17-ß-estradiol and phytoestrogens elicit NO production and vasodilatation through PI3K, PKA and EGF receptors pathways, evidencing functional selectivity.

Endothelial cells express multiple receptors mediating estrogen responses; including the G protein-coupled estrogen receptor (GPER). Past studies on nitric oxide (NO) production elicited by estrogens raised the question whether 17-ß-estradiol (E2) and natural phytoestrogens activate equivalent mechanisms. We hypothesized that E2 and phytoestrogens elicit NO production via coupling to distinct intracellular pathways signalling. To this aim, perfusion of E2 and phytoestrogens to the precontracted rat mesentery bed examined vasorelaxation, while fluorescence microscopy on primary endothelial cells cultures quantified single cell NO production determined following 4-amino-5-methylamino-2',7'-difluoroescein diacetate (DAF) incubation. Daidzein (DAI) and genistein (GEN) induced rapid vasodilatation associated to NO production. Multiple estrogen receptor activity was inferred based on the reduction of DAF-NO signals; G-36 (GPER antagonist) reduced 75 % of all estrogen responses, while fulvestrant (selective nuclear receptor antagonist) reduced significantly more the phytoestrogens responses than E2. The joint application of both antagonists abolished the E2 response but not the phytoestrogen-induced DAF-NO signals. Wortmannin or LY-294002 (PI3K inhibitors), reduced by 90% the E2-evoked signal while altering significantly less the DAI-induced response. In contrast, H-89 (PKA inhibitor), elicited a 23% reduction of the E2-induced signal while blocking 80% of the DAI-induced response. Desmethylxestospongin-B (IP3 receptor antagonist), decreased to equal extent the E2 or the DAI-induced signal. Epidermal growth factor (EGF) induced NO production, cell treatment with AG-1478, an EGF receptor kinase inhibitor reduced 90% DAI-induced response while only 53% the E2-induced signals; highlighting GPER induced EGF receptor trans-modulation. Receptor functional selectivity may explain distinct signalling pathways mediated by E2 and phytoestrogens.

Exploring the multi-targeting phytoestrogen potential of Calycosin for cancer treatment: A review.

Cancer remains a significant challenge in the field of oncology, with the search for novel and effective treatments ongoing. Calycosin (CA), a phytoestrogen derived from traditional Chinese medicine, has garnered attention as a promising candidate. With its high targeting and low toxicity profile, CA has demonstrated medicinal potential across various diseases, including cancers, inflammation, and cardiovascular disease. Studies have revealed that CA possesses inhibitory effects against a diverse array of cancers. The underlying mechanism of action involves a reduction in tumor cell proliferation, induction of tumor cell apoptosis, and suppression of tumor cell migration and invasion. Furthermore, CA has been shown to enhance the efficacy of certain chemotherapeutic drugs, making it a potential component in treating malignant tumors. Given its high efficacy, low toxicity, and multi-targeting characteristics, CA holds considerable promise as a therapeutic agent for cancer treatment. The objective of this review is to present a synthesis of the current understanding of the antitumor mechanism of CA and its research progress.

Comparison of in vitro Toxicities of 8-Prenylnaringenin, Tartrazine and 17ß-Estradiol, Representatives of Natural and Synthetic Estrogens, in Rat and Human Hepatoma Cell Lines.

BACKGROUND: Phytoestrogens have been praised for their beneficial health effects, whereas synthetic xenoestrogens have been connected to ailments. AIMS: To ascertain whether the toxicities of natural and synthetic estrogens differ, we examined the potent phytoestrogen 8-prenylnaringenin (8-PN), the common synthetic xenoestrogen tartrazine, and the physiological estrogen 17ß-estradiol (E2). METHODS: These three compounds were tested for cytotoxicity, cell proliferation and genotoxicity in human HepG2 and rat H4IIE hepatoma cells. RESULTS: All three estrogens elicited cytotoxicity at high concentrations in both cell lines. They also inhibited cell proliferation, with E2 being the most effective. They all tended to increase micronuclei formation. CONCLUSION: Natural estrogens were no less toxic than a synthetic one.

Sex-specific pharmacokinetic response to phytoestrogens in Drosophila melanogaster.

Drosophila melanogaster, or the fruit fly, is widely used for modeling numerous human diseases, such as neurodegeneration, tumor development, cachexia, and intestinal dysfunction. It is a suitable model organism for research targeting the physiology and pathophysiology of the intestinal epithelial barrier and has also been used as a model organism for preliminary drug and bioactive nutrient screening. However, the application of D. melanogaster in research on drug bioavailability and pharmacokinetic properties has not yet been well explored. In this study, we applied D. melanogaster to investigate the absorption and excretion of the orally administered phytoestrogens daidzein, glycitein, genistein, and their glycosides. Therefore, we established a quick, noninvasive method to quantify compound retention in D. melanogaster, suitable for the investigation of a broad variety of potentially bioactive substances. We showed that fruit fly sex plays a key role in the metabolization, transportation, and excretion of phytoestrogenic isoflavones. In particular, female fruit flies retained significantly more isoflavones than male fruit flies, which was reflected in the greater metabolic impact of isoflavones on females. Male fruit flies excreted more isoflavones than females did, which was linked to the upregulation of the xenobiotic transporter gene Mdr50. We also demonstrated that micellized isoflavones were more bioavailable than powdered isoflavones, independent of sex, age or the addition of dietary fibers.

The effect of soy isoflavones in brain development: the emerging role of multiple signaling pathways and future perspectives.

Soybean is a source of protein, fibers, and phytochemical isoflavones which are considered to have numerous health benefits for children and adulthood. On the other hand, isoflavones are widely known as phytoestrogens that exert their action via the estrogen signaling pathway. With this regard, isoflavones are also considered as endocrine-disrupting chemicals. Endogenous estrogen plays a crucial role in brain development through binding to estrogen receptors (ERs) or G protein-coupled estrogen receptors 1 (GPER1) and regulates morphogenesis, migration, functional maturation, and intracellular metabolism of neurons and glial cells. Soy isoflavones can also bind to ERs, GPER1, and, furthermore, other receptors to modulate their action. Therefore, soy isoflavone consumption may affect brain development during the pre-and post-natal periods. This review summarizes the current knowledge on the mechanisms of isoflavone action, particularly in the early stages of brain development by introducing representative human, and animal models, and in vitro studies, and discusses their beneficial and adverse impact on neurobehavior. As a conclusion, the soy product consumption during the pre-and post-natal periods under proper range of dose showed beneficial effects in neurobehavior development, including improvement of anxiety, aggression, hyperactive behavior, and cognition, whereas their adverse effect by taking higher doses cannot be excluded. We also present novel research lines to further assess the effect of soy isoflavone administration during brain development.

Modulatory effects of estrous cycle on ingestive behaviors and energy balance in young adult C57BL/6J mice maintained on a phytoestrogen-free diet.

The estrous cycle is known to modify food, fluid, and electrolyte intake behaviors and energy homeostasis in various species, in part through fluctuations in estrogen levels. Simultaneously, commonly commercially available rodent dietary formulations greatly vary in soy protein content, and thereby the delivery of biologically active phytoestrogens. To explore the interactions among the estrous cycle, sodium, fluid, and caloric seeking behaviors, and energy homeostasis, young adult C57BL/6J female mice were maintained on a soy protein-free 2920x diet and provided water, or a choice between water and 0.15 mol/L NaCl drink solution. Comprehensive metabolic phenotyping was performed using a multiplexed Promethion (Sable Systems International) system, and estrous stages were determined via daily vaginal cytology. When provided food and water, estrous cycling had no major modulatory effects on intake behaviors or energy balance. When provided a saline solution drink choice, significant modulatory effects of the transition from diestrus to proestrus were observed upon fluid intake patterning, locomotion, and total energy expenditure. Access to saline increased total daily sodium consumption and aspects of energy expenditure, but these effects were not modified by the estrous stage. Collectively, these results indicate that when supplied a phytoestrogen-free diet, the estrous cycle has minor modulatory effects on ingestive behaviors and energy balance in C57BL/6J mice that are sensitive to sodium supply.NEW & NOTEWORTHY When provided a phytoestrogen-free diet, the estrous cycle had very little effect on food and water intake, physical activity, or energy expenditure in C57BL/6J mice. In contrast, when provided an NaCl drink in addition to food and water, the estrous cycle was associated with changes in intake behaviors and energy expenditure. These findings highlight the complex interactions among estrous cycling, dietary formulation, and nutrient presentation upon ingestive behaviors and energy homeostasis in mice.

Estrogen receptor-mediated health benefits of phytochemicals: a review.

Estrogen receptors (ERs) are transcription factors with two subtypes: estrogen receptor alpha (ERα) and estrogen receptor beta (ERß), which are essential for the maintenance of human health and play a regulatory role in common diseases such as breast cancer, osteoporosis, neurodegenerative disorders, liver injuries and lung cancers. A number of phytochemicals extracted from various fruits and vegetables have been demonstrated to exhibit estrogenic effects and are termed phytoestrogens. As modulators of ERs, phytoestrogens can be involved in the prevention and treatment of multiple diseases as complementary or alternative therapeutic agents and have a variety of health benefits for humans. This article reviews the health benefits of phytoestrogens in clinical and epidemiologic studies for several diseases and also provides a detailed description of the molecular mechanisms of their action. A brief comparison of the advantages and disadvantages of natural phytochemicals compared to synthetic drugs is also presented. The role of phytoestrogens in the treatment of diseases and human health requires further research to fully realize their therapeutic potential.

Effects of Isoflavonoid and Vitamin D Synergism on Bone Mineral Density-A Systematic and Critical Review.

Phytoestrogens are non-steroidal plant compounds, which bind to α and ß estrogen receptors, thereby causing specific effects. The best-known group of phytoestrogens are flavonoids, including isoflavonoids-genistein and daidzein. They play a role in the metabolism of bone tissue, improving its density and preventing bone loss, which contributes to reducing the risk of fractures. Vitamin D is found in the form of cholecalciferol (vitamin D3) and ergocalciferol (vitamin D2) and is traditionally recognized as a regulator of bone metabolism. The aim of this review was to evaluate the synergistic effect of isoflavonoids and vitamin D on bone mineral density (BMD). The MEDLINE (PubMed), Scopus and Cochrane databases were searched independently by two authors. The search strategy included controlled vocabulary and keywords. Reference publications did not provide consistent data regarding the synergistic effect of isoflavonoids on BMD. Some studies demonstrated a positive synergistic effect of these compounds, whereas in others, the authors did not observe any significant differences. Therefore, further research on the synergism of isoflavonoids and vitamin D may contribute to a significant progress in the prevention and treatment of osteoporosis.

The Role of Genistein in Mammalian Reproduction.

Genistein is a natural compound belonging to flavonoids, having antioxidant, anti-inflammatory, and anti-neoplastic properties. Genistein is considered a phytoestrogen. As such, genistein can bind estrogen receptors (ERα and ERß), although with a lower affinity than that of estradiol. Despite considerable work, the effects of genistein are not well established yet. This review aims to clarify the role of genistein on female and male reproductive functions in mammals. In females, at a high dose, genistein diminishes the ovarian activity regulating several pathway molecules, such as topoisomerase isoform I and II, protein tyrosine kinases (v-src, Mek-4, ABL, PKC, Syk, EGFR, FGFR), ABC, CFTR, Glut1, Glut4, 5α-reductase, PPAR-γ, mitogen-activated protein kinase A, protein histidine kinase, and recently circulating RNA-miRNA. The effect of genistein on pregnancy is still controversial. In males, genistein exerts an estrogenic effect by inducing testosterone biosynthesis. The interaction of genistein with both natural and synthetic endocrine disruptors has a negative effect on testis function. The positive effect of genistein on sperm quality is still in debate. In conclusion, genistein has a potentially beneficial effect on the mechanisms regulating the reproduction of females and males. However, this is dependent on the dose, the species, the route, and the time of administration.

Multi-Anticancer Activities of Phytoestrogens in Human Osteosarcoma.

Phytoestrogens are plant-derived bioactive compounds with estrogen-like properties. Their potential health benefits, especially in cancer prevention and treatment, have been a subject of considerable research in the past decade. Phytoestrogens exert their effects, at least in part, through interactions with estrogen receptors (ERs), mimicking or inhibiting the actions of natural estrogens. Recently, there has been growing interest in exploring the impact of phytoestrogens on osteosarcoma (OS), a type of bone malignancy that primarily affects children and young adults and is currently presenting limited treatment options. Considering the critical role of the estrogen/ERs axis in bone development and growth, the modulation of ERs has emerged as a highly promising approach in the treatment of OS. This review provides an extensive overview of current literature on the effects of phytoestrogens on human OS models. It delves into the multiple mechanisms through which these molecules regulate the cell cycle, apoptosis, and key pathways implicated in the growth and progression of OS, including ER signaling. Moreover, potential interactions between phytoestrogens and conventional chemotherapy agents commonly used in OS treatment will be examined. Understanding the impact of these compounds in OS holds great promise for developing novel therapeutic approaches that can augment current OS treatment modalities.

Investigations of Electronic, Structural, and In Silico Anticancer Potential of Persuasive Phytoestrogenic Isoflavene-Based Mannich Bases.

Isoflavenes have received the greatest research attention among the many groups of phytoestrogens. In this study, various isoflavene-based Mannich bases were selected for their theoretical studies. The purpose of this research was to discover the binding potential of all the designated Mannich bases acting as inhibitors against cancerous proteins EGFR, cMet, hTrkA, and HER2 (PDB codes: 5GTY, 3RHK, 6PL2, and 7JXH, respectively). For their virtual screening, DFT calculations and molecular docking studies were undertaken using in silico software. Docking studies predicted that ligands 5 and 15 exhibited the highest docking score by forming hydrogen bonds within the active pocket of protein 6PL2, ligands 1 and 15 both with protein 3RHK, and 7JXH, 12, and 17 with protein 5GTY. Rendering to the trends in polarizability and dipole moment, the energy gap values (0.2175 eV, 0.2106 eV) for the firm conformers of Mannich bases (1 and 4) replicate the increase in bioactivity and chemical reactivity. The energy gap values (0.2214 eV and 0.2172 eV) of benzoxazine-substituted isoflavene-based Mannich bases (9 and 10) reflect the increase in chemical potential due to the most stable conformational arrangements. The energy gap values (0.2188 eV and 0.2181 eV) of isoflavenes with tertiary amine-based Mannich bases (14 and 17) reflect the increase in chemical reactivity and bioactivity due to the most stable conformational arrangements. ADME was also employed to explore the pharmacokinetic properties of targeted moieties. This study revealed that these ligands have a strong potential to be used as drugs for cancer treatment.

Icariin, a phytoestrogen, exerts rapid estrogenic actions through crosstalk of estrogen receptors in osteoblasts.

Icariin, a flavonoid glycoside derived from Epimedium brevicornum Maxim, exerts bone protective effects via estrogen receptors (ERs). This study aimed to investigate the role of ER-α66, ER-α36, and GPER in bone metabolism in osteoblasts following treatment with icariin. Human osteoblastic MG-63 cells and osteoblast-specific ER-α66 knockout mice were employed. The ERs crosstalk in the estrogenic action of icariin was evaluated in ER-α66-negative human embryonic kidney HEK293 cells. Icariin, like E2, regulated ER-α36 and GPER protein expression in osteoblasts by downregulating them and upregulating ER-α66. ER-α36 and GPER suppressed the actions of icariin and E2 in bone metabolism. However, the in vivo administration of E2 (2 mg/kg/day) or icariin (300 mg/kg/day) restored bone conditions in KO osteoblasts. ER-α36 and GPER expression increased significantly and rapidly activated and translocated in KO osteoblasts after treatment with E2 or icariin. ER-α36 overexpression in KO osteoblasts further promoted the OPG/RANKL ratio induced by E2 or icariin treatment. This study showed icariin and E2 elicit rapid estrogenic responses in bone through recruiting ER-α66, ER-α36, and GPER. Notably, in osteoblasts lacking ER-α66, ER-α36, and GPER mediate the estrogenic effects of icariin and E2, while in intact osteoblasts, ER-α36 and GPER act as negative regulators of ER-α66.

Rutin Promotes Proliferation and Orchestrates Epithelial-Mesenchymal Transition and Angiogenesis in MCF-7 and MDA-MB-231 Breast Cancer Cells.

Rutin has been reported as a potential anti-cancer agent for several decades. This study evaluated the effects of rutin on the proliferation, metastasis, and angiogenesis of MDA-MB-231 and MCF-7 breast cancer cell lines. Increasing concentrations of rutin significantly stimulated the proliferation of MDA-MB-231 and MCF-7 cells compared to controls. Wound scratch assay demonstrated that rutin had an inducing effect on the migration of the cells. In MDA-MB-231 and MCF-7 cells, rutin upregulated MKI67, VIM, CDH2, FN1, and VEGFA and downregulated CDH1 and THBS1 genes. It also increased N-cadherin and VEGFA and decreased E-cadherin and thrombospondin 1 protein expression. Our data indicated that rutin could stimulate proliferation, migration, and pro-angiogenic activity in two different breast cancer cell lines. This phytoestrogen induced invasion and migration of both cell lines by a mechanism involving the EMT process. This suggests that rutin may act as a breast-cancer-promoting phytoestrogen.

Genistein: a promising modulator of apoptosis and survival signaling in cancer.

Genistein, a commonly occurring isoflavone, has recently gained popularity owing to its ever-expanding spectrum of pharmacological benefits. In addition to health benefits such as improved bone health and reduced postmenopausal complications owing to its phytoestrogen properties, it has been widely evaluated for its anti-cancer potential. Several studies have established the potential for its usage in the management of breast, lung, and prostate cancers, and its usage has significantly evolved from early applications in traditional systems of medicine. This review offers an insight into its current status of usage, the chemistry, and pharmacokinetics of the molecule, an exploration of its apoptotic mechanisms in cancer management, and opportunities for synergism to improve therapeutic outcomes. In addition to this, the authors have presented an overview of recent clinical trials, to offer an understanding of contemporary studies and explore prospects for a greater number of focused trials, moving forward. Advancements in the application of nanotechnology as a strategy to improve safety and efficacy have also been highlighted, with a brief discussion of results from safety and toxicology studies.

The Phytoestrogenic Effect of Hibiscus sabdariffa Involves Estrogen Receptor α in Ovariectomized Wistar Rats.

The calyxes of Hibiscus sabdariffa present multiple pharmacological effects primarily attributed to their high anthocyanin content; however, little is known about their phytoestrogenic effect. Ovarian hypofunction (OH) is a process characterized by the rapid detention of the production of ovarian hormones, which compromises reproductive and cognitive functions. Hormone replacement therapy (HRT) efficiently compensates for OH; nevertheless, questions have been raised on its secondary effects and safety. One of the alternatives to tackling OH involves using phytoestrogens such as anthocyanins for their structural similarity to natural estrogens. In a Wistar rat model of ovariectomy (OVX), we recently reported the beneficial properties of an anthocyanin-rich extract prepared from the calyces of H. sabdariffa (HSE) in hindering the adverse effects of OH on memory performance and highlighted a possible phytoestrogenic impact through the modulation of estrogen receptor (ER) expression. We now report that HSE and estradiol differentially affected the expression of ERα and ERß. ERα was more sensitive to HSE; meanwhile, estradiol preferentially modulated ERß. Thus, our study leads to further research on using H. sabdariffa as a nutrition-based alternative to HRT.

A review on phytoestrogens: Current status and future direction.

Phytoestrogens are plant secondary metabolite that is structurally and functionally similar to mammalian estrogens, which have been shown to have various health benefits in humans. Isoflavones, coumestans, and lignans are the three major bioactive classes of phytoestrogens. It has a complicated mechanism of action involving an interaction with the nuclear estrogen receptor isoforms ERα and ERß, with estrogen agonist and estrogen antagonist effects. Depending on their concentration and bioavailability in various plant sources, phytoestrogens can act as estrogen agonist or antagonists. Menopausal vasomotor symptoms, breast cancer, cardiovascular disease, prostate cancer, menopausal symptoms, and osteoporosis/bone health have all been studied using phytoestrogens as an additional standard hormone supplemental remedy. The botanical sources, techniques of identification, classification, side effects, clinical implications, pharmacological and therapeutic effects of their proposed mode of action, safety issues, and future directions for phytoestrogens have all been highlighted in this review.

Structural mechanism of human oncochannel TRPV6 inhibition by the natural phytoestrogen genistein.

Calcium-selective oncochannel TRPV6 is the major driver of cell proliferation in human cancers. While significant effort has been invested in the development of synthetic TRPV6 inhibitors, natural channel blockers have been largely neglected. Here we report the structure of human TRPV6 in complex with the plant-derived phytoestrogen genistein, extracted from Styphnolobium japonicum, that was shown to inhibit cell invasion and metastasis in cancer clinical trials. Despite the pharmacological value, the molecular mechanism of TRPV6 inhibition by genistein has remained enigmatic. We use cryo-EM combined with electrophysiology, calcium imaging, mutagenesis, and molecular dynamics simulations to show that genistein binds in the intracellular half of the TRPV6 pore and acts as an ion channel blocker and gating modifier. Genistein binding to the open channel causes pore closure and a two-fold symmetrical conformational rearrangement in the S4-S5 and S6-TRP helix regions. The unprecedented mechanism of TRPV6 inhibition by genistein uncovers new possibilities in structure-based drug design.

Molecular mechanisms underlying the potential neuroprotective effects of Trifolium pratense and its phytoestrogen-isoflavones in neurodegenerative disorders.

Neurodegenerative disorders are heterogeneous, debilitating, and incurable groups of brain disorders that have common features including progressive degeneration of the structure and function of the nervous system. Phytoestogenic-isoflavones have been identified as active compounds that can modulate different molecular signaling pathways related to the nervous system. The main aim is to shed the light on the molecular mechanisms followed by phytoestrogen-isoflavones profound in the Trifolium pratense and discuss the latest pharmacological findings in the treatment of neurodegenerative disorders. Data were collected using different databases. The search terms used included "Phytoestrogens," "Isoflavones," "neurodegenerative disorders," "Neuronal plasticity," etc., and combinations of these keywords. As a result, this review article mainly demonstrates the potential neuroprotective properties of phystoestrogen-isoflavones present in the Trifolium pratense (Red clover), particularly in neurodegenerative disorders. Phytochemical studies have shown that Trifolium pratense mainly includes more than 30 isoflavone compounds. Among them, phytoestrogen-isoflavones, such as biochanin A, daidzein, formononetin, genistein (Gen), etc.,are characterized by potent neuroprotective properties against different neurodegenerative disorders. There are preclinical and clinical scientific evidence on their mechanisms of action involve molecular interaction with estrogenic receptors, anti-inflammatory, anti-oxidative, antiapoptotic, autophagic inducing, and so on. phytoestrogen-isoflavones are the major bioactive components in the Trifolium pratense that exhibit therapeutic efficacy in the case of neurodegenerative disorders. This review provides detailed molecular mechanisms targeted by phytoestrogen-isoflavones and experimental key findings for the clinical use of prescriptions containing Trifolium pratense-derived isoflavones for the treatment of neurodegenerative disorders.

Soy and Gastrointestinal Health: A Review.

Soybean is the most economically important legume globally, providing a major source of plant protein for millions of people; it offers a high-quality, cost-competitive and versatile base-protein ingredient for plant-based meat alternatives. The health benefits of soybean and its constituents have largely been attributed to the actions of phytoestrogens, which are present at high levels. Additionally, consumption of soy-based foods may also modulate gastrointestinal (GI) health, in particular colorectal cancer risk, via effects on the composition and metabolic activity of the GI microbiome. The aim of this narrative review was to critically evaluate the emerging evidence from clinical trials, observational studies and animal trials relating to the effects of consuming soybeans, soy-based products and the key constituents of soybeans (isoflavones, soy proteins and oligosaccharides) on measures of GI health. Our review suggests that there are consistent favourable changes in measures of GI health for some soy foods, such as fermented rather than unfermented soy milk, and for those individuals with a microbiome that can metabolise equol. However, as consumption of foods containing soy protein isolates and textured soy proteins increases, further clinical evidence is needed to understand whether these foods elicit similar or additional functional effects on GI health.

Coumestrol induces oxidative stress and impairs migration and embryonic growth.

In brief: Healthy development of the placenta is dependent on trophoblast cell migration and reduced oxidative stress presence. This article describes how a phytoestrogen found in spinach and soy causes impaired placental development during pregnancy. Abstract: Although vegetarianism has grown in popularity, especially among pregnant women, the effects of phytoestrogens in placentation lack understanding. Factors such as cellular oxidative stress and hypoxia and external factors including cigarette smoke, phytoestrogens, and dietary supplements can regulate placental development. The isoflavone phytoestrogen coumestrol was identified in spinach and soy and was found to not cross the fetal-placental barrier. Since coumestrol could be a valuable supplement or potent toxin during pregnancy, we sought to examine its role in trophoblast cell function and placentation in murine pregnancy. After treating trophoblast cells (HTR8/SVneo) with coumestrol and performing an RNA microarray, we determined 3079 genes were significantly changed with the top differentially changed pathways related to the oxidative stress response, cell cycle regulation, cell migration, and angiogenesis. Upon treatment with coumestrol, trophoblast cells exhibited reduced migration and proliferation. Additionally, we observed increased reactive oxygen species accumulation with coumestrol administration. We then examined the role of coumestrol within an in vivo pregnancy by treating wildtype pregnant mice with coumestrol or vehicle from day 0 to 12.5 of gestation. Upon euthanasia, fetal and placental weights were significantly decreased in coumestrol-treated animals with the placenta exhibiting a proportional decrease with no obvious changes in morphology. Therefore, we conclude that coumestrol impairs trophoblast cell migration and proliferation, causes accumulation of reactive oxygen species, and reduces fetal and placental weights in murine pregnancy.