Suspect and non-target screening of ovarian follicular fluid and serum - identification of anthropogenic chemicals and investigation of their association to fertility

In this work, ultra-high performance liquid chromatography-high resolution (Orbitrap) mass spectrometry-based suspect and non-target screening was applied to follicular fluid (n = 161) and serum (n = 116) from women undergoing in vitro fertilization in order to identify substances that may be associated with decreased fertility. Detected features were prioritized for identification based on (i) hazard/exposure scores in a database of chemicals on the Swedish market and an in-house database on per- and polyfluoroalkyl substances (PFAS); (ii) enrichment in follicular fluid relative to serum; and (iii) association with treatment outcomes. Non-target screening detected 20 644 features in follicular fluid and 13 740 in serum. Two hundred and sixty-two features accumulated in follicular fluid (follicular fluid: serum ratio >20) and another 252 features were associated with embryo quality. Standards were used to confirm the identities of 21 compounds, including 11 PFAS. 6-Hydroxyindole was associated with lower embryo quality and 4-aminophenol was associated with higher embryo quality. Overall, we show the complexity of follicular fluid and the applicability of suspect and non-target screening for discovering both anthropogenic and endogenous substances, which may play a role in fertility in women

P16-30: Examining the need for an ovarian-specific steroidogenic assay to prioritize potential endocrine disrupting chemicals

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Estrogen-like effects of diet-derived cadmium differ from those of orally administered CdCl(2) in the ERE-luc estrogen reporter mouse model

Cadmium (Cd), an environmental and dietary contaminant, has been described to mimic the effects of 17\u3b2-estradiol (E(2)) in selected model systems when studied as an inorganic salt. However, inorganic Cd salts do not represent the main form of Cd exposure in general human populations. The aims of this study were to compare the estrogen-like effects and the bioavailability of dietary Cd to inorganic CdCl(2). Adult ovariectomized ERE-luc reporter mice were administered two bread based diets containing different concentrations of Cd (17.57 and 49.22\u3bcg/kg, corresponding to oral intakes of 1.8 and 5.1\u3bcg/kg body weight (bw) per day, respectively), inorganic CdCl(2) (1\u3bcg/kg bw per day by gavage) or E(2) (5\u3bcg/kg bw per day pellet) for 21 days. The effects on estrogen signaling were investigated by studying the uterine weights, luciferase activation, and expression of endogenous estrogen target genes. The uterine weight was significantly increased by both CdCl(2) and E(2) but not by the Cd containing diets. All treatments modulated the expression of luciferase and the endogenous estrogen target genes; however, there was no consistent overlap between the responses triggered by the bread diets and the responses stimulated by CdCl(2) or E(2). Oral exposure to Cd was calculated and the concentrations in liver and kidneys quantified to estimate the amount of absorbed Cd retained in tissues. The results suggest significantly lower absorption and/or tissue retention of dietary Cd compared to CdCl(2) following oral exposure. Altogether, our results support previous reports on in vivo estrogenicity of CdCl(2) but do not suggest the same activity for diet bound Cd. This study calls for caution when extrapolating results from pure compound studies (e.g. estrogenicity of CdCl(2)) to dietary exposure scenarios (e.g. estrogenicity of diet bound Cd). Further basic research is needed on the mechanisms of interaction between Cd and the estrogen signaling, biologically active species of Cd, and biomarkers of estrogen-like effects of Cd in vivo before human health risk assessment on the hormone disruptive effects of Cd can be carried out

Persistent organic pollutants, pre-pregnancy use of combined oral contraceptive and time-to-pregnancy in SELMA cohort

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Dietary sources of lignans and isoflavones modulate responses to estradiol in estrogen reporter mice

Dietary phytoestrogens, such as the lignan metabolite enterolactone (ENL) and the isoflavone genistein (GEN), are suggested to modulate the risk of estrogen-dependent disease (e.g., breast cancer) through regulation of estrogen signaling. However, the effects of complex food items containing lignans or isoflavones on estrogen receptor (ER) transactivation have not been assessed so far. In this study, the modulation of ER-mediated signaling by dietary sources of lignans (cereals and flaxseed) and isoflavones (soy) was studied in vivo. Adult ovariectomized 3 x ERE-luciferase (luc) reporter mice received isocaloric diets supplemented with flaxseed, rye, wheat, or soy for 40 h or two weeks, and an additional group of mice was challenged with 17beta-estradiol (E(2)) following the two-week dietary intervention. In non-E(2)-treated mice, soy diet induced luc expression in liver, mammary gland, and pituitary gland while the other diets had no effects. Interestingly, all diets modulated the E(2)-induced luc expression. In particular rye diet efficiently reduced E(2)-induced luc expression as well as uterine growth, the hallmark of estrogen action in vivo. It is concluded that dietary sources of lignans and isoflavones can modulate estrogen signaling in vivo. The results suggest intriguing possibilities for the modulation of the risk of estrogen-dependent diseases by dietary means.

Development and application of a health-based framework for informing regulatory action in relation to exposure of microplastic particles in California drinking water

Microplastics have been documented in drinking water, but their effects on human health from ingestion, or the concentrations at which those effects begin to manifest, are not established. Here, we report on the outcome of a virtual expert workshop conducted between October 2020 and October 2021 in which a comprehensive review of mammalian hazard studies was conducted. A key objective of this assessment was to evaluate the feasibility and confidence in deriving a human health-based threshold value to inform development of the State of California’s monitoring and management strategy for microplastics in drinking water. A tiered approach was adopted to evaluate the quality and reliability of studies identified from a review of the peer-reviewed scientific literature. A total of 41 in vitro and 31 in vivo studies using mammals were identified and subjected to a Tier 1 screening and prioritization exercise, which was based on an evaluation of how each of the studies addressed various quality criteria. Prioritized studies were identified largely based on their application and reporting of dose–response relationships. Given that methods for extrapolating between in vitro and in vivo systems are currently lacking, only oral exposure in vivo studies were identified as fit-for-purpose within the context of this workshop. Twelve mammalian toxicity studies were prioritized and subjected to a Tier 2 qualitative evaluation by external experts. Of the 12 studies, 7 report adverse effects on male and female reproductive systems, while 5 reported effects on various other physiological endpoints. It is notable that the majority of studies (83%) subjected to Tier 2 evaluation report results from exposure to a single polymer type (polystyrene spheres), representing a size range of 0.040 to 20 µm. No single study met all desired quality criteria, but collectively toxicological effects with respect to biomarkers of inflammation and oxidative stress represented a consistent trend. While it was possible to derive a conservative screening level to inform monitoring activities, it was not possible to extrapolate a human–health-based threshold value for microplastics, which is largely due to concerns regarding the relative quality and reliability of current data, but also due to the inability to extrapolate data from studies using monodisperse plastic particles, such as polystyrene spheres to an environmentally relevant exposure of microplastics. Nevertheless, a conservative screening level value was used to estimate a volume of drinking water (1000 L) that could be used to support monitoring activities and improve our overall understanding of exposure in California’s drinking water. In order to increase confidence in our ability to derive a human–health-based threshold value in the future, several research recommendations are provided, with an emphasis towards strengthening how toxicity studies should be conducted in the future and an improved understanding of human exposure to microplastics, insights critically important to better inform future risk assessments

The hydroxysteroid (17β) dehydrogenase family gene HSD17B12 is involved in the prostaglandin synthesis pathway, the ovarian function, and regulation of fertility.

The HSD17B12 gene belongs to the hydroxysteroid (17β) dehydrogenase superfamily, and it has been implicated in the conversion of estrone to estradiol as well as in the synthesis of arachidonic acid (AA). AA is a precursor of prostaglandins, which are involved in the regulation of female reproduction, prompting us to study the role of HSD17B12 enzyme in the ovarian function. We found a broad expression of HSD17B12 enzyme in both human and mouse ovaries. The enzyme was localized in the theca interna, corpus luteum, granulosa cells, oocytes and surface epithelium. Interestingly, haploinsufficiency of the HSD17B12 gene in female mice resulted in subfertility, indicating an important role for HSD17B12 enzyme in the ovarian function. In line with significantly increased length of the diestrus phase, the HSD17B(+/-) females gave birth less frequently than WT females, and the litter size of HSD17B12(+/-) females was significantly reduced. Interestingly, we observed meiotic spindle formation in immature follicles, suggesting defective meiotic arrest in HSD17B12(+/-) ovaries. The finding was further supported by transcriptome analysis showing differential expression of several genes related to the meiosis. In addition, polyovular follicles and oocytes trapped inside the corpus luteum were observed, indicating a failure in the oogenesis and ovulation, respectively. Intraovarian concentrations of steroid hormones were normal in HSD17B12(+/-) females, whereas the levels of AA and its metabolites (6-keto PGT1a, PGD2, PGE2, PGF2alfa and TXB2) were decreased. In conclusion, our study demonstrates that HSD17B12 enzyme plays an important role in female fertility through its role in AA metabolism