Combined exposure to engineered nanoparticles (ENPs) and anthropogenic contaminants can lead to changes in bioavailability, uptake and thus effects of both groups of contaminants. In this study we investigated effects of single and combined exposures of silver (Ag) nanoparticles (AgNPs) and the synthetic hormone 17a-ethinylestradiol (EE2) on tissue uptake of both contaminants in juvenile turbot (Scophthalmus maximus). Silver uptake and tissue distribution (gills, liver, kidney, stomach, muscle and bile) were analyzed following a 14-day, 2-h daily pulsed exposure to AgNPs (2 mg L 1 and 200 mg L 1), Agþ (50 mg L 1), EE2 (50 ng L 1) and AgNP þ EE2 (2 or 200 mg L 1þ50 ng L 1). Effects of the exposures on plasma vitellogenin (Vtg) levels, EE2 and steroid hormone concentrations were investigated. The AgNP and AgNP þ EE2 exposures resulted in similar Ag concentrations in the tissues, indicating that combined exposure did not influence Ag uptake in tissues. The highest Ag concentrations were found in gills. For the Agþ exposed fish, the highest Ag concentrations were measured in the liver. Our results show dissolution processes of AgNPs in seawater, indicating that the tissue concentrations of Ag may partly originate from ionic release. Plasma EE2 concentrations and Vtg induction were similar in fish exposed to the single contaminants and the mixed contaminants, indicating that the presence of AgNPs did not significantly alter EE2 uptake. Similarly, concentrations of most steroid hormones were not significantly altered due to exposures to the combined contaminants versus the single compound exposures. However, high concentrations of AgNPs in combination with EE2 caused a drop of estrone (E1) (female fish) and androstenedione (AN) (male and female fish) levels in plasma below quantification limits. Our results indicate that the interactive effects between AgNPs and EE2 are limited, with only high concentrations of AgNPs triggering synergistic effects on plasma steroid hormone concentrations in juvenile turbots. | acceptedVersion

Perfluorooctane sulfonate (PFOS) is associated with male reproductive disorder, but the related mechanisms are still unclear. In this study, we used in vivo and in vitro models to explore the role of Sertoli cell-derived exosomes (SC-Exo)/miR-9-3p/StAR signaling pathway on PFOS-induced suppression of testosterone biosynthesis. Forty male ICR mice were orally administrated PFOS (0.5–10 mg/kg/bw) for 4 weeks. Bodyweight, organ index, sperm count, reproductive hormones were evaluated. Primary Sertoli cells and Leydig cells were used to delineate the molecular mechanisms that mediate the effects of PFOS on testosterone biosynthesis. Our results demonstrated that PFOS dose-dependently induced a decrease in sperm count, low levels of testosterone, and damage in testicular interstitium morphology. In vitro models, PFOS significantly increased miR-9-3p levels in Sertoli cells and SC-Exo, accompanied by a decrease in testosterone secretion and StAR expression in Leydig cells when Leydig cells were exposed to SC-Exo. Meanwhile, inhibition of SC-Exo or miR-9-3p by their inhibitors significantly rescued PFOS-induced decreases in testosterone secretion and the mRNA and protein expression of the StAR gene in Leydig cells. In summary, the present study highlights the role of the SC-Exo/miR-9-3p/StAR signaling pathway in PFOS-induced suppression of testosterone biosynthesis, advancing our understanding of molecular mechanisms for PFOS-induced male reproductive disorders.

Bisphenol A (BPA) is widely used by manufacturers and in consumer products. Its release in the environment may affect male reproductive function. In this study, we examined the effect of low dose (0.1 mg/kg BW), short term exposure during puberty (PD21-35) on adult rat male reproduction. The results indicated that such exposure reset growth hormone (GH) and follicular stimulating hormone (FSH) homeostasis and resulted in a significantly higher level of serum testosterone without affecting serum luteinizing hormone level. QPCR and Western blot results showed that BPA significantly up-regulated selective genes/proteins in the Leydig cell steroidogenic pathway, including steroidogenic acute regulatory protein, cytochrome P450 11A1, cytochrome P450 17A, and low-density lipoprotein receptor. RNA-Seq analysis of testicular RNAs showed that BPA significantly affected the gene profiles of multiple testicular interstitial populations without affecting germ cells. Also, GO- and KEGG-analysis suggested that IGF1-related PI3K/AKT signaling was activated, which was confirmed by the increased phosphorylation of IRS1, AKT1 and CREB. The results indicated that a low-dose, short-term BPA exposure during puberty affected the adult male rat pituitary (GH and FSH) and testis (testosterone) homeostasis.

Accumulating evidences indicated that heavy metals may disrupt human sex hormones. However, the combined effects of heavy metals on sex hormones remain to be clarified. To explore the independent and combined associations between heavy metal exposure and serum sex hormones among adults, data of 2728 adults from the National Health and Nutrition Examination Survey (NHANES) was applied. We examined independent and combined associations of fourteen urinary heavy metals and three serum sex steroid hormones (total testosterone (TT), estradiol (E2) and sex hormone-binding globulin (SHBG)). Multivariate linear regression was used to evaluate the independent associations between metal exposure and sex hormone alterations. Principle component analysis -weighted quantile sum regression (PCA-WQSR) model was performed to estimate the combined associations in our individuals. In the co-exposure model, we determined that weighted quantile sum (WQS) index of industrial pollutants was negatively associated with E2 in females (WQS Percent change₈₋ₘₑₜₐₗ = -20.6%; 95% CI: -30.1%, -9.96%), while in males WQS index of water pollutants was negatively related to SHBG (WQS Percent change₈₋ₘₑₜₐₗ = -5.35%; 95% CI: -9.88%, -0.598%). Cadmium (Cd), tin (Sn) and lead (Pb) were the dominating metals of female E2-negative association while Ba was the leading contributor related to male SHBG reduction, which was consistent with the results of multivariate linear regression. Additionally, in postmenopausal women, the associations of E2 decrease with heavy metal co-exposure remained significant while Cd and monomethylarsonic acid (MMA) were identified as hazardous metals in the mixture. We concluded that the exposure to heavy metals was associated with human sex hormone alterations in independent or combined manners. Considering the design of NHANES study, further studies from other national-representative surveys are necessary.

Steroid hormones are prevalent in the environment and have become emerging pollutants, but little is known about their effects on soil microbial community composition and function. In the present study, three representative soils in China were amended with environmentally relevant concentrations of testosterone and responses of soil bacterial community composition and soil function were assessed using high-throughput sequencing and nontargeted metabolomics. Our results showed that testosterone exposure significantly shifted bacterial community structure and metabolic profiles in soils at Ningbo (NB) and Kunming (KM), which may reflect high bioavailability of the hormone. Abundances of several bacterial taxa associated with nutrient cycling were reduced by testosterone and metabolites related to amino acid metabolism were downregulated. A close connection between bacterial taxa and specific metabolites was observed and confirmed by Procrustes tests and a co-occurrence network. These results provide an insight into the effects of steroid hormones on soil microbial community and highlight that nontargeted metabolomics is an effective tool for investigating the impacts of pollutants.

N⁶-methyladenosine (m6A) modification, the most prevalent form of RNA methylation, modulates gene expression post-transcriptionally. Di-(2-ethylhexyl) phthalate (DEHP) is a common environmental endocrine disrupting chemical that induces testicular injury due to the inhibition of the demethylase fat mass and obesity-associated protein (FTO) and increases the m6A modification. How FTO-mediated m6A modification in testicular Leydig cell injury induced by DEHP remains unclear. Here, the TM3 Leydig cell line was treated with mono-(2-ethylhexyl) phthalate (MEHP), the main metabolite of DEHP in the body, as well as FB23-2, an inhibitor of FTO. Decreased levels of testosterone in the culture supernatant, significantly increased apoptosis, and a remarkable upregulation of global m6A modification were found in both TM3 cells treated with MEHP and FB23-2. Transcriptome sequencing showed that both treatments significantly induced apoptosis-associated gene expression. Methylated RNA immunoprecipitation sequencing showed that the Leydig cell injury induced by upregulated m6A modification could be associated with multiple physiological disorders, including histone acetylation, reactive oxygen species biosynthesis, MAPK signaling pathway, hormone secretion regulation, autophagy regulation, and male gonadal development. Overall, the inhibition of FTO-mediated up-regulation of m6A could be involved in MEHP-induced Leydig cell apoptosis.

Recently, bioaccumulation of dietary organic selenium (Se) in the ovaries and inhibition of reproduction in female aquatic animals have been reported. However, there is limited data on the subtle reproductive impacts of waterborne exposure to inorganic Se in fish. Here, zebrafish embryos (2 h post-fertilization) were exposed to solutions with environmentally relevant levels of Na₂SeO₃ with concentrations of 0 (control), 7.98 ± 0.31, 25.14 ± 0.15, and 79.60 ± 0.81 μg Se/L for 120 d until they reached sexual maturity. Female zebrafish were selected for reproductive toxicity assessment. In the early embryonic stage, whole-mount in situ hybridization of zebrafish embryos showed that waterborne Na₂SeO₃ exposure did not affect the observed location of vasa expression in primordial germ cells at 24, 48, and 72 h post-fertilization. Life-cycle exposure to 25.14 ± 0.15 and 79.60 ± 0.81 μg Se/L Na₂SeO₃ did not change the testosterone and 17β-estradiol contents in female zebrafish at the endpoint of exposure, but significantly reduced the proportion of early vitellogenic oocytes and mature oocytes. Follicle maturity retardation was accompanied by changes in transcriptional levels of the genes related to the hypothalamus-pituitary-gonad-liver (HPGL) axis. Transcriptional levels of cyp19a and lhr in the ovary were down-regulated, while the transcriptional level of fshr in the ovaries was up-regulated. In the 21-day cumulative spawning experiment, Na₂SeO₃ (25.14 ± 0.15 and 79.60 ± 0.81 μg Se/L) caused fewer eggs to be produced. Additionally, the malformation of zebrafish offspring significantly increased in the group exposed to 79.60 ± 0.81 μg Se/L. In conclusion, for the first time, this study shows that life-cycle exposure to environmentally relevant concentrations of waterborne Na₂SeO₃ significantly delays ovarian maturation and reduces the fertility of the female zebrafish.

As a crucial factor in male reproduction, androgens may represent an intermediate biological mechanism linking metal exposure with effects on semen quality. This study aimed to investigate the association between metal exposure and semen quality, and to assess the mediating role of seminal androgens between metal exposure and semen quality. We investigated the presence of 10 metals in semen and assessed their effect on semen quality in 1136 men recruited from a hospital in Shenzhen, China. Of these, 464 subjects were randomly selected for 4 androgens detection in semen. Cross-sectional associations between single/multiple metals, androgen levels and semen quality were explored by multivariable linear regressions. Mediation analysis was performed to detect the role of seminal androgens on the association between metal exposure and semen quality. Seminal selenium and iron were positively associated with both sperm concentration and total sperm count. Negative associations were observed between both manganese and zinc and sperm concentration, molybdenum and total sperm count, copper and sperm motility. Furthermore, we found significant dose-dependent relationships between both iron and selenium levels and dihydrotestosterone (DHT), arsenic levels and testosterone, as well as zinc and dehydroepiandrosterone. Mediation analysis indicated that higher seminal iron and selenium were associated with an increasing sperm concentration after controlling for DHT, with 10.32% and 12.89% of these associations were mediated by DHT, respectively. A similar mediation effect of DHT was observed in the associations between iron and selenium levels and total sperm count (13.39% and 21.57% mediation, respectively). Our findings suggested that the presence of selenium and iron in semen was beneficial to sperm concentration and total count. Seminal manganese, zinc, molybdenum and copper may be associated with reduced semen quality. The associations between seminal selenium and iron and sperm concentration and total count were partially explained by the concomitant variation of seminal DHT.

Tributyltin (TBT), an organotin compound once widely used in agriculture and industry, has been reported to induce obesity and endocrine disruption. Gut microbiota has a strong connection with the host’s physiology. Nevertheless, the influences of TBT exposure on gut microbiota and whether TBT-influenced gut microbiota is related to TBT-induced toxicity remain unclear. To fill these gaps, ICR (CD-1) mice were respectively exposed to TBT at NOEL (L-TBT) and tenfold NOEL (H-TBT) daily by gavage for 8 weeks in the current study. The results showed that TBT exposure significantly increased body weight as well as epididymal fat, and led to adipocyte hypertrophy, dyslipidemia and impaired glucose and insulin homeostasis in mice. Additionally, TBT exposure significantly decreased the levels of T4, T3 and testosterone in serum. Also of note, TBT exposure changed gut microbiota composition mainly by decreasing Bacteroidetes and increasing Firmicutes proportions. To confirm the role of gut microbiota in TBT-induced overweight and hormonal disorders, fecal microbiota transplantation was performed and the mice receiving gut microbiota from H-TBT mice had similar phenotypes with their donor mice including significant body weight and epididymal fat gain, glucose and insulin dysbiosis and hormonal disorders. These results suggested that gut microbiome altered by TBT exposure was involved in the TBT-induced increased body weight, impaired glucose and insulin homeostasis and endocrine disruption in mice, providing significant evidence and a novel perspective for better understanding the mechanism by which TBT induces toxicity.

Controversial glyphosate-based herbicides (GBHs) are the most frequently used herbicides globally. GBH residues in the wild, in animal and human food may expose non-target organisms to health risks, yet the developmental and cumulative effects of GBHs on physiology and reproduction remain poorly understood. We present the first long-term study on the effects of subtoxic GBH exposure (160 mg/kg) on multiple key physiological biomarkers (cellular oxidative status and neurotransmitters), gut microbiome, reproductive hormones, and reproduction in an avian model. We experimentally exposed in Japanese quail females and males (Coturnix japonica) to GBHs and respective controls from the age of 10 days–52 weeks. GBH exposure decreased hepatic activity of an intracellular antioxidant enzyme (catalase), independent of sex, but did not influence other intracellular oxidative stress biomarkers or neurotransmitter enzyme (acetylcholinesterase). GBH exposure altered overall gut microbiome composition, especially at a younger age and in females, and suppressed potentially beneficial microbes at an early age. Many of the microbial groups increased in frequency from 12 to 28 weeks under GBH exposure. GBH exposure decreased male testosterone levels both at sexual maturity and at 52 weeks of exposure, but did not clearly influence reproduction in either sex (maturation, testis size or egg production). Future studies are needed to characterize the effects on reproductive physiology in more detail. Our results suggest that cumulative GBH exposure may influence health and reproduction-related traits, which is important in predicting their effects on wild populations and global poultry industry.