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Early exposure to environmental estrogens may exert lasting impacts on health. In rodents, homeobox A10 (HOXA10) was demonstrated to be a target of early endocrine disruption, as indicated by persistent changes in uterine HOXA10 expression and promoter DNA methylation in the offspring. This study aimed at analyzing long-term effects of estradiol-17β on porcine uterine HOXA10. Therefore, offspring were exposed in utero to low (0.05 and 10 μg/kg body weight/day) and high (1000 μg/kg body weight/day) doses, respectively. We, furthermore, investigated whether promoter DNA methylation was generally involved in regulating HOXA10 expression. Unexpectedly, the maternal estrogen exposure did not distinctly impact HOXA10 expression and promoter DNA methylation in either pre- or postpubertal offspring. Although differential HOXA10 expression was observed in endometrial tissue during the estrous cycle and the pre-implantation period, no concurrent substantial changes occurred regarding promoter DNA methylation. However, by comparing several tissues displaying larger differences in transcriptional abundance, HOXA10 expression correlated with promoter DNA methylation in prepubertal, but not postpubertal, gilts. Thus, promoter DNA methylation could affect gene expression in pigs, depending on their stage of development. Clearly, early estrogen exposure exerted other effects in pigs as known from studies in rodents. This may be due to endocrine differences as well as to species-specific peculiarities of tissue sensitivity to estradiol-17β during critical windows of development.

Endocrine disrupting chemicals with estrogenic activity play an important role as obesogens. However, studies investigating the most potent natural estrogen, estradiol-17β (E2), at low dose are lacking. We examined endocrine and physiological parameters in gilts receiving distinct concentrations of E2 during pregnancy. We then investigated whether adverse effects prevail in progeny due to a potential endocrine disruption. E2 was orally applied to gilts during the entire period of pregnancy. The concentrations represented a daily consumption at the recommended ADI level (0.05 μg/kg body weight/day), at the NOEL (10 μg/kg body weight/day) and at a high dosage (1000 μg/kg body weight/day). Plasma hormone concentrations were determined using enzyme immuno assays. Offspring body fat was assessed by dual-energy X-ray absorptiometry scanning. In treated gilts receiving 1000 μg E2/kg body weight/day we found significantly elevated plasma E2 levels during pregnancy, paralleled by an increased weight gain. While offspring showed similar weight at birth, piglets exhibited a significant reduction in weight at weaning even though their mothers had only received 0.05 μg E2/kg body weight/day. At 8 weeks of age, specifically males showed a significant increase in overall body fat percentage. In conclusion, prenatal exposure to low doses of E2 affected pig offspring development in terms of body weight and composition. In line with findings from other obesogens, our data suggest a programming effect during pregnancy for E2 causative for the depicted phenotypes. Therefore, E2 exposure may imply a possible contribution to childhood obesity.

In utero and early postnatal exposure to exogenous oestrogens may alter the epigenome, possibly influencing health later in life. The reproductive organs are the main target of oestrogens. Uterine HOXA10 expression and promoter DNA methylation have been shown to be sensitive to endocrine disruptors in rodents. In the endometrium of pigs, in vitro exposure to estradiol-17β resulted in an increase of HOXA10 mRNA and protein expression, and a regulation for HOXA10 has also been shown during early pregnancy. However, it is unclear whether modification in promoter DNA methylation was involved herein. In the present study, we investigated the effect of oral estradiol-17β supplementation to pregnant pigs from Day 0 until delivery (0, 0.05, 10, and 1000 µg kg–1 per day; n = 8–12/group) on HOXA10 expression and promoter DNA methylation in the uterus of prepubertal and the endometrium of postpubertal porcine offspring. In addition, endometrial samples from 2 physiological situations were analyzed, namely the oestrous cycle at Day 0, 3, 6, 12, and 18 (n = 6/day), and early pregnancy at Day 10, 12, and 14 (n = 4/day). Furthermore, different reproductive and nonreproductive tissues from pre- and postpubertal untreated animals were investigated (n = 2/tissue). Gene expression was measured using RT-qPCR. The DNA was first bisulfite converted, amplified by using methylation-sensitive high-resolution melting qPCR, followed by pyrosequencing. Statistical analyses were conducted using one-way ANOVA. In utero estradiol-17β exposure did neither alter HOXA10 expression compared to the control group nor HOXA10 promoter DNA methylation in pre- and postpubertal porcine offspring, respectively. Because the prepubertal group displayed higher HOXA10 mRNA expression than did postpubertal animals, while DNA methylation was lower in the younger animals, a potential role of DNA methylation may be assumed. During the oestrous cycle, HOXA10 expression was significantly regulated (P

In male developing piglets, low endogenous oestrogenic hormone levels prevail. Sensitive tissues expressing high levels of oestrogen receptors, such as the epididymis, might thus be of specific relevance for early disturbances by estrogenic endocrine disruptors. We have previously shown that distinct concentrations of the natural oestrogen oestradiol-17β (E2) orally applied during the whole period of pregnancy in sows lead to a distinct shift in body composition. Therefore, we aimed at investigating whether prenatal E2 exposure had the potential to program male reproductive organs toward altered semen quality. The E2 treatment groups (n = 4 sows/group) were designed to represent doses of pharmacological relevance (PR; 1000 µg kg–1 of body weight per day), oral no-effect level (10 µg kg–1 of body weight per day), and acceptable daily intake level (0.05 µg kg–1 of body weight per day). Total RNA was extracted from the epididymis of 8-week-old piglet twins (n = 2 per sow). The RNA of littermates was pooled and subjected to microarray analysis (MA) using Agilent Porcine Genome Arrays (Agilent Technologies Inc., Santa Clara, CA, USA). Ejaculates of adult male pigs from the control group and the pharmacological group (n = 4) were collected and analysed for quantitative and qualitative parameters by using computer-assisted sperm analysis and flow cytometry. In total, 105 transcripts exhibited a significant difference of more than 1.5-fold for at least one of the treatment groups compared with the control group (P