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Perfluorooctane sulfonate (PFOS) is a widespread and persistent chemical in the environment. Reports show that PFOS is a potential endocrine disruptor; however, the possible effects of PFOS on placental endocrine function are unclear. This study aimed to investigate the endocrine-disrupting effects of PFOS on the placenta in pregnant rats and its potential mechanism. Pregnant rats from gestational days 4-20 were exposed to 0, 10, and 50 μg/mL PFOS through drinking water followed by analysis of various biochemical parameters. PFOS dose-dependently decreased fetal and placental weight in both sexes, with a specific decrease in weight of labyrinth but not junctional layer. Plasma progesterone (↑166%), aldosterone (↑201%), corticosterone (↑205%), testosterone (↑45%), luteinizing hormone (↑49%) levels were significantly increased, while estradiol (↓27%), prolactin (↓28%) and hCG (↓62%) levels were reduced in groups exposed to higher doses of PFOS. Real-time quantitative reverse transcriptase-polymerase chain reaction analysis revealed a significant increase in mRNA levels of placental steroid biosynthesis enzymes, including Cyp11A1 and 3β-HSD1 in male placenta and StAR, Cyp11A1, 17β-HSD1 and 17β-HSD3 in female placenta of PFOS dams. Cyp19A1 expression in ovaries was significantly decreased in PFOS dams. mRNA levels for placental steroid metabolism enzyme UGT1A1 increased in male but not in female placenta of PFOS dams. These results suggest that the placenta is a target tissue of PFOS and PFOS-induced dysregulation in steroid hormone production might be related to the altered expression of hormone biosynthesis and metabolism enzyme genes in the placenta. This hormone disruption might affect maternal health and fetal growth., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Background: Triclosan (TCS) has been detected in human tissues. It can disrupt steroidogenesis in vivo. The study on the effects of TCS on ovarian granulosa cells was lacking., Methods: Primary rat granulosa cells (rGCs) were treated with TCS. Concentrations of estradiol (E 2 ), progesterone (P 4 ) in the cell culture supernatants were measured. Microarray was used to measure gene expression profiles. Pathway analysis was performed to identify signaling networks that linked differentially expressed genes (DEGs). Genes related with steroidogenesis were analyzed., Results: TCS increased E 2 and P 4 production. A total of 2006 DEGs were identified. Pathway analysis revealed that ovarian steroidogenesis pathway was upregulated. Both PCR and Western-blot demonstrated that the expressions of key genes involved in this pathway were significantly increased., Conclusions: TCS co-administered with follicle-stimulating hormone (FSH) could increase E 2 and P 4 production in rGCs and up-regulate ovarian steroidogenesis pathway. StAR and aromatase protein were increased by TCS, while P450scc protein wasn't changed significantly., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Glyphosate has been suggested to be an endocrine disrupting chemical capable of disrupting male reproduction. There are conflicting data, however, with studies reporting effects from exposure to either glyphosate alone or to herbicide formulations, making comparisons difficult. We assessed rat testis histopathology and androgen function following two weeks exposure to either glyphosate at 2.5 and 25 mg/kg bw/day (5x and 50x Acceptable Daily Intake, ADI, respectively), or equivalent high dose of glyphosate in a herbicide formulation; Glyfonova. We observed no significant effects on testes or testosterone synthesis in rats exposed to glyphosate. Limited effects were observed in rats exposed to Glyfonova, with a small upregulation of the steroidogenic genes Cyp11a1 and Cyp17a1. We conclude that glyphosate alone has no effect on adult rat testis at exposure levels up to 25 mg/kg bw/day. Glyfonova induced only minor effects on steroidogenic gene expression, likely caused by additives other than glyphosate., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Bisphenol A (BPA) is a commonly used plasticizer. Previous studies show that in utero exposure to BPA affects reproductive outcomes in the F1-F3 generations of mice. However, its multigenerational effects on ovarian histology and steroidogenesis over the reproductive lifespan are unknown. Thus, we tested the hypothesis that BPA has multigenerational effects on follicle numbers and steroidogenesis. Mice were exposed in utero to vehicle control or BPA (0.5, 20, and 50μg/kg/day). Ovaries were collected for histological and gene expression analyses and sera were collected for hormone assays. In utero BPA exposure decreased preantral follicle numbers, cytochrome P450 aromatase mRNA levels, and estradiol levels in the F1 generation, whereas it decreased testosterone levels and altered steroidogenic acute regulatory protein, cytochrome P450 cholesterol side-chain cleavage, 3β-hydroxysteroid dehydrogenase 1, and cytochrome P450 aromatase mRNA levels in the F2 generation. These data suggest that BPA has multigenerational effects on the ovary in mice., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Perfluorooctanoic acid (PFOA) is a synthetic perfluorinated compound, which has been reported to exert adverse effect on the pregnancy. However, whether it is associated with alteration of luteal function remains unknown. Mice were administered PFOA by gavage from gestational days (GD) 1-7 or 13. PFOA treatment did not significantly affect numbers of embryo implantation. Nevertheless, on GD 13, 10mg/kg PFOA treatment significantly increased numbers of resorbed embryo. Furthermore, PFOA exposure markedly reduced serum progesterone levels but did not affect estradiol levels. Treatment also showed concomitant decreases in transcript levels for key steroidogenic enzymes, and reduced numbers and sizes of corpora lutea. In addition, PFOA administration inhibited activities of superoxide dismutase and catalase, and increased generation of hydrogen peroxide and malondialdehyde, and down-regulated level of Bcl-2 and up-regulated p53 and BAX proteins. In conclusion, PFOA exposure significantly inhibits luteal function via oxidative stress and apoptosis in pregnant mice., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Exposure to di-butyl phthalate (DBP) exerts negative effects on female fertility in animal models, but human studies remain limited. Here, the effects of DBP exposure on mural granulosa cell function were investigated in primary cultures from women undergoing in vitro fertilization. Cultured cells treated with various doses of DBP (0, 0.01μg/mL, 0.1μg/mL, 1μg/mL, 10μg/mL, or 100μg/mL) for 48h were assessed using enzyme-linked immunosorbent assay and qRT-PCR. Treatment with 100μg/mL DBP resulted in significantly lower 17β-estradiol and progesterone production (p<0.01). It also resulted in altered mRNA expression of steroidogenic, angiogenic, and epidermal growth factor-like growth factor genes: CYP11A1 (p<0.001), CYP19A1 (aromatase) (p<0.001), VEGF-A (p<0.02), BTC (p=0.009), and EREG (p=0.04). StAR expression was impaired after exposure to both 10 and 100μg/mL (p<0.03 and p<0.001, respectively). Our results indicate that in vitro exposure of granulosa cells to high doses of DBP alters cell functions., (Copyright © 2017 Elsevier Inc. All rights reserved.)

We investigated whether in vivo exposure to herbicide atrazine (ATR) exerts anovulatory effect by direct action in the ovary. Female rats were given ATR (50mg/kg body weight) during equine chorionic gonadotropin (eCG) priming. Forty eight hours after eCG administration, the animals were injected with human CG (hCG) to induce ovulation. ATR blocked ovulation and prevented expression of epiregulin and progesterone receptor mRNA in hCG-treated animals. During eCG-induced follicular growth, ATR suppressed luteinizing hormone receptor (Lhr) and aromatase expression in granulosa cells and decreased estradiol (E2) serum levels. ATR increased cytochrome p450 1b1 (Cyp1b1) mRNA expression after both in vivo and in vitro exposures. In vitro addition of beta-naphthoflavone, a known Cyp1b1 mRNA inductor, suppressed follicle-stimulating hormone-induced Lhr expression. Collectively, these data indicate that under in vivo conditions, ATR may act directly on granulosa cells by decreasing E2 levels and Lhr mRNA, thus leading to inhibition of ovulation., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Effects of triphenyl phosphate (TPP) and tris-(2-chloroethyl) phosphate (TCEP) exposure on induction of oxidative stress and endocrine disruption were investigated in TM3 cells. After 24h exposure, cell growth declined and morphology changed in TPP and TCEP treated groups with high dosages. Significant increases in superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX) and glutathione S-transferase (GST) activities and their respective gene expressions in a dose-dependent and/or time-dependent manner in TPP or TCEP groups. Moreover, the expression of main genes related to testosterone (T) synthesis including cytochrome P450 cholesterol side-chain cleavage enzyme (P450scc), cytochrome P450 17α-hydroxysteroid dehydrogenase (P450-17α), 3β-hydroxysteroid dehydrogenase (3β-HSD) and 17β-hydroxysteroid dehydrogenase (17β-HSD) were dramatically reduced by TPP and TCEP treatments, especially with the high dosage for 24h. TPP and TCEP treatments for 24h caused significant decreases in T levels in the medium. Furthermore, co-treatments of hCG with TPP or TCEP could inhibit hCG-induced changes in the expression of P450scc, P450-17α and 17β-HSD and T levels. Taken together, TPP and TCEP could induce oxidative stress and endocrine disruption in TM3 cells., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Seafood products, including fish and fish oils, are major sources of persistent organic pollutants (POPs) which may cause endocrine disruption related to reproductive dysfunction in males. Primary porcine neonatal Leydig cells were exposed to three extracts of POPs obtained from different stages in production of cod liver oil dietary supplement, in the absence and presence of luteinizing hormone (LH). No reduced viability was observed and all POP extracts showed increased testosterone and estradiol levels in unstimulated cells and decreased testosterone and estradiol secretion in LH-stimulated cells. A decrease in central steriodogenic genes including STAR, CYP11A1, HSD3B and CYP17A1 was obtained in both culture conditions with all POP extracts. We implicate both small differences in composition and concentration of compounds as well as "old" POPs to be important for the observed steroidogenic effects., (Copyright © 2015 Elsevier Inc. All rights reserved.)

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is the most toxic congener of the polyhalogenated aromatic hydrocarbons (PAH), which causes anatomical abnormalities and developmental defects, impairs ovulation and reduces fertility. TCDD's endocrine-disrupting effects are, in part, caused by a direct action at the ovary. Herein we investigated the in-vitro effects of environmentally relevant doses of TCDD on estradiol-17β (E2) production by human luteinizing granulosa cells (hLGC) obtained from women stimulated for in-vitro fertilization (IVF). TCDD at all concentrations tested (3.1fM, 3.1pM and 3.1nM) significantly decreased E2 secretion when assayed for by radioimmunoassay (RIA). Herein we confirm that TCDD alters E2 secretion by hLGC in a time-, not dose-dependent fashion and are the first to show decreases in E2 secretion with fM concentrations of TCDD. Using real-time quantitative PCR (RT-qPCR), the decreased E2 secretion correlates with a decrease in the mRNA expression levels two enzymes in the estrogen biosynthesis pathway: CYP11A1 and CYP19A1., (Copyright © 2015 Elsevier Inc. All rights reserved.)

The aim of this study was to determine whether short-term, in vivo exposure to silver nanoparticles (AgNPs) could be toxic to male reproduction. Low dose (1mg/kg/dose) AgNPs were intravenously injected into male CD1 mice over 12 days. Treatment resulted in no changes in body and testis weights, sperm concentration and motility, fertility indices, or follicle-stimulating hormone and luteinizing hormone serum concentrations; however, serum and intratesticular testosterone concentrations were significantly increased 15 days after initial treatment. Histologic evaluation revealed significant changes in epithelium morphology, germ cell apoptosis, and Leydig cell size. Additionally, gene expression analysis revealed Cyp11a1 and Hsd3b1 mRNA significantly upregulated in treated animals. These data suggest that AgNPs do not impair spermatogonial stem cells in vivo since treatment did not result in significant decreases in testis weight and sperm concentrations. However, AgNPs appear to affect Leydig cell function, yielding increasing testicular and serum testosterone levels., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The commercial bifenthrin (BF) contains two cis isomers. In the present study, a dose of 15mg/kg of 1R-cis-BF or 1S-cis-BF was orally administered for 3 weeks to female mice before or during pregnancy. Then, the expression of steroidogenesis related genes which were considered as effective biomarkers of endocrine disruption were analyzed in the male offspring. Maternal exposure to 1S-cis-BF during pregnancy significantly reduced the mRNA levels of peripheral benzodiazepine receptor (PBR) and steroidogenic acute regulatory protein (StAR) in the testes of 3- or 6-week old male offspring. In addition, a significant decrease of cytochrome P450 17α-hydroxysteroid dehydrogenase (P450-17α) was also observed in the testes of 6-week old male offspring when dams were treated with 1S-cis-BF during pregnancy but not before pregnancy. Moreover, the scavenger receptor class B type 1 (SRB1) and cytochrome P450 cholesterol side-chain cleavage enzyme (P450scc) decreased significantly in the testes of 6-week old male offspring when dams were treated with 1S-cis-BF during and before pregnancy. Thus, oral administration of the maternal mice to cis-BF for 3 weeks, particularly during pregnancy, resulted in endocrine disruption in the male offspring, with the 1S-cis-BF causing more significant alterations than the 1R-cis-BF form., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Cadmium (Cd) is a well-known testicular toxicant. However, the effects of pubertal Cd exposure on testicular development and spermatogenesis remained to be elucidated. The present study investigated the effects of pubertal Cd exposure on testicular development and spermatogenesis. Male CD-1 mice were intraperitoneally injected with CdCl(2) (1mg/kg) daily from postnatal day 35 (PND35) to PND70. As expected, pubertal Cd exposure significantly decreased the number of spermatozoa in epididymides. In addition, pubertal Cd exposure markedly reduced the weights of testes, epididymides and prostate and seminal vesicle in adult mice. A significant decrease in serum and testicular testosterone (T) was observed in mice exposed to Cd during puberty. Moreover, pubertal Cd exposure markedly reduced mRNA and protein levels of testicular StAR, P450scc, P450(17alpha) and 17beta-HSD. Taken together, these results suggest that the decreased testicular T synthesis might partially contribute to pubertal Cd-induced impairment on testicular development and spermatogenesis in mice., (Copyright 2009. Published by Elsevier Inc.)

Exposure to antiandrogens during the critical developmental window (i.e. sexual differentiation) can permanently demasculinize the male phenotype. Here we have investigated the effects of developmental exposure to di-isononylphthalate (DINP) (250 and 750 mg/kg) and 1,1-dichloro-2,2-bis(p-chlorophenyl)ethylene (p,p'-DDE) (50 and 100mg/kg) on 19.5-day-old fetal Sprague-Dawley rat testicular and adrenal steroidogenesis. Maternal exposure to DINP or p,p'-DDE on embryonic days (EDs) 13.5-17.5 did not down-regulate the activity of steroidogenesis in ED 19.5 male rat fetus. Protein expression levels of testicular and adrenal StAR, P450scc, 3beta-HSD and androgen receptor (AR) did not show any changes. However, p,p'-DDE caused clear abnormalities in the ultrastructure of steroidogenic cells in ED 19.5 rat testis and adrenal. These structural alterations can disturb the development and function of fetal testis and adrenal that may become evident later in life.

Perfluorododecanoic acid (PFDoA), one of a number of commercially important perfluoroalkyl acids, has been detected in sera from humans and other animals; however, the effects of PFDoA on female reproduction remain unclear. To assess the impact of PFDoA on puberty and endocrine status, we exposed weaned pre-pubertal female rats to PFDoA, administered orally at doses of 0, 0.5, 1.5 and 3mg/kg-d for 28 days, and measured body weight, reproductive organ weight and morphology, pubertal indicators, endocrine hormones, total serum cholesterol levels and steroidogenic enzyme gene expression. At 3mg/kg-d, PFDoA significantly decreased body weight and serum estradiol levels, increased cholesterol levels (p<0.05), and altered ovarian expression of genes responsible for cholesterol transport and steroidogenesis, including steroidogenic acute regulatory protein, cholesterol side-chain cleavage enzyme and 17-beta-hydroxysteroid dehydrogenase (p<0.05). PFDoA at the highest dose also reduced estrogen receptor alpha and beta expression levels in the ovary (p<0.05), whereas a lower concentration of PFDoA (0.5mg/kg-d) decreased estrogen receptor beta mRNA levels in the uterus (p<0.05). PFDoA treatment did not affect serum follicle-stimulating hormone or luteinizing hormone (LH) levels at any concentration, although PFDoA at 3mg/kg-d reduced LH receptor mRNA levels. There were no marked changes in sexual organ weight, age and weight at vaginal opening or first estrous cycle, or ovarian/uterine histology at any PFDoA concentration. These data show that PFDoA does not affect the endocrine status of pubertal rats, but at higher doses it does impact estradiol production and the expression of some key genes responsible for estrogen synthesis.

Exposure to the pesticide methoxychlor in rodents is linked to impaired steroid production, ovarian atrophy and reduced fertility. Following in vivo administration, it is rapidly converted by the liver to 2,2-bis-(p-hydroxyphenyl)-1,1,1-trichloroethane (HPTE), the reported active metabolite. Both methoxychlor and HPTE have weak estrogenic and antiandrogenic activities, and these effects are thought to be mediated through the estrogen and androgen receptors, respectively. Previous in vivo studies on methoxychlor exposure to female animals have demonstrated decreased progesterone production but no change in serum estrogen levels. We recently showed that HPTE specifically inhibits the P450 cholesterol side-chain cleavage (P450scc, CYP11A1) step resulting in decreased androgen production by cultured rat testicular Leydig cells. The current studies examined the mechanism of action of HPTE on progesterone production by cultured ovarian cells (granulosa and theca-interstitial) from pregnant mare serum gonadotropin-primed immature rats. In addition, we evaluated whether the effects of HPTE on rat ovarian cell progesterone biosynthesis were mediated through the estrogen or androgen receptors. Exposure to HPTE (0, 10, 50 or 100nM) alone progressively inhibited progesterone formation in cultured theca-interstitial and granulosa cells and the P450scc catalytic activity in theca-interstitial cells in a dose-dependent manner with significant declines starting at 50nM. However, HPTE did not change mRNA levels of the P450scc system (P450scc, adrenodoxin reductase and adrenodoxin) as well as P450scc protein levels. Of interest, estradiol, xenoestrogens (bisphenol-A or 4-tert-octylphenol), a pure antiestrogen (ICI 182,780), or antiandrogens (4-hydroxyflutamide or the vinclozolin metabolite M-2), had no effect on progesterone production even at 1000nM. Co-treatment of HPTE with ICI 182,780 did not block the effect of HPTE on progesterone formation. These studies suggest that the decline in progesterone formation following exposure to HPTE in cultured ovarian cells is associated with the inhibition of catalytic activity of P450scc at least in theca-interstitial cells. This action does not appear to be mediated through the estrogen or androgen receptor signaling pathways, and other chemicals exhibiting estrogenic, antiestrogenic or antiandrogenic properties do not mimic its effect on ovarian steroid production.

Mycotoxins are contaminants of animal feed that can impair fertility and cause abnormal fetal development in farm animals. The aim of the present study was to investigate the influence of Fusarium-toxin contaminated feed on cumulus morphology and maturation of pig oocytes. Naturally with the Fusarium-toxins deoxynivalenol (DON) and zearalenone (ZON) contaminated wheat was included in feed for gilts at increasing proportions which resulted in increasing dietary concentrations of both toxins (in mgtoxin/kg feed: Group 1 (control), 0.21 and 0.004; Group 2, 3.07 and 0.088; Group 3, 6.1 and 0.235; Group 4, 9.57 and 0.358, for DON and ZON, respectively). Oocytes were recovered from gilt ovaries by follicle aspiration after ovario-hysterectomy. Granulosa cells were analyzed for the expression of the P450(SCC) and 3beta-HSD mRNA by RT-PCR and additionally for P450(SCC) protein by Western blotting. Neither the expression of the P450(SCC) nor of the 3beta-HSD mRNA or the abundance of the P450(SCC) protein was significantly influenced by the mycotoxin application. The distribution of different cumulus cell morphologies was not influenced by group. At the time of recovery, oocytes with compact cumuli in Groups 3 and 4 showed a reduced proportion having immature chromatin in comparison to that for Groups 1 and 2. The proportion of oocytes having degenerated meiotic chromatin was significantly higher in Group 4 than in the other groups. The proportion of oocytes reaching metaphase II in culture was significantly lower in Groups 3 and 4 than in Group 1, and tended to be lower in Group 2 than in Group 1. We conclude that oocyte quality is significantly reduced by feeding of Fusarium-toxins to gilts.

Mycotoxins as contaminants of animal food can impair fertility and can cause abnormal fetal development in farm animals. Therefore, the present study has investigated whether derivatives of the mycotoxin zearalenone, alpha-zearalenol (alpha-ZOL) and beta-zearalenol (beta-ZOL), influence progesterone synthesis via cytochrome p450 side chain cleavage enzyme (p450scc) and 3beta-hydroxysteroid dehydrogenase/isomerase (3beta-HSD) in cultured porcine granulosa cells. Both enzymes are essential for the conversion of cholesterol to progesterone. No differences in basal progesterone levels and numbers of viable cell were observed between untreated granulosa cells and those treated with alpha- or beta-ZOL (15 and 30 microM). FSH (0.01 microg/ml) or forskolin (10 microM) enhanced the basal progesterone secretion in the absence of mycotoxins. The addition of alpha- or beta-ZOL (7.5, 15 and 30 microM) to cultures stimulated with FSH (0.01 microg) or forskolin (10 microM) reduced progesterone synthesis and the levels of p450scc and 3beta-HSD transcripts in a dose-dependent manner (P<0.05). The enzymatic activity of 3beta-HSD and the abundance of p450scc protein were also reduced by these mycotoxins. In conclusion, effects of mycotoxins on FSH receptor-dependent and receptor-independent pathways indicate that adenylate cyclase activity and/or regulatory pathways further downstream are targets of mycotoxin actions. The apparent dose-dependent reduction of p450scc and 3beta-HSD transcripts implies an effect of alpha- and beta-ZOL on transcriptional regulation of these enzymes.