The steroid hormone estriol (E3) regulates epigenetic programming of fetal mouse brain and reproductive tract.

BACKGROUND: Estriol (E3) is a steroid hormone formed only during pregnancy in primates including humans. Although E3 is synthesized at large amounts through a complex pathway involving the fetus and placenta, it is not required for the maintenance of pregnancy and has classically been considered virtually inactive due to associated very weak canonical estrogen signaling. However, estrogen exposure during pregnancy may have an effect on organs both within and outside the reproductive system, and compounds with binding affinity for estrogen receptors weaker than E3 have been found to impact reproductive organs and the brain. Here, we explore potential effects of E3 on fetal development using mouse as a model system. RESULTS: We administered E3 to pregnant mice, exposing the fetus to E3. Adult females exposed to E3 in utero (E3-mice) had increased fertility and superior pregnancy outcomes. Female and male E3-mice showed decreased anxiety and increased exploratory behavior. The expression levels and DNA methylation patterns of multiple genes in the uteri and brains of E3-mice were distinct from controls. E3 promoted complexing of estrogen receptors with several DNA/histone modifiers and their binding to target genes. E3 functions by driving epigenetic change, mediated through epigenetic modifier interactions with estrogen receptors rather than through canonical nuclear transcriptional activation. CONCLUSIONS: We identify an unexpected functional role for E3 in fetal reproductive system and brain. We further identify a novel mechanism of estrogen action, through recruitment of epigenetic modifiers to estrogen receptors and their target genes, which is not correlated with the traditional view of estrogen potency.

Molecular mechanisms of estrogen action in female genital tract development.

The development of the female reproductive tract can be divided into three parts consisting of Müllerian duct organogenesis, pre-sexual maturation organ development, and post-sexual maturation hormonal regulation. In primates, Müllerian duct organogenesis proceeds in an estrogen independent fashion based on transcriptional pathways that are suppressed in males by the presence of AMH and SRY. However, clinical experience indicates that exposure to xenoestrogens such as diethylstilbestrol (DES) during critical periods including late organogenesis and pre-sexual maturational development can have substantial effects on uterine morphology, and confer increased risk of disease states later in life. Recent evidence has demonstrated that these effects are in part due to epigenetic regulation of gene expression, both in the form of aberrant CpG methylation, and accompanying histone modifications. While xenoestrogens and selective estrogen receptor modulators (SERMS) both can induce non-canonical binding confirmations in estrogen receptors, the primate specific fetal estrogens Estriol and Estetrol may act in a similar fashion to alter gene expression through tissue specific epigenetic modulation.

Fetal Bisphenol-A Induced Changes in Murine Behavior and Brain Gene Expression Persisted in Adult-aged Offspring.

In utero Bisphenol A (BPA) exposure has been linked to many deficits during brain development, including sexual differentiation, behavior, and motor coordination. Yet, how BPA induces these disorders and whether its effects are long lasting are largely unknown. In this study, using a mouse model, we demonstrated that in utero exposure to an environmentally relevant dose of BPA induced locomotor deficits, anxiety-like behavior, and declarative memory impairments that persisted into old age (18 months). Compared to the control animals, the BPA-exposed mice had a significant decrease in locomotor activity, exploratory tendencies, and long-term memory, and an increase in anxiety. The global brain gene expression profile was altered permanently by BPA treatment and showed regional and sexual differences. The BPA-treated male mice had more changes in the hippocampus, while female mice experienced more changes in the cortex. Overall, we demonstrate that in utero exposure to BPA induces permanent changes in brain gene expression in a region-specific and sex-specific manner, including a significant decrease in locomotor activity, learning ability, long-term memory, and an increase in anxiety. Fetal/early life exposures permanently affect neurobehavioral functions that deteriorate with age; BPA exposure may compound the effects of aging.

N6-methyladenine in DNA antagonizes SATB1 in early development.

The recent discovery of N6-methyladenine (N6-mA) in mammalian genomes suggests that it may serve as an epigenetic regulatory mechanism1. However, the biological role of N6-mA and the molecular pathways that exert its function remain unclear. Here we show that N6-mA has a key role in changing the epigenetic landscape during cell fate transitions in early development. We found that N6-mA is upregulated during the development of mouse trophoblast stem cells, specifically at regions of stress-induced DNA double helix destabilization (SIDD)2-4. Regions of SIDD are conducive to topological stress-induced unpairing of the double helix and have critical roles in organizing large-scale chromatin structures3,5,6. We show that the presence of N6-mA reduces the in vitro interactions by more than 500-fold between SIDD and SATB1, a crucial chromatin organizer that interacts with SIDD regions. Deposition of N6-mA also antagonizes SATB1 function in vivo by preventing its binding to chromatin. Concordantly, N6-mA functions at the boundaries between euchromatin and heterochromatin to restrict the spread of euchromatin. Repression of SIDD-SATB1 interactions mediated by N6-mA is essential for gene regulation during trophoblast development in cell culture models and in vivo. Overall, our findings demonstrate an unexpected molecular mechanism for N6-mA function via SATB1, and reveal connections between DNA modification, DNA secondary structures and large chromatin domains in early embryonic development.

Adult bone marrow progenitors become decidual cells and contribute to embryo implantation and pregnancy.

Decidua is a transient uterine tissue shared by mammals with hemochorial placenta and is essential for pregnancy. The decidua is infiltrated by many immune cells promoting pregnancy. Adult bone marrow (BM)-derived cells (BMDCs) differentiate into rare populations of nonhematopoietic endometrial cells in the uterus. However, whether adult BMDCs become nonhematopoietic decidual cells and contribute functionally to pregnancy is unknown. Here, we show that pregnancy mobilizes mesenchymal stem cells (MSCs) to the circulation and that pregnancy induces considerable adult BMDCs recruitment to decidua, where some differentiate into nonhematopoietic prolactin-expressing decidual cells. To explore the functional importance of nonhematopoietic BMDCs to pregnancy, we used Homeobox a11 (Hoxa11)-deficient mice, having endometrial stromal-specific defects precluding decidualization and successful pregnancy. Hoxa11 expression in BM is restricted to nonhematopoietic cells. BM transplant (BMT) from wild-type (WT) to Hoxa11-/- mice results in stromal expansion, gland formation, and marked decidualization otherwise absent in Hoxa11-/- mice. Moreover, in Hoxa11+/- mice, which have increased pregnancy losses, BMT from WT donors leads to normalized uterine expression of numerous decidualization-related genes and rescue of pregnancy loss. Collectively, these findings reveal that adult BMDCs have a previously unrecognized nonhematopoietic physiologic contribution to decidual stroma, thereby playing important roles in decidualization and pregnancy.

N(6)-Methyladenine in eukaryotes.

DNA modifications are a major form of epigenetic regulation that eukaryotic cells utilize in concert with histone modifications. While much work has been done elucidating the role of 5-methylcytosine over the past several decades, only recently has it been recognized that N(6)-methyladenine (N6-mA) is present in quantifiable and biologically active levels in the DNA of eukaryotic cells. Unlike prokaryotes which utilize N6-mA to recognize "self" from "foreign" DNA, eukaryotes have been found to use N6-mA in varying ways, from regulating transposable elements to gene regulation in response to hypoxia and stress. In this review, we examine the current state of the N6-mA in research field, and the current understanding of the biochemical mechanisms which deposit and remove N6-mA from the eukaryotic genome.

Hematogenous Dissemination of Mesenchymal Stem Cells from Endometriosis.

Endometriosis is ectopic growth of endometrial tissue traditionally thought to arise through retrograde menstruation. We aimed to determine if cells derived from endometriosis could enter vascular circulation and lead to hematogenous dissemination. Experimental endometriosis was established by transplanting endometrial tissue from DsRed+ mice into the peritoneal cavity of DsRed- mice. Using flow cytometry, we identified DsRed+ cells in blood of animals with endometriosis. The circulating donor cells expressed CXCR4 and mesenchymal stem cell (MSC) biomarkers, but not hematopoietic stem cell markers. Nearly all the circulating endometrial stem cells originated from endometriosis rather than from the uterus. Cells expressing DsRed, CXCR4, and MSCs markers were identified in the peritoneal wall and surrounding vessels of recipient mice, contributing to both endometriosis and angiogenesis. Cells originating in endometriosis lesions migrated and implanted in lung tissue and displayed makers of differentiation, indicating retained multipotency. In vitro these cells demonstrated multipotency and were able to differentiate into adipogenic, osteogenic, and chondrogenic lineages. Endometriosis lesions also expressed high levels of CXCL12, the CXCR4 receptor ligand. Serum CXCL12 levels were greater than in sham control mice. In humans with endometriosis, serum CXCL12 levels were significantly higher than controls, suggesting that the CXCL12/CXCR4 axis is operational in women with spontaneous endometriosis as well. Stem cells, rather than differentiated cells from endometriosis, enter the circulation in response to CXCL12. We identify an endometriosis-derived stem cell population, a potential mechanism of dissemination of this disease and a potential target for treatment of endometriosis. Stem Cells 2018;36:881-890.

Preferential epigenetic programming of estrogen response after in utero xenoestrogen (bisphenol-A) exposure.

Bisphenol-A (BPA) is an environmentally ubiquitous estrogen-like endocrine-disrupting compound. Exposure to BPA in utero has been linked to female reproductive disorders, including endometrial hyperplasia and breast cancer. Estrogens are an etiological factor in many of these conditions. We sought to determine whether in utero exposure to BPA altered the global CpG methylation pattern of the uterine genome, subsequent gene expression, and estrogen response. Pregnant mice were exposed to an environmentally relevant dose of BPA or DMSO control. Uterine DNA and RNA were examined by using methylated DNA immunoprecipitation methylation microarray, expression microarray, and quantitative PCR. In utero BPA exposure altered the global CpG methylation profile of the uterine genome and subsequent gene expression. The effect on gene expression was not apparent until sexual maturation, which suggested that estrogen response was the primary alteration. Indeed, prenatal BPA exposure preferentially altered adult estrogen-responsive gene expression. Changes in estrogen response were accompanied by altered methylation that preferentially affected estrogen receptor-α (ERα)-binding genes. The majority of genes that demonstrated both altered expression and ERα binding had decreased methylation. BPA selectively altered the normal developmental programming of estrogen-responsive genes via modification of the genes that bind ERα. Gene-environment interactions driven by early life xenoestrogen exposure likely contributes to increased risk of estrogen-related disease in adults.-Jorgensen, E. M., Alderman, M. H., III, Taylor, H. S. Preferential epigenetic programming of estrogen response after in utero xenoestrogen (bisphenol-A) exposure.