Endocrine disruptors are a rising concern for public health due to their ubiquitous presence affecting reproductive development throughout generations. We aim at studying the transgenerational effect of an EDC mixture on female sexual development and reproduction. Female rats (F0 generation) were orally exposed to a mixture of 14 anti-androgenic and estrogenic EDCs or corn oil for 2 weeks before and throughout gestation and until weaning. The mixture was composed of plasticizers (BPA, DBP, DEHP), fungicides/pesticides (Vinclozolin, Procymidon, Prochloraz, Epoxynazole, Linurone, p-p-DDT), UV filters (4-MBC, OMC), Butylparaben and the analgesic Acetaminophen. Doses were in the human exposure range (μg/kg). Sexual development and reproductive parameters (vaginal opening, GnRH secretion, estrous cyclicity and folliculogenesis) were studied from F1 to F3 generations. Maternal behavior was measured from F0 to F2 generations. At PND21, mediobasal hypothalamus of the F1 and F3 were removed for gene expression analysis (RNAseq, RT-PCR) as well as for Chromatin Immunoprecipitation of histone modifications at regulatory regions of target genes. The results show multiple multi- and transgenerational effects after ancestral EDC exposure. While F2 and F3 females showed delayed vaginal opening, decreased percentage of regular estrous cycles, decreased GnRH interpulse interval and altered folliculogenesis, no such changes were detected in F1 animals. These alterations were accompanied with transcriptional and histone posttranslational modifications of key hypothalamic genes involved in puberty and reproduction. We observed a downregulation of estrogen signaling (Esr1), genes involved in the GnRH network (Kisspeptin, Grin2d, Tac3R), maternal behavior (Th, Oxt, Avp, Drd1, Drd2) and stress responsiveness (Nr3c1). Upregulated gens involved glucocorticoid activity (Crh) and metabolism (Pomc, Cart). Concomitantly with transcriptional levels, while downregulated genes present higher levels of repressive histone marks (H3K9me3, H3K27me3) and decreased levels of activational histone marks (H3K4me3, H3K9ac), upregulated genes present the opposite pattern. Such histone marks related to changes in the polycomb/thritorax group of protein balance, involved in the control of female puberty. F1 and F2 females displayed decreased licking while spending more time resting alone. F1 RNAseq showed a reduction in Th, Drd1 and Drd2 mRNA expression. These alterations on maternal behavior are known to cause transgenerational alterations of the development of the corticotropic and gonadotropic axis. In conclusion, exposure to an environmentally relevant EDC mixture transgenerationally affects sexual development and reproduction throughout epigenetic reprogramming of the hypothalamus. While not yet clear, such effects could be mediated by alterations of maternal behavior caused by exposure to the first generation.
18 May 2019 - 21 May 2019