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The aim of our study is to evaluate the antiproliferative effect of tocopherols obtained from alimentary virgin argan oil extracted from the endemic argan tree of Morocco and of saponins extracted from argan press cake on three human prostatic cell lines (DU145, LNCaP, and PC3). The results were compared to 2-methoxyestradiol as antiproliferative drug candidates. Cytotoxicity and antiproliferative effects were investigated after cells' treatment with tocopherols and saponins compared to 2-Methoxyoestradiol as the positive control. Tocopherols and saponins extracted from argan tree and 2-methoxyestradiol exhibit a dose-response cytotoxic effect and an antiproliferative action on the tested cell lines. The best antiproliferative effect of tocopherols is obtained with DU145 and LNCaP cell lines (28 μg/ml and 32 μg/ml, respectively, as GI 50 ). The saponins fraction displayed the best antiproliferative effect on the PC3 cell line with 18 μg/ml as GI 50 . Our results confirm the antiproliferative effect of 2-methoxyestradiol and show for the first time the antiproliferative effect of tocopherols and saponins extracted from the argan tree on hormone-dependent and hormone-independent prostate cancer cell lines. These data suggest that argan oil is of potential interest in developing new strategies for prostate cancer prevention. [ABSTRACT FROM AUTHOR]

Abstract Background In rainbow trout (Oncorhynchus mykiss), the endocrine control of spermiation is not fully understood. Besides 11ketotestosterone (11KT) and 17alpha, 20beta-dihydroxyprogesterone (MIS), the potential physiological ligand of the mineralocorticoid receptor (MR) 11-deoxycorticosterone (DOC), is a credible candidate in O. mykiss spermiation regulation as spermiation is accompanied with changes in aqueous and ionic flows. Methods In this study, we investigated potential roles of DOC during spermiation 1) by describing changes in blood plasma DOC level, MR mRNA abundance during the reproductive cycle and MR localization in the reproductive tract 2) by investigating and comparing the effects of DOC (10 mg/kg) and MIS (5 mg/kg) supplementations on sperm parameters 3) by measuring the in vitro effect of DOC on testis MIS production. Results The plasma concentration of DOC increased rapidly at the end of the reproductive cycle to reach levels that were 10–50 fold higher in mature males than in immature fish. MR mRNA relative abundance was lower in maturing testes when compared to immature testes, but increased rapidly during the spermiation period, immediately after the plasma rise in DOC. At this stage, immunohistochemistry localized MR protein to cells situated at the periphery of the seminiferous tubules and in the efferent ducts. Neither DOC nor MIS had significant effects on the mean sperm volume, although MIS treatment significantly increased the percentage of males producing milt. However, a significant reduction in the spermatocrit was observed when DOC and MIS were administrated together. Finally, we detected an inhibitory effect of DOC on testis MIS production in vitro. Conclusion These results are in agreement with potential roles of DOC and MR during spermiation and support the hypothesis that DOC and MIS mechanisms of action are linked during this reproductive stage, maybe controlling milt fluidity. They also confirm that in O. mykiss MIS is involved in spermiation induction.

Although the ovarian reserve is constituted in utero, the literature on the effects of persistent organic pollutants (POPs) during this vulnerable period on the ovarian reserve later in life is limited. We investigated whether cord blood concentrations of POPs were associated with decreased anti-Müllerian hormone (AMH, a marker of the ovarian reserve) levels in girls at the age of 12. We included 239 girls from the French mother-child PELAGIE cohort. POP concentrations of 14 organochlorine pesticides, 17 polychlorinated biphenyls (PCBs), 5 polybrominated diphenyl ethers, and 9 per-polyfluoroalkyl substances were measured on cord blood sampled at birth. During a follow-up study at 12 years old, blood samples were collected to measure AMH levels. Single-exposure associations were examined with multivariable linear regression models adjusted a priori for potential confounders. Stratification on menarche status was also performed. Mixture effects were investigated using quantile g-computation and Bayesian kernel machine regression. Overall, 16 POPs were measured in at least 30% of samples. No significant associations were found in multivariable linear regressions, except for the third tercile of exposure to PCB 180 which was statistically significantly associated with an increase in AMH levels at 12 years old (Tercile 2 v. Tercile 1: 0.13 ng/mL, 95% CI = -0.29, 0.56; Tercile 3 v. Tercile 1: 0.51 ng/mL, 95% CI = 0.02, 0.99). Additionally, in post-menarcheal girls (N = 104) only, the second tercile of p,p'-DDE was statistically significantly associated with decreased AMH levels at 12 years old (Tercile 2 v. Tercile 1: -0.61 ng/mL, 95% CI = -1.16, -0.05, Tercile 3 v. Tercile 1: 0.02 ng/mL, 95% CI = -0.51, 0.54). Both mixture models returned null associations. Despite the limited associations observed in this study, we recommend exploring these associations in larger mother-child cohorts and at older ages., 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 © 2024 Elsevier Inc. All rights reserved.)

Background and Objective: Failure rates after first-line treatment of localized prostate cancer (PCa) treatment remain high; therefore, it is essential to improve the selection and identification of at-risk patients to reduce mortality. The aim of the ANDROCAN study was to evaluate the biochemical recurrence (BCR) in patients with localized PCa treated by total prostatectomy at 5 yr after surgery, according to their presurgery gonadal status., Methods: A prospective cohort study was conducted including 1318 patients undergoing total prostatectomy for localized PCa with a 5-yr postoperative follow-up. Clinical and hormonal data (assays of total testosterone [TT], bioavailable testosterone [BT], dihydrotestosterone, estrone, and estradiol were performed by gas chromatography/mass spectrometry) as well as metabolic syndrome parameters were collected at baseline before surgery. Pathological data (predominant Gleason grade 4 and stage) were collected and cross-referenced centrally. Factors associated with BCR were assessed by a multivariate analysis, and BCR-free survival was assessed by a Kaplan-Meier analysis., Key Findings and Limitations: Among the 1318 patients, 237 had BCR of PCa. Considering demographic characteristics, populations with and without BCR were similar. However, patients with BCR had cancers with a higher Gleason score (p = 0.0001) and higher prostate-specific antigen (PSA) values (p = 0.0005) at baseline. Gleason score, pT >3a, and PSA level at baseline were positively correlated with BCR (p < 0.0001, p < 0.0001, and p = 0.0048, respectively), while BT and TT levels were not associated with BCR. This study includes patients with varying clinical characteristics, such as cancer history and metabolic syndrome, introducing variability that makes it challenging to isolate the specific effects of gonadal status on BCR. Another limitation is the lack of evaluation of long-term BCR beyond 5 yr, potentially overlooking recurrences that occur between 5 and 15 yr after surgery. This could lead to an underestimation of the actual long-term recurrence rates., Conclusions and Clinical Implications: Overall, PSA levels, high Gleason score, and pT >3a are associated with a greater likelihood of disease recurrence following initial treatment and could serve as important prognostic indicators for predicting the risk of BCR. In this prospective study, biochemical hypogonadism was not associated with a higher occurrence of BCR within 5 yr of prostatectomy. The biological gonadal status of preoperative patients could potentially be useful for therapeutic decisions but does not provide an indication for the oncological follow-up., Patient Summary: Five-year follow up of patients after surgery showed that there is no association between hypogonadism (low levels of total testosterone and bioavailable testosterone) and cancer recurrence. However, cancer recurrence seems to be more associated with aggressiveness of cancer at the time of detection., (Copyright © 2024 European Association of Urology. Published by Elsevier B.V. All rights reserved.)

Background: Advances in chromatography and mass spectrometry have allowed us to develop a novel technique for measuring intraprostatic hormone concentrations directly on prostate needle biopsies, rather than using traditional punch excision. This has significant clinical implications as intraprostatic dihydrotestosterone and testosterone levels could help monitor prostate growth, neoplasia and castration resistance., Methods: Patients undergoing radical cystoprostatectomy for bladder cancer were prospectively included. Each prostate specimen received one 90mg punch excision and six needle biopsies. Intraprostatic hormones were dosed through gas chromatography-mass spectrometry., Results: We included twenty patients, of which eleven were incidentally diagnosed with prostate cancer; four had ISUP 1 (20%) and seven had ISUP 2 (35%). The prostate biopsy technique was unable to obtain measures for testosterone, Delta-4-androsterone and androstenedione. Tissue concentrations of DHEA, DHT, E1 and E2 can be obtained with no significant difference from the reference established on a punch from a single biopsy core sample., Conclusions: Our study demonstrates that intraprostatic concentrations of DHEA, DHT, E1 and E2 can be measured without significant difference from the reference established on a single punch excision. This finding opens the way to research on the interactions between endocrinology and prostate oncogenesis and particularly on the mechanisms of resistance to hormone therapies in vivo., (Copyright © 2024 Société Française du Cancer. Published by Elsevier Masson SAS. All rights reserved.)

Background: Industrial progress has led to the omnipresence of chemicals in the environment of the general population, including reproductive-aged and pregnant women. The reproductive function of females is a well-known target of endocrine-disrupting chemicals. This function holds biological processes that are decisive for the fertility of women themselves and for the health of future generations. However, insufficient research has evaluated the risk of combined mixtures on this function. This study aimed to assess the direct impacts of a realistic exposure to eight combined environmental toxicants on the critical process of folliculogenesis., Methods: Female rabbits were exposed daily and orally to either a mixture of eight environmental toxicants (F group) or the solvent mixture (NE group, control) from 2 to 19 weeks of age. The doses were computed from previous toxicokinetic data to reproduce steady-state serum concentrations in rabbits in the range of those encountered in pregnant women. Ovarian function was evaluated through macroscopic and histological analysis of the ovaries, serum hormonal assays and analysis of the expression of steroidogenic enzymes. Cellular dynamics in the ovary were further investigated with Ki67 staining and TUNEL assays., Results: F rabbits grew similarly as NE rabbits but exhibited higher total and high-density lipoprotein (HDL) cholesterol levels in adulthood. They also presented a significantly elevated serum testosterone concentrations, while estradiol, progesterone, AMH and DHEA levels remained unaffected. The measurement of gonadotropins, androstenedione, pregnenolone and estrone levels yielded values below the limit of quantification. Among the 7 steroidogenic enzymes tested, an isolated higher expression of Cyp19a1 was measured in F rabbits ovaries. Those ovaries presented a significantly greater density/number of antral and atretic follicles and larger antral follicles without any changes in cellular proliferation or DNA fragmentation. No difference was found regarding the count of other follicle stages notably the primordial stage, the corpora lutea or AMH serum levels., Conclusion: Folliculogenesis and steroidogenesis seem to be subtly altered by exposure to a human-like mixture of environmental toxicants. The antral follicle growth appears promoted by the mixture of chemicals both in their number and size, potentially explaining the increase in atretic antral follicles. Reassuringly, the ovarian reserve estimated through primordial follicles number/density and AMH is spared from any alteration. The consequences of these changes on fertility and progeny health have yet to be investigated., (© 2024. The Author(s).)

Hypercholesterolemia is a major risk factor for coronary artery diseases and cardiac ischemic events. Cholesterol per se could also have negative effects on the myocardium, independently from hypercholesterolemia. Previously, we reported that myocardial ischemia-reperfusion induces a deleterious build-up of mitochondrial cholesterol and oxysterols, which is potentiated by hypercholesterolemia and prevented by translocator protein (TSPO) ligands. Here, we studied the mechanism by which sterols accumulate in cardiac mitochondria and promote mitochondrial dysfunction. We performed myocardial ischemia-reperfusion in rats to evaluate mitochondrial function, TSPO, and steroidogenic acute regulatory protein (STAR) levels and the related mitochondrial concentrations of sterols. Rats were treated with the cholesterol synthesis inhibitor pravastatin or the TSPO ligand 4'-chlorodiazepam. We used Tspo deleted rats, which were phenotypically characterized. Inhibition of cholesterol synthesis reduced mitochondrial sterol accumulation and protected mitochondria during myocardial ischemia-reperfusion. We found that cardiac mitochondrial sterol accumulation is the consequence of enhanced influx of cholesterol and not of the inhibition of its mitochondrial metabolism during ischemia-reperfusion. Mitochondrial cholesterol accumulation at reperfusion was related to an increase in mitochondrial STAR but not to changes in TSPO levels. 4'-Chlorodiazepam inhibited this mechanism and prevented mitochondrial sterol accumulation and mitochondrial ischemia-reperfusion injury, underlying the close cooperation between STAR and TSPO. Conversely, Tspo deletion, which did not alter cardiac phenotype, abolished the effects of 4'-chlorodiazepam. This study reveals a novel mitochondrial interaction between TSPO and STAR to promote cholesterol and deleterious sterol mitochondrial accumulation during myocardial ischemia-reperfusion. This interaction regulates mitochondrial homeostasis and plays a key role during mitochondrial injury., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany.)

Estradiol (E2) is a major hormone-controlling folliculogenesis whose dysfunction may participate in polycystic ovary syndrome (PCOS) infertility. To determine whether both the concentration and action of E2 could be impaired in non-hyperandrogenic overweight PCOS women, we isolated granulosa cells (GCs) and follicular fluid (FF) from follicles of women undergoing ovarian stimulation (27 with PCOS, and 54 without PCOS). An analysis of the transcript abundance of 16 genes in GCs showed that androgen and progesterone receptor expressions were significantly increased in GCs of PCOS (by 2.7-fold and 1.5-fold, respectively), while those of the steroidogenic enzymes CYP11A1 and HSD3B2 were down-regulated (by 56% and 38%, respectively). Remarkably, treatment of GC cultures with E2 revealed its ineffectiveness in regulating the expression of several key endocrine genes (e.g., GREB1 or BCL2 ) in PCOS. Additionally, a comparison of the steroid concentrations (measured by GC/MS) in GCs with those in FF of matched follicles demonstrated that the significant decline in the E2 concentration (by 23%) in PCOS FF was not the result of the E2 biosynthesis reduction. Overall, our study provides novel hallmarks of PCOS by highlighting the ineffective E2 signaling in GCs as well as the dysregulation in the expression of genes involved in follicular growth, which may contribute to aberrant folliculogenesis in non-hyperandrogenic women with PCOS.

In females, androgens contribute to ovarian diseases such as polycystic ovarian syndrome (PCOS), but their action is also crucial for ovarian physiology, i.e., follicular growth and estradiol (E2) synthesis during reproductive life, in interaction with the gonadotropins LH and FSH. However, it is unclear whether androgens already play a role in the ovary at mini-puberty, a phase of postnatal development with active follicular growth and high E2 levels. Therefore, we analyzed the potential actions of androgens on the ovary and their possible interaction with gonadotropins during this period in mice. We used molecular-based studies and pharmacological approaches in vivo and on cultured ovaries. We found that mini-pubertal ovaries produce significant amounts of testosterone and display androgen receptor (AR) expression in growing follicles, both under the control of LH. By blocking AR signaling either in vivo or in ovarian cultures, we found that this pathway may participate in the regulation of prepubertal E2 synthesis and follicular growth, possibly by regulating the expression of a number of key intra-ovarian regulators, including FSH receptor ( Fshr ), the aromatase enzyme converting androgens into estrogens ( Cyp19a1) and the cell cycle inhibitor p27KIP1 ( Cdkn1b) . We further showed that AR may stimulate FSH-mediated regulation of Cyp19a1 through its action on Fshr mRNA abundance. Overall, this work supports the idea that AR signaling is already activated in mini-pubertal ovaries to regulate E2 synthesis and follicular growth, at the interplay with LH and FSH signaling. Its early action may, thus, contribute to the implementation of early ovarian function with possible impacts on reproductive function., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Devillers, François, Chester, Corre, Cluzet, Giton, Cohen-Tannoudji and Guigon.)

Introduction: Polychlorobiphenyls (PCBs), organochlorine pesticides (OCPs), and per- and polyfluoroalkyl substances (PFASs) are persistent organic pollutants (POPs) having numerous toxicological properties, including thyroid endocrine disruption. Our aim was to assess the impact of POPs on thyroid hormones among 12-year-old children, while taking puberty into consideration., Methods: Exposure to 7 PCBs, 4 OCPs, and 6 PFASs (in µg/L), and free tri-iodothyronine (fT3, pg/mL), free thyroxine (fT4, ng/dL), and thyroid-stimulating hormones (TSH, mIU/L) were assessed through blood-serum measurements at age 12 years in 249 boys and 227 girls of the PELAGIE mother-child cohort (France). Pubertal status was clinically rated using the Tanner stages. For each POP, associations were estimated using linear regression, adjusted for potential confounders., Results: Among boys, hexachlorobenzene and perfluorodecanoic acid were associated with decreased fT3 (log-scale; β [95% confidence interval] = -0.07 [-0.12,-0.02] and β = -0.03 [-0.06,-0.00], respectively). Intermediate levels of perfluorohexanesulfonic acid (PFHxS) and PCB180 were associated, respectively, with increased and decreased fT4. After stratification on pubertal status, PCBs and OCPs were associated with decreased TSH only in the more advanced Tanner stages (3-5) and with decreased fT3 among early Tanner stages (1-2). Among girls, PFHxS was associated with decreased TSH (log-scale; β = -0.15 [-0.29,-0.00]), and perfluorooctanoic acid was associated with decreased fT3 (β2nd&#95;tercile = -0.06 [-0.10,-0.03] and β3rd&#95;tercile = -0.04 [-0.08,-0.00], versus. 1st tercile)., Discussion: This cross-sectional study highlights associations between some POPs and thyroid function disruption, which appears consistent with the literature. Considering that the associations were sex-specific and moderated by pubertal status in boys, complex endocrine interactions are likely involved., (© 2022 The Author(s). Published by S. Karger AG, Basel.)

Estrogens are steroid hormones produced by the aromatization of androgens by the aromatase enzyme, encoded by the CYP19A1 gene. Although generally referred to as "female sex hormones", estrogen is also produced in the adult testes of many mammals, including humans. To better understand the function of estrogens in the male, we used the rabbit model which is an important biomedical model. First, the expression of CYP19A1 transcripts was localized mainly in meiotic germ cells. Thus, testicular estrogen appears to be produced inside the seminiferous tubules. Next, the cells expressing ESR1 and ESR2 were identified, showing that estrogens could exert their function on post-meiotic germ cells in the tubules and play a role during sperm maturation, since ESR1 and ESR2 were detected in the cauda epididymis. Then, CRISPR/Cas9 CYP19A1 -/- genetically modified rabbits were analyzed. CYP19A1 -/- males showed decreased fertility with lower sperm count associated with hypo-spermatogenesis and lower spermatid number. Germ/sperm cell DNA methylation was unchanged, while sperm parameters were affected as CYP19A1 -/- males exhibited reduced sperm motility associated with increased flagellar defects. In conclusion, testicular estrogens could be involved in the spermatocyte-spermatid transition in the testis, and in the acquisition of sperm motility in the epididymis.

The binding of 17β-oestradiol to oestrogen receptor alpha (ERα) plays a crucial role in the control of reproduction, acting through both nuclear and membrane-initiated signalling. To study the physiological role of membrane ERα in the reproductive system, we used the C451A-ERα mouse model with selective loss of function of membrane ERα. Despite C451A-ERα mice being described as sterile, daily weighing and ultrasound imaging revealed that homozygous females do become pregnant, allowing the investigation of the role of ERα during pregnancy for the first time. All neonatal deaths of the mutant offspring mice resulted from delayed parturition associated with failure in pre-term progesterone withdrawal. Moreover, pregnant C451A-ERα females exhibited partial intrauterine embryo arrest at about E9.5. The observed embryonic lethality resulted from altered expansion of Tpbpa-positive spiral artery-associated trophoblast giant cells into the utero-placental unit, which is associated with an imbalance in expression of angiogenic factors. Together, these processes control the trophoblast-mediated spiral arterial remodelling. Hence, loss of membrane ERα within maternal tissues clearly alters the activity of invasive trophoblast cells during placentogenesis. This previously unreported function of membrane ERα could open new avenues towards a better understanding of human pregnancy-associated pathologies., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2022. Published by The Company of Biologists Ltd.)

Female infertility has a multifactorial origin, and exposure to contaminants, including pesticides, with endocrine-disrupting properties is considered to be involved in this reproductive disorder, especially when it occurs during early life. Pesticides are present in various facets of the environment, and consumers are exposed to a combination of multiple pesticide residues through food intake. The consequences of such exposure with respect to female fertility are not well known. Therefore, we aimed to assess the impact of pre- and postnatal dietary exposure to a pesticide mixture on folliculogenesis, a crucial process in female reproduction. Mice were exposed to the acceptable daily intake levels of six pesticides in a mixture (boscalid, captan, chlorpyrifos, thiacloprid, thiophanate and ziram) from foetal development until 8 weeks old. Female offspring presented with decreased body weight at weaning, which was maintained at 8 weeks old. This was accompanied by an abnormal ovarian ultrastructure, a drastic decrease in the number of corpora lutea and progesterone levels and an increase in ovary cell proliferation. In conclusion, this study shows that this pesticide mixture that can be commonly found in fruits in Europe, causing endocrine disruption in female mice with pre- and postnatal exposure by disturbing folliculogenesis, mainly in the luteinisation process.

Context: Acetaminophen (APAP, paracetamol) is widely used by pregnant women. Although long considered safe, growing evidence indicates that APAP is an endocrine disruptor since in utero exposure may be associated with a higher risk of male genital tract abnormalities. In rodents, fetal exposure has long-term effects on the reproductive function of female offspring. Human studies have also suggested harmful APAP exposure effects., Objective: Given that disruption of fetal ovarian development may impact women's reproductive health, we investigated the effects of APAP on fetal human ovaries in culture., Design and Setting: Human ovarian fragments from 284 fetuses aged 7 to 12 developmental weeks (DW) were cultivated ex vivo for 7 days in the presence of human-relevant concentrations of APAP (10-8 to 10-3 M) or vehicle control., Main Outcome Measures: Outcomes included examination of postculture tissue morphology, cell viability, apoptosis, and quantification of hormones, APAP, and APAP metabolites in conditioned culture media., Results: APAP reduced the total cell number specifically in 10- to 12-DW ovaries, induced cell death, and decreased KI67-positive cell density independently of fetal age. APAP targeted subpopulations of germ cells and disrupted human fetal ovarian steroidogenesis, without affecting prostaglandin or inhibin B production. Human fetal ovaries were able to metabolize APAP., Conclusions: Our data indicate that APAP can impact first trimester human fetal ovarian development, especially during a 10- to 12-DW window of heightened sensitivity. Overall, APAP behaves as an endocrine disruptor in the fetal human ovary., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Endocrine Society.)

Overnutrition is associated with the activation of inflammatory pathways in metabolically linked organs and an early hypothalamic inflammation is now known to disrupt the central control of metabolic function. Because we demonstrated that fatty acids (FA) target the pituitary and affect gonadotropin synthesis, we asked whether overnutrition induces pituitary inflammation that may contribute to obesity-associated disorders in the control of reproduction. We analyzed pituitary inflammation and hypothalamic-pituitary-testicular axis in male rats fed a short- (4 weeks) or long-term (20 weeks) high-fat diet. The effect of diet enrichment with the ω3 polyunsaturated FA, DHA, was also analyzed. After only 4 weeks and before weight gain of rats, high-fat diet caused a significant decrease in pituitary gonadotropin and hypothalamic GnRH transcript levels despite unchanged testosterone and inhibin B levels. Contrasting with the hypothalamus, there was no concomitant increases in gene expression of pituitary inflammatory mediators and even a reduction of prototypical cytokines such as interleukin-1β and TNF-α. No inflammation was still detected in the pituitary after 20 weeks although gonadotropin transcripts and circulating levels were still altered. Gonadotropins were the only pituitary hormones remaining affected at this stage of the regimen, underlying a differential susceptibility of pituitary lineages to metabolic disorders. DHA enrichment of the diet did not prevent alterations of gonadotrope activity due to either a long- or a short-term high-fat diet although it blocked early hypothalamic inflammation and attenuated several metabolic effects. Taken together, our findings suggest that high-fat diet-induced defects in gonadotrope activity in male rats occurred despite a lack of pituitary inflammation., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Garrel, Rouch, L’Hôte, Tazi, Kassis, Giton, Dairou, Dournaud, Gressens, Magnan, Cruciani-Guglielmacci and Cohen-Tannoudji.)

Granulosa cell tumor (GCT) is a form of ovarian tumor characterized by its tendency to recur years after surgical ablation. Little is known about the mechanisms involved in GCT development and progression. GCTs can produce estradiol (E2), but whether this hormone could play a role in this cancer through its nuclear receptors, i.e. ERα and ERβ, remains unknown. Here, we addressed this issue by cell-based and molecular studies on human GCTs and GCT cell lines. Importantly, we observed that E2 significantly increased the growth of GCT cells by promoting cell survival. The use of selective agonists of each type of receptor, together with Esr1 (ERα) or Esr2 (ERβ)-deleted GCT cells, revealed that E2 mediated its effects through ERα-dependent genomic mechanisms and ERβ/ERα-dependent extra-nuclear mechanisms. Notably, the expression of Greb1, a prototypical ER target gene, was dose-dependently upregulated by E2 specifically through ERα in GCT cells. Accordingly, using GCTs from patients, we found that GREB1 mRNA abundance was positively correlated to intra-tumoral E2 concentrations. Tissue microarray analyses showed that there were various combinations of ER expression in primary and recurrent GCTs, and that ERα expression persisted only in combination with ERβ in ~40% of recurrent tumors. Altogether, this study demonstrates that E2 can promote the progression of GCTs, with a clear dependence on ERα. In addition to demonstrating that GCTs can be classified as a hormone-related cancer, our results also highlight that the nature of ER forms present in recurrent GCTs could underlie the variable efficiency of endocrine therapies. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd., (© 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.)

AROMATASE is encoded by the CYP19A1 gene and is the cytochrome enzyme responsible for estrogen synthesis in vertebrates. In most mammals, a peak of CYP19A1 gene expression occurs in the fetal XX gonad when sexual differentiation is initiated. To elucidate the role of this peak, we produced 3 lines of TALEN genetically edited CYP19A1 knockout (KO) rabbits that were devoid of any estradiol production. All the KO XX rabbits developed as females with aberrantly small ovaries in adulthood, an almost empty reserve of primordial follicles, and very few large antrum follicles. Ovulation never occurred. Our histological, immunohistological, and transcriptomic analyses showed that the estradiol surge in the XX fetal rabbit gonad is not essential to its determination as an ovary, or for meiosis. However, it is mandatory for the high proliferation and differentiation of both somatic and germ cells, and consequently for establishment of the ovarian reserve., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Endocrine Society.)

Background: Chlordecone is an endocrine-disrupting chemical with well recognized estrogenic and progestagenic properties. This organochlorine insecticide was extensively used in the French West Indies from 1973 to 1993 to control the banana root borer. Due to its poor degradation in the environment, permanently polluted soil is responsible for the current contamination of the food chain and human beings. We aimed to examine the relationship of in utero exposure to chlordecone and thyroid (thyroid stimulating hormone [TSH], free tri-iodothyronine [FT3], free thyroxine [FT4]), metabolic (insulin growth-factor 1, leptin, adiponectin), and sex-steroid (dehydroepiandrosterone [DHEA], total testosterone [TT], dihydrotestosterone [DHT], estradiol [E2]) hormone levels in children at the age of seven years who participated in TIMOUN, an ongoing birth cohort in Guadeloupe., Methods: Chlordecone concentrations were measured in cord-blood at delivery. Thyroid, metabolic, and sex-steroid hormone levels were determined in the blood of children at seven years of age. Associations between in utero chlordecone exposure and hormone levels at seven years of age were assessed by multiple linear or logistic regression, controlling for confounding factors., Results: Among the study population (210 boys and 228 girls), chlordecone and hormone measurements were available for 124 boys and 161 girls. We found the third quartile of in utero chlordecone exposure relative to the lowest quartile to be associated with elevated TSH levels in girls and elevated DHEA, TT, and DHT levels in both sexes. Complementary non-linear analysis (spline regression) confirmed a significant non-linear trend for TSH in girls and DHEA and DHT in boys., Conclusion: In utero chlordecone exposure was associated with elevated levels of selected thyroid (TSH) and sex-steroid (DHEA, TT, and DHT) hormones at seven years in a non-monotonic dose response (inverted U) relationship. The implications for future health and reproductive function in puberty and adulthood should be determined., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Ayhan, Rouget, Giton, Costet, Michineau, Monfort, Thomé, Kadhel, Cordier, Oliva and Multigner.)

In female mammals, puberty and fertility are regulated by the synthesis of estradiol (E2) by the ovaries at the infantile stage and at the approach of puberty, a process which may be affected by endocrine disrupting chemicals (EDC)s acting through the Aryl hydrocarbon receptor (AhR). However, there is no information on AhR-mediated regulation of ovarian estrogenic activity during these developmental periods. Here, we assessed in mouse models, the intrinsic and exogenous ligand-induced AhR action on E2 synthesis at the infantile stage (14 days postnatal (dpn)) and at the approach of puberty (28 dpn). Intrinsic AhR pathway became activated in the ovary at the approach of puberty, as suggested by the decreased intra-ovarian expression in prototypical and steroidogenesis-related AhR targets and E2 contents in Ahr knockout (Ahr -/- ) mice versus Ahr +/+ mice exclusively at 28 dpn. Accordingly, AhR nuclear localization in granulosa cells, reflecting its activity in cells responsible for E2 synthesis, was much lower at 14 dpn than at 28 dpn in C57BL/6 mice. However, AhR signaling could be activated by exogenous ligands at both ages, as revealed by FICZ- and TCDD-induced Ahrr and Cyp1a1 expression in C57BL/6 mice. Nevertheless, TCDD impacted ovarian estrogenic activity only at 28 dpn. This age-related AhR action may be ligand-dependent, since FICZ had no effect on E2 synthesis at 28 dpn. In conclusion, AhR would not regulate ovarian estrogenic activity before the approach of puberty. Its activation by EDCs may be more detrimental to reproductive health at this stage than during infancy., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

17β-Estradiol (E2) action can be mediated by the full-length estrogen receptor alpha (ERα66), and also by the AF1 domain-deficient ERα (ERα46) isoform, but their respective sensitivity to E2 is essentially unknown. We first performed a dose response study using subcutaneous home-made pellets mimicking either metestrus, proestrus or a pharmacological doses of E2, which resulted in plasma concentrations around 3, 30 and 600 pM, respectively. Analysis of the uterus, vagina and bone after chronic exposure to E2 demonstrated dose-dependent effects, with a maximal response reached at the proestrus-dose in wild type mice expressing mainly ERα66. In contrast, in transgenic mice harbouring only an ERα deleted in AF1, these effects of E2 were either strongly shifted rightward (10-100-fold) and/or attenuated, depending on the tissue studied. Finally, experiments in different cell lines transfected with ERα66 or ERα46 also delineated varying profiles of ERα AF1 sensitivity to E2. Altogether, this work emphasizes the importance of dose in the tissue-specific actions of E2 and demonstrates the key sensitizing role of AF1 in ERα activity., Competing Interests: Declaration of competing interest The authors have nothing to disclose., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Ovarian granulosa cell tumors (GCTs) are indolent tumors of the ovary affecting women at all ages and potentially displaying late recurrence. Even if there is still little information regarding the mechanisms involved in GCT development and progression, FOXL2 would be a major tumor suppressor gene in granulosa cells. We analyzed the mechanisms underlying GCT initiation and progression by using mice with targeted expression of SV40 large T-antigen in granulosa cells (AT mouse), which develop GCTs. Consistent with patients, AT mice with developing GCTs displayed increased levels in circulating anti-Müllerian hormone (AMH), estradiol and androgens, as well as decreased FOXL2 protein abundance. Very few mice developed metastases (1 out of 30). In situ analyses revealed that GCT initiation resulted from both increased granulosa cell survival and proliferation in large antral follicles. Tumorigenesis was associated with the combined inactivation of p53 and Rb pathways, as shown by the impaired expression of respective downstream targets regulating cell apoptosis and proliferation, i.e., Bax, Bak, Gadd45a, Ccna2, Ccne1, E2f1, and Orc1. Importantly, the expression of FOXL2 was still present in newly developed GCTs and its downregulation only started during GCT growth. Collectively, our experiments provide evidence that disrupted p53/Rb signaling can drive tumor initiation and growth. This model challenges the current paradigm that impaired FOXL2 signaling is a major switch of granulosa cell tumorigenesis, albeit possibly contributing to tumor growth.

Anti-Müllerian hormone (AMH) regulates ovarian function in cyclic females, notably by preventing premature follicle-stimulating hormone (FSH)-mediated follicular growth and steroidogenesis. Its expression in growing follicles is controlled by FSH and by estradiol (E2). In infantile females, there is a transient increase in the activity of the gonadotrope axis, as reflected by elevated levels of both gonadotropins and E2. We previously demonstrated in mice that elevated FSH concentrations are necessary to induce E2 production by preantral/early antral follicles through the stimulation of aromatase expression without supporting their growth. However, whether this action of FSH could involve AMH is unknown. Here, we show that Amh mRNA and protein abundance and serum AMH levels are elevated in infantile mouse females, compared with those in adults. By experimentally manipulating FSH and E2 levels in infantile mice, we demonstrate that high FSH concentrations lower Amh expression specifically in preantral/early antral follicles, whereas E2 has no effect. Importantly, treatment of infantile ovaries in organotypic cultures with AMH decreases FSH-mediated expression of Cyp19a1 aromatase, but it does not alter the expression of cyclin D2-mediating granulosa cell proliferation. Overall, our data indicate that the infantile elevation in FSH levels suppresses Amh expression in preantral/early antral follicles, thereby favoring Cyp19a1 aromatase expression and E2 production. Together with recent discoveries that AMH can act on both the hypothalamus and the pituitary to increase gonadotropin levels, this work suggests that AMH is a critical regulator of the gonadotrope axis during the infantile period, thereby contributing to adult reproductive function programming.

Background: Currently, there is no consensus regarding the expected concentration levels of intra-prostatic sex steroids in patients with Prostate Cancer (PCa). Our objective was to assess the concentration levels of sex steroids in prostatic tissue and serum, in two cohorts of patients with localized PCa or benign prostatic hyperplasia (BPH)., Methods: Between September 2014 and January 2017, men selected for radical cystectomy (for bladder cancer) or open prostatectomy (for BPH), and men selected for radical prostatectomy for localized PCa were included. Blood samples were collected at baseline before surgery, and steroid concentrations were assessed following the recommendations of the Endocrine Society. Intra-prostatic samples were collected from fresh surgical samples, and assessed by gas chromatography and mass spectrometry (GC/MS). Permanova analysis was performed. Analyses were adjusted for age, prostate weight, and prostate-specific antigen (PSA) level., Results: A total of 73 patients (41 patients with PCa and 32 patients with BPH) were included in this study. Patients with PCa were younger, and had smaller prostate volumes with higher levels of PSA. The levels of Total Testosterone (TT), Di-Hydro-Testosterone (DHT), and Estradiol (E2) in the serum were not significantly different between PCa and BPH. In PCa tissue, TT concentrations were significantly lower (0.11 ng/g vs 0.47 ng/g, P = 0.0002), however its derivative E2 had significantly higher concentrations (31.0 ng/g vs 22.3 ng/g, P = 0.01). DHT tissue concentrations were not significantly different between the two groups (5.55 ng/g vs 5.42 ng/g, P = 0.70). Intra-prostatic TT concentrations were significantly lower in the peripheral zone than in the central zone for the CaP group (0.07 ng/g vs 0.15 ng/g, P = 0.004)., Conclusions: Patients with PCa had lower intra-prostatic TT and higher E2 concentrations levels compared to the patients with BPH. PCa seem to consume more TT and produce more E2, especially in the peripheral zone., (© 2018 Wiley Periodicals, Inc.)

Failure rates after first-line treatment of localized prostate cancer (PCa) treatment remain high. Improvements to patient selection and identification of at-risk patients are central to reducing mortality. We aimed to determine if cancer aggressiveness correlates with androgen levels in patients undergoing radical prostatectomy for localized PCa. We performed a prospective, multicenter cohort study between June 2013 and June 2016, involving men with localized PCa scheduled to undergo radical prostatectomy. Clinical and hormonal patient data (testosterone deficiency, defined by total testosterone (TT) levels < 300 ng/dL and/or bioavailable testosterone (BT) levels < 80 ng/dL) were prospectively collected, along with pathological assessment of preoperative biopsy and subsequent radical prostatectomy specimens, using predominant Gleason pattern (prdGP) 3/4 grading. Of 1343 patients analyzed, 912 (68%) had prdGP3 PCa and 431 (32%) had high-grade (prdGP4, i.e., ISUP ≥ 3) disease on prostatectomy specimens. Only moderate concordance in prdGP scores between prostate biopsies and prostatectomy specimens was found. Compared with patients with prdGP3 tumors (i.e., ISUP ≤ 2), significantly more patients with prdGP4 cancers had demonstrable hypogonadism, characterized either by BT levels (17.4% vs. 10.7%, p < 0.001) or TT levels (14.2% vs. 9.7%, p = 0.020). BT levels were also lower in patients with prdGP4 tumors compared to those with prdGP3 disease. Testosterone deficiency (defined by TT and/or BT levels) was independently associated with higher PCa aggressiveness. BT is a predictive factor for prdGP4 disease, and evaluating both TT and BT to define hypogonadism is valuable in preoperative assessment of PCa (AndroCan Trial: NCT02235142).

Background: Glycol ethers (GEs) are oxygenated solvents widely found in occupational and consumer water-based products. Some of them are well-known reproductive and developmental toxicants., Objectives: To study the variations in circulating sex steroid hormones, measured in cord blood, according to biomarkers of prenatal GE exposure., Methods: The study population comes from the PELAGIE mother-child cohort, which enrolled pregnant women from Brittany (France, 2002-2006). Maternal urine samples were collected from a random subcohort (n = 338) before 19 weeks' gestation, from which we measured 8 alkoxycarboxylic metabolites of GEs. We subsequently measured 13 sex steroid hormones and sex hormone-binding globulin (SHBG) in cord blood samples. Linear regressions adjusted for potential confounders were used, and nonlinear dose-response associations were investigated., Results: The detection rates of GE metabolites ranged from 4% to 98%; only the 5 most detected (>20%) metabolites were investigated further. Phenoxyacetic acid (detection rate > 95%) was associated with lower levels of SHBG and various steroids (17-alpha-hydroxy-Pregnenolone, delta-5-androstenediol, and dehydroepiandrosterone) among boys and higher SHBG and 16-alpha-hydroxy-dehydroepiandrosterone levels among girls. The two other highly detected metabolites, methoxyetoxyacetic acid and butoxyacetic acid, were associated with variations in estradiol. Butoxyacetic acid was associated with higher delta-5-androstenediol levels while detectable levels of methoxyacetic acid were associated with lower levels of this hormone., Conclusion: Our study suggests that prenatal exposure to GE may affect endocrine response patterns, estimated by determining blood levels of sex steroid hormones in newborns. These results raise questions about the potential role of these changes in the pathways between prenatal GE exposure and previously reported adverse developmental outcomes, including impaired neurocognitive performance., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Background: The specific involvement of the sex steroids in the growth of the prostatic tissue remains unclear. Sex steroid concentrations in plasma and in fresh surgical samples of benign central prostate were correlated to prostate volume., Methods: Monocentric prospective study performed between September 2014 and January 2017. Age, obesity parameters, and both serum and intraprostatic concentrations of sex steroids were collected complying with the latest Endocrine Society guidelines and the steroids assessed by GC/MS. Statistical calculations were adjusted for age and body mass index (BMI)., Results: Thirty-two patients, equally divided between normal- and high-volume prostate groups, were included in the analysis. High-volume prostate patients were older, heavier and had higher BMI. Comparison adjusted for age and BMI showed higher DHT concentrations in high-volume prostate. Both normal- and high-volume prostate tissues concentrate sex steroids in a similar way. Comparison of enzymatic activity surrogate marker ratios within tissue highlighted similar TT/E1 and TT/E2 ratios, and higher DHT/E1 ratio and lower DHT/PSA ratio in the high-volume prostates., Conclusions: STERPROSER trial provides evidence for higher DHT concentration in highvolume prostates, that could reflect either higher 5-alpha reductase expression or lower expression of downstream metabolizing enzymes such as 3a-hydoxysteroid dehydrogenase., (© 2017 Wiley Periodicals, Inc.)

Context: Anti-Müllerian hormone (AMH) and AMH type II receptor (AMHR2) are overexpressed in granulosa cells (GCs) from women with polycystic ovary syndrome (PCOS), the most common cause of female infertility., Objective: The aim of the study was to compare the regulation of the AMH/AMHR2 system by 5α-dihydrotestosterone (5α-DHT) and estradiol (E2) in GCs from control subjects and women with PCOS., Design, Setting, Patients: Experiments were performed on follicular fluids (FF) and GCs from women undergoing in vitro fertilization., Main Outcome Measures: FF steroid levels were measured by mass spectrometry, and messenger RNA (mRNA) accumulation was quantified by reverse transcription real-time polymerase chain reaction., Results: Total testosterone (T), free T, and 5α-DHT FF levels were significantly higher (P < 0.001) in women with PCOS than in controls. However, E2 and sex hormone-binding globulin concentrations were comparable between the two groups. In GCs from control women, the AMH and AMHR2 expression were not affected by 5α-DHT treatment, whereas AMH mRNA levels were upregulated by 5α-DHT in GCs from patients with PCOS (2.3-fold, P < 0.01) overexpressing the androgen receptor (1.4-fold, P < 0.05). E2 downregulated the AMH and AMHR2 expression in GCs from control women (1.4-fold, P < 0.001 and 1.8-fold, P < 0.01, respectively) but had no effect on these genes in GCs from women with PCOS. This differential effect of E2 was associated with a higher estrogen receptor 1 expression in GCs from women with PCOS (1.9-fold, P < 0.05)., Conclusions: In GCs from women with PCOS, the regulation of AMH and AMHR2 expression is altered in a way that promotes the overexpression of the AMH/AMHR2 system, and could contribute to the follicular arrest observed in these patients., (Copyright © 2017 Endocrine Society)

The widely-used herbicide atrazine (ATZ) is detected in ground and surface water in many countries. Several studies in animals have demonstrated that ATZ has endocrine-disrupting effects on male and female reproduction in many vertebrate species. In this study, we investigated the effects of ATZ exposure on meiosis, a key step in gametogenesis in mammals. The treatment was initiated before oocyte entry into meiosis, which occurs during the embryonic period in females. We found that embryonic exposure to ATZ increases the level of 8-oxo-guanine in the nucleus of meiotic cells, reflecting oxidative stress and affecting meiotic double-strand break repair, chromosome synapsis and crossover numbers. Finally, embryonic exposure to ATZ reduces the number of primordial follicles and increases the incidence of multi-oocyte follicles in adult mice. Our data demonstrate that embryonic exposure to ATZ disrupts prophase I of meiosis and affects normal follicle formation in female mice.

In cyclic females, FSH stimulates ovarian estradiol (E2) production and follicular growth up to the terminal stage. A transient elevation in circulating FSH and E2 levels occurs shortly after birth. But what could be the action of FSH on the ovary during this period, and in particular how it stimulates ovarian steroidogenesis without supporting terminal follicular maturation is intriguing. By experimentally manipulating FSH levels, we demonstrate in mice that the mid-infantile elevation in FSH is mandatory for E2 production by the immature ovary, but that it does not stimulate follicle growth. Importantly, FSH increases aromatase expression to stimulate E2 synthesis, however it becomes unable to induce cyclin D2, a major driver of granulosa cell proliferation. Besides, although FSH prematurely induces luteinizing hormone (LH) receptor expression in granulosa cells, LH pathway is not functional in these cells to induce their terminal differentiation. In line with these results, supplying infantile mice with a superovulation regimen exacerbates E2 production, but it does not stimulate the growth of follicles and it does not induce ovulation. Overall, our findings unveil a regulation whereby high postnatal FSH concentrations ensure the supply of E2 required for programming adult reproductive function without inducing follicular maturation before puberty.