α-Lipoic acid eliminates dioxin-induced offspring sexual immaturity by improving abnormalities in folic acid metabolism.

Maternal exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) causes developmental and reproductive disorders in pups due to the attenuated luteinizing hormone (LH) production during the perinatal stage; however, the administration of α-lipoic acid (LA) to TCDD-exposed pregnant rats reversed the attenuated LH production. Therefore, reproductive disorders in pups are expected to be ameliorated with LA supplementation. To address this issue, pregnant rats orally received low dose TCDD at gestational day 15 (GD15) and proceeded to parturition. The control received a corn oil vehicle. To examine the preventive effects of LA, supplementation with LA was provided until postnatal day 21. In this study, we demonstrated that maternal administration of LA restored the sexually dimorphic behavior of male and female offspring. TCDD-induced LA insufficiency is likely a direct cause of TCDD reproductive toxicity. In the analysis to clarify the mechanism of the decrease in LA, we found evidence suggesting that TCDD inhibits the synthesis and increases the utilization of S-adenosylmethionine (SAM), a cofactor for LA synthesis, resulting in a decrease in the SAM level. Furthermore, folate metabolism, which is involved in SAM synthesis, is disrupted by TCDD, which may adversely affect infant growth. Maternal supplementation of LA restored SAM to its original level in the fetal hypothalamus; in turn, SAM ameliorated abnormal folate consumption and suppressed aryl hydrocarbon receptor activation induced by TCDD. The study demonstrates that the application of LA could prevent and recover next-generation dioxin reproductive toxicity, which provides the potential to establish effective protective measures against dioxin toxicity.

Postnatal developmental expression of apelin receptor proteins and its role in juvenile mice testis.

The expression of apelin system has been shown in the adult testis of rat and mice. It has also been emphasized that regulation of testicular activity in early stages is important to sustain normal testicular activity in adulthood. Since the expression of apelin receptor (APJ) has been shown in the adult testis, moreover, developmental expression of APJ and its role has not been explored yet. Thus, we have examined the testicular expression of APJ during postnatal stages with special reference to proliferation, apoptosis and hormone secretion in early postnatal stage. Postnatal analysis showed that circulating apelin was lowest at PND1 and maximum at PND42. Among testosterone, estrogen and androstenedione, only circulating testosterone showed a gradual increase from PND1 to PND42. Testicular expression of APJ was also developmenatly regulated from PND1 to PND42, revealing a positive correlation with circulating apelin, testosterone, and androstenedione. Immunohistochemical study showed that APJ was mainly confined to Leydig cells of early postnatal stages, whereas, seminiferous tubules at PND42 showed immunostaining in the round spermatids. APJ inhibition from PND14-PND20 by ML221 suppressed the testicular proliferation, increased apoptosis and increased estrogen secretion. However, expression of AR was down-regulated by ML221 treatment. Furthermore, ML221 decreased the abundance of p-Akt. In vitro study also showed that APJ antagonist, ML221 decreased AR expression. These results suggests that apelin signaling during early developmental stages might be required to stimulate the germ cell proliferation, and inhibition of apoptosis. Both in vivo and in vitro study have shown that expression of AR was regulated by apelin signaling. Since the first wave spermatogenesis involves proliferation and apoptosis, therefore, further study would be required to unravel the exact mechanism of apelin mediated regulation of testicular activity during early postnatal stages. In conclusion, the present results are an indicative of apelin mediated signaling during early postnatal stage for regulation of germ cell proliferation, apoptosis and AR expression.

Fetal Programming Effects of a Mild Food Restriction During Pregnancy in Mice: How Does It Compare to Intragestational Ghrelin Administration?

To explore in mice if a 15% food restriction protocol during pregnancy programs the offspring postnatal development, with emphasis on reproductive function, and to assess if ghrelin (Ghrl) administration to mouse dams exerts effects that mimic those obtained under mild caloric restriction. Mice were 15% food-restricted, injected with 4 nmol/animal/day of Ghrl, or injected with the vehicle (control) thorough pregnancy. After birth, the pups did not receive further treatment. Pups born from food-restricted dams (FR pups) were lighter than Ghrl pups at birth, but reached normal weight at adulthood. Ghrl pups were heavier at birth and gained more weight than control pups (C pups). This effect was not associated with plasma IGF-1. FR pups showed a delay in pinna detachment and eye opening, while an advance was observed in Ghrl pups. FR pups showed also impairment in the surface-righting reflex. In both female FR and Ghrl pups, there was an advance in vaginal opening and, in adulthood, FR pups showed a significant decrease in their own litter size and plasma progesterone, and an increase in embryo loss. A delay in testicular descent was evident in male Ghrl pups. Changes in puberty onset were not associated with differences in the expression of Kiss1 in hypothalamic nuclei. Finally, in adulthood, FR pups showed a significant decrease in sperm quality. In conclusion, a mild food restriction thorough gestation exerted programming effects on the offspring, affecting also their reproductive function in adulthood. These effects were not similar to those of intragestational Ghrl administration.

Prenatal dexamethasone exposure and developmental programming of the ovary of the offspring: a structural study in the rat.

Overexposure to glucocorticoids during fetal development alters fetal organ growth and maturation patterns, which can result in adverse programming outcomes in adulthood. The aim of this study was to determine whether exposure to dexamethasone (Dx) during the fetal period programmed ovary development and function in infant (16-day-old) and peripubertal (38-day-old) female offspring. Pregnant Wistar rats were separated into control and Dx-treated (0.5mg kg-1) groups and were injected with Dx or an equivalent volume of vehicle on Days 16, 17 and 18 of gestation. Ovaries from 16- and 38-day-old female offspring were prepared for histological and stereological examination. The volume of the ovary and the number of primordial and primary follicles were significantly reduced in prenatally Dx-exposed infant and peripubertal female offspring compared with control offspring. The number of multilaminar follicles was decreased in infant female offspring. In peripubertal females, prenatal exposure to Dx increased the number of multilaminar and large follicles of all classes. Because vaginal opening did not occur up to Day 38 postpartum in the Dx-exposed offspring, the absence of ovulation and corpora lutea is confirmation that the onset of puberty had been delayed. We can conclude that overexposure to glucocorticoids early in life programs ovary development, which may affect fertility in adulthood.

Systematic Review: Puberty suppression with GnRH analogues in adolescents with gender incongruity.

CONTEXT: Gender incongruence is defined as disharmony between assigned gender and gender identity. Several interventions are liable in this case including genital affirming surgery among other surgical interventions such as harmonization, and also the use of gonadotropin-releasing hormone agonists (GnRHa) for gonadal shielding. This aids in preventing the development of secondary sexual characteristics related to the genetic sex. OBJECTIVE: Systematically review the treatment of gender incongruity with GnRHa analogues. DATA SOURCES: The data source of this research is from Pubmed-Medline and Embase. STUDY SELECTION: Articles published between 2009 and 2019 which studied transgender adolescents treated with GnRHa were carefully selected. DATA EXTRACTION: Were extracted: design, sample size, study context, targeted subjects of intervention, outcome measures, and results. RESULTS: Eleven studies were included. The use of GnRHa seems to be well tolerated by the studied population. When started in pubertal transition, it was associated with a more distinct resemblance to body shape than to the affirmed sex. In addition to preventing the irreversible phenotypic changes that occur in cross-hormonal therapy, the use of GnRHa can equally contribute to the mental health of these adolescents. LIMITATION: There are few consistent studies on the use of GnRHa for gender incongruence. CONCLUSION: As the population of transgender children and adolescents grows, they acquire knowledge and greater access to the various forms and stages of treatment for sex reassignment. The medical community needs to be adequately prepared to better serve this population and offer the safest resources available.

Effects of Prepubertal Exposure to Aroclor-1221 on Reproductive Development and Transcriptional Gene Expression in Female Rats.

Polychlorinated biphenyls (PCBs), as persistent organic pollutants, are environmental endocrine-disrupting chemicals (EDCs). We aim to investigate the effects of prepubertal exposure to PCBs on the reproductive development and expression and regulation of related genes in rats. Female rats were treated with Aroclor-1221 (A-1221) (4 mg/kg/day, 0.4 mg/kg/day) or castor oil daily from postnatal day (PND) 28 for 2 weeks by gavage. Morphological, histological, hormonal, and biochemical parameters were studied. Lower weight and relative weight of hypothalamus, earlier puberty onset, a longer length of the estrous cycle, lower serum estradiol and progesterone levels, accelerated ovarian folliculogenesis, and higher apoptotic index in the ovary were found. The in vitro fertilization study showed a lower fertilization rate and cleavage rate. The genetic study revealed higher expression of Kiss-1 mRNA and lower expression of GnRH mRNA in the hypothalamus and higher expression of AMH mRNA and lower expression of C-myc mRNA in the ovary. These confirmed the reproductive damage of A-1221 in rats.

Chronic effects of maternal tobacco-smoke exposure and/or α-lipoic acid treatment on reproductive parameters in female rat offspring.

Prenatal tobacco-smoke exposure negatively affects the reproductive functions of female offspring and oxidative stress plays a major role at this point. Alpha-lipoic acid (ALA), well known as a biological antioxidant, has been used as a nutritional supplement and as a therapeutic agent in the treatment of certain complications during pregnancy. We aimed to investigate the effects of maternal tobacco-smoke exposure and/or ALA administration on puberty onset, sexual behavior, gonadotrophin levels, apoptosis-related genes, apoptotic cell numbers and oxidative stress markers in the adult female rat offspring. Sprague-Dawley rats were divided into four groups; control, tobacco smoke (TS), TS+ALA and ALA groups. Animals were exposed to TS and/or ALA for 8 weeks before pregnancy and throughout pregnancy. All treatments ended with birth and later newborn female rats were selected for each experimental group. The experiment ended at postnatal day 74-77. Maternal tobacco smoke advanced the onset of puberty in the female offspring of the TS group (p < 0.05). In all treatment groups; the mean number of anogenital investigations and lordosis quality scores showed a decline, serum luteinizing hormone levels significantly increased (p < 0.05) and several histopathological changes in ovaries were observed compared to the control group. In addition, an increase in apoptotic marker levels and apoptotic cell numbers was detected in the ovaries of all treatment groups. Decreased TAS and increased TOS levels were detected in all treatment groups compared to control. These findings suggested that maternal tobacco smoke and/or ALA administration may be leading to the impaired reproductive health of female offspring. Abbreviations: ALA: alpha-lipoic acid; LH: luteinizing hormone; FSH: follicle-stimulating hormone; TAS: total antioxidant status; TOS: total oxidant status; Apaf1: apoptotic protease-activating factor 1; Casp3: caspase 3; Casp9: caspase 9; CF: cyst follicles; 4-HNE: 4-Hidroxynonenal; 8-OHdG: 8-hydroxydeoxyguanosine; TUNEL: terminal deoxynucleotidyl transferase-mediated deoxyuridine-biotin nick end labeling; ROS: reactive oxygen species; GnRHR: gonadotropin-releasing hormone receptor; HPG: hypothalamic-pituitary-gonadal; AMPK: AMP-activated protein kinase; ELISA: enzyme-linked immunosorbent assay; cDNA: complementary DNA; qPCR: quantitative real-time PCR; FC: follicular cysts; PF: primary follicle; SF: secondary follicle; GF: graafian follicle; CL: corpus luteum; DF: degenerated follicle; AF: atretic follicle.

Agrochemicals disrupt multiple endocrine axes in amphibians.

Concern over global amphibian declines and possible links to agrochemical use has led to research on the endocrine disrupting actions of agrochemicals, such as fertilizers, fungicides, insecticides, acaricides, herbicides, metals, and mixtures. Amphibians, like other species, have to partition resources for body maintenance, growth, and reproduction. Recent studies suggest that metabolic impairments induced by endocrine disrupting chemicals, and more particularly agrichemicals, may disrupt physiological constraints associated with these limited resources and could cause deleterious effects on growth and reproduction. Metabolic disruption has hardly been considered for amphibian species following agrichemical exposure. As for metamorphosis, the key thyroid hormone-dependent developmental phase for amphibians, it can either be advanced or delayed by agrichemicals with consequences for juvenile and adult health and survival. While numerous agrichemicals affect anuran sexual development, including sex reversal and intersex in several species, little is known about the mechanisms involved in dysregulation of the sex differentiation processes. Adult anurans display stereotypical male mating calls and female phonotaxis responses leading to successful amplexus and spawning. These are hormone-dependent behaviours at the foundation of reproductive success. Therefore, male vocalizations are highly ecologically-relevant and may be a non-invasive low-cost method for the assessment of endocrine disruption at the population level. While it is clear that agrochemicals disrupt multiple endocrine systems in frogs, very little has been uncovered regarding the molecular and cellular mechanisms at the basis of these actions. This is surprising, given the importance of the frog models to our deep understanding of developmental biology and thyroid hormone action to understand human health. Several agrochemicals were found to have multiple endocrine effects at once (e.g., targeting both the thyroid and gonadal axes); therefore, the assessment of agrochemicals that alter cross-talk between hormonal systems must be further addressed. Given the diversity of life-history traits in Anura, Caudata, and the Gymnophiona, it is essential that studies on endocrine disruption expand to include the lesser known taxa. Research under ecologically-relevant conditions will also be paramount. Closer collaboration between molecular and cellular endocrinologists and ecotoxicologists and ecologists is thus recommended.

Application of a combined aggregate exposure pathway and adverse outcome pathway (AEP-AOP) approach to inform a cumulative risk assessment: A case study with phthalates.

Advancements in measurement and modeling capabilities are providing unprecedented access to estimates of chemical exposure and bioactivity. With this influx of new data, there is a need for frameworks that help organize and disseminate information on chemical hazard and exposure in a manner that is accessible and transparent. A case study approach was used to demonstrate integration of the Adverse Outcome Pathway (AOP) and Aggregate Exposure Pathway (AEP) frameworks to support cumulative risk assessment of co-exposure to two phthalate esters that are ubiquitous in the environment and that are associated with disruption of male sexual development in the rat: di(2-ethylhexyl) phthalate (DEHP) and di-n-butyl phthalate (DnBP). A putative AOP was developed to guide selection of an in vitro assay for derivation of bioactivity values for DEHP and DnBP and their metabolites. AEPs for DEHP and DnBP were used to extract key exposure data as inputs for a physiologically based pharmacokinetic (PBPK) model to predict internal metabolite concentrations. These metabolite concentrations were then combined using in vitro-based relative potency factors for comparison with an internal dose metric, resulting in an estimated margin of safety of ~13,000. This case study provides an adaptable workflow for integrating exposure and toxicity data by coupling AEP and AOP frameworks and using in vitro and in silico methodologies for cumulative risk assessment.

Opposite effects of high- and low-dose di-(2-ethylhexyl) phthalate (DEHP) exposure on puberty onset, oestrous cycle regularity and hypothalamic kisspeptin expression in female rats.

Di-(2-ethylhexyl) phthalate (DEHP) is ubiquitous in the environment and has been proposed to lead to reproductive disruption. In this study, we systematically investigated the effects of different doses of DEHP exposure on female hypothalamic-pituitary-gonadal axis development. Female Sprague-Dawley rats were gavaged with vehicle (corn oil) or DEHP (5 or 500mgkg-1 day-1) during postnatal Days (PNDs) 22-28 or PNDs 22-70. Results demonstrated that the low and high doses of DEHP exerted opposite effects on puberty onset, circulating luteinising hormone, serum oestradiol and progesterone levels, with the low dose (5mgkg-1) promoting and the high dose (500mgkg-1) inhibiting these parameters. Significant dose-related differences were also found in the D500 group with longer oestrous cycle duration, lower ovarian/bodyweight ratio, fewer corpus lutea and more abnormal ovarian stromal tissue in comparison with the oil or D5 groups. Molecular data showed that the hypothalamic Kiss1 mRNA expression in the anteroventral periventricular but not in the arcuate nucleus significantly decreased in the D500 rats and increased in the D5 rats relative to the rats in the oil group. These findings suggested that the kisspeptin system is a potential target for DEHP to disrupt reproductive development and function.

Supplementation of various zinc sources modify sexual development and testicular IGF family gene expression in pre-pubertal male Japanese quail.

Zinc plays an important role in the regulation of insulin-like growth factor-I (IGF-I). IGF system, in turn, has a key role in the development and functions of the reproductive organs. This research was performed to investigate the effects of different sources of zinc on IGF-I gene expression and testicular development in pre-pubertal male Japanese quail. A total of 512 unsexed day-old Japanese quail chicks were randomly divided into 16 groups (4 dietary treatments × 4 replicates) and kept for 35 days. The control group diet was not supplemented with zinc whereas the diets of three groups were supplemented with 25 mg kg-1 zinc oxide (ZnO), zinc oxide nanoparticle (ZnON), and zinc-methionine (Zn-Met). On days 28 and 35, one birds from each subgroup were weighed, bled, and euthanized to evaluate gonado-somatic index (GSI), testicular histology, serum testosterone concentration, cloacal gland index (CGI), and the testicular IGF family gene expression. The results showed that GSI was higher in ZnON (2.307) than control (1.619) on day 35 (P < .05). Germinal epithelium thickness was higher in ZnON (78.88 µm) and Zn-Met (79.73 µm) than control (67.73 µm) on day 35 (P < .05). On day 35, the testosterone concentration was lowest in the control (5.830 ng/ml, P < .05). The CGI of 35-day-old birds was higher in Zn-Met (411.28) than the control (307.59, P < .05). IGF-IR mRNA expression was highest in Zn-Met group on day 28. Therefore, supplementation of diet with Zn-methionine is superior to other sources of zinc for diet supplementation in immature Japanese quail.

Prepubertal acrylamide exposure causes dose-response decreases in spermatic production and functionality with modulation of genes involved in the spermatogenesis in rats.

The increase in human infertility prevalence due to male reproductive disorders has been associated with extensive endocrine-disrupting chemical (EDC) exposure. Acrylamide (AA) is a compound formed spontaneously during heat processing of some foods that are mainly consumed by children and adolescents. In this study, we evaluated the prepubertal AA exposure effects on male adult reproductive physiology using a prepubertal experimental model to analyze the pubertal development, spermatogenesis hormones levels and genes expression involved in male reproductive function. This study is the first one to use the validated protocol to correlate the AA exposure with puberty development, as well as the AA-induced endocrine disrupting effects on reproductive axis. AA did not affect the age at puberty, the reproductive organ's weight and serum hormonal levels. AA reduces spermatogenesis, induces morphological and functional defects on sperm and alters transcript expression of sexual hormone receptors (Ar and Esr2), the transcript expression of Tnf, Egr2, Rhcg and Lrrc34. These findings suggest that excessive AA consumption may impair their reproductive capacity at adulthood, despite no changes in hormonal profile being observed.

The XY Female: Exploring Care for Adolescent Girls with Complete Androgen Insensitivity Syndrome.

Differences in Sex Development (DSD) encompasses many diagnoses, where the development of chromosomal make-up, gonadal development or anatomical development is atypical. XY, DSD is a classification under the recent international consensus statement, and XY females commonly encapsulate disorders of androgen synthesis and androgen action. Complete Androgen Insensitivity Syndrome (CAIS) is the most common XY, DSD diagnosis, which results in an individual having XY chromosomes, but the person is phenotypically female. This article explores the care and management of children and young people with a DSD and focuses on the diagnosis of CAIS in adolescence. Medical and surgical management is discussed, alongside sexual function, gender identity and the psychological impact of the diagnosis. The involvement of the multidisciplinary team is stressed, together with an emphasis on the investment that is needed in psychological and nursing support for girls with CAIS, and their families.

A mouse model to investigate the impact of testosterone therapy on reproduction in transgender men.

STUDY QUESTION: Can mice serve as a translational model to investigate the reproductive effects of testosterone (T) therapy commonly used by transgender men? SUMMARY ANSWER: T enanthate subcutaneous injections at 0.45 mg twice weekly can be used in the postpubertal C57BL/6N female mouse to investigate the reproductive effects of T therapy given to transgender men. WHAT IS KNOWN ALREADY: Most models of T treatment in female mice involve prenatal or prepubertal administration, which are not applicable to transgender men who often begin T therapy after puberty. Studies that have looked at the impact of postpubertal T treatment in female mice have generally not investigated reproductive outcomes. STUDY DESIGN, SIZE, DURATION: A total of 20 C57BL/6N female mice were used for this study. Study groups (n = 5 mice per group) included sesame oil vehicle controls and three doses of T enanthate (0.225, 0.45 and 0.90 mg). Mice were injected subcutaneously twice weekly for 6 weeks. PARTICIPANTS/MATERIALS, SETTING, METHODS: Daily vaginal cytology was performed prior to initiation of treatment to confirm that all mice were cycling. At 8-9 weeks of age, therapy with subcutaneous T enanthate (0.225, 0.45 or 0.90 mg) or the vehicle control was begun. T therapy continued for 6 weeks, at which point mice were sacrificed and compared to control mice sacrificed during diestrus/metestrus. Data collected included daily vaginal cytology, weekly and terminal reproductive hormone levels, terminal body/organ weights/measurements, ovarian follicular distribution/morphology and corpora lutea counts. MAIN RESULTS AND THE ROLE OF CHANCE: Of the mice treated with 0.90 mg T enanthate, two of five mice experienced vaginal prolapse, so this group was excluded from further analysis. T enanthate administration twice weekly at 0.225 or 0.45 mg resulted in cessation of cyclicity and persistent diestrus. One of five mice at the 0.225-mg dose resumed cycling after 2.5 weeks of T therapy. As compared to controls, T-treated mice had sustained elevated T levels and luteinizing hormone (LH) suppression in the terminal blood sample. T-treated mice demonstrated increases in clitoral area and atretic cyst-like late antral follicles (0.45 mg only) as compared to controls. No reduction in primordial, primary, secondary or total antral follicle counts was detected in T-treated mice as compared to controls, and T-treated mice demonstrated an absence of corpora lutea. LIMITATIONS, REASONS FOR CAUTION: Mouse models can provide us with relevant key findings for further exploration but may not perfectly mirror human reproductive physiology. WIDER IMPLICATIONS OF THE FINDINGS: To our knowledge, this report describes the first mouse model mimicking T therapy given to transgender men that facilitates analysis of reproductive changes. This model allows for future studies comparing duration and reversibility of T-induced changes, on the reproductive and other systems. It supports a role for T therapy in suppressing the hypothalamic-pituitary-gonadal axis in adult female mice as evidenced by LH suppression, persistent diestrus and absence of corpora lutea. The increase in atretic cyst-like late antral follicles aligns with the increased prevalence of polycystic ovary morphology seen in case series of transgender men treated with T therapy. The results also suggest that T therapy does not deplete the ovarian reserve. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the American Society for Reproductive Medicine/Society of Reproductive Endocrinology and Infertility Grant and NIH R01-HD098233 to M.B.M. and University of Michigan Office of Research funding (U058227). H.M.K. was supported by the Career Training in Reproductive Biology and Medical Scientist Training Program T32 NIH Training Grants (T32-HD079342, T32-GM07863) as well as the Cellular and Molecular Biology Program. The University of Virginia Center for Research in Reproduction Ligand Assay and Analysis Core is supported by the Eunice Kennedy Shriver NICHD/NIH (NCTRI) Grant P50-HD28934. E.E.M. consults for Allergan. No other authors have competing interests.

Benzyl butyl phthalate non-linearly affects rat Leydig cell development during puberty.

Benzyl butyl phthalate (BBP) is a widely used plasticizer and has raised public health concerns. Here, we report the effects of BBP on the testis development during rat puberty. BBP (0, 10, 100 or 1000 mg/kg) was gavaged to 35-day-old male Sprague Dawley rats for 21 days. The serum testosterone levels, Leydig cell number, the expressions of Leydig and Sertoli cell genes and proteins were measured. The in vitro effects on steroidogenesis and gene expression in immature Leydig cells were observed. BBP significantly increased serum testosterone level at 10 mg/kg but lowered its level at 1000 mg/kg without affecting serum luteinizing hormone and follicle-stimulating hormone levels. BBP increased Leydig cell number at all doses but inhibited steroidogenic capacity per Leydig cell at 1000 mg/kg. BBP significantly increased the ratio of phosphos-AKT2 (pAKT2)/AKT2, and phosphos-ERK1/2 (pERK1/2)/ERK1/2 in the testis. Mono-benzyl phthalate (the metabolite of BBP) inhibited steroidogenesis but BBP did not affect androgen production in immature Leydig cells in vitro. In conclusion, BBP non-linearly regulates Leydig cell development by increasing Leydig cell number but inhibiting steroidogenesis.

Molecular cloning, characterization of dax1 gene and its response to progesterone in Misgurnus anguillicaudatus.

Progesterone (P4) are aquatic contaminants that can impair fish reproduction even in low concentrations. The aim of this study was to investigate the effects of P4 on the sex differentiation, by quantitative determination of transcriptional changes of a candidate target gene (dax1, has a function in the sex determination and gonadal differentiation of several vertebrate species) in Misgurnus anguillicaudatus. We first cloned and characterized the full-length cDNAs for the dax1 in M. anguillicaudatus (designated as Ma-dax1). Sequence analysis reveals that Ma-dax1 shares high homology with dax1 in other species. Quantitative real-time PCR (qRT-PCR) and in situ hybridization showed that Ma-dax1 gene was highly conserved during vertebrate evolution and involved in a wide range of developmental processes including embryogenesis, central nervous system development and gonad development. For the P4 administration assay, groups of mature fish were exposed for 1, 7, 14, 21 and 28 days to nominal concentrations of 10, 100, and 1000 ng/L P4 in a flow-through system. Quantification of Ma-dax1 transcripts revealed the expression of Ma-dax1 mRNA is altered after P4 treatment in mature gonads. Those showed that P4 could influence the sexual development and sex differentiation in M. anguillicaudatus by disturbing sex differentiation-associated gene expression, and dax1 can be used as a sensitive molecular biomarker for early warning to monitor the environmental progestins chemicals in fresh water environment.

In utero exposure to simvastatin reduces postnatal survival and permanently alters reproductive tract development in the Crl:CD(SD) male rat.

We showed previously that in utero exposure to the cholesterol-lowering drug simvastatin (SMV) during sex differentiation lowers fetal lipids and testicular testosterone production (T Prod) in Hsd:SD rats. Here, the effects of SMV on fetal lipids and T Prod in Crl:CD(SD) rats were correlated with postnatal alterations in F1 males. The current study was conducted in two parts: 1) a prenatal assessment to confirm and further characterize the dose response relationship among previously reported alterations of SMV on fetal T Prod and the fetal lipid profile and 2) a postnatal assessment to determine the effects of SMV exposure during the periods of major organogenesis and/or sexual differentiation on F1 offspring growth and development. We hypothesized that SMV would have adverse effects on postnatal development and sexual differentiation as a consequence of the disruptions of fetal lipid levels and testicular T Prod since fetal cholesterol is essential for normal intrauterine growth and development and steroid synthesis. In the prenatal assessment, SMV was administered orally at 0, 15.6, 31.25, 62.5, 80, 90, 100, and 110 mg SMV/kg/d from GD 14-18, the period that cover the critical window of sex differentiation in the male rat fetus. T Prod was maximally reduced by ~40% at 62.5 mg/kg/d, and higher doses induced overt maternal and toxicity. In the postnatal assessment, SMV was administered at 0, 15.6, 31.25, and 62.5 mg/kg/d from GD 8-18 to determine if it altered postnatal development. We found that exposure during this time frame to 62.5 mg SMV/kg/d reduced pup viability by 92%, decreased neonatal anogenital distance, and altered testis histology and morphology in 17% of the F1 males. In another group, SMV was administered only during the masculinizing window (GD14-18) at 62.5 mg/kg/d to determine if male rat sexual differentiation and postnatal reproductive development were altered. SMV-exposed F1 males displayed female-like areolae/nipples, delayed puberty, and reduced seminal vesicle and levator ani-bulbocavernosus weights. Together, these results demonstrate that in utero exposure to SMV reduces offspring viability and permanently disrupts reproductive tract development in the male offspring. While the effects of high dose, short term in utero exposure to SMV in the adult male are likely androgen-dependent and consistent with the 40% reduction in T Prod in the fetal testes, long-term, lower dose administration induced some effects that were likely not mediated by decreased T Prod.

Water and soil pollution as determinant of water and food quality/contamination and its impact on male fertility.

Over the past two decades, public health has focused on the identification of environmental chemical factors that are able to adversely affect hormonal function, known as endocrine disruptors (EDs). EDs mimic naturally occurring hormones like estrogens and androgens which can in turn interfere with the endocrine system. As a consequence, EDs affect human reproduction as well as post and pre-natal development. In fact, infants can be affected already at prenatal level due to maternal exposure to EDs. In particular, great attention has been given to those chemicals, or their metabolites, that have estrogenic properties or antagonistic effects on the activity of androgen or even inhibiting their production. These compounds have therefore the potential of interfering with important physiological processes, such as masculinization, morphological development of the urogenital system and secondary sexual traits. Animal and in vitro studies have supported the conclusion that endocrine-disrupting chemicals affect the hormone-dependent pathways responsible for male gonadal development, either through direct interaction with hormone receptors or via epigenetic and cell-cycle regulatory modes of action. In human populations, epidemiological studies have reported an overall decline of male fertility and an increased incidence of diseases or congenital malformations of the male reproductive system. The majority of studies point towards an association between exposure to EDs and male and/or female reproductive system disorders, such as infertility, endometriosis, breast cancer, testicular cancer, poor sperm quality and/or function. Despite promising discoveries, a causal relationship between the reproductive disorders and exposure to specific toxicants has yet to be established, due to the complexity of the clinical protocols used, the degree of occupational or environmental exposure, the determination of the variables measured and the sample size of the subjects examined. Despite the lack of consistency in the results of so many studies investigating endocrine-disrupting properties of many different classes of chemicals, the overall conclusion points toward a positive association between exposure to EDs and reproductive system. Future studies should focus on a uniform systems to examine human populations with regard to the exposure to specific EDs and the direct effect on the reproductive system.

Molecular characterisation of oestrogen receptor ERα and the effects of bisphenol A on its expression during sexual development in the Chinese giant salamander (Andrias davidianus).

The aim of this study was to characterise the molecular structure of the oestrogen receptor ERα and to evaluate the effect of bisphenol A (BPA) on ERα expression during sexual development of the Chinese giant salamander (Andrias davidianus). The ERα cDNA of A. davidianus includes an open reading frame of 1755bp (encoding 584 amino acids), a 219-bp 5' untranslated region (UTR) and a 611-bp 3'UTR. A polyadenylation signal was not found in the 3'UTR. Amino acid sequence analysis showed high homology between ERα of A. davidianus and that of other amphibians, such as Andrias japonicas (99.66% identity) and Rana rugose (81.06% identity). In 3-year-old A. davidianus, highest ERα expression was observed in the liver and gonads. During different developmental stages in A. davidianus (from 1 to 3 years of age), ERα expression in the testes increased gradually. ERα was localised in the epithelial cells of seminiferous lobules and in interstitial cells. ERα-positive cells were more abundant in the interstitial tissue during testicular development. ERα was located in the nucleus of oocytes during ovary development. We found that the sex of 6-month-old A. davidianus larvae could not be distinguished anatomically. The sex ratio did not change after larvae were treated with 10µM BPA for 1 month. However, BPA treatment reduced bodyweight and ERα expression in the gonads in male larvae.

Psychosexual Aspects, Effects of Prenatal Androgen Exposure, and Gender Change in 46,XY Disorders of Sex Development.

CONTEXT: In 46,XY disorders of sexual development (DSD) patients, several factors may affect psychosexual development, leading to gender identity discrepancy and gender change later in life. Prenatal sexual steroid exposure and external genital virilization are considered to influence human psychosexual development, but their roles not completely understood yet. DESIGN: A total of 144 individuals (18 to 60 years of age) with a clinical/molecular diagnosis of 46,XY DSD from a single tertiary center were enrolled. Psychosexual outcomes (gender role, gender identity, and sexual orientation) were assessed using questionnaires and psychological test. The Sinnecker score was used for genital virilization measurement. Prenatal androgen exposure was estimated according to 46,XY DSD etiology. RESULTS: We found a positive association between prenatal androgen exposure and male psychosexual outcomes. Alternatively, prenatal estrogen exposure, age of gonadectomy, and the degree of external genital virilization did not influence any psychosexual outcome. There were 19% (n = 27) with gender change, which was associated with prenatal androgen exposure (P < 0.001) but not with the external genital virilization. The median age of gender change was 15 years, but most of the patients reported the desire for gender change earlier. CONCLUSIONS: Prenatal androgen exposure influenced psychosexual development in 46,XY DSD favoring male psychosexuality in all psychosexual outcomes, whereas the degree of external genital virilization did not influence these outcomes. The organizational effect of sexual steroids on psychosexuality at puberty appears to be weak in comparison with the prenatal effects. Prenatal androgen exposure also influenced female-to-male gender change frequency. All 46,XY DSD conditions with prenatal androgen exposure must be followed for gender issues in their management.