosome dose contributes to higher hermaphrodite X expression, suggesting that lack of dosage compensation in the germline may have a role in supporting higher expression of X chromosomal genes with female-biased functions in the gonad. In the soma, dosage compensation effectively balances X expression between the sexes. As a result, somatic sex-biased expression is almost entirely due to sex-specific gene regulation. These results suggest that lack of dosage compensation in different tissues and developmental stages allow X chromosome copy number to contribute to sex-biased gene expression and function.

DES-related studies

Possible relationship between endocrine disrupting chemicals and hormone dependent gynecologic cancers

Approximately 800 chemicals are known or suspected to have the potential to function as EDC

Abstract

The effects of the natural and synthetic estrogens have been studied for a long time but the data regarding estrogen related chemicals (endocrine disrupting chemicals, EDCs) and their effects on reproductive system are scarce. EDCs are hormone like agents that are readily present in the environment, which may alter the endocrine system of humans and animals.

Approximately 800 chemicals are known or suspected to have the potential to function as EDC.

Medical hypotheses, NCBI PubMed PMID: 27241264, 2016 Jul;92:84-7. doi: 10.1016/j.mehy.2016.04.041. Epub 2016 Apr 25

Expression of Estrogen by Libertas Academica.

Potential role of EDCs on reproductive disease has gained attention in medical literature in recent years. We hypothesize that exposure to low doses of EDCs in a chronic manner could cause hormone dependent genital cancers including ovarian and endometrial cancer. Long term exposure to low concentrations of EDCs may exert potentiation effect with each other and even with endogenous estrogens and could inhibit enzymes responsible for estrogen metabolism. Exposure time to these EDCs is essential as we have seen from Diethylstilbestrol experience. Dose-response curves of EDCs are also unpredictable. Hence mode of action of EDCs are more complex than previously thought. In the light of these controversies lower doses of EDCs in long term exposure is not harmless.

Possibility of this relationship and this hypothesis merit further investigation especially through in vivo studies that could better show the realistic environmental exposure. With the confirmation of our hypothesis, possible EDCs could be identified and eliminated from general use as a public health measure.

DES DiEthylStilbestrol Resources

Pesticide glyphosate disrupts uterine development

Neonatal exposure to a glyphosate based herbicide alters the development of the rat uterus

Glyphosate herbicide disrupts the development of the uterus of female rats when they are exposed for 7 days after birth.

A new study showed glyphosate caused cell proliferation and structural changes in the rats’ uterus, in spite of the fact that no signs of chronic or acute toxicity or differences in weight gain were seen in treated pups.

Glyphosate pesticide also disrupted the expression of proteins involved in uterine development.

The Argentine researchers conclude that glyphosate exposure may affect female fertility ; promote the development of uterine cancer.

Abstract

Neonatal exposure to a glyphosate based herbicide alters the development of the rat uterus, Toxicology NCBI PubMed PMID: 27287056, 2016 Jun 7.

Roundup “probably causing cancer”  image by Global Justice Now.

Glyphosate-based herbicides (GBHs) are extensively used to control weeds on both cropland and non-cropland areas. No reports are available regarding the effects of GBHs exposure on uterine development.

We evaluated if neonatal exposure to a GBH affects uterine morphology, proliferation and expression of proteins that regulate uterine organogenetic differentiation in rats.

Female Wistar pups received saline solution (control, C) or a commercial formulation of glyphosate (GBH, 2mg/kg) by sc injection every 48h from postnatal day (PND) 1 to PND7. Rats were sacrificed on PND8 (neonatal period) and PND21 (prepubertal period) to evaluate acute and short-term effects, respectively.

The uterine morphology was evaluated in hematoxylin and eosin stained sections. The epithelial and stromal immunophenotypes were established by assessing the expression of luminal epithelial protein (cytokeratin 8; CK8), basal epithelial proteins (p63 and pan cytokeratin CK1, 5, 10 and 14); and vimentin by immunohistochemistry (IHC). To investigate changes on proteins that regulate uterine organogenetic differentiation we evaluated the expression of estrogen receptor alpha (ERα), progesterone receptor (PR), Hoxa10 and Wnt7a by IHC.

  • The GBH-exposed uteri showed morphological changes, characterized by an increase in the incidence of luminal epithelial hyperplasia (LEH) and an increase in the stromal and myometrial thickness.
  • The epithelial cells showed a positive immunostaining for CK8, while the stromal cells for vimentin.
  • GBH treatment increased cell proliferation in the luminal and stromal compartment on PND8, without changes on PND21.
  • GBH treatment also altered the expression of proteins involved in uterine organogenetic differentiation. PR and Hoxa10 were deregulated both immediately and two weeks after the exposure. ERα was induced in the stromal compartment on PND8, and was downregulated in the luminal epithelial cells of gyphosate-exposed animals on PND21.
  • GBH treatment also increased the expression of Wnt7a in the stromal and glandular epithelial cells on PND21.

Glyphosate herbicide disrupts uterine development, gmwatch, 21 June 2016.

Neonatal exposure to GBH disrupts the postnatal uterine development at the neonatal and prepubertal period. All these changes may alter the functional differentiation of the uterus, affecting the female fertility and/or promoting the development of neoplasias.

Adverse health effects in children of women exposed in utero to diethylstilbestrol

2016 study results confirmed a transgenerational transmission of defects in male genital tract

Abstract

OBJECTIVE
Exposure to diethylstilbestrol (DES) in utero is associated with adverse health effects, including genital anomalies in women and men, and cancers in women. Animal studies showed birth defects and tumors in the offspring of DES exposed mice, revealing transgenerational transmission of DES effects. In humans, birth defects, such as hypospadias were observed in children of prenatally exposed women. The aim of this research was to further assess the health effects in children of prenatally exposed women.

Adverse health effects in children of women exposed in utero to diethylstilbestrol (DES), US National Library of Medicine, NCBI pubmed/27203157, 2016 Feb 5.

Russian doll beauty and the beast, rawdonfox.

METHODS
In a retrospective cohort study, the reports of women exposed to DES in utero on their 4409 children were compared with those of unexposed women on their 6203 children. Comparisons used odd ratios (OR) between children of exposed and unexposed women and standardized incidence rate (SIR) with the general population. These cohorts were recruited on a voluntary basis to answer questionnaires.

RESULTS
There was a global increase of defects in children born to exposed women when compared with those born to unexposed (OR 2.29, 95% CI: 1.80-2.79, P<0.001) and with the general population (SIR 2.39, 95% CI: 2.11-2.68). Increased defects were observed in male genital tract, esophagus, lip or palate, musculoskeletal and circulatory systems. For female genital tract anomalies, there was no significant increase. However, this cohort being relatively young, further follow-up is needed. An increase of cerebral palsy was revealed. The incidence of cancers was not increased, in particular for breast, uterus and ovary.

CONCLUSION
Our results confirmed a transgenerational transmission of defects in male genital tract. With caution due to possible bias associated with this method, our data suggest an increase of defects for esophagus, lip or palate, musculoskeletal and circulatory system in children of exposed women.

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Prenatal exposure to DiEthylStilbestrol and sexual orientation in men and women

Prenatal endocrine influences on sexual orientation and on sexually differentiated childhood behavior

Abstracts

Introduction

Both sexual orientation and sex-typical childhood behaviors, such as toy, playmate and activity preferences, show substantial sex differences, as well as substantial variability within each sex. In other species, behaviors that show sex differences are typically influenced by exposure to gonadal steroids, particularly testosterone and its metabolites, during early development (prenatally or neonatally). This article reviews the evidence regarding prenatal influences of gonadal steroids on human sexual orientation, as well as sex-typed childhood behaviors that predict subsequent sexual orientation. The evidence supports a role for prenatal testosterone exposure in the development of sex-typed interests in childhood, as well as in sexual orientation in later life, at least for some individuals. It appears, however, that other factors, in addition to hormones, play an important role in determining sexual orientation. These factors have not been well-characterized, but possibilities include direct genetic effects, and effects of maternal factors during pregnancy. Although a role for hormones during early development has been established, it also appears that there may be multiple pathways to a given sexual orientation outcome and some of these pathways may not involve hormones.

PRENATAL EXPOSURE TO DES AND SEXUAL ORIENTATION IN MEN

The possibility that exposure to ovarian hormones before birth influences sexual orientation in males also has been investigated. These studies have produced largely negative results. One study compared two groups of men exposed to the synthetic estrogen, DES, prenatally to matched controls. One group included 17 men exposed to DES alone and the second included 21 men exposed to DES along with natural progesterone. The study also included 10 men exposed prenatally to natural progesterone alone and 13 men exposed prenatally to synthetic progestins alone. Each of these groups was compared to matched controls. None of the four groups of hormone-exposed men differed from their respective controls in sexual orientation in fantasy or behavior. In addition, for all four samples combined, non-heterosexual orientation was reported by 8 of the 61 hormone-exposed men (13%), and by 8 of the 60 control men (13%). Two other research teams also have looked at sexual orientation in men exposed to DES prenatally, and have found no evidence of reduced heterosexual orientation. One studied 46 men exposed to DES and 29 unexposed controls. Men exposed to DES had somewhat more heterosexual coital experience than did controls, but did not differ in the number of heterosexual or homosexual coital partners. The second compared 1,342 DES-exposed men to 1,342 unexposed men, and found no difference in the numbers reporting sexual experience with a partner of the same sex, although, as noted above, this study used a relatively insensitive procedure for assessing sexual orientation. Nevertheless, the findings overall suggest that prenatal exposure to estrogen does not feminize sexual orientation in developing males, and this conclusion is consistent with predictions from results of experimental studies in other species, where early exposure of male animals to estrogen does not promote the development of female-typical behavior.

Prenatal exposure to DES and sexual orientation in women

One research team has studied three samples of women exposed prenatally to DES. The first sample included 30 women exposed to DES, 30 unexposed women recruited from the same gynaecological clinic and 12 unexposed sisters of the DES-exposed women. All of the participants had abnormal PAP smear findings. (Although DES rarely, if ever, causes genital virilization, prenatal exposure is often associated with abnormal PAP smears). Sexual orientation was assessed by interview and rated using Kinsey scale scores, and a global rating for lifelong sexual responsiveness (behavior and fantasy combined) was reported for 29 of the DES-exposed women and 30 of the controls. DES exposure was associated with reduced heterosexual orientation. Although 76% of the DES-exposed women were exclusively or almost exclusively heterosexual for lifetime scores, 24% were not. The comparable figure for the matched controls with abnormal PAP smear findings was 0%. The subset of 12 sister pairs showed a similar pattern with 42% of the DES-exposed sisters being not exclusively or almost exclusively heterosexual for their lifetime in terms of fantasy or behavior, compared to 8% of their unexposed sisters. Among the total group of DES-exposed women, five had experienced homosexual activities involving genital contact and two were living with a female partner. The same research team later reported data from this initial study along with data from two more samples of women exposed to DES prenatally. The first additional sample included 30 DES-exposed women, 30 demographically matched controls, with no history of DES-exposure or abnormal PAP smears, and 8 unexposed sisters. In this sample, a global Kinsey rating for lifelong sexual responsiveness was reported for 29 of the DES-exposed women and 30 of the matched controls. For the exposed group, 35% were not exclusively or almost exclusively heterosexual, whereas for the control group the comparable figure was 13%. Among the 20 sister pairs in the first and second samples, 40% of the DES-exposed group, compared to 5% of their sisters, were not exclusively or almost exclusively heterosexual. The second additional sample included 37 DES-exposed women whose mothers’ obstetrical files indicated prescription of at least 1000 mg of DES during the pregnancy, and age-matched daughters of women from the same obstetrical practice, whose mothers’ files showed that no DES was prescribed. For these women, 16% of the DES-exposed group and 5% of the unexposed group were not exclusively or almost exclusively heterosexual. For all three samples combined, 24% of the DES-exposed women, and 6% of the control women were not exclusively or almost exclusively heterosexual.

Prenatal endocrine influences on sexual orientation and on sexually differentiated childhood behavior, National Institutes of Health, Front Neuroendocrinol; 32(2): 170–182, NCBI PubMed PMC3296090, 2011 Apr

A separate investigation of women exposed to DES prenatally concluded that this exposure did not influence their sexual orientation. This study included 3,946 women exposed prenatally to DES and 1,740 women not exposed to DES. The DES-exposed women were somewhat less likely than the unexposed women to have had sex with a female partner. The DES-exposed women also were more likely than the unexposed women to have ever married, and for those who had had sexual intercourse with a man, were less likely to have had sexual intercourse before age 17 or to have had more than one sexual partner. These last differences raise questions about the comparability of the exposed and unexposed groups, and, although the large sample is impressive, the assessment of sexual orientation, in terms of a single question regarding sexual behavior, is relatively insensitive.

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