Jodi Anne Flaws

Phthalates are plasticizers and solvents used in numerous consumer products. Exposure to a phthalate mixture (Mix) alters estrous cyclicity, fertility indices, ovarian follicle dynamics, and steroidogenesis. In cultured antral follicles, phthalates alter the aryl hydrocarbon receptor (AHR) pathway, which is known for estradiol (E2) and xenobiotic metabolism. However, how phthalates affect the AHR pathway in the ovary in vivo and how this pathway changes throughout the reproductive lifespan are unknown. Thus, we hypothesized that expression and localization of genes and proteins in the AHR pathway change throughout the reproductive lifespan and in response to Mix exposure. We exposed adult female CD-1 mice to corn oil control or varying doses (0.15, 1.5, or 1,500 ppm) of Mix for 1 to 12 months. Sera and ovaries were collected at each time point. Sera were used to determine circulating E2 concentrations. The AHR pathway in the ovaries was examined using quantitative polymerase chain reaction for gene expression, RNAscope in situ hybridization for transcript localization, and immunohistochemistry for protein localization. The AHR pathway was altered throughout the reproductive lifespan, with increased Cyp1a1 expression and decreased CYP1B1 protein localization in the ovary at 13 months of age. Circulating E2 decreased throughout aging. Mix exposure altered Arnt and Ahr expression after 1 and 12 months, respectively. Additionally, Mix exposure increased CYP1A1 protein localization after 6 months, with areas of increased staining intensity in the ovary. Overall, the AHR pathway was altered with reproductive aging, and Mix exposure affected the pathway over time.

Di(2-ethylhexyl) phthalate (DEHP) and diisononyl phthalate (DiNP) are plasticizers used in many consumer products. Both phthalates alter estrous cyclicity, fertility indices, and folliculogenesis throughout the reproductive lifespan. Our companion paper showed that exposure to a phthalate mixture containing DEHP and DiNP altered the aryl hydrocarbon receptor (AHR) signaling pathway in the ovary. However, limited information was available on how DEHP and DiNP individually affect the AHR signaling pathway in the ovary throughout reproductive aging. Thus, we hypothesized that DEHP and DiNP individually affect the gene and protein expression and localization associated with the AHR pathway throughout the reproductive lifespan. We treated adult female CD-1 mice with corn oil control or doses (0.15 ppm, 1.5 ppm, or 1,500 ppm) of DEHP and DiNP in the diet for 1 to 12 mo. Sera were used to measure circulating estradiol (E2) concentrations. The AHR pathway was assessed using quantitative PCR (qPCR) for gene expression, RNAscope in situ hybridization for transcript localization, and immunohistochemistry for protein localization. Exposure to DEHP, but not DiNP, reduced serum E2 after 1 mo compared with control. DEHP exposure increased Cyp1a1 expression and CYP1B1 localization in the ovary after 6 mo compared with control. DiNP also increased Arnt expression and CYP1B1 localization in the ovary after 1 mo compared with control. DiNP exposure decreased Ahr and Cyp1a1 expression as well as CYP1A1 localization in the antral follicle after 12 mo compared with control. Overall, DEHP and DiNP both influence the ovarian AHR pathway, but at different time points and doses.

Adipose tissue plays a critical role in metabolic and endocrine function because it is essential for maintaining systemic energy homeostasis and other related physiological functions. Mammals have 4 types of adipose tissue: white adipose tissue (WAT), brown adipose tissue (BAT), beige or brite adipose tissue (BeAT), and pink adipose tissue (PAT). These adipose tissues release endocrine factors that modulate diverse processes such as energy storage and expenditure, appetite control, glucose homeostasis, insulin sensitivity, inflammation, lipid metabolism, tissue repair, thermogenesis, and milk production. Proper adipose tissue function relies on hormone receptors and signaling pathways that make the adipose tissues susceptible to disruption by endocrine-disrupting chemicals such as phthalates. Here, we review relevant research on the associations between phthalate exposures and abnormalities in WAT and BAT functions, including phthalate-induced changes in morphology, physiology, and gene expression. This review covers in vitro studies, in vivo studies in mammals, and studies in humans. We also discuss important gaps in the literature. Overall, the evidence indicates that phthalates adversely affect WAT and BAT functions. Further studies are needed to better elucidate the mechanisms through which phthalates act in the adipose tissues and to determine the effects of phthalates on human adipose tissues.

Water disinfection can generate disinfection byproducts (DBPs) such as iodoacetic acid (IAA) by reacting with organic matter in water supplies. In vitro studies have shown that IAA is a cytotoxic and genotoxic DBP. In vivo studies using animal models further indicate that IAA has toxic effects on reproductive function and the endocrine system. Yet, its developmental toxicity remains unknown. In this study, we show that exposure to IAA at concentrations ranging from 1 to 100 µM induces dose-dependent embryonic mortality, with the greatest sensitivity observed during the first 24 hours of development. Sublethal concentrations cause delayed or failed hatching and disrupt the development of multiple vital organs. Notably, IAA exposure impairs swim bladder inflation, reduces otolith size, and decreases heart rate. Moreover, IAA reduces intestinal mucus production by suppressing genes essential for mucin production. Given the crucial role of intestinal mucus in defending against pathogens and maintaining host and gut microbiota homeostasis, the deleterious effect of IAA on mucus production highlights its potential pathogenic role in driving disorders linked to epithelial barrier dysfunction and microbiome dysfunction. Together, these findings demonstrate that IAA induces pronounced developmental toxicity affecting multiple organ systems in zebrafish. This study provides the first in vivo evidence that IAA disrupts embryonic development and intestinal function.

The intrauterine microenvironment is highly susceptible to environmental disturbances, which may result in persistent effects on offspring health. Given that plastic-derived contaminants can cross the placental barrier and act as endocrine-disrupting chemicals, this study evaluated the effects of perinatal exposure to an environmentally relevant phthalate mixture (PM) and nanoplastics (NPs) on the prostate of adult rats. Pregnant Sprague–Dawley rats were allocated into 6 experimental groups: Ctrl (vehicle), T1 (20 μg/kg/d PM), T2 (200 mg/kg/d PM), T3 (NPs: 1.0 mg/kg/d), T4 (20 μg PM + NPs), and T5 (200 mg PM + NPs). Treatments were administered orally from gestational day 10 to postnatal day (PND) 21. Male offspring were euthanized on PND120, and ventral prostate samples were collected. Histological evaluation revealed increased inflammatory foci and stromal expansion in all exposed groups (T1–T5), along with reduced luminal compartment in T1, T3, and T4 compared with controls. All treated groups showed increased total and degranulated mast cells and enhanced androgen receptor immunoreactivity. Tumor necrosis factor (Tnf) expression was increased in all exposure groups, whereas Rela gene expression was elevated in the T4 and T5 groups. Oxidative stress analysis demonstrated increased lipid peroxidation and glutathione S-transferase (GST) activity in all treated animals. Catalase (CAT) activity was reduced in T1, T3, and T5, whereas superoxide dismutase (SOD) activity increased in T3, T4, and T5. Elevated levels of reduced, oxidized, and total glutathione (GSH, GSSG, tGSH) were observed in T5. Overall, perinatal exposure to PM and NPs induced persistent inflammation and altered redox status in the prostate, increasing susceptibility to pathological disorders.

Polystyrene nanoplastics (PS-NPs) are small particles derived from plastic degradation that have been detected in several human tissues. Phthalates are ubiquitous plasticizers used to increase flexibility in polymers which act as endocrine disruptors, impacting hormonal homeostasis. Considering that both pollutants have been detected in human follicular fluid, there is increasing concern regarding their potential effects on female reproductive health. This study evaluated the isolated and combined effects of environmentally relevant doses of PS-NPs and a phthalate metabolite mixture (MM) on antral follicle growth, hormone production, and the expression of genes involved in apoptosis, oxidative stress, steroidogenesis, and hormone receptor signaling. Antral follicles from adult CD-1 mice were cultured with vehicle control (DMSO and water), metabolite mixture (0.01, 0.1, 1, and 10 μg/ml), or PSNPs (5, 25, 50, and 100 μg/ml) or MM + PS-NPs (5 µg/ml PS-NPs + 0.01 µg/ml MM; 100 μg/ml PS-NPs + 10 μg/ml MM). Follicle growth was monitored every 24 h for 96 h. PS-NPs and MM were internalized by follicles and they inhibited follicle growth alone and in co-exposure. Both pollutants altered the expression of apoptosis-related (Casp3, Casp8, Bcl2) and oxidative stress-related (Cat, Nrf2, Gpx1) genes without significantly affecting steroid hormone levels. Co-exposure also reduced Esr2 and Ar expression, demonstrating more pronounced effects under low-dose combined exposure. Altogether, these findings indicate that environmentally relevant exposure to PS-NPs and phthalate mixtures impairs antral follicle growth and disrupts molecular pathways essential for ovarian function, highlighting potential pathways and the importance of understanding combined exposures in reproductive toxicity.

Phthalates are ubiquitous endocrine-disrupting chemicals whose exposure is associated with accelerated reproductive aging in humans. We focused on the pituitary gland, the source of the gonadotropins: Follicle-stimulating hormone (FSH) and luteinizing hormone (LH). We hypothesized that the common phthalates di-(2-ethylhexyl) phthalate (DEHP) and diisononyl phthalate (DiNP) modulate inflammation in the pituitary and impact gonadotropin expression acutely and during aging. To test this, female CD-1 mice were orally dosed with corn oil or varying concentrations of DEHP and DiNP for 10 days. Pituitary tissues were collected immediately after dosing or 15 months post-dosing, processed, and analyzed by quantitative real-time PCR (qPCR) and immunohistochemistry. We found that acute phthalate exposure did not alter Fshb and Lhb mRNA expression compared with controls, but both DEHP and DiNP reduced FSH immunopositive cell number. Phthalate exposure also decreased Il1b, and increased Il18 and Tnf mRNA levels compared with controls, suggesting an inflammatory imbalance. At 15 months post-dosing, DiNP exposure increased Lhb and Il1b mRNA levels, but repressed Fshb and Nlrp3 mRNA levels compared with controls. Next, using dissociated cultures, we investigated the impact of phthalates and the proinflammatory stimulus lipopolysaccharide (LPS) on inflammation and gonadotropin gene expression directly at the pituitary. Both the DEHP metabolite MEHP and LPS decreased Fshb, but not Lhb mRNA relative to control. MEHP also repressed the induction of Il1b by LPS. Together, these findings suggest that acute exposure to phthalate alters mRNA expression of inflammatory markers and gonadotropins in the pituitary, which could alter the process of reproductive aging.

Purpose of Review: Accelerated ovarian aging is associated with early infertility as well as other adverse health outcomes. Little is known about the factors that accelerate ovarian aging, but several studies indicate that exposure to phthalates accelerates ovarian aging. This is a significant human health concern because humans are ubiquitously and unavoidably exposed to phthalates. Thus, it is imperative to study the mechanisms of phthalate-induced accelerated ovarian aging so that strategies can be developed to prevent phthalate-induced ovarian aging. This review focuses on the mechanisms by which phthalates cause ovarian aging in non-human experimental models and highlights gaps in the literature. Recent Findings: Phthalate exposure may accelerate ovarian aging and in turn, accelerate female reproductive aging through several mechanisms. Specifically, phthalates can alter steroidogenesis and folliculogenesis, ultimately dysregulating estrous cyclicity and decreasing fertility. Phthalate-induced disruptions in the brain and gut contribute to these changes. Additionally, phthalate exposures increase ovarian inflammation and oxidative stress, which contribute to accelerated ovarian aging. Phthalate exposure also increases ovarian autophagy, mitochondrial dysfunction, and apoptosis, which ultimately increase follicular atresia and accelerate depletion of the follicle reserve. Summary: Phthalates accelerate ovarian aging through numerous interlinked mechanisms that may be used as targets for prevention, inhibition, or reversal of phthalate-induced ovarian aging in patients experiencing infertility. Further studies should investigate the effects of environmentally relevant phthalate exposures on these mechanisms and explore therapies that target these mechanisms.