Background The increasing incidence of reproductive disorders in humans has been attributed to in utero exposure to estrogenic endocrine disruptors. In particular, exposure of the developing testis to exogenous estrogen can negatively impact male reproductive health. To determine how estrogens impact human gonad function, we treated the human testis-derived cell line NT2/D1 with estrogen and examined its impact on SOX9 and the expression of key markers of granulosa (ovarian) and Sertoli (testicular) cell development. Results Estrogen successfully activated its cognate receptor (estrogen receptor alpha; ESR1) in NT2/D1 cells. We observed a significant increase in cytoplasmic SOX9 following estrogen treatment. After 48 h of estrogen exposure, mRNA levels of the key Sertoli cell genes SOX9, SRY, AMH, FGF9 and PTGDS were significantly reduced. This was followed by a significant increase in mRNA levels for the key granulosa cell genes FOXL2 and WNT4 after 96 h of estrogen exposure. Conclusions These results are consistent with estrogen's effects on marsupial gonads and show that estrogen has a highly conserved impact on gonadal cell fate decisions that has existed in mammals for over 160 million years. This effect of estrogen presents as a potential mechanism contributing to the significant decrease in male fertility and reproductive health reported over recent decades. Given our widespread exposure to estrogenic endocrine disruptors, their effects on SOX9 and Sertoli cell determination could have considerable impact on the adult testis.

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Exposure to estrogenic endocrine disrupting chemicals (EDCs) during in utero development has been linked to the increasing incidence of disorders of sexual development (DSDs). Hypospadias, the ectopic placement of the urethra on the ventral aspect of the penis, is one of the most common DSDs affecting men, and can also affect women by resulting in the misplacement of the urethra. This study aimed to comprehensively assess the resulting hypospadias phenotypes in male and female mice exposed in utero from embryonic day 9.5 to 19.5 to the potent estrogenic endocrine disruptor, diethylstilbestrol (DES), at a high, clinically relevant dose, and a low, previously untested dose, administered via water. The anogenital distance of male pups was significantly reduced and hypospadias was observed in males at a high frequency. Females exhibited hypospadias and urethral-vaginal fistula. These results demonstrate the ability of an estrogen receptor agonist to disrupt sexual development in both male and female mice, even at a low dose, administered via drinking water.

Breast cancer (BCa) incidence increases following aberrant hormone exposure, which has been linked to direct effects on estrogen receptor (ER)+ mammary epithelium. While estrogen exposure during mammary involution has been shown to drive tumour growth via neutrophils, the potential for the ER + immune microenvironment to mediate part (in addition to mammary epithelial cells) of hormonally controlled BCa risk during normal development has not been assessed. We collected mammary tissue, lymph nodes and blood from tumour naïve mice treated with, oophorectomy, estrogen (17β estradiol) or Fulvestrant. Flow cytometry was used to examine the impact on the frequency of innate and adaptive immune cells. Oophorectomy and fulvestrant decreased the proportion of macrophages, particularly pro-tumour polarized M2 macrophages and neutrophils. Conversely, dendritic cells were increased by these therapies, as were eosinophils. Estrogen increased the proportion of M2 macrophages and to a lesser extent CD4-CD8- double negative and FoxP3+ regulatory T cells but decreased CD8 + T cells and B cells. Excluding eosinophils, these changes were restricted to the mammary tissue. This suggests that inhibiting estrogen action lowers the immune suppressive myeloid cells, increases in antigen presentation and eosinophil-mediated direct or indirect cytotoxic effects. In contrast, estrogen exposure, which drives BCa risk, increases the suppressive myeloid cells and reduces anti-tumour cytotoxic T cells. The impact of hormonal exposure on BCa risk, may in part be linked to its immune modulatory activity.

The increasing incidence of testicular dysgenesis syndrome-related conditions and overall decline in human fertility has been linked to the prevalence of oestrogenic endocrine disrupting chemicals (EDCs) in the environment. Ectopic activation of oestrogen signalling by EDCs in the gonad can impact testis and ovary function and development. Oestrogen is the critical driver of ovarian differentiation in non-mammalian vertebrates, and in its absence a testis will form. In contrast, oestrogen is not required for mammalian ovarian differentiation, but it is essential for its maintenance, illustrating it is necessary for reinforcing ovarian fate. Interestingly, exposure of the bi-potential gonad to exogenous oestrogen can cause XY sex reversal in marsupials and this is mediated by the cytoplasmic retention of the testis-determining factor SOX9 (sex-determining region Y box transcription factor 9). Oestrogen can similarly suppress SOX9 and activate ovarian genes in both humans and mice, demonstrating it plays an essential role in all mammals in mediating gonad somatic cell fate. Here, we review the molecular control of gonad differentiation and explore the mechanisms through which exogenous oestrogen can influence somatic cell fate to disrupt gonad development and function. Understanding these mechanisms is essential for defining the effects of oestrogenic EDCs on the developing gonads and ultimately their impacts on human reproductive health.