phytoestrogens and daidzein

Daidzein, 7-hydroxy-3-(4-hydroxyphenyl)-4H-chromen-4-one is a naturally occurring compound present in leguminous plants, especially in soybeans. Chemically it belongs to the isoflavone class and possesses high nutritive value. Daidzein acts on estrogen receptor and is non-steroidal in nature hence it can also be called as non-steroidal phytoestrogenic compound. Daidzein has been studied by many researchers for its pharmacological activities. Daidzein metabolites were also studied in detail for their health benefits. Researchers have developed novel formulations of daidzein in the past few years to improve its aqueous solubility and bioavailability. Self-emulsified daidzein, poly(lactic-co-glycolic) acid daidzein nanoparticles, nanoemulsion, nanoemulsion gel, and co-crystals are a few of them. The present review provides detailed information on the chemistry, drug development aspects, pharmacokinetics, and pharmacodynamics of daidzein. A literature search was performed using various datasets like PubMed, EBSCO, ProQuest Scopus, and selected websites including the National Institutes of Health and the World Health Organization. Daidzein has a wide range of pharmacodynamic properties in the treatment of cancer, neurodegenerative disorders, cardiac disorders, diabetes and its complication, osteoporosis, and skin disorders. The pharmacokinetic, pharmacodynamics, and drug development aspects of daidzein will help researchers to design further research work on daidzein in the future.

Topics: Biological Availability; Glycine max; Isoflavones; Phytoestrogens

Phytoestrogens are naturally occurring nonsteroidal phenolic plant compounds that, due to their molecular structure and size, resemble vertebrate steroids estrogens. This review is focused on plant flavonoids isoflavones, which are ranked among the most estrogenic compounds. The main dietary sources of isoflavones for humans are soybean and soybean products, which contain mainly daidzein and genistein. When they are consumed, they exert estrogenic and/or antiestrogenic effects. Isoflavones are considered chemoprotective and can be used as an alternative therapy for a wide range of hormonal disorders, including several cancer types, namely breast cancer and prostate cancer, cardiovascular diseases, osteoporosis, or menopausal symptoms. On the other hand, isoflavones may also be considered endocrine disruptors with possible negative influences on the state of health in a certain part of the population or on the environment. This review deals with isoflavone classification, structure, and occurrence, with their metabolism, biological, and health effects in humans and animals, and with their utilization and potential risks.

Topics: Animals; Equol; Genistein; Humans; Isoflavones; Phytoestrogens

Phytoestrogens may influence prostate cancer development. This study aimed to examine the association between prediagnostic circulating concentrations of isoflavones (genistein, daidzein, equol) and lignans (enterolactone and enterodiol) and the risk of prostate cancer. Individual participant data were available from seven prospective studies (two studies from Japan with 241 cases and 503 controls and five studies from Europe with 2,828 cases and 5,593 controls). Because of the large difference in circulating isoflavone concentrations between Japan and Europe, analyses of the associations of isoflavone concentrations and prostate cancer risk were evaluated separately. Prostate cancer risk by study-specific fourths of circulating concentrations of each phytoestrogen was estimated using multivariable-adjusted conditional logistic regression. In men from Japan, those with high compared to low circulating equol concentrations had a lower risk of prostate cancer (multivariable-adjusted OR for upper quartile [Q4] vs. Q1 = 0.61, 95% confidence interval [CI] = 0.39-0.97), although there was no significant trend (OR per 75 percentile increase = 0.69, 95 CI = 0.46-1.05, p

Topics: Aged; Case-Control Studies; Equol; Europe; Genistein; Humans; Isoflavones; Japan; Lignans; Male; Middle Aged; Phytoestrogens; Prospective Studies; Prostatic Neoplasms; Risk Factors

Isoflavones (genistein, daidzein, and glycitein) are bioactive compounds with mildly estrogenic properties and often referred to as phytoestrogen. These are present in significant quantities (up to 4-5 mg·g

Topics: Biological Availability; Diet; Fermentation; Food Analysis; Food Handling; Food Irradiation; Food Storage; Genistein; Hot Temperature; Humans; Infant Food; Isoflavones; Nutritive Value; Phytoestrogens; Randomized Controlled Trials as Topic; Soy Foods

This updated meta-analysis was performed to clarify the relationship between phytoestrogens and prostate cancer risk. Twenty one case-control and two cohort studies were finally selected for this meta-analysis, totaling 11,346 cases and 140,177 controls. Analytical results showed that daidzein (OR = 0.85; 95% CI: 0.75-0.96), genistein (OR = 0.87; 95% CI: 0.78-0.98), and glycitein (OR = 0.89; 95% CI: 0.81-0.98) were associated with a reduction of prostate cancer risk, but total isoflavones (OR = 0.93; 95% CI: 0.84-1.04), equol (OR = 0.86; 95% CI: 0.66-1.14), total lignans (OROgna.05; 95% CI: 0.54-2.04), secoisolariciresinol (OR = 1.02; 95% CI: 0.83-1.24), matairesinol (OR = 0.91; 95% CI: 0.75-1.11), enterolactone (OR = 0.94; 95% CI: 0.73-1.20), and coumestrol (OR = 0.89; 95% CI: 0.76-1.06) were not. Sensitivity and publication bias analyses demonstrated that the pooled estimates were stable and reliable. The results support the notion that some phytoestrogens may have a role in decreasing the risk of prostate cancer. Additional large and well-designed cohort studies are needed to confirm these relationships.

Topics: Case-Control Studies; Cohort Studies; Diet, Healthy; Evidence-Based Medicine; Genistein; Humans; Isoflavones; Male; Men's Health; Phytoestrogens; Prostatic Neoplasms; Risk

Menopause is associated with increased bone resorption and decreased bone mineral density (BMD). Phytoestrogens are believed to prevent bone loss. This study reviewed relevant randomized, controlled trials to determine the effects of phytoestrogens on BMD in postmenopausal women.. In order to perform this systematic review, PubMed, Science Direct, Scopus, Cochrane Library, ISI Web of Knowledge, and ProQuest databases were searched for articles published during 2005-2016. The main keywords used during the searches were "phytoestrogen" and "bone mineral density" and "menopause". The Cochrane Risk of Bias Assessment Tool was used to evaluate the quality of the selected studies and to assess the risk of bias.. A total of 23 eligible studies were included in this systematic review. Most selected studies used a double-blind, placebo-controlled design. In total, 3494 participants were enrolled in the selected trials. Different types of soy isoflavone extracts, including genistein extracts (either alone or in combination with daidzein), dietary products containing different amounts of phytoestrogens, and red clover extracts were used in the designed interventions. The duration of the interventions ranged from 7 weeks to 3 years. In most studies, the primary outcome was the efficacy of the designed intervention which was assessed through measuring whole body or regional BMD or bone mineral content, T-scores, and biomarkers of bone metabolism.. Isoflavones probably have beneficial effects on bone health in menopausal women. Nevertheless, there were controversial reports about changes in BMD. Supplementation with a phytoestrogen can probably prevent the reduction in BMD and maintain a healthy bone structure during menopause.

Topics: Bone Density; Diet; Double-Blind Method; Female; Genistein; Glycine max; Humans; Isoflavones; Menopause; Phytoestrogens; Phytotherapy; Placebos; Plant Extracts; Randomized Controlled Trials as Topic

The study aimed to examine the association between genes encoding molecules in the ornithine decarboxylase (ODC)-polyamine pathway (ODC1, AMD1, NQO1, NOS2A, and OAZ2) and gastric cancer risk and whether the gene-phytoestrogen interaction modifies gastric cancer risk.. Among 76 gastric cancer cases and their 1:4 matched controls within the Korean Multi-center Cancer Cohort, a total of 30 SNPs in five genes involved in the ODC pathway were primarily analyzed. The second-stage genotyping in 388 matched case-control sets was conducted to reevaluate the significant SNPs interacting with phytoestrogens during the primary analysis. The summary odds ratios (ORs) [95 % confidence intervals (CIs)] for gastric cancer were estimated. Interaction effects between the SNPs and plasma concentrations of phytoestrogens (genistein, daidzein, equol, and enterolactone) were evaluated.. In the pooled analysis, NQO1 rs1800566 showed significant genetic effects on gastric cancer without heterogeneity [OR 0.83 (95 % CI 0.70-0.995)] and a greater decreased risk at high genistein/daidzein levels [OR 0.36 (95 % CI 0.15-0.90) and OR 0.26 (95 % CI 0.10-0.64), respectively; p interaction < 0.05]. Risk alleles of AMD1 rs1279599, AMD1 rs7768897, and OAZ2 rs7403751 had a significant gene-phytoestrogen (genistein and daidzein) interaction effect to modify the development of gastric cancer. They had an increased gastric cancer risk at low isoflavone levels, but a decreased risk at high isoflavone levels (p interaction < 0.01).. Our findings suggest that common variants in the genes involved in the ODC pathway may contribute to the risk of gastric cancer possibly by modulating ODC polyamine biosynthesis or by interaction between isoflavones and NQO1, OAZ2, and AMD1.

Topics: 4-Butyrolactone; Adenosylmethionine Decarboxylase; Asian People; Case-Control Studies; Equol; Gene-Environment Interaction; Genistein; Humans; Isoflavones; Lignans; Multicenter Studies as Topic; NAD(P)H Dehydrogenase (Quinone); Nitric Oxide Synthase Type II; Ornithine Decarboxylase; Phytoestrogens; Polyamines; Polymorphism, Single Nucleotide; Stomach Neoplasms

With 13 million new cases worldwide every year, prostate cancer is as a very real public health concern. Prostate cancer is common in over-50s men and the sixth-leading cause of cancer-related death in men worldwide. Like all cancers, prostate cancer is multifactorial - there are non-modifiable risk factors like heredity, ethnicity and geographic location, but also modifiable risk factors such as diet. Diet-cancer linkages have risen to prominence in the last few years, with accruing epidemiological data pointing to between-population incidence differentials in numerous cancers. Indeed, there are correlations between fat-rich diet and risk of hormone-dependent cancers like prostate cancer and breast cancer. Diet is a risk factor for prostate cancer, but certain micronutrients in specific diets are considered protective factors against prostate cancer. Examples include tomato lycopene, green tea epigallocatechin gallate, and soy phytoestrogens. These micronutrients are thought to exert cancer-protective effects via anti-oxidant pathways and inhibition of cell proliferation. Here, we focus in on the effects of phytoestrogens, and chiefly genistein and daidzein, which are the best-researched to date. Soy phytoestrogens are nonsteroid molecules whose structural similarity lends them the ability to mimic the effects of 17ß-estradiol. On top of anti-oxidant effects, there is evidence that soy phytoestrogens can modulate the epigenetic modifications found in prostate cancer. We also studied the impact of phytoestrogens on epigenetic modifications in prostate cancer, with special focus on DNA methylation, miRNA-mediated regulation and histone modifications.

Topics: Adult; Antioxidants; Diet; DNA Methylation; Epigenesis, Genetic; Genistein; Glycine max; Histone Code; Humans; Isoflavones; Male; MicroRNAs; Middle Aged; Phytoestrogens; Prostatic Neoplasms; Risk Factors

Phytoestrogens are phenolic compounds derived from plants and exert an estrogenic as well as an antiestrogenic effect and also various biological efficacies. Chemopreventive properties of phytoestrogens has emerged from epidemiological observations indicating that the incidence of some cancers including breast and prostate cancers is much lower in Asian people, who consume significantly higher amounts of phytoestrogens than Western people. There are 4 main classes of phytoestrogens: isoflavones, stilbenes, coumestans, and lignans. Currently, resveratrol is recognized as another major phytoestrogen present in grape and red wine and has been studied in many biological studies. Phytoestrogens have biologically diverse profitabilities and advantages such as low cytotoxicity to patients, lack of side effects in clinical trials, and pronounced benefits in a combined therapy. In this review, we highlighted the effects of genistein, daidzein, and resveratrol in relation with their anticarcinogenic activity. A lot of in vitro and in vivo results on their chemopreventive properties were presented along with the underlying mechanisms. Besides well-known mechanisms such as antioxidant property and apoptosis, newly elucidated anticarcinogenic modes of action including epigenetic modifications and topoisomerase inhibition have been provided to examine the possibility of phytoestrogens as promising reagents for cancer chemoprevention and/or treatment and to suggest the importance of plant-based diet of phytoestrogens.

Topics: Antioxidants; Chemoprevention; Genistein; Humans; Isoflavones; Phytoestrogens; Resveratrol; Stilbenes

Isoflavones are phytoestrogens that have been linked to both beneficial as well as adverse effects in relation to cell proliferation and cancer risks. The present article presents an overview of these seemingly contradicting health effects and of mechanisms that could be involved in this dualistic mode of action. One mechanism relates to the different ultimate cellular effects of activation of estrogen receptor (ER) α, promoting cell proliferation, and of ERβ, promoting apoptosis, with the major soy isoflavones genistein and daidzein activating especially ERβ. A second mode of action includes the role of epigenetics, including effects of isoflavones on DNA methylation, histone modification and miRNA expression patterns. The overview presented reveals that we are only at the start of unraveling the complex underlying mode of action for effects of isoflavones, both beneficial or adverse, on cell proliferation and cancer risks. It is evident that whatever model system will be applied, its relevance to human tissues with respect to ERα and ERβ levels, co-repressor and co-activator characteristics as well as its relevance to human exposure regimens, needs to be considered and defined.

Topics: Animals; Apoptosis; Cell Proliferation; Co-Repressor Proteins; DNA Methylation; Estrogen Receptor alpha; Estrogen Receptor beta; Genistein; Glycine max; Histones; Humans; Isoflavones; MicroRNAs; Neoplasms; Phytoestrogens; Plant Extracts; Risk Factors

Diet is believed to play an important role in cancer. It has been revealed by epidemiological studies that Asian populations, who consume phytoestrogens in large amounts, have a lower incidence of prostate cancer in comparison with the Western world, where consumption of soy is lower. Genistein and daidzein, the soy phytoestrogens most widely studied, are believed to be potent anticancer agents and have been shown to possess anticancer properties. It has been shown that these compounds inhibit the growth of cancer cells through the modulation of genes controlling cell-cycle progression. Genistein inhibits the activation of the kappa light polypeptide gene enhancer in B-cells (NF-κB), signaling pathway, which is implicated in the balance between cell survival and programmed cell death (apoptosis). Antioxidant and antiangiogenesis properties of genistein have been also described. Soy isoflavones are also implicated in reversion of epigenetic events observed in prostate cancer. Significant advances have been made for understanding how soy isoflavones are implicated in protection against prostate cancer. However, more studies are needed to better-understand and elucidate all pathways mobilized by genistein and daidzein, in order to fully exploit their anticancer properties.

Topics: Antineoplastic Agents; Apoptosis; Cell Survival; Epigenomics; Feeding Behavior; Genistein; Glycine max; Humans; Isoflavones; Male; NF-kappa B; Phytoestrogens; Prostatic Neoplasms

Isoflavones, a group of phytoestrogens, are selective oestrogen receptor (ER) modulators. They may positively impact endocrine-related conditions but the current evidence is sparse. Equol, a non-steroidal oestrogen, is produced by the metabolism of the isoflavone daidzein by intestinal bacteria. In Western countries, 30-50% of individuals metabolize daidzein into equol and are known as equol producers. Equol production may be the source of benefit from isoflavones in endocrine disease.

Topics: Animals; Dietary Supplements; Endocrine System Diseases; Equol; Female; Functional Food; Humans; Isoflavones; Male; Phytoestrogens; Soy Foods

Vitamin D is known to increase Ca absorption in adults. However, the threshold vitamin D status to benefit Ca absorption is lower than the target vitamin D status for higher bone mineral density and lower fracture risk, pointing to another pathway for vitamin D to benefit bone. One possibility is by affecting osteoblast and osteoclasts directly. Vitamin D-related bone metabolism may also be affected by soy isoflavones, which selectively bind to the estrogen receptor β and may reduce bone loss in postmenopausal women. We discuss a possible synergistic effect of soy isoflavones and vitamin D on bone by affecting osteoblast and osteoclast formation and activity in postmenopausal women.

Topics: Absorption; Bone Density; Calcium; Drug Synergism; Female; Genistein; Glycine max; Hip Fractures; Humans; Isoflavones; Middle Aged; Osteoporosis, Postmenopausal; Phytoestrogens; Postmenopause; Risk Factors; Vitamin D

Changes in the regulation of potassium channels are increasingly implicated in the altered activity of breast cancer cells. Increased or reduced expression of a number of K(+) channels have been identified in numerous breast cancer cell lines and cancerous tissue biopsy samples, compared to normal tissue, and are associated with tumor formation and spread, enhanced levels of proliferation, and resistance to apoptotic stimuli. Through knockout or silencing of K(+) channel genes, and use of specific or more broad pharmacologic K(+) channel blockers, the growth of numerous cell lines, including breast cancer cells, has been modified. In this manner it has been proposed that in MCF7 breast cancer cells proliferation appears to be regulated by the activity of a number of K(+) channels, including the Ca(2+) activated K(+) channels, and the voltage-gated K(+) channels hEAG and K(v)1.1. The effect of phytoestrogens on K(+) channels has not been extensively studied but yields some interesting results. In a number of cell lines the phytoestrogen genistein inhibits K(+) current through several channels including K(v)1.3 and hERG. Where it has been used, structurally similar daidzein has little or no effect on K(+) channel activity. Since many K(+) channels have roles in proliferation and apoptosis in breast cancer cells, the impact of K(+) channel regulation by phytoestrogens is of potentially great relevance.

Topics: Breast Neoplasms; Cell Line, Tumor; Female; Genistein; Humans; Isoflavones; Phytoestrogens; Potassium Channels

Many clinical studies have been carried out to determine the health benefits of soy protein and the isoflavones contained in soy. S-equol is not present in soybeans but is produced naturally in the gut of certain individuals, particularly Asians, by the bacterial biotransformation of daidzein, a soy isoflavone. In those intervention studies in which plasma S-equol levels were determined, a concentration of >5-10 ng/mL has been associated with a positive outcome for vasomotor symptoms, osteoporosis (as measured by an increase in bone mineral density), prostate cancer, and the cardiovascular risk biomarkers low-density lipoprotein cholesterol and C-reactive protein. These studies suggest that S-equol may provide therapeutic benefits for a number of medical needs.

Topics: Biomarkers; Biotransformation; C-Reactive Protein; Cardiovascular Diseases; Equol; Estrogen Receptor beta; Female; Humans; Intestines; Isoflavones; Lipoproteins, LDL; Male; Osteoporosis; Phytoestrogens; Prostatic Neoplasms; Risk Factors

The objective for this paper was to review human studies of O-desmethylangolensin (O-DMA) concentrations and of O-DMA producers compared with nonproducers in the context of results from in vitro studies. O-DMA is an intestinal bacterial metabolite of daidzein, an isoflavone compound observed to have phytoestrogenic properties. Not all individuals harbor bacteria capable of metabolizing daidzein to O-DMA, and individuals can be classified as O-DMA producers and nonproducers. O-DMA is less structurally similar to 17β-estradiol than its parent compound, daidzein; thus, it may exhibit different biological actions than daidzein. Evidence from in vitro studies suggests that O-DMA has several cancer-related biological actions. However, results from human metabolic studies and observational studies of disease risk suggest that these actions may not be physiologically relevant in vivo due to the amount and form (primarily glucuronide) of circulating O-DMA. Apart from circulating O-DMA concentrations, the underlying bacteria may have a distinct physiological role. Urinary excretion of O-DMA in humans is a marker of harboring intestinal bacteria capable of C-ring cleavage. Bacterial C-ring cleavage reactions are relevant to other phytochemicals that may exert biological actions in vivo that are stronger than the actions of O-DMA; thus, the role of the phenotype may extend beyond daidzein metabolism. There are a limited number of studies that have evaluated disease risk factors in relation to being an O-DMA producer, with mixed results. Further research evaluating disease risk in relation to the O-DMA-producer phenotype from the perspective of intestinal microbial composition is recommended.

Topics: Bacteria; Biomarkers; Equol; Humans; Intestines; Isoflavones; Molecular Structure; Phenotype; Phytoestrogens

The amount of soy products consumed in Japan is much greater than that in Western countries. Recent evidence indicates that soy isoflavones play a beneficial role in obesity, cancer, osteoporosis, and cardiovascular disease. The soybean isoflavone genistein is present at high levels in soy products. Genistein is structurally similar to 17beta-estradiol (E2), and genistein has been suggested to be act as E2 or an antagonist against E2. Genistein suppresses antigen-specific immune response in vivo and lymphocyte proliferation response in vitro. However, genistein enhances the cytotoxic response mediated by NK and cytotoxic T cells and the cytokine production from T cells. Thus, the effect of genistein on immunity is immune cell-dependent. Due to its unique effect on immune function, genistein has been used for the treatment of the diseases in animal models and it has been found that genistein inhibits allergic inflammatory responses. In this review, we summarize current studies related to the effect of isoflavone genistein on the immune system.

Topics: Animals; Antibody Formation; Cell Differentiation; Genistein; Glycine max; Humans; Immunity; Immunity, Cellular; Isoflavones; Neoplasms, Experimental; Phytoestrogens; T-Lymphocytes

A consensus view of soyabean phyto-oestrogens in clinical interventions in post-menopausal women is presented that is based on data from the EU-funded project Phytohealth. The phyto-oestrogens, primarily genistein and daidzein, were given as soyabean-protein isolates, whole-soyabean foods or extracts, supplements or pure compounds. A comprehensive literature search was conducted with well-defined inclusion or exclusion criteria. For areas for which substantial research exists only placebo-controlled double-blind randomised controlled trials (RCT) conducted on healthy post-menopausal women were included. For emerging areas all available human studies in post-menopausal women were reviewed. In order to make cross comparisons between studies the doses of isoflavones were calculated as aglycone equivalents. There is a suggestion, but no conclusive evidence, that isoflavones from the sources studied so far have a beneficial effect on bone health. The consumption of whole-soyabean foods and soyabean-protein isolates has some beneficial effects on lipid markers of cardiovascular risk. The consumption of isolated isoflavones does not affect blood lipid levels or blood pressure, although it may improve endothelial function. For menopausal symptoms there is currently limited evidence that soyabean-protein isolates, soyabean foods or red-clover (Trifolium pratense L.) extract are effective but soyabean isoflavone extracts may be effective in reducing hot flushes. There are too few RCT studies to reach conclusions on the effects of isoflavones on breast cancer, colon cancer, diabetes or cognitive function. The health benefits of soyabean phyto-oestrogens in healthy post-menopausal women are subtle and even some well-designed studies do not show protective effects. Future studies should focus on high-risk post-menopausal women, especially in the areas of diabetes, CVD, breast cancer and bone health.

Topics: Anticarcinogenic Agents; Breast Neoplasms; Cardiovascular Diseases; Female; Genistein; Glycine max; Humans; Isoflavones; Neoplasms; Osteoporosis, Postmenopausal; Phytoestrogens; Postmenopause; Randomized Controlled Trials as Topic

Isoflavones are recognized to be estrogenic compounds that are often associated with a reduced risk of cancers. The estrogenic activity can be enhanced after metabolization to more active compounds such as genistein and daidzein by gut microorganisms. The direct use of these metabolites has been investigated in laboratory rats and farm animals over the last decade. This paper reviews the research progress on the effect of isoflavonic compounds including metabolites on the physiology, gut microbiology and performance of farm animals in China.

Topics: Animals; Animals, Domestic; China; Digestive System; Gastrointestinal Tract; Isoflavones; Lactation; Oviposition; Phytoestrogens; Rumen

The impact of soyfood intake on breast cancer risk has been investigated extensively. Much of this focus can be attributed to the soybean being a dietary source that is uniquely rich in isoflavones. The chemical structure of isoflavones is similar to that of estrogen, and isoflavones bind to both estrogen receptors (ER alpha and ER beta) (although they preferentially bind to and activate ER beta) and exert estrogen-like effects under some experimental conditions. Isoflavones also possess nonhormonal properties that are associated with the inhibition of cancer cell growth. Thus, there are several possible mechanisms by which soy may reduce the risk of breast cancer. However, the role of isoflavones in breast cancer has become controversial because, in contrast to the possible beneficial effects, some data from in vitro and animal studies suggest that isoflavones, especially genistein, the aglycone of the main soybean isoflavone genistin, may stimulate the growth of estrogen-sensitive tumors. Limited human data directly address the tumor-promoting effects of isoflavones and soy. Because the use of soyfoods and isoflavone supplements is increasing, it is important from a public health perspective to understand the impact of these products on breast cancer risk in women at high risk of the disease and on the survival of breast cancer patients. To this end, a workshop was held in November 2005 to review the existing literature and to make research recommendations. This paper summarizes the workshop findings and recommendations. The primary research recommendation is that the impact of isoflavones on breast tissue needs to be evaluated at the cellular level in women at high risk for breast cancer.

Topics: Animals; Anticarcinogenic Agents; Breast Neoplasms; Carcinogens; China; Clinical Trials as Topic; Congresses as Topic; Disease Models, Animal; Female; Genistein; Humans; Isoflavones; Los Angeles; Neoplasms, Hormone-Dependent; Phytoestrogens; Population Surveillance; Receptors, Estrogen; Risk Factors; Soy Foods

As alternative medicine gains popularity in the US, a greater understanding of the proven benefits and detriments of the supplements commonly used is needed by physicians. Chemoprevention through the use of supplements or dietary means is one example. Through epidemiological studies, it is clear that there is variation in the geographic incidence of certain cancers. One such variation is in prostate cancer, for which Asian men have a decreased death rate as compared with their Western counterparts. One hypothesis for this reduction in prostate cancer deaths is due to the difference in soy consumption. The purpose of this paper is to review the effects of soy at the molecular level as well as to review the in-vivo effects.. The mechanism by which soy or, more accurately, the isoflavones act is described in this review. Multiple studies attempting to clarify the effects of the isoflavones on molecular pathways will be discussed. Furthermore, recent studies demonstrating the effect of isoflavones on prostate-specific antigen, testosterone, estrogen, and hormone receptor expression in human subjects will be reviewed.. After reading this review, we expect that the reader will understand the background of the isoflavones, the effect they exert at the molecular level, and their proposed benefits and limitations in human patients. However, what will be evident at the conclusion of this manuscript is the need for future studies of the effects of soy in prostate cancer patients.

Topics: Anticarcinogenic Agents; Genistein; Glycine max; Humans; Isoflavones; Male; Phytoestrogens; Phytotherapy; Plant Extracts; Prostatic Neoplasms

Plant extracts containing phytohormones are very popular as 'alternative' medicine for many kinds of diseases. They are especially favored by women who enter menopause and are concerned about the side effects of hormone replacement therapy. However, adverse health effects of phytoestrogens have often been ignored. This review examines the literature on genotoxicity and apoptotic effects of phytohormones. Genistein, coumestrol, quercetin, zearalenone, and resveratrol exerted genotoxic effects in in vitro test systems. Other phytoestrogens such as lignans, the isoflavones daidzein and glycetein, anthocyanidins, and the flavonol fisetin exhibited only weak or no effects in vitro. However, some metabolites of daidzein showed a genotoxic activity in vitro. Practically all of the phytoestrogens exhibit pro-apoptotic effects in some cell systems. Further investigations regarding dose-response-relationships and other aspects relevant for extrapolation to human exposure seem necessary. Until then, care may be advised in taking concentrated phytohormones. Nevertheless, the intake of substantial amounts of plant-food in a normal diet constitutes an important, individual contribution to cancer prevention.

Topics: Animals; Anthocyanins; Cell Proliferation; Flavonoids; Genistein; Genomic Instability; Humans; Isoflavones; Lignans; Mutagens; Phytoestrogens; Resveratrol; Stilbenes; Zearalenone

The soy-isoflavones genistein and daidzein and the flaxseed-lignans secoisolariciresinol and matairesinol belong to the group of phytoestrogens. Epidemiological data suggest that phytoestrogens have a preventive effect against various estrogen-related diseases/symptoms such as breast cancer, menopausal symptoms, cardiovascular diseases, and osteoporosis. To prove these assumptions, available controlled clinical trials have been critically reviewed. Especially soy-isoflavones have been extensively studied. There is no scientific evidence for an effect of phytoestrogens on menopausal symptoms and risk factors of breast cancer. However, isoflavones-containing soy protein can lower total cholesterol, LDL cholesterol, and triglyceride serum levels. The strongest evidence exists for a preventive effect of soy isoflavones on postmenopausal bone loss of the lumbar spine. Distinct effects on estrogen-related diseases can be explained at least in part by the different affinity of isoflavones to estrogen receptors alpha and beta and the distinct tissue distribution of these receptors.

Topics: Anticarcinogenic Agents; Clinical Trials as Topic; Female; Genistein; Humans; Isoflavones; Lipids; Osteoporosis, Postmenopausal; Phytoestrogens; Phytotherapy; Plant Preparations

Epidemiological data of phytoestrogens and prostate cancer strongly supports the cancer protective effects of isoflavones found in soy products. Inhibition of cell proliferation via hormone-dependent and hormone-independent mechanisms by soy phytochemicals has been studied extensively in cell culture and animal studies. Herein, we review the current literature on the epidemiology and effects of two soy phytoestrogens, genistein and daidzein, and would stress the need for controlled human trials to assess the true preventive and therapeutic effects of these compounds.

Topics: Animals; Disease Models, Animal; Estrogens, Non-Steroidal; Genistein; Glycine max; Humans; Isoflavones; Male; Phytoestrogens; Plant Preparations; Prostate-Specific Antigen; Prostatic Neoplasms; Rats; Rats, Wistar; Signal Transduction

Topics: Animals; Estrogens, Non-Steroidal; Genistein; Humans; Isoflavones; Myocardial Contraction; Phytoestrogens; Plant Preparations; Selective Estrogen Receptor Modulators; Vasodilator Agents

One of the problems which may beset epidemiological studies is the difficulty of accurately measuring the dietary intakes of participants. Biomarkers of diet promise to provide a more accurate measure of dietary intake and a more objective one in that they are not reliant on the subject's memory. This review considers some issues of importance in epidemiology when information is obtained from biomarkers. The approach taken is to use examples both of normal dietary constituents and of contaminants in relation to a range of diet and health questions to illustrate these points. A brief overview of the role of sample collection, processing and storage is given including some generic recommendations for maximising the reliability of subsequent analytical data. Using the examples of phytoestrogens and iodine the question of whether biomarkers can accurately reflect the intake of the dietary constituents of interest at the population level or at the individual level is considered. The relationship of the biomarker to the natural history of the disease is exemplified using the role of folate in neural tube defects. Finally, intakes of vitamin D and heterocyclic amines are used to illustrate the integration of biomarkers into epidemiological studies of prostate and colorectal cancer, respectively. It is concluded that biomarkers may provide a more accurate and objective measure of diet than estimates of current or usual intake but that this approach also has limitations. A combination of methods will probably prove to be most valuable and this approach is being taken in current large prospective studies.

Topics: Biomarkers; Colorectal Neoplasms; Cooking; Diet; Epidemiologic Factors; Estrogens, Non-Steroidal; Female; Folic Acid; Genetic Predisposition to Disease; Genistein; Health Surveys; Heterocyclic Compounds; Humans; Iodine; Isoflavones; Male; Meat; Middle Aged; Neural Tube Defects; Nutritional Status; Phytoestrogens; Plant Preparations; Predictive Value of Tests; Pregnancy; Prostatic Neoplasms; Risk Factors; Specimen Handling; Vitamin D

To determine whether genistein and daidzein, the major phytoestrogens in soy, can stimulate breast cancer growth.. Systematic search through primary English-language literature on MEDLINE (1966-January 2001), EMBASE (1982-January 2001) and Current Contents (1998-January 2001).. Genistein and daidzein at low concentrations were found to stimulate breast tumor growth in in vitro and in vivo animal studies, and antagonize the antitumor effect of tamoxifen in vitro. At high concentrations, genistein inhibited tumor growth and enhanced the effect of tamoxifen in vitro.. Genistein and daidzein may stimulate existing breast tumor growth and antagonize the effects of tamoxifen. Women with current or past breast cancer should be aware of the risks of potential tumor growth when taking soy products.

Topics: Animals; Anticarcinogenic Agents; Breast Neoplasms; Dose-Response Relationship, Drug; Estrogens, Non-Steroidal; Female; Genistein; Humans; Isoflavones; Phytoestrogens; Plant Preparations; Soybean Proteins; Tamoxifen

Isoprene is the main component of steroid hormones. It is found in many plants and herbal compounds, e.g. the isoflavonoids are therefore slightly estrogenic. Since they are bound to the estradiol receptors, more active estrogens cannot induce a signal transduction. Hence phytoestrogens may be protective on breast tissue and other hormone dependent organs.

Topics: Aged; Anticarcinogenic Agents; Breast Neoplasms; Estrogen Replacement Therapy; Estrogens, Non-Steroidal; Female; Genistein; Humans; Isoflavones; Middle Aged; Osteoporosis, Postmenopausal; Phytoestrogens; Plant Preparations; Postmenopause; Receptors, Estrogen; Resveratrol; Selective Estrogen Receptor Modulators; Stilbenes

PhytoSERM is a selective estrogen receptor beta (ERβ) modulator comprised of three phytoestrogens: genistein, daidzein, and S-equol. The PhytoSERM formulation promotes estrogenic action in the brain while largely inactive or inhibitory in reproductive tissue. A phase Ib/IIa clinical trial (ClinicalTrial.gov ID: NCT01723917) of PhytoSERM demonstrated safety and pharmacokinetics profile of PhytoSERM. While this study was not powered for efficacy analysis, we conducted a pilot, retrospective analysis to identify potential responders to PhytoSERM treatment, and to determine the optimal populations to pursue in a phase II clinical trial of efficacy of the PhytoSERM formulation.. In this retrospective analysis involving 46 participants (n = 16, placebo; n = 18, 50 mg/d PhytoSERM; and n = 12, 100 mg/d PhytoSERM), the therapeutic effect of PhytoSERM was stratified by 2 genetic risk modulators for Alzheimer's disease: mitochondrial haplogroup and APOE genotype.. Our retrospective responder analysis indicated that participants on 50 mg of daily PhytoSERM (PS50) for 12 weeks significantly reduced hot flash frequency compared with their baseline (mean [95% CI])-1.61, [-2.79, -0.42], P = 0.007). Participants on 50 mg of PhytoSERM also had significantly greater reduction in hot flash frequency at 12 weeks compared with the placebo group (-1.38, -0.17 [median PS50, median placebo], P = 0.04). Fifty milligrams of daily PhytoSERM also preserved cognitive function in certain aspects of verbal learning and executive function. Our analysis further suggests that mitochondrial haplogroup and APOE genotype can modify PhytoSERM response.. Our data support a precision medicine approach for further development of PhytoSERM as a safe and effective alternative to hormone therapy for menopause-associated hot flash and cognitive decline. While definitive determination of PhytoSERM efficacy is limited by the small sample size, these data provide a reasonable rationale to extend analyses to a larger study set powered to address statistical significance.

Topics: Apolipoproteins E; Cognition; Cognitive Dysfunction; Double-Blind Method; Equol; Feasibility Studies; Female; Genistein; Haplotypes; Hot Flashes; Humans; Isoflavones; Menopause; Middle Aged; Mitochondria; Phytoestrogens; Pilot Projects; Retrospective Studies; Selective Estrogen Receptor Modulators; Treatment Outcome

Topics: Absorptiometry, Photon; Adult; Bone Density; Calcium; Double-Blind Method; Female; Genistein; Humans; Isoflavones; Lumbar Vertebrae; Pelvic Bones; Phytoestrogens; Placebos; Premenopause

Soy phytoestrogens are potential alternatives to postmenopausal hormone replacement therapy (HRT). Adverse effects of HRT such as myocardial infarction, stroke, and pulmonary embolism are mediated by calcium-induced signaling.. To determine whether soy isoflavones affect serum calcium in healthy female subjects.. In a double-blind trial, 197 premenopausal women were randomly assigned to either isoflavone (N = 99) or placebo pills (N = 98) 5 days per week for up to 2 years, plus prenatal vitamins. Isoflavone pills contained 60 mg genistein, 60 mg daidzein and 16.6 mg glycitein (expressed as aglycone equivalents). All pills contained 15 mg riboflavin as an adherence marker. Blood chemistries and urinary daidzein, genistein and riboflavin were measured multiple times during the luteal phase before and during treatment.. Analysis of the adherent population (N = 83 per group), revealed significantly strong associations between urinary levels of isoflavones and serum concentrations of calcium (regression coefficients 0.082 for daidzein and 0.229 for genistein, all P < 0.01) and chloride (regression coefficient, -1.537 for genistein, P < 0.0001), mediated in part by albumin. The effects amounted to mean changes of +0.24 mg/dL for calcium and -1.45 mEq/L for chloride, with each visit for subjects excreting the most vs. the least amounts of isoflavones. These associations were not evident in the intention-to-treat analysis (N = 197) that did not assess expected variations in isoflavone levels within and between subjects from metabolism and adherence.. These novel and strong effects of soy isoflavones on calcium homeostasis have important implications for long term effects of these natural substances on cardiovascular diseases.

Topics: Adult; Calcium; Chlorides; Double-Blind Method; Female; Genistein; Glycine max; Humans; Isoflavones; Phytoestrogens; Placebos; Premenopause; Riboflavin

There is much speculation with regard to the potential cardioprotective benefits of equol, a microbial-derived metabolite of the isoflavone daidzein, which is produced in the large intestine after soy intake in 30% of Western populations. Although cross-sectional and retrospective data support favorable associations between the equol producer (EP) phenotype and cardiometabolic health, few studies have prospectively recruited EPs to confirm this association.. The aim was to determine whether the acute vascular benefits of isoflavones differ according to EP phenotype and subsequently investigate the effect of providing commercially produced S-(-)equol to non-EPs.. We prospectively recruited male EPs and non-EPs (n = 14/group) at moderate cardiovascular risk into a double-blind, placebo-controlled crossover study to examine the acute effects of soy isoflavones (80-mg aglycone equivalents) on arterial stiffness [carotid-femoral pulse-wave velocity (cfPWV)], blood pressure, endothelial function (measured by using the EndoPAT 2000; Itamar Medical), and nitric oxide at baseline (0 h) and 6 and 24 h after intake. In a separate assessment, non-EPs consumed 40 mg S-(-)equol with identical vascular measurements performed 2 h after intake.. After soy intake, cfPWV significantly improved in EPs at 24 h (cfPWV change from 0 h: isoflavone, -0.2 ± 0.2 m/s; placebo, 0.6 ± 0.2 m/s; P < 0.01), which was significantly associated with plasma equol concentrations (R = -0.36, P = 0.01). No vascular effects were observed in EPs at 6 h or in non-EPs at any time point. Similarly, no benefit of commercially produced S-(-)equol was observed in non-EPs despite mean plasma equol concentrations reaching 3.2 μmol/L.. Acute soy intake improved cfPWV in EPs, equating to an 11-12% reduced risk of cardiovascular disease if sustained. However, a single dose of commercially produced equol had no cardiovascular benefits in non-EPs. These data suggest that the EP phenotype is critical in unlocking the vascular benefits of equol in men, and long-term trials should focus on confirming the implications of EP phenotype on cardiovascular health. This trial was registered at clinicaltrials.gov as NCT01530893.

Topics: Aged; Bacteria; Cardiovascular Diseases; Cross-Over Studies; Cross-Sectional Studies; Dietary Supplements; Double-Blind Method; Equol; Gastrointestinal Microbiome; Glycine max; Humans; Isoflavones; Male; Middle Aged; Phenotype; Phytoestrogens; Phytotherapy; Plant Extracts; Prospective Studies; Pulse Wave Analysis; Vascular Stiffness

Although higher habitual soy intake is associated with lower blood pressure (BP) and stroke incidence, clinical trials using soy protein or isoflavones on cardiovascular risks yielded inconsistent results. The discrepancies are hypothesized to be due to the individuals' intestinal bacterial capacity to metabolite isoflavones daidzein into equol. Animal and in vitro studies have revealed that equol has stronger estrogen-like and anti-oxidative activity than isoflavones and possesses natriuretic and vasorelaxant properties which may play an important role in the prevention of hypertension. However, no clinical trial has examined the effect of equol on BP. We thus propose a 24-week randomized controlled trial to test the effectiveness of natural S-equol on BP and vascular function among equol non-producers.. This will be a 6-month double-blind, randomized, placebo-controlled trial among 207 non-equol producing postmenopausal women with prehypertension or early untreated hypertension. Eligible participants who have completed a 2-week run-in will be randomized to either one of the 3 groups: placebo group, low-equol group (10 mg/d) and high equol group (20 mg/d). The outcome measures will be conducted at baseline and at the end of the trial including 24 h ambulatory BP, endothelial function (by ultrasound determined brachial flow mediated dilation), arterial stiffness (by pulse wave analysis) and other cardiovascular risk factors (lipid profile, glycemic control and inflammatory biomarkers). Urinary isoflavones will be tested for compliance assessment. One way analysis of variance will be applied to compare the 6-month changes in ambulatory BP or parameters of vascular function among the 3 treatment groups.. This study will be performed in community subjects. If the antihypertensive effect of equol is proven, the provision of natural equol to those high risk adults who are unable to produce equol will have enormous public health implications for the primary and secondary prevention of hypertension and cardiovascular diseases on a population basis. The research efforts will also have significant implications for industry in the provision of suitable soy products for the prevention of hypertension and its related complications.. The trial was registered in ClinicalTrials.gov with identifier of NCT02515682 .

Topics: Aged; Blood Pressure; Clinical Protocols; Double-Blind Method; Equol; Female; Glycine max; Humans; Hypertension; Isoflavones; Middle Aged; Natriuretic Agents; Phenotype; Phytoestrogens; Plant Extracts; Postmenopause; Prehypertension; Research Design; Soybean Proteins; Vasodilation; Vasodilator Agents

Although observational studies suggest that soy foods or isoflavones are cardio-protective, clinical trials on whole soy or isoflavone daidzein (one major isoflavone and the precursor of equol) on blood pressure (BP) and endothelial function (EF) are few and have not been specifically conducted among equol producers, a population most likely to benefit from soy treatment.. We performed a 6-month double-blind, randomized, placebo-controlled trial to examine the effect of whole soy (soy flour) or purified daidzein on BP and EF in prehypertensive or untreated hypertensive postmenopausal women verified to be equol producers. A total of 270 eligible women were recruited and randomized to either one of the three treatment groups, 40 g soy flour (whole soy group), 40 g low-fat milk powder+63 mg daidzein (daidzein group) or 40 g low-fat milk powder (active control group) daily, each given as a solid beverage powder for 6 months. The primary outcome measures were 24 h ambulatory BP (ABP) and EF assessed by flow-mediated dilation using brachial artery ultrasound.. A total of 253 subjects completed the study according to protocol. Urinary isoflavones indicated good compliance with the interventions. Intention to treat and per-protocol analysis indicated that there was no significant difference in the 6-month changes or % changes in parameters of ABP and brachial flow-mediated dilation among the three treatment groups. A further subgroup analysis among hypertensive women (n=138) did not alter the conclusions.. Whole soy and purified daidzein had no significant effect on BP and vascular function among equol-producing postmenopausal women with prehypertension or untreated hypertension.

Topics: Blood Pressure; Blood Pressure Monitoring, Ambulatory; China; Double-Blind Method; Endothelium, Vascular; Equol; Female; Glycine max; Humans; Hypertension; Isoflavones; Middle Aged; Phytoestrogens; Plant Extracts; Postmenopause; Prehypertension; Seeds; Soy Foods; Vasodilation

Postmenopausal estrogen depletion is a major contributing factor to bone loss. Soy isoflavones have variable effects on the prevention of postmenopausal bone loss, which is possibly related to the specific isoflavone content or the variable equol-producing capacity of individuals.. We aimed to determine the effects of the content of isoflavones in a soy supplement and the equol-producing ability of the individual on postmenopausal bone calcium retention.. The study was a blinded, randomized, crossover intervention trial in 24 postmenopausal women who were prescreened for their ability to convert daidzein to equol. Women were equilibrated with (41)Ca before the intervention. Interventions were 5 soy isoflavone oral supplements (2 doses of a genistein-rich soy supplement and 3 doses of mixed isoflavones in various proportions) and a bisphosphonate (risedronate). Each intervention was given sequentially for 50 d followed by a 50-d washout period. The percentage of bone calcium retention was determined from the change in urinary (41)Ca:calcium.. Interventions that ranged from 52 to 220 mg total isoflavones/d increased bone calcium retention between 3.4% and 7.6% (P < 0.05), which was a moderate effect compared with that of risedronate at 15.3% (95% CI: 7.1%, 22.7%; P = 0.0014). The most-effective soy intervention delivered 105.23 mg total isoflavones/d as genistein, daidzein, and glycitein in their natural ratios and increased bone calcium retention by 7.6% (95% CI: 4.9%, 10.2%; P < 0.0001). Genistein, at 52.85 mg/d, increased bone calcium retention by 3.4% (95% CI: 0.5%, 6.2%; P = 0.029); but there was no benefit at higher amounts (113.52 mg/d). There was no difference (P = 0.5) in bone calcium retention between equol producers and nonproducers.. Soy isoflavones, although not as potent as risedronate, are effective bone-preserving agents in postmenopausal women regardless of their equol-producing status, and mixed isoflavones in their natural ratios are more effective than enriched genistein. This trial was registered at clinicaltrials.gov as NCT00244907.

Topics: Administration, Oral; Aged; Bone and Bones; Calcium; Cross-Over Studies; Dietary Supplements; Dose-Response Relationship, Drug; Equol; Female; Genistein; Glycine max; Humans; Isoflavones; Middle Aged; Osteoporosis, Postmenopausal; Phytoestrogens; Postmenopause; Risedronic Acid

Dietary supplements containing soy or isoflavones are widely used as alternatives to hormone therapy. However, their efficacy is still inconclusive, and limited data on equol producers are available. The aim of this study was to examine the effect of whole soy (soy flour) or purified daidzein (one major soy isoflavone and the precursor of equol) on menopausal symptoms in equol-producing postmenopausal women, a population most likely to benefit from soy intervention.. This is a 6-month parallel-group, double-blind, randomized, placebo-controlled trial. Two hundred seventy equol-producing prehypertensive Chinese postmenopausal women were randomized to one of three treatment groups: 40 g of soy flour (whole soy group), 40 g of low-fat milk powder + 63 mg of daidzein (daidzein group), or 40 g of low-fat milk powder (placebo group) daily, each given as a solid beverage for 6 months. Changes in menopausal symptoms were assessed by a validated and structured symptom checklist at baseline and 6 months.. Two hundred fifty-three participants completed the study according to protocol. Urinary isoflavones indicated good compliance with the interventions. Baseline menopausal symptoms were comparable among the three study groups. Intention-to-treat analysis indicated that there was no significant difference in the 6-month changes or percent changes in the total number of menopausal symptoms, in the five dimensions of symptoms, and in the frequencies of individual symptoms among the three treatment groups.. Whole soy and purified daidzein have no significant effect on alleviation of menopausal symptoms among equol-producing postmenopausal women with prehypertension.

Topics: Anxiety; Back Pain; China; Cognition; Cough; Depression; Dietary Supplements; Double-Blind Method; Dyspnea; Equol; Female; Glycine max; Hot Flashes; Humans; Isoflavones; Phytoestrogens; Phytotherapy; Plant Preparations; Postmenopause; Sleep Wake Disorders

Daidzein (one of the major isoflavones) can be metabolized to equol in certain individuals. The effects of isoflavones alone and equol status on lipid profiles are still controversial. To evaluate the 6-mo effects of daidzein on cardiovascular risk factors in hypercholesterolemic individuals and the interactions of these effects with equol status and estrogen receptor (ESR) genotypes, we conducted a randomized, double-blind, placebo-controlled trial consisting of 210 hypercholesterolemic adults (40-65 y old). The participants were randomly assigned (177 completed) to consume placebo, 40 mg daidzein (DAI40), or 80 mg daidzein (DAI80) daily for 6 mo. Daidzein decreased serum triglycerides (TGs) by 0.15 ± 0.62 mmol/L (mean ± SD) and 0.24 ± 0.61 mmol/L and decreased serum uric acid by 23 ± 47 μmol/L and 29 ± 44 μmol/L in the DAI40 and DAI80 groups, respectively. These reductions in the DAI40 and DAI80 groups were greater than those in the placebo group (P < 0.05). Other blood lipids, glucose, insulin, or glycated hemoglobin did not significantly change after daidzein treatment. No dose-dependent effects of daidzein were found. The reduction of TGs was influenced by the ESR genotype, with a greater effect observed in participants with the GA genotype compared with those with the GG genotype of ESR-β RsaI. These effects were not influenced by equol status. Six-month supplementation of daidzein significantly decreased TGs and uric acid. ESR-β RsaI genotype, not equol status, influenced daidzein's effects on TGs. Daidzein consumption may be effective to improve cardiovascular risk factors, especially in adults with the GA genotype of ESR-β RsaI. This trial was registered at the Chinese clinical trial registry as ChiCTR-TRC-10001048.

Topics: Adult; Aged; Asian People; Cardiovascular Diseases; Dietary Supplements; Double-Blind Method; Energy Intake; Estrogen Receptor beta; Female; Genotype; Humans; Hypercholesterolemia; Isoflavones; Male; Middle Aged; Phytoestrogens; Risk Factors; Triglycerides; Uric Acid

This paper reported the effects of commonly used whole soy foods (soy flour) and purified daidzein (one of the major isoflavones and the precursor of equol) on changes in anthropometric measurements and body composition in a 6-month double-blind, randomized, placebo-controlled trial among prehypertensive postmenopausal women who are also equol producers.. 270 eligible women were randomized to either one of the three treatments: 40 g soy flour (whole soy group), 40 g low-fat milk powder + 63 mg daidzein (daidzein group), or 40 g low-fat milk powder (placebo group) daily each for 6 months. Anthropometric indicators and body composition were measured before and after intervention.. 253 subjects completed the study with good compliance. Urinary isoflavones levels suggested good compliance of subjects with supplementation. Whole soy and purified daidzein had no significant effect on body weight, body mass index (BMI), waist and hip circumferences, waist to hip ratio (WHR), body fat percentage, fat mass, and free fat mass.. Six-month consumption of whole soy and purified daidzein at provided dosage had no improvement on body weight and composition compared with isocaloric milk placebo among prehypertensive equol-producing postmenopausal women. This trial is registered with ClinicalTrials.gov NCT01270737.

Topics: Adiposity; Aged; Analysis of Variance; Body Composition; Body Mass Index; Body Weight; Double-Blind Method; Equol; Female; Hong Kong; Humans; Isoflavones; Middle Aged; Overweight; Phytoestrogens; Postmenopause; Predictive Value of Tests; Prehypertension; Soy Foods; Time Factors; Treatment Outcome; Waist Circumference; Waist-Hip Ratio

The treatment of 300-mg/day isoflavones (aglycone equivalents) (172.5 mg genistein + 127.5 mg daidzein) for 2 years failed to prevent lumbar spine and total proximal femur bone mineral density (BMD) from declining as compared with the placebo group in a randomized, double-blind, two-arm designed study enrolling 431 postmenopausal women 45-65 years old.. This study evaluated the effects of soy isoflavones on bone metabolism in postmenopausal women.. Four hundred and thirty-one women, aged 45-65 years, orally consumed 300-mg/day isoflavones (aglycone equivalents) or a placebo for 2 years in a parallel group, randomized, double-blind, two-arm study. Each participant also ingested 600 mg of calcium and 125 IU of vitamin D(3) per day. The BMD of the lumbar spine and total proximal femur were measured using dual-energy X-ray absorptiometry at baseline and every half-year thereafter. Serum bone-specific alkaline phosphatase, urinary N-telopeptide of type 1 collagen/creatinine, and other safety assessments were examined regularly.. Two hundred out of 217 subjects in the isoflavone group and 199 out of 214 cases in placebo group completed the treatment. Serum concentrations of isoflavone metabolites, genistein and daidzein, of the intervention group were remarkably elevated following intake of isoflavones (p < 0.001). However, differences in the mean percentage changes of BMD throughout the treatment period were not statistically significant (lumbar spine, p = 0.42; total femur, p = 0.39) between the isoflavone and placebo groups, according to the generalized estimating equation (GEE) method. A significant time trend of bone loss was observed at both sites as assessed by the GEE method following repeated measurement of BMD (p < 0.001). Differences in bone marker levels were not significant between the two treatment groups.. Treatment with 300-mg/day isoflavones (aglycone equivalents) failed to prevent a decline in BMD in the lumbar spine or total femur compared with the placebo group.

Topics: Absorptiometry, Photon; Bone Density; Bone Density Conservation Agents; Double-Blind Method; Drug Administration Schedule; Drug Combinations; Female; Femur; Genistein; Humans; Isoflavones; Lumbar Vertebrae; Middle Aged; Osteoporosis, Postmenopausal; Phytoestrogens; Placebos; Treatment Outcome

Estrogen therapy is, to date, the most effective treatment of menopausal syndrome and also has a favorable effect on lipid profiles. Because of its potential adverse effects, however, a more acceptable alternative therapy needs to be identified. This study examines the effect of soy germ isoflavones on menopausal symptoms and serum lipids.. Ninety early postmenopausal Chinese women, aged 45 to 60 years, were randomly assigned to three treatment groups (30 each) receiving daily doses of 0 (placebo), 84, and 126 mg of soy germ isoflavones. Hot flush frequency, Kupperman scores, serum 17β-estradiol, follicle-stimulating hormone, luteinizing hormone, and serum lipids, including triglyceride, total cholesterol, low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, apolipoprotein A-I, and apolipoprotein B100, were assessed at baseline and at 12 and 24 weeks after treatment.. Both the frequency of hot flushes and the Kupperman index score decreased in all three treatment groups during the intervention period, but the percentage decreases in both were significantly greater in the two isoflavone groups (44.3 ± 19.1 and 57.8 ± 37.4 [84 mg isoflavones]; 48.5 ± 27.2 and 56.7 ± 26.7 [126 mg isoflavones]) than in the placebo group (27.8 ± 15.5 and 34.6 ± 46.2; p < 0.01). There was no significant difference in the changes in estradiol, follicle-stimulating hormone, and luteinizing hormone among the three treatment groups during the study, and no significant differences were observed in the lipid components.. A daily supplement of 84 or 126 mg soy germ isoflavones may improve menopausal symptoms, although neither dose was found to affect lipid profiles in early postmenopausal Chinese women after 24 weeks of treatment. The favorable effects are unlikely to be associated with female hormones.

Topics: China; Estradiol; Estrogens; Female; Follicle Stimulating Hormone; Glycine max; Hot Flashes; Humans; Isoflavones; Lipids; Luteinizing Hormone; Middle Aged; Phytoestrogens; Postmenopause; Severity of Illness Index; Single-Blind Method; Treatment Outcome

To investigate the effects of standardized soy extract on climacteric symptoms, lipid profiles, bone markers, and serum isoflavone concentration in healthy Taiwanese postmenopausal women.. A multicenter, open-labeled, randomized, prospective, comparative study design was used. A total of 130 outpatients who had undergone natural menopause were randomly administered either 70 mg or 35 mg soy extract daily for 24 weeks.. The evidence suggests that the soy extract treatment that was administered to both groups for 1 month could help reduce climacteric scores (reductions of 19.66% [p<0.01] and 18.85% [p<0.01] in the 35 mg and 70 mg groups compared with baseline, respectively), and the efficacy was more potent after 6 months of treatment. Soy isoflavone significantly reduced the total cholesterol (reductions of 4.50% [p<0.01] and 3.06% [p<0.05] in the 35 mg and 70 mg groups, respectively) and low density lipoprotein cholesterol levels (reductions of 4.67% [p<0.05] and 5.09% [p<0.05] in the 35 mg and 70 mg groups, respectively) in patients with total cholesterol > 200 mg/dL after 6 months of treatment. In patients with high bone turnover (urinary deoxypyridinoline/creatinine > 7.4 nM/mM), soy extract treatment reduced the deoxypyridinoline/creatinine level by 10.53% (p<0.05) and 11.58% (p<0.05) in the 35 mg and 70 mg groups, respectively. Serum levels of isoflavone increased in both groups after 6 months of treatment.. Soy extract is highly efficacious at relieving menopausal symptoms and demonstrates a positive effect on the cardiovascular system and skeleton.

Topics: Amino Acids; Analysis of Variance; Cholesterol; Cholesterol, LDL; Creatinine; Female; Genistein; Glycine max; Hot Flashes; Humans; Isoflavones; Middle Aged; Phytoestrogens; Postmenopause; Severity of Illness Index

Shell quality decreases as laying hens age and the aim of present study was to investigate how a supplement of daidzein, a natural phytoestrogen in soya, affects key factors in the shell gland and eggshell quality in late-stage laying hens. Hybrids of Lohmann Selected Leghorn (LSL) and Lohmann Brown (LB), received either a daidzein diet (50 mg/kg feed) or a control diet from 60 to 72 weeks of age. Both the total number of capillaries and capillaries with carbonic anhydrase (CA) activity were higher in the LSL hybrid than in the LB. After daidzein supplementation the number of CA positive capillaries was unaffected in the LSL but increased in the LB hybrid indicating a higher sensitivity to daidzein in this hybrid. Estrogen receptor alpha and beta (ERα, ERβ) were localized and the complete picture of the two ERs can now be described in shell gland of domestic hens. Nuclear and cytoplasmic staining was generally stronger for ERβ, while membrane associated staining was present only for ERα. Interestingly, capillary endothelium contained only ERβ and since estrogen regulation of CA is well documented, the presence of an endothelial ER provides one possible route for the increase in CA positive capillaries found in LB hybrids. Eggshell quality or egg production was not affected by daidzein supplementation. The hybrids used in this study showed anatomical differences and reacted differently to daidzein supplementation, but if this can be explained by the divergences in ERβ localization noted between the hybrids remains to be clarified.

Topics: Animal Feed; Animals; Chickens; Diet; Dietary Supplements; Drug Administration Schedule; Egg Shell; Female; Genitalia, Female; Isoflavones; Oviposition; Phytoestrogens

Evaluate the effect of diet, physical exercise, and a daily oral intake of a soy isoflavones extract (Fisiogen(®)) contained 200 mg of Glycine max, which corresponded to 80 mg of isoflavone (60.8 mg of genistein, 16 mg of daidzein and 3.2 mg of glicitein) on leptin and other adipokines plasma levels in healthy obese postmenopausal women.. A multicentric randomized longitudinal prospective cohort study was conducted in a sample of 87 healthy obese postmenopausal women. Patients were randomly assigned to a 1200 kcal diet and exercise group (control group) or a group of 1200 kcal diet, exercise, and daily oral intake of daily oral intake of a soy isoflavones extract (Fisiogen(®)) contained 200 mg of Glycine max, which corresponded to 80 mg of isoflavone (60.8 mg of genistein, 16 mg of daidzein and 3.2 mg of glicitein) (soy isoflavones group) along 6 months. Main outcome measures were: anthropometric measures, body composition, leptin, adiponectin, TNF-alpha, homocysteine, C-reactive protein, glucose, insulin, lipid profile and oestradiol serum levels, Kupperman index and Cervantes Scale.. Mean serum leptin and TNF-alpha levels declined after 6 months in both groups of the study, but only women in the soy isoflavones group showed a significant increase of mean serum levels of adiponectin.. Diet, physical exercise and daily oral intake of a soy isoflavones extract (Fisiogen(®)) contained 200 mg of Glycine max, which corresponded to 80 mg of isoflavone (60.8 mg of genistein, 16 mg of daidzein and 3.2 mg of glicitein) have a beneficial effect on serum leptin, adiponectin and TNF-α in healthy obese postmenopausal women after 6 months of treatment.

Topics: Adipokines; Cytokines; Diet; Exercise; Female; Genistein; Glycine max; Humans; Isoflavones; Leptin; Middle Aged; Obesity; Phytoestrogens; Plant Extracts; Postmenopause; Prospective Studies; Tumor Necrosis Factor-alpha; Women's Health

Concerns regarding the risk of estrogen replacement have resulted in a significant increase in the use of soy products by menopausal women who, despite the lack of evidence of the efficacy of such products, seek alternatives to menopausal hormone therapy. Our goal was to determine the efficacy of soy isoflavone tablets in preventing bone loss and menopausal symptoms.. The study design was a single-center, randomized, placebo-controlled, double-blind clinical trial conducted from July 1, 2004, through March 31, 2009. Women aged 45 to 60 years within 5 years of menopause and with a bone mineral density T score of -2.0 or higher in the lumbar spine or total hip were randomly assigned, in equal proportions, to receive daily soy isoflavone tablets, 200 mg, or placebo. The primary outcome was changes in bone mineral density in the lumbar spine, total hip, and femoral neck at the 2-year follow-up. Secondary outcomes included changes in menopausal symptoms, vaginal cytologic characteristics, N -telopeptide of type I bone collagen, lipids, and thyroid function.. After 2 years, no significant differences were found between the participants receiving soy tablets (n = 122) and those receiving placebo (n = 126) regarding changes in bone mineral density in the spine (-2.0% and -2.3%, respectively), the total hip (-1.2% and -1.4%, respectively), or the femoral neck (-2.2% and -2.1%, respectively). A significantly larger proportion of participants in the soy group experienced hot flashes and constipation compared with the control group. No significant differences were found between groups in other outcomes.. In this population, the daily administration of tablets containing 200 mg of soy isoflavones for 2 years did not prevent bone loss or menopausal symptoms.. clinicaltrials.gov Identifier: NCT00076050.

Topics: Dietary Supplements; Double-Blind Method; Estrogen Replacement Therapy; Female; Genistein; Glycine max; Hot Flashes; Humans; Isoflavones; Logistic Models; Middle Aged; Osteoporosis, Postmenopausal; Phytoestrogens; Phytotherapy

Despite decades of research on the relation between soy and breast cancer, questions regarding the absorption, metabolism, and distribution of isoflavones in breast tissue largely remain unanswered.. We evaluated the potential health effects of isoflavone consumption on normal breast tissue; isoflavone concentrations, metabolites, and biodistribution were investigated and compared with 17beta-estradiol exposure.. In this dietary intervention study, healthy women were randomly allocated to a soy milk (n = 11; 16.98-mg genistein and 5.40-mg daidzein aglycone equivalents per dose), soy supplement (n = 10; 5.27-mg genistein and 17.56-mg daidzein aglycone equivalents per dose), or control (n = 10) group. After a run-in period > or = 4 d, 3 doses of soy milk or soy supplements were taken daily for 5 d before an esthetic breast reduction. Blood and breast biopsies were collected during surgery and analyzed with liquid chromatography-tandem mass spectrometry.. After soy administration, genistein and total daidzein concentrations, which were expressed as aglycone equivalents, ranged from 135.1 to 2831 nmol/L and 105.1 to 1397 nmol/L, respectively, in hydrolyzed serum and from 92.33 to 493.8 pmol/g and 22.15 to 770.8 pmol/g, respectively, in hydrolyzed breast tissue. The major metabolites identified in nonhydrolyzed samples were genistein-7-O-glucuronide and daidzein-7-O-glucuronide, with an overall glucuronidation of 98%. Total isoflavones showed a breast adipose/glandular tissue distribution of 40:60, and their mean (+/-SEM) derived 17beta-estradiol equivalents toward estrogen receptor beta were 21 +/- 4-fold and 40 +/- 10-fold higher than the 17beta-estradiol concentrations in adipose (0.283 +/- 0.089 pmol/g, P < 0.001) and glandular (0.246 +/- 0.091 pmol/g, P = 0.001) fractions, respectively.. After intake of soy milk and soy supplements, isoflavones reach exposure levels in breast tissue at which potential health effects may occur.

Topics: Adipose Tissue; Adolescent; Adult; Breast; Chromatography, Liquid; Dietary Supplements; Estradiol; Estrogen Receptor beta; Female; Genistein; Glycine max; Humans; Isoflavones; Mammary Glands, Human; Middle Aged; Phytoestrogens; Plant Extracts; Reference Values; Soy Foods; Tandem Mass Spectrometry; Tissue Distribution; Young Adult

Effective dietary intervention strategies that can be widely disseminated and have the potential for sustainable dietary modifications are needed. The purpose of this study was to describe and evaluate the effectiveness of a telephone-based soy intervention.. A randomized controlled trial comparing self-reported intake and serum measures of soy during a 1-year dietary soy (Soy) to fruit and vegetable (Placebo) intervention conducted in two of five arms from the Herbal Alternatives Trial between May 2001 and September 2004.. One hundred sixty-three peri- and postmenopausal women (mean age=52 years) consuming self-selected diets in the Pacific Northwest, United States.. Five telephone contacts with a registered dietitian during a 12-month intervention with the goal to increase soy food consumption to two servings daily.. Change from baseline in self-reported soy servings and serum isoflavone (daidzein and genistein) concentrations were estimated using analysis of variance and generalized estimating equations. Proportions of participants achieving the intervention goal were compared using chi(2) tests.. Ninety-four percent (n=74) of participants in the Soy arm and 89% (n=75) in the Placebo arm completed the trial, and slightly more than one third (n=27) received five phone contacts. Mean (+/-standard deviation) intakes of soy were similar for the Soy and Placebo arms at baseline (0.6+/-1.0 vs 0.4+/-0.8 servings/day; P>0.05). At 12-month follow-up visit, mean+/-standard deviation servings of soy per day were 1.6+/-1.4 for the Soy intervention compared to 0.5+/-0.9 within the Placebo arm (P<0.001). There were concomitant increases in serum isoflavones at 3 and 6 months from baseline in the Soy arm only, with approximately twofold increases in both daidzein (mean=66.4 nmol/L, 95% confidence interval [CI]: 39.0 to 93.9 [mean 16.9 ng/mL, 95% CI: 9.9 to 23.8]) and genistein (mean=100.4 nmol/L, 95% CI: 60.9 to 139.9 [mean 27.1 ng/mL, 95% CI: 16.5 to 37.8]) concentrations. Mean weight changed by <1 kg during the 12-month period in each group and physical activity remained stable, suggesting that participants incorporated soy foods into their diet by substituting for non soy foods rather than adding them to their diet.. A brief telephone-based intervention with a focused message delivered by a registered dietitian is a feasible approach for encouraging targeted dietary changes, such as an increase in soy intake among peri- and postmenopausal women.

Topics: Analysis of Variance; Chi-Square Distribution; Counseling; Dietetics; Female; Fruit; Genistein; Health Promotion; Humans; Isoflavones; Middle Aged; Perimenopause; Phytoestrogens; Postmenopause; Self Disclosure; Soy Foods; Telephone; United States; Vegetables

Seaweed and soy foods are consumed daily in Japan, where breast cancer rates for postmenopausal women are significantly lower than in the West. Likely mechanisms include differences in diet, especially soy consumption, and estrogen metabolism. Fifteen healthy postmenopausal women participated in this double-blind trial of seaweed supplementation with soy challenge. Participants were randomized to 7 wk of either 5 g/d seaweed (Alaria) or placebo (maltodextrin). During wk 7, participants also consumed a daily soy protein isolate (2 mg isoflavones/kg body weight). After a 3-wk washout period, participants were crossed over to the alternate supplement schedule. There was an inverse correlation between seaweed dose (mg/kg body weight) and serum estradiol (E2) (seaweed-placebo = y = -2.29 x dose + 172.3; r = -0.70; P = 0.003), [corrected] which was linear across the range of weights. Soy supplementation increased urinary daidzein, glycitein, genistein, and O-desmethylangolensin (P = 0.0001) and decreased matairesinol and enterolactone (P < 0.05). Soy and seaweed plus soy (SeaSoy) increased urinary excretion of 2-hydroxyestrogen (2-OHE) (P = 0.0001) and the ratio of 2-OHE:16alpha-hydroxyestrone (16alphaOHE(1)) (P = 0.01). For the 5 equol excretors, soy increased urinary equol excretion (P = 0.0001); the combination of SeaSoy further increased equol excretion by 58% (P = 0.0001). Equol producers also had a 315% increase in 2:16 ratio (P = 0.001) with SeaSoy. Seaweed favorably alters estrogen and phytoestrogen metabolism and these changes likely include modulation of colonic bacteria.

Topics: Aged; Cross-Over Studies; Diet; Dietary Supplements; Double-Blind Method; Equol; Estradiol; Estrogens; Female; Genistein; Humans; Isoflavones; Middle Aged; Phytoestrogens; Placebos; Postmenopause; Seaweed; Sex Hormone-Binding Globulin; Soybean Proteins

Isoflavone intake is associated with various properties beneficial to human health which are related to their antioxidant activity, for example, to their ability to increase LDL oxidation resistance. However, the distribution of isoflavones among plasma lipoproteins has not yet been elucidated in vivo. Therefore, the objective of the present study was to investigate the association between daidzein (DAI) and lipoproteins in human plasma upon administration of the aglycone and glucoside form. Five men aged 22-30 years participated in a randomised, double-blind study in cross-over design. After ingestion of DAI and daidzein-7-O-beta-D-glucoside (DG) (1 mg DAI aglycone equivalents/kg body weight) blood samples were drawn before isoflavone administration as well as 1, 2, 3, 4.5, 6, 8, 10, 12, 24 and 48 h post-dose. Concentrations of DAI in the different lipoprotein fractions (chylomicrons, VLDL, LDL, HDL) and in the non-lipoprotein fraction were analysed using isotope dilution capillary GC/MS. The lipoprotein fraction profiles were similar for all subjects and resembled those obtained for plasma in our previously published study. The lipoprotein distribution based on the area under the concentration-time profiles from 0 h to infinity in the different fractions were irrespective of the administered form: non-lipoprotein fraction (53%) > LDL (20%) > HDL (14%) > VLDL (9.5%) > chylomicrons (2.5%). Of DAI present in plasma, 47 % was associated to lipoproteins. Concentrations in the different lipoprotein fractions as well as in the non-lipoprotein fraction were always higher after the ingestion of DG than of DAI. Taken together, these results demonstrate an association between isoflavones and plasma lipoproteins in vivo.

Topics: Adult; Biological Transport; Cholesterol, HDL; Cholesterol, LDL; Chylomicrons; Cross-Over Studies; Double-Blind Method; Gas Chromatography-Mass Spectrometry; Glycine max; Humans; Isoflavones; Lipoproteins; Lipoproteins, VLDL; Male; Phytoestrogens; Young Adult

We hypothesized that a dietary combination of soy with either a probiotic (yoghurt) or a prebiotic (resistant starch) would result in enhanced lipid-lowering effects compared with a control soy diet, possibly via improvements in isoflavone bioavailability.. Mildly hypercholesterolaemic subjects (men and post-menopausal women) older than 45 years were recruited via the local media. Thirty-six subjects commenced the study; five withdrew.. Soy+probiotic significantly decreased total cholesterol (4.7+/-2.0%; P=0.038) and soy+prebiotic significantly decreased total and low-density lipoprotein cholesterol (5.5+/-1.6%; P=0.003 and 7.3+/-2.2%; P=0.005, respectively). The bioavailabilities of daidzein, genistein or equol were not affected by probiotic or prebiotic consumption or associated with lipid changes.. Dietary combination of soy with either a probiotic or a prebiotic resulted in significant lipid lowering, not related to isoflavone bioavailability.

Topics: Aged; Bread; Cholesterol; Cholesterol, HDL; Cholesterol, LDL; Cross-Over Studies; Diet; Dietary Carbohydrates; Female; Genistein; Humans; Hypercholesterolemia; Isoflavones; Male; Middle Aged; Phytoestrogens; Probiotics; Soy Foods; Starch; Triglycerides; Triticum; Yogurt

Soya isoflavones, genistein and daidzein, are the focus of numerous studies investigating their potential effects on health and results remain controversial. Bioavailability is clearly a crucial factor influencing their bioefficacy and could explain these discrepancies. This study aimed at assessing: (1) the isoflavone content of sixty-nine European soya-derivative products sold on the French market; (2) the bioavailability of isoflavones comparing supplement with food. Twelve healthy volunteers were recruited in a randomized two-way crossover trial and received 35 mg isoflavones equivalent aglycone either through supplements or through cheese, both containing different patterns of isoflavone conjugates and different daidzein:genistein ratios. A specific ELISA method was used to assess the plasma and urinary concentrations of isoflavones and thus the pharmacokinetic parameters, which were then normalized to mg of each isoflavone ingested. Results showed that the normalized Cmax of daidzein (P = 0.002) and similarly the normalized AUC0 --> infinity and Cmax of genistein (P = 0.002) from soya-based capsules were higher than that from soya-based cheese. In conclusion, this work completes studies on isoflavone bioavailability and presents new data regarding isoflavone concentrations in soya-derivative products. Assuming that isoflavone conjugation patterns do not influence isoflavone bioavailability, this study shows that isoflavones contained in capsules are more bioavailable than those contained in soya-based cheese. Although the supplement is more bioavailable, the relative importance of this is difficult to interpret as there is little evidence that supplements are biologically active in human subjects to date and further studies will be necessary for this specific supplement to prove its efficacy.

Topics: Adult; Biological Availability; Chromatography, High Pressure Liquid; Cross-Over Studies; Dietary Supplements; Enzyme-Linked Immunosorbent Assay; Genistein; Glycine max; Humans; Isoflavones; Male; Phytoestrogens; Soy Foods

We evaluated the effects of soy isoflavone supplementation on hemostasis in healthy postmenopausal women.. In this double-blinded, placebo-controlled study, 47 postmenopausal women 47-66 y of age received 40 mg of soy isoflavone (n = 25) or 40 mg of casein placebo (n = 22) once a day for 6 mo. Levels of factors VII and X, fibrinogen, thrombin-antithrombin complex, prothrombin fragments 1 plus 2, antithrombin, protein C, total and free protein S, plasminogen, plasminogen activator inhibitor-1, and D-dimers were measured at baseline and 6 mo. Urinary isoflavone concentrations (genistein and daidzein) were measured as a marker of compliance and absorption using high-performance liquid chromatography. Baseline characteristics were compared by unpaired Student's t test. Within-group changes and comparison between the isoflavone and casein placebo groups were determined by a mixed effects model.. The levels of hemostatic variables did not change significantly throughout the study in the isoflavone group; however, the isoflavone group showed a statistically significant reduction in plasma concentration of prothrombin fragments 1 plus 2; both groups showed a statistically significant reduction in antithrombin, protein C, and free protein S levels. A significant increase in D-dimers was observed only in the isoflavone group. Plasminogen activator inhibitor-1 levels increased significantly in the placebo group. However, these changes were not statistically different between groups.. The results of the present study do not support a biologically significant estrogenic effect of soy isoflavone on coagulation and fibrinolysis in postmenopausal women. However, further research will be necessary to definitively assess the safety and efficacy of isoflavone.

Topics: Aged; Antithrombin III; Biomarkers; Blood Coagulation; Dietary Supplements; Double-Blind Method; Factor VII; Factor X; Female; Fibrinogen; Fibrinolysis; Genistein; Hemostasis; Humans; Isoflavones; Middle Aged; Patient Compliance; Peptide Fragments; Peptide Hydrolases; Phytoestrogens; Postmenopause; Protein Precursors; Prothrombin

To study the effects of consuming isoflavone aglycone-enriched soymilk fermented by bifidobacteria on urinary excretion of equol with respect to fermentation, daidzein dose, supplementation duration and background diet.. Double-blind crossover pilot study comprising three 14-day supplementation periods separated by a washout.. Victoria University, Melbourne, Australia.. Sixteen postmenopausal women.. SUBJECTS randomized into two groups consuming either fermented (FS) or non-fermented soymilk (NFS), ingested three daily dosages of daidzein via soymilk and collected pooled urine specimens. Daidzein and equol were quantified using high-performance liquid chromatography.. After 14-days supplementation six women (38%) excreted equol (>1 micromol equol/day), including four from the FS group, two of whom were classified as non-producers at day 4. Bifidobacteria ingestion, composition of daidzein and its glucosides, and carbohydrate intake appeared to influence equol formation among equol producers.. Pilot-study group mean urinary equol excretion results provided insufficient evidence (P>0.05) that FS consumption instigates equol production in women predetermined as non-producers.

Topics: Bifidobacterium; Chromatography, High Pressure Liquid; Cross-Over Studies; Dose-Response Relationship, Drug; Equol; Female; Fermentation; Food Handling; Humans; Isoflavones; Middle Aged; Phytoestrogens; Pilot Projects; Postmenopause; Probiotics; Soy Milk

Equol production, isoflavone excretion, and the salivary estradiol profile among 36 females, native Irish Caucasian volunteers following ingestion of 200mL soymilk is reported. The soymilk contained daidzein (73+/-6.7mg) and genistein (86+/-10.2mg). Volunteers provided personal and family medical history. Dietary analysis revealed that all volunteers regularly consumed soy-based or soy-supplemented food products. The mean age, mean age at menarche, and body mass index of volunteers were 46.6+/-12.3 years, 13.1 years and 26.1, respectively. The average number of children per volunteer was 2.13. Twelve (34%) of the volunteers were found to be first-degree relatives of breast cancer patients. Following consumption of the soymilk, equol was detected in the urine of 18 (51%) of the volunteers. Mean urinary daidzein and genistein concentrations during the hours following soymilk ingestion were 13.5 and 16.7microg/mg creatinine, respectively, however, some volunteers excreted little (less than 4.0microg/mg) or no isoflavone. Salivary estradiol in most (24) volunteers had decreased from 51.5+/-28.67pmol/L pre-ingestion to 29.75+/-16.13pmol/L 5h after drinking the soymilk. However, the salivary estradiol in 12 subjects (34%) increased from 33.76+/-13.4pmol/L to 137.4+/-65.64pmol/L over the same period. Individuals whose salivary estradiol increased had significantly less children (1.58 (P<0.05)), were more likely to (a) return urine samples with low isoflavone content (50.3% compared to 25%), (b) to be equol producers (67% compared to 41.7%), and (c) to be first-degree relatives of breast cancer patients (41.7% compared to 25%). Volunteers who reported a first-degree link to breast cancer were more likely to have a higher body mass index (29.0 compared to 26.1 (P<0.05)), to be equol producers (75% compared to 51%), and to excrete isoflavones in low quantities only (60% compared to 50%). First-degree relatives also had fewer children (1.75 (P<0.05)). The results indicate a significant, distinctive variation in equol production, isoflavone excretion and salivary estradiol profile among individual volunteers following ingestion of soymilk.

Topics: Adult; Aged; Chromatography, High Pressure Liquid; Dietary Supplements; Equol; Estradiol; Female; Genistein; Humans; Isoflavones; Middle Aged; Phytoestrogens; Plant Preparations; Saliva; Soy Milk; White People

Equol, a microbial metabolite of daidzein, has been hypothesized as a clue to the effectiveness of soy and its isoflavones but is excreted by only 33% of Caucasians. Microbial and dietary factors associated with the ability to harbor equol-producing bacteria were studied in a randomized dietary intervention trial with 100 healthy postmenopausal women. After a 4-d baseline period, subjects delivered first-void urine, fecal, and breath samples. During the 5-d treatment period, 3 portions of either soymilk or soy germ containing 28.51 and 37.99 mg isoflavone aglycone equivalents/portion, respectively, were administered daily, and on the last day, 24-h urine samples were collected. The urinary recoveries of genistein and daidzein from soymilk were significantly higher than those from soy germ tablets. Because the proportion of equol:(daidzein + metabolites) in the urine did not differ between the treatment groups, subjects were pooled and classified into poor, moderate, and strong equol producers based on this criterion. The strong equol producer phenotype correlated negatively [in vivo, r = -0.478 (-0.256 to -0.893), P = 0.021; in vitro, r = -0.576 (-0.350 to -0.949), P = 0.030] with Clostridium coccoides-Eubacterium rectale counts and positively [in vivo, r = 1.158 (0.971-1.380), P = 0.048; in vitro, r = 1.156 (1.007-1.327), P = 0.039] with the abundance of sulfate-reducing bacteria. Furthermore, persons with a higher PUFA [in vivo, r = 2.150 (1.058-4.371), P = 0.034; in vitro, r = 2.131 (1.144-3.967), P = 0.017] and alcohol [in vivo, r = 1.166 (0.721-1.887), P = 0.050; in vitro, r = 1.850 (1.215-2.817), P = 0.004] intake were more likely to be strong equol producers. Finally, we validated the daidzein metabolism by fecal cultures as screening assay to identify equol producers without dietary intervention.

Topics: Adult; Aged; Diet; Equol; Feces; Female; Gastrointestinal Tract; Health; Humans; Isoflavones; Middle Aged; Phenotype; Phytoestrogens; Postmenopause; Soy Milk

Phytoestrogens have been suggested to reduce the risk of prostate cancer (CaP), but no data exists on how oral phytoestrogen supplementation influences phytoestrogen concentrations in prostate tissue.. Forty men with CaP, assigned for radical prostatectomy, received 240 mg of clover phytoestrogens or placebo daily for a 2-week period before their operation in a prospective and randomized study. Phytoestrogens were measured in plasma and prostate tissue by time-resolved fluoroimmunoassay (TR-FIA).. All patients had low baseline phytoestrogen concentrations and only 35% had a detectable plasma concentration of equol. Oral supplementation with phytoestrogens induced a statistically significant (P<0.001) 23- and 7-fold increase in prostate tissue concentrations of the phytoestrogens genistein and daidzein, respectively. Supplemented patients demonstrated prostate tissue genistein and daidzein concentrations that were over twofold higher than their plasma. Interestingly, even though the placebo group did not receive phytoestrogen challenge, they also demonstrated twofold prostate tissue genistein and daidzein concentrations compared to their plasma values, suggesting that the prostate can concentrate available phytoestrogens. In addition, after the supplementation, 90% of the supplemented patients had a detectable plasma equol concentration.. We conclude that prostate tissue can concentrate genistein and daidzein. Significant elevation of intraprostatic genistein and daidzein concentrations can be achieved with a short-term dietary phytoestrogen supplementation.

Topics: Administration, Oral; Aged; Dietary Supplements; Genistein; Humans; Isoflavones; Male; Middle Aged; Phytoestrogens; Phytotherapy; Placebos; Prostatectomy; Prostatic Neoplasms

To observe the effect of daidzein on serum inflammatory factors of senile patients with coronary heart disease (CHD).. Forty senile patients with CHD were randomly assigned into the control group and the daidzein group, 20 in each group. Patients in both groups were treated with conventional medicine, while those in the daidzein group were given daidzein Tablets additionally for 6 weeks. The levels of resting heart rate (RHR), blood pressure (BP), fasting plasma glucose (FPG), blood lipids and inflammatory factors, including hsCRP, IL-6 and TNF-alpha, were measured before and after treatment.. In the control group, the levels of RHR, BP and hsCRP changed after conventional medicinal treatment (P < 0.05 or P < 0.01) but other parameters unchanged (P > 0.05). While in the daidzein group, all the parameters measured were decreased in different degrees after 6 weeks treatment (P < 0.05 or P < 0.01), and also showed significant difference as compared with those in the control group respectively (P < 0.05 or P < 0.01).. Daidzein can effectively decrease the levels serum inflammatory factors in senile patients with CHD, the fact proved that isoflavone has anti-inflammatory effect in patients with coronary atherosclerosis.

Topics: Aged; C-Reactive Protein; Coronary Artery Disease; Female; Humans; Interleukin-6; Isoflavones; Male; Middle Aged; Phytoestrogens; Tumor Necrosis Factor-alpha

7-Hydroxy-3-(4'-hydroxyphenyl)-chroman (S-equol) is a specific end-metabolite formed in the biotransformation of the dietary soy isoflavones daidzin and daidzein by intestinal bacteria. The frequency of equol production varies among individuals and populations, and it is suggested that the efficacy of soy foods differs depending on the ability of an individual to produce equol. To develop a standardized approach to define equol-producer status that can be universally adopted to differentiate these 2 distinct populations, we measured isoflavones in serum and urine collected from a cohort of 41 healthy adults, comprising 29 vegetarians and 12 nonvegetarians, after consuming 2 x 250 mL/d soy milk on 3 consecutive days. Serum and urinary daidzein and S-equol concentrations were analyzed by MS. Serum S-equol and daidzein concentrations ranged from 10.3-139 nmol/L (2.5-33.6 microg/L) and 16-1401 nmol/L (4.0-356 microg/L), respectively, whereas in urine the corresponding concentrations ranged from 16-12,574 nmol/L (4-3043 microg/L) and 539-26,834 nmol/L (137-6816 microg/L), respectively. The log10-transformed urinary S-equol:daidzein ratio provided a clearer distinction of equol-producer status than the absolute serum or urinary S-equol concentrations because it is independent of isoflavone intake and minimizes interindividual variation in isoflavone pharmacokinetics or differences in analytical methodologies. A threshold value for the log10-transformed urinary S-equol:daidzein ratio of -1.75 provided a demarcation to define equol-producer status. The frequency of equol producers in the vegetarians was 59%, similar to the reported frequency in Japanese adults consuming soy, and much higher than for nonvegetarian adults (25%), suggesting that dietary components other than soy influence S-equol synthesis by intestinal bacteria.

Topics: Adult; Diet, Vegetarian; Equol; Female; Gas Chromatography-Mass Spectrometry; Humans; Isoflavones; Male; Middle Aged; Phytoestrogens; Soybean Proteins

The aim of this study was to determine whether equol excretion status and plasma hormone and leptin concentrations can be influenced by consumption of a probiotic supplement. A secondary focus was to investigate whether male equol excretors have a hormone profile consistent with reduced prostate cancer risk.. The design was a randomized, single-blinded, placebo-controlled, parallel-arm trial.. Thirty-one (31) of the initially enrolled 39 subjects, 18 to 37 years old, completed all study requirements.. Subjects consumed either probiotic capsules (containing Lactobacillus acidophilus and Bifidobacterium longum) or placebo capsules for 2 months. Fasting plasma concentrations of testosterone (T), dihydrotestosterone (DHT), androstanediol glucuronide (AAG), androstenedione (A), dehydroepiandrosterone sulfate (DHEAS), sex hormone-binding globulin (SHBG), and leptin were measured on days 1 and 57. Urinary excretion of genistein, glycitein, daidzein, O-desmethylangolensin (O-Dma), and equol was measured on days 4 and 61 following a 4-day soy challenge.. Probiotic consumption did not significantly alter equol excretor status, plasma hormone, or leptin concentrations in these subjects. At baseline, there were no differences in plasma hormone concentrations between equol excretors and nonexcretors; however, the low number of equol excretors included in this study limits the strength of this finding.. The 2-month intervention with probiotic capsules did not significantly alter equol excretion, plasma hormone, or leptin concentrations in these subjects. A secondary finding was that male equol excretors in this study did not exhibit a hormone profile consistent with reduced prostate cancer risk, although this result should be interpreted with caution.

Topics: Adrenal Cortex Hormones; Adult; Androstane-3,17-diol; Androstenedione; Bifidobacterium; Dehydroepiandrosterone Sulfate; Dihydrotestosterone; Equol; Genistein; Hormones; Humans; Isoflavones; Lactobacillus acidophilus; Leptin; Male; Phytoestrogens; Pilot Projects; Probiotics; Sex Hormone-Binding Globulin; Single-Blind Method; Testosterone

The effect of a phyto-oestrogen, daidzein, on the laying performance of Shaoxing female ducks was examined in three experiments performed at different stages of the egg production cycle. Egg-laying rate, egg weight, egg composition, feed conversion ratio, hatchability characteristics of eggs and body weight, ovary and oviduct weight, as well as changes in serum concentrations of T3, T4 and E2 were recorded as response criteria. In the first experiment, 68 young ducks, 100 d of age, were given a basal diet (maize-soybean meal) with or without 3 mg of daidzein/kg diet for 42 d. Daidzein did not affect the onset of lay but apparently decreased egg-laying rate and mean egg weight as well as the feed conversion ratio. In the second experiment, 240 breeding ducks, 402 d of age, were allotted at random to three groups and given the basal diet containing daidzein at 0 (control), 3 (Da1) and 5mg/kg (Da2) for 35d. Egg-laying rate, mean egg weight and feed conversion ratio increased in both Da1 and Da2 groups. However, an adverse effect of daidzein on fertility and hatchability was observed. In the third experiment, 320 breeding ducks, 415 d of age, were fed on the basal diet with or without 5mg of daidzein/kg diet for 63 d. Egg-laying rate increased by 7.7%, average egg weight tended to increase, whereas yolk/albumen ratio decreased. Daidzein-treated ducks had higher body weight and oviduct weight compared with their controls. Elevated plasma T4 and E2 concentrations accompanied these phenotypic changes, but serum T3 was not affected. It is suggested that daidzein exerts divergent effects on the egg-laying performance of Shaoxing ducks under different physiological conditions and this action is dose-dependent. The changes in circulating E2 imply possible participation of endogenous oestrogen in the mechanism of daidzein action.

Topics: Animals; Drug Administration Schedule; Ducks; Female; Fertility; Isoflavones; Oviposition; Phytoestrogens

Up to 80 % of the Western female population experience premenstrual syndrome (PMS). Long-term pharmacological therapy is unacceptable to most women, and is not warranted for moderate symptoms. Nutritional therapies are popular, but lack a clear evidence base. Anecdotal evidence suggests beneficial effects of soy isoflavones because of their influence on endogenous oestrogen and actions on specific tissues. The effect of isolated soya protein (ISP) containing 68 mg/d (aglycone equivalents) soy isoflavones (IF) on premenstrual symptom severity was studied in a seven-menstrual cycle, double-blind, placebo-controlled, crossover intervention study in twenty-three women with prospectively confirmed PMS aged 18-35 years and BMI 19-30 kg/m(2). ISP containing IF or milk protein placebo was consumed for two complete menstrual cycles. ISP containing IF (genistein, daidzein, equol) were measured in 24 h urine samples. After two cycles of ISP containing IF intervention, total symptoms (F(2,36) 8.20, P=0.000) and physical symptoms (F(2,36) 8.18, P=0.000) were significantly reduced compared with baseline after both active and placebo treatments, although differences between active and placebo treatment were non-significant. Specific premenstrual symptoms, headache (F(2,32) 4.10, P=0.026) and breast tenderness (F(2,32) 4.59, P=0.018), were reduced from baseline after soy IF, but not milk protein placebo. Cramps (F(2,32) 4.15, P=0.025) and swelling (F(2,32) 4.64, P=0.017) were significantly lower after active treatment compared with placebo. Concentrations of genistein and daidzein were increased following soy IF consumption, but equol production did not enhance symptom reduction. The present study showed that ISP containing IF may have potential to reduce specific premenstrual symptoms via non-classical actions.

Topics: Adolescent; Adult; Breast Diseases; Cross-Over Studies; Double-Blind Method; Female; Genistein; Glycine max; Headache; Humans; Isoflavones; Phytoestrogens; Premenstrual Syndrome; Prospective Studies

We investigated the effects of consuming an isoflavone aglycone-enriched soya milk containing viable bifidobacteria on urinary isoflavone excretion and percentage recovery. Sixteen postmenopausal women were randomly divided into two groups to consume either fermented or non-fermented soya milk. Each group participated in a double-blind, crossover study with three 14 d supplementation periods, separated by a 14 d washout. Subjects ingested three daily dosages of isoflavone via the soya milk and collected four 24 h pooled urine specimens per supplementation period. Soya milks were prepared with soya protein isolate and soya germ, followed by fermentation with bifidobacteria. Isoflavone levels were quantified using HPLC. Non-fermented soya milks at 20, 40 and 80 mg isoflavone/200 ml contained 10 %, 9 % and 7 % aglycone, respectively, with their fermented counterparts containing 69 %, 57 % and 36 % aglycone (P<0.001). A trend to a greater percentage urinary recovery of daidzein and glycitein was observed among women consuming fermented soya milk at a dosage of 40 mg isoflavone (P=0.13). A distinct linear dose response for the fermented soya milk group (R2=0.9993) compared with the non-fermented group (R2=0.8865) suggested less interindividual variation in isoflavone absorption. However, total urinary isoflavone excretion was similar for both groups (P>0.05), with urinary isoflavone recovery at approximately 31 %. Increasing the isoflavone dosage correlated positively with its urinary excretion, but urinary percentage recovery of isoflavone was inversely related to dosage level. Hence, a modest dosage ranging from 20 to 30 mg/d may provide the most bioavailable source of isoflavone, regardless of whether it is via an aglycone-rich fermented soya milk or a glucoside-rich soya milk.

Topics: Bifidobacterium; Biological Availability; Body Mass Index; Body Weight; Cross-Over Studies; Double-Blind Method; Eating; Enzyme Inhibitors; Female; Fermentation; Genistein; Humans; Isoflavones; Middle Aged; Phytoestrogens; Postmenopause; Probiotics; Soy Milk

Soy-derived phytoestrogens may exert several health-beneficial effects. Although plasma and urine levels of these compounds after ingestion have been thoroughly investigated, little is known about their tissue distribution, which is particularly important for tissues with high endogenous estrogen and estrogen receptor concentrations. We aimed to investigate the concentrations of genistein, daidzein, and equol in human breast tissue homogenate and to compare these with the corresponding values in serum and urine.. A randomized, double-blind, placebo-controlled study was undertaken to evaluate the concentrations of soy-derived phytoestrogens achieved in breast tissue homogenate, serum, and urine after ingestion of either a soy-based food supplement (n = 9) or a placebo tablet (n = 19) for 5 consecutive evenings before aesthetic breast surgery. To account for the heterogeneity of the breast tissue samples, markers for cellularity, epithelial content, blood vessel content, and total fat were determined.. Urine concentrations of genistein, daidzein, and equol were significantly higher in the soy-supplemented subjects than in the subjects ingesting the placebo (P <.05). Only genistein was found to be significantly higher in serum of the soy group than in the placebo group, and no significant differences were found in breast tissue homogenate concentrations of all analytes between the 2 groups.. Intake of soy-based food supplements for 5 consecutive days did not result in significantly higher genistein, daidzein, and equol concentrations in breast tissue homogenate when compared with the placebo group. The concentrations were in the low nanomolar range, whereas in the corresponding serum samples, concentrations were a hundred-fold higher.

Topics: Adult; Body Fluids; Breast; Dietary Supplements; Double-Blind Method; Equol; Genistein; Humans; Isoflavones; Middle Aged; Phytoestrogens; Plant Preparations; Soy Foods; Tissue Distribution

A phase I clinical trial was conducted to determine the safety, pharmacokinetic parameters, and efficacy of orally administered isoflavones (genistein and daidzein, potential cancer chemotherapeutic agents) over a 3-mo period in men with prostate neoplasia. Twenty men, ages 40 and above, with stage B, C, or D adenocarcinoma of the prostate were treated with a multiple-dose regimen of a soy isoflavone formulation (delivering approximately 300 or 600 mg/day genistein and half this much daidzein) for 84 days. The delivered dose of isoflavones was more than 10-fold higher than that typically taken by prostate cancer patients. In men with prostate cancer, relatively minor side effects of chronic isoflavone treatment were observed including some estrogenic effects (breast changes, increased frequency of hot flashes). Serum dehydroepiandrosterone was decreased by 31.7% (P = 0.0004) at the end of treatment. Except for those subjects whose prostate-specific antigen (PSA) values were below 0.4 ng/ml, subjects had a history of increasing PSA levels prior to the trial. This increase continued during the trial both while on soy isoflavones and after treatment was discontinued. On average the rate of rise accelerated after soy isoflavones were discontinued, but that difference did not attain statistical significance. Genistein and daidzein were rapidly cleared from plasma and excreted in urine. Pharmacokinetic data for chronic dose administration were similar to single-dose administration for the isoflavones investigated except that we observed slightly longer circulation time for daidzein.

Topics: Adenocarcinoma; Administration, Oral; Aged; Aged, 80 and over; Antineoplastic Agents, Phytogenic; Dose-Response Relationship, Drug; Genistein; Glycine max; Half-Life; Humans; Isoflavones; Male; Middle Aged; Phytoestrogens; Prostatic Neoplasms; Safety; Treatment Outcome

To determine the effects of diets rich in soy and linseed compared with a control diet on biochemical markers of prostate cancer in men diagnosed with prostate cancer.. Twenty-nine men diagnosed with prostate cancer and scheduled to undergo a radical prostatectomy were randomized to one of three groups: soy (high phytoestrogen), soy and linseed (high phytoestrogen), or wheat (low phytoestrogen). A bread was specially manufactured to incorporate 50 g of heat-treated (HT) soy grits or 50 g of HT soy grits and 20 g of linseed as part of the study participant's daily diet. Baseline and preoperative levels of prostate-specific antigen (PSA), free PSA, testosterone, sex hormone-binding globulin, free androgen index, and dihydrotestosterone were measured.. Statistically significant differences were detected between the HT soy grits group and the control wheat group for the percentage of change in total PSA (-12.7% versus 40%, P = 0.02) and the percentage of change in free/total PSA ratio (27.4% versus -15.6%, P = 0.01); and between the HT soy grits group and the HT soy grits and linseed group for the percentage of change in free androgen index (16.4% versus -15.5%, P = 0.04) and the percentage of change in free/total PSA ratio (27.4% versus -10%, P = 0.007).. The data from this study indicate that a daily diet containing four slices of a bread rich in HT soy grits favorably influences the PSA level and the free/total PSA ratio in patients with prostate cancer. This work provides some evidence to support epidemiologic studies claiming that male populations who consume high phytoestrogen diets have a reduced risk of prostate cancer development and progression.

Topics: Adenocarcinoma; Aged; Androgens; Biomarkers, Tumor; Bread; Dihydrotestosterone; Double-Blind Method; Flax; Genistein; Gonadal Steroid Hormones; Humans; Isoflavones; Male; Middle Aged; Neoplasm Proteins; Phytoestrogens; Phytotherapy; Prostate-Specific Antigen; Prostatic Neoplasms; Sex Hormone-Binding Globulin; Soy Foods; Testosterone

Phytoestrogens are thought to be beneficial to vascular health. Possible mechanisms of action could involve C-reactive protein (CRP), endothelial E-selectin, and nitric oxide. We therefore designed a randomized, placebo-controlled, double-blind trial in which we studied the effects of isoflavonoids on CRP, E-selectin, and nitrate-nitrite (NO(x); reflecting the release of nitric oxide) in postmenopausal women. Fifty-six postmenopausal women (FSH > 30 U/liter) with a history of breast cancer used (in a randomized order) phytoestrogen (114 mg isoflavonoids) or placebo tablets daily for 3 months; the treatment regimens were crossed over after a 2-month washout period. The serum levels of CRP and E-selectin, and plasma levels of NO(x) were measured before and on the last day of each treatment. The phytoestrogen regimen did not affect the levels of either CRP (P = 0.584) or NO(x) (P = 0.270), but the levels of E-selectin were reduced by 4.0% (2.9 ng/ml; P = 0.031) during phytoestrogen use and by 2.2% (1.3 ng/ml; P = 0.023) during placebo use. No difference was found at any marker at 3 months between the groups. In conclusion, our data, suggesting neutral effects of phytoestrogens on CRP, E-selectin, and nitric oxide, fail to support a vasoprotective role of phytoestrogens.

Topics: Adult; Aged; Breast Neoplasms; C-Reactive Protein; Cross-Over Studies; E-Selectin; Estrogens, Non-Steroidal; Female; Humans; Isoflavones; Middle Aged; Nitrates; Phytoestrogens; Phytotherapy; Plant Preparations

In order to evaluate the efficacy of soya isoflavones (genistein and daidzein) in the treatment of the principal menopausal disorders, a double blind randomized study was performed on a sample of 50 women (with an average age of 53.3 +/- 3.1 years) with Climacteric syndrome referred to the I Clinica Ostetrica e Ginecologica, Policlinico Umberto I, Roma. The research protocol involved the random subdivision of the enrolled sample into two groups of 25 women, group 1 (with an average age of 53.3 +/- 3.5 years, and an average menopausal age of 51.6 +/- 1.8 years) and group 2 (with an average age of 53.1 +/- 2.9 years, and an average menopausal age of 51.3 +/- 1.2 years), who were to receive treatment for three months with the product being studied and with a placebo. After the three-month period, as an additional check, the group initially treated with the placebo would move to the phyto-oestrogens and viceversa. All of the patients were subjected to a series of clinical and instrumental examinations and were asked to fill in a questionnaire concerning their complaints, at the start, at halfway (third month) and at the end (sixth month) of the trial. The results of the evaluation of the questionnaires performed on the 47 patients who had completed the trial showed, in the first three months, an improvement in the symptoms (hot flushes) in 11 patients treated with phyto-oestrogens against 6 patients from the group that received only the placebo. In the second three-month period the hot flushes reappeared in 4 of the 11 patients who had previously seen improvements and had then passed to the placebo. In contrast, the group that passed to the phyto-oestrogens, after treatment with the placebo, experienced the disappearance of hot flushes in 11 women, including the 6 who had already improved in the first three months. There was no significant reduction in anxiety, insomnia or vaginal dryness. None of the enrolled patients indicated complaints linked to the treatment. It can be concluded that the use of a product based on phyto-oestrogens, such as the one experimented, can lead to a significant reduction in some of the disorders linked with the menopause, especially hot flushes.

Topics: Climacteric; Cross-Over Studies; Double-Blind Method; Estrogens, Non-Steroidal; Female; Genistein; Hot Flashes; Humans; Isoflavones; Middle Aged; Phytoestrogens; Plant Preparations; Surveys and Questionnaires; Treatment Outcome

Genistein, a phytoestrogen, may have estrogenic cardioprotective actions. We investigated whether genistein influences endothelium-dependent vasodilation in forearm vasculature of healthy human subjects and compared the effects of genistein with those of 17beta-estradiol.. The brachial arterial was cannulated with a 27-gauge needle for drug infusion. Forearm blood flow responses were measured with strain-gauge plethysmography. Genistein (10 to 300 nmol/min, each dose for 6 minutes) produced a dose-dependent increase in forearm blood flow from 3.4+/-0.3 to 9.6+/-1.3 mL x min(-1) x 100 mL forearm(-1) (mean+/-SEM) in men (n=9, P:<0.0001 by ANOVA). The mean forearm venous concentration of genistein during infusion of the highest dose was 1.8+/-0.3 micromol/L in 6 additional men. Genistein produced a similar increase in blood flow in premenopausal women. Daidzein, another phytoestrogen, was ineffective, but equimolar concentrations of 17beta-estradiol caused similar vasodilation to genistein. Responses to genistein and 17beta-estradiol were inhibited to the same degree by the NO synthase inhibitor N:(G)-monomethyl-L-arginine. A threshold dose of genistein potentiated the endothelium-dependent vasodilator acetylcholine but not the endothelium-independent vasodilator nitroprusside.. Genistein causes L-arginine/NO-dependent vasodilation in forearm vasculature of human subjects with similar potency to 17beta-estradiol and potentiates endothelium-dependent vasodilation to acetylcholine.

Topics: Acetylcholine; Adult; Blood Vessels; Cardiotonic Agents; Dose-Response Relationship, Drug; Enzyme Inhibitors; Estradiol; Estrogens, Non-Steroidal; Female; Forearm; Genistein; Humans; Isoflavones; Male; Middle Aged; Nitric Oxide; Nitric Oxide Donors; Nitroprusside; omega-N-Methylarginine; Phytoestrogens; Plant Preparations; Vasodilation; Vasodilator Agents

Animal studies and human intervention trials have demonstrated the cancer chemopreventive properties of plant phytoestrogens, and phytoestrogen supplements are now widely available 'over-the-counter'. However, consumption of phytoestrogen-rich diets can cause impaired fertility and reproductive tract disorders in some animals and the apparent decline in human sperm quality over recent decades may be related to increased exposure to environmental endocrine disruptors. The present study determines the effects of a short-term phytoestrogen supplement on semen quality and serum sex steroid and gonadotrophin levels in human males. Healthy volunteers took a supplement containing 40 mg of isoflavones daily for 2 months and donated blood and semen samples monthly for 2 months before and 4 months after supplementation. Semen samples were analysed for ejaculate volume, sperm concentration, total sperm count, motility and morphology. Blood samples were analysed for sex hormone and gonadotrophin levels and phytoestrogen concentrations, and testicular volume was measured using an orchidometer. The phytoestrogen supplement increased plasma genistein and daidzein concentrations to approx. 1 microM and 0.5 microM respectively; yet, there was no observable effect on endocrine measurements, testicular volume or semen parameters over the study period. This is the first study to examine the effects of a phytoestrogen supplement on reproductive health in males. We conclude that the phytoestrogen dose consumed had no effect on semen quality.

Topics: Adult; Dietary Supplements; Estrogens, Non-Steroidal; Fertility; Genistein; Glycine max; Gonadal Steroid Hormones; Gonadotropins, Pituitary; Humans; Isoflavones; Male; Phytoestrogens; Plant Preparations; Semen; Sperm Count; Sperm Motility; Spermatozoa; Testis

The transdermal absorption of the isoflavones, daidzein and genistein, applied on the skin in olive oil was studied in vivo. The concentrations of the isoflavones and their metabolites were monitored in plasma and urine by GC-MS methods. It was found that the concentration of genistein in plasma was 3-fold higher than the plasma concentration of daidzein. In contrast, daidzein excretion was 2-3-fold higher than that of genistein in urine. The excretion rate of the studied phytoestrogens in urine and their concentration in plasma was significantly decreased after repeated transdermal application. The urinary recovery of administered daidzein and genistein after the first application was 15.9% and 7.7%, respectively and this dropped to 1.6% and 0.7% after the second application. The results obtained might suggest that daidzein and genistein are captured in the skin following repeated transdermal application.

Topics: Administration, Cutaneous; Adult; Estrogens, Non-Steroidal; Female; Gas Chromatography-Mass Spectrometry; Genistein; Humans; Isoflavones; Ointment Bases; Olive Oil; Phytoestrogens; Plant Oils; Plant Preparations; Skin Absorption; Suspensions

To analyze the impact of soy protein dietary supplements containing phytoestrogens on menopausal symptoms in healthy postmenopausal women.. A double-blind, placebo-controlled trial was conducted in 94 healthy postmenopausal women aged 50-75 years, with 44 randomized to soy supplements containing 118 mg of isoflavones (daidzein, genistein, glycitein and their respective glycosides), and 50 to an identically presented casein placebo. A validated questionnaire on menopausal symptoms was administered at baseline and at 3 months of treatment. Compliance was assessed by high-performance liquid chromatography assay of urinary phytoestrogens. Statistical analysis was completed using non-parametric statistical methods and multivariate analysis.. At baseline 80% of women recruited were experiencing menopausal symptoms, although symptom severity was mild. Those consuming phytoestrogen supplements had 13- and 17-fold increases in urinary excretion of genistein and daidzein, respectively, with no change in the placebo group. Active soy supplements did not significantly alter either individual symptoms or specific symptom category scores when compared to placebo. Within-group comparisons revealed that the active group reported a significant improvement in vaginal dryness (p = 0.01), libido (p = 0.009), facial hair (p = 0.04) and dry skin (p = 0.027). However, similarly, those on placebo reported an improvement in libido (p = 0.015), facial hair (p = 0.014) and dry skin (p = 0.011) but not vaginal dryness.. In this group of 94 older postmenopausal women with a high frequency of mild menopausal symptoms, 3 months of soy supplements containing phytoestrogens did not provide symptomatic relief compared with placebo.

Topics: Aged; Double-Blind Method; Estrogens, Non-Steroidal; Female; Genistein; Hirsutism; Humans; Isoflavones; Libido; Menopause; Middle Aged; Phytoestrogens; Placebos; Plant Preparations; Postmenopause; Skin Diseases; Soybean Proteins; Surveys and Questionnaires; Treatment Outcome; Vaginal Diseases

An association has been reported between consumption of a high soy diet and a low incidence of breast cancer within populations of Southeast Asia. Phytoestrogens present in soy act as partial estrogen agonists or antagonists and can inhibit breast cancer cell proliferation in vitro. The effect of 14-day dietary soy supplementation with 60 g (45 mg isoflavones) on the normal breast of 84 premenopausal patients was determined. Serum concentrations of the isoflavanoids, genistein, daidzein, and equol, were raised in patients after soy supplementation (P < or = 0.025). Nipple aspirate (NA) levels of genistein and daidzein were higher than paired serum levels, both before (P < 0.001 and P = 0.001, respectively) and after soy supplementation (P < 0.001 and P = 0.049, respectively); however, there was no significant increase in NA isoflavone levels in response to soy. NA levels of apolipoprotein D were significantly lowered and pS2 levels raised in response to soy supplementation (P < or = 0.002), indicative of an estrogenic stimulus. No effect of soy supplementation on breast epithelial cell proliferation, estrogen and progesterone receptor status, apoptosis, mitosis, or Bcl-2 expression was detected. In conclusion, short term dietary soy has a weak estrogenic response on the breast, as measured by nipple aspirate apolipoprotein D and pS2 expression. No antiestrogenic effect of soy on the breast was detected.

Topics: Adult; Apolipoproteins; Apolipoproteins D; Body Fluids; Chromans; Diet; Equol; Estrogens, Non-Steroidal; Female; Genistein; Glycine max; Humans; Isoflavones; Nipples; Phytoestrogens; Plant Preparations; Proteins; Soybean Proteins; Suction; Trefoil Factor-1; Tumor Suppressor Proteins

The aim of this study was to test the hypothesis that increased dietary intake of phytoestrogens reduces the health impact of the menopause. To test this hypothesis, a double-blind, randomized, entry-exit, cross-over study was conducted to assess the effects of three dietary manipulations--soy and linseed diets (high in phytoestrogens) and a wheat diet (low in phytoestrogens). Postmenopausal women were recruited and randomly assigned to one of the three dietary regimens. Urinary phytoestrogen concentrations, hot flush rate, vaginal smears, bone mineral density and bone mineral content were assessed for two 12-week periods. Comparative analysis showed no significant differences, but, when analyzed separately, groups consuming high phytoestrogen diets had between 10 and 30 times higher urinary excretion of phytoestrogens compared to those consuming the low phytoestrogen diet (p < 0.01). Study participants consuming soy, linseed and wheat diets had a 22% (not significant, n.s.), 41% (p < 0.009) and 51% (p < 0.001) reduction in hot flush rate; a 103% (p < 0.04), 5.5% (n.s.) and 11% (n.s.) increase in vaginal cytology maturation index; and a 5.2% (p < 0.04), 5.2% (n.s.) and 3.8% (n.s.) increase in bone mineral content, respectively. No changes were detected in bone mineral density. The differential effects of high phytoestrogen dietary manipulations on outcomes may represent tissue-specific responses to isoflavones and lignans contained in soy and linseed, respectively. Whilst health outcome measures were not significantly different between groups, the data obtained from separate analysis suggest that phytoestrogens in soy and linseed may be of use in ameliorating some of the symptoms of menopause. Furthermore, the significant decrease in hot flush rate in the wheat group cannot be attributable to phytoestrogens measured in this study. Due to subject variability, larger studies are still needed to evaluate population benefit.

Topics: 4-Butyrolactone; Aged; Bone Density; Diet; Double-Blind Method; Estrogens, Non-Steroidal; Female; Genistein; Glycine max; Hot Flashes; Humans; Isoflavones; Lignans; Linseed Oil; Middle Aged; Phytoestrogens; Plant Preparations; Postmenopause; Triticum

The effects of consuming a soy protein isolate beverage powder (60 g/d for 28 d) vs. a casein supplement was evaluated in 20 male subjects who were randomly allocated into the two groups. A dramatic rise in plasma isoflavone concentrations was observed after supplementation in the soy protein group, the levels reaching 907 +/- 245 nmol/L for genistein (a 110-fold increase) and 498 +/- 102 nmol/L for daidzein (a 150-fold increase) as measured by isotope dilution gas chromatography - mass spectrometry. These concentrations are higher than previously reported for the plasma of Japanese subjects consuming a traditional diet (276 nmol/L and 107 nmol/L, respectively). No significant differences in collagen- or 9,11-dideoxy-11alpha, 9alpha-epoxymethanoprostaglandin F2alpha (U46619)-induced platelet aggregation were observed in platelet-rich plasma from the two groups; the increase in plasma isoflavonoids from soy protein supplementation is not sufficient to significantly inhibit platelet aggregation ex vivo. Similarly, plasma total and HDL-cholesterol were not affected by protein supplementation, possibly because the men were normocholesterolemic at entry. Analysis of plasma phospholipid polyunsaturated fatty acid composition showed no differences between soy protein and casein supplementation. Previous investigations reported a significant alteration in fatty acid status in animals fed soy protein relative to those fed casein. The present studies indicate that although soy protein supplementation to a typical Western diet can increase plasma concentrations of isoflavones, this may not necessarily be sufficient to counter heart disease risk factors such as high plasma cholesterol and platelet aggregation.

Topics: Adult; Caseins; Cholesterol; Cholesterol, HDL; Collagen; Estrogens, Non-Steroidal; Fatty Acids; Gas Chromatography-Mass Spectrometry; Genistein; Humans; Isoflavones; Lipids; Male; Phospholipids; Phytoestrogens; Plant Preparations; Plant Proteins, Dietary; Platelet Aggregation; Soybean Proteins

To compare the effects of consumption of fermented and unfermented soy products on excretion of urinary isoflavonoid phytoestrogens and lignans in healthy men.. A randomized, crossover trial consisting of two 9-day feeding periods following 5 days of baseline data collection.. Healthy men, aged 20 to 40 years, were recruited from the University of Minnesota Twin Cities community. Of the 22 subjects who began the study, 17 completed all feeding periods.. Fermented soy product (112 g tempeh) or unfermented soy (125 g soybean pieces) was consumed during each controlled feeding period.. Urine samples collected while subjects consumed their habitual diets and on the last 3 days of each feeding period were analyzed for isoflavonoid and lignan content by isotope dilution gas chromatography-mass spectrometry.. Comparisons of isoflavonoid and lignan excretion were analyzed using the general linear model procedure. Orthogonal contrasts were used to determine treatment differences of interest.. Urinary excretion of isoflavonoids (equol, O-desmethylangolensin [O-DMA], daidzein, genistein) was higher and excretion of lignans (enterodiol, enterolactone) was lower when subjects consumed soy-supplemented diets than when they consumed their habitual diets (P < .05). Urinary isoflavonoid excretion and lignan excretion were similar when subjects consumed tempeh and soybean pieces diets; however, recovery of daidzein and genistein was significantly higher when subjects consumed the tempeh diet than when they consumed the soybean pieces diet (P < .002). When fed soy, 5 of 17 subjects excreted high amounts of equol. These five subjects tended to excrete less O-DMA and daidzein than the 12 subjects who excreted low amounts of equol (P < .06).. Fermentation of soy decreased the isoflavone content of the product fed but increased the urinary isoflavonoid recovery. This finding suggests that fermentation increases availability of isoflavones in soy.

Topics: 4-Butyrolactone; Adult; Chromans; Cross-Over Studies; Defecation; Diet; Equol; Estrogens; Estrogens, Non-Steroidal; Fermentation; Food Handling; Genistein; Glycine max; Humans; Isoflavones; Lignans; Male; Phytoestrogens; Plant Preparations; Taste

Equol, which is produced by enteric bacteria from soybean isoflavones, has a chemical structure similar to estrogen. Both in vivo and in vitro studies have shown the beneficial metabolic effects of equol. However, its effects on type 2 diabetes remain unclear. We investigated the association between the equol producers/non-producers and type 2 diabetes.. The participants included 147 patients with type diabetes mellitus aged 70-89 years, and 147 age- and sex-matched controls. To ascertain the equol producers or non-producers, we used the comparative logarithm between the urinary equol and daidzein concentrations (cut-off value -1.75).. The urinary equol concentration was significantly lower in the diabetes group compared with the non-diabetes group (P = 0.01). A significant difference in the proportion of equol producers was observed among all participants (38.8% in the diabetes group and 53.1% in the non-diabetes group; P = 0.01). The proportion of equol producers among women was significantly lower in the diabetes group (31.4%) than in the non-diabetes group (52.8%; P < 0.01). Additionally, the frequency of dyslipidemia in female equol producers was significantly lower than that in female non-equol producers (P < 0.01). Among men, no such differences were observed. We found a significant positive correlation between the urinary equol and daidzein concentrations among equol producers (r = 0.55, P < 0.01).. Our study findings showed that postmenopausal women had a low proportion of equol producers with diabetes and dyslipidemia.

Topics: Aged; Aged, 80 and over; Diabetes Mellitus, Type 2; Dyslipidemias; East Asian People; Equol; Female; Gastrointestinal Microbiome; Glycine max; Humans; Isoflavones; Male; Phytoestrogens; Postmenopause; Sex Factors

Soy isoflavones belong to the group of phytoestrogens and are associated with beneficial health effects but are also discussed to have adverse effects. Isoflavones are intensively metabolized by the gut microbiota leading to metabolites with altered estrogenic potency. The population is classified into different isoflavone metabotypes based on individual metabolite profiles. So far, this classification was based on the capacity to metabolize daidzein and did not reflect genistein metabolism. We investigated the microbial metabolite profile of isoflavones considering daidzein and genistein.. Isoflavones and metabolites were quantified in the urine of postmenopausal women receiving a soy isoflavone extract for 12 weeks. Based on these data, women were clustered in different isoflavone metabotypes. Further, the estrogenic potency of these metabotypes was estimated.. Based on the excreted urinary amounts of isoflavones and metabolites, the metabolite profiles could be calculated, resulting in 5 metabotypes applying a hierarchical cluster analysis. The metabotypes differed in part strongly regarding their metabolite profile and their estimated estrogenic potency.

Topics: Female; Genistein; Glycine max; Humans; Isoflavones; Phytoestrogens; Postmenopause

Phytoestrogens are considered to be endocrine disruptors, since they can alter the endocrine system, thus disturbing many reproductive events. The intake of diets containing a high content of phytoestrogens has increased worldwide in human populations and in domestic animals. Phytoestrogens in maternal blood can pass through the placenta to the fetus in high amounts and can have long-term organizational effects. Mesquite (Prosopis sp) is a leguminous plant widely used to feed several livestock species, and is also used in the human diet. In this study we assessed the effects of exposure to mesquite pod extract during the periconception and pregnancy periods on the reproduction of male and female descendants. The females of three experimental groups received one of the following treatments: 1) vehicle injection; 2) mesquite pod extract or 3) the isoflavone daidzein during the periconception and pregnancy periods. Estrous cyclicity, sexual behavior and hormones, as well as uterine and vaginal epithelia were evaluated in the female descendants. In the males, sexual behavior and hormones, apoptosis in testicular cells and sperm quality were evaluated. In females the following was observed: alterations in estrous cycles, decreased sexual behavior, estradiol and progesterone levels, increased uterine and vaginal epithelia. In males, we observed a decrease in sexual behavior, testosterone and sperm quality, and apoptosis increased in testicular cells. All these effects were similar to those caused by daidzein. These results indicate that prenatal exposure to mesquite pod extract or daidzein, administered to females before and during pregnancy, can disrupt normal organizational-activational programming of reproductive physiology in female and male descendants.

Topics: Animals; Estradiol; Female; Humans; Isoflavones; Male; Phytoestrogens; Plant Extracts; Pregnancy; Prosopis; Rats; Reproduction; Seeds

Phytoestrogens can control high-fat diet-induced hypothalamic inflammation that is associated with severe consequences, including obesity, type 2 diabetes, cardiovascular and neurodegenerative diseases. However, the phytoestrogen anti-neuroinflammatory action is poorly understood. In this study, we explored the neuroprotection mediated by daidzein in hypothalamic neurons by using a membrane-based model of obesity-related neuroinflammation. To test the daidzein therapeutic potential a biohybrid membrane system, consisting of hfHypo GnRH-neurons in culture on PLGA membranes, was set up. It served as reliable in vitro tool capable to recapitulate the in vivo structure and function of GnRH hypothalamic tissue. Our findings highlighted the neuroprotective role of daidzein, being able to counteract the palmitate induced neuroinflammation. Daidzein protected hfHypo GnRH cells by downregulating cell death, proinflammatory processes, oxidative stress, and apoptosis. It also restored the proper cell morphology and functionality through a mechanism which probably involves the activation of ERβ and GPR30 receptors along with the expression of GnRH peptide and KISS1R.

Topics: Anti-Inflammatory Agents; Antioxidants; Apoptosis; Cells, Cultured; Encephalitis; Gonadotropin-Releasing Hormone; Humans; Hypothalamus; Isoflavones; Membranes, Artificial; Models, Biological; Neurons; Neuroprotective Agents; Palmitates; Phytoestrogens; Polylactic Acid-Polyglycolic Acid Copolymer

At present, concerns are pointing to "tasteful" high-fat diets as a cause of conditioning physical-social states that through alterations of some key emotional- and nutritional-related limbic circuits such as hypothalamic and amygdalar areas lead to obesity states. Feeding and energetic homeostatic molecular mechanisms are part of a complex neuronal circuit accounting for this metabolic disorder. In an attempt to exclude conventional drugs for treating obesity, daidzein, a natural glycosidic isoflavone, which mimics estrogenic neuroprotective properties against increased body weight, is beginning to be preferred. In this study, evident anxiolytic-like behaviors were detected following treatment of high-fat diet hamsters with daidzein as shown by extremely evident (p < 0.001) exploration tendencies in novel object recognition test and a notably greater amount of time spent (p < 0.01) in open arms of elevated plus maze. Moreover, the isoflavone promoted a protective role against neurodegeneration processes as shown by few, if any, amino cupric silver granules in amygdalar, hypothalamic and hippocampal neuronal fields when compared with obese hamsters. Interestingly, elevated expression levels of the anorexic neuropeptide receptor neurotensin1 in the above limbic areas of obese hamsters were extremely reduced by daidzein, especially during recovery of cognitive events. Contextually, such effects were strongly paralleled by increased levels of the anti-neuroinflammatory cytokine, interleukin-10. Our results corroborate a neuroprotective ability of this natural glycosidic isoflavone, which through its interaction with the receptor neurotensin1 and interleukin-10 pathways is correlated not only to improved feeding states, and subsequently obesity conditions, but above all to cognitive performances.

Topics: Animals; Brain; Cricetinae; Diet, High-Fat; Exploratory Behavior; Gene Expression; Interleukin-10; Isoflavones; Mesocricetus; Nootropic Agents; Obesity; Phytoestrogens; Receptors, Neurotensin

Low risk of breast cancer is observed among females consuming a moderate quantity of soy throughout their life. The present study was conducted to evaluate the anticancer potential of Daidzein, one of the major Isoflavones in soy using Human breast cancer cells MCF-7.. MCF-7 were subjected to various doses of Daidzein treatment to determine the IC50 value. Onset of apoptosis was ascertained by AnnexinV assay and caspase 3/7 activity post treatment. Expression of pro-apoptotic protein Bax and anti-apoptotic protein Bcl2 was also assessed to further confirm apoptotic mode of cell death. ROS production post treatment with Daidzein was assessed to ascertain the apoptosis via intrinsic pathway. Expression of ER α and ER β was evaluated by western blot analysis.. Human breast cancer cells MCF-7 were found to be sensitive to Daidzein treatment, with an IC50 value of 50µM. Increased percentage of treated cells stained with Annexin V confirmed apoptosis mediated cell death. Activity of Caspase 3/7 activity was found to be 1.4-fold higher in Daidzein treated cells than control cells, confirming apoptosis. Daidzein caused over expression of Bax and down-regulated expression of Bcl2. There has been an outburst of ROS in Daidzein treated cells indicating that Daidzein induces apoptosis via intrinsic pathway. A decrease in the expression of ER α and increase in levels of ER β has been observed which are conducive indicator of apoptosis.. In conclusion, the present study suggests that Daidzein induces apoptosis in MCF-7 cells by mitochondrial pathway along with lowering the ratio of ER α/β and an outburst of Reactive Oxygen Species(ROS).

Topics: Adenocarcinoma; Apoptosis; bcl-2-Associated X Protein; Breast Neoplasms; Cell Culture Techniques; Estrogen Receptor alpha; Estrogen Receptor beta; Humans; Inhibitory Concentration 50; Isoflavones; MCF-7 Cells; Phytoestrogens; Proto-Oncogene Proteins c-bcl-2; Reactive Oxygen Species

Cocaine addiction is a chronic relapsing disorder that lacks of an effective treatment. Isoflavones are a family of compounds present in different plants and vegetables like soybeans that share a common chemical structure. Previous studies have described that synthetic derivatives from the natural isoflavone daidzin can modulate cocaine addiction, by a mechanism suggested to involve aldehyde-dehydrogenase (ALDH) activities.. Based on these previous studies, we investigated the effects of three natural isoflavones, daidzin, daidzein, and genistein, on the modulation of the cocaine reinforcing effects and on cue-induced reinstatement in an operant mouse model of cocaine self-administration.. Chronic treatment with daidzein or genistein decreased operant responding to obtain cocaine intravenous infusions. On the other hand, daidzein and daidzin, but not genistein, were effective in decreasing cue-induced cocaine reinstatement. Complementary studies revealed that daidzein effects on cocaine reinforcement were mediated through a mechanism that involved dopamine type-2/3 receptors (DA-D2/3) activities.. Our results suggest that these natural compounds alone or in combination can be a potential therapeutic approach for cocaine addiction. Further clinical studies are required in order to ascertain their potential therapeutic use.

Topics: Animals; Cocaine; Cocaine-Related Disorders; Conditioning, Operant; Cues; Dopamine Uptake Inhibitors; Dose-Response Relationship, Drug; Extinction, Psychological; Isoflavones; Locomotion; Male; Mice; Phytoestrogens; Reinforcement, Psychology; Self Administration

The incidences of intracranial aneurysm and aneurysmal subarachnoid hemorrhage are high in postmenopausal women. Although population-based studies suggest that hormone replacement therapy is beneficial for postmenopausal women with intracranial aneurysms, estrogen replacement may no longer be recommended for the prevention of chronic diseases given its association with adverse outcomes, such as cancer and ischemic stroke. The isoflavone daidzein and its intestinal metabolite equol are bioactive phytoestrogens and potent agonists of estrogen receptors. Given their estrogenic properties, we investigated whether the isoflavones daidzein and equol are protective against the formation and rupture of intracranial aneurysms in a mouse model of the postmenopausal state.. We induced intracranial aneurysms in ovariectomized adult female mice using a combination of induced systemic hypertension and a single injection of elastase into the cerebrospinal fluid. We fed the mice with an isoflavone-free diet with/without daidzein supplementation, or in a combination of intraperitoneal equol, or oral vancomycin treatment. We also used estrogen receptor beta knockout mice.. Both dietary daidzein and supplementation with its metabolite, equol, were protective against aneurysm formation in ovariectomized mice. The protective effects of daidzein and equol required estrogen receptor-β. The disruption of the intestinal microbial conversion of daidzein to equol abolished daidzein’s protective effect against aneurysm formation. Mice treated with equol had lower inflammatory cytokines in the cerebral arteries, suggesting that phytoestrogens modulate inflammatory processes important to intracranial aneurysm pathogenesis.. Our study establishes that both dietary daidzein and its metabolite, equol, protect against aneurysm formation in ovariectomized female mice through the activation of estrogen receptor-β and subsequent suppression of inflammation. Dietary daidzein’s protective effect required the intestinal conversion to equol. Our results indicate the potential therapeutic value of dietary daidzein and its metabolite, equol, for the prevention of the formation of intracranial aneurysms and related subarachnoid hemorrhage.

Topics: Animals; Equol; Female; Inflammation Mediators; Intracranial Aneurysm; Isoflavones; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovariectomy; Phytoestrogens

Phytoestrogens comprise biologically active constituents of human and animal diet that can impact on systemic and local estrogen functions in the brain. Here we report on the importance of dietary phytoestrogens for maintaining activity in a brain circuit controlling aggressive and social behavior of male mice. After six weeks of low-phytoestrogen chronic diet (diadzein plus genistein <20 μg/g) a reduction of intermale aggression and altered territorial marking behavior could be observed, compared to littermates on a standard soy-bean based diet (300 μg/g). Further, mice on low-phyto diet displayed a decrease in sociability and a reduced preference for social odors, indicating a general disturbance of social behavior. Underlying circuits were investigated by analysing the induction of the activity marker c-Fos upon social encounter. Low-phyto diet led to a markedly reduced c-Fos induction in the medial as well as the cortical amygdala, the lateral septum, medial preoptic area and bed nucleus of the stria terminalis. No difference between groups was observed in the olfactory bulb. Together our data suggest that dietary phytoestrogens critically modulate social behavior circuits in the male mouse brain.

Topics: Aggression; Animals; Behavior, Animal; Brain; Corticomedial Nuclear Complex; Diet; Isoflavones; Male; Mice; Mice, Inbred C57BL; Nerve Net; Phytochemicals; Phytoestrogens; Preoptic Area; Proto-Oncogene Proteins c-fos; Social Behavior; Territoriality

Isoflavones in soybeans are well-known phytoestrogens. Soy isoflavones present in conjugated forms are converted to aglycone forms during processing and storage. Isoflavone aglycones (IFAs) of soybeans in human diets have poor solubility in water, resulting in low bioavailability and bioactivity. Enzyme-mediated glycosylation is an efficient and environmentally friendly way to modify the physicochemical properties of soy IFAs. In this study, we determined the optimal reaction conditions for

Topics: beta-Glucans; Biological Availability; Chromatography, High Pressure Liquid; Deinococcus; Escherichia coli; Genetic Vectors; Genistein; Glucosyltransferases; Glycine max; Glycosylation; Isoflavones; Mass Spectrometry; Phytoestrogens; Plant Extracts

Phytoestrogens are plant compounds that are structurally similar to estrogen and that possess anti-cancer properties. Previous studies have reported that coumestrol, daidzein and genistein could induce cell death by reducing Annexin A1 protein in leukemic cell lines. Annexin A1 (ANXA1) is involved in cell progression, metastasis, and apoptosis in several types of cancer cells. The present study sought to investigate if the effects of phytoestrogens on apoptosis, cell cycle arrest and phagocytosis in ANXA1-knockdown leukemic cells are mediated through ANXA1 or occurred independently.. Transfection of ANXA1 siRNA was conducted to downregulate ANXA1 expression in Jurkat, K562 and U937 cells. Apoptosis and cell cycle assays were conducted using flow cytometry. Western blot was performed to evaluate ANXA1, caspases and Bcl-2 proteins expression. Phagocytosis was determined using hematoxylin and eosin staining.. The expression of ANXA1 after the knockdown was significantly downregulated in all cell lines. Genistein significantly induced apoptosis associated with an upregulation of procaspase-3, -9, and - 1 in Jurkat cells. The Bcl-2 expression showed no significant difference in Jurkat, K562 and U937 cells. Treatment with phytoestrogens increased procaspase-1 expression in Jurkat and U937 cells while no changes were detected in K562 cells. Flow cytometry analysis demonstrated that after ANXA1 knockdown, coumestrol and genistein caused cell cycle arrest at G2/M phase in selected type of cells. The percentage of phagocytosis and phagocytosis index increased after the treatment with phytoestrogens in all cell lines.. Phytoestrogens induced cell death in ANXA1-knockdown leukemia cells, mediated by Annexin A1 proteins. Graphical abstract.

Topics: Annexin A1; Apoptosis; Caspases; Cell Cycle; Cell Death; Coumestrol; Gene Knockdown Techniques; Genistein; Humans; Isoflavones; Jurkat Cells; K562 Cells; Leukemia; Phagocytosis; Phytoestrogens; Proto-Oncogene Proteins c-bcl-2; RNA, Small Interfering; THP-1 Cells; U937 Cells

Daidzein application may represent an effective and less harmful alternative to indicated, classical estrogenization of ageing men. The aim of this study was to perform structural and hormonal analysis of the adrenal cortex, after estradiol or daidzein supplementation in a rat model of the andropause.. Middle-aged Wistar rats were divided into sham operated (SO; n = 8), orchidectomized (Orx; n = 8), estradiol treated orchidectomized (Orx + E; n = 8) and daidzein treated orchidectomized (Orx + D; n = 8) groups. Estradiol (0.625 mg/kg b.m./day) or daidzein (30 mg/kg b.m./day) were administered subcutaneously for three weeks, while the SO and Orx groups received the vehicle alone. Set objectives were achieved using stereology, histochemistry/immunohistochemistry, immunoassays and ultrastructural analysis.. Both estradiol and daidzein treatment significantly increased volumes of the zona glomerulosa cell and nuclei, but decreased circulating aldosterone levels. Estradiol markedly increased volumes of the zona fasciculata cell and nuclei in parallel with significant decrease of the adrenal tissue level of corticosterone, while daidzein significantly decreased both the adrenal and circulating levels of corticosterone. Serum DHEA level and volumes of the zona reticularis cell and nuclei significantly increased upon estradiol treatment, whereas daidzein even stronger increased the circulating level of DHEA. Shunting of the corticosteroidogenesis pathways towards adrenal androgens production, after the treatments, corresponded to the ultrastructural findings and zonal capillary network rearrangements.. Given the coherence of its effects and relative safety, daidzein could be the remedy of choice for the treatment of ageing-caused androgen deprivation and the hypothalamo-pituitary-adrenal axis hyperfunction/related metabolic issues in males.

Topics: Adrenal Cortex; Aldosterone; Andropause; Animals; Body Weight; Corticosterone; Dehydroepiandrosterone; Enzyme-Linked Immunosorbent Assay; Immunohistochemistry; Isoflavones; Male; Microscopy, Electron, Transmission; Mitochondria; Orchiectomy; Organ Size; Phytoestrogens; Potassium; Random Allocation; Rats; Rats, Wistar; Sodium

Efficient aquaculture is depending on sustainable protein sources. The shortage in marine raw materials has initiated a shift to "green aquafeeds" based on staple ingredients such as soy and wheat. Plant-based diets entail new challenges regarding fish health, product quality and consumer risks due to the possible presence of chemical contaminants, natural toxins and bioactive compounds like phytoestrogens. Daidzein (DAI), genistein (GEN) and glycitein (GLY) are major soy isoflavones with considerable estrogenic activities, potentially interfering with the piscine endocrine system and affecting consumers after carry-over. In this context, information on isoflavone biotransformation in fish is crucial for risk evaluation. We have therefore isolated hepatic fractions of Atlantic salmon (Salmo salar), the most important species in Norwegian aquaculture, and used them to study isoflavone elimination and metabolite formation. The salmon liver microsomes and primary hepatocytes were characterized with respect to phase I cytochrome P450 (CYP) and phase II uridine-diphosphate-glucuronosyltransferase (UGT) enzyme activities using specific probe substrates, which allowed comparison to results in other species. DAI, GEN and GLY were effectively cleared by UGT. Based on the measurement of exact masses, fragmentation patterns, and retention times in liquid chromatography high-resolution mass spectrometry, we preliminarily identified the 7-O-glucuronides as the main metabolites in salmon, possibly produced by UGT1A1 and UGT1A9-like activities. In contrast, the production of oxidative metabolites by CYP was insignificant. Under optimized assay conditions, only small amounts of mono-hydroxylated DAI were detectable. These findings suggested that bioaccumulation of phytoestrogens in farmed salmon and consumer risks from soy-containing aquafeeds are unlikely.

Topics: Animals; Aquaculture; Biotransformation; Chromatography, Liquid; Genistein; Glucuronides; Glucuronosyltransferase; Glycine max; Hepatocytes; Isoflavones; Microsomes, Liver; Phytoestrogens; Salmo salar; UDP-Glucuronosyltransferase 1A9

Few potentially modifiable risk factors of male infertility have been identified, and while different diets and food groups have been associated with male infertility, evidence linking dietary factors including phytoestrogens and semen quality is limited and contradictory.. To study the associations between phytoestrogen intake and other dietary factors and semen quality.. High consumption of daidzein (≥13.74 μg/d), a phytoestrogen found in soy products, was a protective factor for MSC with an odds ratio (95%CI) of 0.58 (0.42-0.82) after adjustment for clustering and potential confounding. Dietary risk factors for PM after similar adjustment showed that drinking whole milk (OR 0.67, 95%CI 0.47-0.96) and eating red meat were protective with an OR 0.67 (0.46-0.99) for eating red meat >3 times/wk.. In this case-referent study of men attending an infertility clinic for fertility diagnosis, we have identified that low MSC is inversely associated with daidzein intake. In contrast, daidzein intake was not associated with PM but eating red milk and drinking whole milk were protective.. Dietary factors associated with semen quality were identified, suggesting that male fertility might be improved by dietary changes.

Topics: Case-Control Studies; Diet; Food Preferences; Humans; Infertility, Male; Isoflavones; Male; Meat; Phytoestrogens; Risk Factors; Semen Analysis; Soy Foods; Sperm Count; Spermatozoa; Surveys and Questionnaires

Soy isoflavones are bioactive phytoestrogens with known health benefits. Soybean embryo extract (SEE) has been consumed as a source of isoflavones, mainly daidzein, glycitein, and genistein. While previous studies have reported the anti-obesity effects of SEE, this study investigates their molecular mechanisms and the synergistic effects of co-treatment with SEE and enzymatically modified isoquercitrin (EMIQ). SEE upregulated genes involved in lipolysis and brown adipocyte markers and increased mitochondrial content in differentiated C3H10T1/2 adipocytes in vitro. Next, we use a high-fat diet-induced obesity mouse model to determine the anti-obesity effect of SEE. Two weeks of single or combined treatment with SEE and EMIQ significantly reduced body weight gain and improved glucose tolerance. Mechanistically, SEE treatment increased mitochondrial content and upregulated genes involved in lipolysis in adipose tissue through the cAMP/PKA-dependent signaling pathway. These effects required a cytosolic lipase adipose triglyceride lipase (ATGL) expression, confirmed by an adipocyte-specific ATGL knockout mouse study. Collectively, this study demonstrates that SEE exerts anti-obesity effects through the activation of adipose tissue metabolism and exhibits a synergistic effect of co-treatment with EMIQ. These results improve our understanding of the mechanisms underlying the anti-obesity effects of SEE related to adipose tissue metabolism.

Topics: Adipocytes; Adipose Tissue; Animals; Cell Differentiation; Diet, High-Fat; Genistein; Glycine max; Humans; Isoflavones; Lipolysis; Mice; Obesity; Phytoestrogens; Plant Extracts; Quercetin; Seeds

Daidzein (DAI) is a kind of natural isoflavonic phytoestrogen with estrogenic activity. However, little is known about its influence on early fetal growth in mammalian animals. The current study aimed to explore the characteristics of amniotic fluid exposure to dietary DAI using

Topics: Amniotic Fluid; Animals; Antioxidants; Cytokines; Diet; Dietary Supplements; Embryo, Mammalian; Female; Fetal Development; Glutathione Peroxidase; Hormones; Insulin-Like Growth Factor I; Isoflavones; Metabolome; Phytoestrogens; Pregnancy; Progesterone; Swine

Leucaena feed has been reported to cause disruptive effects on livestock reproduction, such as low calving percentages in cows, abortion in female goats and pigs, dead fetuses and fetal resorption in pregnant rats. In this study, the effects of Leucaena on different female reproductive variables were analyzed in two different reproductive conditions: gonadally intact and ovariectomized (OVX) female rats. Leucaena (LEU) was administered to females in both experimental conditions for 30 consecutive days. The effects of the legume extract were compared with those of Daidzein (DAI), a phytoestrogen, and of the female hormone estradiol (E

Topics: Animals; Estradiol; Estrogens; Estrous Cycle; Fabaceae; Female; Isoflavones; Organ Size; Ovarian Follicle; Ovary; Phytoestrogens; Plant Extracts; Progesterone; Rats; Rats, Wistar; Reproduction; Sexual Behavior, Animal; Uterus; Vagina

Phytoestrogens are nonsteroidal plant compounds with similar chemical structures to mammalian estrogen capable of mimicking the effect of estrogen in selective tissues. A diet rich in phytoestrogens is associated with a variety of health benefits including decreased risks for heart disease, breast cancer, and osteoporosis. Obesity has long thought to be associated with improved bone density due to increased mechanical loading, but recent literature suggests obesity may actually decrease bone health. Daidzein, a soy-derived phytoestrogen, has been shown to improve parameters of bone health in lean animal models of osteoporosis but has not been tested in obese animals. Following a one-week acclimation to a standard AIN-93G diet, 19 five-week-old female obese Zucker rats (OZR) were randomly assigned to a modified AIN-93G diet containing either high daidzein (HD, 0.121 g kg-1 feed) or low daidzein (LD, 0.01 g kg-1 feed). After 8 weeks, tibias and femurs were removed to assess true density (Archimedes principal), mechanical strength (three-point bending test), and femoral osteogenic gene expression. Serum was collected to assess osteocalcin and deoxypyridinoline. Our results indicated that there were no significant differences between the measures for tibial or femoral true density or mechanical strength for the rats in the HD and LD diet groups. Similarly, there were no significant differences in gene expressions related to osteogenic pathways, or serum biomarkers of bone formation and resorption. Overall, an increased dose of daidzein from soy protein supplementation does not elicit an improvement in markers of bone health in obese Zucker rats.

Topics: Amino Acids; Animals; Biomarkers; Body Weight; Bone and Bones; Bone Density; Diet; Dietary Supplements; Disease Models, Animal; Energy Intake; Female; Femur; Gene Expression; Isoflavones; Obesity; Osteocalcin; Osteogenesis; Osteoporosis; Phytoestrogens; Rats; Rats, Zucker

The aim of this research was to isolate bacteria capable of biotransforming daidzein from fresh feces from pregnant horses. A Hungate anaerobic roller tube was used for anaerobic culture. Single colonies were picked at random and incubated with daidzein. High performance liquid chromatography was used to detect whether the isolated bacteria were able to biotransform the substrate. A strain capable of reducing daidzein was selected and characterized using sequence analysis of 16S rDNA, and a phylogenetic tree was constructed. The morphological physiological and biochemical characteristics of the strain were investigated. A facultative anaerobic, Gram-positive bacterium capable of converting daidzein to dihydrodaidzein was isolated and named HXBM408 (MF992210). A BLAST search of HXBM408's 16S rDNA sequence against the GenBank database suggested that the strain has 99% similarity with Pediococcus acidilactici strain DSM (NR042057). The morphological, physiological, and biochemical characteristics of HXBM408 are very similar to those of Pediococcus. Based on these characteristics, the strain was identified as Pediococcus acidilactici. The bacterial strain HXBM408 isolated from the feces of pregnant horses was able to reduce the isoflavone daidzein to dihydrodaidzein.

Topics: Animals; Bacteria; Biotransformation; DNA, Bacterial; DNA, Ribosomal; Feces; Female; Glycine max; Horses; Isoflavones; Pediococcus acidilactici; Phylogeny; Phytoestrogens; Pregnancy; RNA, Ribosomal, 16S

The soy isoflavone genistein has been described to up-regulate breast cancer resistance protein (BCRP) and, thus, enhance chemoresistance in breast cancer cells. The aim of this work was to assess the effect of long- and short-term incubation with daidzein, the second most abundant soy isoflavone and its metabolite equol on the expression and activity of P-glycoprotein, multidrug resistance-associated proteins 1 and 2 (MRP1 and MRP2) and BCRP in breast cancer cells.. MCF-7 and MDA-MB-231 cells were treated with phytoestrogen concentrations within the range achieved in individuals with a high isoflavone intake. Transporter expression was evaluated at protein and mRNA level through western blot and qRT-PCR, respectively. Transporter activity was determined using doxorubicin, mitoxantrone and carboxy-dichlorofluorescein as substrates.. Daidzein (5 µM) up-regulated MRP2- and down-regulated MRP1 protein expressions in MCF-7 and MDA-MB-231 cells, respectively. Both effects were ER-dependent, as determined using the antagonist ICI 182,780. The decrease in MRP1 mRNA in MDA-MB-231 cells indicates a transcriptional mechanism. On the contrary, MRP2 induction in MCF-7 cells takes place post-transcriptionally. Whereas changes in the transporter expression had a minor effect on the transporter activity, acute incubation with daidzein, R-equol and S-equol led to a strong inhibition of BCRP activity and an increase in the IC. In contrast to previous reports for genistein, daidzein and equol do not provoke a major up-regulation of the transporter expression but instead an inhibition of BCRP activity and sensitization to BCRP substrates.

Topics: ATP Binding Cassette Transporter, Subfamily G, Member 2; Blotting, Western; Breast Neoplasms; Cell Line, Tumor; Down-Regulation; Equol; Humans; Isoflavones; Neoplasm Proteins; Phytoestrogens; Polymerase Chain Reaction; Up-Regulation

Pueraria mirifica (PM) is a medicinal plant native to Thailand contained high amount of phytoestrogen and possesses anticancer activity. This study reports the effect of P. mirifica extract, phytoestrogen of diadzein and genistein for its benign prostate hyperplasia properties in testosterone-induced prostate hyperplasia in male Sprague Dawley rats. The P. mirifica extract was evaluated for its total phenols, flavonoid and antioxidant activity using DPPH, FRAP and metal chelating assay. The assessment of P. mirifica, diadzein and genistein against benign prostate hyperplasia was determined in testosterone-induced prostate hyperplasia in male Sprague Dawley rats. The total phenol was higher than flavonoid but showed low antioxidant activity of DPPH, FRAP and metal chelating. The aqueous PM extract at 1000 mg/kg significantly increased testosterone levels in testosterone-induced rats by 13% while diadzein and genistein increased it by 11% and 17% respectively. However, levels of FSH, LH, triglyceride and HDL are not affected by the oral administration of PM, diadzein and genistein to the rats. Similarly, total protein, albumin, globulin, total bilirubin, conjugated bilirubin, alkaline phosphatase, alanine aminotransferase, AST, and G-glutamyltransferase showed no significant difference as compared with negative control rats. The body weight of the rats, testis, kidney and liver showed no toxic effect. The zinc content increased significantly and the zinc transporter gen of ZnT4 and ZIP4 highly expressed suggesting that the PM, diadzein and genistein plays essential role in modulating prostate zinc homeostasis. Similarly, the expression of IL-6, AR and ER was significantly reduced indicating functioning in regulation of prostate growth and acts as anti-inflammatory role in preventing BPH. In conclusion, the results indicated that PM reduced BPH and contributed to the regulation in the zinc transport expression of the prostate cells in the benign prostate hyperplasia (BPH).

Topics: Animals; Antioxidants; Genistein; Hyperplasia; Isoflavones; Male; Membrane Transport Proteins; Phytoestrogens; Plant Extracts; Prostate; Prostatic Hyperplasia; Pueraria; Rats; Rats, Sprague-Dawley; Testis; Testosterone; Thailand; Zinc

Equol is a metabolite of isoflavone daidzein and has an affinity to estrogen receptors. Although equol is produced by intestinal bacteria, the association between the status of equol production and the gut microbiota has not been fully investigated. The aim of this study was to compare the intestinal bacteria responsible for equol production in gut microbiota between equol producer and non-producer subjects regarding the intake of daidzein. A total of 1044 adult subjects who participated in a health survey in Hirosaki city were examined. The concentration of equol in urine was measured by high-performance liquid chromatography. The relative abundances of 8 bacterial species responsible for equol production in the gut microbiota was assessed using 16S rRNA amplification. There were 458 subjects identified as equol producers. The proportion of equol production status and the intake of daidzein increased with age. Daily intake of daidzein was larger in equol-producer. The intestinal bacteria, which convert daidzein to equol were present in both equol producers and non-producers. However, the relative abundance and the prevalence of

Topics: Adult; Aged; Bacteria; Equol; Female; Gastrointestinal Microbiome; Humans; Isoflavones; Male; Middle Aged; Phytoestrogens; RNA, Ribosomal, 16S

Phytoestrogens, such as daidzein and genistein, may be used to treat various hormone-dependent disorders. Daidzein can be metabolized by intestinal microbes to S-equol. However, not all individuals possess bacteria producing this metabolite, resulting in categorization of equol vs nonequol producers. Past human and rodent studies have suggested that supplementation of this compound might yield beneficial metabolic and behavioral effects. We hypothesized that administration of S-equol to diet-induced obese male and female mice would mitigate potential diet-induced metabolic and comorbid neurobehavioral disorders. To test this possibility, we placed 5-week-old C57 mice on a high-fat diet (HFD) to mimic the diet currently consumed by many Western adults. Animals were randomly assigned to S-equol supplementation (10 mg/kg body weight) or vehicle control group. After 4 weeks on HFD with or without S-equol supplementation, metabolic and behavioral phenotyping was performed. Although the initial hypothesis proposed that S-equol treatment would improve metabolic and neurobehavioral outcomes, this supplementation instead exacerbated aspects of HFD-induced metabolic disease, as indicated by suppressed physical activity in treated individuals, reduced energy expenditure in treated males, and serum chemistry changes (hyperglycemia in treated individuals; hyperinsulinemia and hypoleptinemia in treated males). Conversely, S-equol individuals exhibited less anxiety-like and depressive-like behaviors, as evidenced by increased exploratory time in the elevated plus maze by treated males and increased time spent mobile in the tail suspension test for treated individuals. In summary, S-equol may be beneficial in mitigating depression and anxiety disorders in individuals, but for indeterminate reasons, supplementation may worsen facets of metabolic disorders in obese individuals.

Topics: Animals; Anxiety; Anxiety Disorders; Behavior, Animal; Blood Glucose; Depression; Depressive Disorder; Dietary Supplements; Equol; Female; Hindlimb Suspension; Insulin; Isoflavones; Leptin; Male; Maze Learning; Metabolic Diseases; Metabolic Syndrome; Mice, Inbred C57BL; Phytoestrogens; Sex Factors

Phytoestrogens, naturally occurring plant chemicals, have long been thought to confer beneficial effects on human cardiovascular and metabolic health. However, recent epidemiological studies, have yielded conflicting outcomes, in which phytoestrogen consumption was both positively and negatively correlated with adiposity. Interestingly, several dietary phytoestrogens are known to stimulate or inhibit the activity of the peroxisome proliferator-activated receptor gamma (PPARγ), a key physiological regulator of adipogenesis.. The objective of this study was to test the hypothesis that the pro- or anti-adipogenic activity of phytoestrogen chemicals is related to the ability to activate PPARγ in adipocytes.. The effects of resveratrol and the soy isoflavones genistein and daidzein on adipogenesis were examined in cell-based assays using the 3T3-L1 cell model. In parallel, ligand-mediated alterations in PPARγ target gene expression were measured by quantitative polymerase chain reaction. The agonist/antagonist activities of phytoestrogens on PPARγ were further assessed by quantifying their ability to affect recruitment of transcriptional cofactors to the receptor.. Resveratrol displayed significant anti-adipogenic activities as exhibited by the ability to antagonize PPARγ-dependent adipocyte differentiation, down-regulate genes involved in lipid metabolism, block cofactor recruitment to PPARγ, and antagonize the effects of the PPARγ agonist rosiglitazone. In contrast, genistein and daidzein functioned as PPARγ agonists while also displaying pro-adipogenic activities.. These data provide biological evidence that the pro- or anti-obesity effects of phytoestrogens are related to their relative agonist/antagonist activity on PPARγ. Thus, PPARγ-activation assays may enable the screening of dietary components and identification of agents with adipogenic activities. https://doi.org/10.1289/EHP3444.

Topics: 3T3-L1 Cells; Adipogenesis; Animals; Diet; Genistein; Isoflavones; Mice; Phytoestrogens; PPAR gamma; Resveratrol

Daidzein (DAZ), a dominant isoflavone in various natural products such as soybeans, has been gaining attention due to the beneficial health effects (e.g., protection against cancer and diabetes) of its metabolites. Our major hypothesis was that dietary exposure to the soy phytoestrogen DAZ could modulate the immune responses toward a protective effect and lead to improved metabolic functions (such as glucose metabolism). In this study, we applied complementary mouse models, the hybrid B6C3F1 and inbred type 1 diabetes prone non-obese diabetic (NOD) mice, to investigate if DAZ exposure modulated the immune responses. The animals were orally administered DAZ at various physiological doses (2-20 mg/kg body weight) during adulthood. DAZ significantly altered the relative organ weights in female B6C3F1 mice and decreased the B cell population (represented by CD3

Topics: Animals; Antibody Formation; B-Lymphocytes; Diabetes Mellitus, Type 1; Dietary Supplements; Female; Glucose; Glycine max; Homeostasis; Immunomodulation; Isoflavones; Male; Mice; Mice, Inbred C3H; Mice, Inbred C57BL; Mice, Inbred NOD; Phytoestrogens; T-Lymphocytes

Equol improves menopausal symptoms and it is synthesized from daidzein, one of the isoflavonoids in soybeans, by the bacteria in the large intestines of some people. The purpose of this study was to isolate equol-producing bacteria using daidzein from the intestinal microflora and to produce equol-containing chungkookjang (short-term fermented soybean). Equol-producing bacteria from the feces of Sprague-Dawley female rats were isolated using media containing daidzein. The isolated bacteria were cultured in thioglycollate media and equol production was identified through thin-layer chromatography and ultraperformance liquid chromatography-mass spectrometry. The bacteria were identified by 16S rRNA sequencing. The rate of equol production in different concentrations of daidzein was assessed. The expression of genes that code for enzymes associated with the production of equol from daidzein was detected through reverse transcription quantitative PCR. The bacterium we isolated was Lactobacillus intestinalis (LC096206.1, 99%). L. intestinalis was found to express daidzein reductase, dihydrodaidzein reductase, and tetrahydrodaidzein reductase, the enzymes involved in producing equol from daidzein. The conversion rate of equol from daidzein was highest (29.5%) using 200 μM daidzein for 48 h of incubation. When chungkookjang fermented with Bacillus amyloquencies SRCM100001 was incubated with L. intestinalis, 0.32 ± 0.04 mg equol/g chungkookjang was produced. In conclusion, L. intestinalis efficiently produces equol from not only daidzein but also in chungkookjang.

Topics: Animals; Bacillus; Equol; Feces; Female; Fermentation; Fermented Foods; Gastrointestinal Microbiome; Glycine max; Humans; Isoflavones; Lactobacillus; Oxidoreductases; Phytoestrogens; Rats; Rats, Sprague-Dawley; RNA, Ribosomal, 16S; Soybean Proteins

Topics: Animals; Estradiol; Estrogens; Female; Genistein; Glycine max; Inflammation; Isoflavones; Ovary; Oxidative Stress; Phytoestrogens; Polycystic Ovary Syndrome; Rats

To investigate whether phytoestrogens (genistein and daidzein) alter in vitro decidualization of human endometrial stromal cells (ESCs).. Isolated primary ESCs were exposed to phytoestrogens and decidualized in vitro.. Academic fertility center.. Twenty fertile oocyte donors attending the IVI Valencia clinic.. Treatment of ESC with phytoestrogens at 0, 10, 20, 50, and 100 μM.. The ESC proliferation was analyzed by MTS assay. In vitro decidualization was induced in the presence of phytoestrogens by medroxyprogesterone acetate/cyclic adenosine 3':5' monophosphate and evaluated by prolactin (PRL) ELISA and F-actin immunostaining. The Ki67 proliferative marker was analyzed by immunofluorescence. The ESC apoptosis was assessed by annexin V/propidium iodide detection using flow cytometry. Estrogen (ERβ) and P receptor (PR) localization were evaluated by immunofluorescence.. The ESC exposed to 0, 19, 20, 50, and 100 μM of genistein, daidzein, and genistein + daidzein showed a dose-dependent proliferation decrease. After 48-96 hours of culture, this reduction was significant in the presence of 50 μM of phytoestrogens versus 10 μM untreated ESC. The ESC decidualized in the presence of phytoestrogens did not rearrange their cytoskeletons and showed a significant decrease in PRL secretion compared with untreated decidualized ESCs (dESCs). However, phytoestrogens did not alter proliferative status or the percentage of viable/apoptotic cells in dESC compared with untreated dESC. During decidualization, phytoestrogens induced the same nuclear translocation of ERβ and PR as the control dESC.. This study reveals that high doses of phytoestrogens could affect the in vitro decidualization process.

Topics: Cell Proliferation; Cells, Cultured; Decidua; Dose-Response Relationship, Drug; Endometrium; Female; Genistein; Humans; Isoflavones; Phytoestrogens; Signal Transduction; Stromal Cells

To evaluate the relationship between phytoestrogen and colon cancer risk, we quantified plasma isoflavones (Genistein and Daidzein) and lignan (enterolactone) in a Korean nested case-control study and conducted replication study in a Vietnamese case-control study.. Study populations of 101 cases and 391 controls were selected from the Korean Multicenter Cancer Cohort which was constructed from 1993 to 2004. For replication study, Vietnamese hospital-based case-control subjects of 222 cases and 206 controls were selected from 2003 to 2007. The concentrations of plasma genistein, daidzein, and enterolactone were quantified by liquid chromatography-mass spectrometry. Logistic regression models were used to compute odds ratios (ORs) and 95% confidence intervals (CIs), and meta-analysis was conducted to estimate combined ORs (CORs) and 95% Cis of Korean and Vietnamese population in 2014.. Genistein showed a continual decrease in colorectal cancer risk according to level up of the concentration categories in Korean and Vietnamese population (P for trend = 0.032, and 0.001, respectively) and a significantly decreased risk was found at the highest concentration of genistein and daidzein (for the highest category compared to the lowest: COR (95% CI) = 0.46 (0.30-0.69), and COR (95% CI) = 0.54 (0.36-0.82)). When the study population was stratified, the beneficial relationship of genistein with colorectal cancer was observed regardless of sex and anatomical subtype. However, enterolacton level was not associated with colorectal cancer risk.. High plasma levels of isoflavones had relationship with a decreased risk of colorectal cancer, regardless of different ethnic background.

Topics: 4-Butyrolactone; Adult; Aged; Asian People; Case-Control Studies; Colorectal Neoplasms; Female; Genistein; Humans; Isoflavones; Lignans; Male; Middle Aged; Phytoestrogens; Republic of Korea; Risk Factors; Vietnam

Selected estrogen receptor β-selective phytoestrogen (phytoSERM), a preparation of genistein, daidzein, and S-equol, has an 83-fold selective affinity for estrogen receptor (ER) β, and may promote neuronal survival and estrogenic mechanisms in the brain without exerting feminizing activity in the periphery. The aim of this study was to assess the safety, tolerability, and single-dose pharmacokinetics of the phytoSERM formulation in perimenopausal and postmenopausal women.. Eighteen women aged 45 to 60 years from a 12-week clinical trial evaluating cognitive performance and vasomotor symptoms were randomly assigned to placebo, 50 mg, or 100 mg phytoSERM treatment groups. Plasma levels of the three parent phytoestrogens and their metabolites were measured before and at 2, 4, 6, 8, and 24 hours after ingestion by isotope dilution high-performance liquid chromatography (HPLC) electrospray ionization tandem mass spectrometry.. Plasma concentrations of genistein, daidzein, and S-equol peaked at 9, 6, and 4 hours, respectively, for the 50-mg dose, and at 6, 6, and 5 hours, respectively, for the 100-mg dose. The maximum concentration (Cmax) and area under the curve (AUC) for the three parent compounds were greater in the 100-mg dose group, indicating a dose-dependent change in concentration with the phytoSERM treatment. No adverse events were elicited.. A single-dose oral administration of the phytoSERM formulation was well-tolerated and did not elicit any adverse events. It was rapidly absorbed, reached high plasma concentrations, and showed a linear dose-concentration response in its pharmacokinetics. These findings are consistent with previously reported parameters for each parent compound (Clinicaltrials.gov NCT01723917).

Topics: Area Under Curve; Drug Combinations; Equol; Estrogen Receptor beta; Female; Genistein; Humans; Isoflavones; Middle Aged; Phytoestrogens; Selective Estrogen Receptor Modulators

This study aimed to investigate the effects of soy isoflavones, genistein (GEN) and daidzein, (DAI) on the uterine function in ovary-intact middle-aged rats. GEN and DAI (35mg/kg) were subcutaneously administrated to acyclic (12-month-old) Wistar females, daily, for 4weeks. Control group received either vehicle (olive oil and ethanol, 9:1) or remained intact. We found that GEN and DAI differently affect uterine morphophysiology. GEN significantly increased the uterine wet weight which was associated with hyperplastic changes, revealed by stereological and histomorphometrical analyses. Also, PCNA immunoexpression was increased, whereas expression of apoptotic marker (caspase-3) was decreased. Protein and gene expressions of ERα were down-regulated, while PR and ERβ were up-regulated after GEN application. Also, GEN caused an increase of LAC and VEGF mRNA expression, together with an up-regulation of Akt activity. In contrast, DAI did not change the uterine wet weight and stereological features of the main uterine compartments as well as LAC and VEGF gene expression. Absence of hyperplastic changes were illustrated by an increase in caspase-3 immunoexpression, associated with reduced PCNA expression. DAI up-regulated only the expression of ERβ, while the expression levels of ERα and PR remain unaffected. Also, DAI inhibited the activation of Akt due to down-regulation of phosphorylated and total form of Akt protein expression. Compared to GEN, DAI did not promote events associated with the endometrial cell proliferation in the conducted study, figuring as the compound with a potential safety profile, which justifies further investigation.

Topics: Age Factors; Animals; Anticarcinogenic Agents; Cell Proliferation; Female; Genistein; Homeostasis; Injections, Subcutaneous; Isoflavones; Ovary; Phytoestrogens; Rats; Rats, Wistar; Uterus

Daidzein, the main bioactive soy isoflavone in Nature, has been found to possess many biological functions. It has been investigated in particular as a phytoestrogen owing to the similarity of its structure with that of the human hormone estrogen. Due to the lack of comprehensive studies on daidzein metabolism, further research is still required to clarify its in vivo metabolic fate and intermediate processes. In this study, an efficient strategy was established using UHPLC-LTQ-Orbitrap mass spectrometry to profile the metabolism of daidzein in rats. Meanwhile, multiple data-mining methods including high-resolution extracted ion chromatogram (HREIC), multiple mass defect filtering (MMDF), neutral loss fragment (NLF), and diagnostic product ion (DPI) were utilized to investigate daidzein metabolites from the HR-ESI-MS¹ to ESI-MS

Topics: Animals; Chromatography, High Pressure Liquid; Data Mining; Isoflavones; Male; Metabolome; Metabolomics; Molecular Structure; Phytoestrogens; Rats, Sprague-Dawley; Spectrometry, Mass, Electrospray Ionization; Tandem Mass Spectrometry

Topics: 3T3-L1 Cells; Adipocytes; Animals; Cell Differentiation; Genistein; Isoflavones; Mice; Obesity; Phytoestrogens; Phytotherapy; Transcription Factors

Endometriosis is an estrogen-dependent disease, and isoflavones interact with estrogen receptors. The purposes of this study are to investigate the in vitro and in vivo effects of daidzein-rich isoflavone aglycones (DRIAs), dietary supplements, on cellular proliferation in endometriosis. Stromal cells isolated from ovarian endometrioma (OESCs) and normal endometrium (NESCs) were cultured with DRIAs, i.e., each of the DRIA components (daidzein, genistein, or glycitein), or isoflavone glycosides (IG; DRIA precursors). A mouse model of endometriosis was established by transplanting donor-mouse uterine fragments into recipient mice. Our results showed that DRIAs (0.2-20 μM) inhibited the proliferation of OESCs (P < 0.05 for 0.2 μM; P < 0.01 for 2 and 20 μM) but not of NESCs. However, daidzein, genistein, glycitein, and IG did not inhibit their proliferation. DRIA-induced suppression was reversed by inhibition of the estrogen receptor (ER)β by an antagonist, PHTPP, or by ERβ siRNA (P < 0.05), but not by MPP, an ERα antagonist. In OESCs, DRIAs led to reduced expression of IL-6, IL-8, COX-2, and aromatase, as well as reduced aromatase activity, serum glucocorticoid-regulated kinase levels, and PGE

Topics: Animals; Cell Proliferation; Endometriosis; Female; Humans; Inflammation; Isoflavones; Mice; Phosphorylation; Phytoestrogens; Signal Transduction

Postmenopausal osteoporosis is a common disorder accompanied with estrogen deficiency in women. Plants containing phytoestrogens and amino acids have been used in the osteoporosis treatment. The present study aims to evaluate the estrogen-like activity of the Cicer arietinum extract (CAE) and its ability to inhibit osteoclastogenesis process. These achieved by investigating the binding of its active phytoestrogens (genistein, daidzein, formononetin and biochanin A) to the estrogen receptors (ER) α and β of rats and human in silico. In addition, in vivo study on ovariectomized (OVX) rats is performed. For in vivo study, twenty four rats were divided into four groups (n= 6). Group I is the sham control rats which administered distilled water. Groups II, III, and IV are OVX groups which administered distilled water, CAE (500 mg/kg), and alendronate; respectively. The docking study revealed that the phytoestrogens docked into the protein active site with binding energies comparable with that of estrogens (estriol and β-estradiol) which means the similarity between the estrogenic contents of CAE and the ensogenous ones. Additionally, in vivo study revealed that CAE reverse TRAP5b and RANKL levels that drastically increased in the untreated OVX group. But, it trigger upregulation of OPG, enhance the OPG/RANKL ratio and modulate the bone and uterus alterations of OVX group. Phytoestrogens and the bone-protective amino acids contents of CAE could be responsible for their estrogen-like effect and antiosteoporotic activity. These results concluded that CAE is an attractive candidate for developing a potential therapeutic cheap agent used as an alternative to the synthetic estrogen replacement therapy. Further, in vivo validation is required for its clinical application.

Topics: Alendronate; Animals; Bone Density Conservation Agents; Calcium-Binding Proteins; Cicer; Disease Models, Animal; Estradiol; Estrogen Receptor alpha; Estrogen Receptor beta; Female; Gene Expression Regulation; Genistein; Humans; Isoflavones; Membrane Glycoproteins; Molecular Docking Simulation; Osteogenesis; Osteoporosis; Osteoprotegerin; Ovariectomy; Phytoestrogens; Phytotherapy; Protein Structure, Secondary; RANK Ligand; Rats; Receptors, Cytoplasmic and Nuclear; Receptors, Peptide

Based on the assumed oestrogenic and apoptotic properties of soya isoflavones (genistein, daidzein), and following the current OECD test-guidelines and principle of 3Rs, we have studied the potential toxicity of phytochemicals on the zebrafish embryos test (ZFET). For this purpose, zebrafish embryos at 2-3 h post-fertilisation (hpf) were exposed to both soya isoflavones (from 1.25 mg/L to 20 mg/L) and assayed until 96 hpf. Lethal and sub-lethal endpoints (mortality, hatching rates and malformations) were estimated in the ZFET, which was expanded to potential gene expression markers, determining the lowest observed effect (and transcriptional) concentrations (LOEC, LOTEC), and the no-observable effect (and transcriptional) concentrations (NOEC, NOTEC). The results revealed that genistein is more toxic (LC50-96 hpf: 4.41 mg/L) than daidzein (over 65.15 mg/L). Both isoflavones up-regulated the oestrogen (esrrb) and death receptors (fas) and cyp1a transcript levels. Most thyroid transcript signals were up-regulated by genistein (except for thyroid peroxidase/tpo), and the hatching enzyme (he1a1) was exclusively up-regulated by daidzein (from 1.25 mg/L onwards). The ZFET proved suitable for assessing toxicant effects of both isoflavones and potential disruptions (i.e. oestrogenic, apoptotic, thyroid, enzymatic) during the embryogenesis and the endotrophic larval period.

Topics: Animals; Apoptosis; Cytochrome P-450 CYP1A1; Dietary Supplements; Ectogenesis; Embryo, Nonmammalian; Endocrine Disruptors; fas Receptor; Gene Expression Regulation, Developmental; Genistein; Glycine max; Isoflavones; Larva; Lethal Dose 50; Phytoestrogens; Receptors, Estrogen; Seeds; Signal Transduction; Thyroid Gland; Toxicity Tests, Acute; Zebrafish

It is currently unclear as to whether sex hormones are significantly affected by soy or whey protein consumption. Additionally, estrogenic signaling may be potentiated via soy protein supplementation due to the presence of phytoestrogenic isoflavones. Limited also evidence suggests that whey protein supplementation may increase androgenic signaling. Therefore, the purpose of this study was to examine the effects of soy protein concentrate (SPC), whey protein concentrate (WPC), or placebo (PLA) supplementation on serum sex hormones, androgen signaling markers in muscle tissue, and estrogen signaling markers in subcutaneous (SQ) adipose tissue of previously untrained, college-aged men (n = 47, 20 ± 1 yrs) that resistance trained for 12 weeks. Fasting serum total testosterone increased pre- to post-training, but more so in subjects consuming WPC (p < 0.05), whereas serum 17β-estradiol remained unaltered. SQ estrogen receptor alpha (ERα) protein expression and hormone-sensitive lipase mRNA increased with training regardless of supplementation. Muscle androgen receptor (AR) mRNA increased while ornithine decarboxylase mRNA (a gene target indicative of androgen signaling) decreased with training regardless of supplementation (p < 0.05). No significant interactions of supplement and time were observed for adipose tissue ERα/β protein levels, muscle tissue AR protein levels, or mRNAs in either tissue indicative of altered estrogenic or androgenic activity. Interestingly, WPC had the largest effect on increasing type II muscle fiber cross sectional area values (Cohen's d = 1.30), whereas SPC had the largest effect on increasing this metric in type I fibers (Cohen's d = 0.84). These data suggest that, while isoflavones were detected in SPC, chronic WPC or SPC supplementation did not appreciably affect biomarkers related to muscle androgenic signaling or SQ estrogenic signaling. The noted fiber type-specific responses to WPC and SPC supplementation warrant future research.

Topics: Adipose Tissue; Adult; Dietary Supplements; Estradiol; Estrogen Receptor alpha; Estrogen Receptor beta; Genistein; Humans; Isoflavones; Male; Muscle Fibers, Fast-Twitch; Muscle Fibers, Slow-Twitch; Muscle, Skeletal; Ornithine Decarboxylase; Phytoestrogens; Plant Extracts; Receptors, Androgen; Resistance Training; Soybean Proteins; Sterol Esterase; Testosterone; Whey Proteins; Young Adult

Equol is one of the most active soy isoflavones. When the association between soy food intake in daily life and health outcomes is examined in epidemiological studies, it is important to define the equol-producing status of each individual. However, few studies have assessed equol-producing status without a soy challenge test. To determine a robust cutoff criterion for equol producer classification in observational studies, we conducted a urinary isoflavone concentration survey in daily life among women. Furthermore, we examined the association between eating habits regarding soy foods and equol-producing status. A total of 4,412 participants were included in the analyses. Urinary isoflavones were analyzed using a high-performance liquid chromatography method. We examined the distribution of the log10 equol/daidzein ratios, finding a mixture of two normal distributions, corresponding to equol producer and non-producer subpopulations. Applying a finite mixture model, we estimated the means, standard deviations, and mixing proportions of these two distributions. The estimation was carried out using the SAS NLIN procedure. The optimal cutoff point for the log10 equol/daidzein ratio in the study population was determined to be -1.42, according to the estimated parameters of the mixture distribution. Based on this criterion, 1,830 (41.5%) of the participants were identified as equol producers. Compared with non-consumers of soy foods, consumers of soy foods had significantly higher odds of being equol producers. Using log10-transformed equol/daidzein ratios ≥ -1.42 to define equol producers among Japanese women is reasonable and suitable for determining equol-producing status in epidemiological studies. We found that soy food eating habits were associated with equol-producing status. Further investigation is required to evaluate associations between equol-producing status in daily life and health outcomes. The results of this study suggest the best cutoff point to use in the definition of equol-producing status in daily life.

Topics: Adult; Cohort Studies; Equol; Feeding Behavior; Female; Humans; Isoflavones; Japan; Middle Aged; Phytoestrogens; Soy Foods

Phytoestrogens and xenoestrogens act as agonists/antagonists in bone formation and differentiation. Strong bones are depending of the ability of osteoblasts to form new tissue and to mineralize the newly formed tissue. Dysfunctional or loss of mineralization leads to weak bone and increased fracture risk. In this study, we reported the effect of different types of phytoestrogens (daidzein, genistein and equol) on mineralization in hFOB 1.19 cells stimulated with bisphenol A (BPA).. Cell mineralization capacity of phytoestrogens was investigated by evaluating calcium, phosphate content and alkaline phosphatase activity. Bone related markers, osteocalcin and osteonectin, responsible in maintaining mineralization were also measured.. BPA is significantly interfering with bone mineralization in hFOB 1.19 cells. However, the enhanced mineralization efficacy of daidzein and genistein (particularly at a dose of 5 and 40 μg/mL, respectively) was evidenced by increasing calcium and phosphate content, higher ALP activity, compared to the untreated BPA group. The quantitative analyses were confirmed through morphological findings. Osteocalcin and osteonectin levels were increased in phytoestrogens-treated cells. These findings revealed the potential effect of phytoestrogens in reverting the demineralization process due to BPA exposure in hFOB 1.19 cells.. We found that osteoblast differentiation and mineralization were maintained following treatment with phytoestrogens under BPA exposure.

Topics: Benzhydryl Compounds; Calcification, Physiologic; Cell Differentiation; Cells, Cultured; Equol; Estrogens, Non-Steroidal; Genistein; Humans; Isoflavones; Osteoblasts; Osteogenesis; Phenols; Phytoestrogens

Estrogen receptors (ERs) α and β are distributed in most tissues of women and men. ERs are bound by estradiol (E2), a natural hormone, and mediate the pleiotropic and tissue-specific effects of E2, such as proliferation of breast epithelial cells or protection and differentiation of neuronal cells. Numerous environmental molecules, called endocrine disrupting compounds, also interact with ERs. Phytoestrogens belong to this large family and are considered potent therapeutic molecules that act through their selective estrogen receptor modulator (SERM) activity. Using breast cancer cell lines as a model of estrogen-dependent proliferation and a stably ER-expressing PC12 cell line as a model of neuronal differentiating cells, we studied the SERM activity of major dietary compounds, such as apigenin, liquiritigenin, daidzein, genistein, coumestrol, resveratrol and zearalenone. The ability of these compounds to induce ER-transactivation and breast cancer cell proliferation and enhance Nerve Growth Factor (NGF) -induced neuritogenesis was assessed. Surprisingly, although all compounds were able to activate the ER through an estrogen responsive element reporter gene, they showed differential activity toward proliferation or differentiation. Apigenin and resveratrol showed a partial or no proliferative effect on breast cancer cells but fully contributed to the neuritogenesis effect of NGF. However, daidzein and zearalenone showed full effects on cellular proliferation but did not induce cellular differentiation. In summary, our results suggest that the therapeutic potential of phytoestrogens can diverge depending on the molecule and the phenotype considered. Hence, apigenin and resveratrol might be used in the development of therapeutics for breast cancer and brain diseases.

Topics: Adrenal Gland Neoplasms; Animals; Antineoplastic Agents, Phytogenic; Apigenin; Breast Neoplasms; Cell Proliferation; Chemokine CXCL12; Diet; Dose-Response Relationship, Drug; Estrogen Receptor alpha; Female; Gene Expression Regulation, Neoplastic; Humans; Isoflavones; MCF-7 Cells; Nerve Tissue Proteins; Neurites; Neurogenesis; PC12 Cells; Pheochromocytoma; Phytoestrogens; Rats; Response Elements; Resveratrol; Selective Estrogen Receptor Modulators; Stilbenes; Transcription, Genetic; Transfection; Zearalenone

Daidzein has become increasingly popular as a dietary supplement, particularly for postpeak-estrus animals, as a safe and natural alternative estrogen-like compound. However, there is little available safety data of daidzein in laying hens. A study was conducted to examine if high-dose daidzein affected the safety of hens, including mortality, laying performance, egg quality, hematological parameters, clinical chemical parameters, organ development parameters, and hatchability. A total of 2,448 42-wk-old Rugao laying hens were randomly assigned to 4 groups with 6 replicates of 102 birds each (612 laying hens per group). After a 2-wk acclimation period, the birds were fed diets supplemented with 0, 10, 100, or 200 mg/kg of daidzein for 12 wk. The hatchability of setting eggs increased linearly with increasing dietary daidzein supplementation (P = 0.034), while the hatchability of fertile eggs also tended to increase linearly (P = 0.069). The red cell distribution width (RCDW) and coefficient variation of RCDW showed an increasing and then decreasing quadratic response to increasing dietary daidzein supplementation (P = 0.001 and 0.002, respectively). No statistically significant changes were observed in mortality, laying performance, egg quality, clinical chemistry parameters, or organ development parameters (P > 0.05). The magnitude of these hematological changes was such that they were considered to be of no toxicological significance. Therefore, a nominal daidzein concentration of 200 mg/kg is not expected to cause adverse effects following daily administration to laying hens for 84 d.

Topics: Animal Feed; Animals; Chickens; Diet; Dietary Supplements; Dose-Response Relationship, Drug; Female; Isoflavones; Ovum; Phytoestrogens; Random Allocation; Reproduction

Isoflavones are a class of polyphonic compounds present in legumes and are called phytoestrogens because of their estrogen-like activity. Estrogen influences the behavior of mammary epithelial cells (MECs) during pregnancy and lactation. In this study, we investigated the direct influences of isoflavones and their metabolites in milk production ability of MECs.. Mouse MECs were cultured with prolactin and dexamethasone (glucocorticoid analog) to induce milk production ability. Subsequently, lactating MECs were treated with each isoflavone. Coumestrol, biochanin A, genistein, and formononetin decreased the intracellular and secreted β-casein. On the other hand, p-ethylphenol, daidzein, and equol did not significantly influence β-casein production at any concentration. Coumestrol, biochanin A and genistein down-regulated the mRNA expression of whey acidic protein (WAP), lactoferrin and α-lactalbumin. In contrast, p-ethylphenol, daidzein and equol up-regulated β-casein and/or WAP with α-lactalbumin. Furthermore, coumestrol and genistein down-regulated the expression of prolactin receptor and signal transducer and activator of transcription 5 (STAT5) accompanied by a decrease in STAT5 phosphorylation.. Isoflavones and their metabolites influence the milk production ability of MECs through different interactions with prolactin/STAT5 signaling. Simultaneous intake of multiple isoflavones by consumption of legumes may induce promotive or adverse effects on lactating MECs.

Topics: Animals; Caseins; Cells, Cultured; Coumestrol; Epithelial Cells; Female; Genistein; Isoflavones; Mammary Glands, Animal; Mice; Mice, Inbred ICR; Milk; Phenols; Phosphorylation; Phytoestrogens; Prolactin; STAT5 Transcription Factor

The study is aimed at evaluating the chemosensitization and apoptotic effect of aglycone rich extracts of dietary phytoestrogens (derived from soybean and flaxseed) on estrogen receptor positive, MCF-7 and estrogen receptor negative, MDA-MB-231 cells. The extracts show potent activity on both the cell lines, hence, in silico studies have been carried out to find the possible reason for their activity.. MTT assay was carried to assess chemosensitization effect and activated caspase-3/7 activity was studied using flow-cytometry and western blotting. In silico studies were carried out using PharmMapper and the top hits were taken up for docking using the Schrödinger software. Top molecular targets were subjected to gene expression studies by qPCR and protein expression using Western blot analysis.. This study reports the apoptotic activity and chemosensitization effect of the phytoestrogens. Molecular docking studies predict AKR1B1 (aldose reductase), HRAS (Harvey rat sarcoma) and GSTP1 (glutathione s-transferase pi) as potential molecular targets for genistein, daidzein and secoisolariciresinol, respectively. Gene and protein expression studies show down-regulation of AKR1BI, HRAS and GSTP1 by the extracts.. The qPCR and western blot analysis results support the computational analyses, and hence genistein, daidzein and secoisolariciresinol may be considered as good candidates for future development into potent inhibitors of the respective protein targets through medicinal chemistry optimization.

Topics: Antineoplastic Agents, Phytogenic; Apoptosis; Blotting, Western; Breast Neoplasms; Butylene Glycols; Caspase 3; Caspase 7; Cell Line, Tumor; Computer Simulation; Down-Regulation; Female; Flow Cytometry; Gene Expression Regulation, Neoplastic; Genistein; Humans; Isoflavones; Lignans; MCF-7 Cells; Molecular Docking Simulation; Phytoestrogens

Ubiquitin-specific protease 19 (USP19) is a key player in the negative regulation of muscle mass during muscle atrophy. Loss-of-function approaches demonstrate that 17β-estradiol (E2) increases USP19 expression through estrogen receptor (ER) α and consequently decreases soleus muscle mass in young female mice under physiological conditions. Daidzein is one of the main isoflavones in soy, and activates ERβ-dependent transcription. Here, we investigated the effects of daidzein on E2-increased USP19 expression and E2-decreased soleus muscle mass in young female mice. Daidzein stimulated the transcriptional activity of ERβ in murine C2C12 cells and down-regulated USP19 expression. Consistently, daidzein inhibited E2-induced USP19 expression in a reporter activity using a functional half-estrogen response element (hERE) from Usp19. Daidzein inhibited E2-induced recruitment of ERα and promoted recruitment of ERβ to the Usp19 hERE. Dietary daidzein down-regulated the expression of USP19 at the mRNA and protein levels and increased soleus muscle mass in female mice, but not in males. In soleus muscle from ovariectomized (OVX) female mice, dietary daidzein inhibited E2-increased USP19 mRNA expression and E2-decreased muscle mass. Furthermore, E2 induced the recruitment of ERα and ERβ to the hERE, whereas daidzein inhibited E2-induced recruitment of ERα, and enhanced E2-increased recruitment of ERβ, to the Usp19 hERE. These results demonstrate that dietary daidzein decreases USP19 mRNA expression through ERβ and increases soleus muscle mass in young female mice, but not in male mice, under physiological conditions.

Topics: Active Transport, Cell Nucleus; Animals; Animals, Outbred Strains; Cell Line; Dietary Supplements; Endopeptidases; Enzyme Repression; Estrogen Receptor beta; Female; Genes, Reporter; Isoflavones; Male; Mice; Muscle, Skeletal; Myoblasts, Skeletal; Ovariectomy; Phytoestrogens; Random Allocation; Response Elements; Sarcopenia; Sex Characteristics; Signal Transduction; Ubiquitin-Specific Proteases

During lactation, mammary epithelial cells (MECs) form the blood-milk barrier by less-permeable tight junctions (TJs) to prevent the leakage of milk components. Phytoestrogens affect the proliferation, differentiation, and apoptosis of MECs. However, it remains unclear whether phytoestrogens are involved in the blood-milk barrier. Therefore, we investigated the influence of phytoestrogens (coumestrol, genistein, and daidzein) by using an in vitro mouse-MEC-culture model. The results showed that coumestrol and genistein changed the expression of TJ proteins (claudins-3 and -4 and occludin), weakened barrier function, and reduced β-casein production. Daidzein also weakened barrier function without inhibiting β-casein production. Additionally, coumestrol and genistein induced apoptosis in MECs. These results indicate that phytoestrogens weaken the blood-milk barrier by directly affecting TJs and the cellular viability of lactating MECs in different ways.

Topics: Animals; Caseins; Cell Survival; Coumestrol; Epithelial Cells; Female; Genistein; Humans; Isoflavones; Lactation; Mammary Glands, Animal; Mice; Mice, Inbred ICR; Milk; Phytoestrogens; Tight Junctions

The key biological active molecule of soya is the isoflavone daidzein, which possesses phytoestrogenic activity. The direct effect of soya and daidzein on ovarian cell functions is not known. This study examined the effect of daidzein on basic porcine ovarian granulosa cell functions and the response to follicle-stimulating hormone (FSH). We studied the effects of daidzein (0, 1, 10 and 100 μm), FSH (0, 0.01, 0.1, 1 IU/ml) and combinations of FSH (0, 0.01, 0.1, 1 IU/ml) + daidzein (50 μm) on proliferation, apoptosis and hormone release from cultured porcine ovarian granulosa cells and ovarian follicles. The expression of a proliferation-related peptide (PCNA) and an apoptosis-related peptide (Bax) was analysed using immunocytochemistry. The release of progesterone (P4) and testosterone (T) was detected using EIA. Leptin output was analysed using RIA. Daidzein administration increased granulosa cell proliferation, apoptosis and T and leptin release but inhibited P4 output. Daidzein also increased T release and decreased P4 release from cultured ovarian follicles. Follicle-stimulating hormone stimulated granulosa cell proliferation, apoptosis and P4, T and leptin release. The addition of daidzein promoted FSH-stimulated apoptosis (but not proliferation) but suppressed FSH-stimulated P4, T and leptin release. Our observations of FSH action confirm previous data on the stimulatory effect of FSH on ovarian cell proliferation, apoptosis and steroidogenesis and demonstrate for the first time the involvement of FSH in the upregulation of ovarian leptin release. Our observations of daidzein effects demonstrated for the first time that this soya isoflavone affected basic ovarian cell functions (proliferation, apoptosis and hormones release) and modified the effects of FSH. Daidzein promoted FSH action on ovarian cell proliferation and apoptosis and suppressed, and even inverted, FSH action on hormone release. The direct action of daidzein on basic ovarian cell functions and the ability of these cells to respond to FSH indicate the potential influence of soya-containing diets on female reproductive processes via direct action on the ovary.

Topics: Animals; Apoptosis; Cell Proliferation; Cells, Cultured; Dose-Response Relationship, Drug; Drug Interactions; Female; Follicle Stimulating Hormone; Granulosa Cells; Isoflavones; Phytoestrogens; Swine

Compounds with estrogenic potencies and their adverse effects in surface waters have received much attention. Both anthropogenic and natural compounds contribute to overall estrogenic activity in freshwaters. Recently, estrogenic potencies were also found to be associated with cyanobacteria and their blooms in surface waters. The present study developed and compared the solid phase extraction and LC-MS/MS analytical approaches for determination of phytoestrogens (8 flavonoids - biochanin A, coumestrol, daidzein, equol, formononetin, genistein, naringenin, apigenin - and 5 sterols - ergosterol, β-sitosterol, stigmasterol, campesterol, brassicasterol) and cholesterol in water. The method was used for analyses of samples collected in stagnant water bodies dominated by different cyanobacterial species. Concentrations of individual flavonoids ranged from below the limit of detection to 3.58 ng/L. Sterols were present in higher amounts up to 2.25 μg/L. Biological potencies of these phytoestrogens in vitro were characterized using the hERα-HeLa-9903 cell line. The relative estrogenic potencies (compared to model estrogen - 17β-estradiol) of flavonoids ranged from 2.25E-05 to 1.26E-03 with coumestrol being the most potent. None of the sterols elicited estrogenic response in the used bioassay. Estrogenic activity was detected in collected field water samples (maximum effect corresponding to 2.07 ng/L of 17β-estradiol equivalents, transcriptional assay). At maximum phytoestrogens accounted for only 1.56 pg/L of 17β-estradiol equivalents, contributing maximally 8.5% of the total estrogenicity of the water samples. Other compounds therefore, most likely of anthropogenic origin such as steroid estrogens, are probably the major drivers of total estrogenic effects in these surface waters.

Topics: Cholestadienols; Cholesterol; Cyanobacteria; Estradiol; Estrogens; Estrone; Fresh Water; Genistein; HeLa Cells; Humans; Isoflavones; Phytoestrogens; Phytosterols; Receptors, Estrogen; Sitosterols; Sterols; Tandem Mass Spectrometry; Water; Water Pollutants, Chemical

High dietary intake of plant estrogens (phytoestrogens) can affect brain structure and function. The effects of phytoestrogen intake within the range of normal animal and human dietary consumption, however, remain uncertain. The aim of the present study was to determine the effects of the isoflavonoids present in a standard low phytoestrogen laboratory rat chow on spine synapse density in the stratum radiatum of area CA1 of the hippocampus. Weanling rats (22days old) were fed either standard chow (Teklad 2018), a nutritionally comparable diet without soy (Teklad 2016) or a custom diet containing Teklad 2016 supplemented with the principal soy isoflavonoids, daidzein and genistein, for 40days. Rats were ovariectomized at 54days of age. Eight days later, spine synapse density on the apical dendrites of hippocampal pyramidal neurons in the stratum radiatum of area CA1 was measured by electron microscopic stereological analysis. Animals maintained on Teklad 2016 exhibited an approximately 60% lower CA1 spine synapse density than animals consuming Teklad 2018. Replacing genistein and daidzein in Teklad 2016 returned synapse density to levels indistinguishable from those in animals on Teklad 2018. These results indicate that the isoflavonoids in a standard laboratory rat diet exert significant effects on spine synapse density in the CA1 region of the hippocampus. Since changes in spine synapse density in this region of the hippocampus have been linked to cognitive performance and mood state, these data suggest that even relatively low daily consumption of soy phytoestrogens may be sufficient to influence hippocampal function.

Topics: Animal Feed; Animals; CA1 Region, Hippocampal; Dendritic Spines; Diet; Female; Genistein; Isoflavones; Microscopy, Electron; Ovariectomy; Phytoestrogens; Pyramidal Cells; Rats, Sprague-Dawley; Soybean Proteins; Synapses

Advantages of the supercritical fluid (SCF) process compared to the conventional solution stirring method (CSSM) in the preparation of daidzein-hydroxypropyl-β-cyclodextrin (HPβCD) complexes were investigated. Formation of daidzein/ HPβCD inclusion complexes was confirmed by Fourier transformed-infrared spectroscopy (FTIR), differential scanning calorimetry (DSC), X-ray diffraction (XRD) and scanning electron microscopy (SEM). Particle size, inclusion yield, drug solubility and dissolution of daidzein/HPβCD complexes were evaluated. Compared to CSSM, the SCF process resulted in higher inclusion yield and higher solubility. Also, extended dissolution of daidzein from the SCF processed HPβCD inclusion complexes was observed, with only 22.94 % released in 45 min, compared to its rapid release from those prepared by CSSM, with 98.25 % drug release in 15 min. This extended release of daidzein from SCF prepared inclusion complexes was necessary to avoid drug precipitation and improve drug solubilisation in the gastrointestinal tract. The results showed that the SCF process is a superior preparation method for daidzein-hydroxypropyl-β-cyclodextrin complexes.

Topics: 2-Hydroxypropyl-beta-cyclodextrin; beta-Cyclodextrins; Calorimetry, Differential Scanning; Chromatography, Supercritical Fluid; Crystallography, X-Ray; Delayed-Action Preparations; Drug Compounding; Excipients; Feasibility Studies; Isoflavones; Kinetics; Microscopy, Electron, Scanning; Particle Size; Phytoestrogens; Powder Diffraction; Solubility; Spectroscopy, Fourier Transform Infrared; Technology, Pharmaceutical; Thermogravimetry

Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) microsphere loaded 45S5 bioactive glass (BG) based scaffolds with drug releasing capability have been developed. PHBV microspheres with a mean particle size 4 ± 2 μm loaded with daidzein were obtained by oil-in-water single emulsion solvent evaporation method and applied to the surface of BG scaffolds by dip coating technique. The morphology, in vitro bioactivity in simulated body fluid (SBF), mechanical properties and drug release kinetics of microsphere loaded scaffolds were studied. The microspheres were shown to be homogeneously dispersed on the scaffold surfaces. It was confirmed that hydroxyapatite crystals homogeneously grew not only on the surface of the scaffold but also on the surface of the microspheres within 3 days of immersion in SBF. The daidzein release from the microsphere loaded scaffolds lasted almost 1 month and was determined to be diffusion controlled. The microsphere loaded BG scaffolds with daidzein releasing capability obtained in this study are a candidate for bone tissue engineering. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1765-1774, 2017.

Topics: Ceramics; Compressive Strength; Drug Delivery Systems; Drug Liberation; Glass; Isoflavones; Microspheres; Phytoestrogens; Polyesters; Tissue Scaffolds

Experimental studies suggest that phytoestrogen intake alters cancer and cardiovascular risk. This study investigated the associations of urinary phytoestrogens with total cancer (n = 79), cardiovascular (n = 108), and all-cause (n = 290) mortality among 5179 participants in the continuous National Health and Nutrition Examination Survey (1999-2004).. Urinary phytoestrogens were measured using high-performance liquid chromatography with tandem mass spectrometric detection. Survival analysis was performed to evaluate hazard ratios (HRs) and 95 % confidence intervals (CIs) for each of the three outcomes in relation to urinary phytoestrogens.. After adjustment for confounders, higher urinary concentrations of total enterolignans were associated with a reduced risk of death from cardiovascular disease (HR for tertile 3 vs. tertile 1 0.48; 95 % CI 0.24, 0.97), whereas higher urinary concentrations of total isoflavones (HR for tertile 3 vs. tertile 1 2.14; 95 % CI 1.03, 4.47) and daidzein (HR for tertile 3 vs. tertile 1 2.05; 95 % CI 1.02, 4.11) were associated with an increased risk. A reduction in all-cause mortality was observed for elevated urinary concentrations of total enterolignans (HR for tertile 3 vs. tertile 1 0.65; 95 % CI 0.43, 0.96) and enterolactone (HR for tertile 3 vs. tertile 1 0.65; 95 % CI 0.44, 0.97).. Some urinary phytoestrogens were associated with cardiovascular and all-cause mortality in a representative sample of the US population. This is one of the first studies that used urinary phytoestrogens as biomarkers of their dietary intake to evaluate the effect of these bioactive compounds on the risk of death from cancer and cardiovascular disease.

Topics: 4-Butyrolactone; Adult; Biomarkers; Body Mass Index; Cardiovascular Diseases; Cross-Sectional Studies; Female; Follow-Up Studies; Humans; Isoflavones; Lignans; Male; Middle Aged; Mortality; Neoplasms; Nutrition Surveys; Phytoestrogens; Proportional Hazards Models; Risk Factors; Treatment Outcome

Ischemic heart disease is the leading cause of death in postmenopausal women. The expression of caveolin, a membrane protein and a negative regulator of nitric oxide (NO), increases after menopause. The present study was designed to determine the effect of daidzein (DDZ), a phytoestrogen in attenuated cardioprotective effect of ischemic preconditioning (IPC) in ovariectomized rat heart.. Heart was isolated from ovariectomized rat and mounted on Langendorff's apparatus, subjected to 30 min ischemia and 120 min reperfusion. IPC was mediated by four cycles of 5 min ischemia and 5 min reperfusion. The infarct size was estimated using triphenyltetrazolium chloride stain, and coronary effluent was analyzed for lactate dehydrogenase and creatine kinase MB (CK-MB) release to assess the degree of myocardial injury. The release of NO was estimated indirectly by measuring the release of nitrite in coronary effluent.. IPC-induced cardioprotection was significantly attenuated in ovariectomized rats as compared to normal rats, which was restored by treatment of DDZ, a caveolin inhibitor (0.2 mg/kg subcutaneously) for 1 week. However, this observed cardioprotection was significantly attenuated by perfusion of l-nitroarginine methyl ester, an endothelial nitric oxide synthase (eNOS) inhibitor (100 µM/L) and glibenclamide, an adenosine triphosphate-sensitive potassium ion channel blocker (10 µM/L) alone or in combination, noted in terms of increase in myocardial infarct size, release of LDH and CK-MB, and also decrease in the release of NO.. Thus, it is suggested that DDZ restores the attenuated cardioprotective effect in ovariectomized rat heart, which may be due to downregulation of caveolin and subsequent increase in the activity of eNOS.

Topics: Animals; Caveolin 1; Creatine Kinase, MB Form; Disease Models, Animal; Female; Heart; In Vitro Techniques; Ischemic Preconditioning, Myocardial; Isoflavones; L-Lactate Dehydrogenase; Myocardial Infarction; Nitrites; Ovariectomy; Phytoestrogens; Rats, Wistar

Equol is a metabolite of daidzein that is produced by intestinal microbiota. The oestrogenic activity of equol is stronger than daidzein. Equol-producing bacteria are believed to play an important role in the gut. The rod-shaped and Gram-positive anaerobic equol-producing intestinal bacterium Slackia TM-30 was isolated from healthy human faeces and its effects on urinary phyto-oestrogen, plasma and faecal lipids were assessed in adult mice.. The urinary amounts of equol in urine were significantly higher in mice receiving the equol-producing bacterium TM-30 (BAC) group than in the control (CO) group (P < 0.05). However, no significant differences were observed between the urinary amounts of daidzein, dihydrodaidzein, enterodiol, and enterolactone between the BAC and CO groups. No significant differences in the plasma lipids were observed between the two groups. The lipid content (% dry weight) in the faeces sampled on the final day of the experiment tended to be higher in the BAC group than in the CO group (P = 0.07).. Administration of equol-producing bacterium TM-30 affected the urinary amounts of phyto-oestrogens and the faecal lipid contents of mice. The equol-producing bacterium TM-30 likely influences the metabolism of phyto-oestrogen via changes in the gastrointestinal environment. © 2015 Society of Chemical Industry.

Topics: 4-Butyrolactone; Actinobacteria; Animals; Equol; Feces; Female; Gastrointestinal Microbiome; Humans; Isoflavones; Lignans; Lipids; Mice; Mice, Inbred ICR; Phytoestrogens

Phytoestrogens are popular alternatives to estrogen therapy however their effects on hemostasis in postmenopausal women are unknown. This chapter describes a protocol to determine the effect of the phytoestrogens genistein, daidzein and equol, on the expression of key genes from the hemostatic system in human hepatocyte cell models and to determine the role of estrogen receptors in mediating any response seen using in vitro culture systems and Taqman(®) gene expression analysis.

Topics: Carcinoma, Hepatocellular; Equol; Estradiol; Estrogen Receptor alpha; Gene Expression Regulation, Neoplastic; Genistein; Hep G2 Cells; Humans; Isoflavones; Liver Neoplasms; Phytoestrogens; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; Taq Polymerase; Transfection; Workflow

The amount of secondary metabolites in plants can be enhanced or reduced by various external factors. In this study, the effect of strontium ions on the production of phytoestrogens in soybeans was investigated. The plants were treated with Hoagland's solution, modified with Sr(2+) with concentrations ranging from 0.5 to 3.0 mM, and were grown for 14 days in hydroponic cultivation. After harvest, soybean plants were separated into roots and shoots, dried, and pulverized. The plant material was extracted with methanol and hydrolyzed. Phytoestrogens were quantified by HPLC. The significant increase in the concentration of the compounds of interest was observed for all tested concentrations of strontium ions when compared to control. Sr(2+) at a concentration of 2 mM was the strongest elicitor, and the amount of phytoestrogens in plant increased ca. 2.70, 1.92, 3.77 and 2.88-fold, for daidzein, coumestrol, genistein and formononetin, respectively. Moreover, no cytotoxic effects were observed in HepG2 liver cell models after treatment with extracts from 2 mM Sr(2+)-stressed soybean plants when compared to extracts from non-stressed plants. Our results indicate that the addition of strontium ions to the culture media may be used to functionalize soybean plants with enhanced phytoestrogen content.

Topics: Cations, Divalent; Cell Survival; Chromatography, High Pressure Liquid; Coumestrol; Genistein; Glycine max; Hep G2 Cells; Humans; Hydroponics; Isoflavones; Methanol; Phytoestrogens; Plant Extracts; Plant Roots; Plant Shoots; Solvents; Strontium

Isoflavones genistein and daidzein are nonsteroidal phytoestrogens occurring mainly in soybean foods. These phytoestrogens possess estrogenic properties and show a variety of health benefits as anti-inflammatory agents. However, the mechanism of their action has not been identified in detail. The aim of this study is to characterize the antioxidant powers of genistein, daidzein and daidzein metabolite-equol through their activities to scavenge superoxide anion radical (O(•) 2 (-) ), hydroxyl radical (HO(•) ), 2,2-diphenyl-1-picrylhydrazyl radical (DPPH(•) ) and hydrogen peroxide (H2 O2 ) using chemiluminescence and spectrophotometry techniques. Potassium superoxide in dimethyl sulphoxide (DMSO) and 18-crown-6 ether were used as a source of O(•) 2 (-) . Hydroxyl radicals were produced using the Fenton reaction. In free radical assays, genistein had the IC50 values (an amount of antioxidant concentration required to decrease the initial radical concentration by 50%) 0.391 ± 0.012 mM for O(•) 2 (-) , 0.621 ± 0.028 mM for HO(•) and 1.89 ± 0.16 mM for DPPH(•) . The IC50 values for daidzein for these free radicals were 1.924 ± 0.011 mM, 0.702 ± 0.012 mM and 2.81 ± 0.03 mM, respectively. Equol was the most active the free radical scavenger with IC50  = 0.451 ± 0.018 mM for HO(•) and IC50  = 1.36 ± 0.11 mM for DPPH(•) . All tested compounds exerted a significant effect on the H2 O2 : IC50  = 18.1 ± 1.1 μM for genistein, IC50  = 2.1 ± 0.5 μM for daidzein, and IC50  = 1.06 ± 0.2 μM for equol. These findings show that genistein, daidzein and equol are effective free radical scavengers and possess high antioxidant power in vitro. Copyright © 2016 John Wiley & Sons, Ltd.

Topics: Antioxidants; Genistein; Isoflavones; Luminescence; Phytoestrogens

We evaluated the relationship between urine concentrations of phyto-oestrogens (isoflavones and lignans) and risk of incident type 2 diabetes in middle-aged and elderly Chinese residing in Singapore. Urine metabolites of isoflavones and lignans were assayed by HPLC among 564 diabetes cases and 564 matched controls in a case-control study nested within the Singapore Chinese Health Study cohort. Participants were free of diagnosed diabetes, CVD and cancer at morning urine collections during 1999-2004. Cases were participants who reported to have physician-diagnosed diabetes at follow-up visits during 2006-2010, whereas controls were randomly selected among those who remained free of diabetes and were matched to the index cases by age, sex, dialect group and date of urine collection. Conditional logistic regression models were used to calculate OR and 95 % CI with adjustment for potential confounders. The mean age of the participants at the time of urine collection was 59·8 years, and the average interval between urine collection and diabetes diagnosis was 4·0 years. The multivariate-adjusted OR for diabetes were 1·00 (reference), 0·76 (95 % CI 0·52, 1·11), 0·78 (95 % CI 0·53, 1·14) and 0·79 (95 % CI 0·54, 1·15) across quartiles of urine isoflavones (P for trend=0·54), and were 1·00 (reference), 0·87 (95 % CI 0·60, 1·27), 1·10 (95 % CI 0·77, 1·56) and 0·93 (95 % CI 0·63, 1·37) for lignans (P for trend=0·93). The results were similar in men and women, as well as for individual metabolites of isoflavones (genistein, daidzein, glycitin and equol) or lignans (enterodiol and enterolactone). The present study did not find a significant association between urine phyto-oestrogen metabolites and risk of type 2 diabetes in Chinese adults.

Topics: Asian People; Case-Control Studies; Diabetes Mellitus, Type 2; Equol; Female; Genistein; Humans; Isoflavones; Lignans; Logistic Models; Male; Middle Aged; Odds Ratio; Phytoestrogens; Risk Factors; Singapore

Epidemiological studies indicate lower prevalences of breast and prostate cancers and cardiovascular disease in Southeast Asia where vegetarianism is popular and diets are traditionally high in phytoestrogens. This study assessed plasma isoflavones in vegetarian and non-vegetarian Malaysian men according to age. Daidzein, genistein, equol (a daidzein metabolite), formononetin, biochanin A, estrone, estradiol and testosterone were measured by validated liquid chromatography tandem mass spectrometry (LCMSMS). Plasma isoflavone and sex hormone concentrations were measured in 225 subjects according to age (18-34, 35-44 and 45-67 years old). In all age groups, vegetarians had a higher concentration of circulating isoflavones compared with non-vegetarians especially in the 45-67 year age group where all isoflavones except equol, were significantly higher in vegetarians compared with omnivores. By contrast, the 18-34 year group had a significantly higher concentration of daidzein in vegetarians and significantly higher testosterone and estrone concentrations compared with non-vegetarians. In this age group there were weak correlations between estrone, estradiol and testosterone with some of the isoflavones. This human study provides the first Malaysian data for the phytoestrogen status of vegetarian and nonvegetarian men.. 流行病学研究表明东南亚乳腺癌、前列腺癌和心血管疾病的发病率较低，素食 在这些地区备受欢迎，传统饮食中富含植物雌激素。本研究根据年龄评估了马 来西亚素食和非素食男性血浆中异黄酮的含量。通过高效液相色谱-串联质谱 法（LCMSMS）测定血浆中大豆异黄酮、染料木素、雌马酚（一种大豆异黄 酮的代谢产物）、芒柄花素、鹰嘴豆芽素A、雌酮、雌二醇和睾酮的浓度。按 照年龄（18-34 岁、35-44 岁和45-67 岁）测定了225 名研究对象的血浆异黄酮 和性激素的浓度。在所有年龄组，尤其是45-67 岁组，素食者循环异黄酮浓度 高于非素食者，在45-67 岁组，除雌马酚外的所有异黄酮，素食者均显著高于 杂食者。相反，在18-34 岁组，素食者的大豆异黄酮、睾酮和雌酮的浓度显著 高于非素食者。在这个年龄组中，雌酮、雌二醇和睾酮与一些异黄酮之间有弱 的相关性。本研究提供了马来西亚素食和非素食男性植物雌激素状态的第一手 资料。.

Topics: Adolescent; Adult; Aged; Aging; Cross-Sectional Studies; Dairy Products; Diet, Vegetarian; Eggs; Equol; Estradiol; Estrone; Genistein; Humans; Isoflavones; Malaysia; Male; Middle Aged; Phytoestrogens; Testosterone

Mesquite (Prosopis sp.) and Leucaena leucocephala are widespread legumes, widely used to feed several livestock species and as food source for human populations in several countries. Both mesquite and Leucaena contain several phytoestrogens which might have potential estrogenic effects. Thus, the aim of this study was to evaluate the effects of mesquite pod and Leucaena extracts on several aspects of behavior and reproductive physiology of the male rat. The effects of the extracts were compared with those of estradiol (E2) and of two isoflavones: daidzein (DAI) and genistein (GEN). The following treatments were given to groups of intact male rats: vehicle; mesquite pod extract; Leucaena extract; E2; DAI; GEN. The results indicate that mesquite pod and Leucaena extracts disrupt male sexual behavior in a similar way to DAI and GEN, but less than E2. The main disruptor of sexual behavior was E2, however after 40 and 50days of administration, both extracts and phytoestrogens disrupted sexual behavior in a similar way to E2. The extracts also increased testicular germ cell apoptosis, decreased sperm quality, testicular weight, and testosterone levels, as phytoestrogens did, although these effects were less than those caused by estradiol. The number of seminiferous tubules with TUNEL-positive germ cells increased in extracts treated groups in a similar way to phytoestrogens groups, and E2 caused the greatest effect. The number of TUNEL-positive cells per tubule increased only in Leucaena extract and E2 groups, but not in mesquite- and phytoestrogens-treated groups. Spermatocytes and round spermatids were the TUNEL-positive cells observed in all experimental groups. This effect was associated with smaller testicular weights without atrophy in experimental groups compared with control. Testicular atrophy was only observed in estradiol-treated males. Testosterone decreased in males of all experimental groups, compared with control, this androgen was undetectable in E2 treated males. These results suggest that mesquite pod and Leucaena extracts cause effects similar to those of phytoestrogens in male rat reproduction, these effects were lower than those caused by E2.

Topics: Analysis of Variance; Animals; Body Weight; Genistein; In Situ Nick-End Labeling; Isoflavones; Male; Mimosine; Phytoestrogens; Plant Extracts; Prosopis; Rats; Rats, Wistar; Reproduction; Sexual Behavior, Animal; Spermatozoa; Testis; Testosterone; Time Factors

Urine and serum biomonitoring was used to measure internal exposure to selected dietary estrogens in a cohort of 30 pregnant women. Exposure was measured over a period comprising one-half day in the field (6 h) and one day in a clinic (24 h). Biomonitoring of the dietary phytoestrogens genistein (GEN), daidzein (DDZ) and equol (EQ), as well as the mycoestrogen, zearalenone (ZEN) and its congeners, was conducted using UPLC-MS/MS. Biomonitoring revealed evidence of internal exposure to naturally occurring dietary estrogens during pregnancy. Urinary concentrations of total GEN, DDZ and EQ were similar to levels reported for general adult U.S.. Measurable concentrations of total (parent and metabolites) GEN, DDZ and EQ were present in 240, 207 and 2 of 270 serum samples, respectively. Six out of 30 subjects had measurable concentrations of unconjugated GEN and/or DDZ in serum between 0.6 and 7.1 nM. Urine to serum total isoflavone ratios for GEN, DDZ and EQ were 13, 47, and 180, respectively. ZEN and its reductive metabolite, α-zearalenol (α-ZEL), were present in pregnant women (11 out of 30 subjects) as conjugates at levels near the limit of quantification. The average total urinary concentration was 0.10 μg/L for ZEN and 0.11 μg/L for α-ZEL.

Topics: Adult; Cohort Studies; Diet; Environmental Exposure; Environmental Monitoring; Equol; Estrogens; Female; Genistein; Glycine max; Humans; Isoflavones; Phytoestrogens; Pregnancy; Tandem Mass Spectrometry; Zearalenone

Daidzein is an isoflavone derived from soybeans that exerts preventive effects on bone loss in ovariectomized (OVX) animals. These effects have been correlated with increasing serum equol levels. In the present study, we investigated the effects of antibiotic intake on equol metabolism from daidzein, and the corresponding levels of bone loss in OVX mice.. Eight-week-old female ddY mice (n = 42) were either ovariectomized (OVX) or subjected to a sham operation (sham). OVX mice were then divided into six dietary subgroups: control diet (control), 0.3 % kanamycin diet (KN), 0.1 % daidzein diet (Dz), 0.1 % daidzein and 0.0375 % kanamycin diet (Dz+KN3.75), 0.1 % daidzein and 0.075 % kanamycin diet (Dz+KN7.5), and 0.1 % daidzein and 0.3 % kanamycin diet (Dz+KN30). The mice were fed their respective diets for 4 weeks.. Uterine weight and femoral bone mineral density (BMD) were significantly lower in the OVX mice compared in the sham mice. No significant differences in uterine weight were observed among all OVX dietary subgroups. The Dz subgroup was found to exhibit higher plasma equol and O-desmethylangolensin (O-DMA) concentrations, as well as greater femoral BMD, compared to all other OVX subgroups. Furthermore, when compared to the Dz group, kanamycin intake decreased plasma equol and O-DMA concentrations, as well as femoral BMD in the OVX mice.. These results suggest that kanamycin intake inhibited the conversion of daidzein to equol and O-DMA, blocking the preventive effects of daidzein on bone loss in OVX mice. Therefore, the bone-protective effects of daidzein intake may be predominantly associated with increased plasma concentrations of either equol or O-DMA.

Topics: Administration, Oral; Animals; Biotransformation; Body Weight; Bone Density; Diet; Disease Models, Animal; Equol; Female; Femur; Humans; Isoflavones; Kanamycin; Mice; Organ Size; Osteoporosis; Ovariectomy; Phytoestrogens; Uterus

Daidzein, the most widely studied soy phytoestrogen, is not only a potential antiosteoporosis agent owing to its possible osteogenic activity, but also shows anticancer activity. However, the mechanisms through which daidzein affects osteoblast function have not been investigated thoroughly. Here, we show that daidzein stimulated cell proliferation and differentiation of osteoblasts, demonstrated by upregulation of XTT activity, enhancement of alkaline phosphatase (ALP) activity, and upregulation of osteoblast-specific marker genes, including Runt-related transcription factor 2 (Runx2) and Smad1, as well as upregulation of Runx2 and Smad1 protein expression. To determine the mechanisms underlying daidzein's effects on osteoblast differentiation, we first tested the role of daidzein in bone morphogenetic protein (BMP)-2 gene expression in OCT1 cells, and found that it significantly upregulated the expression of BMP-2. Furthermore, it significantly enhanced the phosphorylated protein level of Smad1/5/8 and the protein level of Osterix and increased the activity of 12xSBE-OC-Luc. Finally, we demonstrated that daidzein stimulated Col I, Runx2, and ALP expression, while these effects were significantly blocked by the BMP signaling inhibitor noggin. Together, our data indicate that daidzein acts through stimulating the activation of BMP-2/Smads pathway to promote osteoblast proliferation and differentiation.

Topics: Animals; Bone Morphogenetic Protein 2; Cell Differentiation; Cell Proliferation; Cells, Cultured; Core Binding Factor Alpha 1 Subunit; Isoflavones; Mice; Octamer Transcription Factor-1; Osteoblasts; Phytoestrogens; Smad Proteins; Up-Regulation

Endocrine-disrupting chemicals (EDCs) are increasingly thought to be involved in the rising prevalence of disorders such as obesity, diabetes, and some hormone-dependent cancers. Several lines of evidence have indicated that vegetarian and vegan diets may offer some protection from such diseases. We hypothesized that exposure to selected EDCs among residents of the unique vegetarian/vegan community of Amirim would be lower than what has recently been reported for the omnivorous population in the first Israel Biomonitoring Study (IBMS).. We studied 42 Amirim residents (29 vegetarians/13 vegans; 24 women/18men, aged 50.7±13.7y). Subjects answered detailed lifestyle, and multipass, memory-based 24-hr dietary recall questionnaires. Concentrations of bisphenol A (BPA), 11 phthalate metabolites, and the isoflavone phytoestrogens (genistein and daidzein) were determined by GC or LC tandem mass-spectrometry on a spot urine sample. The results were compared to those obtained following the same methodology in the Jewish subgroup of the IBMS (n=184).. While a vegetarian/vegan nutritional pattern had no effect on exposure to BPA, it seemed to confer a modest protection (~21%) from exposure to high molecular weight phthalates. Furthermore, the summed metabolites of the high molecular weight phthalate DiNP were 36% lower in vegans compared to vegetarians (P<0.05). In contrast, Amirim residents exhibited a level of exposure to isoflavone phytoestrogens about an order of magnitude higher than in the IBMS (P<0.001).. In Israel, a country whose inhabitants demonstrate exposure to EDCs comparable to that of the US and Canada, a voluntary lifestyle of vegetarianism and preference for organic food has a modest, but possibly valuable, impact on exposure to phthalates, while it is associated with a very steep increase in the exposure to phytoestrogens. Major reduction in exposure to EDCs will require regulatory actions.

Topics: Adult; Benzhydryl Compounds; Cross-Sectional Studies; Diet, Vegetarian; Endocrine Disruptors; Environmental Monitoring; Female; Food, Organic; Genistein; Humans; Isoflavones; Israel; Life Style; Male; Middle Aged; Phenols; Phthalic Acids; Phytoestrogens; Rural Population; Surveys and Questionnaires; Vegans; Vegetarians

Increased serum norepinephrine level is one of pathological processes relating to heart disease (HD). Estrogens\ are considered as potential therapeutics for the treatment of HD; however, estrogen supplementation\ shows some side-effects, such as increasing the risk of developing breast, endometrial and ovarian cancers.\ This study investigated the cardio-protective effects of daidzein (Dai), a selective estrogen receptor\ modulator (SERM) from soy bean extract, in H9c2 cardiomyoblast cells treated with isoproterenol (ISO),\ a norepinephrine analog. In this in vitro model, H9c2 cells treated with Dai at different concentrations\ showed no statistical difference in cell viability. TdT-mediated digoxigenin-dUTP nick-end labeling (TUNEL) data and western blotting results indicated\ that Dai treated-H9c2 cells recovered from the damage induced by ISO. The recovery effects of Dai\ on ISO-induced damage were blocked by inhibition of Akt activation through adding Akt inhibitor.\ On the other hand, the fold changes of phosphorylated Akt (p-Akt)/Akt normalized with the control for con, 0.25, 0.5, 1, 3\ and 24 h of treatment were 1, 2, 5, 13, 11 and 10, respectively. In conclusion, Dai ameliorates apoptosis\ of cardiomyoblasts induced by ISO through Akt signaling pathway.

Topics: Animals; Apoptosis; Cell Line; In Situ Nick-End Labeling; Isoflavones; Isoproterenol; Myoblasts, Cardiac; Phytoestrogens; Proto-Oncogene Proteins c-akt; Rats

The objective of this study was to investigate the effects of daidzein on collagen metabolism and its underlying mechanism in cultured skin fibroblast and nude mouse skin.. Skin fibroblasts were exposed to different concentrations of daidzein (0.5-50 μg/mL) for 24 h or 48 h, respectively. Female nude mice were treated topically with 200 μg/mL daidzein once a day for 6 weeks. Cell viability and cell cycle were determined by MTT and flow cytometer. The transcriptional activity of collagen type I was evaluated and the expression of procollagen, matrix metalloproteinase-1 (MMP1) and MMP2 were measured by real-time polymerase chain reaction. A Western blot analysis was applied to detect the levels of phosphorylated-Smad2 and Smad3.. In the daidzein-treated cells the expression of type I procollagen increased markedly while the expressions of MMP1, and MMP2 was significantly inhibited. Additionally, the mouse skin showed more collagen deposition after daidzein treatment. The levels of transforming growth factor (TGF)-β, phosphorylated-smad2 and smad3 were also higher in the daidzein treated skin fibroblasts than in the controls.. The results showed that daidzein treatment can increase skin collagen synthesis and inhibit collagen degradation in vitro and in vivo. It seems that TGF-β/smad signalling pathways play an important role in daidzein-induced collagen accumulation.

Topics: Animals; Cell Line; Cell Survival; Collagen Type I; Female; Fibroblasts; Humans; Isoflavones; Matrix Metalloproteinase 1; Matrix Metalloproteinase 2; Mice; Mice, Inbred BALB C; Mice, Nude; Phosphorylation; Phytoestrogens; Procollagen; Signal Transduction; Smad2 Protein; Smad5 Protein; Transcription, Genetic; Transforming Growth Factor beta

Soy isoflavones (IF) are phytoestrogens, which interact with estrogen receptors. They are extensively metabolized by glucuronosyltransferases and sulfotransferases, leading to the modulation of their estrogenic activity. It can be assumed that this biotransformation also has a crucial impact on the uptake of IF by active or passive cellular transport mechanisms, but little is known about the transport of IF phase II metabolites into the cell. Therefore, transport assays for phase II metabolites of daidzein (DAI) were carried out using HEK293 cell lines transfected with five human candidate carriers, i.e., organic anion transporter OAT4, sodium-dependent organic anion transporter (SOAT), Na(+)-taurocholate cotransporting polypeptide (NTCP), apical sodium-dependent bile acid transporter ASBT, and organic anion transporting polypeptide OATP2B1. Cellular uptake was monitored by UHPLC-DAD. DAI monosulfates were transported by the carriers NTCP and SOAT in a sodium-dependent manner, while OAT4-HEK293 cells revealed a partly sodium-dependent transport for these compounds. In contrast, DAI-7,4'-disulfate was only taken up by NTCP-HEK293 cells. DAI-7-glucuronide, but not DAI-4'-glucuronide, was transported exclusively by OATP2B1 in a sodium-independent manner. DAI-7-glucuronide-4'-sulfate, DAI-7-glucoside, and DAI were no substrate of any of the tested carriers. In addition, the inhibitory potency of the DAI metabolites toward estrone-sulfate (E1S) uptake of the above-mentioned carriers was determined. In conclusion, human SOAT, NTCP, OATP2B1, and OAT4 were identified as carriers for the DAI metabolites. Several metabolites were able to inhibit carrier-dependent E1S uptake. These findings might contribute to a better understanding of the bioactivity of IF especially in case of hormone-related cancers.

Topics: Biological Transport; Chromatography, High Pressure Liquid; HEK293 Cells; Humans; Isoflavones; Organic Anion Transporters; Organic Anion Transporters, Sodium-Dependent; Organic Anion Transporters, Sodium-Independent; Phytoestrogens; Sterol O-Acyltransferase; Symporters

Contamination with bacterial endotoxin causes the disruption of the tight junction (TJ) barrier. We investigated the ameliorative effect of dietary flavonoids genistein (Ge) and daidzein (Di) in normal or lipopolysaccharide (LPS)-induced disruption of epithelial barrier function of the endometrium. Using the immortalized porcine glandular endometrial epithelial cells (PEG), transepithelial electrical resistance (TER) and FITC-dextran flux (FD-4) across the monolayer were measured. The mRNA expression of TJ proteins, zona occludens-1 (ZO1), and claudin-1, -3, -4, -7 and -8 was evaluated by real-time RT-PCR for coinciding effect of Ge or Di occurred at the gene transcription level. The results revealed that Ge and Di altered the TER, depending on times and concentrations. Low concentration (10(-10) M) of both compounds decreased the TER, whereas higher concentrations (10(-8) and 10(-6) M) increased the TER which was not related to the FD-4 flux. The increased TER by Ge or Di was parallel to the induction of claudin-3 and -4 or -8 mRNA expression respectively. With LPS inoculation, all isoflavone treatments inhibited the decreased TER induced by LPS, but only Ge (10(-8) or 10(-6) M) or Di (10(-10) or 10(-6) M) was coincidence with the decreased FD-4 flux. Under this LPS-stimulated condition, some or all examined TJ gene expressions appeared to be promoted by specific concentration of Ge or Di respectively. Our findings suggest that the soy isoflavones treatment could promote and restore the impaired endometrial barrier function caused by LPS contamination.

Topics: Animals; Cell Line; Claudins; Endometrium; Female; Gene Expression; Genistein; Isoflavones; Lipopolysaccharides; Phytoestrogens; Swine; Tight Junctions; Zonula Occludens-1 Protein

This study determined whether estradiol (E2) or the phytoestrogens genistein and daidzein regulate expression of growth-related and lipogenic genes in rainbow trout. Juvenile fish (5 mon, 65.8±1.8 g) received intraperitoneal injections of E2, genistein, or daidzein (5 μg/g body weight) or a higher dose of genistein (50 μg/g body weight). Liver and white muscle were harvested 24h post-injection. In liver, expression of vitellogenin (vtg) and estrogen receptor alpha (era1) increased in all treatments and reflected treatment estrogenicity (E2>genistein (50 μg/g)>genistein (5 μg/g)=daidzein (5 μg/g)). Estradiol and genistein (50 μg/g) reduced components of the growth hormone (GH)/insulin-like growth factor (IGF) axis in liver, including increased expression of IGF binding protein-2b1 (igfbp2b1) and reduced igfbp5b1. In liver E2 and genistein (50 μg/g) affected expression of components of the transforming growth factor beta signaling mechanism, reduced expression of ppar and rxr transcription factors, and increased expression of fatty acid synthesis genes srebp1, acly, fas, scd1, and gpat and lipid binding proteins fabp3 and lpl. In muscle E2 and genistein (50 μg/g) increased era1 and erb1 expression and decreased erb2 expression. Other genes responded to phytoestrogens in a manner that suggested regulation by estrogen receptor-independent mechanisms, including increased ghr2, igfbp2a, igfbp4, and igfbp5b1. Expression of muscle regulatory factors pax7 and myod was increased by E2 and genistein. These data indicate that genistein and daidzein affect expression of genes in rainbow trout that regulate physiological mechanisms central to growth and nutrient retention.

Topics: Animals; Estradiol; Female; Genistein; Isoflavones; Liver; Muscle Fibers, Fast-Twitch; Oncorhynchus mykiss; Phytoestrogens; Receptors, Estrogen

Two cationic derivatives of γ-cyclodextrin (GCD) were synthesized by functionalization with glycidyltrimethylammonium chloride (GTMAC) and ethylenediamine (EDA). Both these derivatives (GCD-GTMAC and GCD-EDA) have been shown to interact strongly with anionic biopolymers, unfractionated heparin (UFH) and mucin, the latter showing their mucoadhesive properties. They form inclusion complexes with daidzein (DAI), an isoflavone displaying a multitude of physiological effects, much more efficiently than the unmodified GCD. It was also shown that the complexes of these GCD derivatives with DAI and Nile Red penetrate human fibroblasts and murine hippocampal neuronal cells indicating that cationic GCD derivatives can be considered as potential delivery systems for isoflavones and other poorly water soluble compounds. Moreover, it was found that DAI delivered in cationic GCD complexes decreased the level of the cellular glycosaminoglycans (GAGs) in normal fibroblasts suggesting their possible application in the control of GAGs in mucopolysaccharidoses, lysosomal storage diseases caused by pathological accumulation of GAGs in the cells.

Topics: Absorption, Physiological; Animals; Cell Survival; Cells, Cultured; Drug Delivery Systems; Epoxy Compounds; Ethylenediamines; Fibroblasts; gamma-Cyclodextrins; Glycosaminoglycans; Hippocampus; Humans; Indicators and Reagents; Isoflavones; Lysosomal Storage Diseases; Mice; Mucopolysaccharidoses; Neurons; Phytoestrogens; Quaternary Ammonium Compounds; Solubility

Soy flour diet (MS) prevented isoflavones from stimulating MCF-7 tumor growth in athymic nude mice, indicating that other bioactive compounds in soy can negate the estrogenic properties of isoflavones. The underlying signal transduction pathways to explain the protective effects of soy flour consumption were studied here.. Ovariectomized athymic nude mice inoculated with MCF-7 human breast cancer cells were fed either Soy flour diet (MS) or purified isoflavone mix diet (MI), both with equivalent amounts of genistein. Positive controls received estradiol pellets and negative controls received sham pellets. GeneChip Human Genome U133 Plus 2.0 Array platform was used to evaluate gene expressions, and results were analyzed using bioinformatics approaches. Tumors in MS-fed mice exhibited higher expression of tumor growth suppressing genes ATP2A3 and BLNK and lower expression of oncogene MYC. Tumors in MI-fed mice expressed a higher level of oncogene MYB and a lower level of MHC-I and MHC-II, allowing tumor cells to escape immunosurveillance. MS-induced gene expression alterations were predictive of prolonged survival among estrogen-receptor-positive breast cancer patients, whilst MI-induced gene changes were predictive of shortened survival.. Our findings suggest that dietary soy flour affects gene expression differently than purified isoflavones, which may explain why soy foods prevent isoflavones-induced stimulation of MCF-7 tumor growth in athymic nude mice.

Topics: Antineoplastic Agents, Phytogenic; Breast Neoplasms; Cluster Analysis; Computational Biology; Female; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Genistein; Humans; Isoflavones; MCF-7 Cells; Neoplasm Proteins; Oligonucleotide Array Sequence Analysis; Ovariectomy; Phytoestrogens; Random Allocation; Soy Foods; Tumor Burden; Xenograft Model Antitumor Assays

The process of soybean biotransformation increases the quantity of isoflavones (daidzein and genistein), which besides being considered an alternative to estroprogestive hormone replacement therapy (HRT), are able of hindering the growth and development of tumor cells. We investigated the effects of soybean extract biotransformed by fungus on estrogen-dependent (MCF-7) and nondependent (SK-BR-3) breast cell lines. Cells were treated with different concentrations of biotransformed (BSE) and nonbiotransformed soybean extract (SE), or daidzein (D) and genistein (G) patterns isolated and in combination (D + G). Afterwards, we analyzed cell viability by MTT assay, phosphatidylserine exposure and cell permeability by flow cytometry; expression of apoptotic proteins by Western blotting. BSE promoted reduction in cell viability and increase in DNA degradation in both cell lines. In addition, we verified increase in cell permeability and in the expression of phosphatidylserine, as well as modulation in the expression of apoptotic proteins in MCF-7 cells. The cells did not show any signs of cell death when incubated with the controls (D, G, and D + G). Unknown components found in the BSE induce cell death by apoptosis and necrosis, mainly in MCF-7 cells. These processes depend on the activation of caspase-3 and involve an increase in the expression of proapoptotic molecules.

Topics: Apoptosis; Aspergillus; Biotransformation; Caspase 3; Cell Death; Cell Line, Tumor; Cell Survival; Chromatography, High Pressure Liquid; Genistein; Glycine max; Humans; Isoflavones; MCF-7 Cells; Phytoestrogens; Plant Extracts; Proto-Oncogene Proteins c-bcl-2

In prostate cancer, DNA methylation is significantly associated with tumor initiation, progression, and metastasis. Previous studies have suggested that soy phytoestrogens might regulate DNA methylation at individual candidate gene loci and that they play a crucial role as potential therapeutic agents for prostate cancer. The purpose of our study was to examine the modulation effects of phytoestrogens on a genome-wide scale in regards to DNA methylation in prostate cancer. Prostate cancer cell lines DU-145 and LNCaP were treated with 40 μM of genistein and 110 μM of daidzein. DNMT inhibitor 5-azacytidine (2 μM) and the methylating agent budesonide (2 μM) were used to compare their demethylation/methylation effects with phytoestrogens. The regulatory effects of phytoestrogens on DNA methylation were analyzed by using a methyl-DNA immunoprecipitation method coupled with Human DNA Methylation Microarrays (MeDIP-chip). We observed that the methylation profiles of 58 genes were altered by genistein and daidzein treatments in DU-145 and LNCaP prostate cancer cells. In addition, the methylation frequencies of the MAD1L1, TRAF7, KDM4B, and hTERT genes were remarkably modified by genistein treatment. Our results suggest that the modulation effects of phytoestrogens on DNA methylation essentially lead to inhibition of cell growth and induction of apoptosis. Genome-wide methylation profiling reported here suggests that epigenetic regulation mechanisms and, by extension, epigenetics-driven novel therapeutic candidates warrant further consideration in future "omics" studies of prostate cancer.

Topics: Antimetabolites, Antineoplastic; Apoptosis; Azacitidine; Budesonide; Cell Line, Tumor; Cell Proliferation; DNA Methylation; DNA, Neoplasm; Epigenesis, Genetic; Gene Expression Regulation, Neoplastic; Genistein; Glycine max; Humans; Isoflavones; Male; Oligonucleotide Array Sequence Analysis; Phytoestrogens; Prostatic Neoplasms

The aim of the present study was to assess the effect of seven days daidzein pretreatment in cecal ligation and puncture (CLP) model of sepsis.. We assessed the survival benefit of daidzein and its effect on lung injury in CLP-induced sepsis in mice and determined the bacterial load in peritoneal fluid, blood, and lung homogenates. Tumor necrosis factor α (TNF-α) and corticosterone levels were measured by enzyme-linked immunosorbent assay; relative mRNA expression was estimated by real-time polymerase chain reaction, and standard biochemical techniques were used to measure nitrite level, myeloperoxidase activity, and vascular permeability.. Daidzein pretreatment for seven days at a dose of 1 mg/kg body weight subcutaneously increased the survival time of septic mice. Daidzein decreased the bacterial load in peritoneal fluid, blood, and lungs, reduced the tumor necrosis factor α and nitrite level in plasma, and partially suppressed lung injury by reducing vascular permeability and myeloperoxidase activity in septic mice. Further, it restored the relative mRNA expressions of inducible nitric oxide synthase, glucocorticoid receptor α, and glucocorticoid receptor β genes in septic lungs were restored by daidzein pretreatment.. Daidzein pretreatment for 7 d in sepsis increased the survival time in mice, which may be relate to decrease in bacterial load, anti-inflammatory effect, and protection from lung injury.

Topics: Acute Lung Injury; Animals; Bacterial Load; Biomarkers; Cecum; Corticosterone; Drug Administration Schedule; Enzyme-Linked Immunosorbent Assay; Injections, Subcutaneous; Isoflavones; Male; Mice; Nitric Oxide; Nitrites; Peroxidase; Phytoestrogens; Real-Time Polymerase Chain Reaction; Sepsis; Treatment Outcome; Tumor Necrosis Factor-alpha

Daidzein (DZ), an isoflavone with the potential to interfere with estrogen signaling, is found in soy products, which have gained popularity due to purported beneficial effects on the cardiovascular and skeletal systems and potential antineoplastic properties. However, the ingestion of phytoestrogens has been associated with impaired reproductive function in many species. The aim of this study was to determine the long-term effects on the ovaries of rat offspring exposed to DZ or ethinyl estradiol (EE) during prenatal development. Gravid rats were administered either vehicle or 5 or 60 mg DZ/kg body weight/d or 0.002 mg 17-α EE /kg body weight/d on gestational days 6-21. Ovarian-related endpoints were investigated during adulthood in female offspring. The mean cell height of the ovarian surface epithelium was significantly reduced in all treated groups. Alterations in folliculogenesis included increased follicular atresia, a reduction in secondary and tertiary follicle numbers, and cyst formation. An elevated prevalence of a slightly prolonged estrus phase was also observed. The morphological changes to the ovarian surface epithelium are consistent with an antiproliferative effect, while ovarian folliculogenesis was adversely affected. The effects of the high dose DZ were similar to those observed with 17-α EE.

Topics: Animals; Epithelial Cells; Estrogens; Estrous Cycle; Ethinyl Estradiol; Female; Isoflavones; Ovarian Follicle; Ovary; Phytoestrogens; Pregnancy; Prenatal Exposure Delayed Effects; Rats; Rats, Sprague-Dawley

In the present study, the in vitro cytotoxicity of daidzein was evaluated in human BEL-7402, A549, HeLa, HepG-2 and MG-63 cancer cell lines. BEL-7402 cells were sensitive to daidzein treatment, with an IC50 value of 59.7±8.1 µM. Daidzein showed no cytotoxic activity toward A549, HeLa, HepG-2 and MG-63 cells. Daidzein increased the levels of reactive oxygen species (ROS) and induced a decrease in mitochondrial membrane potential. Morphological and comet assays showed that daidzein effectively induced apoptosis in BEL-7402 cells. Additionally, daidzein caused cell cycle arrest at the G2/M phase in the BEL-7402 cell line. Daidzein downregulated the expression of Bcl-2, Bcl-x and Baid proteins and upregulated the levels of Bim protein in the BEL-7402 cells. The results demonstrated that daidzein induced BEL-7402 cell apoptosis through an ROS-mediated mitochondrial dysfunction pathway.

Topics: Apoptosis; Blotting, Western; Caspases; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Proliferation; Comet Assay; Flow Cytometry; Humans; In Vitro Techniques; Isoflavones; Membrane Potential, Mitochondrial; Phytoestrogens; Proto-Oncogene Proteins c-bcl-2; Reactive Oxygen Species

The aim of this study was to assess the effects of genistein (G) and daidzein (D) on the histological, hormonal, and functional parameters of the pituitary-ovarian axis in middle-aged female rats, and to compare these effects with the effects of estradiol (E), commonly used in the prevention and treatment of menopausal symptoms. Middle-aged (12 month old) Wistar female rats subcutaneously received 35 mg/kg of G, or 35 mg/kg of D, or 0.625 mg/kg of E every day for 4 weeks. Each of the treated groups had a corresponding control group. An intact control group was also established. G and D did not change the intracellular protein content within gonadotropic and lactotropic cells, but vacuolization was observed in all the cell types. In contrast, E caused an inhibition of gonadotropic and stimulation of lactotropic cells. Also, ovaries of middle-aged female rats exposed to G or D have more healthy primordial and primary follicles and less atretic follicles. E treatment in the ovaries had a mostly negative effect, which is reflected by the increased number of atretic follicles in all tested classes. G and D provoked decrease in CuZnSOD and CAT activity, while E treatment increased MnSOD and decreased CuZnSOD and GSHPx activity. All the treatments increased serum estradiol and decreased testosterone levels, while D and E increased the serum progesterone level. In conclusion, soy phytoestrogens exhibited beneficial effects on pituitary-ovarian function in middle-aged female rats, as compared to estradiol.

Topics: Animals; Antioxidants; Disease Models, Animal; Drug Evaluation, Preclinical; Estrogen Replacement Therapy; Female; Genistein; Hormones; Isoflavones; Menopause; Ovary; Phytoestrogens; Pituitary Gland; Rats, Wistar

Phytoestrogens have been associated with subtle hormonal changes, although effects on male fecundity are largely unknown.. We evaluated associations between male urinary phytoestrogen (isoflavone and lignan) concentrations and semen quality.. This study was a prospective cohort study of 501 male partners of couples desiring pregnancy and discontinuing contraception. Each participant provided up to 2 semen samples that were analyzed for 35 semen quality endpoints the following day. Linear mixed-effects models were used to estimate associations between baseline urinary phytoestrogen concentrations and semen quality parameters, adjusted for age, body mass index (BMI), research site, and serum lipid and cotinine concentrations.. Most associations between urinary phytoestrogens and semen quality parameters were null. However, select individual phytoestrogens were associated with semen quality parameters, with associations dependent on the class of phytoestrogens and modified by BMI. Specifically, genistein and daidzein were associated with a lower percentage of normal sperm and increased abnormalities in semen morphology, with reduced associations observed as BMI increased (P < 0.05) [percentages (95% CIs) of normal morphology by WHO traditional criteria: genistein, main effect: -5.61% (-9.42%, -1.79%); interaction: 0.19% (0.06%, 0.31%) per log unit increase; daidzein, main effect: -5.35% (-9.36%, -1.34%); interaction: 0.18% (0.05%, 0.32%) per log unit increase]. Enterolactone was associated with fewer abnormalities in semen morphometry and morphology and decreased DNA fragmentation, with reduced associations observed as BMI increased (P < 0.05) [percentages (95% CIs) of abnormalities in the neck and midpiece: enterolactone, main effect: -3.35% (-6.51%, -0.19%); interaction: 0.11% (0.01%, 0.21%) per log unit increase].. These results suggest that male urinary phytoestrogen concentrations characteristic of the US population may be associated with subtle indicators of male fecundity and semen quality but were not associated with couple fecundity.

Topics: 4-Butyrolactone; Adult; Body Mass Index; Cholesterol; Cotinine; DNA Fragmentation; Endpoint Determination; Female; Fertility; Genistein; Humans; Isoflavones; Lignans; Linear Models; Male; Phytoestrogens; Pregnancy; Prospective Studies; Semen Analysis

BRF2 is a transcription factor required for synthesis of a small group of non-coding RNAs by RNA polymerase III. Overexpression of BRF2 can transform human mammary epithelial cells. In both breast and lung cancers, the BRF2 gene is amplified and overexpressed and may serve as an oncogenic driver. Furthermore, elevated BRF2 can be independently prognostic of unfavorable survival. Dietary soy isoflavones increase metastasis to lungs in a model of breast cancer and a recent study reported significantly increased cell proliferation in breast cancer patients who used soy supplementation. The soy isoflavone daidzein is a major food-derived phytoestrogen that is structurally similar to estrogen. The putative estrogenic effect of soy raises concern that high consumption of soy foods by breast cancer patients may increase tumor growth.. Expression of BRF2 RNA and protein was assayed in ER-positive or -negative human breast cancer cells after exposure to daidzein. We also measured mRNA stability, promoter methylation and response to the demethylating agent 5-azacytidine. In addition, expression was compared between mice fed diets enriched or deprived of isoflavones.. We demonstrate that the soy isoflavone daidzein specifically stimulates expression of BRF2 in ER-positive breast cancer cells, as well as the related factor BRF1. Induction is accompanied by increased levels of non-coding RNAs that are regulated by BRF2 and BRF1. Daidzein treatment stabilizes BRF2 and BRF1 mRNAs and selectively decreases methylation of the BRF2 promoter. Functional significance of demethylation is supported by induction of BRF2 by the methyltransferase inhibitor 5-azacytidine. None of these effects are observed in an ER-negative breast cancer line, when tested in parallel with ER-positive breast cancer cells. In vivo relevance is suggested by the significantly elevated levels of BRF2 mRNA detected in female mice fed a high-isoflavone commercial diet. In striking contrast, BRF2 and BRF1 mRNA levels are suppressed in matched male mice fed the same isoflavone-enriched diet.. The BRF2 gene that is implicated in cancer can be induced in human breast cancer cells by the isoflavone daidzein, through promoter demethylation and/or mRNA stabilization. Dietary isoflavones may also induce BRF2 in female mice, whereas the converse occurs in males.

Topics: Animals; Breast Neoplasms; Cell Line, Tumor; Disease Models, Animal; DNA Methylation; Female; Humans; Isoflavones; Male; Neoplasm Proteins; Phytoestrogens; Promoter Regions, Genetic; Proto-Oncogene Mas; RNA, Messenger; RNA, Neoplasm; TATA-Binding Protein Associated Factors; Transcription Factor TFIIIB

Exposure to artificial or natural endocrine disruptors, such as bisphenol A (BPA) and phytoestrogens has been demonstrated to have health effects, especially in children. Biomonitoring of BPA and phytoestrogens in human urine can be used to assess the intake levels of these compounds.. In this study, BPA and phytoestrogens in urine specimens (n = 256) collected from children in China were measured by liquid chromatography (LC)-tandem mass spectrometry (MS/MS).. BPA was detected in most specimens, with a geometric mean concentration of 1.58 ng/mL. For the first time, levels of urinary phytoestrogens in Chinese children were reported. Daidzein and enterolactone are the typical isoflavones and lignans compounds in urine, respectively.. Relatively high levels of urinary BPA indicate an increasing risk of BPA exposure to Chinese children. Urinary concentrations of daidzein in Chinese children are higher when compared with those reported in the U.S. children, while concentrations of urinary enterolactone and enterodiols are significantly lower. This suggests a significant difference in phytoestrogen intake between the children from China and from the U.S.

Topics: 4-Butyrolactone; Benzhydryl Compounds; Child; China; Chromatography, Liquid; Endocrine Disruptors; Environmental Monitoring; Female; Humans; Isoflavones; Lignans; Male; Phenols; Phytoestrogens; Risk Assessment; Tandem Mass Spectrometry; United States

Phytoestrogens are of interest because of their reported beneficial effects on many human maladies including cancer, neurodegeneration, cardiovascular disease and diabetes. As data on phytoestrogens continues to accumulate, it is clear that there is significant overlap in the cellular effects elicited by these various compounds. Here, we show that one mechanism by which a number of phytoestrogens achieve their growth inhibitory and cytoprotective effects is via induction of the mitochondrial manganese superoxide dismutase (MnSOD). Eight phytoestrogens, including resveratrol, coumestrol, kaempferol, genistein, daidzein, apigenin, isoliquirtigenin and glycitin, were tested for their ability to induce MnSOD expression in mouse C2C12 and primary myoblasts. Five of these, resveratrol, coumestrol, kaempferol, genistein and daidzein, significantly increased MnSOD expression, slowed proliferative growth and enhanced stress resistance (hydrogen peroxide LD50) . When siRNA was used to prevent the MnSOD induction by genistein, coumestrol or daidzein, none of these compounds exerted any effect on proliferative growth, and only the effect of coumestrol on stress resistance persisted. The estrogen antagonist ICI182780 prevented the increased MnSOD expression and also the changes in cell growth and stress resistance, indicating that these effects are mediated by estrogen receptors (ER). The absence of effects of resveratrol or coumestrol, but not genistein, in ERβ-null cells further indicated that this ER in particular is important in mediating these effects. Thus, an ER-mediated induction of MnSOD expression appears to underlie the growth inhibitory and cytoprotective activities of multiple phytoestrogens.

Topics: Animals; Cell Cycle; Cell Line; Cell Proliferation; Coumestrol; Cytoprotection; Estradiol; Fulvestrant; Genistein; Isoflavones; Kaempferols; Mice; Myoblasts; Phytoestrogens; Receptors, Estrogen; Resveratrol; Stilbenes; Stress, Physiological; Superoxide Dismutase

Heat stress decreases natural immunity making cows more vulnerable to diseases. A previous study reported that daidzein can enhance animal resistance to heat stress and regulate animal immunocompetence. However, it is unclear whether daidzein regulates the immune performance of late lactation cows under heat stress. In this study, late lactation cows in four groups were raised in hot weather and fed with basic diet, basic diet plus 200, 300, 400 mg/day daidzein, respectively, and the experimental period was 60 days. Blood was collected to examine the changes of serum total protein (TP), albumin (ALB), immunoglobulin G (IgG), interferon alpha (IFN-α), and interleukin-2 (IL-2). We found the levels of serum IgG and INF-α were significantly higher in late lactation cows after 300 and 400 mg/day daidzein treatment compared to those in the control group and 200 mg/day daidzein treatment (P < 0.05 or P < 0.01). Moreover, 300 and 400 mg/day daidzein treatment markedly increased serum IL-2 (P < 0.01), while the levels of serum TP and ALB were not changed by any concentration of daidzein treatment (P > 0.05). Daidzein can enhance the immunocompetence of late lactation cows and strengthen cow resistance to heat stress.

Topics: Animals; Blood Proteins; Cattle; Dose-Response Relationship, Drug; Female; Hot Temperature; Immunity, Innate; Immunocompetence; Immunoglobulin G; Interferon-alpha; Interleukin-2; Isoflavones; Lactation; Phytoestrogens; Serum Albumin; Stimulation, Chemical; Stress, Physiological

Phytoestrogens are popular alternatives to estrogen therapy however their effects on hemostasis in post-menopausal women are unknown. The aim of this study was to determine the effect of the phytoestrogens, genistein, daidzein and equol on the expression of key genes from the hemostatic system in human hepatocyte cell models and to determine the role of estrogen receptors in mediating any response seen. HepG2 cells and Hep89 cells (expressing estrogen receptor alpha (ERα)) were incubated for 24 h with 50 nM 17β-estradiol, genistein, daidzein or equol. Tissue plasminogen activator (tPA), plasminogen activator inhibitor-1 (PAI-1), Factor VII, fibrinogen γ, protein C and protein S mRNA expression were determined using TaqMan PCR. Genistein and equol increased tPA and PAI-1 expression in Hep89 cells with fold changes greater than those observed for estradiol. In HepG2 cells (which do not express ERα), PAI-1 and tPA expression were unchanged. Increased expression of Factor VII was observed in phytoestrogen treated Hep89 cells but not in similarly treated HepG2s. Prothrombin gene expression was increased in equol and daidzein treated HepG2 cells in the absence of the classical estrogen receptors. These data suggest that phytoestrogens can regulate the expression of coagulation and fibrinolytic genes in a human hepatocyte cell line; an effect which is augmented by ERα.

Topics: Blood Coagulation; Equol; Estradiol; Estrogen Receptor alpha; Factor VII; Fibrin; Gene Expression; Genistein; Hemostatics; Hep G2 Cells; Hepatocytes; Humans; Isoflavones; Phytoestrogens; Plant Extracts; Plasminogen Activator Inhibitor 1; Prothrombin; RNA, Messenger; Tissue Plasminogen Activator

Naturally occurring phytoestrogens may mimic biogenic estrogens and modulate endocrine action in vertebrates. Little is known, however, about their temporal and spatial variability in the environment and the biological effects associated with exposures. The present study assessed the environmental presence of phytoestrogens in human-impacted and relatively pristine areas. The response in larval and sexually mature fathead minnows to environmentally relevant concentrations of 3 common phytoestrogens (genistein, daidzein, and formononetin), both singly and in mixture, was also quantified. Phytoestrogens were only present in the human-impacted surface waters. When detected, mean concentrations were low (± standard deviation) in an urban lake: 1.4 ± 0.5 ng/L, 1.6 ± 0.7 ng/L, and 1.1 ± 0.2 ng/L for genistein, daidzein, and formononetin, respectively, and in treated wastewater effluent: 1.6 ± 0.4 ng/L, 1.8 ± 1.3 ng/L, and 2.0 ng/L. Biochanin A was detected twice, whereas zearalenone and coumestrol were never detected. No clear temporal trends of aqueous phytoestrogen concentration were evident. Larval survival was significantly reduced in genistein, formononetin, and mixture treatments, whereas adult male fish only exhibited subtle changes to their anatomy, physiology, and behavior. Daidzein-exposed adult females produced greater quantities of eggs. The present study indicates that genistein, daidzein, and formononetin are likely attenuated rapidly and are unlikely to cause widespread ecological harm in the absence of other stressors.

Topics: Animals; Cyprinidae; Female; Genistein; Isoflavones; Larva; Male; Phytoestrogens; Water Pollutants, Chemical

Phytoestrogens are endocrine active compounds derived from plants, including the isoflavones genistein and daidzein, and their methylated derivatives biochanin A and formononetin. These compounds have been detected at the µg/L level in the effluents of plant-processing industries and municipal treatment plants and at the ng/L level in surface waters worldwide. The present study assessed the persistence of genistein and daidzein in natural aquatic systems, specifically riverine samples. Initial concentration, temperature, sample location, and time of sample collection varied. Genistein and daidzein were found to be readily biodegradable at all tested concentrations, at both 10 °C and 20 °C, in samples collected during different seasons, and in samples from 3 different rivers. In addition, organismal responses in larval and sexually mature fathead minnows (Pimephales promelas) were quantified following exposure to microbiologically degraded phytoestrogens (genistein, daidzein, and formononetin). Products of the microbiological degradation of parent phytoestrogens did not affect larval survival, growth, or predator avoidance. Female adult fathead minnows exposed to these degradation products produced significantly fewer eggs than those exposed to a control, but no other morphological, physiological, or behavioral changes were observed with male or female minnows. The present research suggests that although phytoestrogens are not likely to persist in aquatic systems, they may pseudo-persist if discharges are continuous; in addition, caution should be exercised with respect to high-concentration effluents because of the potentially antiestrogenic effects of phytoestrogen degradates.

Topics: Animals; Cyprinidae; Female; Genistein; Isoflavones; Larva; Male; Phytoestrogens; Rivers; Water Microbiology; Water Pollutants, Chemical

In this study, we investigated the synergistic effects of daidzein (Dz) and kiwifruit on bone and equol production in ovariectomised (OVX) rats. Female Sprague-Dawley rats were randomly assigned to one of five groups: sham operated, OVX control, OVX fed 0.1% Dz-supplemented diet (OVX + Dz), OVX fed 0.1% Dz and green kiwifruit (GRK)-supplemented diet (OVX + Dz + GRK) and OVX fed 0.1% Dz and gold kiwifruit (GOK)-supplemented diet (OVX + Dz + GOK). There were no significant differences in whole body and femur bone mineral density (BMD) among groups at week 8. BMD in the OVX group significantly decreased at week 8; however, BMD in the OVX + Dz + GRK was not significantly different from baseline in the end of the study. However, supplementation with kiwifruit did not affect urinary equol concentrations, urinary ratios of equol to Dz and the composition of caecal microbiota. These results suggest that the combination of Dz and GRK may slightly reduce bone loss caused by oestrogen deficiency but does not affect equol production.

Topics: Actinidia; Animals; Bone Density; Cecum; Dietary Supplements; Drug Synergism; Equol; Female; Fruit; Humans; Isoflavones; Osteoporosis, Postmenopausal; Ovariectomy; Phytoestrogens; Plant Preparations; Rats; Rats, Sprague-Dawley

Some evidence suggests that phytoestrogens, such as soy-derived isoflavones, may have beneficial effects on cardiovascular health and glycemic control. These data are mainly limited to postmenopausal women or individuals at elevated cardiometabolic risk. There is a lack of data for pregnant women who have elevated estrogen levels and physiologically altered glucose and lipid metabolism. We analyzed data from 299 pregnant women who participated in the NHANES 2001-2008 surveys. Multivariable linear regression analyses were used to examine the association between urinary concentrations of isoflavonoids and cardiometabolic risk markers, adjusted for body mass index, pregnancy trimester, total energy intake, dietary intake of protein, fiber, and cholesterol, and demographic and lifestyle factors. Cardiometabolic risk markers were log-transformed, and geometric means were calculated by quartiles of urinary concentrations of isoflavonoids. Comparing women in the highest vs. lowest quartiles of urine total isoflavone concentrations, we observed significant, inverse associations with circulating concentrations of fasting glucose (79 vs. 88 mg/dL, P-trend = 0.0009), insulin (8.2 vs. 12.8 μU/mL, P-trend = 0.03), and triglyceride (156 vs. 185 mg/dL, P-trend = 0.02), and the homeostasis model assessment of insulin resistance (1.6 vs. 2.8, P-trend = 0.01), but not for total, low-density lipoprotein cholesterol and high-density lipoprotein cholesterol. The concentrations of individual isoflavonoids, daidzein, equol, and O-desmethylangolensin were inversely associated with some cardiometabolic risk markers, although no clear pattern emerged. These data suggest that there may be a relation between isoflavone intake and cardiometabolic risk markers in pregnant women.

Topics: Adult; Biomarkers; Blood Glucose; Body Mass Index; Cardiovascular Diseases; Cholesterol, HDL; Cross-Sectional Studies; Energy Intake; Equol; Fasting; Female; Humans; Insulin; Isoflavones; Life Style; Linear Models; Multivariate Analysis; Nutrition Surveys; Phytoestrogens; Pregnancy; Risk Factors; Triglycerides

Soybean isoflavonoids have received significant attention due to their potential anticarcinogenic and antiproliferative effects and possible role in many signal transduction pathways. However, their mechanisms of action and their molecular targets remain to be further elucidated. In this paper, we demonstrated that two soybean isoflavones (genistein and daidzein) reduced the proliferation of the human colon adenocarcinoma grade II cell line (HT-29) at concentrations of 25 and 50-100 μM, respectively. We then investigated the effects of genistein and daidzein by RT-PCR on molecules that involved in tumor development and progression by their regulation of cell proliferation. At a concentration of 50 μM genistein, there was suppressed expression of β-catenin (CTNNBIP1). Neither genistein nor daidzein affected APC (adenomatous polyposis coli) or survivin (BIRC5) expression when cells were treated with concentrations of 10 or 50 μM. These data suggest that the down-regulation of β-catenin by genistein may constitute an important determinant of the suppression of HT-29 cell growth and may be exploited for the prevention and treatment of colon cancer.

Topics: Adaptor Proteins, Signal Transducing; Adenocarcinoma; Adenomatous Polyposis Coli Protein; Anticarcinogenic Agents; Cell Growth Processes; Colonic Neoplasms; Dose-Response Relationship, Drug; Gene Expression; Genistein; Glycine max; HT29 Cells; Humans; Inhibitor of Apoptosis Proteins; Intracellular Signaling Peptides and Proteins; Isoflavones; Phytoestrogens; Phytotherapy; Survivin

Phytoestrogens (PEs), including genistein and daidzein, are plant-derived substances that mimic or antagonize estrogen action in animals. The majority of studies investigated the effects of PEs on reproduction in humans and laboratory animals. The mechanisms of phytoestrogen action on reproductive processes in domesticated animals, including pigs, are garnering increasing attention. However, very few in vivo and in vitro studies investigating the effects of PEs on adrenal glands have been carried out on models other than humans and rats. The aim of the present study was to determine whether the effects of genistein and daidzein on adrenal in vitro steroidogenesis are accompanied by changes in expression of genes encoding key steroidogenic enzymes in porcine adrenocortical cells. The following genes were analyzed: cholesterol side-chain cleavage enzyme (P450scc, CYP11A1 gene), 3β-hydroxysteroid dehydrogenase (3β-HSD, HSD3B1 gene), 17α-hydroxylase/C17-20 lyase (P450c17, CYP17A1 gene) and 21-hydroxylase (P450c21, CYP21A2 gene). Porcine adrenocortical cells collected from both luteal- and follicular-phase gilts were exposed for eight hours to genistein (10 μM), or daidzein (10 μM), in the absence or presence of ACTH (5 nM). Genistein and daidzein inhibited basal and ACTH-stimulated secretion of cortisol and corticosterone and stimulated secretion of androstenedione. PEs did not affect the expression of CYP11A1, HSD3B1, CYP17A1 and CYP21A2 in the adrenocortical cells of luteal- and follicular-phase gilts. It can be concluded that the influence of PEs on steroid secretion in porcine adrenal glands is not mediated by changes in the expression of genes encoding major steroidogenic enzymes. More studies are needed to elucidate the intracellular mechanisms leading to the PE-induced changes in adrenal steroidogenesis in pigs.

Topics: 3-Hydroxysteroid Dehydrogenases; Adrenal Cortex; Androstenedione; Animals; Cells, Cultured; Cholesterol Side-Chain Cleavage Enzyme; Corticosterone; Female; Follicular Phase; Gene Expression; Genistein; Hydrocortisone; Isoflavones; Luteal Phase; Phytoestrogens; Steroid 17-alpha-Hydroxylase; Steroid 21-Hydroxylase; Swine

Major phytoestrogens genistein and daidzein have been reported to have the ability to reverse DNA methylation in cancer cell lines. The mechanism by which genistein and daidzein have an inhibiting action on DNA methylation is not well understood. The aim of this study was to investigate the effects of soy phytoestrogens and the natural estrogen 17β-estradiol (E2) to determine whether one of the estrogen receptors is mobilized for the action of these compounds on DNA methylation. We also made a comparative study with a DNA methylation inhibitor (5-azacytidine) and a DNA methylation activator (budesonide). Three prostate cell lines, PC-3, DU-145, and LNCaP, were treated with 40 μM genistein, 110 μM daidzein, 2 μM budesonide, 2 μM 5-azacytidine, and 10 μM E2. In these 3 human prostate cancer cell lines, we performed methylation quantification using methyl-profiler-DNA-methylation analysis. Soy phytoestrogens and E2 induced a demethylation of all the promoter regions studied except for those that were unmethylated in control cells. Our results showed that E2 induces, like soy phytoestrogen, a decrease in DNA methylation in prostate cancer cell lines. This action may be mediated through ERβ.

Topics: Azacitidine; Budesonide; Cell Line, Tumor; DNA Methylation; Estradiol; Estrogen Receptor beta; Gene Expression Regulation, Neoplastic; Genistein; Glycine max; Humans; Isoflavones; Male; Phytoestrogens; Promoter Regions, Genetic; Prostatic Neoplasms

Soybeans and other legumes investigated as fishmeal replacements in aquafeeds contain phytoestrogens capable of binding to and activating estrogen receptors. Estradiol has catabolic effects in salmonid white muscle, partially through increases in protein turnover. The current study determines whether phytoestrogens promote similar effects. In rainbow trout (Oncorhynchus mykiss) primary myocyte cultures, the phytoestrogens genistein, daidzein, glycitein, and R- and S-equol reduced rates of protein synthesis and genistein, the phytoestrogen of greatest abundance in soy, also increased rates of protein degradation. Increased expression of the ubiquitin ligase fbxo32 and autophagy-related genes was observed with high concentrations of genistein (100 μM), and R- and S-equol (100 μM) also up-regulated autophagy-related genes. In contrast, low genistein concentrations in vitro (0.01-0.10 μM) and in vivo (5 μg/g body mass) decreased fbxo32 expression, suggesting a potential metabolic benefit for low levels of genistein exposure. Phytoestrogens reduced cell proliferation, indicating that effects of phytoestrogens extend from metabolic to mitogenic processes. Co-incubation of genistein with the estrogen receptor (ER) antagonist, ICI 182,780, ameliorated effects of genistein on protein degradation, but not protein synthesis or cell proliferation, indicating that effects of genistein are mediated through ER-dependent and ER-independent mechanisms. Collectively, these data warrant additional studies to determine the extent to which dietary phytoestrogens, especially genistein, affect physiological processes that impact growth and nutrient retention.

Topics: Animals; Cell Proliferation; Cells, Cultured; Equol; Female; Gene Expression; Genistein; Isoflavones; Muscle Cells; Muscle Proteins; Muscles; Oncorhynchus mykiss; Phytoestrogens; Protein Biosynthesis; Proteolysis; Receptors, Estrogen

S-(-)7-hydroxy-3-(4'-hydroxyphenyl)-chroman, or S-(-)equol, a biologically active intestinally derived bacterial metabolite of the soy isoflavones daidzin/daidzein, is not produced in neonatal life. Because its synthesis is dependent on equol-producing bacteria, we hypothesized that early nutrition may influence equol production. This prospective 2.5-year study determined the frequency of S-(-)equol production in healthy infants (n = 90) fed breast milk, soy infant formula, or cow's milk formula in their first year. Urinary S-(-)equol and daidzein were quantified by mass spectrometry after a standardized 3.5-day soy isoflavone challenge. Infants were tested at 6, 9, 12, 18, 24, and 36 months of age, and 3-day diet records were obtained at each visit to explore the effect of early and postweaning (>12 months) macronutrient and micronutrient dietary composition and S-(-)equol production. Use of antibiotics was also recorded. At age 6 months, none of the breast-fed infants produced S-(-)equol, whereas 3.8% and 6.0%, respectively, of soy and cow's milk formula-fed infants were equol producers. By age 3 years, 50% of the formula-fed infants were equol producers, compared with 25% of breast-fed infants. Use of antibiotics was prevalent among infants and may have impacted the stability of S-(-)equol production. No significant differences among the groups were observed in postweaning dietary intakes of total energy, carbohydrate, fiber, protein, fat, saturated fatty acids, or polyunsaturated fatty acids and the propensity to make S-(-)equol. In conclusion, S-(-)equol production is developmentally regulated and initially related to diet composition with the proportion of equol producers increasing over the first 3 years of life, with a trend for formula feeding favoring S-(-)equol production.

Topics: Animals; Bottle Feeding; Breast Feeding; Child, Preschool; Diet; Equol; Feeding Behavior; Humans; Infant; Infant Formula; Infant Nutritional Physiological Phenomena; Intestinal Mucosa; Intestines; Isoflavones; Male; Milk; Milk, Human; Phytoestrogens; Prospective Studies; Soy Foods; Weaning

Isoflavones are the most common phytoestrogens found in human diets. However, it is still not clear whether isoflavones have effects on the reproductive and the endocrine systems under normal dietary intake and overdose. The aim of this study was to determine how the most important isoflavones, genistein and daidzein, affect androgen and glucorticoid levels on male prepuberal rats. A hundred and seventy-five 30-day-old male Wistar rats were dosed orally by stomach tube every day for 35 days, with saline solution, low and high doses of genistein, daidzein and a mixture of both. Serum samples were analysed by an enzyme immunoassay for hormone determinations. In control group, there was a peak of testosterone (T) and dihydrotestosterone levels associated to the onset of puberty, at the third week. However, in low-dose groups, the same peak was found at the fourth week (p < 0.05), indicating a delay in the onset of puberty in these groups. Moreover, high doses groups serum androgen levels were significantly lower (p < 0.05) than the control group from the first week until fifth week. This fact was supported by a epididymal histological analysis that indicate in low doses there were several content of spermatozoa at fourth week and in high doses there were few content of spermatozoa. Besides, corticosterone levels followed the same pattern of androgens in all groups. We can conclude that oral administration of isoflavones in male rats decreased the secretion of androgens and glucocorticoids causing a delay in the onset of puberty and may cause physiological and developmental problems.

Topics: Androgens; Animals; Corticosterone; Dihydrotestosterone; Dose-Response Relationship, Drug; Epididymis; Genistein; Glucocorticoids; Isoflavones; Male; Phytoestrogens; Rats; Rats, Wistar; Sexual Maturation; Sperm Count; Testosterone

The inflow, transformation, and attenuation of natural steroid hormones and phytoestrogens and estrogenic activity were assessed across the lagoon/sprayfield system of a prototypical commercial swine sow operation. Free and conjugated steroid hormones (estrogens, androgens, and progesterone) were detected in urine and feces of sows across reproductive stages, with progesterone being the most abundant steroid hormone. Excreta also contained phytoestrogens indicative of a soy-based diet, particularly, daidzein, genistein, and equol. During storage in barn pits and the anaerobic lagoon, conjugated hormones dissipated, and androgens and progesterone were attenuated. Estrone and equol persisted along the waste disposal route. Following application of lagoon slurry to agricultural soils, all analytes exhibited attenuation within 2 days. However, analytes including estrone, androstenedione, progesterone, and equol remained detectable in soil at 2 months postapplication. Estrogenic activity in the yeast estrogen screen and T47D-KBluc in vitro bioassays generally tracked well with analyte concentrations. Estrone was found to be the greatest contributor to estrogenic activity across all sample types. This investigation encompasses the most comprehensive suite of natural hormone and phytoestrogen analytes examined to date across a livestock lagoon/sprayfield and provides global insight into the fate of these analytes in this widely used waste management system.

Topics: Agriculture; Androgens; Androstenedione; Animals; Diet; Environmental Monitoring; Equol; Estrogens; Estrone; Feces; Genistein; Hormones; Isoflavones; Phytoestrogens; Progesterone; Steroids; Swine; Urine

S-Equol is enantioselectively produced from the isoflavone daidzein by gut microflora and is absorbed by the body. An increase of pancreatic β-cell death is directly associated with defects in insulin secretion and an increased risk of type 2 diabetes mellitus. In the present study, we demonstrate that only the S-enantiomer has suppressive effects against alloxan-induced oxidative stress in INS-1 pancreatic β-cells. S-Equol reduced alloxan-induced cell death in a dose-dependent manner, whereas R-equol had no effects. In contrast, no significant differences were observed between the enantiomers in estrogenic activity. The cytoprotective effects of S-equol were stronger than those of its precursor daidzein and were blocked by the protein synthesis inhibitor cycloheximide. The cytoprotection was diminished when cells were incubated with a protein kinase A (PKA) inhibitor (H89), but not an estrogen receptor inhibitor. S-Equol increased intracellular cAMP levels in an enantioselective manner. S-Equol, but not R-equol, induced phosphorylation of cAMP-response element-binding protein at Ser 133, and induced cAMP-response element-mediated transcription, both of which were diminished in the presence of H89. Taken together, these results show that S-equol enantioselectively increases the survival of INS-1 cells presumably through activating PKA signaling. Thus, S-equol might have applications as an anti-type 2 diabetic agent.

Topics: Alloxan; Animals; Bacteria; Cell Death; Cell Line; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Diabetes Mellitus, Type 2; Equol; Insulin; Insulin-Secreting Cells; Isoflavones; Isomerism; Oxidative Stress; Phosphorylation; Phytoestrogens; Plant Extracts; Rats; Signal Transduction

Dietary phytoestrogens are metabolized or converted in the gastrointestinal tract of ruminants, only limited knowledge exists on the extent and location of this conversion in vivo. The objective of this study was to quantify the gastro-intestinal metabolism of phytoestrogens in lactating dairy cows fed silages with different botanical composition. Four lactating rumen cannulated Norwegian Red cattle were assigned to a 4 × 4 Latin square with 1 cow per treatment period of 3 wk. The 4 treatment silages were prepared from grasslands with different botanical compositions: organically managed short-term timothy (Phleum pratense L.) and red clover (Trifolium pratense L.) ley (2 yr old: ORG-SG); organically managed long-term grassland with a high proportion of unsown species (6 yr old; ORG-LG); conventionally managed perennial ryegrass (Lolium perenne L.) ley (CON-PR); and conventionally managed timothy ley (CON-TI). The herbages were cut, wilted, and preserved with additive in round bales, fed as a mix of the first and third cut at 90% of ad libitum intake, and contributed to 70% of the total dry matter intake. Milk, feed, omasal digesta, urine, and feces were collected at the end of each period and analyzed for the concentrations of phytoestrogens by using a liquid chromatography-tandem mass spectrometry technique. Concentration of total isoflavones was highest in ORG-SG and lowest in CON-TI silage, whereas the content of total lignans was highest in the grass silages. The isoflavones were extensively metabolized in the rumen on all diets, and the recovery of formononetin and daidzein in omasum, mainly as equol, averaged 0.11 mg/mg. The apparent intestinal metabolism was less severe as, on average, 0.29 mg/mg of the omasal flow was recovered in feces. The plant lignans were also strongly degraded in the rumen. However, the flow of lignans to omasum and excretion in feces were, on average, 7.2- and 5.2-fold higher, respectively, than the intake of the plant lignans matairesinol and secoisolariciresinol, known as precursors of mammalian lignans. Thus, excretion to milk could not be directly related to intake, implying that plant lignans other than matairesinol and secoisolariciresinol in forage are precursors for enterolactone production in the rumen and for its content in milk. Equol followed mainly the flow of large particles out of the rumen, whereas the mammalian lignans were distributed between phases proportional to dry matter flow. The main metabolism of

Topics: Animals; Cattle; Chromatography, Liquid; Diet; Feces; Female; Furans; Isoflavones; Lactation; Lignans; Lolium; Mass Spectrometry; Milk; Omasum; Phleum; Phytoestrogens; Poaceae; Rumen; Silage; Trifolium

To investigate the effect of Bushengzhuyang Fang (Yangjing Capsule, YJC) on penile erectile function and its action mechanisms in rats.. Fifty-six male SD rats were randomly divided into seven groups of equal number: blank control, daidzein, daidzein + testosterone, daidzein + sildenafil, daidzein + low-dose YJC, daidzein + medium-dose YJC, and daidzein + high-dose YJC. The rats in the blank control group were treated intragastrically with normal saline and those in the other groups with daidzein at the dose of 100 mg per kg per day for 30 days. Then the last five groups received additionally testosterone (4 mg per kg per day), sildenafil (2.5 mg per kg per day), low-dose YJC, (0.315 mg per kg per day), medium-dose YJC (0.63 mg per kg per day), and high-dose YJC (1. 26 mg per kg per day), respectively. At 0, 30 and 60 days of treatment, we observed the apomorphine-induced spontaneous erectile response and pathological changes in the corpus cavernosum of the rats, recorded the number of penile erection and erectile incubation period, and determined the serum levels of testosterone (T) and luteinizing hormone (LH).. At 30 days of treatment, the number of apomorphine-induced erections was decreased, the erectile incubation period prolonged, and the serum levels of T and LH reduced remarkably in all groups of rats (P < 0.05). Compared with the findings at 30 days, the number of penile erections was significantly decreased at 60 days in the daidzein group (1.39 ± 0.42 vs 2.67 ± 0.33, P < 0.05) and daidzein + low-dose YJC group (1.33 ± 0.49 vs 2.83 ± 0.61, P < 0.05); the erectile incubation period was markedly ex- tended ([16.33 ± 3.11] vs [8.50 ± 0.93] min and [15.50 ± 3.21] vs [8.63 ± 1.54] min, P < 0.05); and the serum levels of T ([5.34 ± 0.89] vs [1.24 ± 0.30] ng/ml and [5.28 ± 1.12] vs [2.07 ± 0.76] ng/ml, P < 0.05) and LH ([3.62 ± 0.37] vs [2.09 ± 0.12] ng/ml and [3.79 ± 0.28] vs [2.17 ± 0.33] ng/ml, P < 0.05) were significantly reduced in the daidzein and daidzein + low-dose YJC groups, respectively. Pathological examination revealed slightly decreased cavernous sinuses and blood vessels in the corpus cavernosum of the rats in the daidzein + testosterone, daidzein + sildenafil, daidzein + medium-dose YJC, and daidzein + high-dose YJC groups as compared with those in the blank control group.. High-dose Yangjing Capsule is efficacious for the recovery of erectile function in rats, especially for phytoestrogen-induced erectile dysfunction.

Topics: Animals; Apomorphine; Drugs, Chinese Herbal; Erectile Dysfunction; Humans; Isoflavones; Luteinizing Hormone; Male; Penile Erection; Penis; Phytoestrogens; Phytotherapy; Piperazines; Purines; Rats; Rats, Sprague-Dawley; Sildenafil Citrate; Sulfonamides; Testosterone; Vasodilator Agents

Soy products, commonly used as a protein source in farm animals' diets, contain considerable quantities of non-nutrient constituents such as phytoestrogens. Genistein and daidzein are known to affect the reproductive processes in humans and animals. However, reports concerning phytoestrogens and porcine adrenal steroidogenesis are scarce, and the adrenal mechanism of phytoestrogen action in species other than humans and rodents is poorly recognized. The goal of the present paper was to examine the in vitro effects of genistein and daidzein on the activity of key enzymes for cortisol and corticosterone synthesis in porcine adrenocortical cells harvested during the luteal or follicular phase of the porcine estrous cycle. The cells were treated with genistein or daidzein (10 μM), with or without ACTH (5 nM), in the presence or absence of precursors (1 μM) of cortisol (pregnenolone, P5; progesterone, P4; 17-hydroxyprogesterone, 17OH-P4; or 11-deoxycortisol, 11d-cortisol) or corticosterone: (P5 or P4) synthesis. The supplementation of a medium with P5, P4, 17OH-P4 or 11d-cortisol enabled us to measure the activity of cholesterol side-chain cleavage enzyme (P450scc), 3β-hydroxysteroid dehydrogenase (3β-HSD), 17α-hydroxylase/C17-20 lyase (P450c17) or 21-hydroxylase (P450c21) and 11β-hydroxylase (P45011β), respectively. We demonstrated that in sexually mature, cyclic pigs, regardless of the phase of the estrous cycle, phytoestrogens genistein and daidzein suppressed basal and ACTH-stimulated in vitro secretion of cortisol and corticosterone via progesterone synthesis inhibition. This indicates that phytoestrogens specifically inhibit the 3β-HSD activity in porcine adrenocortical cells. We suggest that genistein and daidzein present in soy products may negatively affect glucocorticoid synthesis of mature gilts by disrupting adrenal steroidogenesis at the 3β-HSD level.

Topics: 3-Hydroxysteroid Dehydrogenases; Adrenal Cortex; Adrenocorticotropic Hormone; Animals; Corticosterone; Estrous Cycle; Female; Genistein; Glucocorticoids; Hydrocortisone; Isoflavones; Phytoestrogens; Pregnenolone; Swine

1. The effect of daidzein, a naturally occurring phytoestrogen, on the reproductive performance of 120 female Zhedong White geese was determined. The geese were divided into 4 groups which were fed on diets containing 0 (Control), 10 (Da1), 20 (Da2) and 30 (Da3) mg daidzein per kg diet. Egg production and weight, fertility and hatchability rates, concentrations of estradiol (E2), triiodothyronine (T3), progesterone (P4), thyroxine (T4) and growth hormone (GH) in serum, and mRNA levels of gonadotropin releasing hormone (GnRH), β-follicle stimulating hormone (FSHβ), follicle stimulating hormone receptor (FSHR), oestrogen receptor1 (ESR1), oestrogen receptor2 (ESR2), prolactin (PRL), prolactin receptor (PRLR), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in the hypothalamus-pituitary-gonadal axis (HPGA) were measured. 2. Daidzein increased egg weight and fertility but had no detectable effect on egg production and hatchability. 3. Daidzein affected serum P4 and GH concentrations and T4 rhythm, up-regulated GnRH mRNA and PRLR mRNA levels in the hypothalamus, down-regulated PRLR mRNA in the hypothalamus, PRL mRNA in the pituitary, and ESR2 mRNA levels in the ovary, respectively. The mRNA rhythms of PRLR in the hypothalamus, PRL, PRLR and FSHβ in the pituitary, FSHR, ESR1 and ESR2 in the ovary were significantly changed in the Da2 group. 4. It is suggested that an appropriate dose of daidzein might improve reproductive performance by affecting serum hormone concentrations and rhythms and regulating gene mRNA levels in the HPGA of female Zhedong White geese.

Topics: Animal Feed; Animal Husbandry; Animals; Diet; Dose-Response Relationship, Drug; Female; Geese; Gene Expression Regulation; Hormones; Hypothalamo-Hypophyseal System; Isoflavones; Ovary; Phytoestrogens; Radioimmunoassay; Real-Time Polymerase Chain Reaction; Reproduction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Seasons

Experimental data indicate that gestational exposures to estrogenic compounds impact risk of hypospadias. We examined whether risk of hypospadias (i.e., a congenital malformation in which the opening of the penile urethra occurs on the ventral side of the penis) was associated with maternal intake of phytoestrogens, given their potential impact on estrogen metabolism. The analysis included data on mothers of 1,250 hypospadias cases and 3,118 controls who delivered their infants from 1997 to 2005 and participated in the National Birth Defects Prevention Study, a multistate, population-based, case-control study. After adjustment for several covariates, high intakes of daidzein, genistein, glycetin, secoisolariciresinol, total isoflavones, total lignans, and total phytoestrogens were associated with reduced risks; odds ratios comparing intakes ≥90th percentile with intakes between the 11th and 89th percentiles ranged from 0.6 to 0.8. For example, the odds ratio for total phytoestrogen intake was 0.7 (95% confidence interval: 0.5, 1.0). This study represents the first large-scale analysis of phytoestrogen intake and hypospadias. The observed associations merit investigation in additional populations before firm conclusions can be reached.

Topics: Adult; Butylene Glycols; Case-Control Studies; Diet, Vegetarian; Female; Genistein; Humans; Hypospadias; Infant, Newborn; Isoflavones; Lignans; Male; Odds Ratio; Phytoestrogens; Pregnancy; Prenatal Exposure Delayed Effects; Risk Assessment; Risk Factors; Surveys and Questionnaires

Phytoestrogens (PEs) are naturally occurring chemical constituents of certain plants. The internal PE exposures, mainly from diet, vary among different populations and in different regions due to various eating habits. To investigate the potential relationship between urinary PE levels and idiopathic male infertility and semen quality in Chinese adult males, 608 idiopathic infertile men and 469 fertile controls were recruited by eligibility screening procedures. Individual exposure to PEs was measured using UPLC-MS/MS as spot urinary concentrations of 6 PEs (daidzein, DAI; equol, EQU; genistein, GEN; naringenin, NAR; coumestrol, COU; and secoisolariciresinol, SEC), which were adjusted with urinary creatinine (CR). Semen quality was assessed by sperm concentration, number per ejaculum and motility. We found that exposures to DAI, GEN and SEC were significantly associated with idiopathic male infertility (P-value for trend=0.036; 0.002; and 0.0001, respectively), while these exposures had stronger association with infertile subjects with at least one abnormal semen parameter than those with all normal semen parameters. Exposures to DAI, GEN and SEC were also related to idiopathic male infertility with abnormal sperm concentration, number per ejaculum and motility (P-value for trend<0.05), while these exposures had stronger association with the infertile men with abnormal sperm number per ejaculum. These findings provide the evidence that PE exposures are related to male reproductive function and raise a public health concern because that exposure to PEs is ubiquitous in China.

Topics: Adult; Butylene Glycols; China; Diet; Feeding Behavior; Genistein; Humans; Infertility, Male; Isoflavones; Lignans; Male; Phytoestrogens; Semen; Semen Analysis; Sperm Count; Tandem Mass Spectrometry

The Ministry of Health Biomonitoring Study estimated exposure of individuals in the Israeli population to bisphenol A (BPA), organophosphate (OP) pesticides, phthalates, cotinine, polycyclic aromatic hydrocarbons (PAHs), and the phytoestrogenic compounds genistein and daidzein.. In 2011, 250 individuals (ages 20-74) were recruited from five different regions in Israel. Urine samples were collected and questionnaire data were obtained, including detailed dietary data (food frequency questionnaire and 24hour recall). Urinary samples were analyzed for BPA, OP metabolites (dialkyl phosphates), phthalate metabolites, cotinine, PAH metabolites, genistein, and daidzein.. BPA urinary concentrations were above the limit of quantification (LOQ) in 89% of the samples whereas urinary concentrations of phthalate metabolites were above the LOQ in 92-100% of the samples. PAH metabolites were above the LOQ in 63-99% of the samples whereas OP metabolites were above the LOQ in 44-100% of the samples. All non-smoking participants had detectable levels of cotinine in their urine; 63% had levels above the LOQ, and the rate of quantification was high compared to the general non-smoking population in Canada. Median creatinine adjusted concentrations of several OP metabolites (dimethyl phosphate, dimethyl thiophosphate) were high in our study population compared to the general US and Canadian populations. Median creatinine adjusted urinary BPA concentrations in the study population were comparable to those in Belgium and Korea; higher than those reported for the general US, German, and Canadian populations; and very low compared to health-based threshold values. Phthalate concentrations were higher in our study population compared to the general US population but values were very low compared to health-based threshold values. Median creatinine adjusted PAH concentrations were generally comparable to those reported for the general US population; median creatinine adjusted daidzein concentrations were high in our population compared to the general US population whereas genistein concentrations were comparable.. We interpreted observed urinary contaminant levels observed in our study by comparing values with health-based threshold values and/or values from international human biomonitoring studies. Using this data interpretation scheme, we identified two contaminants as being of potential public health concern and high priority for public health policy intervention: environmental tobacco smoke (ETS) and OP pesticides. We used the data collected in this study to support public health policy interventions. We plan to conduct a follow-up biomonitoring study in 2015 to measure ETS and OP exposure in the general population in Israel, to evaluate the effectiveness of relevant policy interventions.

Topics: Adult; Aged; Benzhydryl Compounds; Cotinine; Environmental Exposure; Environmental Monitoring; Environmental Pollutants; Female; Genistein; Humans; Insecticides; Isoflavones; Israel; Male; Middle Aged; Organophosphorus Compounds; Phenols; Phthalic Acids; Phytoestrogens; Polycyclic Aromatic Hydrocarbons; Tobacco Smoke Pollution; Young Adult

To determine whether daidzein improves insulin resistance by modifying weight gain, visceral fat accumulation, blood lipids and serum cytokines levels in ovariectomized Sprague-Dawley rats.. Twenty-eight 12-week-old female rats were divided into three groups: the sham-operated group (SHAM) (n =10), the ovariectomized group receiving daidzein therapy (DAID) (n =10), and the ovariectomized control group (Control) (n =8). The rats in the DAID group received 50 mg/kg daidzein via gavage daily. Weight and food intake were recorded every 2 weeks. All of the animals were euthanized 12 weeks after ovariectomy, after which their fasting insulin, glucose, blood lipids, estradiol, interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), adiponectin and leptin levels were measured.. After 12 weeks, the ovariectomized rats demonstrated an increase in their body weight and visceral fat; compared to the SHAM rats, the ovariectomized rats also experienced a significant increase in their serum IL-6 levels and insulin resistance, which was calculated using the homeostatic model assessment of insulin resistance (HOMA-IR) (p <0.05). Daidzein therapy decreased weight gain, visceral fat, the HOMA-IR index and IL-6 levels that were induced by ovariectomy. Rats which had received daidzein therapy had lower levels of TNF-α, leptin and blood lipids (except for high density lipoprotein cholesterol) than the other two groups. IL-6 levels positively correlated with the HOMA-IR index in all of the rats after adjustment for body weight (r =0.495; p =0.016).. We conclude that daidzein can improve insulin resistance induced by ovariectomy by decreasing weight gain, visceral fat accumulation, blood lipids, TNF-α, leptin and IL-6 levels.

Topics: Analysis of Variance; Animals; Blood Glucose; Cholesterol, HDL; Cholesterol, LDL; Female; Insulin Resistance; Interleukin-6; Intra-Abdominal Fat; Isoflavones; Leptin; Ovariectomy; Phytoestrogens; Rats; Rats, Sprague-Dawley; Triglycerides; Tumor Necrosis Factor-alpha; Weight Gain

To evaluate whether soybean extracts and estrogens present additive effects on adult rat uterus.. Fifty ovariectomized rats were randomly divided into five equal groups of ten animals: Control, treated with vehicle; SE46 and SE120, treated with 46 and 120 mg/kg soybean concentrated extract (SE), respectively; EE, treated with conjugated equine estrogens (CE) 50 μg/kg; SE120 + EE, treated with 50 μg/kg (CE) plus 120 mg/kg SE. The substances were administered daily by gavage for 21 consecutive days. Thereafter the animals were weighed and killed by decapitation; trunk blood was collected for hormone determinations. Uteri were removed immediately and fixed in 10% formaldehyde, followed by dehydration, embedding in paraffin and 6-m sections staining with hematoxylin and eosin for histomorphometric analyses of myometrium and endometrium. After ANOVA analysis of the data, the study was complemented with the Tukey-Kramer test for multiple comparisons.. The concentrated extract of soybean at high concentration (SE 120 kg/mg) and estrogens proved to have a trophic effect on the uterus (endometrium and myometrium) of castrated rats. In groups SE120, EE and SE120 + EE, all morphometric parameters examined (number of glands, eosinophils, blood vessels and the glandular area) were increased. No significant addictive effects of soybean extract plus estrogens were detected in the SE120 + EE group.. Our results indicate that soy extract has a trophic effect on rat uterine structures. Treatment of ovariectomized rats with a concentrated soy extract in combination with conjugated estrogens had no addictive effect on the uterine response.

Topics: Analysis of Variance; Animals; Endometrium; Estradiol; Estrogens; Estrogens, Conjugated (USP); Female; Genistein; Glycine max; Isoflavones; Myometrium; Organ Size; Ovariectomy; Phytoestrogens; Plant Extracts; Progesterone; Rats; Uterus

Daidzein belongs to the group of isoflavones, found in a wide variety of plant-derived foods, especially in soybeans and soy-based foods. In this study, the effect of daidzein on human gastric carcinoma cells (BGC-823) and its mechanism were investigated. MTT assay was applied in the detection of the inhibitory effects of daidzein on cell proliferation. Hoechst-propidium iodide staining and flow cytometry were used to examine the apoptosis as well as the mitochondrial transmembrane potential. Western blotting was performed to detect the expression of apoptosis-associated proteins: cleaved PARP, cleaved caspase-9, cleaved caspase-3, Bcl-2, and Bax. Daidzein significantly inhibited the growth and proliferation of human gastric carcinoma cells (BGC-823) in a concentration- and time-dependent manner. Furthermore, it was found that an insult of daidzein to BGC-823 cells caused them to die by disruption of mitochondrial transmembrane potential, demonstrated not only by staining dead cells for phosphatidylserine but also by the up-regulation (cleaved PARP, cleaved caspase-9, cleaved caspase-3, Bax) and down-regulation (Bcl-2) of proteins associated with apoptosis and survival; whereas, the pan-caspase inhibitor z-VAD-fmk could partially rescue cells against damage of daidzein. Taken together, the results of this study demonstrate that daidzein significantly induces apoptosis via a mitochondrial pathway. Specifically, daidzein induced a change in the Bax/Bcl-2 ratios and activation of caspases-3 and -9 and the cleavage of PARP. Therefore, daidzein has the potential for use as a therapeutic agent for the treatment of gastric carcinoma.

Topics: Apoptosis; bcl-2-Associated X Protein; Blotting, Western; Caspase 3; Caspase 9; Cell Line, Tumor; Cell Proliferation; Dose-Response Relationship, Drug; Down-Regulation; Enzyme Activation; Flow Cytometry; Humans; Isoflavones; Membrane Potential, Mitochondrial; Mitochondria; Phytoestrogens; Poly(ADP-ribose) Polymerases; Proto-Oncogene Proteins c-bcl-2; Signal Transduction; Stomach Neoplasms; Time Factors

The growth and development of prostate gland is governed by testosterone. Testosterone helps in maintaining the adipose tissue stores of the body. It is well documented that with advancing age there has been a gradual decline in testosterone levels. Our aim was to study the protective role of daidzein on flutamide-induced androgen deprivation on matrix degrading genes, lipid profile and oxidative stress in Wistar rats. Sub-chronic (60 days) flutamide (30 mg/kg b.wt) administration resulted in marked increase in expressions of matrix degrading genes [matrix metalloproteases 9 and urokinase plasminogen activation receptor]. Additionally, it increased the levels of low density lipoproteins, total cholesterol, triglycerides, and lowered the levels of high density lipoproteins and endogenous antioxidant levels. Oral administration of daidzein (20 and 60 mg/kg b.wt) restituted the levels to normal. Daidzein administration resulted in amelioration of the prostate atrophy, degeneracy and invasiveness induced by flutamide. Our findings suggest that the daidzein may be given as dietary supplement to patients who are on androgen deprivation therapy, to minimize the adverse effects related to it and also retarding susceptibility of patients to cardiovascular diseases.

Topics: Androgen Antagonists; Animals; Atrophy; Catalase; Cholesterol; Flutamide; Glutathione; Glutathione Peroxidase; Glutathione Reductase; Isoflavones; Lipid Metabolism; Lipids; Lipoproteins, HDL; Lipoproteins, LDL; Male; Matrix Metalloproteinase 9; Orchiectomy; Oxidative Stress; Phytoestrogens; Prostate; Rats; Rats, Wistar; Receptors, Urokinase Plasminogen Activator; Triglycerides

Daidzein (Daid) has been implicated in bone health for its estrogen-'like' effects but low bioavailability, unfavorable metabolism and uterine estrogenicity impede its clinical potential. This study was aimed at assessing isoformononetin (Isoformo), a naturally occurring methoxydaidzein, for bone anabolic effect by overcoming the pitfalls associated with Daid.. Sprague-Dawley ovariectomized (OVx) rats with established osteopenia were administered Isoformo, 17β-oestradiol (E2) or human parathyroid hormone. Efficacy was evaluated by bone microarchitecture using microcomputed tomography and determination of new bone formation by fluorescent labeling of bone. Osteoblast apoptosis was measured by co-labeling of bone sections with Runx-2 and TUNEL. Biochemical markers of bone metabolism were measured by ELISA. Plasma and bone marrow levels of Isoformo and Daid were determined by LC-MS-MS. Rat bone marrow stromal cells were harvested to study osteoblastic differentiation by Isoformo and Daid. New born rat pups were injected with Isoformo and Daid to study the effect of the compounds on the expression of osteogenic genes in the calvaria by real time PCR.. In osteopenic rats, Isoformo treatment restored trabecular microarchitecture, increased new bone formation, increased the serum osteogenic marker (procollagen N-terminal propeptide), decreased resorptive marker (urinary C-terminal teleopeptide of type I collagen) and diminished osteoblast apoptosis in bone. At the most effective osteogenic dose of Isoformo, plasma and bone marrow levels were comprised of ~90% Isoformo and the rest, Daid. Isoformo at the concentration reaching the bone marrow achieved out of its most effective oral dosing induced stromal cell mineralization and osteogenic gene expression in the calvaria of neonatal rats. Isoformo exhibited uterine safety.. Our study demonstrates that Isoformo reverses established osteopenia in adult OVx rats likely via its pro-survival effect on osteoblasts. Given its bone anabolic and anti-catabolic effects accompanied with safety at uterine level we propose its potential in the management of postmenopausal osteoporosis.

Topics: Animals; Apoptosis; Biomarkers; Bone and Bones; Bone Density Conservation Agents; Bone Diseases, Metabolic; Bone Resorption; Calcification, Physiologic; Female; Isoflavones; Metabolism; Osteoblasts; Osteogenesis; Osteoporosis; Ovariectomy; Phytoestrogens; Phytotherapy; Plant Extracts; Rats; Rats, Sprague-Dawley; Stromal Cells; Uterus

Cytochrome c (CytC) released from mitochondria induces apoptosis in both normal and tumor cells. Expression of Fas ligand (FasL) helps maintain tumor cell survival by inducing apoptosis of Fas-bearing anti-tumor immune cells. A risk of endometrial cancer has been reported to associate with phytoestrogen consumption. Therefore the effects of phytoestrogens, genistein and daidzein, on FasL and CytC protein expression were examined in primary cultured porcine endometrial cells (PE) and human cancerous endometrial cells (RL95-2) by Western blot analysis. Both cells were cultured in standard medium (SM) and switched to estrogen-deprived medium (SF) with or without 17beta-estradiol (E, 1 nM), genistein (10 microM) or daidzein (10 microM) for 48 h. FasL (25 kDa) which was found only in RL95-2 cells was upregulated in SF compared to SM. Treatment of RL95-2 cells with E, daidzein or genistein significantly increased the FasL expression by 7-10 folds. In the present study, low level of CytC was detected in both cells cultured in SM but markedly increased in SF by 1.5-2 folds. The SF-induced increase in CytC level was reversed by genistein or daidzein while E suppressed CytC in PE cells, but not in RL95-2 cells. The findings suggest that genistein and daidzein appear to act as a survival factor by inhibiting intracellular apoptogenic initiator in both normal and cancer endometrial cells. In addition, estrogen and phytoestrogens inducing the death signal FasL expressed by cancerous endometrial cells may cause the tumor progression. Thus, consuming phytoestrogen as a supplement should be awareness in patient with endometrial cancer.

Topics: Analysis of Variance; Animals; Apoptosis; Blotting, Western; Cells, Cultured; Cytochromes c; Endometrial Neoplasms; Estradiol; Fas Ligand Protein; Female; Genistein; Humans; Isoflavones; Phytoestrogens; Swine

The role of peroxisome proliferator-activated receptors (PPARs) as anti-inflammatory mediators has been established, and the fact that some isoflavones are dual agonists of PPARα/γ indicates the involvement of PPARα and/or PPARγ in the anti-inflammatory action of certain isoflavones. However, the dependency of isoflavones on PPARs in their anti-inflammatory action has not been demonstrated. Here, we report the dependency of an isoflavone biochanin A and the independency of another isoflavone genistein in relation to PPARγ to ameliorate the cytokine secretion profile of lipopolysaccharide (LPS)-stimulated mouse RAW264.7 macrophages. A total amount of 10 µmol/l of biochanin A or genistein significantly suppressed the secretion of tumor necrosis factor α (TNFα) and interleukin-6 (IL-6) in LPS-induced RAW264.7 cells, whereas another two isoflavones, formononectin and daidzein, only significantly suppressed the secretion of IL-6. Their anti-inflammatory efficiencies were not in correspondence with their PPARα/γ agonist activities. Inhibition of PPARγ activity by its antagonist GW9662 significantly reversed the anti-inflammatory effect of biochanin A but not genistein, which demonstrated the dependency of biochanin A and the independency of genistein on PPARγ in their anti-inflammatory actions. Meanwhile, the PPARγ-dependency of biochanin A was further confirmed by the result that the suppression of LPS-induced NF-κB activation by biochanin A was reversed following GW9662 co-treatment. Moreover, inhibition of PPARα activity by its antagonist MK886 did not significantly reverse the anti-inflammatory effects of biochanin A and genistein, indicating that their anti-inflammatory properties were PPARα-independent.

Topics: Anilides; Animals; Anticarcinogenic Agents; Cell Line; Genistein; Humans; Indoles; Interleukin-6; Isoflavones; Lipopolysaccharides; Lipoxygenase Inhibitors; Macrophages; Mice; NF-kappa B; Phytoestrogens; PPAR alpha; PPAR gamma; Tumor Necrosis Factor-alpha

Host modulatory agents directed at inhibiting specific proinflammatory mediators could be beneficial in terms of attenuating periodontal disease progression and potentially enhancing therapeutic responses. The aim of this study was to investigate whether daidzein could modulate the production inflammatory mediators in macrophages stimulated with lipopolysaccharide (LPS) from Prevotella intermedia, a pathogen implicated in periodontal disease, and to delineate underlying mechanisms of action.. LPS was extracted from P. intermedia ATCC 25611 cells by the standard hot phenol-water method. The amounts of nitric oxide (NO) and interleukin-6 (IL-6) secreted into the culture medium were assayed. A real-time PCR was performed to quantify inducible nitric oxide synthase (iNOS) and IL-6 mRNA expression. We used immunoblot analysis to characterize iNOS protein expression, phosphrylation of c-Jun N-terminal kinase (JNK) and p38, degradation of inhibitory κB-α (IκB-α), nuclear translocation of nuclear factor-κB (NF-κB) subunits and phosphorylation of signal transducer and activator of transcription 1 (STAT1). The DNA-binding activity of NF-κB was assessed by using ELISA-based kits.. Daidzein significantly inhibited the production of NO and IL-6, as well as their mRNA expression, in P. intermedia LPS-treated RAW264.7 cells. The JNK and p38 pathways were not involved in the regulation of LPS-induced NO and IL-6 release by daidzein. Daidzein inhibited the degradation of IκB-α induced by P. intermedia LPS. In addition, daidzein suppressed NF-κB transcriptional activity via regulation of the nuclear translocation and DNA-binding activity of NF-κB p50 subunit and blocked STAT1 phosphorylation.. Although additional studies are required to dissect the molecular mechanism of action, our results suggest that daidzein could be a promising agent for treating inflammatory periodontal disease. Further research in animal models of periodontitis is necessary to better evaluate the potential of daidzein as a novel therapeutic agent to treat periodontal disease.

Topics: Animals; Anti-Inflammatory Agents; Bacteriological Techniques; Cell Culture Techniques; Cell Line; Growth Inhibitors; I-kappa B Kinase; Inflammation Mediators; Interleukin-6; Isoflavones; Janus Kinase 2; JNK Mitogen-Activated Protein Kinases; Lipopolysaccharides; Macrophages; Mice; NF-kappa B; NF-kappa B p50 Subunit; Nitric Oxide Synthase Type II; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Phytoestrogens; Prevotella intermedia; STAT1 Transcription Factor; Transcription Factor RelA

In the present work the feasibility of β-cyclodextrin in complexation was explored, as a tool for improving the solubility and biological ability of daidzein derivatives. A series of phosphorylated daidzein derivatives featuring different chain lengths were synthesized through a modified Atherton-Todd reaction and their inclusion complexes with βCD were prepared by coprecipitation method. The inclusion complexation behavior was studied by fluorescence, UV, FT-IR, MS and (1)H NMR. The results showed that only phosphorylated daidzein derivative carrying small substituent group ((C(2)H(5)O)(2)PO) entered the cavity of βCD and formed 1:1 inclusion complex. The formation constant was 175(mol/L)(-1).

Topics: beta-Cyclodextrins; Isoflavones; Magnetic Resonance Spectroscopy; Phosphorylation; Phytoestrogens; Solubility; Spectrophotometry, Ultraviolet; Spectroscopy, Fourier Transform Infrared

The risks of hormone replacement therapy have led to a search for new alternatives such as phytoestrogens, plant compounds with estrogen-like biological activity. Isoflavones are the phytoestrogens most extensively studied and can be found in soybean, red clover and other plants. Due to this estrogen-like activity, phytoestrogens can have some effect on atherosclerosis. Human umbilical vein endothelial cells (HUVEC) have been extensively used to study the biology and pathobiology of human endothelial cells and most of the knowledge acquired is due to experiments with cultures of these cells.. To evaluate the effects of the phytoestrogen extracts from Glycine max soy bean, genistein, formononetin, biochanin A and daidzein, as well as a mixture of these extracts (Mix), on expression of adhesion molecules, VCAM-1, ICAM-1 and E-selectin, by endothelial cell HUVEC, stimulated with lipopolysaccharide.. HUVEC were cultured in medium EBM(2), pretreated with isoflavones for 24 and 48 h and then stimulated with lipopolysaccharide; in addition, isoflavones were added, after stimulation by lipopolysaccharide, to HUVEC. We evaluated the production of VCAM-1, ICAM-1 and E-selectin on cell surface, by cell-based enzyme immunoassay, and of sVCAM-1, sICAM-1 and sE-selectin in culture supernatant, by ELISA.. Genistein, formononetin, biochanin A and daidzein, as well as the Mix were able to reduce VCAM-1, ICAM-1 and E-selectin on cell surface and in culture supernatant. Conclusion Isoflavones extracted from Glycine max soy bean, in vitro, presented antiatherogenic effects, reducing the expression of adhesion molecules and acting as preventive agents as well as therapeutic agents.

Topics: Cells, Cultured; Drug Therapy, Combination; E-Selectin; Genistein; Glycine max; Human Umbilical Vein Endothelial Cells; Humans; Intercellular Adhesion Molecule-1; Isoflavones; Phytoestrogens; Time Factors; Vascular Cell Adhesion Molecule-1

Phytochemicals found in soy and other legumes have been speculated to reduce the risk of endometrial cancer; however, inconsistent findings have been reported in the few epidemiological studies conducted to date.. We conducted a prospective analysis of 46 027 nonhysterectomized postmenopausal women who were recruited into the Multiethnic Cohort (MEC) Study between August 1993 and August 1996 and provided detailed baseline information on diet and other endometrial cancer risk factors. A total of 489 women diagnosed with incident endometrial cancer were identified through the Surveillance, Epidemiology, and End Results tumor registry linkages during a median follow-up period of 13.6 years. Cox proportional hazards models were used to estimate multivariable-adjusted relative risks (RRs) and 95% confidence intervals (CIs) for endometrial cancer associated with dietary intake of legumes, soy, and tofu, and for total isoflavones and specific isoflavones (daidzein, genistein, or glycitein). Truncated (age 50-89 years) age-adjusted incidence rates were calculated by applying age-specific rates within isoflavone quintiles to the overall MEC population eligible for endometrial cancer. To estimate the percentage of endometrial cancers that may have been prevented by consuming the highest quintile of total isoflavones, the partial population attributable risk percent was calculated.. A reduced risk of endometrial cancer was associated with total isoflavone intake (highest vs lowest quintile, ≥7.82 vs <1.59 mg per 1000 kcal/d, RR = 0.66, 95% CI = 0.47 to 0.91), daidzein intake (highest vs lowest quintile, ≥3.54 vs <0.70 mg per 1000 kcal/d, RR = 0.64, 95% CI = 0.46 to 0.90), and genistein intake (highest vs lowest quintile, ≥3.40 vs <0.69 mg per 1000 kcal/d, RR = 0.66, 95% CI = 0.47 to 0.91). No statistically significant association with endometrial cancer risk was observed for increasing intake of legumes, soy, tofu, or glycitein. Truncated age-adjusted incidence rates of endometrial cancer for the highest vs lowest quintile of total isoflavone intake were 55 vs 107 per 100 000 women per year, respectively. The partial population attributable risk percent for total isoflavone intake lower than the highest quintile was 26.7% (95% CI = 5.3% to 45.8%).. This study suggests that greater consumption of isoflavone-containing foods is associated with a reduced risk of endometrial cancer in this population of nonhysterectomized postmenopausal women.

Topics: Aged; Aged, 80 and over; Cohort Studies; Endometrial Neoplasms; Fabaceae; Feeding Behavior; Female; Genetic Predisposition to Disease; Genistein; Glycine max; Growth Inhibitors; Humans; Incidence; Isoflavones; Life Style; Middle Aged; Phytoestrogens; Postmenopause; Proportional Hazards Models; Prospective Studies; Risk Assessment; Risk Factors; SEER Program; Soy Foods; Surveys and Questionnaires; United States

Due to the health risks attributed to perimenopausal hormone therapy, phytoestrogens such as flavonoids are receiving widespread attention to help alleviate menopausal symptoms, including hormone-driven mood disorders. Based on our previous reporter gene study regarding their transactivational activity in raphe nuclei cells from a brain region involved in regulation of mood disturbances, we herein study their effects on the regulation of expression of 17β-estradiol (E2)-regulated genes. DNA microarray was used to globally assess E2-induced gene expression in RNDA cells, a rat raphe nuclei-derived cellular model expressing oestrogen receptor β. Out of 212 regulated genes, six were selected for verification and as endpoints for the effect of flavonoids on the regulation of mRNA expression in proliferating as well as differentiating RNDA cells. Under proliferative conditions, E2 up-regulated mRNA expression of Cml-5, Sox-18 and Krt-19. Similar effects were observed in response to 8-prenylnaringenin (8-PN), genistein (GEN), daidzein (DAI) and equol (EQ). In line with E2, mRNA expression of Nefm and Zdhhc-2 was down-regulated following 8-PN, GEN, DAI, EQ and naringenin treatment. No regulation was observed on Slc6a4 mRNA expression in response to E2 or the flavonoids in proliferating RNDA cells. When cells were shifted to conditions promoting differentiation, changes in cell morphology, in mRNA expression levels and in responsiveness towards E2 and the tested flavonoids were noticed. These expression studies additionally highlighted some of the genes as markers for RNDA cellular differentiation. RNDA cells should prove useful to elucidate molecular and cellular mechanisms of exogenous oestrogen receptor ligands with neural cell populations.

Topics: Animals; Cell Differentiation; Cell Line; Cell Proliferation; Down-Regulation; Equol; Estradiol; Estrogen Receptor beta; Estrogens; Flavanones; Flavonoids; Gene Expression Profiling; Gene Expression Regulation; Genistein; Isoflavones; Phytoestrogens; Rats; RNA, Messenger; SOXF Transcription Factors

The captive southern white rhinoceros (SWR; Ceratotherium simum simum) population serves as an important genetic reservoir critical to the conservation of this vulnerable species. Unfortunately, captive populations are declining due to the poor reproductive success of captive-born females. Captive female SWR exhibit reproductive problems suggested to result from continual ovarian follicular activity and prolonged exposure to endogenous estrogen. However, we investigated the potential role of exogenous dietary phytoestrogens in the reproductive failure of SWR by cloning and characterizing in vitro phytoestrogen binding and activation of recombinant SWR estrogen receptors (ESR). We compared those characteristics with recombinant greater one-horned rhinoceros (GOHR; Rhinoceros unicornis) ESR, a species that receives similar captive diets yet reproduces relatively well. Our results indicate that phytoestrogens bind rhino ESR in a manner similar to other vertebrate species, but there are no differences found in phytoestrogen binding affinity of SWR ESR compared with GOHR ESR. However, species-specific differences in ESR activation by phytoestrogens were detected. The phytoestrogen coumestrol stimulated greater maximal activation of SWR ESR1 than GOHR ESR1. SWR ESR2 were also more sensitive to phytoestrogens and were activated to a greater extent by both coumestrol and daidzein. The concentrations in which significant differences in ESR activation occurred (10(-7) to 10(-5) m) are consistent with circulating concentrations measured in other vertebrate species. Taken together, these findings suggest that phytoestrogens potentially pose a risk to the reproductive health of captive SWR. However, additional studies are needed to further clarify the physiological role of dietary phytoestrogens in the reduced fertility of this species.

Topics: Animals; Cloning, Molecular; Coumestrol; Estrogens; Female; HEK293 Cells; Humans; Isoflavones; Perissodactyla; Phytoestrogens; Protein Binding; Protein Structure, Tertiary; Receptors, Estrogen; Reproduction

Estrogen deficiency was produced in female Sprague-Dawley rats by surgical removal of both the ovaries and these animals were used 4 weeks later. Endothelium-dependent and endothelium-independent relaxations due to acetylcholine and sodium nitroprusside were observed respectively, in isolated rat thoracic aortic ring preparation. Extent of lipid peroxidation was measured by estimating serum TBARS. Integrity of vascular endothelium was assessed using hematoxylin and eosin staining. Generation of nitric oxide was measured indirectly, by estimating serum and urinary nitrite/nitrate concentration. Ovariectomy produced significant vascular endothelial dysfunction, measured in terms of reduced acetylcholine-induced endothelium-dependent vasorelaxation, serum and urinary nitrite/nitrate concentration and impairment of integrity of vascular endothelium. Administration of daidzein (0.2 mgkg(-1)day(-1), sc 0.4 mgkg(-1)day(-1), sc and 0.8 mgkg(-1)day(-1), sc) and Atorvastatin (30 mgkg(-1)day(-1), po Positive Control) for one week markedly improved vascular endothelial dysfunction due to increase in nitric oxide bioavailability perhaps by inhibiting caveolin-1 and activation of PI3K-AKT pathway.

Topics: Acetylcholine; Animals; Aorta, Thoracic; Aortic Diseases; Caveolin 1; Endothelium, Vascular; Female; Isoflavones; Nitrates; Nitrites; Nitroprusside; Ovariectomy; Phytoestrogens; Rats; Rats, Sprague-Dawley; Vasodilation; Vasodilator Agents

Osteoblast and osteoclast activity is disrupted in post-menopausal osteoporosis. Thus, to fully address this imbalance, therapies should reduce bone resorption and promote bone formation. Dietary factors such as phyto-oestrogens and Zn have beneficial effects on osteoblast and osteoclast activity. However, the effect of combinations of these factors has not been widely studied. We therefore examined the effect of coumestrol, daidzein and genistein in the presence or absence of zinc sulphate (Zn) on osteoclast and osteoblast activity. Osteoclast differentiation and bone resorption were significantly reduced by coumestrol (10- 7 m), daidzein (10- 5 m) and genistein (10- 7 m); and this direct anti-osteoclastic action was unaffected by Zn (10- 5 m). In addition, Zn augmented the inhibitory effect of phyto-oestrogens on the osteoblast-derived stimulus for osteoclast formation, significantly reducing the ratio of receptor activator of NF-κB ligand (RANKL)-to-osteoprotegerin mRNA expression in human osteoblast. We then examined the effect of these compounds on osteoblast activity. Mineralisation was enhanced by coumestrol (10- 5 to 10- 7 m), daidzein (10- 5 to 10- 6 m) and genistein (10- 5 m); and Zn significantly augmented this response. Zn and phyto-oestrogens also significantly enhanced alkaline phosphatase activity and Runt-related transcription factor 2 (Runx2) mRNA expression. On the other hand, Zn blunted phyto-oestrogen-induced type I collagen and osteocalcin expression and suppressed coumestrol and daidzein-stimulated osterix expression. Zn may therefore modify the anabolic action of phyto-oestrogens, promoting characteristics associated with early rather than late stages of osteoblast differentiation. Our data suggest that while Zn enhances the anti-osteoclastic effect of phyto-oestrogens, it may limit aspects of their anabolic action on bone matrix formation.

Topics: Acid Phosphatase; Animals; Bone Resorption; Cell Line; Cell Proliferation; Collagen Type I; Core Binding Factor Alpha 1 Subunit; Coumestrol; Gene Expression Regulation; Genistein; Humans; Isoenzymes; Isoflavones; Ligands; Macrophages; Mice; NF-kappa B; Osteoblasts; Osteoclasts; Osteoprotegerin; Phytoestrogens; Sp7 Transcription Factor; Tartrate-Resistant Acid Phosphatase; Transcription Factors; Zinc

Although soy phytoestrogens have been postulated to exert a protective effect against breast cancer, the attendant mechanisms, in particular epigenetics underpinnings, have remained elusive. We investigated the putative effects on DNA methylation by two naturally occurring isoflavones, genistein and daidzein, in a study of the BRCA1 and BRCA2 oncosuppressor genes in breast cancer cell lines (MCF-7, MDA-MB 231, and MCF10a). A demethylant agent, the 5-azacytidine, and a methylant, the budesonide, were used as treatment controls. DNA methylation of BRCA1 and BRCA2 was investigated with methylated DNA immunoprecipitation coupled with PCR. In parallel, protein expression was determined by Western blot, immunohistochemistry, and confocal microscopy. Our results suggest that treatment with 18.5 μM Genistein or 78.5 μM Daidzein might reverse DNA hypermethylation and restore the expression of the oncosuppressor genes BRCA1 and BRCA2. 5-Azacitydine also enhanced the reexpression of these genes while budesonide had an opposite effect. To the best of our knowledge, these observations, while requiring replication, provide new evidence on potential epigenetic mechanisms by which genistein and daidzein might contribute to regulation of the BRCA1 and BRCA2. Future studies are warranted on whether the demethylating effect of genistein and daidzein is global or focused on select candidate genes.

Topics: BRCA1 Protein; BRCA2 Protein; Breast Neoplasms; Budesonide; Cell Line, Tumor; DNA Methylation; Female; Gene Expression; Gene Expression Regulation, Neoplastic; Genistein; Glycine max; Humans; Immunohistochemistry; Isoflavones; Phytoestrogens

Phytoestrogens are non steroidal compounds that can bind to estrogen receptors, mimicking some effects of estradiol (E(2)). These compounds are widespread among legumes, which are used as pasture, and their importance in animal agriculture has increased. Mesquite (Prosopis sp) is a widespread legume, widely used to feed several livestock species in Mexico. The main product of mesquite is the pod, which is considered high quality food. As a legume, it could be assumed that mesquite contains some amounts of phytoestrogens which might induce potential estrogenic effects. However, to our knowledge, there are no reports regarding the possible estrogenic activity of this legume either in livestock or in animal models such as the rat. Therefore, in this study, we evaluated the potential estrogenic effects of mesquite pod extract on several aspects of behavior and reproductive physiology of the female rat. The effects of the extract were compared with those of E(2) and two isoflavones: daidzein (DAI) and genistein (GEN). The following treatments were given to groups of intact and ovariectomized (OVX) female rats: vehicle; mesquite pod extract; E(2); GEN; DAI. Compared to vehicle groups, mesquite pod extract, DAI, GEN, and E(2) increased uterine weight and induced growth in vaginal and uterine epithelia. In intact rats, mesquite pod extract, GEN and DAI altered estrous cyclicity, decreased lordotic quotient and intensity of lordosis. In OVX rats, mesquite pod extract, DAI and GEN induced vaginal estrus, increased vaginal epithelium height, and induced lordosis, although its intensity was reduced, compared with intact rats in estrus and E2-treated rats. These results suggest that mesquite pod extract could have estrogenic activity. However, the presence of phytoestrogens in this legume remains to be confirmed.

Topics: Animals; Epithelium; Estradiol; Estrous Cycle; Female; Genistein; Isoflavones; Organ Size; Ovariectomy; Phytoestrogens; Plant Extracts; Prosopis; Rats; Rats, Wistar; Reproduction; Seeds; Sexual Behavior, Animal; Uterus; Vagina

Previous studies have suggested that the daidzein metabolite equol rather than daidzein itself contributes to the beneficial effect of soya foods in the prevention of CVD. The aim of the present study is to examine the proportion of equol excretion in Chinese adults and compare plasma lipids and carotid artery intima-media thickness (IMT) between equol excretors and non-excretors, and to evaluate the effect of soya isoflavone intakes on serum lipids and IMT in either equol excretors or non-excretors. Subjects (n 572; women n 362, men n 210) were recruited for the present study. An overnight urine sample was provided by each subject on their usual diet to quantify urinary concentrations of daidzein and equol. Far-wall IMT was determined by B-mode ultrasound in the right carotid at two sites, carotid bulb (CB-IMT) and common carotid artery (CCA-IMT), and fasting serum lipids were measured. Habitual dietary intakes were estimated with a FFQ, and soya isoflavone intake derived from the FFQ was assessed. Of the 572 subjects, the proportion of equol excretors on their usual diet was 25·0 % (n 143). Compared with non-excretors, equol excretors showed significantly lower serum TAG (-38·2 (95 % CI -70·4, -5·9) %, P = 0·012) and CCA-IMT (-4·9 (95 % CI -9·7, -0·3) %, P = 0·033). Equol excretors with higher daily isoflavone intakes (-5·4 mg/d) had significantly lower IMT (-16·2 %, P = 0·035) and tended to have higher HDL-cholesterol (P = 0·055) than did those with lower daily isoflavone intakes (1·5 mg/d), while no association was observed between soya isoflavone intakes and serum lipids or IMT in non-excretors. In conclusion, the benefits of soya isoflavones in preventing CVD may be apparent among equol excretors only.

Topics: Adult; Aged; Cardiovascular Diseases; Carotid Arteries; Carotid Intima-Media Thickness; China; Diet; Equol; Female; Glycine max; Humans; Isoflavones; Lipids; Male; Middle Aged; Phytoestrogens; Risk Factors; Seeds; Surveys and Questionnaires

Genistein and daidzein are two estrogenic compounds derived from plants, especially legumes. This research begins to explore their environmental fate, focusing on direct and indirect photolysis. UV-visible spectra for both compounds at varying pH values were taken, the pK(a) values for both compounds were measured, and UV-visible spectra for each protonation state were determined. The loss of both compounds in deionized water was observed upon exposure to natural sunlight, and the quantum yields were determined for each protonation state. In Mississippi River water, direct photolysis does not account for all of the loss of genistein and daidzein. The mechanism of indirect photolysis was probed using quenchers and sensitizers, and results suggest that daidzein is transformed mainly via direct photolysis and singlet oxygenation, while genistein is transformed mainly via reaction with triplet-state natural organic matter. The parameters determined in this study will allow for estimation of the concentration of genistein and daidzein in sunlit surface waters, which will allow for assessment of any risks posed to aquatic wildlife.

Topics: Furans; Genistein; Half-Life; Hydrogen-Ion Concentration; Hydroxyl Radical; Isoflavones; Kinetics; Mississippi; Photolysis; Phytoestrogens; Protons; Quantum Theory; Rivers; Singlet Oxygen; Spectrophotometry, Ultraviolet; Ultraviolet Rays

Estrogen replacement therapy (ERT) reduces the risk of Alzheimer's disease and symptoms in postmenopausal and elderly women. However, ERT is associated with increased risk of uterine and breast cancer. Dietary phytoestrogens have been suggested as a potential alternative to ERT, while little information is available regarding the effects and the underlying mechanisms of such treatment on central neuron function. The present study aimed to determine the effects of phytoestrogens including genistein and daidzein on the proliferation and survival of the hippocampus neural cells, which are of importance in learning and memory function.. H19-7/IGF-IR neural cell line was cultured in DMEM absented of serum for 72 h, and treated with various concentrations of genistein, daidzein or 17β-estradiol. Neuronal cell viability and proliferation were determined by MTT and BrdU assay, respectively Cell cycle analysis was performed using flow cytometry. The effects of genistein and daidzein on brain-derived neurotrophic factor (BDNF) mRNA and protein expression were determined by RT-PCR and ELISA, respectively. The effect of Trk receptors inhibitor on genistein and daidzein - induced hippocampus neuronal cell proliferation was also examined.. 17β-estradiol, genistein and daidzein ranged from 20 nM to 2000 nM significantly promoted hippocampus neuronal cell viability and proliferation. Similar to the effect of 17β-estradiol, genistein and daidzein induced an increase in the percentage of cells in S phase. Genistein and daidzein significantly increased the expression of BDNF mRNA and protein levels. The effect of genistien and daidzein on hippocampus neuronal proliferation was blocked by K252a, a selective Trk receptors inhibitor.. This study concluded that genistein and daidzein improved hippocampus neuronal cell viability and proliferation in vitro. These neuroprotective effects might be mediated by BDNF-Trk pathway.

Topics: Animals; Brain-Derived Neurotrophic Factor; Cell Cycle; Cell Line; Cell Proliferation; Cell Survival; Estradiol; Estrogen Replacement Therapy; Gene Expression Regulation; Genistein; Hippocampus; Isoflavones; Neurons; Neuroprotective Agents; Phytoestrogens; Rats

HIV-1 viral protein Tat partially mediates the neural dysfunction and neuronal cell death associated with HIV-1 induced neurodegeneration and neurocognitive disorders. Soy isoflavones provide protection against various neurotoxic insults to maintain neuronal function and thus help preserve neurocognitive capacity.. We demonstrate in primary cortical cell cultures that 17β-estradiol or isoflavones (genistein or daidzein) attenuate Tat(1-86)-induced expression of apoptotic proteins and subsequent cell death. Exposure of cultured neurons to the estrogen receptor antagonist ICI 182,780 abolished the anti-apoptotic actions of isoflavones. Use of ERα or ERβ specific antagonists determined the involvement of both ER isoforms in genistein and daidzein inhibition of caspase activity; ERβ selectively mediated downregulation of mitochondrial pro-apoptotic protein Bax. The findings suggest soy isoflavones effectively diminished HIV-1 Tat-induced apoptotic signaling.. Collectively, our results suggest that soy isoflavones represent an adjunctive therapeutic option with combination anti-retroviral therapy (cART) to preserve neuronal functioning and sustain neurocognitive abilities of HIV-1 infected persons.

Topics: Animals; Apoptosis; Down-Regulation; Estrogen Receptor alpha; Estrogen Receptor beta; Genistein; HIV-1; Isoflavones; Neurons; Peptide Fragments; Phytoestrogens; Rats; Rats, Sprague-Dawley; Receptors, Estrogen; tat Gene Products, Human Immunodeficiency Virus

Phytoestrogens, such as the soy isoflavones genistein and daidzein, are suggested to beneficially affect lipid metabolism in humans and thereby contribute to healthy ageing. New evidences show that phytoestrogens might slow ageing processes also by affecting immune processes.. We tested in the nematode Caenorhabditis elegans the effects of 17β-estradiol, genistein, and daidzein on resistance versus the nematode pathogen Photorhabdus luminescens with focus on vitellogenins, which are invertebrate estrogen-responsive genes that encode homologues to ApoB100 with impact on immune functions. Here, we show that the estrogen 17β-estradiol increases the resistance of C. elegans versus P. luminescens by enhancing vitellogenin-expression at the mRNA and protein level. Knockdown of single out of five functional vits by RNA-interference blunted the life-extending effects under heat-stress of 17β-estradiol, demonstrating a lack of redundancy for the vitellogenins. RNAi for nhr-14, a suggested nuclear hormone receptor for estrogens, displayed no influence on 17β-estradiol effects. The soy isoflavone genistein reduced vitellogenin-expression and also resistance versus P. luminescens whereas daidzein increased resistance versus the pathogen in a vitellogenin-dependent manner.. Our studies show that induction of estrogen-responsive vitellogenin(s) by the phytoestrogen daidzein potently increases resistance of C. elegans versus pathogenic bacteria and heat whereas genistein acts in an antiestrogenic manner.

Topics: Animals; Caenorhabditis elegans; Caenorhabditis elegans Proteins; Diet; Down-Regulation; Genistein; Heat-Shock Response; Humans; Immunity, Innate; Immunologic Factors; Isoflavones; Photorhabdus; Phytoestrogens; Protein Isoforms; Receptors, Estrogen; RNA Interference; RNA, Messenger; Up-Regulation; Vitellogenins

In mice, exposure to isoflavones (ISO), abundant in soy infant formula, during the first 5 d of life alters structural and functional development of reproductive organs. Effects of longer exposures are unknown. The study objective was to evaluate whether exposure to a combination of daidzein and genistein in the first 10 compared to 5 d of life results in greater adverse effects on ovarian and uterine structure in adult mice. Thirteen litters of 8-12 pups were cross-fostered and randomized to corn oil or ISO (2 mg daidzein + 5 mg genistein/kg body weight/d) for the first 5 or 10 d of life. The 10-d protocol mimicked the period when infants are fed soy protein formula (SPF) but avoids the time when suckling pups can consume mother's diet. Body and organ weights, and histology of ovaries and uteri were analyzed. There were no differences in the ovary or uterus weight, number of ovarian follicles, number of multiple oocyte follicles, or percent of ovarian cysts with 5 or 10 d ISO intervention compared to respective controls. The 10-d ISO group had higher body weights from 6 d to 4 mo of age and a higher percent of hyperplasia in the oviduct than the respective control. Lower number of ovarian corpus lutea and a higher incidence of abnormal changes were reported in the uteri of both ISO groups compared to their respective controls. Five and 10-d exposure to ISO had similar long-lasting adverse effects on the structure of ovaries and uterus in adult mice. Only the 10-d ISO exposure resulted in greater body weight gain at adulthood.

Topics: Animals; Animals, Suckling; Female; Genistein; Humans; Hyperplasia; Infant; Infant Formula; Isoflavones; Lactation; Maternal Nutritional Physiological Phenomena; Mice; Mice, Inbred Strains; Ovary; Oviducts; Phytoestrogens; Random Allocation; Soy Foods; Uterus; Weight Gain

Some bladder disorders, such as obstructive bladder and hyperactivity, may be caused partly by ischemia/reperfusion injury (I/R). The neuroprotective effects of estrogens were demonstrated in in vitro studies and a great interest in soy isoflavones (genistein and daidzein) as alternative to the synthetic estrogen receptor modulators for therapeutic use has been pointed out. The aim of this study was to investigate the effect of genistein and daidzein, on rat detrusor smooth muscle contractility and their possible neuroprotective role against I/R-like condition. Whole rat urinary bladders were subjected to in vitro anoxia-glucopenia (A-G) and reperfusion (R) in the absence or presence of drugs and response to electrical field stimulation (EFS) of intrinsic nerves evaluated. Furthermore rats were treated in vivo for 1 week with the phytoestrogens and the same in vitro protocol was applied to the ex vivo bladders. Antioxidant activity of genistein and daidzein on the A-G/R model was determined by measuring malonyldialdehyde (MDA). Moreover, hormones plasma levels were determined by radioimmunoassay. Genistein and daidzein administered either in vitro or in vivo showed significant neuroprotective effect and antioxidant activity. Testosterone and 17β-estradiol plasma levels were not modified by daidzein, while a significant decrease of testosterone in genistein treated rats was evident. Moreover both phytoestrogens significantly decreased detrusor contractions induced by EFS in a concentration-dependent manner. For being either neuroprotective and myorelaxant, genistein and daidzein could be considered a good lead for new therapeutic agents to protect the urinary bladder from hyperactivity and nerve damage.

Topics: Animals; Antioxidants; Electric Stimulation; Estradiol; Genistein; Glycine max; Hypoxia; Isoflavones; Lipid Peroxidation; Male; Muscle Contraction; Neuroprotective Agents; Phytoestrogens; Rats; Rats, Wistar; Reperfusion Injury; Testosterone; Urinary Bladder

Bone density has been suggested as a marker of cumulative hormone exposure. Small studies also suggest that patterns of daidzein metabolism may be related to hormone concentrations. To our knowledge, no studies in premenopausal women have compared bone density by daidzein-metabolizing phenotypes in the absence of a soy intervention.. The objective was to evaluate the relationship between daidzein-metabolizing phenotypes [equol and O-desmethylangolensin (ODMA) production] and bone density and body composition in premenopausal women in the United States.. Two hundred and three women attended a clinic visit during which their bone density and body composition were measured by DXA, and 200 (99 %) provided a urine sample following a 3-day soy challenge. Samples were analyzed for isoflavones to determine daidzein-metabolizing phenotypes.. In adjusted analyses, there were no differences in hip, spine, femoral neck, or head bone mineral density (BMD) or body composition between producers and non-producers of either equol or ODMA (P > .05).. In this population of low-soy consuming premenopausal women, there were no associations between daidzein-metabolizing phenotypes and hip, spine, femoral neck, or head BMD or body composition, suggesting that these phenotypes per se do not influence premenopausal bone density or body composition.

Topics: Absorptiometry, Photon; Adult; Body Composition; Bone Density; Cross-Sectional Studies; Equol; Female; Femur Neck; Glycine max; Humans; Ilium; Isoflavones; Lumbar Vertebrae; Middle Aged; Phenotype; Phytoestrogens; Premenopause; Skull; United States

Daidzein is a naturally occurring compound and has various health benefits. However, its effects on intestinal smooth muscle contractility remain unknown.. The present study was to characterize the effects of daidzein on the contractility of isolated jejunal smooth muscle and its underlying mechanisms.. Ex vivo assay was selected as the major method to determine the effects of daidzein on the contractility of isolated jejunal smooth muscle fragment (JSMF).. Daidzein (5-160 µmol/L) inhibited the contractility of JSMF in normal contractile state and in a dose-dependent manner. Daidzein also inhibited the contractility of JSMF induced by ACh, histamine, erythromycin and high Ca²⁺, respectively, and decreased charcoal propulsion in the small intestine in vivo. The inhibitory effects of daidzein were partially blocked by phentolamine or propranolol and were abolished in the presence of varapamil or at Ca²⁺-free assay condition. However, the inhibitory effects of daidzein on jejunal contraction were not significantly influenced by nitric oxide (NO) synthase inhibitor L-NG-nitro-arginine (L-NNA). Daidzein was also found to directly inhibit the phosphorylation and Mg²⁺-ATPase activity of smooth muscle myosin.. The results implicated that α- and β-adrenergic receptors were involved in the inhibitory effects produced by daidzein rather than via NO pathway. As a phytoestrogen, daidzein has shown its potential value in relieving the hypercontractility of small intestine.

Topics: Animals; Ca(2+) Mg(2+)-ATPase; Dose-Response Relationship, Drug; Enzyme Inhibitors; Gastrointestinal Agents; Gastrointestinal Motility; Gastrointestinal Transit; Isoflavones; Jejunum; Mice; Muscle Contraction; Muscle, Smooth; Myosins; Nitric Oxide; Phosphorylation; Phytoestrogens; Rats; Rats, Sprague-Dawley; Receptors, Adrenergic, alpha; Receptors, Adrenergic, beta; Signal Transduction; Time Factors

To investigate whether genes involved in AKT/nuclear factor kappa B signaling and/or gene-environment interactions between the genes and phytoestrogens may be susceptible factors for gastric cancer.. The representative single nucleotide polymorphisms (SNPs) identified during the primary analysis (screening a total of 622 SNPs within ± 5 kbp of the 51 target gene locations) were further investigated in 317 matched case-control sets. The summary odds ratios (ORs) and 95% confidence intervals (CIs) for gastric cancer were calculated. Interaction effects between the SNPs and phytoestrogen biomarkers (genistein, daidzein, equol, and enterolactone) were computed. CDK1 rs4145643, FAS rs6586161, and FAS rs1468063 in the AKT signaling pathway presented significant genetic effects on gastric cancer (OR = 0.81 (95% CI: 0.66-0.99) for CDK1 rs4145643; OR = 1.27 (95% CI: 1.03-1.58) for FAS rs6586161; OR = 1.29 (95% CI: 1.03-1.56) for FAS rs1468063; Cochran Q statistics > 0.10). Risk alleles of FAS rs6586161, FAS rs1468063, MAP3K1 rs16886448, and MAP3K1 rs252902 showed significant interaction effects with enterolactone (p(interaction) < 0.05).. CDK1 and FAS genes involved in AKT signaling and influenced by anti-carcinogenic property of phytoestrogens can play a role as susceptible genetic factors in gastric carcinogenesis. FAS and MAP3K1 genes significantly interact with enterolactone, thereby modifying the individual's risk for gastric cancer.

Topics: 4-Butyrolactone; Aged; Anticarcinogenic Agents; Asian People; Case-Control Studies; CDC2 Protein Kinase; Equol; fas Receptor; Female; Gene-Environment Interaction; Genetic Predisposition to Disease; Genistein; Humans; Isoflavones; Lignans; Male; MAP Kinase Kinase Kinase 1; Middle Aged; Odds Ratio; Phytoestrogens; Polymorphism, Single Nucleotide; Proto-Oncogene Proteins c-akt; Republic of Korea; Signal Transduction; Stomach Neoplasms

Dietary soy is thought to be cancer-preventive; however, the beneficial effects of soy on established breast cancer is controversial. We recently demonstrated that dietary daidzein or combined soy isoflavones (genistein, daidzein, and glycitein) increased primary mammary tumor growth and metastasis. Cancer-promoting molecules, including eukaryotic protein synthesis initiation factors (eIF) eIF4G and eIF4E, were up-regulated in mammary tumors from mice that received dietary daidzein. Herein, we show that increased eIF expression in tumor extracts of mice after daidzein diets is associated with protein expression of mRNAs with internal ribosome entry sites (IRES) that are sensitive to eIF4E and eIF4G levels. Results with metastatic cancer cell lines show that some of the effects of daidzein in vivo can be recapitulated by the daidzein metabolite equol. In vitro, equol, but not daidzein, up-regulated eIF4G without affecting eIF4E or its regulator, 4E-binding protein (4E-BP), levels. Equol also increased metastatic cancer cell viability. Equol specifically increased the protein expression of IRES containing cell survival and proliferation-promoting molecules and up-regulated gene and protein expression of the transcription factor c-Myc. Moreover, equol increased the polysomal association of mRNAs for p 120 catenin and eIF4G. The elevated eIF4G in response to equol was not associated with eIF4E or 4E-binding protein in 5' cap co-capture assays or co-immunoprecipitations. In dual luciferase assays, IRES-dependent protein synthesis was increased by equol. Therefore, up-regulation of eIF4G by equol may result in increased translation of pro-cancer mRNAs with IRESs and, thus, promote cancer malignancy.

Topics: Animals; Breast Neoplasms; Cell Line, Tumor; Dietary Supplements; Equol; Eukaryotic Initiation Factor-4E; Eukaryotic Initiation Factor-4G; Female; Gene Expression Regulation, Neoplastic; Glycine max; Humans; Isoflavones; Mice; Mice, Nude; Neoplasm Transplantation; Phytoestrogens; Protein Biosynthesis; Proto-Oncogene Proteins c-myc; RNA, Messenger; RNA, Neoplasm; Transplantation, Heterologous; Up-Regulation

Sex hormone replacement therapy provides several advantages in the quality of life for climacteric women. However, estrogen-induced cell proliferation in the uterus and mammary gland increases the risk of cancer development in these organs. The lower incidence of mammary cancer in Asian women as compared with Western women has been attributed to high intake of soy isoflavones, including genistein. We have previously shown that genistein induces an estradiol-like hypertrophy of uterine cells, but does not induce cell proliferation, uterine eosinophilia, or endometrial edema. It also inhibits estradiol-induced mitosis in uterine cells and hormone-induced uterine eosinophilia and endometrial edema. Nevertheless, genistein stimulates growth of human breast cancer cells in culture; therefore, it is not an ideal estrogen for use in hormone replacement therapy (HRD). The present study investigated the effect of another soy isoflavone, daidzein (subcutaneous, 0.066 mg/kg body weight), in the same animal model, and its effect on responses induced by subsequent treatment (1 h later) with estradiol-17β (E(2); subcutaneous, 0.33 mg/kg body weight). In addition, we investigated the effects of daidzein (1 μg/mL) or E(2) on the growth of human breast cancer cells in culture. Results indicate that daidzein stimulates growth of breast cancer cells and potentiates estrogen-induced cell proliferation in the uterus. We suggest caution for the use of daidzein or formulas containing this compound in HRD. Future research strategies should be addressed in the search for new phytoestrogens that selectively inhibit cell proliferation in the uterus and breast.

Topics: Animals; Antineoplastic Agents, Hormonal; Breast Neoplasms; Cell Line, Tumor; Cell Proliferation; Drug Interactions; Estradiol; Estrogens; Female; Glycine max; Humans; Isoflavones; MCF-7 Cells; Phytoestrogens; Plant Extracts; Rats; Rats, Sprague-Dawley; Uterus

The current investigations were undertaken to study the mechanism of the adverse effect of phytoestrogens on the function of bovine granulosa (follicles >1< cm in diameter) and luteal cells from day 1-5, 6-10, 11-15, 16-19 of the oestrous cycle. The cells were incubated with genistein, daidzein or coumestrol (each at the dose of 1 × 10(-6) m). The viability and secretion of estradiol (E2), progesterone (P4) and oxytocin (OT) were measured after 72 h of incubation. Moreover, the expression of mRNA for neurophysin-I/OT (NP-I/OT; precursor of OT) and peptidyl-glycine-α-amidating monooxygenase (PGA, an enzyme responsible for post-translational OT synthesis) was determined after 8 h of treatment. None of the phytoestrogens used affected the viability of cells except for coumestrol. The increased secretion of E2 and P4 was only obtained by coumestrol (p<0.05) from granulosa cells from follicles <1cm in diameter and decreased from luteal cells on days 11-15 of the oestrous cycle, respectively. All three phytoestrogens stimulated (p<0.05) OT secretion from granulosa and luteal cells in all stages of the oestrous cycle and the expression of NP-I/OT mRNA in the both types of cells. The expression of mRNA for PGA was stimulated (p<0.05) by daidzein and coumestrol in granulosa cells, and by genistein and coumestrol in luteal cells. In conclusion, our results demonstrate that these phytoestrogens can impair the ovary function in cattle by adversely affecting the synthesis of OT in follicles and in corpus luteum. However, their influence on the ovarian steroids secretion was less evident.

Topics: Animals; Cattle; Cells, Cultured; Coumestrol; Female; Genistein; Granulosa Cells; Isoflavones; Luteal Cells; Mixed Function Oxygenases; Multienzyme Complexes; Neurophysins; Ovary; Oxytocin; Phytoestrogens; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger

We demonstrated previously that phytoestrogens and vitamin D analogs like estradiol-17β (E2) modulate bone morphology in rat female model.. We now analyze the effects of phytoestrogens, E2, selective E2 re ceptor modulators, and the less-calcemic analogs of vitamin D: JKF1624F2-2 (JKF) or QW1624F2-2 (QW) on fat content in bone marrow (BM) from long bones in ovariectomized female rats (OVX).. OVX rats were injected with treatments known to affect bone formation, 5 days per week for 2.5 month for analysis of fat content in BM.. In OVX young adults there is a decreased bone formation and a 10-fold increase in fat cells content in BM. Treatment with E2, raloxifene (Ral) or DT56a resulted in almost completely abolishment of fat cells content. Daidzein (D) decreased fat cells content by 80%, genistein (G) or biochainin A (BA) did not change fat cells content and carboxy BA (cBA) had a small but significant effect. JKF or QW did not affect fat cells content, whereas combined treatment of JKF or QW with E2 resulted in complete abolishment of fat cells content. These changes in fat cells content are inversely correlated with changes in bone formation.. Our results demonstrate that adipogenesis induced by OVX is a reversible process which can be corrected by hormonal treatments. The awareness of a relationship between fat and bone at the marrow level might provide a better understanding of the pathophysiology of bone loss as well as a novel approach to diagnosis and treatment of postmenopausal osteoporosis.

Topics: Adipocytes; Animals; Bone Marrow Cells; Calcitriol; Estradiol; Estrogen Receptor Modulators; Estrogens; Female; Genistein; Isoflavones; Ovariectomy; Phytoestrogens; Raloxifene Hydrochloride; Rats; Rats, Wistar

Phytooestrogens are known to cause anti-cancer effects on mamma carcinoma cells. In this study, the effects of the lignan secoisolariciresinol and the isoflavone glycosides and aglycones genistein, genistin, daidzein and daidzin were tested on MCF-7 and BT20 cells in vitro.. First, the cellular expression of hormone receptors was examined by immunohistochemical procedures. The effects of the phytooestrogens on the cells were detected by using three different assays measuring cell letality, viability and proliferation. The phytooestrogens were tested in concentrations of 1, 5, 10 and 50 μg/mL, respectively. 17β-oestradiol and tamoxifen were used as controls, respectively, in the same concentrations as the phytooestrogens.. The immunohistochemistry showed evidence of oestrogen- and progesterone receptors at the MCF-7 cell line, whereas no expression could be seen at the BT20 cells. Among the phytooestrogens, genistein and secoisolariciresinol showed various anti-cancerogenic effects on both cell lines, respectively, but only in the highest concentration. Regarding the controls, tamoxifen showed a stronger antivital and anti-proliferative effect on BT20 than on MCF-7. Oestradiol caused sporadic anti-cancer effects on both cell lines, respectively, at its highest concentration.. Genistein and Secoisolariciresinol have anti-cancer properties on MCF-7 and BT20 in vitro. There are differences in the effects of isoflavones depending on the glycolysation status. The role of the oestrogen receptors in the mechanisms of action of both the phytooestrogens and controls is of less importance. Further investigations have to be carried out, especially concerning the mechanisms of action. Phytooestrogens may be potential substances in the therapy of mamma carcinomas.

Topics: Breast Neoplasms; Butylene Glycols; Carcinoma; Cell Line, Tumor; Cell Proliferation; Cell Survival; Female; Genistein; Humans; Immunohistochemistry; Isoflavones; L-Lactate Dehydrogenase; Lignans; Phytoestrogens; Receptors, Estrogen; Receptors, Progesterone

In light of concerns about hormonally active agents, it is important to assess human exposure to such compounds, especially in children as a susceptible subgroup. Estrogenic plant constituents are present in the human diet in varying levels, in particular the isoflavones daidzein (DAI) and genistein (GEN). We aimed to examine age-dependent and secular trends in phytoestrogen exposures and to investigate equol (EQ) excretion of German children using biomarker analysis in 24-h urine samples from a longitudinally designed study.. The concentrations of DAI, its metabolite EQ and GEN were determined by GC-MS analysis in 24-h urines (510 samples) collected between 1985 and 2000 in 90 (47 boys) German children (6-18 years old), who are participants in the Dortmund Nutritional and Anthropometric Longitudinally Designed study. The results from the urinary biomarker analysis indicate isoflavone exposures at quite variable levels in German children: Analyte concentrations in over 500 urine samples cover the range reported previously in adults on typical German diet and with soy intake. EQ, the DAI metabolite produced by the gastrointestinal microflora, was detected in a high fraction of all samples, with 28/90 children (31%) excreting EQ in all their urines, and 62/90 children (68%) in at least one sample. Interestingly, when multiple urines obtained from individuals at different ages (6-18 years) were analyzed, EQ formation did not appear to be a constant trait over time. When stratified by sex, DAI, EQ and GEN concentrations (ng/mL) in urines and excretion rates (μg/day) were similar in boys and girls. Total isoflavone excretion rates (μg/day) increased during childhood (6-12 years) (p=0.02) and were constant during adolescence (13-18 years) (p=0.6). No clear trend for changes in dietary isoflavone exposure over the total study period was seen (p=0.7).. In conclusion, biomarkers in urine of German children and adolescents indicate a frequent, but widely variable dietary isoflavone intake and suggest no secular increase (1985-2000) in the exposure to isoflavone phytoestrogens among German children and adolescents.

Topics: Adolescent; Biomarkers; Child; Diet; Equol; Female; Gas Chromatography-Mass Spectrometry; Genistein; Germany; Glycine max; Humans; Isoflavones; Linear Models; Longitudinal Studies; Male; Phytoestrogens; White People

The effects of genistein, a protein tyrosine kinase (PTK) inhibitor, on voltage-dependent K(+) (Kv) 4.3 channel were examined using the whole cell patch-clamp techniques. Genistein inhibited Kv4.3 in a reversible, concentration-dependent manner with an IC(50) of 124.78 μM. Other PTK inhibitors (tyrphostin 23, tyrphostin 25, lavendustin A) had no effect on genistein-induced inhibition of Kv4.3. Orthovanadate, an inhibitor of protein phosphatases, did not reverse the inhibition of Kv4.3 by genistein. We also tested the effects of two inactive structural analogs: genistin and daidzein. Whereas Kv4.3 was unaffected by genistin, daidzein inhibited Kv4.3, albeit with a lower potency. Genistein did not affect the activation and inactivation kinetics of Kv4.3. Genistein-induced inhibition of Kv4.3 was voltage dependent with a steep increase over the channel opening voltage range. In the full-activation voltage range positive to +20 mV, no voltage-dependent inhibition was found. Genistein had no significant effect on steady-state activation, but shifted the voltage dependence of the steady-state inactivation of Kv4.3 in the hyperpolarizing direction in a concentration-dependent manner. The K(i) for the interaction between genistein and the inactivated state of Kv4.3, which was estimated from the concentration-dependent shift in the steady-state inactivation curve, was 1.17 μM. Under control conditions, closed-state inactivation was fitted to a single exponential function, and genistein accelerated closed-state inactivation. Genistein induced a weak use-dependent inhibition. These results suggest that genistein directly inhibits Kv4.3 by interacting with the closed-inactivated state of Kv4.3 channels. This effect is not mediated via inhibition of the PTK activity, because other types of PTK inhibitors could not prevent the inhibitory action of genistein.

Topics: Animals; CHO Cells; Cricetinae; Cricetulus; Dose-Response Relationship, Drug; Genistein; Ion Channel Gating; Isoflavones; Kinetics; Phosphorylation; Phytoestrogens; Potassium Channel Blockers; Protein Kinase Inhibitors; Shal Potassium Channels; Tyrosine

Globally, non-alcoholic fatty liver disease (NAFLD) continues to rise and isoflavones exert antisteatotic effects by the regulation of hepatic lipogenesis/insulin resistance or adiposity/a variety of adipocytokines are related to hepatic steatosis. However, there is very little information regarding the potential effects of daidzein, the secondary abundant isoflavone, on NAFLD. Here, we have assessed the hepatic global transcription profiles, adipocytokines and adiposity in mice with high fat-induced NAFLD and their alteration by daidzein supplementation.. C57BL/6J mice were fed with normal fat (16% fat of total energy), high fat (HF; 36% fat of total energy) and HF supplemented with daidzein (0.1, 0.5, 1 and 2 g per kg diet) for 12 weeks.. Daidzein supplementation (≥ 0.5 g per kg diet) reduced hepatic lipid concentrations and alleviated hepatic steatosis. The hepatic microarray showed that daidzein supplementation (1 g per kg diet) downregulated carbohydrate responsive element binding protein, a determinant of de novo lipogenesis, its upstream gene liver X receptor β and its target genes encoding for lipogenic enzymes, thereby preventing hepatic steatosis and insulin resistance. These results were confirmed by lower insulin and blood glucose levels as well as homeostasis model assessment insulin resistance scores. In addition, daidzein supplementation inhibited adiposity by the upregulation of genes involved in fatty acid β-oxidation and the antiadipogeneis, and moreover augmented antisteatohepatitic leptin and adiponectin mRNA levels, whereas it reduced the mRNA or concentration of steatotic tumor necrosis factor α and ghrelin.. These findings show that daidzein might alleviate NAFLD through the direct regulation of hepatic de novo lipogenesis and insulin signaling, and the indirect control of adiposity and adipocytokines by the alteration of adipocyte metabolism.

Topics: Adipocytes; Adipokines; Adipose Tissue; Animals; Body Weight; Diet; Fatty Liver; Gene Expression Profiling; Insulin; Insulin Resistance; Isoflavones; Lipogenesis; Mice; Mice, Inbred C57BL; Non-alcoholic Fatty Liver Disease; Phytoestrogens; Reverse Transcriptase Polymerase Chain Reaction

This study aimed to determine the influence of high-dose soy isoflavones (daidzein and genistein) administered from prenatal life to sexual maturity on testosterone and estradiol levels, testicular and epididymal morphology, the number of epididymal spermatozoa, and mineral metabolism in rats.. Pregnant Wistar rats received orally soy isoflavones, daidzein, and genistein at a dose of 200 mg/kg of body weight per day. After separating sucklings from their mothers, male rats received the same dose of isoflavones until reaching the age of sexual maturity, i.e., for 3 mo.. In the isoflavone-treated group, statistically significant decreased concentrations of zinc (determined using atomic absorption spectrophotometry) in blood serum and increased concentrations in bone were observed. The isoflavones induced changes in the morphology of the seminiferous epithelium of rat testes. However, there were no significant changes in the number of spermatozoa in the epididymis. The levels of estradiol in serum and cauda epididymis homogenates of rats receiving phytoestrogens were significantly higher than in the control group. No differences were observed in testosterone concentrations in the serum of treated and control rats. The testosterone levels in the homogenates of the treated rat testes were significantly lower than in the control group.. The relatively mild effects of phytoestrogen administration on the morphology of testes and epididymides and the number of epididymal spermatozoa were observed despite the high dose used. The exposure of rats to genistein and daidzein during intrauterine life until sexual maturity influenced the mineral metabolism of the organism by significant decreases of Zn concentration in serum and increased Zn concentration in bones.

Topics: Animals; Bone and Bones; Epididymis; Estradiol; Female; Genistein; Genitalia, Male; Glycine max; Gonadal Steroid Hormones; Isoflavones; Male; Phytoestrogens; Plant Extracts; Pregnancy; Prenatal Exposure Delayed Effects; Rats; Rats, Wistar; Seminiferous Epithelium; Testis; Testosterone; Zinc

A series of didzein derivatives were synthesized and assessed for stimulation of osteoblast differentiation using primary cultures of rat calvarial osteoblasts. Data suggested that three synthetic analogs, 1c, 3a and 3c were several folds more potent than daidzein in stimulating differentiation and mineralization of osteoblasts. Further, these three compounds did not show any estrogen agonistic activity, however had mild estrogen antagonistic effect. Out of the three compounds, 3c was found to maximally increase the mineralization of bone marrow osteoprogenitor cells. Compound 3c also robustly increased the mRNA levels of osteogenic genes including bone morphogenetic protein-2 and osteocalcin in osteoblasts. Unlike daidzein, 3c did not inhibit osteoclastogenesis. Collectively, we demonstrate osteogenic activity of daidzein analogs at significantly lower concentrations than daidzein.

Topics: Animals; Bone Marrow Cells; Bone Morphogenetic Protein 2; Calcification, Physiologic; Cell Differentiation; Cells, Cultured; Gene Expression Regulation; Isoflavones; Osteoblasts; Osteocalcin; Phytoestrogens; Rats; RNA, Messenger; Skull

We developed and validated three different sample preparation and extraction methods followed by HPLC-MS/MS (negative electrospray ionization) analysis for the quantification of estrogenic isoflavones (formononetin, daidzein, equol, biochanin A, and genistein) and coumestrol in red clover, soil, and manure. Plant and manure samples were solid-liquid extracted, whereas soil was extracted with accelerated solvent extraction. Absolute recoveries were between 80 and 93%, 20 and 30%, and 14 and 91% for plant, soil, and manure samples, respectively. Relative recoveries ranged from 75 to 105% for all matrices, indicating that isotope-labeled internal standards (¹³C₃-formononetin, ¹³C₃-daidzein, ¹³C₃-equol, ¹³C₃-biochanin A, and ¹³C₃-genistein) were capable to compensate for losses during analysis. The limits of detection in red clover, soil, and manure were 3-9 μg/g(dryweight(dw)), 0.6-8.2 ng/g(dw), and 34.2 ng/g(dw) to 17.0 μg/g(dw), respectively. Formononetin was the most dominant compound in red clover plants (up to 12.5 mg/g(dw)) and soil (up to 3.3 μg/g(dw)), whereas equol prevailed in manure (up to 387 μg/g(dw)).

Topics: Chromatography, High Pressure Liquid; Coumestrol; Equol; Genistein; Isoflavones; Manure; Phytoestrogens; Soil; Tandem Mass Spectrometry; Trifolium

To evaluate the effect of different cyclodextrins (β-cyclodextrin [β-CD], methyl-β-cyclodextrin [Mβ-CD], or hydroxypropyl-β-cyclodextrin [HPβ-CD]) and/or hydrophilic polymers (carboxymethylcellulose, hydroxypropylmethylcellulose [HPMC], polyethyleneglycol, or polyvinylpyrrolidone [PVP]) on daidzein solubility in water.. The corresponding associations were characterized in aqueous media using phase-solubility studies. The morphology of daidzein/cyclodextrin freeze-dried complexes was characterized using scanning electron microscopy, and their spatial configuration was proposed by means of nuclear magnetic resonance spectroscopy.. In the presence of 6 mM of cyclodextrins, the solubility of daidzein in water was significantly enhanced: 5.7-fold (β-CD), 7.2-fold (Mβ-CD), and 9.4-fold (HPβ-CD). The analysis of the three solid complexes proved that the formation of inclusion complexes occurred through the insertion of the B and C rings of daidzein molecule into the cyclodextrins cavity. The association of daidzein/cyclodextrin complexes to the hydrophilic polymers HPMC or PVP (1%, w/w) was able to improve the solubility of daidzein even further.. The highest solubilizing effect was obtained for daidzein/HPβ-CD/PVP ternary system (12.7-fold).

Topics: Chemistry, Pharmaceutical; Cyclodextrins; Hydrophobic and Hydrophilic Interactions; Isoflavones; Magnetic Resonance Spectroscopy; Phytoestrogens; Polymers; Solubility; Water

TNF-α-induced osteoclastogenesis is central to post-menopausal and inflammatory bone loss, however, the effect of phytoestrogens on TNF-α-induced bone resorption has not been studied. The phytoestrogens genistein, daidzein, and coumestrol directly suppressed TNF-α-induced osteoclastogenesis and bone resorption. TRAP positive osteoclast formation and resorption area were significantly reduced by genistein (10(-7)  M), daidzein (10(-5)  M), and coumestrol (10(-7)  M), which was prevented by the estrogen antagonist ICI 182,780. TRAP expression in mature TNF-α-induced osteoclasts was also significantly reduced by these phytoestrogen concentrations. In addition, in the presence of ICI 182,780 genistein and coumestrol (10(-5) -10(-6)  M) augmented TNF-α-induced osteoclast formation and resorption. However, this effect was not observed in the absence of estrogen antagonist indicating that genistein's and coumestrol's ER-dependent anti-osteoclastic action normally negates this pro-osteoclastic effect. To determine the mechanism mediating the anti-osteoclastic action we examined the effect of genistein, coumestrol, and daidzein on caspase 3/7 activity, cell viability and expression of key genes regulating osteoclast differentiation and fusion. While anti-osteoclastic phytoestrogen concentrations had no effect on caspase 3/7 activity or cell viability they did significantly reduce TNF-α-induced c-fos and NFATc1 expression in an ER dependent manner and also inhibited NFATc1 nuclear translocation. Significant decreases in NFκB and DC-STAMP levels were also noted. Interestingly, constitutive c-fos expression prevented the anti-osteoclastic action of phytoestrogens on differentiation, resorption and NFATc1. This suggests that phytoestrogens suppress TNF-α-induced osteoclastogenesis via inhibition of c-fos-dependent NFATc1 expression. Our data provides further evidence that phytoestrogens have a potential role in the treatment of post-menopausal and inflammatory bone loss directly inhibiting TNF-α-induced resorption.

Topics: Animals; Bone Resorption; Cell Differentiation; Cell Line; Cell Survival; Coumestrol; Genistein; Isoflavones; Mice; NFATC Transcription Factors; Phytoestrogens; Polymerase Chain Reaction; Proto-Oncogene Proteins c-fos; Tumor Necrosis Factor-alpha

Equol and O-desmethylangolensin (ODMA) are products of gut bacterial metabolism of daidzein, a phytochemical found predominantly in soy. Dietary sources of equol from animal products have been identified, which has raised the question of the relative contributions of daidzein intake and gut metabolism to equol and of equol intake from animal products in low-soy-consuming populations.. The objective was to evaluate the contribution of dietary food groups to urinary isoflavone and daidzein metabolite concentrations in a representative sample of US adults.. A cross-sectional analysis of dietary and urinary isoflavonoid data from 3115 individuals in the 2001-2002 and 2003-2004 data cycles of the National Nutrition and Health Examination Survey (NHANES) was conducted.. Daidzein intake and consumption frequency of grain products and legumes, nuts, and seeds were significant correlates of daidzein, genistein, and ODMA concentrations; and soy legumes were a stronger correlate than were nonsoy legumes. Milk and milk product consumption and daidzein intake, but not legumes, were significant correlates of urinary equol concentrations; milk products were more strongly correlated (P for trend < 0.001) than was daidzein intake (P = 0.011).. These results suggest that dietary daidzein and legumes may contribute to urinary daidzein, genistein, and ODMA concentrations in this low-soy-consuming population. These results also suggest that equol concentrations in low-soy-consuming populations may reflect equol intakes from mammalian milk sources and may not reflect the endogenous production of equol from the microbial metabolism of daidzein-an observation not yet documented in the US population. These results support the careful design and interpretation of urinary isoflavonoid excretion studies, particularly bacterial metabolites, in low-soy-consuming populations.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Cross-Sectional Studies; Dairy Products; Diet; Equol; Female; Genistein; Humans; Isoflavones; Male; Middle Aged; Nutrition Surveys; Phytoestrogens; Soy Foods; United States; Young Adult

Phytoestrogens are compounds that have moderate estrogenic or anti-estrogenic activity toward mammals. Although genistein and daidzein, the main phytoestrogens of soybean, have been the subject of thousands of studies that address their benefit to human health, relatively little is known about their benefits to plants that produce them. It has been suggested that genistein and daidzein protect plants against arthropod herbivores, but direct tests of this hypothesis are rare. In this study, we evaluated the effect of genistein and daidzein on the survivorship, growth, and fecundity of the gypsy moth, a generalist insect herbivore that does not encounter phytoestrogens in its normal diet. We compared survivorship, egg-to-pupa growth rate, and 4th instar performance of gypsy moth caterpillars on artificial diets containing no phytoestrogen, genistein, daidzein, or a combination of genistein and daidzein. Our results indicate that genistein and daidzein do not decrease survivorship, growth, or fecundity of this insect herbivore. Therefore, it seems unlikely that the primary function of these compounds in aboveground plant tissues is anti-herbivore defense.

Topics: Animals; Genistein; Host-Parasite Interactions; Isoflavones; Moths; Phytoestrogens; Plants

Daidzein is a potential natural alternative to estradiol during therapy of some malignancies in men. Besides weak inhibition of tyrosine kinase activity, daidzein has a sizeable inhibitory effect on calcium channels. The aim of this study was to examine the effects of daidzein on the immunohistomorphometric features of pituitary adrenocorticotropes (ACTH cells) and circulating levels of ACTH and corticosterone, in comparison with estradiol, in an animal model of the andropause. Sixteen-month-old Wistar rats were divided into sham operated (SO), orchidectomized (Orx), estradiol treated orchidectomized (Orx+E) and daidzein treated orchidectomized (Orx+D) groups. Estradiol (0.625 mg/kg/day) and daidzein (30 mg/kg/day) were administered subcutaneously for three weeks, while the SO and Orx groups received the vehicle alone. ACTH cells were identified by the peroxidase-antiperoxidase (PAP) immunohistochemical procedure. Peripheral circulating concentrations of ACTH and corticosterone were measured by immunoassay. Orchidectomy reduced (p<0.05) the cell volume and volume density of adrenocorticotropes by 11% and 16%, respectively, in comparison to SO rats. In Orx+E rats, the volume density of ACTH cells decreased (p<0.05) by 25%, but the circulating level of ACTH increased (p<0.05) by 29%, compared to Orx rats. Daidzein treatment significantly decreased (p<0.05): volume density of ACTH cells, circulating ACTH and corticosterone by 24%, 48% and 33%, respectively, compared to the Orx group. In conclusion, this study revealed that daidzein negatively modulated the immunohistomorphometric features of ACTH cells and, unlike estradiol, decreased ACTH and corticosterone secretion, in an animal model of the andropause.

Topics: Andropause; Animals; Corticosterone; Corticotrophs; Disease Models, Animal; Estradiol; Estrogens; Immunohistochemistry; Isoflavones; Male; Orchiectomy; Phytoestrogens; Rats; Rats, Wistar

The present study compared the changes in isoflavone (daidzein and genistein) and their metabolite (equol and para-ethyl-phenol) concentrations in the blood plasma of cows with induced mastitis and metritis after feeding with soy bean. Sixteen cows were divided into four groups: control for mastitis group, cows with induced mastitis group, control for metritis group, and cows with induced metritis group. All cows were fed a single dose of 2.5 kg of soy bean and then blood samples were taken from the jugular vein for 8 h at predetermined intervals. The concentrations of soy bean-derived isoflavones and their active metabolites were measured in the blood plasma on HPLC system. β-Glucuronidase activity in the blood plasma of cows was measured by fluorometric method. In the blood plasma of cows with induced mastitis and metritis, we found considerably higher concentrations and time-dependent increase in isoflavone metabolites (equol and para-ethyl-phenol) with reference to cyclic cows (P < 0.05). Moreover, we noticed significant decrease of genistein in the blood plasma of the cows with induced metritis compared with control cows (P < 0.05). In addition, in the blood plasma of the cows with induced metritis, we found an increase in β-glucuronidase activity compared with control cows (P < 0.05). In conclusion, health status of the females influenced the concentrations of isoflavone metabolites in the blood plasma of the cows. Experimentally induced mastitis and metritis increased isoflavone absorption, biotransformation and metabolism. Therefore, we suggest that cows with induced mastitis and metritis are more exposed to active isoflavone metabolite actions than healthy cows.

Topics: Animals; Biotransformation; Cattle; Cattle Diseases; Equol; Female; Genistein; Glucuronidase; Glycine max; Isoflavones; Mastitis, Bovine; Metrial Gland; Phytoestrogens; Uterine Diseases

Isoflavones are ubiquitous compounds in foods and in the environment in general. Daidzein and genistein, the best known of isoflavones, are structurally similar to 17β-estradiol and known to exert estrogenic effects. They also evidence a broad variety of biological properties, including antioxidant, anti-carcinogenic, anti-atherogenic and anti-osteoporotic activities. Previously, daidzein was reported to increase the phagocytic activity of peritoneal macrophages and splenocyte proliferation, and to inhibit nitric oxide (NO) production in macrophages. However, its potential impacts on immune response in dendritic cells (DCs), antigen-presenting cells that link innate and adaptive immunity, have yet to be clearly elucidated. In this study, we evaluated the effects of isoflavones on the maturation and activation of DCs. Isoflavones (formononetin, daidzein, equol, biochanin A, genistein) were found to differentially affect the expression of CD86, a costimulatory molecule, on lipopolysaccharide (LPS)-stimulated DCs. In particular, daidzein significantly and dose-dependently inhibited the expression levels of maturation-associated cell surface markers including CD40, costimulatory molecules (CD80, CD86), and major histocompatibility complex class II (I-A(b)) molecule on LPS-stimulated DCs. Daidzein also suppressed pro-inflammatory cytokine production such as IL-12p40, IL-6 and TNF-α, whereas it didn't affect IL-10 and IL-1β expression. Furthermore, daidzein enhanced endocytosis and inhibited the allo-stimulatory ability of LPS-stimulated DCs on T cells, indicating that daidzein treatment can inhibit the functional maturation of DCs. These results demonstrate that daidzein may exhibit immunosuppressive activity by inhibiting the maturation and activation of DCs.

Topics: Animals; Cell Enlargement; Dendritic Cells; Female; Growth Inhibitors; Immunosuppressive Agents; Isoflavones; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Phytoestrogens

DNA hypermethylation is an epigenetic mechanism which induces silencing of tumor-suppressor genes in prostate cancer. Many studies have reported that specific components of food plants like soy phytoestrogens may have protective effects against prostate carcinogenesis or progression. Genistein and daidzein, the major phytoestrogens, have been reported to have the ability to reverse DNA hypermethylation in cancer cell lines. The aim of this study was to investigate the potential demethylating effects of these two soy compounds on BRCA1, GSTP1, EPHB2 and BRCA2 promoter genes.. Prostate cell lines DU-145 and PC-3 were treated with genistein 40 µM, daidzein 110 µM, budesonide (methylating agent) 2 µM and 5-azacytidine (demethylating agent) 2 µM. In these two human prostate cancer cell lines we performed methylation quantification by using Methyl Profiler DNA methylation analysis. This technique is based on a methylation-specific digestion followed by quantitative PCR. We analyzed the corresponding protein expression by western blotting.. Soy phytoestrogens induced a demethylation of all promoter regions studied except for BRCA2, which is not methylated in control cell lines. An increase in their protein expression was also demonstrated by western blot analysis and corroborated the potential demethylating effect of soy phytoestrogens.. This study showed that the soy phytoestrogens, genistein and daidzein, induce a decrease of methylation of BRCA1, GSTP1 and EPHB2 promoters. Therefore, soy phytoestrogens may have a protective effect on prostate cancer. However, more studies are needed in order to understand the mechanism by which genistein and daidzein have an inhibiting action on DNA methylation.

Topics: Blotting, Western; BRCA1 Protein; BRCA2 Protein; Cell Line, Tumor; DNA Methylation; Flow Cytometry; Genes, Tumor Suppressor; Genistein; Glutathione S-Transferase pi; Glycine max; Humans; Isoflavones; Male; Phytoestrogens; Promoter Regions, Genetic; Prostatic Neoplasms; Receptor, EphB2

Daidzein-loaded chitosan microspheres were prepared by emulsification/chemical-crosslinking technique. The dialysis bag method determined the release of daidzein from the microspheres. It demonstrated that the accumulative release curve in vitro was fit for the zero-order release equation and had good correlation with the absorptive fraction in vivo, suggesting the dialysis bag method evaluated the release of the microspheres well. The release of chitosan determined by the ninhydrin assay in vitro was very slow, less than 3 percent at 35 day. The pathological section by hematoxylin-eosin staining found the good biocompatibility of the prepared microspheres in the injective site. Combining the degradation photos by scanning electron microscopy with the plasma concentration-time data, it was speculated that the drug on the surface of the microspheres firstly released, then the major of drug near the surface and the inner of the microspheres released by diffusion through the shallow cavities and crack, lastly the drug released rapidly and completely being companied with the beginning of polymer degradation.

Topics: Absorbable Implants; Animals; Biocompatible Materials; Biological Availability; Chitosan; Delayed-Action Preparations; Drug Compounding; Female; In Vitro Techniques; Infusion Pumps, Implantable; Injections, Intramuscular; Isoflavones; Mice; Mice, Inbred ICR; Microspheres; Phytoestrogens; Rats; Time Factors

The status of glycoconjugates (protein bound hexose, hexosamine, sialic acid and fucose) in plasma or serum serve as potential biomarkers for assessing tumor progression and therapeutic interventions. Aim of the present study was to investigate the protective effect of two major soy isoflavones, genistein and daidzein, in combination on the status of glycoconjugates in plasma, erythrocyte membrane and mammary tissues during 7,12-dimethylbenz[a]anthracene (DMBA) induced mammary carcinogenesis in female Sprague-Dawley rats. A single subcutaneous injection of DMBA (25 mg rat(-1)) in the mammary gland developed mammary carcinoma in female Sprague-Dawley rats. Elevated levels of plasma and mammary tissue glycoconjugates accompanied by reduction in erythrocyte membrane glycoconjugates were observed in rats bearing mammary tumors. Oral administration of genistein + daidzein (20 mg + 20 mg kg(-1) bw/day) to DMBA treated rats significantly (p< 0.05) brought back the status of glycoconjugates to near normal range. The present study thus demonstrated that genistein and daidzein in combination protected the structural integrity of the cell surface and membranes during DMBA-induced mammary carcinogenesis.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Biomarkers, Tumor; Cell Membrane; Drug Therapy, Combination; Female; Genistein; Glycoconjugates; Isoflavones; Mammary Glands, Animal; Mammary Neoplasms, Experimental; Phytoestrogens; Rats; Rats, Sprague-Dawley

Phytoestrogens have attracted attention as being safer alternatives to hormone replacement therapy (HRT) and as chemopreventive reagents for breast cancer because dietary soy isoflavone intake has been correlated with reduction in risk. To identify safe and effective phytoestrogen candidates for HRT and breast cancer prevention, we investigated the effects of daidzein, genistein, coumestrol, resveratrol and glycitein on cell growth, cell cycle, cyclin D1 expression, apoptosis, Bcl-2/Bax expression ratio and p53-dependent or NF-kappaB-dependent transcriptional activity in MCF-7 breast cancer cells. Phytoestrogens, except for glycitein, significantly enhanced estrogen-response-element-dependent transcriptional activity up to a level similar to that of 17beta-estradiol (E(2)). E(2) increased cell growth significantly, coumestrol increased cell growth moderately, and resveratrol and glycitein reduced cell growth. Phytoestrogens, except for glycitein, stimulated the promotion of cells to G(1)/S transition in cell cycle analysis, similar to E(2). This stimulation was accompanied by transient up-regulation of cyclin D1. While genistein, resveratrol and glycitein all increased apoptosis and reduced the Bcl-2/Bax ratio, resveratrol reduced this ratio more than either genistein or glycitein. Moreover, resveratrol significantly enhanced p53-dependent transcriptional activity, but slightly reduced NF-kappaB-dependent transcriptional activity. On knockdown analysis, genistein, resveratrol and glycitein all reduced the Bcl-2/Bax ratio in the presence of apoptosis-inducing stimuli, and estrogen receptor (ER) alpha silencing had no effect on these reductions. In contrast, in the absence of apoptosis-inducing stimuli, only resveratrol reduced the ratio, and ERalpha silencing abolished this reduction. Thus, resveratrol might be the most promising candidate for HRT and chemoprevention of breast cancer due to its estrogenic activity and high antitumor activity.

Topics: Apoptosis; bcl-2-Associated X Protein; Breast Neoplasms; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Coumestrol; Estradiol; Estrogen Receptor alpha; Female; Genistein; Humans; Isoflavones; Phytoestrogens; Proto-Oncogene Proteins c-bcl-2; Resveratrol; Stilbenes

In order to study the anticancer effects and cellular apoptosis pathways induced by daidzein.. We used the human MCF-7 breast cancer cell line as a model and examined the apoptosis by Hoechst-propidium iodide staining fluorescence imaging and flow cytometry.. Our data indicated that daidzein induces antiproliferative effects in a concentration- and time-dependent manner. We demonstrated that daidzein-induced apoptosis in MCF-7 cells was initiated by the generation of reactive oxygen species (ROS). Furthermore, we showed that this daidzein-induced ROS generation was accompanied by disruption of mitochondrial transmembrane potential, down-regulation of bcl-2, and up-regulation of bax, which led to the release of cytochrome C from the mitochondria into the cytosol, which, in turn, resulted in the activation of caspase-9 and caspase-7, and ultimately in cell death. The induction of the mitochondrial caspase-dependent pathway was confirmed by pretreatment with pan-caspase inhibitor z-VAD-fmk and antioxidant N-acetyl-L-cysteine.. Accordingly, daidzein could induce breast cancer cell apoptosis through the mitochondrial caspase-dependent cell death pathway.

Topics: Antineoplastic Agents, Phytogenic; Apoptosis; Breast Neoplasms; Cell Line, Tumor; Cell Proliferation; Dose-Response Relationship, Drug; Drug Evaluation, Preclinical; Female; Humans; Isoflavones; Membrane Potential, Mitochondrial; Mitochondria; Phytoestrogens; Reactive Oxygen Species; Signal Transduction; Time Factors; Up-Regulation

Two in vitro systems were employed to delineate the estrogenic activity of daidzein (Da), alone or in combination with high or low concentrations of estrogen in two cell types possessing different estrogen-receptor (ER) isoforms, ERalpha and/or ERbeta: (1) vitellogenin II (VTG), the egg yolk precursor protein and the endpoint biomarker for estrogenicity, in chicken primary hepatocytes, and (2) CHO-K1 cells transiently co-transfected with ERalpha or ERbeta and estrogen-response elements (ERE) linked to a luciferase reporter gene. Da (100 microM) alone induced VTG mRNA expression in chicken hepatocytes, albeit with much less potency compared to estradiol (E(2)). Da exhibited different effects in the presence of 1 microM and 10 microM E(2). At a concentration of 100 microM, Da enhanced 1 microM E(2)-induced VTG transcription by 2.4-fold, but significantly inhibited 10 microM E(2)-induced VTG mRNA expression in a dose-dependent fashion from 1 to 100 microM. Tamoxifen completely blocked the estrogenic effect of daidzein, alone or in combination with 1 microM of E(2), but did not influence its anti-estrogenic effect on 10 microM E(2)-induced VTG mRNA expression. Furthermore, neither E(2) nor daidzein, alone or in combination, affected ERalpha mRNA expression, yet all the treatments significantly up-regulated ERbeta mRNA expression in chicken hepatocytes. E(2) effectively triggered estrogen-response elements (ERE)-driven reporter gene transactivation in CHO-K1 cells expressing ERalpha or ERbeta and showed much greater potency with ERalpha than with ERbeta. In contrast, daidzein was 1000 times more powerful in stimulating ERbeta- over ERalpha-mediated transactivation. Daidzein, in concentrations ranging from 5 nM to 50 microM, did not affect ERbeta-mediated transactivation induced by 1 nM E(2), but it significantly inhibited ERbeta-mediated transactivation induced by 10 nM E(2) at 500 nM. Despite the tremendous difference in sensitivity between the two in vitro systems, daidzein exhibited greater potency as an estrogen-antagonist for ERbeta-mediated activity.

Topics: Animals; Chickens; CHO Cells; Cricetinae; Cricetulus; Estradiol; Estrogen Receptor alpha; Estrogen Receptor beta; Humans; Isoflavones; Liver; Phytoestrogens; Response Elements; Transcriptional Activation; Vitellogenins

Daidzein, a phytoestrogen, has been reported to produce vasodilation via inhibition of Ca(2+) inflow. However, the involvement of large-conductance Ca(2+)-activated K(+) (BK(Ca)) channels in the effect of daidzein is debated. Therefore, the present study was designed to investigate the effect of daidzein on the rat cerebral basilar artery and the underlying molecular mechanisms. Isolated cerebral basilar artery rings and single vascular smooth muscle cells (VSMCs) were used for vascular reactivity and electrophysiology measurements, to investigate the effect of daidzein on BK(Ca) channels in cerebral basilar artery smooth muscle. In addition, the human BK(Ca) channel alpha-subunit gene (hslo) was transfected into HEK293 cells, to directly assess whether daidzein activates BK(Ca) channels. The results showed that daidzein produced a concentration-dependent but endothelium-independent relaxation in rat cerebral basilar arteries. Paxilline, a selective BK(Ca) channel blocker, significantly inhibited the daidzein-induced vasodilation, whereas NS1619, a selective BK(Ca) channel opener, enhanced the vasodilation. In the whole-cell configuration, daidzein increased noisy oscillation currents in cerebral basilar artery VSMCs in a concentration-dependent manner, and washout of daidzein or blockade of BK(Ca) channels with paxilline fully reversed the increase. However, daidzein did not substantially affect hSlo currents in HEK293 cells when applied to the outside of the cell membrane. In conclusion, these results indicate that the activation of BK(Ca) channels in VSMCs at least partly contributes to the daidzein-induced vasodilation of the rat cerebral basilar artery. The beta1-subunit of BK(Ca) channels plays a critical role in the activation of BK(Ca) currents by daidzein.

Topics: Animals; Basilar Artery; Cerebral Arteries; Dose-Response Relationship, Drug; Electrophysiology; Isoflavones; Male; Muscle, Smooth, Vascular; Patch-Clamp Techniques; Phytoestrogens; Potassium Channels, Calcium-Activated; Rats; Rats, Sprague-Dawley; Vasodilation

Breast cancer is a public health problem in the Western countries. Several studies have shown that BRCA2, like BRCA1 oncosuppressors, are strongly involved in hereditary and sporadic mammary carcinogenesis. It has also been suggested that soy has a protective effect against breast cancer in Asia and, more particularly, phytoestrogens such as daidzein and genistein. Thus, phytoestrogens may have an impact on the expression of BRCA2 gene, and there is a possible link between BRCA2 and genes acting around the BRCA2. To focus on these processes, we set up the BRCA2 specific knockdown by RNA interference in two breast tumor cell lines (MCF-7 and MDA-MB-231) and also in a non-tumorigenic breast cell line (MCF-10a). After inhibition of BRCA2 expression, cells were maintained in different conditions and treated with either daidzein or genistein or left untreated. Microarray analysis of mRNAs isolated from the BRCA2 knocked down MCF-7, MDA-MB-231, and MCF-10a cell lines after being treated with phytoestrogens showed 35 differentially expressed genes between positive-ERbeta cells and negative-ERbeta cells. After genistein or daidzein treatments, BRCA1 was found to be up-regulated when knocked down with BRCA2-siRNA MCF-7 and BRCA2 was found to be up-regulated when knocked down with BRCA2-siRNA MDA-MB 231 cells. In MCF-10a, we observed a significant decrease in BAX and BCL2 expressions with a greater effect of daidzein. We also found an increase in BRIP expression between genistein and daidzein treatment knocked down with BRCA2-siRNA MCF-7 and MDA-MB-231 cell lines.

Topics: bcl-2-Associated X Protein; Breast Neoplasms; Cell Line, Tumor; DNA-Binding Proteins; Fanconi Anemia Complementation Group Proteins; Female; Gene Expression Regulation, Neoplastic; Genes, bcl-2; Genes, BRCA2; Genistein; Humans; Isoflavones; Phytoestrogens; Polymerase Chain Reaction; Receptors, Estrogen; RNA Helicases

Previous studies have suggested that daidzein's metabolites, equol and O-desmethylangolensin (O-DMA), rather than daidzein itself may contribute to the beneficial effects of soya foods in the prevention of CVD. The present study aims to assess the proportion of equol and O-DMA producers, and to compare differences in anthropometric factors, serum lipids, glucose and uric acid between producers and non-producers in Chinese adults aged 20-69 years. For the present cross-sectional study, 202 subjects (100 women and 102 men) were recruited. Twenty-four-hour urinary daidzein and its metabolites were determined in these subjects while on their usual diet and again after a 3-d isoflavone challenge. Fasting serum lipids, glucose and uric acid were examined on their usual diet. Three days of 24 h dietary recalls were used to assess dietary intakes. Of the 202 subjects, 27 (13.4 %) and 27 (13.4 %) excreted equol and O-DMA on their usual diet, and 101 (50 %) and 94 (46.5 %) produced equol and O-DMA after a load of 80 mg/d isoflavones. Equol producers showed lower serum uric acid ( - 10.2 %, P = 0.001), TAG ( - 29.5 %, P = 0.007) and waist:hip ratio ( - 2.6 %, P = 0.032), and tended to have higher HDL cholesterol (6.3 %, P = 0.069) compared with equol non-producers. There were no significant differences in serum lipids, glucose and uric acid between O-DMA producers and non-producers. In conclusion, equol phenotypes might influence cardiovascular risk.

Topics: Adult; Cardiovascular Diseases; China; Cholesterol, HDL; Cross-Sectional Studies; Diet Records; Equol; Female; Humans; Isoflavones; Lipids; Male; Middle Aged; Phenotype; Phytoestrogens; Risk Factors; Triglycerides; Uric Acid; Waist-Hip Ratio; Young Adult

Although an immunomodulatory role of the soy isoflavone genistein has been demonstrated, the effects of other soy isoflavones on induction of antigen (Ag)-specific immune responses are not known. In this study, we therefore investigated the effects of daidzein and equol on ovalbumin (OVA)-specific T cell and B cell responses in BALB/c mice. Mice that had been treated with 20 mg/kg equol showed a significantly higher level of OVA-specific IgE than control mice. Levels of interferon (IFN)-gamma and interleukin (IL)-4 production were not different between the control and equol groups. However, IL-13 production level in mice administered 20 mg/kg equol was significantly higher than that in control mice. Strong induction of OVA-specific IgE production by equol was also observed in ovariectomized BALB/c mice, suggesting that the immunomodulatory effect of equol is not affected by endogenous estrogen.

Topics: Animals; B-Lymphocytes; Equol; Estrogens; Female; Glycine max; Hypersensitivity, Immediate; Immunization; Immunoglobulin E; Immunologic Factors; Interleukin-13; Isoflavones; Mice; Mice, Inbred BALB C; Ovalbumin; Ovariectomy; Phytoestrogens; T-Lymphocytes

Plant-derived estrogen-like compounds such as isoflavones (IF) especially daidzein and genistein are said to be preserving the bone in the osteoporotic conditions. However, it is not known whether a combination of IF and calcium (Ca) supplementation attenuates losses in bone mass and prevents the loss of vitamin D (VD). The present study addresses the role of phytoestrogens (PE) and Ca supplementation in low Ca and low VD diet induced osteoporosis (OSP). Cowpea (CP) which has high amount of the IF was selected to study its effect on diet induced osteoporotic conditions. Female weanling WNIN rats (total of 68) were divided into five groups and fed for five weeks on semisynthetic diet with low Ca (0.15%) and low VD (0.1IU/day/rat) in combination with low (10 mg/kg) or high (25 mg/kg) concentrations of PEs derived from CPIF. The study groups are: (I) normal Ca(0.47%) and normal VD (25IU/day/rat), (II) low Ca+low VD, (III) low Ca+low VD+low CPIF (10 mg/kg diet), (IV) low Ca+low VD+high CPIF (25 mg/kg diet) and (V) low Ca+low VD+17-(-estradiol (3.2 mg/kg diet). After the development of OSP the group II was subgrouped into: (SG I) continued on low Ca+VD, (SG II) low CPIF, (SG III) high CPIF, (SG IV) 17-beta-estradiol and (SG V) normal Ca and VD. Serum 25-VD levels were in the range of 14-38 ng/ml in groups I, III, IV and V, where as the values were very low in the group II (5.8 ng/ml). These were partially reversed upon supplementation of CPIF. The results correlated with altered Ca levels, body weight, bone mineral density and content and other related biochemical parameters. The paper further explains the possibility of protective and therapeutic role of VD in the presence of CPIF in osteoporotic health manifestations.

Topics: Animals; Bone Density; Calcium; Cholecalciferol; Dietary Supplements; Disease Models, Animal; Female; Genistein; Isoflavones; Osteoporosis; Phosphorus; Phytoestrogens; Plants; Rats; Vitamin D

Previous studies have suggested that soy isoflavones exert anticarcinogenic effects against prostate cancer. We propose that soy extracts, containing a mixture of soy isoflavones and other bioactive components, would be a more potent chemo-preventive agent than individual soy isoflavones. We compared the apoptotic effects of whole soy extracts and individual soy isoflavones, genistein and daidzein, on prostate cancer cells. The soy extract contained 50% w/w of total isoflavones with approximately 1:5.5:3.5 ratios of genistin, daidzin and glycitin, respectively. Benign prostate hyperplasia (BPH-1), LnCap and PC3 cells were treated with varying concentrations of soy extract, genistein or daidzein and analyzed for cell cycle alterations and induction of apoptosis. At equal concentrations (25 micromol/L), soy extract induced a significantly higher percentage of cells undergoing apoptosis than genistein or daidzein (P < 0.001). No significant changes in cell cycle arrest or apoptosis were observed in non-cancerous BPH-1 cells treated with soy extract, suggesting that the effects of soy extract may be tumor cell specific. On the contrary, both genistein and daidzein induced apoptosis in BPH-1 cells, suggesting that individual isoflavones may have cytotoxicity in non-cancerous cells. Soy extracts also increased Bax expression in PC3 cells, but no significant changes in nuclear factor kappaB (NF kappaB) activation were detected, suggesting that the induction of apoptosis was independent of the NF kappaB pathway. Food products that bear a combination of active compounds may be more efficacious and safer as chemo-preventive agents than individual compounds. This 'whole-food'-based approach is significant for the development of public health recommendations for prostate cancer prevention.

Topics: Anticarcinogenic Agents; Apoptosis; bcl-2-Associated X Protein; Caspases; Cell Cycle; Cell Line, Tumor; Dietary Supplements; Enzyme Activation; Glycine max; Humans; Isoflavones; Male; NF-kappa B; Phytoestrogens; Plant Extracts; Prostatic Neoplasms

Transformed hairy roots of Psoralea corylifolia were established by infection with Agrobacterium rhizogenes LBA 9402. The aim of this work was to elucidate the effects of media constituents on production of the phytoestrogenic isoflavones daidzein and genistein. A. rhizogenes strain LBA 9402 harboring Ri plasmid was used to transform stem segments of in vitro seedlings. The resultant hairy roots were confirmed by polymerase chain reaction (PCR) and exhibited Ri T-DNA. Transformed hairy root clones were cultured in Murashige and Skoog's (MS) medium altered with different concentrations of NH(4) (+) and NO(3) (-) and their growth and production of isoflavones were assessed. Biomass and productivity increased when MS medium was supplemented with NH(4) (+) and NO(3) (-) at a ratio of 20:10. Increased yield of daidzein was obtained when sucrose level in the culture medium increased, whereas decreased level of sucrose favored genistein production. The hairy roots produced the highest levels of daidzein (2.06% dry wt.) and genistein (0.37% dry wt.) in the presence of low concentrations of PO(4) (3-). Hairy roots secreted trace amounts of daidzein and genistein into the culture medium. The present results demonstrated that the productivity of daidzein was 2.2-fold more than that of untransformed roots.

Topics: Culture Media; Genistein; Isoflavones; Nitrates; Phosphates; Phytoestrogens; Plant Roots; Polymerase Chain Reaction; Psoralea; Quaternary Ammonium Compounds; Rhizobium

The androgen receptor (AR) plays a critical role in prostate cancer development and progression. This study aimed to use a computerized docking approach to examine the interactions between the human AR and phyto-oestrogens (genistein, daidzein, and flavone) and xeno-oestrogens (bisphenol A, 4-nonylphenol, dichlorodiphenyl trichloroethane [DDT], diethylstilbestrol [DES]). The predicted three-dimensional structure of AR and androgens was established using X-ray diffraction. The binding of four xeno-oestrogens and three phyto-oestrogens to AR was analysed. The steroids estradiol and dihydrotestosterone (DHT) were used as positive controls and thyroxine as negative control. All the ligands shared the same binding site except for thyroxine. The endogenous hormones DHT and 17beta-oestradiol showed the strongest binding with the lowest affinity energy (< -10 kcal mol(-1)). All three phyto-oestrogens and two xeno-oestrogens (bisphenol A and DES) showed strong binding to AR. The affinities of flavone, genistein, and daidzein were between -8.8 and -8.5 kcal mol(-1), while that of bisphenol A was -8.1 kcal mol(-1) and DES -8.3 kcal mol(-1). Another two xeno-oestrogens, 4-nonylphenol and DDT, although they fit within the binding domain of AR, showed weak affinity (-6.4 and -6.7 kcal mol(-1), respectively). The phyto-oestrogens genistein, daidzein and flavone, and the xeno-oestrogens bisphenol A and DES can be regarded as androgenic effectors. The xeno-oestrogens DDT and 4-nonylphenol bind only weakly to AR.

Topics: Benzhydryl Compounds; Computer Simulation; DDT; Diethylstilbestrol; Flavones; Genistein; Humans; Isoflavones; Ligands; Phenols; Phytoestrogens; Receptors, Androgen

Although the rate of breast cancer differs between women in Asian and Western countries, molecular genetics/genomics basis of this epidemiological observation remains elusive. Moreover, the intake of phytoestrogens is associated with a lower incidence of breast cancer. Genistein and daidzein are the primary soy isoflavones with a chemical structure similar to estrogens. Conceivably, the actions of phytoestrogens on gene expression signatures might mediate their postulated effects on breast cancer pathogenesis. The present study evaluated the transcriptional responsiveness of breast cancer cells to soy phytoestrogens using a whole-genome microarray-based approach. Human breast cancer cell lines and a fibrocystic breast cell line were treated with genistein or daidzein. We identified 278 and 334 differentially expressed genes after genistein or daidzein treatment, respectively, in estrogen-positive (MCF-7) and estrogen-negative (MDA-MB-231, MCF-10a) cells. Hierarchical clustering of this finding revealed a significant modulation, respectively, of 246 or 169 genes after genistein or daidzein exposures. Importantly, the molecular pathways for the differentially expressed genes included those that relate to cell communication, biodegradation of xenobiotics, lipid metabolism, signal transduction, and cell growth/death. These molecular observations collectively contribute to a growing knowledgebase on the putative mechanism(s) of action of phytoestrogens in breast cancer pathogenesis and chemoprevention.

Topics: Breast Neoplasms; Cell Line, Tumor; Cells, Cultured; Female; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Genistein; Genome, Human; Humans; Isoflavones; Microarray Analysis; Molecular Sequence Data; Multigene Family; Phytoestrogens

To enhance oral absorption of poorly water-soluble daidzein, self-microemulsifying drug delivery system (SMEDDS) composed of oil, surfactant and cosurfactant for oral administration of daidzein was formulated, and its physicochemical properties and pharmacokinetic parameters were evaluated. Solubility of daidzein was determined in various vehicles. Pseudo-ternary phase diagrams were constructed to identify the efficient self-microemulsification region and particle size distributions of the resultant microemulsions were determined using a laser diffraction sizer. From these studies, an optimized formulation consisting of Ethyl oleate (10%), Cremophor RH 40 (60%), and polyethylene glycol 400 (PEG400) (30%) was selected. The dissolution rate of daidzein from SMEDDS was significantly higher than the conventional tablet. Relative bioavailability of SMEDDS was enhanced about 2.5-fold compared with that of the control group. The data suggest that the use of SMEDDS provide a potential way of daidzein administered orally.

Topics: Administration, Oral; Animals; Biological Availability; Drug Delivery Systems; Emulsions; Isoflavones; Male; Microchemistry; Microscopy, Electron, Transmission; Phytoestrogens; Rats; Rats, Sprague-Dawley; Solubility

Intervention studies in man have shown that dietary soy isoflavones may provide certain health benefits. One of the possible reasons for this benefit is that the daidzein contained in soy is converted to S-equol. As part of a drug development program for S-equol, three genotoxicity studies were conducted. The studies comprised bacterial mutation, chromosomal aberration, and in vivo bone marrow micronucleus tests conducted according to Good Laboratory Practices (GLP). No evidence of genotoxic activity was observed in the in vitro tests at concentrations up to those associated with cell toxicity. In addition, no evidence of cytotoxicity or genotoxicity was seen in the rat bone marrow micronucleus test in animals dosed at levels up to the standard limit of 2000 mg/kg. It is concluded that S-equol is not active in the standard battery of genotoxicity assays recommended by the International Conference on Harmonisation (ICH) for registration of new pharmaceuticals. The current results support the further development of S-equol.

Topics: Animals; Bacteria; Bone Marrow Cells; Chromosome Aberrations; Equol; Female; Isoflavones; Male; Micronucleus Tests; Mutagenicity Tests; Mutagens; Phytoestrogens; Rats; Rats, Sprague-Dawley; Salmonella typhimurium

Phytoestrogens are plant compounds that mimic the actions of endogenous estrogens. The abundance of these chemicals in nature and their potential effects on health require the development of a convenient method to detect phytoestrogens. We have developed a nanoparticle (NP)-conjugated FRET probe based on the human estrogen receptor α (ER) ligand-binding domain (LBD) to detect phytoestrogens. The NP-conjugated FRET probe showed fluorescence signals for genistein, resveratrol and daidzein compounds with Δ ratios of 1.65, 2.60 and 1.37 respectively, which are approximately six times greater compared to individual FRET probes. A significantly higher signal for resveratrol versus genistein and daidzein indicates that the probe can differentiate between antagonistic phytoalexin substances and agonistic isoflavone compounds. NP-conjugated probes demonstrated a wide dynamic range, ranging from 10(-18) to 10(-1) M with EC(50) values of 9.6 × 10(-10), 9.0 × 10(-10) and 9.2 × 10(-10) M for genistein, daidzein and resveratrol respectively, whereas individual probes detected concentrations of 10(-13) to 10(-4) M for phytoestrogens compounds. The time profile revealed that the NP-conjugated probe is stable over 30 h and there is not a significant deviation in the FRET signal at room temperature. These data demonstrate that conjugation of a FRET probe to nanoparticles is able to serve as an effective FRET sensor for monitoring bioactive compounds with significantly increased sensitivity, dynamic range and stability.

Topics: Binding Sites; Estrogen Receptor alpha; Fluorescence Resonance Energy Transfer; Fluorescent Dyes; Genistein; Humans; Isoflavones; Ligands; Nanoparticles; Phytoestrogens; Resveratrol; Sensitivity and Specificity; Stilbenes

We evaluated the relationship of spot urinary concentrations of phytoestrogens with total prostate cancer and tumor grade in a hospital-based case-control study in Jamaica. Urine samples were analyzed for genistein, daidzein, equol (isoflavones), and enterolactone (lignan) among newly diagnosed cases (n = 175) and controls (n = 194). Urinary concentrations of enterolactone (lignan) were higher among cases. There were no significant differences in median concentrations of isoflavone excretion. Compared with non-producers of equol (reference tertile), men who produced equol were at decreased risk of total prostate cancer (tertile 2: OR, 0.42; CI, 0.23-0.75) (tertile 3: OR, 0.48; CI, 0.26-0.87) (p (trend), 0.020) and high-grade disease (tertile 2: OR, 0.31; CI, 0.15-0.61) (tertile 3: OR, 0.29; CI, 0.13-0.60) (p (trend), 0.001). Higher concentrations of enterolactone were positively related to total prostate cancer (OR, 1.85; CI, 1.01-3.44; p (trend), 0.027) as well as high-grade disease (OR, 2.46; CI, 1.11-5.46; p (trend), 0.023). There were no associations between urinary excretion of genistein and daidzein with risk of prostate cancer. Producers of equol (isoflavone) may be at reduced risk of total- and high-grade prostate cancer whereas enterolactone may increase the likelihood of disease.

Topics: Aged; Carcinoma; Case-Control Studies; Equol; Genistein; Humans; Isoflavones; Jamaica; Lignans; Male; Middle Aged; Phytoestrogens; Prostatic Neoplasms; Risk

In this study, the effect of daidzin or daidzein isolated from Pueraria lobata on the memory impairments induced by scopolamine was assessed in male mice using the passive avoidance and the Morris water maze tasks. Administration of daidzin (5 mg/kg) or daidzein (5 mg/kg) significantly reversed the scopolamine (1 mg/kg)-induced cognitive impairments in male mice as evidenced by the passive avoidance test (p < 0.05) and on the Morris water maze test (p < 0.05). Moreover, the ameliorating effects of daidzin or daidzein were antagonized by tamoxifen (1 mg/kg), the nonspecific estrogen receptor antagonist. These results indicate that daidzin or daidzein may be useful in cognitive impairment induced by cholinergic dysfunction, and this beneficial effect is mediated, in part, via estrogen receptor.

Topics: Animals; Avoidance Learning; Cholinergic Antagonists; Estrogen Antagonists; Glucosides; Isoflavones; Learning Disabilities; Male; Maze Learning; Memory Disorders; Mice; Mice, Inbred ICR; Phytoestrogens; Receptors, Estrogen; Scopolamine; Swimming; Tamoxifen

In the United States, about 25% of infant formula sold is based on soy protein, which is an important source of estrogenic isoflavones in the human food supply. Nevertheless, few studies report isoflavone levels in infants. We did a partly cross-sectional and partly longitudinal pilot study to examine children's exposure to isoflavones from different feeding methods. A total of 166 full-term infants between birth and 1 year of age were recruited into soy formula, cow milk formula, or breast milk regimens according to their feeding histories. A total of 381 urine, 361 saliva, and 88 blood samples were collected at 382 visits. We used automated online solid-phase extraction coupled to high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) for measuring three isoflavones (daidzein, genistein, and equol) in urine, and used similar LC/MS/MS techniques for saliva and blood spots. Concentrations of daidzein and genistein were undetectable in most blood or saliva samples from children fed breast milk or cow milk formula. The proportion of non-detectable values was somewhat lower in urine than in the other matrices. Concentrations of equol were detectable only in a few urine samples. For both daidzein and genistein, urine contained the highest median concentrations, followed by blood and then saliva. Urinary concentrations of genistein and daidzein were about 500 times higher in the soy formula-fed infants than in the cow milk formula-fed infants. The correlations between matrices for either analyte were strikingly lower than the correlation between the two analytes in any single matrix. We did not find significant correlations between isoflavone concentrations and the levels of certain hormones in children fed soy formula. Our results, based on much larger numbers of infants, strongly confirm previous reports, but whether phytoestrogens in soy formula are biologically active in infants is still an open question. We plan further longitudinal studies focusing on physical and developmental findings reflecting the effects of estrogen exposure.

Topics: Animals; Chromatography, High Pressure Liquid; Cross-Sectional Studies; Female; Genistein; Humans; Infant; Infant Formula; Infant, Newborn; Isoflavones; Longitudinal Studies; Male; Milk; Milk, Human; Phytoestrogens; Pilot Projects; Saliva; Soy Milk; Tandem Mass Spectrometry

Mammographic breast density is an established marker of breast cancer risk, and is hormonally sensitive. Studies suggest that production of the daidzein metabolites equol and O-Desmethylangolensin (ODMA) may be associated with hormones and hormonally mediated factors, but few studies have assessed relationships between the capacity to produce these metabolites and breast density.. To evaluate the relationship between equol- and ODMA-producer phenotypes and breast density in premenopausal women in the United States.. Two hundred and three women attended a clinic visit and 200 provided a urine sample following a 3 day soy challenge. Samples were analyzed for isoflavones by GC-MS to determine daidzein-metabolizing phenotypes. Percent density on recent (<14 month prior to their clinic visit) mammograms was assessed by one reader using a computer-assisted method. Multiple regression analysis was used to assess relationships between the production of equol and ODMA and breast density. Results 55(27.5%) and 182(91%) women were classed as equol- and ODMA-producers (>87.5 ng/ml urine), respectively. In unadjusted and adjusted analyses, there were no differences in breast density between producers and non-producers of either equol or ODMA (P > 0.05).. In this population of low-soy consuming premenopausal women, there were no associations between daidzein-metabolizing phenotypes and breast density, suggesting that these phenotypes per se do not influence premenopausal breast density.

Topics: Adult; Breast; Breast Neoplasms; Equol; Female; Gas Chromatography-Mass Spectrometry; Humans; Hydroxyestrones; Isoflavones; Mammography; Middle Aged; Phenotype; Phytoestrogens; Soy Foods

This study proposed secondary metabolites incremental yield due to manipulation of nutrient components into the culture medium. To validate this, the effects of nutrients such as carbon, phosphate and nitrogen on growth and production of phytoestrogens daidzein and genistein by suspension cultures of Psoralea corylifolia was investigated for the first time. The maximum production of daidzein and genistein was achieved when sucrose and maltose used as a sole source of carbon. Suspension cell cultures enriched with sucrose (3%) stimulated accumulation of isoflavones daidzein (1.76% dry wt) and genistein (0.25% dry wt) compared to glucose, fructose and maltose. Sucrose feeding strategy significantly stimulated biomass growth and isoflavones (2.79% dry wt of daidzein and 0.32% dry wt of genistein) production rate. Reduced concentrations of phosphate (0.625 mM) promoted daidzein (1.89% dry wt) and genistein (0.26% dry wt) production by suspension cell cultures, whereas high amount (5mM) in medium was inhibited isoflavones production. It was observed that medium fortified with NH(4)(+) and NO(3)(-) alone inhibited production of isoflavones. The maximum production obtained of daidzein (2.20% dry wt) and genistein (0.29% dry wt) when medium comprised with NH(4)(+)/NO(3)(-) at ratio 20:40 mM as a nitrogen source. Similar nutrient components ratio when altered NH(4)(+)/NO(3)(-); 40:20mM) resulted in approximately 3-fold decrease in production. HPLC analysis revealed that suspension cells cultures leached out trace amount of daidzein and genistein into the culture medium.

Topics: Biomass; Biotechnology; Carbon; Chromatography, High Pressure Liquid; Culture Media; Genistein; Isoflavones; Nitrogen; Phosphates; Phytoestrogens; Psoralea

Neonatal exposure to genistein (GEN), an isoflavone abundant in soy, favorably modulates bone mineral density (BMD) and bone strength in mice at adulthood. The study objective was to determine whether early exposure to a combination of the soy isoflavones daidzein (DAI) and GEN that naturally exists in soy protein-based infant formula results in greater benefits to bone at adulthood than either treatment alone. Male and female CD-1 mice (n = 8-16 pups per group per gender) were randomized to subcutaneous injections of DAI (2 mg x kg body weight(-1) x d(-1)), GEN (5 mg x kg body weight(-1) x d(-1)), DAI+GEN (7 mg x kg body weight(-1) x d(-1)), diethylstilbesterol (DES; positive control) (2 mg x kg body weight(-1) x d(-1)), or control (CON) from postnatal d 1-5 and were studied to 4 mo of age. BMD, biomechanical bone strength, and bone microarchitecture were assessed at the femur and lumbar vertebrae (LV). Females treated with DAI, GEN, DAI+GEN, or DES had greater (P < 0.05) BMD at the LV compared with CON and vertebra in the DAI and DES group were more resistant to compression fractures. Microstructural analyses demonstrated that treatment with DAI and GEN resulted in greater (P < 0.05) trabecular connectivity and trabecular thickness, respectively, than the CON. In conclusion, neonatal exposure to DAI and/or GEN had a positive effect on the skeleton of female mice at adulthood, but, compared with individual treatments, DAI+GEN did not have a greater benefit to bone in females or males.

Topics: Animals; Animals, Newborn; Body Weight; Bone Density; Bone Development; Drug Therapy, Combination; Female; Femur; Genistein; Isoflavones; Male; Mice; Phytoestrogens; Sex Characteristics; Spine

Phytoestrogens are a diverse group of nonsteroidal plant compounds that occur naturally in many plants. Because they possess a ring system similar to estrogens they are able to bind on estrogen receptors alpha and beta in humans. The effects of the phytoestrogens genistein and daidzein on the production of progesterone and estrogen in isolated human term trophoblast cells in vitro were tested in this study.. Cytotrophoblast cells were isolated from human term placentas. Phytoestrogens genistein and daidzein were incubated in different concentrations with trophoblast cells. Untreated cells were used as controls. After 24 h aliquots were removed and tested for progesterone and estrogen production.. The production of the steroid hormones progesterone and estrogen are influenced by phytoestrogens genistein and daidzein in human term trophoblast cells. A strong inhibition effect of both phytoestrogens tested in the production of progesterone was demonstrated. In addition, a significant stimulating effect on estrogen production by genistein and daidzein was observed.. Results obtained with this study show that phytoestrogens (genistein and daidzein) sufficiently reduce progesterone production in human term trophoblast cells. Because blockade of progesterone is a possible mechanism involved in initiation of labor, we may speculate that high doses of phytoestrogens at the feto-maternal interphase could play a negative role in maintenance of pregnancy. Stimulation of estrogen production by genistein and daidzein in trophoblast cells is probably due to estrogen receptor blocking effects of both phytoestrogens. Trophoblast cells seem to compensate blocking of its estrogen receptors by higher estrogen production.

Topics: Cells, Cultured; Dose-Response Relationship, Drug; Estradiol; Female; Genistein; Humans; Isoflavones; Phytoestrogens; Pregnancy; Progesterone; Term Birth; Trophoblasts

The present study was undertaken to determine whether treatment with genistein, the plant-derived estrogen-like compound influences agonist-induced vascular smooth muscle contraction and, if so, to investigate related mechanisms. The measurement of isometric contractions using a computerized data acquisition system was combined with molecular experiments. Genistein completely inhibited KCl-, phorbol ester-, phenylephrine-, fluoride- and thromboxane A(2)-induced contractions. An inactive analogue, daidzein, completely inhibited only fluoride-induced contraction regardless of endothelial function, suggesting some difference between the mechanisms of RhoA/Rho-kinase activators such as fluoride and thromboxane A(2). Furthermore, genistein and daidzein each significantly decreased phosphorylation of MYPT1 at Thr855 had been induced by a thromboxane A(2) mimetic. Interestingly, iberiotoxin, a blocker of large-conductance calcium-activated potassium channels, did not inhibit the relaxation response to genistein or daidzein in denuded aortic rings precontracted with fluoride. In conclusion, genistein or daidzein elicit similar relaxing responses in fluoride-induced contractions, regardless of tyrosine kinase inhibition or endothelial function, and the relaxation caused by genistein or daidzein was not antagonized by large conductance K(Ca)-channel inhibitors in the denuded muscle. This suggests that the RhoA/Rho-kinase pathway rather than K(+)-channels are involved in the genistein-induced vasodilation. In addition, based on molecular and physiological results, only one vasoconstrictor fluoride seems to be a full RhoA/Rho-kinase activator; the others are partial activators.

Topics: Animals; Aorta; Blotting, Western; Carcinogens; Cardiotonic Agents; Genistein; Isoflavones; Male; Muscle Contraction; Muscle Relaxation; Peptides; Phenylephrine; Phorbol Esters; Phosphorylation; Phytoestrogens; Potassium Channel Blockers; Potassium Channels, Calcium-Activated; Potassium Chloride; Protein Phosphatase 1; Rats; Rats, Sprague-Dawley; rhoA GTP-Binding Protein; Sodium Fluoride; Thromboxane A2; Toxins, Biological

In previous studies on HeLa cells we demonstrated estrogen-responsiveness of the epidermal growth factor receptor (EGFR) gene, as 17beta-estradiol (E(2)) and selective estrogen receptor modulators (SERMs) genistein (G), daidzein (D), and 4-hydroxytamoxifen (4OH-T) modulated its transcription in a ligand- and estrogen receptor (ER) isoform-specific way. This study describes further investigations into the role of ERs in mediating the effects induced by E(2) and SERMs on EGFR expression, and the relationship between the actions of ERs and EGFR in U2OS osteosarcoma cells stably expressing ERalpha or ERbeta. Cell number and DNA content determination revealed that E(2), G, and D inhibited proliferation and cell cycle progression and promoted apoptosis in both cell lines. In parallel, changes in cell morphology typical of osteoblast maturation were observed via optical microscopy. Consistently, quantitative PCR and Western blot analysis showed an up-regulation of markers of osteoblast differentiation and bone repair, and a decrease in EGFR expression. The transfection of specific antisense (AS) oligonucleotides strengthened our hypothesis that EGFR reduction caused changes in the proliferation/differentiation pattern comparable to those induced by ER ligands. The link between the ER and EGFR pathways was confirmed by treatment with 4OH-T, which decreased the EGFR level and produced differentiation effects via ERalpha, but induced both EGFR expression and proliferation effects via ERbeta. In conclusion, we show that also in U2OS cells, E(2) and SERMs are able to modulate the expression of the EGFR gene and can affect events strictly controlled by its signaling pathway, such as the maturation of osteoblasts.

Topics: Apoptosis; Biomarkers; Cell Cycle; Cell Differentiation; Cell Line, Tumor; Cell Proliferation; Cell Shape; Down-Regulation; ErbB Receptors; Estradiol; Estrogen Receptor alpha; Estrogen Receptor beta; Genistein; Humans; Isoflavones; Osteoblasts; Osteosarcoma; Phytoestrogens; RNA, Messenger; Selective Estrogen Receptor Modulators; Tamoxifen; Time Factors

Gut bacterial modification of soy isoflavones produces metabolites that differ in biological activity from the parent compounds. Hydrolysis of glycosides results in more active compounds. In contrast, further degradation and transformation of aglycones produce more or less active compounds, depending on the substrate metabolized and the product formed. Bacterial metabolism of soy isoflavones varies among individuals. The predominant daidzein metabolites produced by human intestinal bacteria are equol and O-desmethylangolensin. Among humans, 30-50% have the bacteria capable of producing equol and 80-90% harbor O-desmethylangolensin-producing bacteria. Factors that influence the capacity to produce equol and O-desmethylangolensin are not clearly established; however, gut physiology, host genetics, and diet are reported to contribute to interindividual differences in conversion of daidzein to equol. Effects of these phenotypes on human health are poorly characterized. Some studies in high soy-consuming populations reported an inverse association between urinary and serum equol concentrations and breast and prostate cancer risk. Furthermore, several studies of soy supplementation and bone density suggest that soy products may be more effective in maintaining bone density in equol-producing individuals. Factors that contribute to the phenotypes and the relation of these specific phenotypes to human health need to be further elucidated. The extent to which isoflavone metabolism is key to the efficacy of soy foods remains to be established.

Topics: Bacteria; Equol; Feeding Behavior; Female; Humans; Hydrolysis; Intestines; Isoflavones; Male; Phytoestrogens

We investigated the effects of daidzein on the antioxidant defence system in mice with 7,12-dimethylbenz[a]-anthracene (DMBA)-induced oxidative stress. Daidzein was administered orally at 5 and 25 mg/kg body weight for 5 weeks. Subsequently, mice pretreated with daidzein received DMBA intragastrically twice a week for 2 weeks. As controls, mice were given vehicle or DMBA alone. In the DMBA group, biomarkers of oxidative stress (thiobarbituric acid reactive substances value, carbonyl content) were significantly increased. However, the rise in oxidative damage was significantly reduced by daidzein at the higher dose. In addition, several antioxidant enzymes were downregulated in the DMBA-treated mice. Catalase and superoxide dismutase activity was increased by daidzein in a dose-dependent manner. Although the reduced/oxidized glutathione ratio was unaffected, glutathione peroxidase and reductase were activated by daidzein, and the effect was significant at the higher dose. Further, in the DMBA-treated mice, apoptosis was induced by a decrease in Bcl-2 and an increase in Bax. These changes were restored to their normal values in the daidzein-treated mice. Upregulation of caspase-3 was also decreased by daidzein. These results suggest that daidzein exerts a hepatoprotective effect on mice with DMBA-induced oxidative stress through its antioxidant activity and the reduction of apoptosis.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Antioxidants; Apoptosis; Carcinogens; Cytoprotection; Drug Evaluation, Preclinical; Equol; Female; Glutathione; Isoflavones; Liver; Mice; Mice, Inbred ICR; Oxidative Stress; Phytoestrogens

Because of the complexity of estrogen receptor (ER) physiological activity, the interaction of pure isoflavones or soy-based diets on ER needs to be clearly demonstrated.. To investigate the effects of the administration of isoflavones as a pure compound or as a component of diet on the ER transcriptional activity in adult mice.. Effects of acute (6 h) and chronic (21 days) oral administration of soy milk, pure genistein and a mix of genistein and daidzein was studied in living ERE-Luc mice. In this animal model, the synthesis of luciferase is under the state of ER transcriptional activity. Luciferase activity was measured in living mice by daily bioluminescence imaging sessions and in tissue extracts by enzymatic assay.. Acute, oral administration of genistein or soymilk caused a significant increase of ER activity in liver. In a 20 day long treatment, soymilk was more potent than genistein in liver and appeared to extend its influence on ER transcriptional activity in other tissues, such as the digestive tract. A mixture of pure genistein and daidzein at the same concentration as in soymilk failed to induce significant changes during acute and chronic studies suggesting an important, uncharacterized role of the soymilk matrix. Consistent with this observation, synergistic effects of the matrix plus isoflavones were observed in MCF-7 cells stably transfected with the ERE-luc construct.. This study underlines the limitations of the analysis of single food components in the evaluation of their effects on estrogen receptor activity and advocates the necessity to use complex organisms for the full comprehension of the effects of compounds altering the endocrine balance.

Topics: Administration, Oral; Animals; Cell Line; Female; Genistein; Isoflavones; Liver; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Phytoestrogens; Receptors, Estrogen; Soy Milk

Dietary phytoestrogens may be involved in the occurrence of chronic diseases. Reliable information on the phytoestrogen content in foods is required to assess dietary exposure and disease risk in epidemiological studies. However, there is little information on isoflavone, lignan, and coumestrol content of cereals and cereal-based foods, leading to an underestimation of intake. This is the first study of phytoestrogens (isoflavones: biochanin A, daidzein, formononetin, genistein, glycitein; lignans: matairesinol, secoisplariciresinol; coumestrol) in a comprehensive selection of 101 cereals and cereal-based foods-including breads, breakfast cereals, biscuits, pasta and rice-consumed in the UK using a sensitive LCMS technique with 13C-labelled internal standards. Phytoestrogens were detected in all foods analyzed; bread contained the highest amount of phytoestrogens-many as isoflavones-with an average content of 375 +/- 67 microg/100 g wet weight (excluding soya-linseed bread with 12,000 microg/100 g). Most other foods contained less than 100 microg/100 g, many as lignans. Our study shows that all foods analyzed contained phytoestrogens, with the highest amount found in breads, making them one of the main sources of dietary phytoestrogens in the UK. These results will allow a more accurate estimation of exposure to dietary phytoestrogens.

Topics: Edible Grain; Food Analysis; Genistein; Isoflavones; Phytoestrogens

As a new sample preparation technique, pressurized liquid extraction (PLE), in combination with reversed-phase liquid chromatography (RP-LC) and photodiode-array (PDA) detection were used for the isolation and determination of phytoestrogenic isoflavones in hydrolyzed extracts obtained from aerial parts of five Trifolium L. (clover) species. To optimize the effectiveness of PLE procedure, variable extraction parameters: methanol and acetone (or their 75% aqueous solutions), as extraction solvents, temperatures (75, 100 and 125 degrees C) and the changeable number of static extraction cycles were tested. Additionally, two other micropreparative techniques: ultrasound-assisted extraction (UAE), and conventional solvent extraction (CSE), under optimized conditions, were also used and compared. Optimum extraction efficiency, expressed in the highest yield of biochanin A, formononetin, daidzein and genistein from plant material, with PLE, using methanol-water (75:25, v/v) as an extraction solvent, an oven temperature of 125 degrees C and three 5-min static extraction cycles, was obtained.

Topics: Chromatography, High Pressure Liquid; Genistein; Isoflavones; Phytoestrogens; Solvents; Trifolium; Ultrasonics

There are limited reports on the bioavailability and pharmacokinetics of isoflavones in elderly humans and aged animals. The present study was conducted to assess the effect of glycosidation of isoflavones on their bioavailability and pharmacokinetics in aged (20 month old) male Fischer-344 (F-344) rats. The F-344 rat, developed by the National Institute on Aging, is an inbred rat model that is commonly used for aging studies and resembles many features of aging humans. Three sources of isoflavones; Novasoy (a commercial supplement), a mixture of synthetic aglycons (daidzein, genistein and glycitein), and a mixture of synthetic glucosides (daidzin, genistin, and glycitin) were tested. Following administration, blood samples were collected at different times (0-48 h post-oral gavage and 0-8 h post-IV dosing). Plasma isoflavones and 7-hydroxy-3-(4'-hydroxyphenyl)-chroman (a metabolite of daidzein) were measured by LC/MS. The extent of absorption was determined by comparing the area under the curve (AUC) of the plasma-concentration time curve after intravenous (IV) administration with that following oral administration. The extent of bioavailability was then calculated as: %bioabailability = (AUC(or)/AUC(IV))x(Dose(IV)/Dose(or))x100. Bioavailabilities for genistein were significantly (p = 0.013) higher for the aglycon (35 +/- 9%) compared with the glucoside forms (11 +/- 3%). In contrast, the bioavailabilities for glycitein were significantly (p = 0.011) higher in Novasoy (27 +/- 13%) and the glucoside form (21 +/- 10%) compared with the aglycon (8 +/- 3%). No significant differences in the bioavailability of daidzein were observed in aged rats dosed with aglycon, glucoside or Novasoy. However, aged rats were able to produce equol as early as 8 h post-dosing. In summary, the source of isoflavones had significant effects on genistein and glycitein bioavailability in aged male rats.

Topics: Aging; Animals; Biological Availability; Diet; Equol; Genistein; Glucosides; Glycine max; Glycosylation; Injections, Intravenous; Isoflavones; Male; Phytoestrogens; Rats; Rats, Inbred F344

To investigate the effects of phytoestrogens (daidzein and genistein) on the testosterone production of rat Leydig cells and the possible mechanisms.. Primary Leydig cells were obtained from 3-month old male SD rats using discontinuous Percoll density gradient centrifugation. The effects of phytoestrogens at various concentrations were evaluated by ELISA, with hCG as the positive control. The mRNA expression of P450 side-chain cleavage enzyme (P450scc) was analyzed by semi-quantitative RT-PCR.. Genistein at 0.1 micromol/L obviously promoted the secretion of testosterone and upregulated the mRNA level of P450scc. At a higher concentration of 5 micromol/L, however, both daidzein and genistein significantly inhibited the testosterone production of Leydig cells (P > 0.05).. Genistein can promote the testosterone production of Leydig cells at a low concentration (0.1 micromol/L), but both daidzein and genistein can inhibit it at a higher concentration ( >5 micromol/L).

Topics: Animals; Cells, Cultured; Genistein; Isoflavones; Leydig Cells; Male; Phytoestrogens; Rats; Rats, Sprague-Dawley; Testosterone

Phytoestrogens are of special interest in prostate cancer research because populations in Asia with a high consumption of phytoestrogens have a lower incidence of the disease than comparable populations in Western countries.. This case-control study is nested within a large multiethnic cohort in Hawaii and California. Urine samples were analysed for daidzein, genistein, equol, and enterolactone among 249 incident prostate cancer cases and 404 controls matched on age, race/ethnicity, date/time of specimen collection, and fasting status.. The median excretion of daidzein was 0.173 nmol mg(-1) creatinine in cases and 0.291 in controls (P=0.01), and the median excretion of genistein was 0.048 in cases and 0.078 in controls (P=0.05). An inverse association was seen for daidzein overall (odds ratio for the highest vs lowest quintile=0.55, 95% confidence interval=0.31-0.98, P(trend)=0.03) and seemed to apply to localized (P(trend)=0.08) as well as advanced or high-grade cancer (P(trend)=0.09). This association was consistent across the four ethnic groups examined. Although the relationship was weaker for genistein, the odds ratios and trends were similarly inverse. Urinary excretion of equol and enterolactone was not significantly related to prostate cancer risk.. Our findings suggest that high intake of isoflavones, as reflected by urinary excretion of daidzein and genistein, may be protective against prostate cancer.

Topics: Aged; California; Case-Control Studies; Cohort Studies; Genistein; Hawaii; Humans; Isoflavones; Male; Phytoestrogens; Prostatic Neoplasms

Chemical compounds, including plant-based phytoestrogens, can function as hormone mimics and alter endocrine signaling in wildlife. In the present study, the waste streams from 19 plant-processing industries, including biofuel manufacturers, were sampled and analyzed for the phytoestrogens genistein, daidzein, coumestrol, formononetin, biochanin A, and zearalenone, via liquid chromatography/mass spectrometry. Eight of these industries contained phytoestrogens at environmentally relevant levels (≥1,000 ng/L), with the highest at approximately 250,000 ng/L. The influent and effluent streams of three municipal wastewater treatment plants receiving flow from some of these industries also were sampled and analyzed for the same phytoestrogens. It appeared that aerobic biological treatment, such as activated sludge, was able to remove these compounds from the liquid stream. Nevertheless, the effluent stream from one of the wastewater treatment plants had a phytoestrogen concentration above 1,000 ng/L. Results of the present study indicate the need for caution when designing facilities to treat the effluents from biofuel and other plant-processing industries.

Topics: Biofuels; Chromatography, Liquid; Coumestrol; Environmental Monitoring; Genistein; Industrial Waste; Isoflavones; Mass Spectrometry; Phytoestrogens; Sewage; Water Pollutants, Chemical

The aims of the study were to compare the in vitro effects of daidzein or 17beta-estradiol (E(2)) on: 1) progesterone (P(4)) secretion by luteinized granulosa cells harvested from large porcine follicles, as well as 2) estrogen receptor alpha and beta (ERalpha and ERbeta) mRNA and protein expression in the cells. In addition, the effect of daidzein on E(2) secretion and viability of the granulosa cells was examined. We found that basal and gonadotropin-stimulated P(4) secretion were inhibited in granulosa cells cultured in the presence of daidzein either for 24 or 48 hours. In contrast to daidzein, E(2) reduced P(4) secretion only during 24-hour cell cultures increasing it during longer cultures. Daidzein did not affect E(2) secretion by granulosa cells. The expression of ERalpha and ERbeta mRNA, as well as ERbeta protein, was up-regulated by daidzein but unaffected by E(2). To conclude, the soy estrogen daidzein acts directly on the porcine ovary to decrease progesterone production and to increase expression of ERbeta mRNA and protein. Daidzein actions in porcine luteinized granulosa cells differ from those of estradiol and it may suggest disadvantageous effects of the phytoestrogen on reproductive processes in females.

Topics: Animals; Cell Survival; Cells, Cultured; Dose-Response Relationship, Drug; Estradiol; Estrogen Receptor alpha; Estrogen Receptor beta; Female; Gene Expression; In Vitro Techniques; Isoflavones; Luteal Cells; Ovarian Follicle; Phytoestrogens; Progesterone; Receptors, Estrogen; RNA, Messenger; Swine

Several studies have demonstrated a protective effect of estrogen against the risk of developing neurodegenerative diseases; however, the molecular mechanisms involved have not been fully addressed. Isoflavones have been proposed as potential alternatives to estrogen replacement therapy. Therefore, in the present study, we investigated effects of isoflavones on cell death and tau phosphorylation in SH-SY5Y human neuroblastoma cells. Cells were treated with tunicamycin (TM) to induce endoplasmic reticulum (ER) stress-mediated toxicity, which is involved in development of neurodegenerative diseases. Treatment of cells with either 17beta-estradiol or isoflavones (either genistein or daidzein) significantly protected cells against cell death. The protective effect against cell death was blocked by a specific estrogen receptor blocker, ICI 182,780, suggesting that isoflavones protect against cell death via estrogen receptor-dependent pathways. Isoflavones also suppressed ER stress as determined by decreased expressions of the immunoglobulin binding protein (BiP) mRNA, spliced X-box binding protein-1 (Xbp-1) mRNAs, and C/EBP homologous protein (CHOP). TM activated glycogen synthase kinase 3beta (GSK3beta), a kinase involved in tau phosphorylation; in contrast, isoflavones inactivated GSK3beta and decreased tau hyperphosphorylation. In conclusion, our results clearly demonstrate that isoflavones prevent ER stress-mediated neurotoxicity by inhibiting tau hyperphosphorylation in SH-SY5Y cells.

Topics: Cell Death; Cell Line, Tumor; Endoplasmic Reticulum; Estradiol; Female; Genistein; Glycine max; Glycogen Synthase Kinase 3; Humans; Isoflavones; Nerve Tissue Proteins; Neuroblastoma; Neurons; Neuroprotective Agents; Phosphorylation; Phytoestrogens; Plant Extracts; Receptors, Estrogen; tau Proteins; Transcription Factor CHOP; Tunicamycin

Prostate cancer (PCa) has a high propensity to metastasize to the bone. PCa cells produce several bone-related factors, namely parathyroid hormone related protein (PTHrP), its PTH type 1 receptor (PTH1R), osteoprotegerin (OPG), and receptor activator of NF-kappa B ligand (RANKL). The effects of these factors might explain, at least in part, the ability of PCa cells to grow in and interact with bone.. We first analyzed the expression of the aforementioned factors (by western blot and flow cytometry), and their modulation by the phytoestrogens genistein and daidzein (as potential anti-tumoral agents), in human PCa cells in vitro. We also assessed the impact of these osteomimetic factors on PCa cell viability (by propidium iodide staining and flow cytometry, and trypan blue staining).. Genistein and daidzein, at nM range, increased both the PTHrP/PTH1R system and the OPG/RANKL protein ratio, while genistein and, to a lesser extent, daidzein, at >microM doses, inhibited cell viability in PCa cells. Both N- and C-terminal domains of PTHrP inhibited genistein-induced cell death by modulating transcription factor Runx-2 and the Bcl-2/Bax protein ratio in PCa cells.. Our findings indicate that high doses of genistein and daidzein cause PCa cell death. On the other hand, low doses of these phytoestrogens induce some osteomimetic features in PCa cells with putative impact on PCa development.

Topics: Blotting, Western; Bone and Bones; Cell Death; Cell Line, Tumor; Cell Survival; Cytokines; Flow Cytometry; Genistein; Humans; Isoflavones; Male; Osteoprotegerin; Parathyroid Hormone-Related Protein; Phytoestrogens; Prostatic Neoplasms; RANK Ligand; Receptor, Parathyroid Hormone, Type 1; Reverse Transcriptase Polymerase Chain Reaction; RNA, Neoplasm; Transfection

Antiangiogenic compounds are gaining more and more interest as a new approach in the prevention and treatment of cancer and inflammatory diseases. The objective of this study was the evaluation of the antiangiogenic effect of a red clover extract (RCE) used in food supplements for menopausal complaints as well as of its main isoflavones in an in vivo system, the chorioallantoic membrane assay of fertilized hen's eggs. At a dosage of 250 microg/pellet the red clover extract showed excellent inhibition of angiogenesis. The antiangiogenic activity of the non-methylated isoflavones daidzein and genistein was higher than that of the methylated compounds formononentin and biochanin A. The results demonstrate that RCE is not only suitable for menopausal complaints, but might also be a powerful chemopreventive agent against chronic diseases e.g. which have a high incidence especially in elderly female.

Topics: Angiogenesis Inhibitors; Animals; Chick Embryo; Chorioallantoic Membrane; Genistein; Inflammation; Isoflavones; Methylation; Phytoestrogens; Plant Extracts; Trifolium

The microbial metabolism of dietary phytoestrogens varies considerably among individuals and influences the final exposure to bioactive compounds. In view of the increasing number of food supplements combining several classes of phytoestrogens, the microbial potential to activate various proestrogens within an individual was evaluated in 3 randomized dietary crossovers. Treatment allocation was based on participants' eligibility (>45% in vitro bioactivation of >or=2 separate proestrogens by fecal cultures; n = 40/100). After a run-in of >or=4 d, participants were given soy-, hop-, and/or flax-based food supplements dosed either separately (SOY: 2.83 mg daidzein aglycone equivalents/supplement, HOP: 1.20 mg isoxanthohumol (IX)/supplement, or FLAX: 2.08 mg secoisolariciresinol (SECO) aglycone equivalents/supplement; reference intervention) or simultaneously (MIX; test intervention) 3 times/d for 5 d, followed by a wash-out period (>or=7 d) and the second intervention. Before and after each (co)supplementation, spot urine and serum were collected. In total, 22 equol, 19 8-prenylnaringenin (8-PN), and 21 enterolactone (ENL) producers completed the SOY+MIX, HOP+MIX, and FLAX+MIX trials, respectively. The microbial bioactivation of daidzein, IX, and SECO, generally decreased upon coincubation in vitro (equol: 4.4%, P = 0.164; 8-PN: 20.5%, P < 0.001; ENL: 44.3%, P < 0.001) and cosupplementation in vivo (equol: 28.3%, P = 0.009; 8-PN: 35.4%, P = 0.107; ENL: 35.9%, P = 0.003). Although the bioavailabilities of total isoflavones, prenylflavonoids, and lignans were not significantly affected upon coadministration, participants were exposed to lower phytoestrogen-derived 17beta-estradiol equivalents. In conclusion, the bioavailability of phytoestrogens, especially when given in mixtures, is subject to high interindividual variation. These findings support the importance of personalized screening when assessing the efficacy of such products and mixtures.

Topics: Dietary Supplements; Equol; Estradiol; Feces; Flavanones; Flavonoids; Genistein; Humans; Isoflavones; Lignans; Phytoestrogens

Soy phytoestrogen is often used as hormone replacement therapy to alleviate the symptoms of menopause in postmenopausal women. Since estrogen has been considered as an important risk factor for the development of breast carcinoma, we need to know whether it is safe for these postmenopausal women with breast cancer to take soy foods that are rich in phytoestrogen. In the present study, we investigated the efficacy of soy phytoestrogen on tumor proliferation, apoptosis, and angiogenesis in mammary tumors that had already formed in ovariectomized rats. We found that soy phytochemical extraction inhibited proliferation and induced apoptosis in vitro and in vivo, and it demonstrated better antitumor effects than single phytoestrogen. Soy phytochemical extraction also produced surprisingly good antiangiogenic effects, which were evidenced by lower microvascular density, reduced plasma vascular endothelial growth factor, and increased plasma endostatin levels. Our findings suggest that soy phytochemical extraction exerts significant antitumor and antiangiogenic activity in a postmenopausal animal model with breast cancer.

Topics: Angiogenesis Inducing Agents; Angiogenesis Inhibitors; Animals; Antineoplastic Agents; Apoptosis; Cell Line, Tumor; Cell Proliferation; Cells, Cultured; Dose-Response Relationship, Drug; Endothelial Cells; Female; Genistein; Glycine max; Humans; Isoflavones; Mammary Neoplasms, Experimental; Microvessels; Phytoestrogens; Phytotherapy; Plant Extracts; Random Allocation; Rats; Rats, Sprague-Dawley; Tumor Burden

Exposure to xenoestrogens in humans and animals has gained increasing attention due to the effects of these compounds on reproduction. The present study was undertaken to investigate the influence of low-dose dietary phytoestrogen exposure, i.e. a mixture of genistein, daidzein, biochanin A and formononetin, on the establishment of testosterone production during puberty in male goat kids.. Goat kids at the age of 3 months received either a standard diet or a diet supplemented with phytoestrogens (3-4 mg/kg/day) for approximately 3 months. Plasma testosterone and total and free triiodothyronine (T3) concentrations were determined weekly. Testicular levels of testosterone and cAMP were measured at the end of the experiment. Repeated measurement analysis of variance using the MIXED procedure on the generated averages, according to the Statistical Analysis System program package (Release 6.12, 1996, SAS Institute Inc., Cary, NC, USA) was carried out.. No significant difference in plasma testosterone concentration between the groups was detected during the first 7 weeks. However, at the age of 5 months (i.e. October 1, week 8) phytoestrogen-treated animals showed significantly higher testosterone concentrations than control animals (37.5 nmol/l vs 19.1 nmol/l). This elevation was preceded by a rise in plasma total T3 that occurred on September 17 (week 6). A slightly higher concentration of free T3 was detected in the phytoestrogen group at the same time point, but it was not until October 8 and 15 (week 9 and 10) that a significant difference was found between the groups. At the termination of the experiment, testicular cAMP levels were significantly lower in goats fed a phytoestrogen-supplemented diet. Phytoestrogen-fed animals also had lower plasma and testicular testosterone concentrations, but these differences were not statistically significant.. Our findings suggest that phytoestrogens can stimulate testosterone synthesis during puberty in male goats by increasing the secretion of T3; a hormone known to stimulate Leydig cell steroidogenesis. It is possible that feedback signalling underlies the tendency towards decreased steroid production at the end of the experiment.

Topics: Animals; Cyclic AMP; Diet; Genistein; Goats; Isoflavones; Male; Phytoestrogens; Sexual Maturation; Testis; Testosterone; Triiodothyronine

To investigate the intestinal absorption and metabolism of daidzein in Caco-2 cell model.. The damage of daidzein to Caco-2 cell was evaluated by MTT value and alkaline phosphatase (ALP) and lactate dehydrogenase (LDH) activity. With the reference to propranolol, the apparent permeability coefficient (Papp) of daidzein was measured through the monolayer under the different concentrations and pH media. The metabolites were also measured by enzyme hydrolysis.. Daidzein had no damage to the growth of the Caco-2 cells in the concentration of 1-50 microg/mL. The apparent permeability coefficient (Papp) of daidzein across the monolayer showed concentration- and pH- independent manner, which was similar to that of transcellcular marker-propranolol, suggesting the good absorption of daidzein in vivo. By hydrolysis with beta-glucuronidase, low metabolites were detected in monolayer and transport medium, verifying the existence of glucuronides and sulfates.. The daidzein absorption in Caco-2 cell model is a passive and transcellular transport and quite good in the intestines.

Topics: Absorption; Biological Transport; Caco-2 Cells; Cell Membrane Permeability; Glycine max; Humans; Hydrogen-Ion Concentration; Isoflavones; Models, Biological; Phytoestrogens; Plants, Medicinal

Clinical observations have suggested a relationship between osteoarthritis and a changed estrogen metabolism in menopausal women. Phytoestrogens have been shown to ameliorate various menopausal symptoms. Proteoglycans (PG) consisting of low and high sulfated glycosaminoglycans (GAG) are the main components of articular cartilage matrix, and their synthesis is increased by insulin in growth plate cartilage. We have investigated whether GAG synthesis and sodium [35S]sulfate incorporation in female bovine articular chondrocytes are affected by daidzein, genistein, and/or insulin. For comparative purposes, estradiol incubations were performed. Articular chondrocytes were cultured in monolayers at 5% O2 and 5% CO2 in medium containing serum for 7 days followed by the addition of 10(-11) M-10(-4) M daidzein, genistein, 17beta-estradiol, or 5 microg/ml insulin in a serum-free culture phase of 2 days. Photometrically analyzed GAG synthesis was significantly suppressed by high doses (10(-5) M-10(-4) M) of daidzein, genistein, and 17beta-estradiol. Although insulin raised the sodium [35S]sulfate uptake significantly, different concentrations of daidzein, genistein, or 17beta-estradiol showed no significant effects. However, the stimulating effect of insulin on sulfate incorporation was enhanced significantly after preincubation of cells with 10(-11) M-10(-5) M daidzein or 10(-9) M-10(-5) M genistein but not by 17beta-estradiol. In view of the risks of long-term estrogen replacement therapy, further experiments should clarify the potential benefit of phytoestrogens and insulin in articular cartilage metabolism.

Topics: Animals; Cartilage, Articular; Cattle; Cells, Cultured; Chondrocytes; Culture Media, Serum-Free; Dose-Response Relationship, Drug; Female; Genistein; Hypoglycemic Agents; Insulin; Isoflavones; Phytoestrogens; Sulfuric Acid Esters; Time Factors

We report here the effects of estrogens and phytoestrogens on catecholamine signaling in cultured bovine adrenal medullary cells used as a model of catecholaminergic neurons in the brain. Treatment of the cells for 20 min with 17beta-estradiol (E(2)) (0.3-100 nM) or phytoestrogens such as daidzein (0.01-1.0 microM), a soy isoflavone, and resveratrol (0.1-1.0 microM), a grape polyphenol, stimulated (14)C-catecholamine synthesis from [(14)C]tyrosine, which was associated with the activation of tyrosine hydroxylase. The stimulatory effect of E(2) and phytoestrogens was not inhibited by ICI182,780, a nuclear estrogen receptor inhibitor, but abolished by U0126, an inhibitor of extracellular signal-regulated kinase1/2 (ERK1/2) kinase. E(2) enhanced the phosphorylation of ERK1/2. The plasma membrane isolated from the adrenal medulla showed two classes of specific binding sites of [(3)H]E(2). Resveratrol and daidzein at high concentrations (> or =1.0 microM) inhibited catecholamine secretion induced by various secretagogues. The present findings suggest that estrogens and phytoestrogens most likely stimulate catecholamine synthesis via estrogen receptors in the plasma membrane, but in high concentrations phytoestrogens inhibit catecholamine secretion induced by secretagogues in adrenal medullary cells, and probably in brain neurons.

Topics: Animals; Cardiovascular System; Catecholamines; Cattle; Cell Membrane; Cells, Cultured; Dose-Response Relationship, Drug; Estradiol; Intracellular Space; Isoflavones; Models, Biological; Phytoestrogens; Protective Agents; Receptors, Estrogen; Resveratrol; Signal Transduction; Stilbenes

Since soy isoflavones may influence the thyroid hormone feedback system by interference with their biosynthesis, secretion and metabolism, we tested whether their controlled shortterm consumption affects thyroid function.. Eighty six volunteers--university students (32 males and 54 females) were eating unprocessed boiled natural soybeans (2 g/kg body weight/day) for 7 consecutive days. Thyrotropin, free thyroid hormones, antibodies to thyroid peroxidase and to thyroglobulin, and actual levels of unconjugated major soy phytoestrogens, daidzein and genistein, were measured in sera collected before, at the end and one week after finishing soy meal consumption.. Both phytoestrogens increased significantly (p<0.0001) at the end of soy-diet and fell down after its termination nearly back to the initial values. No significant changes were found in female group, while in males a significant transitory increase of thyrotropin (p<0.0001) was recorded. When actual levels of phytoestrogens were related to thyroid parameters, the only significant correlations were found between basal levels of daidzein and thyrotropin, daidzein and antithyroglobulin at the end of soy consumption in males, and between daidzein and free thyroxine at the end of the soy ingestion in females.. Though only modest and transitory effects on thyroid parameters occurred after controlled short-term soy consumption, some actual thyroid hormone parameters do correlate with actual isoflavone levels.

Topics: Adolescent; Adult; Autoantibodies; Diet; Female; Genistein; Glycine max; Humans; Iodide Peroxidase; Isoflavones; Male; Phytoestrogens; Thyroglobulin; Thyroid Hormones; Thyrotropin; Thyroxine; Triiodothyronine

Daidzein is a very good candidate for treating cardio-cerebrovascular diseases, but its poor oral absorption and bioavailability limit its curative efficacy. In this work, daidzein-loaded solid lipid nanoparticles (SLNs) with PEGylated phospholipid as stabilizer were successfully prepared by hot homogenization method. SLNs showed the mean particle size 126+/-14 nm with entrapment efficiency 82.5+/-3.7%. In vitro release of SLNs demonstrated a sustained release manner with cumulative release over 90% within 120 h in bovine serum albumin solution (4%, w/v). The pharmacokinetic behavior showed that SLNs loading daidzein could significantly increase circulation time compared with orally administrated daidzein suspension or intravenously delivered daidzein solution. SLNs showed the better effect on cardiovascular system of the anesthetic dogs by reducing the myocardial oxygen consumption (MOC) and the coronary resistance (CR) in heart compared with oral suspension or intravenous solution. The SLNs demonstrated the best effect on cerebrovascular system by increasing cerebral blood flow (CeBF) and reducing cerebrovascular resistance (CeR) in anesthetized dogs, and the protective effect on rats with ischemia-reperfusion injury model among three formulations. These results suggested that SLNs could be a potential candidate for the treatment of cardio-cerebrovascular diseases.

Topics: Animals; Blood Flow Velocity; Brain Ischemia; Dogs; Drug Carriers; Isoflavones; Lipids; Male; Nanoparticles; Phytoestrogens; Rats; Rats, Sprague-Dawley

Data regarding convenient, valid methods for measuring U.S. isoflavone intake are limited. We evaluated a soy food questionnaire (SFQ), the Willett food frequency questionnaire (FFQ), and overnight urine samples relative to excretion in 24-h urine samples. We also described intake among women in a high-risk program for breast or ovarian cancer. Between April 2002 and June 2003, 451 women aged 30 to 50 yr with a family history of breast or ovarian cancer completed the SFQ and FFQ. Of them, 27 provided four 24-h and overnight urine specimens. In these women, 24-h sample measures were correlated with SFQ estimates of daidzein (Spearman r = .48) and genistein (r = .54) intake, moderately correlated with the Willett FFQ (daidzein r = .38, genistein r = .33), and strongly correlated with overnight urine excretion (daidzein r = .84, genistein r = 0.93). Among all 451 SFQ respondents, mean (median) daidzein and genistein intakes were 2.8 (0.24) and 3.9 (0.30) mg/day. Primary sources of both were soymilk, soy nuts, and tofu. We conclude that targeted soy food questionnaires, comprehensive FFQs, and multiple overnight urines are all reasonable options for assessing isoflavone intake in epidemiologic studies.

Topics: Adult; Biomarkers; Circadian Rhythm; Diet Records; Female; Genistein; Humans; Isoflavones; Middle Aged; Philadelphia; Phytoestrogens; Reproducibility of Results; Soy Foods; Surveys and Questionnaires; United States

The goal of this study was to evaluate how oral antibiotics (OABX) change the appearance of isoflavones (IFLs) in adults and children after soy consumption. The urinary IFL excretion rate (UIER) known to reflect circulating IFLs was hypothesized to change due to intestinal microflora changes by OABX. Subjects provided urine collections in pairs of a baseline urine and an overnight urine before and after consuming soy nuts first during OABX treatment and then again when healthy. During OABX versus when healthy, UIER (nmol/h/kg) in adults (n = 12) was increased (P < 0.05) for daidzein (35.2 +/- 7.2 vs. 18.9 +/- 2.4), daidzein + genistein + glycitein [nonmetabolites (NM); 42.6 +/- 8.0 vs. 23.6 +/- 2.9), and total isoflavonoids (Total IFLs; daidzein + genistein + glycitein + dihydrodaidzein + dihydrogenistein + equol + O-desmethylangolensin) (51.5 +/- 10.3 versus 29.6 +/- 4.7). In contrast, children (n = 7) showed reduced UIER (P < 0.05) when on OABX versus when healthy for daidzein (36.3 +/- 6.4 vs. 46.8 +/- 4.7), dihydrodaidzein (1.2 +/- 0.6 vs. 3.0 +/-1.1), NM (46.3 +/- 8.2 vs. 59.5 +/- 6.0), dihydrodaidzein + dihydrogenistein + equol + O-desmethylangolensin (1.0 +/- 0.8 vs. 4.3 +/- 1.3), and Total IFLs (48.2 +/- 8.5 vs. 63.8 +/- 6.4).

Topics: Administration, Oral; Adolescent; Adult; Age Factors; Anti-Bacterial Agents; Biomarkers; Child; Child, Preschool; Female; Follow-Up Studies; Genistein; Glycine max; Humans; Isoflavones; Male; Phytoestrogens; Soy Foods

To investigate the preventive effect of the phytoestrogen daidzein on prostatic hyperplasia induced by testosterone in rats.. Thirty-six male adult Sprague-Dawley rats were equally randomized into 6 groups: Group I and II (normal control and model, treated with 1 ml distilled water by oral gavage), and Group III-VI (low-, medium- and high-dose daidzein and positive control, respectively given daidzein at 2, 20 and 100 mg/kg and diethylstilbestrol at 0.1 mg/kg once a day for 90 days). From the 91st day , Group III-VI were treated with subcutaneous injection of testosterone at 7.5 mg/kg/d for 10 days to induce prostatic hyperplasia. The wet weight and the index of the prostate were obtained, its morphological changes detected and the changes of ERalphaa and ERbeta expressions in the prostate observed by immunohistochemistry.. Compared with the model group, the wet weight and the index of the prostate were significantly reduced in the 3 daidzein groups and the positive control group (P < 0.05 or P < 0.01). Medium- and high-dose daidzein induced a more obvious alleviation of prostate hyperplasia, characterized by thinner epithelia, decreased secretions in the glandular cavity and reduced interstitial tissues. The expression of ERalpha showed no significant difference between the model group and the other groups, while that of ERbeta was markedly decreased in the daidzein-treated groups as compared with the normal control or the model group.. The phytoestrogen daidzein has some preventive effect on prostatic hyperplasia induced by testosterone in rats.

Topics: Animals; Disease Models, Animal; Dose-Response Relationship, Drug; Isoflavones; Male; Phytoestrogens; Prostate; Prostatic Hyperplasia; Random Allocation; Rats; Rats, Sprague-Dawley; Testosterone

The HPLC system separated completely isofavonoids such as daidzein (15.2 min) and genistein (17.3 min). Initially, the concentrations of major isoflavone Genistein and Daidzein in the tested soy milk were determined. Commercial soy milk samples were analyzed for isoflavones and two major isoflavones detected: genistein 25.86 (mg L(-1)) +/- 0.66 SD and daidzein 8.25 (mg L(-1)) +/- 1.13 SD. Concentrations of genistein in soy milk were higher than daidzein. The results obtained in this study can serve as a basis for estimating amount of soy milk can be consumed by people as related to its main isoflavone content.

Topics: Chromatography, High Pressure Liquid; Genistein; Iran; Isoflavones; Phytoestrogens; Soy Milk

Studies have shown that soya consumption has been associated with low incidence of CVD. Because the chemical structures of soya isoflavones are similar to oestrogen, the beneficial outcome may be attributed to the oestrogenicity of these compounds. In this study, effect of the soya isoflavone genistein on the mRNA expression of apoA-1 in the human hepatoma HepG2 cell was investigated. Without oestrogen receptor (ER) alpha transfection, soya isoflavones in the physiological range had no effect on the apoA-1 transcription. Once ERalpha was ectopically expressed in these cells, soya isoflavone dramatically increased the apoA-1 mRNA abundance quantified by real-time PCR. ApoA-1-reporter assays with plasmid constructed from the 5'-flanking segment upstream to the coding region revealed that the transactivation of the apoA-1 promoter was induced by the soya isoflavone in HepG2 cells expressing ERalpha. This induction was reduced by the anti-oestrogen ICI 182780, but not the inhibitors of protein kinase (PK) C, PKA, or mitogen-activated PK. Based on the previously identified response elements on the promoter, a series of truncated promoter reporter plasmids were then constructed. An induction profile of genistein was built and insulin response core element at -411 to -404 appeared to be a potential site of interaction. This study illustrated that soya isoflavones at physiological concentrations could up regulate apoA-1 mRNA expression in ERalpha-transfected HepG2 cells.

Topics: Apolipoprotein A-I; Dose-Response Relationship, Drug; Estradiol; Estrogen Antagonists; Estrogen Receptor alpha; Fulvestrant; Genistein; Humans; Isoflavones; Phytoestrogens; Promoter Regions, Genetic; Protein Kinase Inhibitors; RNA, Messenger; Transcriptional Activation; Tumor Cells, Cultured; Up-Regulation

Soy-based formulas are consumed by growing numbers of infants and used as regular food supplements in livestock production. Moreover, constituent dietary phytoestrogens may compete with endogenous estrogens and affect individual growth. This study aimed to investigate the in vitro effects of isoflavones in comparison with estrogens on the proliferation of porcine satellite cells derived from neonatal muscle. After 7 h of exposure in serum-free medium, 17beta-estradiol (1 nM, 1 microM), estrone (1 microM), and daidzein (1, 100 microM) slightly decreased whereas 100 microM genistein substantially lowered DNA synthesis. Declines in DNA amount were observed with genistein (1, 100 microM) and daidzein (100 microM). After 26 h of exposure, 100 microM genistein reduced DNA synthesis, whereas it was increased by 10 microM genistein and 10 and 100 microM daidzein. In the case of 10 microM genistein and 100 microM daidzein, these increases apparently resulted from the repair of damaged DNA. Genistein and daidzein (100 microM) reduced protein synthesis, caused a G2/M phase block, and decreased DNA amount in association with higher rates of cell death partially resulting from apoptosis. Conclusively, isoflavones at concentrations of greater than 1 muM act as inhibitors of porcine skeletal muscle cell proliferation.

Topics: Animals; Animals, Newborn; Apoptosis; Cell Cycle; Cell Proliferation; Cells, Cultured; DNA Damage; DNA Replication; Dose-Response Relationship, Drug; Estradiol; Estrone; Genistein; Isoflavones; Myoblasts, Skeletal; Phytoestrogens; Protein Biosynthesis; Swine; Time Factors

Ovariectomy of immature female rats, results in significant decrease of trabecular bone volume and in cortical bone thickness. Previously, we found that estradiol-17beta (E(2)) restored bone structure of ovariectomized (Ovx) female rats to values obtained in intact sham-operated female rats. E(2) also selectively stimulated creatine kinase (CK) specific activity a hormonal-genomic activity marker. In the present study, we compared the effects of E(2) and the phytoestrogens: daidzein (D), biochainin A (BA), genistein (G), carboxy-derivative of BA (cBA), and the SERM raloxifene (Ral) in Ovx, on both histological changes of bones and CK, when administered in multiple daily injections for 2.5 months. Bone from Ovx rats, showed significant disrupted architecture of the growth plate, with fewer proliferative cells and less chondroblasts. The metaphysis underneath the growth plate, contained less trabeculae but a significant increased number of adipocytes in the bone marrow. D like E(2) and Ral but not G, BA, or cBA, restored the morphology of the tibiae, similar to that of control sham-operated animals; the bony trabeculeae observed in the primary spongiosa was thicker, with almost no adipocytes in bone marrow. Ovariectomy resulted also in reduced CK, which in both epiphysis and diaphysis was stimulated by all estrogenic compounds tested. In summary, only D stimulated skeletal tissues growth and differentiation as effectively as E(2) or Ral, suggesting that under our experimental conditions, D is more effective in reversing menopausal changes than any of the other isolated phytoestrogens which cannot be considered as one entity.

Topics: Adipocytes; Animals; Bone and Bones; Bone Marrow; Creatine Kinase; Estradiol; Female; Genistein; Growth Plate; Isoflavones; Ovariectomy; Phytoestrogens; Raloxifene Hydrochloride; Rats; Rats, Wistar; Tibia; Trabecular Meshwork

We aim to study the mechanisms underlying the neurotrophic effect of daidzein (Dz) in hippocampal neurons. Dz-enhanced axonal outgrowths manifested growth cone formation and increased immunostaining intensity of growth-associated protein 43 (GAP-43) in growth cones. Consistent with this, Dz increased GAP-43 phosphorylation and its membrane translocation without affecting total GAP-43 levels. In the presence of Dz, significant increase in the immunoreactivity for estrogen receptor (ER) beta, but not ERalpha, was observed on the membrane of cell bodies and growing axons. Dz also induced the activation of protein kinase C alpha (PKCalpha), which was inhibited by the ICI182,780 pretreatment. Similarly, Dz-promoted axonal elongation was blocked by ICI182,780 and Gö6976. Moreover, Dz-stimulated activation of GAP-43 was specifically abolished by Gö6976, suggesting PKCalpha being the upstream effector of GAP-43. Taken together, our data suggest that Dz triggers an ERbeta/PKCalpha/GAP-43 signaling cascade to promote axonal outgrowths in cultured hippocampal neurons.

Topics: Animals; Axons; Cell Enlargement; Cells, Cultured; Dose-Response Relationship, Drug; Hippocampus; Isoflavones; Nerve Tissue Proteins; Neurons; Phytoestrogens; Rats; Rats, Wistar; Signal Transduction

Red clover, known for its estrogenic activity due to its isoflavones content (biochanin A, genistein, daidzein and formononetin), was inoculated with the arbuscular mycorrhizal fungus Glomus mosseae. Once the symbiotic fungus was well established, plants were harvested and we determined the root and shoot dry weight as well as the P-content. In roots and leaves the levels of biochanin A, genistein, daidzein and formononetin were quantified by reversed-phase HPLC and the estrogenic activity of the leaves was measured by a transactivation assay using a yeast two-plasmid system. Mycorrhization increased the levels of biochanin A in the root and the shoot and reduced the levels of genistein in the shoot of red clover. The levels of the other isoflavones were not affected. The shoot biomass of mycorrhizal plants more than doubled compared with non-mycorrhizal control plants, and this growth-stimulating effect of arbuscular mycorrhiza did not affect the estrogenic activity of red clover. In a control P treatment, the biomass of red clover was greatly enhanced. However, the estrogenic activity was reduced. These results suggest that, in contrast to an enhanced shoot biomass production after P application with a reduced estrogenic activity, with arbuscular mycorrhiza the shoot biomass of red clover can be enhanced without a negative effect on estrogenic activity.

Topics: Biomass; Genistein; Glomeromycota; Isoflavones; Mycorrhizae; Phosphates; Phytoestrogens; Plant Leaves; Plant Roots; Trifolium

Only 30% to 50% of people produce the daidzein-metabolite equol after eating soy. We conducted a cross-sectional study of the associations between equol status, intake of soy foods, and mammographic density in a sample of postmenopausal women recruited at a radiology clinic near Buffalo, New York. Participants were 48 to 82 years old, had no history of cancer or breast reduction/augmentation, and no recent use of antibiotics or hormones. Percent density was measured by computer-assisted analysis of digitized images of craniocaudal films. Equol status was assessed using a soy-challenge protocol and usual soy intake by questionnaire. General linear models were used to assess independent and joint effects of equol status and intake of soy on multivariate adjusted percent density (covariates included age, body mass index, parity, age at first birth, and ever use of combined hormone therapy). Of 325 enrolled, 232 (71%) participants completed study assessments and are included in the present analysis. Mean percent density was 34% (+/-18%). Seventy-five (30%) participants were producers of equol. Forty-three (19%) participants reported regularly eating >1 soy food or supplement/wk. There were no significant independent associations of equol status or soy intake with percent density, but the interaction between these factors was significant (P < 0.01). Among equol producers, those with weekly soy intake had lower percent density (30.7% in weekly consumers of soy versus 38.9% in others; P = 0.08); among nonproducers, weekly soy intake was associated with higher percent density (37.5% in weekly soy consumers versus 30.7% in others; P = 0.03). Results suggest that equol producers and nonproducers may experience different effects of dietary soy on breast tissue.

Topics: Aged; Aged, 80 and over; Body Mass Index; Breast; Chromatography, Gas; Chromatography, High Pressure Liquid; Cross-Sectional Studies; Diet; Equol; Female; Humans; Isoflavones; Mammography; Middle Aged; Phytoestrogens; Postmenopause; Soy Foods; Soybean Proteins; Surveys and Questionnaires

Based on the hypothesis that isoflavones are absorbed more efficiently from fermented than from non-fermented soya foods, we compared the urinary isoflavonoid excretion (UIE) after intake of miso soup or soya milk. We recruited twenty-one women with Japanese ancestry who consumed standardized soya portions containing 48 mg isoflavones. On day 1, half the women consumed soya milk, the other half started with miso soup. On day 3, the subjects ate the other soya food and on day 5, they repeated the first food. Each participant collected a spot urine sample before and an overnight urine sample after soya food intake. All urine samples were analysed for daidzein, genistein and equol using LC-MS and were expressed as nmol/mg creatinine. We applied mixed models to evaluate the difference in UIE by food while including the baseline values and covariates. Relative to baseline, both groups experienced significantly higher UIE after consuming any of the soya foods. We observed no significant difference in UIE when soya milk was compared to miso soup (P = 0.87) among all women or in the seven equol producers (P = 0.88). Repeated intake of the same food on different days showed high reproducibility within subjects. These preliminary results indicate similar UIE after consuming a fermented soya food (miso) as compared to a non-fermented soya food (soya milk). Therefore, recommendations favouring fermented soya foods are not justified as long as the intestinal microflora is capable of hydrolysing the isoflavone glucosides from non-fermented soya foods.

Topics: Adult; Aged; Aged, 80 and over; Asian; Equol; Female; Fermentation; Genistein; Humans; Isoflavones; Middle Aged; Nutritional Physiological Phenomena; Phytoestrogens; Soy Foods; Soy Milk; Specimen Handling

The nutritional intake of phytoestrogens seems to reduce the risk of breast cancer or other neoplastic diseases. However, these epidemiological findings remain controversial because low doses of phytoestrogens, achievable through soy-rich diets, stimulate the proliferation of estrogen-sensitive tumor cells. The question of whether such phytochemicals prevent cancer or rather pose additional health hazards prompted us to examine global gene expression programs induced by a typical soy product. After extraction from soymilk, phytoestrogens were deconjugated and processed through reverse- and normal-phase cartridges. The resulting mixture was used to treat human target cells that represent a common model system for mammary tumorigenesis. Analysis of mRNA on high-density microarrays revealed that soy phytoestrogens induce a genomic fingerprint that is indistinguishable from the transcriptional effects of the endogenous hormone 17beta-estradiol. Highly congruent responses were also observed by comparing the physiologic estradiol with daidzein, coumestrol, enterolactone, or resveratrol, each representing distinct phytoestrogen structures. More diverging transcriptional profiles were generated when an inducible promoter was used to reconstitute the expression of estrogen receptor beta (ERbeta). Therefore, phytoestrogens appear to mitigate estrogenic signaling in the presence of both ER subtypes but, in late-stage cancer cells lacking ERbeta, these phytochemicals contribute to a tumor-promoting transcriptional signature.

Topics: 4-Butyrolactone; Animals; Biomarkers, Tumor; Breast Neoplasms; Cattle; Estradiol; Estrogen Receptor beta; Female; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Genistein; Humans; Isoflavones; Lignans; Milk; Neoplasm Proteins; Oligonucleotide Array Sequence Analysis; Phytoestrogens; Promoter Regions, Genetic; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Soy Milk; Tumor Cells, Cultured

There is evidence that metabolic activation can increase the estrogenic activity of the phytoestrogen-rich herb in tests with HepG2 cells. Variation in both plant genetics and harvest season may also influence estrogenic activity of the plant materials. We evaluated the influence of in vitro metabolic activation by S9 mixture on the estrogenic activity of tuberous samples of different cultivars of the phytoestrogen-rich herb, Pueraria mirifica, harvested in different seasons.. Plant extracts were derived from the tubers of five plant cultivars collected during summer, rainy season and winter and administered to MCF-7 cultures, an ERalpha-positive human mammary adenocarcinoma cell line for 3 days at dosages of 0.1, 1, 10, 100 and 1000microg/ml. These data were compared with the major plant isoflavonoids puerarin, daidzin, genistin, daidzein and genistein and with 17beta-estradiol, at concentrations of 10(-12) to 10(-6)M. The test system was done in the absence and presence of the S9 mixture.. The major plant isoflavonoids and the plant extracts exhibited variable degrees of estrogenic activities as evaluated by altered proliferation of the MCF-7 cell line which were significantly enhanced in the presence of the S9 mixture.. Metabolic activation of plant isoflavonoids at least in vitro by S9 mixture plays a significant role in amplification of the estrogenic activity of the phytoestrogen-rich plant. In addition, the estrogenic activities of the plant samples were potentially influenced by both seasonal changes and plant genetics.

Topics: Adenocarcinoma; Breast Neoplasms; Cell Line, Tumor; Cell Proliferation; Estradiol; Female; Genistein; Humans; Isoflavones; Phytoestrogens; Plant Extracts; Pueraria; Seasons

Daidzein is a soy isoflavone with estrogenic activity present in plant-based food items and health foods and used as an alternative therapy for cancer and cardiovascular diseases. The present study was designed to investigate the effects of daidzein on the cavernosal components, including smooth muscle cells, collagen fibers, and elastic fibers, that are the key structures fundamental for erection.. A total of 30 adult male Sprague-Dawley rats were equally divided into a normal control group, three experimental groups, and one positive control group. The three experimental groups were given daidzein at a dose of 2, 20, and 100 mg/kg body weight daily, and the positive control group received 0.1 mg diethylstilbestrol per animal daily for 90 days. The collagen fibers and elastic fibers in the corpora cavernosa were measured using histochemical or immunohistochemical techniques, and their relative contents were evaluated quantitatively or semiquantitatively.. The relative content of collagen fibers in the corpus cavernosa in rats treated with low-dose daidzein (2 mg/kg) was not significantly different from that of controls, as was the case for the smooth muscle and elastic fiber content (all P >0.05). However, the relative content of the collagen fibers was significantly increased in rats treated with a medium dose (20 mg/kg) and a high dose (100 mg/kg) of daidzein. The smooth muscle cell and elastic fiber content was reduced significantly compared with that of the controls (all P <0.01). Similar alterations were observed in the diethylstilbestrol-treated rats.. These results suggest that daidzein, if ingested in a relatively large amount, could induce histopathologic alterations in the penile cavernosal structures characterized by an increase in the collagen content and a reduction in smooth muscle cell and elastic fiber content, which might be suggestive of erectile dysfunction.

Topics: Animals; Collagen; Diethylstilbestrol; Elastic Tissue; Estrogens, Non-Steroidal; Isoflavones; Male; Muscle, Smooth; Penis; Phytoestrogens; Rats; Rats, Sprague-Dawley

The neuroprotective action of oestrogens and oestrogen-like compounds is in the focus of basic and clinical research. Although such action has been shown to be associated with neuronal plasma membranes, the implication of G-proteins remains to be elucidated. This study revealed that micromolar concentrations (microM) of 17beta-oestradiol and phytoestrogens, genistein and daidzein, significantly (P < 0.05) stimulate G-proteins ([(35)S]GTP gamma S binding) in the post-mortem hippocampal membranes of age-matched control women with the respective maximum effects of 28, 20 and 15% at 10 microM. In the frontocortical membranes, the stimulation of G-proteins did not differ significantly from that in hippocampal membranes. Although in the hippocampus and frontal cortex of the Alzheimer's disease (AD) women's brain, 10 microM 17beta-oestradiol produced significantly (P < 0.05) lower stimulation of G-proteins than in the control regions, stimulation by phytoestrogens revealed no remarkable decline. 17beta-Oestradiol, genistein and daidzein revealed a selective effect on various G-proteins (G(alphas), G(alpha o), G(alpha i1) or G(alpha 11) plus G(beta 1 gamma 2)) expressed in Sf9 cells. At a concentration of 10 microM, 17beta-oestradiol suppressed the H(2)O(2) and homocysteine stimulated G-proteins in the frontocortical membranes of control women to a greater extent than phytoestrogens. In AD, the suppressing effect of each compound was lower than in the controls. In the cell-free systems, micromolar concentrations of phytoestrogens scavenged OH(*) and the 2.2-diphenyl-1-picrylhydrazyl free radical (DPPH(*)) more than 17beta-oestradiol did. In the frontocortical membranes of control women, the 20 microM 17beta-oestradiol stimulated adenylate cyclase with 20% maximal effect, whereas, in AD, the effect was insignificant. Genistein did not stimulate enzyme either in control or AD frontocortical membranes. Our data confirm that the agents stimulate G-proteins in control and AD women's brains, although 17beta-oestradiol and phytoestrogens have similarities and differences in this respect. We suggest that, besides the ER-dependent one, the ER-independent antioxidant mechanism is responsible for the oestrogen stimulation of G-proteins in the brain membranes. Both of these mechanisms could be involved in the neuroprotective signalling of oestrogens that contributes to their preventive/therapeutic action against postmenopausal neurological disorders.

Topics: Adenylyl Cyclases; Aged; Aged, 80 and over; Alzheimer Disease; Brain; Case-Control Studies; Cell Membrane; Cytoprotection; Estradiol; Female; Free Radical Scavengers; Genistein; GTP-Binding Proteins; Guanosine 5'-O-(3-Thiotriphosphate); Humans; Isoflavones; Male; Models, Biological; Phytoestrogens; Protein Binding

Natural phytoestrogens such as the isoflavones genistein and daidzein, and the flavones quercetin exhibit anti-cancer properties. This study was purpose to investigate the anti-proliferative effect of phytoestrogens on acute myeloid leukemia (AML) and chronic myeloid leukemia (CML) cells, and their synergistic antileukemic effect in combination with chemotherapeutic drugs. Optimal dosage of genistein, quercetin and in combination with chemicals for leukemia cells were determined by experiments. Cell viability, apoptosis induction and cell cycle arrest were detected by trypan blue staining, MTT assay, optical microscopy, flow cytometry (FCM). The schedule treatment of combination of genistein and chemicals was determined. The results showed that genistein exhibited a dose- and time-dependent inhibitory effect on cell proliferation in NB4 and HL-60 cells, induced apoptosis and cell cycle arrest in G2/M phase. Quercetin had evident inhibitory effect on the proliferation of K562 and K562/A cells. The combination of genistein and chemicals exerted a synergistic effect on cell growth inhibition. In conclusion, this study demonstrated the synergistic antileukemic effect of genistein with chemotherapeutic drugs on leukemic cells. This combination appears to be a new idea for the clinical novel treatment of leukemia.

Topics: Antineoplastic Agents; Apoptosis; Cell Proliferation; Drug Synergism; Genistein; HL-60 Cells; Humans; Isoflavones; Leukemia, Monocytic, Acute; Leukemia, Myeloid, Acute; Phytoestrogens; Quercetin

The soyabean isoflavones genistein and daidzein, which may protect against some cancers, cardiovascular disease and bone mineral loss, undergo substantial Phase 2 metabolism, predominantly glucuronidation. We observed a correlation between rates of metabolism of marker substrates of specific UGTs and rates of glucuronidation of genistein and daidzein in vitro by a panel of human liver microsomes, demonstrating that UGT1A1 and UGT1A9, but not UGT1A4, make a major contribution to the metabolism of these isoflavones by human liver. These findings were substantiated by observations that recombinant human UGT1A1 and UGT1A9, but not UGT1A4, catalysed the production of the major glucuronides of both genistein and daidzein in vitro. Recombinant human UGT1A8 also metabolised both genistein and daidzein, whereas UGT1A6 was specific to genistein and UGTs 2B7 and 2B15 were inactive, or only marginally active, with either isoflavone as substrate. The intestinal isoform UGT1A10 metabolised either both isoflavones or genistein only, depending on the commercial supplier of the recombinant enzyme, possibly as a result of a difference in amino acid sequence, which we were unable to confirm. Daidzein (16 microM) increased cell death in the MCF-7 human breast cancer cell line and this effect was reversed by glucuronidation. In view of a well-characterised functional polymorphism in UGT1A1, these observations may have implications for inter-individual variability in the potential health-beneficial effects of isoflavone consumption.

Topics: Anticarcinogenic Agents; Breast Neoplasms; Cell Line, Tumor; Chromatography, High Pressure Liquid; Genistein; Glucuronides; Glucuronosyltransferase; Glycine max; Humans; Isoflavones; Liver; Models, Chemical; Phytoestrogens; UDP-Glucuronosyltransferase 1A9

There is considerable evidence that exogenous estrogenic compounds can have adverse effects on fertility. The main reason cited in literature for hyperestrogenism in pigs is contamination of feedstuffs by the mycotoxin zearalenone (Boehm, 2000), but further estrogenically active substances might also be involved in cases of impaired fertility with symptoms like enlarged, red-coloured vulvae in piglets, irregular estrus cycles and anestrus of sows (Bennetts et al., 1946; Drane et al., 1981). It is well known that soy used in diets for pigs as a main protein source contains phytoestrogens. Amongst them, isoflavones like genistein and daidzein are of particular interest. Aim of this study was to optimize and use an established bioassay (Kluczka, 2003) to determine estrogenic activity in feedstuffs for pigs related to isoflavones and further substances with estrogenic potential. This bioassay is a reporter gene assay based on stably transfected human embryonal kidney cells (HEK 293) that contains either alpha or beta estrogen receptor (alpha- or beta-HEK). The estrogenic activity measured in the luciferase assay was expressed in estradiol-equivalents (EEQ) and the results were compared with the isoflavone content (genistein, daidzein) obtained by chemical analysis using high performance liquid chromatography-Ultraviolet (HPLC-UV). Mean estrogenic activity in diets fed to sows in herds with altered fertility was 275.8 microg EEQ/kg feed in alpha-HEK cells and 295.0 microg EEQ/kg feed in beta-HEK cells. Feedstuffs from herds without any altered fertility showed an average estrogenic activity of 204.9 microg EEQ/kg feed in alpha-HEK and 213.3 microg EEQ/kg feed in beta-HEK. The estrogenic activity was strongly related to the concentration of the isoflavones (alpha-HEK, r(2)=0.9488; beta-HEK, r(2)=0.9427). Clinically relevant zearalenone concentrations (>50-150 microg/kg feed) displayed estrogenic effects in the bioassay that did not differ significantly from those caused by high isoflavone concentration because of the use of soy as protein source.

Topics: Animal Feed; Animal Nutritional Physiological Phenomena; Animals; Biological Assay; Chromatography, High Pressure Liquid; Estrogen Receptor alpha; Estrogen Receptor beta; Female; Fertility; Food Contamination; Genistein; Humans; Isoflavones; Phytoestrogens; Receptors, Estrogen; Swine; Transfection; Tumor Cells, Cultured; Zearalenone

The use of daunomycin against neoplasms is limited due to its severe cardiotoxicity. The cytotoxicity of daunomycin can be minimized by linking it to an affinity tag. Since ovarian cancer cells are sensitive to isoflavone action, we synthesized a daidzein daunomycin conjugate. In MLS human ovarian cancer cells, the conjugate was shown to have a larger cytotoxic effect than daunomycin per se at a low concentration. The conjugate was then tested in vivo in mice carrying MLS xenografts. Tumour growth in the groups of conjugate and daunomycin was inhibited by >50% as compared to vehicle treated mice. In contrast to daunomycin treated mice, no weight reduction or death was seen in mice treated with the conjugate. In vivo imaging of the fluorescence signal generated by daunomycin indicated uptake of both conjugate and daunomycin by the tumour. Tumour fluorescence was, however, higher in the conjugate treated mice than in the daunomycin treated mice, thus suggesting specific delivery of the drug to the tumour. Histological examination of myocardial tissue indicated that only the daunomycin, but not conjugate treated mice showed cardiac damage. These results indicate that targeting of daunomycin via carboxymethyldaidzein retains daunomycin's cytotoxic effects while averting its toxicity in an ovarian xenograft.

Topics: Animals; Antibiotics, Antineoplastic; Cell Line, Tumor; Cell Survival; Daunorubicin; Female; Humans; Isoflavones; Mice; Mice, Nude; Molecular Structure; Ovarian Neoplasms; Phytoestrogens; Tumor Burden; Xenograft Model Antitumor Assays

The phytoestrogens genistein, daidzein and the daidzein metabolite equol have been shown previously to possess oestrogen agonist activity. However, following consumption of soya diets, they are found in the body not only as aglycones but also as metabolites conjugated at their 4'- and 7-hydroxyl groups with sulphate. This paper describes the effects of monosulphation on the oestrogen agonist properties of these three phytoestrogens in MCF-7 human breast cancer cells in terms of their relative ability to compete with [(3)H]oestradiol for binding to oestrogen receptor (ER), to induce a stably transfected oestrogen-responsive reporter gene (ERE-CAT) and to stimulate cell growth. In no case did sulphation abolish activity. The 4'-sulphation of genistein reduced oestrogen agonist activity to a small extent in whole-cell assays but increased the relative binding affinity to ER. The 7-sulphation of genistein, and also of equol, reduced oestrogen agonist activity substantially in all assays. By contrast, the position of monosulphation of daidzein acted in an opposing manner on oestrogen agonist activity. Sulphation at the 4'-position of daidzein resulted in a modest reduction in oestrogen agonist activity but sulphation of daidzein at the 7-position resulted in an increase in oestrogen agonist activity. Molecular modelling and docking studies suggested that the inverse effects of sulphation could be explained by the binding of daidzein into the ligand-binding domain of the ER in the opposite orientation compared with genistein and equol. This is the first report of sulphation enhancing activity of an isoflavone and inverse effects of sulphation between individual phytoestrogens.

Topics: Breast Neoplasms; Cell Line, Tumor; Cell Proliferation; Equol; Estradiol; Estrogen Receptor alpha; Female; Genistein; Humans; Isoflavones; Phytoestrogens; Recombinant Proteins; Structure-Activity Relationship; Sulfates

The activation of 17beta-estradiol (E2) to 2-hydroxyestradiol (2-HO-E2), the more genotoxic 4-hydroxyestradiol (4-HO-E2), and the oxidation to the respective quinones constitutes a risk factor in hormonal carcinogenesis. 2-HO-E2 is formed by cytochrome P450 CYP1A1, and 4-HO-E2 is formed by CYP1B1. Both are detoxified by catechol-O-methyltransferase (COMT), whereas their quinones are inactivated by NADPH-quinone-oxidoreductase (QR). Since the soy isoflavones genistein (GEN) and daidzein (DAI) are widely consumed due to their putative protective function in breast carcinogenesis, we examined the influence of E2, GEN, and DAI on CYP1A1/1B1, COMT, and QR expression in MCF-7 cells by reverse transcription/competitive PCR. CYP1A1 and COMT enzyme activity were determined using ethoxyresorufin and quercetin as substrates. Furthermore, estrogen receptor (ER)-regulated cell proliferation was determined by E-screen. E2, GEN, and DAI inhibited the expression of CYP1A1, COMT, and QR. The maximum effect (reduction by 40-80%, depending on the gene product and compound) was obtained at 100 pM E2, 1 microM GEN, and 10 microM DAI, which also induced the most pronounced cell proliferation in the E-screen. In contrast, expression of CYP1B1 was only slightly affected. CYP1A1 and COMT mRNA levels correlated with enzyme activities. The ER antagonist ICI 182,780 reversed the E2- and isoflavone-mediated effects. Thus, GEN and DAI at estrogen-active concentrations stimulate the formation of the more E2 genotoxic metabolites and inhibit the detoxification of catechol and quinone estrogens in estrogen-responsive tumor cells.

Topics: Aryl Hydrocarbon Hydroxylases; Breast Neoplasms; Catechol O-Methyltransferase; Cell Proliferation; Cytochrome P-450 CYP1A1; Cytochrome P-450 CYP1B1; Estradiol; Female; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; Genistein; Humans; Isoflavones; NAD(P)H Dehydrogenase (Quinone); Phytoestrogens; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tumor Cells, Cultured

Pueraria mirifica tubers collected from 28 out of 76 provinces of Thailand and Pueraria lobata tubers collected from Guangzhou province, China were submitted to HPLC analysis with the established gradient system comprising 1.5% acetic acid and acetonitrile. Five major isoflavonoids, including puerarin, daidzin, genistin, daidzein and genistein, were adopted as authentic standards. P. mirifica tubers showed intra- as well as inter-provincial differences in isoflavonoid and total isoflavonoid contents. The difference in both cases should be mostly influenced by genetic and environmental factors. In comparison with P. lobata, P. mirifica population exhibited differences only with a lower amount of daidzein.

Topics: Calibration; China; Chromatography, High Pressure Liquid; Drugs, Chinese Herbal; Genistein; Isoflavones; Molecular Structure; Phytoestrogens; Plant Tubers; Pueraria; Quality Control; Seasons; Thailand

It has been suggested that the consumption of a diet rich in phytoestrogens might protect against a variety of diseases common in Western societies. However, there are little available data on the food sources or distribution of intake in the UK diet.. To estimate the average intake and range of soya foods and isoflavones in a population-based cohort and to provide data on isoflavone consumption by food group.. Men and women (11,843) from the Norfolk arm of the European Prospective Investigation into Cancer and Nutrition (EPIC).. Dietary daidzein and genistein intakes were obtained from 7-day food diaries, completed by participants between 1993 and 1998 and calculated from an in-house food composition database. Energy and anthropometric measurements were also carried out.. Average daily isoflavone intakes for both men and women were less than 1 mg (interquartile range (IQR) men: 0.39-0.82 mg; women: 0.30-0.64 mg). However, in soya-consumers, average daily intakes were higher: 8.6 mg in women (IQR: 2.28-10.72 mg) and 7.5 mg in men (IQR: 2.22-9.17 mg). In both men and women, bread and bread rolls made the highest contribution to isoflavone intake - 62.5 and 53.0%, respectively. In soya-consuming men and women, vegetable dishes and milks were the main contributors - 25.0 and 38.5% in men and 38.5% and 26.0% in women, respectively.. Isoflavone intake is low in the UK but may be an underestimate due to soya added to commercial products. Future analyses of the isoflavone and lignan content of basic ingredient foods and commercial items commonly consumed in the UK diet will enable more accurate estimates of phytoestrogen intake to be made. The ability to estimate isoflavone intake in Western populations more accurately will enable investigations to be conducted into the suggested beneficial effects of phytoestrogens on health.

Topics: Adult; Aged; Cohort Studies; Confidence Intervals; Databases, Factual; Diet Records; Diet Surveys; Female; Food Analysis; Genistein; Humans; Isoflavones; Male; Middle Aged; Odds Ratio; Phytoestrogens; Soy Foods; United Kingdom

HOXA10 is necessary for normal development of the Müllerian duct, and continued adult expression in the uterus is necessary for female fertility. HOXA10 expression is altered by diethylstilbestrol, leading to uterine anomalies. Other endocrine disruptors may potentially lead to reproductive anomalies or dysfunction by altering HOXA10 expression. Here we investigated the effect of isoflavones on HOXA10 expression after in utero or adult exposure in the mouse. Genistein, but not diadzein, regulated HOXA10 mRNA and protein expression in the adult mouse uterus. In contrast, in utero genistein or diadzein exposure had no lasting effect on HOXA10 expression in the exposed offspring. Reporter gene expression driven by the HOXA10 estrogen response element was increased in a dose-responsive manner by genistein, but not daidzein. Neither estrogen receptor-alpha nor estrogen receptor-beta binding to the HOXA10 estrogen response element was affected by genistein or daidzein. In utero exposure to isoflavones is unlikely to result in HOXA10-mediated developmental anomalies. Adult genistein exposure alters uterine HOXA10 expression, a potential mechanism by which this agent affects fertility.

Topics: Animals; Cells, Cultured; DNA-Binding Proteins; Female; Gene Expression Regulation; Genes, Reporter; Genistein; Homeobox A10 Proteins; Homeodomain Proteins; Humans; Isoflavones; Maternal-Fetal Exchange; Mice; Mice, Inbred Strains; Phytoestrogens; Pregnancy; Pregnancy, Animal; Prenatal Exposure Delayed Effects; Receptors, Estrogen; Response Elements; Uterus

An anaerobic incubation mixture of two bacterial strains Eggerthella sp. Julong 732 and Lactobacillus sp. Niu-O16, which have been known to transform dihydrodaidzein to S-equol and daidzein to dihydrodaidzein respectively, produced S-equol from daidzein through dihydrodaidzein. The biotransformation kinetics of daidzein by the mixed cultures showed that the production of S-equol from daidzein was significantly enhanced, as compared to the production of S-equol from dihydrodaidzein by Eggerthella sp. Julong 732 alone. The substrate daidzein in the mixed culture was almost completely converted to S-equol in 24 h of anaerobic incubation. The increased production of S-equol from daidzein by the mixed culture is likely related to the increased bacterial numbers of Eggerthella sp. Julong 732. In the mixture cultures, the growth of Eggerthella sp. Julong 732 was significantly increased while the growth of Lactobacillus sp. Niu-O16 was suppressed as compared to either the single culture of Eggerthella sp. Julong 732 or Lactobacillus sp. Niu-O16. This is the first report in which two metabolic pathways to produce S-equol from daidzein by a mixed culture of bacteria isolated from human and bovine intestinal environments were successfully linked under anaerobic conditions.

Topics: Animals; Bacteria; Bacteria, Anaerobic; Equol; Humans; Intestines; Isoflavones; Metabolic Networks and Pathways; Microbiological Techniques; Phytoestrogens

The ovariectomized (OVX) rat, as an established animal model of human osteoporosis, was adopted in the present experiment to study the protective effects of sodium daidzein sulfonate (SDS) on trabecular bone. Six-month-old Sprague-Dawley rats were sham-operated or ovariectomized. Five days later, the OVX rats were randomly assigned to one of three experimental groups and treated for 90 days with vehicle, 17beta-estradiol (E(2)) or SDS. Compared with OVX rats, SDS administration (15 mg/kg) prevented OVX-induced decrease in lumbar vertebral and femoral bone mineral density (BMD), and significantly increased bone mechanical strength parameters, including ultimate stress and elastic modulus. In the OVX group, the structure of trabecular plate in the femoral head was absorbed and became progressively thinner or was removed completely, accompanied by enlargement of marrow cavities and amalgamation of two or more marrow cavities. Administration of SDS and E(2 )prevented the change of trabecular bone microarchitecture induced by OVX, increasing the trabecular bone area and trabecular thickness, while decreasing the trabecular separation. These results indicate that SDS administration prevents OVX-induced decrease in BMD and bone mechanical strength, and has a moderate protective effect on the microarchitecture of trabecular bone in aged Sprague-Dawley rats.

Topics: Amino Acids; Animals; Bone and Bones; Bone Density; Bone Resorption; Dose-Response Relationship, Drug; Estradiol; Female; Femur Head; Isoflavones; Lumbar Vertebrae; Microscopy, Electron, Scanning; Organ Size; Osteocalcin; Ovariectomy; Phytoestrogens; Random Allocation; Rats; Rats, Sprague-Dawley; Tensile Strength; Uterus; Weight Gain

To identify the phytoandrogen from phytohormone, we established an assay to assess the androgenicity of phytoestrogens by using androgen receptor (AR) cofactors to modulate the AR transcriptional activity.. A Dual-luciferase reporter assay was used to evaluate the transcriptional activity of AR stimulated by the phytoestrogen daidzein.. The Dual luciferase data showed that daidzein can enhance androgenic effects in AR negative PC-3 cells cotransfected with AR and AR cofactors. In AR and ARA70 positive LNCaP cells, daidzein can enhance ARA55-mediated induction of AR transcriptional activity. With increasing amounts of transfected ARA55, AR transcriptional activity was enhanced by daidzein in a dose-dependent manner.. Although daidzein is a phytoestrogen, it can create androgenic effects when cells are cotransfected with AR cofactors. When screening for phytoandrogens, the modulating effects of AR cofactors with AR should be considered in the assay system.

Topics: Cell Line, Tumor; Dihydrotestosterone; Estradiol; Humans; Intracellular Signaling Peptides and Proteins; Isoflavones; LIM Domain Proteins; Male; Nuclear Receptor Coactivators; Oncogene Proteins; Phytoestrogens; Prostatic Neoplasms; Receptors, Androgen; Transcription Factors; Transcription, Genetic; Transfection

Plant-derived phytoestrogens and estrogens in hormone replacement therapies have overlapping yet sometimes divergent effects on the incidence of breast cancer and osteoporosis. Using human MCF-7 breast carcinoma and G-292 osteosarcoma cell lines, it was investigated whether the phytoestrogens genistein and daidzein affect reporter gene transcription via the estrogen receptors (ERs) ERalpha and ERbeta1 as well as whether they affect the expression of estrogen-responsive genes in MCF-7 cells and the secretion of the cytokine IL-6 from G-292 cells. The results showed that genistein and daidzein potently trigger transactivation with ERbeta1 from estrogen response element-reporter genes (EC50s of 1.7-16 nM) although they were 400- to 600-fold less potent than 17beta-estradiol (E2) (EC50 of 0.02-0.04 nM). E2 was the only potent activator of ERalpha (EC50 of 0.1-0.4 nM). The rank order potency (E2 > genistein > daidzein) is maintained in MCF-7 cells as well as G-292 cells with both receptor subtypes, with a strong receptor selectivity of the phytoestrogens for ERbeta1 over ERalpha. Genistein and daidzein increased the expression of estrogen-responsive genes in MCF-7 cells. Daidzein, like E2, inhibited IL-1beta- and hormone-mediated IL-6 secretion from G-292 cells. The results provide a basis for understanding how dietary phytoestrogens protect bone without increasing the risks for breast cancer.

Topics: Breast Neoplasms; Cell Line, Tumor; Estrogen Receptor beta; Estrogens; Female; Gene Expression Profiling; Genes, BRCA1; Genistein; Humans; Interleukin-6; Isoflavones; Osteosarcoma; Phytoestrogens; Response Elements

Equol is the main active intestinal metabolite of the isoflavone daidzein and is postulated to be responsible for the cardiovascular benefits of soy. Cerebral vascular effects of equol are unknown. We compared the vasorelaxant and antioxidant effects of equol and daidzein in carotid and basilar artery of normal and hypertensive rats.. Relaxant responses to equol and daidzein were measured in the isolated carotid artery and in the basilar artery in vivo. Effects of nitric oxide synthase (NOS) inhibition, high extracellular K(+), endothelial removal and gender on responses to equol were investigated in carotid arteries. Antioxidant activity was assessed as the reduction of NADPH-induced superoxide levels. Hypertension was induced using angiotensin II (0.7 mg/kg per day for 14 days).. In normotensive rats, equol displayed vasorelaxant activity similar to daidzein. The relaxant effect of equol was independent of an intact endothelium, NOS activity, K(+) channels and gender. In the basilar artery, where superoxide levels are higher, equol exerted weak antioxidant effects, whereas effects of daidzein were insignificant. During hypertension, equol-induced vasorelaxation was preserved, whereas relaxant responses to daidzein were impaired.. Equol possesses substantial vasodilator and weak antioxidant activity in cerebral arteries, with similar activity to daidzein, whereas in hypertension the vasorelaxant response to equol, but not daidzein, is preserved. However, daidzein possesses comparable direct vascular effects with equol, without the need for intestinal conversion to equol. Nevertheless, equol may represent a more useful therapeutic agent during cerebral vascular disease.

Topics: Analysis of Variance; Angiotensin II; Animals; Antioxidants; Blood Pressure; Carotid Arteries; Cerebral Arteries; Dose-Response Relationship, Drug; Drug Interactions; Enzyme Inhibitors; Equol; Female; Hypertension; In Vitro Techniques; Isoflavones; Male; Phytoestrogens; Rats; Superoxides; Vasodilation

Soy products are of particular interest because of their potential health benefits in a range of hormonal conditions, such as osteoporosis, due to their high content in phytoestrogens. Because equol, the main metabolite from soy isoflavones, is thought to be powerful, the present study was designated to evaluate the bone-sparing effects of equol by either providing the molecule through the diet or by eliciting its endogenous production by modulating intestinal microflora by short-chain fructooligosaccharides (sc-FOS) or live microbial (Lactobacillus casei) together with daidzein, its precursor.. A comparison with daidzein and genistein was also performed. Rats (3 months old) were ovariectomised (OVX) or sham-operated (SH). Ovariectomised rats were randomly assigned to six experimental diets for 3 months: a control diet (OVX), the control diet supplemented with either genistein (G), or daidzein (D), or equol (E) at the level of 10 microg/g body weight/d. The remaining OVX rats were given daidzein at the dose of 10 mug/g body weight/d, simultaneously with short-chain FOS (Actilight, Beghin-Meiji) (D+FOS) or Lactobacillus casei (Actimel, Danone) (D+L). The SH rats were given the same control diet as OVX.. Genistein, daidzein or equol exhibited a bone sparing effect. Indeed, total femoral bone mineral density (BMD) was significantly enhanced (compared to that of OVX rats), as was the metaphyseal compartment. Bone strength was improved by E consumption, but not by genistein or daidzein given alone. As far as the FOS diet is concerned, the addition of prebiotics significantly raised efficiency of the daidzein protective effect on both femoral BMD and mechanical properties. The effects of lactobacillus were similar, except that the increase in metaphyseal-BMD was not significant.. In conclusion, long-term equol consumption, like genistein and daidzein, in the ovariectomized rat, provides bone sparing effects. Adding indigestible sugars, such as FOS or live microbial as L. casei, in the diet significantly improves daidzein protective effects on the skeleton.

Topics: Animals; Biomarkers; Body Weight; Bone Density; Bone Resorption; Disease Models, Animal; Equol; Female; Femur; Genistein; Glycine max; Isoflavones; Organ Size; Osteocalcin; Osteoporosis; Ovariectomy; Phytoestrogens; Rats; Rats, Wistar; Uterus

A population-based case-control study of diet, inherited susceptibility and prostate cancer was undertaken in the lowlands and central belt of Scotland to investigate the effect of phyto-oestrogen intake and serum concentrations on prostate cancer risk. A total of 433 cases and 483 controls aged 50-74 years were asked to complete a validated FFQ and provide a non-fasting blood sample. Multivariate logistic regression analysis found significant inverse associations with increased serum concentrations of enterolactone (adjusted OR 0.40, 95 % CI 0.22, 0.71] and with the consumption of soy foods (adjusted OR 0.52, 95 % CI 0.30, 0.91). However, no significant associations were observed for isoflavone intake or serum genistein, daidzein and equol. This study supports the hypotheses that soy foods and enterolactone metabolised from dietary lignans protect against prostate cancer in older Scottish men.

Topics: 4-Butyrolactone; Aged; Case-Control Studies; Diet; Diet Surveys; Energy Intake; Equol; Genistein; Humans; Isoflavones; Lignans; Male; Middle Aged; Phytoestrogens; Prostatic Neoplasms; Risk Assessment; Scotland

Dietary phytoestrogens may play a role in chronic disease occurrence. The aim of our study was to assess the variability of plasma concentrations in European populations. We included 15 geographical regions in 9 European countries (Denmark, France, Germany, Greece, Italy, Spain, Sweden, The Netherlands, and UK) and a 16th region, Oxford, UK, where participants were recruited from among vegans and vegetarians. All subjects were participants of the European Prospective Investigation into Cancer and Nutrition (EPIC). Plasma concentrations of 3 isoflavones (daidzein, genistein, and glycitein), 2 metabolites of daidzein [O-desmethylangolensin (O-DMA) and equol] and 2 mammalian lignans (enterodiol and enterolactone) were measured in 1414 participants. We computed geometric means for each region and used multivariate regression analysis to assess the influence of region, adjusted for gender, age, BMI, alcohol intake, smoking status, and laboratory batch. Many subjects had concentrations below the detection limit [0.1 microg/L (0.4 nmol/L)] for glycitein (80%), O-DMA (73%) and equol (62%). Excluding subjects from Oxford, UK, the highest concentrations of isoflavones were in subjects from the Netherlands and Cambridge, UK [2-6 microg/L (7-24 nmol/L); P < 0.05], whereas concentrations for lignans were highest in Denmark [8 microg/L (27 nmol/L); P < 0.05]. Isoflavones varied 8- to 13-fold, whereas lignans varied 4-fold. In the vegetarian/vegan cohort of Oxford, concentrations of isoflavones were 5-50 times higher than in nonvegetarian regions. Region was the most important determinant of plasma concentrations for all 7 phytoestrogens. Despite the fact that plasma concentrations of phytoestrogens in Europe were low compared with Asian populations, they varied substantially among subjects from the 16 different regions.

Topics: Cohort Studies; Diet, Vegetarian; Europe; Female; Humans; Isoflavones; Lignans; Male; Middle Aged; Multivariate Analysis; Osmolar Concentration; Phytoestrogens; Prospective Studies

Soy phytoestrogens are popular, but information on their coronary effects in patients with suspected ischemic heart disease is limited. Accordingly, we investigated the relationship between blood phytoestrogen levels and coronary reactivity in women with suspected myocardial ischemia referred for coronary angiography.. Coronary flow velocity reserve (CFVR) and volumetric flow reserve (VFR) to adenosine (ADO) and nitroglycerin (NTG) (nonendothelial-dependent responses) and acetylcholine (ACH) (endothelial-dependent response) were assessed in 106 women from the Women's Ischemia Syndrome Evaluation (WISE). Blood phytoestrogen (daidzein and genistein) and estrogen (estradiol) levels were correlated with coronary reactivity measures.. Participants were mostly postmenopausal (79%), mean age 56 years, and 24% had obstructive coronary artery disease (CAD) at angiography. Genistein blood levels were negatively correlated with nonendothelial-dependent coronary flow responses. The highest genistein tertile (>6.1 ng/mL) had a CFVR of 2.1 +/- 0.5 (mean +/- SD) and VFRADO of 1.0 +/- 0.6, and both were significantly (p= 0.0001) lower compared with the other genistein tertiles combined. Similar associations were noted for CFVR(NTG) and VFR(NTG) (p = 0.03 and p = 0.01, respectively). The highest genistein tertile was associated with lower CFVR(ACH) compared with the other tertiles (p = 0.03). In multivariable modeling, blood genistein levels were significant independent predictors of coronary flow responses to ADO. There were no significant correlations between coronary reactivity variables and daidzein or endogenous estrogen.. In women with suspected myocardial ischemia, higher genistein blood levels are associated with impaired nonendothelial-dependent and endothelial-dependent coronary microvascular function.

Topics: Acetylcholine; Adenosine; Adult; Aged; Blood Flow Velocity; Coronary Angiography; Coronary Artery Disease; Coronary Vessels; Estradiol; Female; Genistein; Humans; Isoflavones; Middle Aged; Myocardial Ischemia; Nitroglycerin; Phytoestrogens; Predictive Value of Tests

Human diet contains weakly estrogenic compounds such as daidzein (DAI) and genistein (GEN), phytoestrogens present in soy and many vegetables as well as bisphenol A (BPA), a contaminant from packing materials and plastic containers for foods and beverages. In light of concerns about hormonally active agents, biomonitoring methods are needed to assess human exposure to such compounds. A method for simultaneous determination of DAI, its metabolite equol (EQ), GEN, and BPA by GC-MS analysis was established, validated and applied to measure concentrations in human urine. Sample preparation involves enzymatic conjugate cleavage, SPE and derivatization by silylation. For GC/MS analysis, deuterated DAI and GEN and( 13)C-BPA are used as internal standards. LOD are 4, 4, 5 and 3 ng/mL urine for DAI, EQ, GEN and BPA, respectively. Interassay variations were 9% for DAI, 15% for EQ, 18% for GEN and 10% for BPA. Simple workup and accuracy of the method are suited for biomonitoring. An analysis of urine samples from 15 adults consuming typical German food revealed dietary exposure to phytoestrogens in all samples: GEN concentrations ranged between 13 and 238 ng/mL, those for DAI ranged from 12 to 356 ng/mL. More than half of the individuals excreted also the more estrogenic metabolite EQ, at levels of 8-128 ng/mL. Higher concentrations (GEN: 820, DAI: 960 and EQ: 1740 ng/mL) were measured in a 24 h urine sample upon ingestion of soy protein (50 g with 12.9 mg DAI and 25.2 mg GEN). Only urine collected after some days on strict phytoestrogen-free diet had undetectable isoflavone levels. BPA was detected in 9 of 15 urine samples ranging from 3 to 11 ng/mL, and at 55 ng/mL in one sample. In conclusion, a reliable method to determine BPA and isoflavones in urine was established and applied in a pilot study: Biomonitoring results show much higher dietary exposure to phytoestrogens than to BPA in German adults.

Topics: Adult; Benzhydryl Compounds; Diet; Equol; Gas Chromatography-Mass Spectrometry; Genistein; Humans; Isoflavones; Phenols; Phytoestrogens; Reproducibility of Results

Daidzein (4',7-dihydroxyisoflavone), a soy phytoestrogen, is a weakly estrogenic compound that may have potential health benefits. Biotransformation of daidzein by the human gut microflora after ingestion converts it to either the highly estrogenic metabolite equol or to nonestrogenic metabolites. We investigated the metabolism of daidzein by colonic microflora of rats. Fecal samples, obtained before and after rats were exposed to daidzein at 250 or 1000 parts per million, were incubated in brain-heart infusion (BHI) broth with daidzein under anaerobic conditions. Samples were removed from the cultures daily and analyzed by high-performance liquid chromatography (HPLC) and mass spectrometry. The fecal bacteria of all rats, regardless of prior daidzein exposure, metabolized the added daidzein to dihydrodaidzein. Both compounds disappeared rapidly from BHI cultures incubated for more than 24 h, but no other daidzein metabolites were detected. Only daidzein and dihydrodaidzein were found in a direct analysis of the feces of rats that had consumed daidzein in their diets. Unlike the fecal bacteria of humans and monkeys, the rat flora rapidly metabolized daidzein to aliphatic compounds that could not be detected by HPLC or mass spectral analysis.

Topics: Animals; Animals, Genetically Modified; Bacteria; Biotransformation; Chromatography, High Pressure Liquid; Dose-Response Relationship, Drug; Feces; Gastrointestinal Tract; Isoflavones; Phytoestrogens; Rats; Rats, Inbred F344; Spectrometry, Mass, Electrospray Ionization

This study investigated the effects of soy isoflavone intake on uterotrophic responses in growing (juvenile) and adult female rats. In the growing rats, feed intake showed a decreasing trend as the level of dietary isoflavones increased to 0.02%, 0.1%, and 0.2% of the diets. However, in the case of the adult rats there were no significant differences among groups. Weight gains were significantly lower in the rats fed 0.1% and 0.2% isoflavones than the controls in both juvenile and adult rats. The urinary excretion of daidzein and genistein was significantly increased with increasing levels of dietary isoflavones. The calculated urinary recoveries of daidzein and genistein were significantly lower in the groups fed 0.1% and 0.2% isoflavones compared to the juvenile and adult rat groups fed 0.02% isoflavones; no significant difference was observed between the 0.1% and 0.2% groups. The calculated urinary recoveries of daidzein and genistein in the adult rats were significantly higher than in the juvenile rats. The differences in the urinary recoveries between ages may be due to greater availability of the isoflavones in the adult rats. Isoflavone supplementation did not alter the histological phenotype of endometrial cells in growing rats, but a hyperplastic response of endometrium was shown in the adult rats. Dietary isoflavones, therefore, may not have an estrogenic effect on the uterus at these dose levels during the growth period, but this organ would be expected to be a likely target for isoflavone action in adults. We observed in the present study that isoflavones are more bioavailable in adult rats than in the juvenile rats. Therefore, soy isoflavone supplementation may not act as an endocrine disrupter during the growth period but may exert a phytoestrogenic effect on the uterus of adult rats.

Topics: Aging; Animals; Biological Availability; Body Weight; Diet; Eating; Female; Genistein; Glycine max; Isoflavones; Phytoestrogens; Rats; Rats, Sprague-Dawley; Uterus

The use of soy isoflavones is a potential alternative to hormone replacement therapy in post-menopausal bone-loss prevention. Nevertheless, phytoestrogens can target other organs and may disrupt cell proliferation, or could modify endogenous steroid hormone metabolism. These mechanisms could be linked to an increased risk of developing cancer. We therefore studied the possible side effects of such treatments in an experimental model of menopause. Forty adult female Wistar rats were ovariectomized and fed with a genistein-, daidzein- or equol-supplemented diet at bone-sparing levels (10 mg/kg BW/day) for 3 months. The estrogenic effects were assessed by histological and molecular analyses on reproductive organs. The impact on the oxidative metabolism of estradiol and on associated cytochrome P450 (CYP) activities was evaluated in liver microsomes. The relative wet weights of both the uterus and the vagina were increased in the equol group, but no significant changes in proliferating cell nuclear antigen or hormone receptor mRNA expression were noticed. In contrast, genistein and daidzein did not induce uterotrophy but caused an overexpression of estrogen receptor alpha mRNA which could correspond to a long-lasting effect of physiological concentrations of estrogens. The hepatic metabolism of estradiol was influenced by daidzein which increased the synthesis of putative mutagenic derivatives. At the same time, genistein favored estrogen 2-hydroxylation, and equol decreased 4-hydroxyestrogen production. Surprisingly, no significant alteration in hepatic CYP activities was detected. Taken together, these results demonstrate that isoflavonoid-based bone-sparing treatments are able to cause side effects on other estrogen-sensitive target organs when given in the long-term.

Topics: Animals; Bone Density; Cytochrome P-450 Enzyme System; Disease Models, Animal; Equol; Estradiol; Female; Gene Expression Regulation; Genistein; Isoflavones; Menopause; Microsomes, Liver; Organ Size; Osteoporosis; Ovariectomy; Phytoestrogens; Proliferating Cell Nuclear Antigen; Rats; Rats, Wistar; Uterus; Vagina

Soybean isoflavones have multiple beneficial health effects especially on estrogen-deficient diseases such as menopausal symptoms. In this study, isoflavones were produced from soybean flour, and the extraction and purification parameters were optimized to give a high yield of total isoflavones, about 0.62 mg of aglycones/g of soybean flour, which is >2 times the initial yield. HPLC analysis and MTT cell proliferation assay using MCF-7 cells revealed that the product thus obtained not only contained a high content of isoflavone aglycones but also had estrogenic activity. MTT data also revealed that both genistein and daidzein exhibited estrogenic effects at lower concentrations and antiproliferative effects at higher concentrations, and 1 microM genistein and 10 microM daidzein exerted significant estrogenic activities, which were not more than that of the endogenous level of 17beta-estradiol (E2). The production method developed can be used as a guideline for manufacturing soy isoflavones, and the MTT assay was demonstrated to be suitable for quality control on isoflavone products. The results on the estrogenic properties of isoflavones can be used as reference data for their effective and safe usages in estrogenic therapy.

Topics: Cell Division; Cell Line; Chromatography, High Pressure Liquid; Genistein; Glycine max; Isoflavones; Phytoestrogens

To compare endogenous and exogenous beta-glucosidases for the hydrolysis of the predominant isoflavone glucosides in soymilk in order to improve the biological activity.. beta-glucosidase activity of probiotic organisms, including Bifidobacterium animalis ssp. lactis Bb12, Lactobacillus acidophilus ATCC 4461 and Lactobacillus casei 2607 in soymilk, was evaluated and was related to the increase in the concentration of isoflavone aglycones during fermentation. The concentrations of isoflavone compounds in soymilk were monitored using a Varian model HPLC with an Amperometric electrochemical detector. The aglycone composition, also known as aglycone equivalent ratio, has been considered to be important for the delivery of health benefits of isoflavones, and was monitored during the fermentation of soymilk. Comparison of the hydrolytic effectiveness of both exogenous and endogenous enzyme during 4-h incubation in soymilk was conducted using the Otieno-Shah (O-S) index. Results showed that exogenous enzyme exhibited faster rate of isoflavone glucoside hydrolysis than that by endogenous enzyme. Highest O-S indices were obtained after 4, 3 and 2 h of incubation with enzyme solution having beta-glucosidase activity of 0.288 U ml(-1), 0.359 U ml(-1) and 0.575 U ml(-1), resulting into aglycone concentration increments of 5.87-, 6.07- and 5.94-fold, respectively. Conversely, aglycone concentration in the soymilk with B. animalis ssp. lactis Bb12, L. casei 2607 and L. acidophilus 4461 increased by 3.43-, 2.72- and 3.03-fold, respectively, after 4 h of fermentation at 37 degrees C. In addition, the O-S index of endogenous enzyme was much lower than that of the exogenous enzyme over the same 4-h incubation period. Optimum aglycone equivalent ratios coincided with highest O-S indices and highest aglycone concentrations in soymilk hydrolysed with exogenous enzyme. The same correlation of O-S indices and highest aglycone concentrations occurred for endogenous enzyme during the 24 h of fermentation.. Obtaining highest aglycone concentration and optimum aglycone equivalent ratio could provide a critical beginning point in clinical trials for the realization of unique health benefits of soy isoflavones.. Screening for beta-glucosidase activities of probiotics in soymilk and comparing their hydrolytic potentials with that of exogenous beta-glucosidase could find wide applications in the development of different aglycone-rich functional soy beverages.

Topics: Bifidobacterium; Cellulases; Chromatography, High Pressure Liquid; Culture Media; Fermentation; Food Microbiology; Genistein; Hydrolysis; Isoflavones; Lacticaseibacillus casei; Lactobacillus acidophilus; Phytoestrogens; Probiotics; Soy Milk

Topics: Anticarcinogenic Agents; Equol; Feeding Behavior; Genistein; Humans; Intestines; Isoflavones; Japan; Male; Phytoestrogens; Prostatic Neoplasms; Soy Foods

To observe the transcriptional regulation of the two isoflavones genistein and daidzein on target genes.. In this study, we used ERalpha or ERbeta over-expressing Hela cells to observe the transcriptional regulation of genistein and daidzein on ERE reporter gene with calcium-phosphate method, and furthermore observing the effects of phytoestrogen antagonist ICI 182780 on their activation.. Our results showed that both genistein and daidzein could activate ERE receptor gene through ERalpha and ERbeta, and these effects could be blocked by ICI 182780.. Both genistein and daidzein can mimic estrogen's effect to activate the transcription of target genes through binding to the ERs.

Topics: Estrogen Receptor alpha; Estrogen Receptor beta; Gene Expression Regulation; Genistein; HeLa Cells; Humans; Isoflavones; Phytoestrogens; Transcription, Genetic

Nutritional supplements containing soybean phytoestrogens, the isoflavones genistein (G) and daidzein (D), are increasingly used as alternative therapy for osteoporosis, cancer, and cardiovascular and other diseases with a frequency that increases with advancing age. In this study we examined the effects of subcutaneous administration of either G or D on serum lipid levels in orchidectomized (Orx) and intact (IA) middle-aged male rats, which are experimental models of andropause. Sixteen-month-old Wistar rats were treated with 10 mg/kg and 30 mg/kg of either G or D. The control groups received testosterone, estradiol, or vehicle for 3 weeks, after which the total serum cholesterol (TC), low-density lipoprotein (LDL-C), high-density lipoprotein (HDL-C), and total triglycerides (TT) were measured. Compared with the matching vehicle-treated controls, the higher doses of G and D and testosterone treatment significantly (P < 0.05) lowered the TC and lipoprotein cholesterol levels. The greatest effect was observed regarding LDL-C in both Orx and IA males after G and D treatments, in which LDL-C decreased by more than 30%. The lower isoflavone doses induced a significant cholesterol-lowering effect (P < 0.05) only in the Orx group. Like the estradiol treatment, the higher doses of G and D increased the TT levels in both rat models by more than 50% (P < 0.05). The lower doses of isoflavones increased TT only in the Orx group. In male middle-aged rats, injections of higher doses of G and D decreased the serum cholesterol levels, as did testosterone injection, and brought about an increase in serum triglycerides similar to that observed after estradiol treatment.

Topics: Age Factors; Animals; Cholesterol; Genistein; Growth Inhibitors; Injections, Subcutaneous; Isoflavones; Male; Orchiectomy; Phytoestrogens; Rats; Rats, Wistar; Testosterone; Triglycerides

To investigate the impact of exposure to the phytoestrogen daidzein on erectile function and sexual hormones. The negative effects of phytoestrogens on the male reproductive system, particularly on penile erection, have hardly been evaluated.. Thirty adult male Sprague-Dawley rats were equally divided into a normal control group, three experimental groups, and one positive control group. The three experimental groups were given daidzein at doses of 2, 20, and 100 mg/kg body weight daily, and the positive control group received 0.1 mg diethylstilbestrol per animal daily for 90 days. The apomorphine-induced erection test was performed 0, 30, 60, and 90 days after daidzein administration to evaluate for erectile function or dysfunction. After each test, blood samples were collected for plasma testosterone and luteinizing hormone measurement.. High-dose daidzein (100 mg/kg) decreased erectile responses to apomorphine from the 30th day of daidzein treatment and lasted to the 90th day without significant differences compared with the diethylstilbestrol-treated rats. However, similar changes were observed in the medium-dose daidzein (20 mg/kg) group from the 60th day. Low-dose daidzein (2 mg/kg) had no significant effect on the erectile responses to apomorphine compared with the normal control group (P >0.05). The plasma testosterone and luteinizing hormone levels showed a declining trend similar to that of the apomorphine-induced erections.. The phytoestrogen daidzein has the potential to adversely affect erectile function in a dose and time-related manner that is at least partly attributable to androgen deficiency. These findings implicate that phytoestrogens, especially isoflavones, if overconsumed for a long period, might be a novel risk factor for erectile dysfunction.

Topics: Androgen Antagonists; Animals; Apomorphine; Diethylstilbestrol; Dose-Response Relationship, Drug; Drug Administration Schedule; Isoflavones; Luteinizing Hormone; Male; Penile Erection; Phytoestrogens; Random Allocation; Rats; Rats, Sprague-Dawley; Testosterone; Yawning

The purpose of this study was to explore the neuroprotective effects of daidzein on the apoptotic pathway in the hippocampus and cortex of D-galactose treated mice. For this purpose we have examined the expression of bcl-2 mRNA, bax mRNA and caspase-3 in the hippocampus and cortex of D-galactose-treated mice after fed with 10 or 5 mg/kg of daidzein. The results of in situ hybridization experiments indicate that daidzein could help increase the transcriptions of bcl-2 and decrease the transcriptions of bax in those brain regions of D-galactose-treated mice. Furthermore, immunohistochemical studies showed that daidzein could reduce the expression of caspase-3 in both brain regions. These results suggest that daidzein in soybean can inhibit the D-gal induced apoptosis via Bcl-2/Bax apoptotic pathway and be a potential medical candidate for neurodegeneration therapy.

Topics: Animals; Apoptosis; bcl-2-Associated X Protein; Brain; Caspase 3; Estradiol; Female; Galactose; Gene Expression Regulation; Hippocampus; In Situ Hybridization; Isoflavones; Mice; Phytoestrogens; RNA, Messenger

Pueraria mirifica is a tuberous plant enriched with active phytoestrogens. There is no established information about the factors influencing isoflavonoid storage in the tubers. We investigated the tuberous storage of the major isoflavonoids of 1-year-old plants. Four cultivars of P. mirifica were cultivated in the same field trial during the same period to establish a unique plant age and differentiation under the same environment and soil conditions. The tubers collected from the 1-year-old plants in the summer, rainy season and winter were submitted to an HPLC analysis with a gradient system comprising 0.1% acetic acid and acetonitrile. Five major isoflavonoids, puerarin, daidzin, genistin, daidzein and genistein, were adopted as standards. P. mirifica tubers of different cultivars collected in the same season exhibited significant differences in individual and total isoflavonoid contents, showing chemovariety. P. mirifica tubers of the same cultivar collected from different seasons also exhibited significant differences in individual and total isoflavonoid contents, showing the influence of season. In conclusion, the tuberous storage of major isoflavonoids in 1-year-cultivated plants was greatly diverse and was strongly influenced by the season and plant genetics.

Topics: Chromatography, High Pressure Liquid; Drugs, Chinese Herbal; Genistein; Isoflavones; Models, Biological; Molecular Structure; Phytoestrogens; Plant Tubers; Pueraria; Rain; Seasons; Time Factors

To investigate the protective effects of daidzein (DD) on ventricular remodeling in rats with myocardial hypertrophy induced by pressure overload and its mechanism.. Myocardial hypertrophy model of rats induced by pressure overload was prepared by constricting abdominal aorta. The operated rats were randomly divided into sham operated control group, aorta-constricted model group and three DD groups (30, 60, 120 mg kg(-1)). Four weeks later, the heart-weight (HW), left ventricular weight (LVW), the ratio of HW/BW and LVW/BW (LVI) and the cardio-myocyte diameters (MD) after dyeing by HE colar were measured. The hydroxyroline, nitric oxide (NO) and the activity of nitric oxide synthetase (NOS) and Na+ -K+ -ATPase, Ca2+ -ATPase in left ventricle were quantified with spectrophotometry and the angiotension II (Ang II) in left ventricle and serum was messured with radioimmunoassay.. After treatment of the left ventricular with DD, vs aorta-contricted model group, NO content, cNOS and Na+ -K+ -ATPase, Ca2+ -ATPase activity were significantly increased, the content of AngII in left ventricle and serum and iNOS activity and the ratio of HW/BW, LVI, MD were significantly reduced.. DD has protective effects on ventricular remodeling in rats with myocardial hypertrophy induced by pressure overload and its mechanism may be related to raising NO content and reducing the level of Ang II.

Topics: Angiotensin II; Animals; Hypertrophy, Left Ventricular; Isoflavones; Male; Myocardium; Nitric Oxide; Phytoestrogens; Random Allocation; Rats; Rats, Wistar; Ventricular Remodeling

We compared the effects of genistein with its structural derivatives daidzein and equol on excitation of pulmonary artery and vein. The concentration of genistein necessary to inhibit contractions evoked by U46619 (1nM-100 microM) ranged from 10 to 100 microM. Genistein (55 microM) reduced KCl-responses by approximately 50% and essentially abolished those evoked by U46619. Daidzein was much less effective against either agonist, and equol was ineffective against U46619. A23187-evoked contractions were markedly reduced by all 3 isoflavones, but caffeine-evoked contractions were not. Using the Western blot technique, we found many proteins were tyrosine phosphorylated within 30 seconds after stimulation with U46619, reaching a peak at 120 seconds and then falling at 300 seconds. One band at 110 kD was increased nearly 300% above baseline, while 3 others ranging from 60 to 80 kD were more than doubled in intensity. Genistein had little effect on baseline levels of phosphorylation but largely prevented the U46619-induced change; daidzein was much less effective in this respect, and equol did not significantly affect this phosphorylation. We conclude that these isoflavones provide powerful tools in the study of excitation-contraction coupling of pulmonary vasculature and that inhibition of tyrosine kinase activity may be useful clinically against pulmonary hypertension.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Animals; Blotting, Western; Caffeine; Calcimycin; Cattle; Dose-Response Relationship, Drug; Equol; Genistein; In Vitro Techniques; Isoflavones; Muscle, Smooth, Vascular; Phosphorylation; Phosphotyrosine; Phytoestrogens; Potassium Chloride; Protein-Tyrosine Kinases; Proteins; Pulmonary Artery; Pulmonary Veins; Tyrosine; Vasoconstriction

The optimum test diet and rodent species/strain for evaluating endocrine-disrupting compounds (EDCs) are critical.. We conducted studies to evaluate rodent species sensitivity and the effects of diets varying in phytoestrogen content on the time of vaginal opening (VO) in CD-1 mice, Fischer 344 (F344) rats, and CD Sprague-Dawley (S-D) rats.. Mice were weaned on postnatal day (PND) 15 and rats on PND19 and randomly assigned to control or test diets. Body weights, food consumption, and time of VO were recorded.. The time of VO was significantly advanced in F344 rats fed diets containing daidzein and genistein, whereas these same diets did not advance VO in S-D rats. When animals were fed the AIN-76A diet spiked with genistein, time of VO was significantly advanced at all doses in CD-1 mice, at the two highest doses in F344 rats, and at the highest dose in S-D rats. The time of VO in F344 rats was more highly correlated with the phytoestrogen content than with the total metabolizable energy (ME) of 12 diets.. The S-D rat is less sensitive to dietary phytoestrogens compared with the F344 rat or the CD-1 mouse, suggesting that the S-D rat is not the ideal model for evaluating estrogenic activity of EDCs. The profound effects of dietary phytoestrogens on the time of VO, an estrogen-sensitive marker, indicate that a standardized open-formula phytoestrogen-free diet containing a low ME level should be used to optimize the sensitivity of estrogenic bioassays.

Topics: Animals; Body Weight; Diet; Energy Metabolism; Feeding Behavior; Female; Genistein; Isoflavones; Mice; Phytoestrogens; Rats; Rats, Inbred F344; Rats, Sprague-Dawley; Vagina

Intestinal bacterial metabolize the soy isoflavone daidzein to O-desmethylangolensin (O-DMA) or equol. Some individuals do not excrete O-DMA or equol after soy consumption, suggesting they do not harbor bacteria capable of producing these metabolites. The aim of this study was to evaluate bone mineral density (BMD) in relation to presence of these urinary metabolites.. BMD, determined by whole-body dual X-ray absorptiometry scan, was age-adjusted and evaluated in relation to O-DMA-producer and equol-producer phenotypes in 92 postmenopausal women, aged 50-75 years. Women consumed supplemental soy foods (daidzein source) for 3 days and collected a first-void urine sample on the fourth day in order to determine metabolic phenotypes.. In O-DMA producers (n=76) compared to O-DMA non-producers (n=16), greater total, leg and head BMD (p<0.05) were observed. Total BMD among the O-DMA producers (geometric mean=1.04 g/cm2) was 6% greater than total BMD among the O-DMA non-producers (geometric mean=0.98 g/cm2). Total and site-specific BMD did not differ between equol producers (n=24) and non-producers (n=68) (p>0.05). In exploratory analyses, among regular soy consumers, spinal BMD was 20% lower among the equol producers than non-producers, whereas, among soy non-consumers, no such difference was observed (p-interaction<0.05). Among equol producers, circulating estrone and free estradiol concentrations were inversely or not associated with total BMD, whereas, among equol non-producers, these hormones were positively associated (p-interaction<0.05).. Our results provide evidence that intestinal bacterial composition may influence BMD in postmenopausal women. Further studies characterizing associations of intestinal bacterial profiles with BMD are warranted.

Topics: Absorptiometry, Photon; Aged; Biomarkers; Bone Density; Equol; Female; Humans; Intestines; Isoflavones; Middle Aged; Osteoporosis, Postmenopausal; Phenotype; Phytoestrogens; Postmenopause; Soybean Proteins

We found detectable levels of three phytoestrogens (enterolactone, daidzein and genistein) and bisphenol A (BPA) in 21 residual amniotic fluid specimens that were collected before 20 weeks gestation. Samples were obtained by amniocentesis from women who were referred to the Mount Sinai Medical center because of advanced maternal age. Phytoestrogens were present in higher concentrations than BPA. Enterolactone was detected at the highest concentration (median 95.9 microg/L), followed by daidzein and genistein (9.5 and 1.4 microg/L, respectively). BPA was present at very low concentrations (10%>LOD of 0.5 microg/L). The relative concentration of the chemicals measured in amniotic fluid were identical to those in urine reported by other studies, i.e. enterolactone>daidzein>genistein>>BPA. Amniotic fluid is a source of fetal exposure to polar xenobiotics that come from the mother.

Topics: 4-Butyrolactone; Amniotic Fluid; Benzhydryl Compounds; Estrogens, Non-Steroidal; Female; Genistein; Humans; Isoflavones; Lignans; Maternal Exposure; Maternal-Fetal Exchange; Phenols; Phytoestrogens; Pregnancy; Xenobiotics

Phytoestrogens are plant compounds that have been proposed to have a variety of health benefits. The aim of this study was to assess the effects of these compounds on a number of physiological endpoints. Subjects were given a single intake of a phytoestrogen-rich (80 mg total phytoestrogens) supplement containing soy, rye and linseed (Phase 1), followed by a week-long intervention using the same supplement (Phase 2) (80 mg total phytoestrogens daily). A number of biochemical endpoints were assessed including urinary phytoestrogen metabolites, lipids, antioxidant status, DNA damage and insulin-like growth factor-1 (IGF-1) and IGF binding protein-1 (IGFBP-1) and -3 (IGFBP-3). Ten healthy female subjects took part in the study. Excretion of the isoflavones genistein, daidzein and equol in urine increased in both phases of the study. No other endpoint was altered in Phase 1. However, in Phase 2, concentrations of IGF-1 and IGFBP-3 were increased by phytoestrogen supplementation [IGF-1, median (IQ range), baseline 155 (123, 258), postweek 265 (228, 360) ng/ml, P<.05; IGFBP-3, baseline 3725 (3631, 4196), postweek 4420 (4192, 4935) ng/ml, P<.05]. There was no effect of supplementation on lipids or markers of antioxidant status. Short-term phytoestrogen supplementation increases urinary phytoestrogen excretion and increases IGF-1 and IGFBP-3. These results require elucidation in further controlled studies.

Topics: Adult; Antioxidants; Diet; DNA Damage; Equol; Female; Flax; Genistein; Glycine max; Humans; Insulin-Like Growth Factor Binding Protein 3; Insulin-Like Growth Factor I; Isoflavones; Kinetics; Lignans; Lipids; Middle Aged; Phytoestrogens

To study dietary intake and serum concentrations of isoflavones in order to provide relative validation of isoflavone intake estimates from the Scottish Collaborative Group - Food-Frequency Questionnaire (SCG-FFQ).. Validation study.. Southern Scotland.. Dietary intake of isoflavones was estimated using the semiquantitative SCG-FFQ and rank correlation and Kappa statistics were used for the relative validation of intakes against serum isoflavone concentrations in 203 male participants who were population controls in a case-control study of diet and prostate cancer.. The median intake of isoflavones (daidzein and genistein) was 1.0mg/day (l-QR 0.6-1.8). The median serum concentration of genistein was 33.79 nmol/l (I-QR 14.12-64.93), nearly twice that of daidzein (18.00 nmol/l, I-QR 8.26-29.45). Equol was detected in 49% of subjects; in these subjects the median was 0.67 nmol/l (I-QR 0.34-1.51). Isoflavone intake was significantly correlated with serum concentrations of daidzein (p = 0.24, P = 0.001), genistein (p = 0.26, P < 0.001) and total isoflavonoids (sum of daidzein, genistein and equol) ( p = 0.27, P < 0.001). Whereas values for weighted Kappa ranged from 0.16 (P = 0.002) for daidzein and equol combined to 0.22 (P < 0.001) for genistein.. These results demonstrate the suitability of the SCG-FFQ to rank usual isoflavone intakes in older Scottish men, a population observed to have low consumption of soy foods.

Topics: Aged; Biomarkers; Cross-Sectional Studies; Diet Surveys; Equol; Genistein; Glycine max; Humans; Isoflavones; Male; Middle Aged; Phytoestrogens; Reproducibility of Results; Scotland; Surveys and Questionnaires

The estrogenic and antioxidant effects of the phytoestrogen daidzein (DAI) on germ cell proliferation were evaluated by a chicken ovarian germ-somatic cell coculture model. Ovarian cells were dispersed from 18-day-old embryos, cultured in serum-free McCoy's 5A medium and challenged with DAI alone or in combinations with estrogen receptor antagonist tamoxifen for 48 h. The number of germ cells was counted and the proliferating cells were identified by immunocytochemistry of proliferating cell nuclear antigen (PCNA). The labeling index (LI) was determined for germ cells. Results showed that DAI significantly increased the number of germ cells (P<0.05) and this stimulating effect was inhibited by tamoxifen in a dose-dependent manner. Furthermore, PCNA-LI of germ cells displayed similar changes with the number of germ cells. To estimate the antioxidant action of DAI, ovarian cells were exposed to the reactive oxygen species (ROS)-producing system hypoxanthine/xanthine oxidase (HX/XO). The changes of superoxide dismutase (SOD) activity and glutathione (GSH) level were measured for estimation of the antioxidant status. Ovarian cells were severely damaged by free radicals and this deteriorating effect could be prevented by DAI. Moreover, HX/XO-induced decrease in SOD activity and GSH level was restored by DAI (P<0.05). These results indicated that DAI promoted proliferation of cultured ovarian germ cells by estrogenic action and attenuated ROS-induced toxicity by antioxidant action in embryonic chickens.

Topics: Animals; Antioxidants; Cell Growth Processes; Chick Embryo; Estrogen Antagonists; Female; Glutathione; Immunohistochemistry; Isoflavones; Microscopy, Phase-Contrast; Ovary; Ovum; Phytoestrogens; Proliferating Cell Nuclear Antigen; Reactive Oxygen Species; Superoxide Dismutase; Tamoxifen; Xanthine Oxidase

High consumption of soy isoflavones in Asian diets has been correlated with a lower incidence of clinically important cases of prostate cancer. The chemopreventive properties of these diets may result from an interaction of several types of isoflavones, including genistein and daidzein. The present study investigated the effects of a soy isoflavone concentrate (ISF) on growth and gene expression profiles of PC-3 human prostate cancer cells. Trypan blue exclusion and [3H]-thymidine incorporation assays showed that ISF decreased cell viability and caused a dose-dependent inhibition of DNA synthesis, respectively, with 50% inhibition (IC50) of DNA synthesis at 52 mg/L (P = 0.05). The glucoside conjugates of genistein and daidzein in ISF were converted to bioactive free aglycones in cell culture in association with the inhibition of DNA synthesis. Flow cytometry and Western immunoblot analyses showed that ISF at 200 mg/L caused an accumulation of cells in the G2/M phase of the cell cycle (P < 0.05) and decreased cyclin A by 20% (P < 0.05), respectively. The effect of ISF on the gene expression profile of PC-3 cells was analyzed using Affymetrix oligonucleotide DNA microarrays that interrogate approximately 17,000 human genes. Of the 75 genes altered by ISF, 28 were upregulated and 47 were downregulated (P < 0.05). Further analysis showed that IL-8, matrix metalloproteinase 13, inhibin beta A, follistatin, and fibronectin mRNA levels were significantly reduced, whereas the expression of p21(CIP1), a major cell cycle inhibitory protein, was increased. The effects of ISF on the expression of IL-8 and p21(CIP1) mRNA and protein were validated at high and low ISF concentrations. Our data show that ISF inhibits the growth of PC-3 cells through modulation of cell cycle progression and the expression of genes involved in cell cycle regulation, metastasis, and angiogenesis.

Topics: DNA; Gene Expression Regulation, Neoplastic; Genistein; Humans; Interleukin-8; Isoflavones; Male; Oligonucleotide Array Sequence Analysis; Phytoestrogens; Prostatic Neoplasms; Soybean Proteins; Tumor Cells, Cultured

Genistein and daidzein are the main isoflavones in legumes. Equol is an intestinal bacterial metabolite of daidzein. In this study, we evaluated the estrogenic potential of daidzein and synthetic (+/-)-equol to stimulate growth of estrogen-dependent breast cancer (MCF-7) in vitro and in vivo. We hypothesize that estrogenic effects of daidzein and (+/-)-equol could modulate the growth of MCF-7 cells both in vitro and also once implanted into ovariectomized athymic mice. At concentrations between 0.001 and 50 microM, daidzein and (+/-)-equol stimulated the growth of MCF-7 cells with maximal stimulation at 1 muM in vitro. To evaluate their effects on the growth of MCF-7 cells implanted in ovariectomized athymic mice, two dietary dose-response studies [daidzein (125, 250, 500 and 1000 p.p.m.) and (+/-)-equol (250, 500 and 1000 p.p.m.)] were conducted. Tumor size and body weight were monitored weekly during the study. At completion of the study, we analyzed cellular proliferation of tumors using immunohistochemical staining (ki-67), pS2 expression in tumors using a real time quantitative reverse transcription-polymerase chain reaction (qRT-PCR), and total daidzein and (+/-)-equol levels in plasma using liquid chromatography-electrospray tandem mass spectrometry (LC-ES/MS/MS). Dietary daidzein had a slight but significant stimulatory effect on MCF-7 tumor growth in mice. No significant induction of pS2 mRNA (an estrogen-responsive marker) in tumors by dietary daidzein was observed. Total plasma daidzein concentrations in plasma were between 0.25 and 1.52 microM. Dietary equol treatment (for 37 weeks) did not stimulate MCF-7 tumor growth. There were no statistical differences in tumor size, proliferation and pS2 expression among any treatment groups. Total equol concentrations in plasma were 2.10-3.21 microM. In conclusion, daidzein and (+/-)-equol have proliferative effects on MCF-7 cell growth in vitro within the concentration range tested. Dietary daidzein had a slight but significant stimulatory effect on tumor growth, whereas (+/-)-equol did not stimulate the growth of estrogen-dependent breast tumor growth in athymic mice, increase the cell proliferation in tumors, or induce an estrogen-responsive pS2 expression. Total daidzein or (+/-)-equol plasma levels in mice fed the isoflavones were in the range that stimulated MCF-7 cell growth in vitro. These results suggest that pharmacokinetic and/or metabolic factors attenuate the estrogenic effects of daidzein

Topics: Animals; Breast Neoplasms; Cell Proliferation; Disease Progression; Dose-Response Relationship, Drug; Equol; Female; Humans; Isoflavones; Mice; Mice, Nude; Ovariectomy; Phytoestrogens; Transplantation, Heterologous

The principal phyto-oestrogens (PO) in food are isoflavones, lignans, coumestans and prenylated flavonoids, with isoflavones and lignans being the most commonly found in UK diets. Until recently obtaining accurate data on the PO content of foods was hampered by lack of suitable analytical methods and validation techniques. Furthermore, although PO data exist for some foods, these foods may not be available in the UK. The aim of the present study was to construct a new, comprehensive isoflavone (total genistein + daidzein) database. Using data, mainly from recent GC-MS analysis, for approximately 300 foods available in the UK, and extensive recipe calculations, a new database was constructed containing approximately 6000 foods allocated an isoflavone value. By analysing 7 d weighed food diaries, the database was subsequently used to estimate isoflavone intake in two groups of healthy volunteers, omnivores (n 9) and vegetarians (n 10). Mean isoflavone intake in the vegetarian and omnivorous group was 7.4 (sem 3.05) and 1.2 (sem 0.43) mg/d, respectively. Mean intake for the total group was 4.5 (sem 1.89) mg/d. Main food sources of isoflavones for the vegetarian group were soya milk (plain), meat-substitute foods containing textured vegetable protein and soya protein isolate, soya mince, wholemeal bread and rolls, white bread and rolls, croissants and pitta breads, beans, raisins and soya sauce. Main food sources of isoflavones for the omnivorous group were soya yogurts, wholemeal bread and rolls, white bread and rolls, garlic bread, nan bread and brown bread, sultanas and scones.

Topics: Adult; Aged; Bread; Cooking; Databases, Factual; Diet, Vegetarian; Female; Food Analysis; Genistein; Humans; Isoflavones; Male; Middle Aged; Patient Compliance; Phytoestrogens; Soy Foods

To investigate human sperm responsiveness to the estrogenic xenobiotic genistein and seek further information regarding the mechanism of action of estrogenic xenobiotics using mouse spermatozoa.. Uncapacitated human spermatozoa were incubated with genistein and assessed using chlortetracycline (CTC) fluorescence. CTC was also used to evaluate mouse sperm responses to daidzein and combinations of genistein, 8-prenylnaringenin and nonylphenol. Several steroids were tested to determine structure-function relationships, and possible involvement of cAMP and G proteins in responses was also investigated.. Genistein significantly accelerated capacitation and acrosome loss in human spermatozoa, with 1, 10 and 100 nmol/l being equally effective. In mouse spermatozoa, daidzein produced significant responses, and combinations of xenobiotics at low concentrations were more effective than used singly. The compounds appear to act at the cell surface, and responses to three different steroids were nonidentical. A protein kinase-A inhibitor blocked responses to xenobiotics, while genistein and nonylphenol significantly stimulated cAMP production. Pertussis toxin and dideoxyadenosine blocked responses, suggesting involvement of inhibitory G proteins and membrane-associated adenylyl cyclases.. Human and mouse sperm responses to genistein are very similar, but human gametes appear to be even more sensitive. The mechanism of action may involve unregulated stimulation of cAMP production, leading to significant acrosome loss, undesirable because already acrosome-reacted cells are nonfertilizing. Xenobiotics were even more effective in combination. Since simultaneous exposure to low concentrations of multiple xenobiotics is likely to occur in animals and humans, further investigation is needed to determine whether this could impair fertility.

Topics: Acrosome; Animals; Cyclic AMP; Dideoxyadenosine; Estradiol; Flavanones; Genistein; Humans; Isoflavones; Male; Mice; Pertussis Toxin; Phenols; Phytoestrogens; Protein Kinase Inhibitors; Protein-Tyrosine Kinases; Sperm Capacitation; Spermatozoa; Steroids; Xenobiotics

Breast cancer is the most common cancer in women and a significant cause of death. Mutations of the oncosuppressor genes BRCA1 and BRCA2 are associated with a hereditary risk of breast cancer, and dysregulation of their expression has been observed in sporadic cases. Soya isoflavones have been shown to inhibit breast cancer in studies in vitro, but associations between the consumption of isoflavone-containing foods and breast cancer risk have varied in epidemiological studies. Soya is a unique source of the phytoestrogens daidzein (4',7-dihydroxyisoflavone) and genistein (4',5,7-trihydroxyisoflavone), two molecules that are able to inhibit the proliferation of human breast cancer cells in vitro. The aim of the present study was to determine the effects of genistein (5 microg/ml) and daidzein (20 microg/ml) on transcription in three human breast cell lines (one dystrophic, MCF10a, and two malignant, MCF-7 and MDA-MB-231) after 72 h treatment. The different genes involved in the BRCA1 and BRCA2 pathways (GADD45A, BARD1, JUN, BAX, RB1, ERalpha, ERbeta, BAP1, TNFalpha, p53, p21Waf1/Cip1, p300, RAD51, pS2, Ki-67) were quantified by real-time quantitative RT-PCR, using the TaqMan method and an ABI Prism 7700 Sequence Detector (Applied Biosystems). We observed that, in response to treatment, many of these genes were overexpressed in the breast cancer cell lines (MCF-7 and MDA-MB-231) but not in the dystrophic cell line (MCF10a).

Topics: Anticarcinogenic Agents; Breast Neoplasms; Cell Line, Tumor; Female; Gene Expression Regulation, Neoplastic; Genes, BRCA1; Genes, BRCA2; Genes, Tumor Suppressor; Genistein; Glycine max; Humans; Isoflavones; Phytoestrogens; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; RNA, Neoplasm; Transcription, Genetic

Thyroid hormones and thyroid autoantibodies, along with serum concentrations of two phytoestrogens of the isoflavone series, daidzein and genistein, were measured in 268 children without overt thyroid diseases, screened for iodine deficiency in one region of the Czech Republic. Since both phytoestrogens have been reported to inhibit thyroid hormone biosynthesis and in high concentrations to exert goitrogenic effects, we investigated whether their presence in the circulation could influence thyroid hormone function in a population where soy consumption is not common. Correlation analysis revealed a significant positive association of genistein with thyroglobulin autoantibodies and a negative correlation with thyroid volume. Multiple regression analysis of the relationships between actual phytoestrogen levels and measured thyroid parameters revealed only a weak but significant association between genistein and thyroid variables. Higher levels of free thyroxine were found in a subgroup of 36 children who ate soy food in the previous 24 h. In conclusion, only modest association was found between actual phytoestrogen levels and parameters of thyroid function. On the other hand, even small differences in soy phytoestrogen intake may influence thyroid function, which could be important when iodine intake is insufficient.

Topics: Adolescent; Autoantibodies; Child; Czech Republic; Deficiency Diseases; Female; Genistein; Humans; Iodine; Isoflavones; Male; Mass Screening; Phytoestrogens; Soy Foods; Thyroid Function Tests; Thyroid Gland

Estrogens modulate the transcription of sensitive genes either via binding of the activated ER to responsive elements in their promoter region or via binding of the activated ER to transcription factors like NFkappaB. In this study we have analyzed the effects of the phytoestrogens daidzein, coumestrol and genistein in promoter-specific reporter gene systems. The dose-dependent ability to stimulate an ERE-bearing reporter in MVLN breast cancer cells was compared to the dose-dependent ability to repress the IL-1beta-stimulated reporter in U2OS osteosarcoma cells. Coumestrol, daidzein and genistein stimulate the expression of the ERE-dependent reporter in MVLN cells and repress the activity of the IL-6 promoter in U2OS cells in a dose-dependent manner. Interestingly, the relative potency of all phytoestrogens to repress the activity of the IL-6 promoter in U2OS cells was much higher than their potency to stimulate the ERE-dependent reporter in MVLN cells. We assume that the demonstrated promoter-specific potency therefore could be an important mechanism to explain a tissue-specific action of some of these compounds.

Topics: Cell Line, Tumor; Coumestrol; Dose-Response Relationship, Drug; Genes, Reporter; Genistein; Humans; Isoflavones; Phytoestrogens; Promoter Regions, Genetic; Transcription, Genetic

Women approaching menopause increasingly investigate alternatives to hormone replacement therapy. Plant phytoestrogens are being promoted as "natural" alternatives but there is a lack of substantive data to advocate their safe use in breast cancer patients receiving tamoxifen (TAM), or in those who have relapsed. The aim of our study was to investigate the proliferative effects and mode of action of the phytoestrogens genistein, daidzein and coumestrol on TAM-sensitive (-s) and resistant (-r) breast cancer cells under in vitro conditions designed to mimic the hormonal environment of the pre- and post-menopausal breast. At physiological concentrations (<10 microM) and under reduced estrogen (E2) conditions, genistein was mitogenic to TAM-s cells with TAM-r cells generally refractory. Daidzein and coumestrol were growth stimulatory irrespective of TAM sensitivity. Transcriptional activity was ERE-mediated. Combining phytoestrogens with E2 (simulating the pre-menopausal breast environment) had no effect on growth of TAM-s or TAM-r cells. Addition of 4-HT mimicked the hormonal environment in post-menopausal breast cancer patients receiving TAM. The growth inhibitory effects of 4-HT were abrogated in TAM-s cells when combined with genistein and coumestrol, and to a lesser extent, daidzein, where significant growth stimulatory effects were observed. In TAM-r cells, proliferation did not exceed control values. At phytoestrogen concentrations above 10 microM, growth inhibitory effects were seen, irrespective of estrogenic environment or cell sensitivity to TAM. Our in vitro data suggests that phytoestrogens could have potentially adverse mitogenic effects on tumour cells and should probably be avoided by patients who remain sensitive to TAM or in those with pre-existing and possibly undiagnosed breast tumours.

Topics: Antineoplastic Agents, Hormonal; Breast Neoplasms; Cell Line, Tumor; Cell Proliferation; Coumestrol; Dose-Response Relationship, Drug; Drug Resistance, Neoplasm; Estrogen Receptor alpha; Estrogen Receptor beta; Estrogens; Female; Gene Expression; Genistein; Humans; Isoflavones; Luciferases; Phytoestrogens; Receptors, Progesterone; Response Elements; Tamoxifen; Transfection

The aim of this study was to examine ganders at different stages of the reproductive season and the effect of: (1) diets with high phytoestrogen content and (2) in vitro phytoestrogen treatment on testosterone secretion by isolated Leydig cells. Thirty-six male Biłgoraj geese were fed control diets with low phytoestrogen content (containing grass meal) and diets with high phytoestrogens (containing alfalfa meal and soy). Testes were obtained from both groups of ganders at three different times of the breeding season: peak of reproductive activity (March), second half of reproductive activity (May) and beginning of photorefractoriness (July). Isolated Leydig cells were incubated with LH as well as genistein, daidzein, equol and coumestrol and the concentration of testosterone in the medium was determined by radioimmunoassay. The mean weight of testes from ganders in May and July decreased relative to their weights in March, but no significant differences among experimental groups were noted. No differences were observed in basal and LH-stimulated testosterone secretion by Leydig cells of ganders fed the control diets and the diets with higher phytoestrogen content. In July, LH did not stimulate testosterone secretion in either group. In vitro treatment with genistein, daidzein and equol (5 and 50 microM) inhibited basal and LH-stimulated testosterone production by Leydig cells from both groups. Coumestrol (5 and 50 microM) inhibited basal testosterone secretion only in March in the control group. Dietary exposure to phytoestrogens had a slight effect on in vitro testicular secretion in ganders. In vitro treatment with phytoestrogen inhibited testosterone production by Leydig cells. Genistein showed the strongest effect and coumestrol had the weakest influence on testicular secretion.

Topics: Animal Feed; Animals; Dose-Response Relationship, Drug; Equol; Geese; Genistein; Isoflavones; Leydig Cells; Male; Organ Size; Phytoestrogens; Radioimmunoassay; Random Allocation; Reproduction; Seasons; Testosterone

This study examines whether anti-diabetic effects of genistein and daidzein are mediated by hepatic glucose and lipid regulating enzyme activities in type 2 diabetic animals. Male C57BL/KsJ-lepr(db)/lepr(db) (db/db) mice and age-matched non-diabetic littermates (db/+) were used in this study. The db/db mice were divided into control, genistein (0.02%, w/w) and daidzein (0.02%, w/w) groups. The blood glucose and HbA(1c) levels were significantly lower in the genistein and daidzein groups than in the control group, while glucose tolerance only was significantly improved in the genistein-supplemented group. The plasma insulin and C-peptide levels did not differ significantly between groups, yet the glucagon level was lower in the genistein and daidzein groups compared to that in the control db/db or db/+ group. The genistein and daidzein supplements increased the insulin/glucagon ratio in the type 2 diabetic animals. While the hepatic glucokinase activity was significantly lower in the db/db control group, the glucose-6-phosphatase and phosphoenolpyruvate carboxykinase activities were significantly higher in the control group compared to the db/+ group. Interestingly, these hepatic glucose metabolizing enzyme activities were reversed by the genistein and daidzein supplementation in db/db mice compared to the control group. The hepatic fatty acid synthase, beta-oxidation and carnitine palmitoyltransferase activities were all significantly lower in the genistein and daidzein groups than in the control group. The genistein and daidzein supplements also improved the plasma total cholesterol, triglyceride, HDL-cholesterol/total cholesterol, free fatty acid and hepatic triglyceride concentrations in the db/db mice. These results suggest that genistein and daidzein exert anti-diabetic effect in type 2 diabetic conditions by enhancing the glucose and lipid metabolism.

Topics: Animals; Body Weight; C-Peptide; Diabetes Mellitus, Type 2; Diet; Eating; Genistein; Glucagon; Glucose; Glucose Tolerance Test; Glycated Hemoglobin; Insulin; Isoflavones; Leptin; Lipid Metabolism; Liver; Liver Glycogen; Mice; Mice, Inbred C57BL; Phytoestrogens; Receptors, Cell Surface; Receptors, Leptin

In veal calf production plant-based proteins are frequently included in milk replacer fed to the animals. Since soy products, which are mostly used, are known for their high levels of phyto-oestrogens, the effects of these feeds on the veal calf prostate were examined. Goal was to determine whether these compounds could interfere with histological screening for oestrogenic growth promoters. In a feeding experiment, four groups of veal calves fed plant-based protein-supplemented milk replacer (PBM), containing 5% soy concentrate, 5% soy isolate, 5% wheat gluten and 2% potato protein, for 4 weeks were compared to animals fed dairy-based control feed (DBM); animals treated with estradiol benzoate, diethylstilbestrol and ethinylestradiol served as positive controls. Daidzein and genistein levels measured in feed and urine showed high levels of genistein and daidzein in the soy isolate and soy concentrate supplemented feeds. Genistein and daidzein were also found in the urine of the animals that were fed these feeds. Haematoxylin-eosin-stained prostate sections of PBM-fed animals showed slight hyperplasia and some dilated tubules as compared to the DBM-fed group, but no metaplasia, which is used for screening for oestrogenic hormones. The positive controls showed extensive squamous metaplasia. Immunohistochemical staining for cytokeratin 5 (using RCK 103 monoclonal antibody) in basal cells showed a normal staining pattern of basal cells in the DBM-fed calves and extensive basal cell proliferation and squamous metaplasia in the oestrogen-treated positive control animals. PBM-fed calves showed no increase of basal cell staining but showed elongations of the basal cells in most animals, sometimes resulting in circular figures. It is concluded that the feeds examined in this study did not interfere with histological screening for oestrogens in male veal calves.

Topics: Animal Feed; Animal Nutritional Physiological Phenomena; Animals; Animals, Newborn; Cattle; Estrogens, Non-Steroidal; Genistein; Glycine max; Immunohistochemistry; Isoflavones; Male; Phytoestrogens; Prostate; Random Allocation; Weight Gain

The present study utilized microarray technology as a tool to elucidate the molecular signatures of soy-derived phytochemicals in the human androgen-responsive prostate cancer cell line LNCaP. Global gene expression pattern analysis of LNCaP cells exposed to 0, 1, 5, or 25 microM of the soy-derived phytochemicals equol and daidzein were conducted and compared. The data were further compared with previously generated data from exposure of LNCaP cells to the same doses of genistein, a soy isoflavone. Multidimensional scaling (MDS) analyses of the expression patterns suggest that these compounds exerted differential effects on gene expression in LNCaP cells. Further examination of specific gene changes revealed that these compounds differentially modulated genes in multiple cellular pathways, including the cell-cycle pathway genes. However, the three compounds also exerted similar effect on genes belonging to several other important cellular pathways. A universal effect of the three compounds on androgen-responsive genes, IGF-1 pathway gene, and MAP kinase-related pathway gene was observed. These results provide the foundation for establishing molecular signatures for equol, daidzein, and genistein. Moreover, these results also allow for the identification of candidate mechanism(s) by which soy phytochemicals and soy may act in prostate cancer cells.

Topics: Androgens; Cell Cycle; Cell Line, Tumor; Equol; Gene Expression; Gene Expression Profiling; Genistein; Glycine max; Humans; Insulin-Like Growth Factor I; Isoflavones; Male; Neoplasms, Hormone-Dependent; Oligonucleotide Array Sequence Analysis; Phytoestrogens; Prostatic Neoplasms; Receptor, IGF Type 1

Epidemiological studies indicate that Asian women have a lower incidence of breast cancer compared with their counterparts in the West, and soya consumption has been suggested as a contributory factor. Clinical and animal studies have revealed that cyclooxygenase-2 (COX-2) expression is associated with a risk of breast cancer. In the present study, we investigated the effect of soya isoflavones on the expression of COX-2 in the breast cell line MCF-7. Genistein, daidzein and equol were found to inhibit COX-2 expression induced by phorbol 12-myristate 13-acetate (PMA). Similar findings were observed in the COX-2 protein analysis. In order to study transcriptional control, a fragment of the 5'-flanking region of the hCOX-2 gene was amplified and inserted into a firefly luciferase reporter plasmid. The reporter assay indicated that the transactivation of the hCOX-2 promoter was induced by PMA, and activity was inhibited with the co-administration of genistein, daidzein or equol. An activator protein-1 (AP-1)/cyclic AMP response element binding protein (CREB) binding site (-59/-53) was identified in hCOX-2 promoter, and this could be critical in PMA-induced COX-2 expression. Truncation reporter plasmids with (-70/-36) and without (-51/-36) AP-1/CREB were constructed for subsequent analysis. The results revealed that the hCOX-2 promoter transactivation suppressed by isoflavone could be dependent on AP-1/CREB binding. Nonetheless, this study illustrated that the soya isoflavones reduced COX-2 expression, which could be important in the post-initiation events of breast carcinogenesis.

Topics: Anticarcinogenic Agents; Breast Neoplasms; Carcinogens; Cell Line, Tumor; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; Equol; Female; Genistein; Glycine max; Humans; Isoflavones; Membrane Proteins; Phytoestrogens; Plant Extracts; Promoter Regions, Genetic; Tetradecanoylphorbol Acetate

The effect of daidzein, an important isoflavone, on the differentiation and mineralization in MC3T3-E1 cells, a mouse calvaria osteoblast-like cell line, was investigated. The MTT assay, the alizarin red S and von Kossa staining, the measurement of calcium (Ca) and phosphorus (P) concentrations by inductively coupled plasma-atomic emission spectrometry and the nitrophenol liberation method were used to determine the cell proliferation, mineralization and intracellular alkaline phosphatase (ALP) activity, respectively. Daidzein enhanced the cell proliferation after the culture for 2 days and the effect reached maximum on day 6. ALP activity and cellular Ca and P contents were increased time- and dose-dependently when the cells were treated with daidzein in the presence of disodium beta-glycerophosphate and L-ascorbic acid. Differentiation of the cells to the mature osteoblasts was prompted under incubation in the presence of daidzein for 21 days, by the time the mineralized nodules formed. The calcium depositions of the cells by alizarin red S staining were increased significantly after the culture with daidzein as long as 28 days. It has been demonstrated that daidzein may be able to enhance the bone differentiation and mineralization and prompt the bone formation in the early growing stage and the late growing stage of osteoblasts.

Topics: Alkaline Phosphatase; Animals; Calcification, Physiologic; Calcium; Cell Differentiation; Cells, Cultured; Dose-Response Relationship, Drug; Isoflavones; Mice; Osteoblasts; Phosphorus; Phytoestrogens; Stimulation, Chemical; Time Factors

The soy isoflavones genistein and daidzein, as well as the daidzein metabolite equol, have structural similarities to mammalian estrogens and bind with varying affinity to both known subtypes of the estrogen receptor. Consequently, prospective studies in both humans and animals have begun to evaluate the potential effects of isoflavones on estrogen receptor-mediated phenomena. However, many diets of laboratory-housed animals derive their protein from soy and thus likely contain substantial quantities of isoflavones. Exposing experimental subjects to these isoflavones via such diets could confound studies, particularly those evaluating the effects of estrogen or estrogen-like ligands. The aim of this study was to compare the levels of circulating concentrations of isoflavones and their metabolites in monkeys fed either a soy-free diet, a soy-based diet providing 130 mg of isoflavone (daidzein, genistein, and glycitein aglycon equivalents) daily, or a commercially available 'chow' diet containing an unspecified amount of soybean meal. Animals consuming the commercial diet had serum concentrations of daidzein, genistein, and glycitein that were significantly higher than those of animals fed a soy-free diet but similar to those of monkeys fed a soybased diet formulated to be high in isoflavones. Notably, animals fed the commercial diet also had serum equol concentrations that were similar to or, in some cases, in excess of serum concentrations in the animals fed the soy diet. These data argue for the use of soy-free diets in studies investigating estrogenic effects on physiologic or behavioral endpoints.

Topics: Animal Nutritional Physiological Phenomena; Animals; Diet; Female; Genistein; Isoflavones; Laboratory Animal Science; Macaca fascicularis; Macaca mulatta; Male; Phytoestrogens; Soy Foods

It has been postulated that the R- and S-equol enantiomers have different biological properties given their different binding affinities for the estrogen receptor. S-(-)equol is produced via the bacterial conversion of the soy isoflavone daidzein in the gut. We have compared the biological effects of purified S-equol to that of racemic (R and S) equol on breast and prostate cancer cells of varying receptor status in vitro. Both racemic and S-equol inhibited the growth of the breast cancer cell line MDA-MB-231 (> or = 10 microM) and the prostate cancer cell lines LNCaP (> or = 5 microM) and LAPC-4 (> or = 2.5 microM). The compounds also showed equipotent effects in inhibiting the invasion of MDA-MB-231 and PC-3 cancer cells through matrigel. S-equol (1, 10, 30 microM) was unable to prevent DNA damage in MCF-7 or MCF-10A breast cells following exposure to 2-hydroxy-4-nonenal, menadione, or benzo(a)pyrene-7,8-dihydrodiol-9,10-epoxide. In contrast, racemic equol (10, 30 microM) prevented DNA damage in MCF-10A cells following exposure to 2-hydroxy-4-nonenal or menadione. These findings suggest that racemic equol has strong antigenotoxic activity in contrast to the purified S-equol enantiomer implicating the R-, rather than the S-enantiomer as being responsible for the antioxidant effects of equol, a finding that may have implications for the in vivo chemoprotective properties of equol.

Topics: Antineoplastic Agents; Breast Neoplasms; Cell Division; Cell Line, Tumor; Comet Assay; DNA Damage; DNA, Neoplasm; Dose-Response Relationship, Drug; Equol; Female; Humans; Isoflavones; Male; Phytoestrogens; Prostatic Neoplasms; Receptors, Estrogen; Stereoisomerism

Tumor metastasis is associated with integrin-mediated adhesion and hyaluronan receptor expression. Accumulating evidence suggests that phytoestrogens, which are naturally occurring, plant-derived phytochemicals, could inhibit tumorigenesis during the development of breast cancer. Less is known, however, about the regulation of adhesion receptors by phytoestrogens and, particularly, their potency to influence proliferation of primary human breast cells in comparison with the steroid hormone 17beta-estradiol. Throughout the proliferation experiments, we used primary human mammary epithelial cells from normal tissue that was derived from plastic surgery. For receptor expression (beta1, alpha2, alpha3, CD44), we used the cell line MCF-7. Both investigations were carried out by flow cytometry. The phenotype of primary human mammary epithelial cells was microscopically characterized by analyzing the distribution of ZO-1, cytokeratin and the estrogen receptors alpha and beta. The integrins and the hyaluronan receptor were significantly up-regulated with 17beta-estradiol in human MCF-7 cells. In contrast, genistein and daidzein did not affect the expression at a concentration of 100 micromol/l. In all proliferation experiments with a significant stimulation of the primary human mammary epithelial cell growth due to 17beta-estradiol, in general, genistein and daidzein did not influence S-phase and G2/M-phase cells. Additionally, the stimulative effect of 17beta-estradiol could be inhibited. As the phytoestrogens do not up-regulate adhesion receptors in human breast cells and, regarding proliferation, are able to abolish the stimulatory effect of 17beta-estradiol, we suggest that phytoestrogens could have beneficial effects for the prevention or inhibition of carcinogenesis in hormone-dependent malignancies.

Topics: Cell Line, Tumor; Cell Proliferation; Cells, Cultured; Epithelial Cells; Female; Genistein; Humans; Isoflavones; Keratins; Mammary Glands, Human; Membrane Glycoproteins; Membrane Proteins; Phosphoproteins; Phytoestrogens; Platelet Glycoprotein GPIb-IX Complex; Receptors, Estrogen; Tight Junctions; Zonula Occludens-1 Protein

Dietary isoflavones, such as genistein and daidzein, are metabolised by the human gut microflora. Case-control studies have disclosed a link between the formation of the daidzein metabolite equol and prostate cancer risk. We evaluated the effects of genistein, daidzein and five metabolites on two prostate cancer cell lines by determining DNA integrity and cell growth. LNCaP cells contain the T877A androgen receptor mutation whereas Los Angeles prostate cancer (LAPC)-4 cells express the wild-type receptor, both of which may affect responses to isoflavones. DNA integrity was determined using the comet assay. Cell growth was assessed by staining DNA with 4',6'-diamidino-2-pheylindole hydrochloride. Endogenous steroid hormones, but not isoflavones, induced DNA strand breaks. Dihydrotestosterone stimulated the growth of both cell lines. 17beta-Oestradiol increased the growth of LNCaP but not LAPC-4 cells, pointing to an involvement of the T877A androgen receptor. Isoflavones did not stimulate growth in either prostate cancer cell line. However, the growth of LNCaP and LAPC-4 cells was suppressed by genistein (inhibitory concentration 50 % (IC50) 39.7 mumol/l, 37.2 mumol/l) and by equol (IC50 53.8 mumol/l, 35.1 mumol/l). O-desmethylangolensin inhibited the growth of LAPC-4 cells (IC50 45.2 mumol/l), but not of LNCaP cells. In conclusion, isoflavones do not damage DNA or promote growth of androgen-dependent prostate cancer cells. Several isoflavones, including the reduced daidzein metabolites equol and O-desmethylangolensin, suppress cancer cell growth. Taken together, these data suggest a contribution of gut-formed isoflavone metabolites to the beneficial effects of dietary isoflavones on prostate cancer risk.

Topics: Androgens; Antineoplastic Agents; Cell Division; Cell Line, Tumor; Comet Assay; Dihydrotestosterone; DNA Damage; DNA, Neoplasm; Estradiol; Genistein; Humans; Intestines; Isoflavones; Male; Phytoestrogens; Prostatic Neoplasms

Equol, a derivative of daidzein produced by enterobacteria, has greater activity as a phyto-oestrogen compared with daidzein. Difructose anhydride III (DFAIII) is a newly manufactured non-digestible disaccharide with unique fermentation properties. The present study evaluated the prebiotic effects of DFAIII on equol production and on plasma cholesterol concentrations related to the changes in equol production. We compared plasma equol concentrations at 10.00 and 18.00 hours and faecal isoflavone excretion in three groups of seven rats (male Wistar-ST strain, 6 weeks old) fed a basal diet or a DFAIII or fructooligosaccharide (15 g/kg diet) diet containing 1 g soya isoflavones/kg diet for 20 d. Equol concentrations in the DFAIII group were higher than in the control and fructooligosaccharides groups, especially in the later phase of the light period (18.00 hours) throughout the experiment. Daizein and genistein concentrations did not change between the diet groups. The faecal ratios of equol:daidzein were very high in all groups, but the ratios were higher in the DFAIII group than the control and fructooligosaccharide groups on day 3, and this tendency continued throughout the experiment. On day 20, the plasma total cholesterol concentration was lowest in the DFAIII group. Additionally, the cholesterol concentrations were inversely correlated to plasma equol concentration in all the rats. In conclusion, DFAIII efficiently enhanced plasma equol concentrations, which may be associated with an increase in equol production and a decrease in equol degradation by enterobacteria. Higher plasma equol concentrations may contribute to the hypocholesterolaemic effect of DFAIII feeding.

Topics: Animals; Body Weight; Cecum; Cholesterol; Diet; Dietary Supplements; Disaccharides; Equol; Feces; Genistein; Isoflavones; Male; Oligosaccharides; Phytoestrogens; Rats; Rats, Wistar

Although estrogen replacement has been the main therapy to prevent and treat osteoporosis, there are concerns about its safety. Phytoestrogens have attracted attention to their potential impacts in osteoporosis prevention and treatment. Among phytoestrogens, the isoflavone daidzein (Dz) acts on transcription via the intracellular estrogen receptors (ER), mainly ERbeta, in osteoblasts, but mimics only part of the estrogen effects. Since estradiol also exerts rapid effects in osteoblasts, we investigated the multistep processes involved in the rapid actions of low (1-100 pM) doses of daidzein. Dz bound to a membrane moiety, related to ERbeta since the calcium response to Dz was blocked by an anti-ERbeta antibody directed against the C-terminus, but not by a double-stranded siRNA specific for ERbeta. This protein was coupled to a pertussis toxin (PTX)-sensitive Gbeta1 subunit whose transducer was PLC-beta2, which triggered a rapid (5 sec) mobilization of calcium from the endoplasmic reticulum. Dz phosphorylated within 15 sec ERK1/2 whose phosphorylation involved two routes: Gbeta1/PLC-beta2/PKC/c-Raf-1/MEK1/2 and Gbeta1/PI3K/cSrc/c-Raf-1/MEK1/2 as shown using several inhibitors. Dz induced rapid (1 min) changes in the actin cytoskeleton via the two routes. The rapid (20 sec) phosphorylation of Elk-1 and CREB by Dz involved Gbeta1 and ERK1/2. All the processes were insensitive to the estradiol antagonist ICI 182,780. In conclusion, the rapid effects of Dz seem to be biologically relevant for the function of osteoblast in bone since the isoflavone activates transcription factors linked to early genes controlling cellular proliferation and differentiation, and modulates actin cytoskeleton which controls cell adhesion, division, or secretion.

Topics: Actins; Animals; Calcium; Cell Membrane; Cytoskeleton; Estrogen Receptor beta; Estrogens; Extracellular Signal-Regulated MAP Kinases; Female; GTP-Binding Protein beta Subunits; Isoenzymes; Isoflavones; Osteoblasts; Phytoestrogens; Protein Kinase C; Rats; Rats, Wistar; Signal Transduction; Transcription Factors; Type C Phospholipases

Ethanol causes extensive damage to the intestinal tract from the oropharynx to the rectum. The jejunum has also been shown to be particularly vulnerable to the deleterious effects of ethanol. We hypothesized that (I) the pathogenesis of acute alcohol-mediated injury in the small intestine involves generation of reactive oxygen species, and consequentially, enhanced lipid peroxidation; (II) the pathogenic changes due to alcohol can be ameliorated with daidzein pretreatment. To test these hypotheses male Wistar rats (n=24) were divided into four groups as follows (pretreatment followed by treatment): [A] carrier+saline (control); [B] daidzein+saline; [C] carrier+ethanol; [D] daidzein+ethanol. Daidzein (100 mg/kg) or carrier (Intralipid) pretreatment was twice administered as a single dose, whereas ethanol (75 mmol/kg) or saline (0.15 mol/l NaCl) treatment was administered once only. At 24 h after ethanol or saline was administered, rats were sacrificed. The analytes 7alpha-and 7beta-hydroperoxycholest-5-en-3beta-ol (7alpha-OOH and 7beta-OOH), 7alpha-and 7beta-hydroxycholesterol (7alpha-OH and 7beta-OH), and 7-ketocholesterol (7-keto) in jejunum were analyzed by HPLC. The data showed that daidzein per se did not affect levels of cholesterol hydroperoxides nor oxysterols. However, there were significant increases in 7alpha- and 7beta-OOHs, 7alpha- and 7beta-OHs, and 7-keto after ethanol dosage compared to controls. Daidzein ameliorated these effects, i.e., values in the daidzein+ethanol group were similar to those in the carrier+saline (control) group. This is the first report showing that (1) cholesterol-derived markers of oxidative stress are increased in the rat jejunum in response to ethanol, indicative of metabolic damage; (2) daidzein pretreatment has protective effects against ethanol-induced injury.

Topics: Animals; Ethanol; Isoflavones; Jejunum; Lipid Peroxidation; Male; Phytoestrogens; Rats; Rats, Wistar

Isoflavones (mainly daidzein and genistin) belong to the flavonoid group of compounds and are classified as phytoestrogens. In the intestine, daidzin is converted to daidzein by beta-glucosidase, and then daidzein is converted to O-desmethylangolensin (O-DMA) or equol via dihydrodaidzein by enzymes of intestinal bacteria. We isolated, for the first time, an anaerobic gram-positive rod-shaped strain capable of producing equol from daidzein. Its 16S rDNA gene sequence (1428 bp) showed 99% similarity with that of the human intestinal bacterium SNU-Julong 732 (AY310748) and 93% similarity with that of Eggerthella lenta ATCC 25559(T) (AF292375). This strain converted daidzein to equol via dihydrodaidzein in an equol-assay medium anaerobically. The addition of butyric acid and arginine increased the conversion ratio of daidzein to equol 4.7- and 4.5-fold, respectively.

Topics: Actinobacteria; Animals; Base Sequence; Equol; Humans; Intestines; Isoflavones; Molecular Sequence Data; Phytoestrogens; Rats; RNA, Ribosomal, 16S

The major constituents of isoflavones, daidzein (DZ) and genistein (GE) are known to interact with the alpha and beta estrogen receptors (ERalpha/beta) in several tissues including mammary. In this study, we used ovariectomy (OVX) to model menopause and determined the effects of DZ, GE or 17beta-estradiol (E2) exposures on chemically induced mutagenesis and carcinogenesis in the mammary glands of female Big Blue (BB) transgenic rats. The rats were fed control diet containing the isoflavones and E2 and treated with a single oral dose of 7,12-dimethylbenz[a]anthracene (DMBA) at PND 50. Animals were sacrificed at 16 or 20 weeks post-carcinogen treatment to assess mutant frequencies (MFs) and histopathological parameters, respectively. The isoflavones or E2 supplementation alone resulted in modest increases in the lacI MF that were not significantly different from the MFs measured in rats fed the control diet alone. DMBA exposure, however, induced significant increases in the lacI MFs in the mammary of both OVX and ovary intact (INT) rats and Hprt MFs in spleen lymphocytes (P

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Animals, Genetically Modified; Female; Genistein; Glycine max; Isoflavones; Mammary Neoplasms, Experimental; Mutagenesis; Ovariectomy; Phytoestrogens; Rats

The aim of this study is to investigate the hypotensive and vasodilator effects of daidzein sulfates, a water-solubility derivative of daidzein. Tail cuff blood pressure of spontaneously hypertensive rat (SHR) was measured with non-invasive Electro-Sphygmomanometer. An isometric tension of rat mesenteric artery ring segments was recoded in vitro on a myograph. The results showed that daidzein sulfates (20 and 40 mg/kg) could decrease blood pressure of SHR in single dose and multi-doses. Daidzein sulfates (1-100 microM) inhibited the contraction of rat mesenteric arterial ring segments induced by norepinephrine (NA) and 5-hydroxytryptamine (5-HT). Daidzein sulfates (100-1000 microM) inhibited arterial segment's contraction induced by KCl and CaCl(2). The concentration- contractive curves were shifted toward right in a non-parallel manner with decreased E(max.) Daidzein sulfaltes inhibited the extracellular Ca(2+)-dependent contraction. Daidzein sulfates of 100 and 300 microM significantly inhibited the contraction induced by CaCl(2) in Ca(2+)-free solution, which is an extracellular Ca(2+)-dependent contraction; but daidzein sulfates did not inhibit the intracellular Ca(2+)-dependent NA-induced contraction, in Ca(2+)-free solution. The results suggest that daidzein sulfates possess significant hypotensive and vasodilator effects which mainly derive from artery smooth muscle cells by inhibiting the receptor-mediated Ca(2+)-influx.

Topics: Animals; Blood Pressure; Calcium; Calcium Chloride; Hypertension; In Vitro Techniques; Isoflavones; Male; Mesenteric Arteries; Norepinephrine; Phytoestrogens; Potassium Chloride; Rats; Rats, Inbred SHR; Rats, Sprague-Dawley; Serotonin; Sulfuric Acid Esters; Vasodilator Agents

To identify the hydroxylate metabolites and its sulfate conjugates of daidzein in rat urine.. Urine samples from 0 - 24 h were collected after single ig dose of 500 mg x kg(-1) daidzein to each of six rats. The urine samples were purified by SPE column (SPE C18) and analyzed with liquid chromatographic-tandem electrospray ionization ion trap mass spectrometry (LC-ESI/MS(n)) for potential metabolites.. Several new hydroxylate metabolites and its sulfate conjugates were found and identified in rat urine.. LC-ESI/MS(n) is proved to be a simple, rapid, sensitive and specific technique for identification of the hydroxylate metabolites and its sulfate conjugates of daidzein in rat urine.

Topics: Animals; Chromatography, Liquid; Glycine max; Hydroxylation; Isoflavones; Male; Molecular Structure; Phytoestrogens; Rats; Rats, Sprague-Dawley; Reproducibility of Results; Seeds; Spectrometry, Mass, Electrospray Ionization; Sulfates; Tandem Mass Spectrometry

The mechanism of phytoestrogen action in gonadal cells of ganders has not been elucidated. The aim of the study was to investigate in Biłgoraj ganders the possibility of phytoestrogen action via estrogen or androgen receptors or via protein tyrosine kinase pathways in Leydig cells. Genistein and daidzein (5 and 50 microM) as well as equol (50 microM) inhibited testosterone (T) secretion by incubated Leydig cells (1x10(5)/ml; 20 h; 37 degrees C). The effects of hydroxytamoxifen (estrogen receptor inhibitor) and cyproterone acetate (androgen receptor antagonist) on phytoestrogen inhibition of T release by Leydig cells were not observed. Lavendustin A (protein tyrosine kinases inhibitor) did not change T production. The influence of phytoestrogens seems not to be conducted via estrogen and androgen receptors or protein tyrosine kinases system in these cells, but further studies are required to completely examine the mechanism of phytoestrogens action in testes of ganders.

Topics: Animals; Equol; Geese; Genistein; Isoflavones; Leydig Cells; Male; Phenols; Phytoestrogens; Poland; Protein-Tyrosine Kinases; Radioimmunoassay; Tamoxifen; Testosterone

The main objective of this study was to compare the protective effect of daidzein and genistein against induced oxidative damage in Jurkat T-cell line and in peripheral blood lymphocytes of healthy subjects. After supplementation of cells with isoflavones (from 2.5 to 20micromol/L in Jurkat T-cell and from 0.01 to 2.5micromol/L in primary lymphocytes, 24h), we determined DNA damage induced by hydrogen peroxide using the comet assay and lipid peroxidation evaluating malondialdehyde (MDA) production after ferrous ion treatment. Supplementation of Jurkat cells and primary lymphocytes with both isoflavones significantly increased DNA protection from oxidative damage at concentrations between 0.1 and 5micromol/L (P<0.05), and with just daidzein, at concentrations higher than 2.5micromol/L, there was a decrease in the production of MDA (P<0.05). Our results seem to support that daidzein is just as effective as genistein in protecting cells against oxidative damage especially with respect to DNA. Moreover, since the protective effect was found at concentrations reachable in plasma after soy consumption (less than 2micromol/L), it can be assumed that the antioxidant activity of isoflavones could really contribute to the healthy properties of soy.

Topics: Antioxidants; Cell Line, Transformed; Cells, Cultured; Comet Assay; Cytotoxicity Tests, Immunologic; DNA Damage; Dose-Response Relationship, Drug; Ferrous Compounds; Genistein; Glycine max; Humans; Hydrogen Peroxide; Isoflavones; Jurkat Cells; Leukocytes, Mononuclear; Lipid Peroxidation; Malondialdehyde; Oxidative Stress; Phytoestrogens; T-Lymphocytes

The study was conducted to investigate the effects of phytoestrogen daidzein on blastocyst implantation in rats. Following successful mating, female rats were given daidzein by subcutaneous administration at the dose of 0 (vehicle control, n=15), 50 mg/kg body weight (n=15) and 150 mg/kg body weight (n=15) daily on day 1-7 of pregnancy and were sacrificed on day 8 of gestation. The results revealed that high-dose treatment (150 mg/kg body weight) significantly diminished the rate of blastocyst implantation and serum levels of gonadotropin-releasing hormone (GnRH), progesterone, and gonadotropins (FSH and LH), meanwhile the serum level of beta endorphin increased significantly. These effects were not observed in the low-dose treatment group (50 mg/kg body weight). The results of this study suggested that the anti-implantation effects of daidzein are probably caused by the interference of the hypothalamus-pituitary-gonadal axis which is involved in the implantation process.

Topics: Animals; Blastocyst; Dose-Response Relationship, Drug; Embryo Implantation; Female; Follicle Stimulating Hormone; Gonadotropin-Releasing Hormone; Isoflavones; Luteinizing Hormone; Phytoestrogens; Pregnancy; Pregnancy Rate; Progesterone; Random Allocation; Rats; Rats, Sprague-Dawley

Aromatase is an enzyme responsible for the conversion of androgen to estrogen. We genetically engineered an aromatase-deficient mouse (Ar(-/-) mouse) to express an enhanced green fluorescent protein (EGFP) gene in the uterus, ovary, adrenal and pituitary glands in a 17beta-estradiol (E2)-inducible manner. In this study, we analyzed estrogenic activities of diethylstilbestrol, genistein, daidzein and E2 in the Ar(-/-) tissues by using the EGFP expression as an indicator. These analyses manifest differential responses of the tissues to the compounds and also allow to determine the relative estrogenic potency of the compounds to that of E2 in vivo. Furthermore, analyses of the EGFP expression in ERalpha-deficient mice suggested that the expression is ERalpha-dependent in the uterus and pituitary gland. In conclusion, the Ar(-/-) mouse carrying the E2-inducible EGFP gene is a valuable tool for quantitative analyses of natural and synthetic estrogenic compounds in vivo.

Topics: Adrenal Glands; Animals; Antineoplastic Agents; Aromatase; Diethylstilbestrol; Estradiol; Estrogen Receptor alpha; Estrogens, Non-Steroidal; Female; Genistein; Green Fluorescent Proteins; Homozygote; Isoflavones; Mice; Mice, Knockout; Ovary; Phytoestrogens; Pituitary Gland; Uterus

We studied estrogen effects on osteoclastic differentiation using RAW264.7, a murine monocytic cell line. Differentiation, in response to RANKL and colony-stimulating factor 1, was evaluated while varying estrogen receptor (ER) stimulation by estradiol or nonsteroidal ER agonists was performed. The RAW264.7 cells were found to express ERalpha but not ERbeta. In contrast to RANKL, which decreased ERalpha expression and induced osteoclast differentiation, 10 nm estradiol, 3 microm genistein, or 3 microm daidzein all increased ERalpha expression, stimulated cell proliferation, and decreased multinucleation, with the effects of estrogen > or = daidzein > genistein. However, no estrogen agonist reduced RANKL stimulation of osteoclast differentiation markers or its down-regulation of ERalpha expression by more than approximately 50%. Genistein is also an Src kinase antagonist in vitro, but it did not decrease Src phosphorylation in RAW264.7 cells relative to other estrogen agonists. However, both phytoestrogens and estrogen inhibited RANKL-induced IkappaB degradation and NF-kappaB nuclear localization with the same relative potency as seen in proliferation and differentiation assays. This study demonstrates, for the first time, the direct effects of estrogen on osteoclast precursor differentiation and shows that, in addition to effecting osteoblasts, estrogen may protect bone by reducing osteoclast production. Genistein, which activates ERs selectively, inhibited osteoclastogenesis less effectively than the nonselective phytoestrogen daidzein, which effectively reproduced effects of estrogen.

Topics: Animals; Apoptosis; Carrier Proteins; Cell Cycle; Cell Differentiation; Cell Line; Cell Proliferation; Estrogen Receptor alpha; Estrogen Receptor beta; Estrogens; Extracellular Signal-Regulated MAP Kinases; Genistein; Isoflavones; Macrophage Colony-Stimulating Factor; Membrane Glycoproteins; Mice; NF-kappa B; Osteoclasts; Phytoestrogens; RANK Ligand; Receptor Activator of Nuclear Factor-kappa B; Signal Transduction; Transcription Factor RelA

Widespread epidemiological data support the notion that high isoflavone intake is safe and may provide health benefits similar to estrogen. Evidence from rodents shows that certain phytoestrogens can act as estrogen receptor (ER) ligands in the brain. This study sought to determine the estrogenic profile of food-borne phytoestrogens in neuronal cell lines using physiologically attainable concentrations. At sub-micromolar concentrations genistein, daidzein, and zearalenone stimulated ERalpha and ERbeta-dependent transcription in Neuro2A cells co-transfected with ERs and simple and complex estrogen-response-element (ERE) containing promoters, although compounds were more active in the presence of ERbeta. In SN56, neuronblastoma cells expressing endogenous ERs, only genistein mimicked estrogen regulation of progesterone receptor steady state mRNA levels. Unlike pharmaceutical SERMs, phytoestrogens did not stimulate an AP-1-dependent promoter. Micromolar concentrations of phytoestrogens did not antagonize physiological estrogen concentrations or antagonist activation of an AP-1-dependent promoter. These results demonstrate that food-borne phytoestrogens, particularly those found in soy, act as ERE-, but not AP-1-dependent transcriptional activators in neurons in the absence of estrogen, and dietary levels of these compounds do not act as antagonists to physiological estrogen concentrations.

Topics: Animals; Cell Line; Gene Expression Regulation; Genistein; Isoflavones; Neurons; Phytoestrogens; Rats; Receptors, Estrogen; Response Elements; RNA, Messenger; Selective Estrogen Receptor Modulators; Transcription Factor AP-1; Transcription, Genetic; Zearalenone

In this study, we examined the effects of exposure to phytoestrogen (daidzein), 17beta-estradiol (E2), diethylstilbestrol (DES) and staurosporin on the TM4 testicular cell line, using comprehensive analysis, such as cDNA microarray and two-dimension polyacrylamide gel electropholesis (2D-PAGE) analysis, and we demonstrated if these toxicogenomic analyses could classify the chemical compounds. First, RNA was extracted from TM4 cells that had been treated with daidzein (80 microM), DES, E2 (40 microM) and stauroporin (100 nM) for 30 min. We performed cDNA microarray analysis, and the expression ratio data thus obtained were then analyzed using hierarchical clustering. This hierarchical clustering showed that daidzein exposure induced a different effect on gene expression change from that of E2, DES and staurosporin. Next, protein extracted from TM4 cells also underwent cDNA microarray analysis for 3 h. We performed 2D-PAGE analysis, and the spot intensity ratio data thus obtained were analyzed using hierarchical clustering. As with cDNA microarray, the hierarchical clustering of protein spot ratios showed that daidzein exposure induced a different effect on gene expression change from that of the other substances. In conclusion, we have demonstrated for the first time that classification of these chemicals can be performed by clustering analysis, using data from cDNA microarray and 2D-PAGE analyses, and that exposure to daidzein induces effects different from those of E2, DES and staurosporin.

Topics: Cell Line; Diethylstilbestrol; Electrophoresis, Gel, Two-Dimensional; Estradiol; Estrogens, Non-Steroidal; Gene Expression Profiling; Humans; Isoflavones; Male; Oligonucleotide Array Sequence Analysis; Phytoestrogens; RNA, Messenger; Sertoli Cells; Testis

The aim of this study was to determine how the efficacy of tamoxifen is affected when combined with soy isoflavones. To address this, female Sprague-Dawley rats were placed on diets supplemented with tamoxifen, genistein, daidzein, or a combination of each isoflavone with tamoxifen; a week later mammary tumours were induced by 7,12 dimethylbenzanthracene. The most effective diet was the tamoxifen/daidzein combination. It reduced tumour multiplicity by 76%, tumour incidence by 35%, tumour burden by over 95%, and increased tumour latency by 62% compared with positive controls. The tamoxifen/daidzein combination diet was in all aspects more effective while the tamoxifen/genistein combination was less effective than the tamoxifen diet. The tamoxifen/daidzein diet significantly decreased 8-oxo-deoxyguanosine levels (an indicator of oxidative DNA damage) in the mammary glands. This study conclusively shows for the first time the combination of daidzein with tamoxifen produces increased protection against mammary carcinogenesis, while the combination of genistein with tamoxifen produces an opposing effect when compared with tamoxifen alone.

Topics: Animals; Antineoplastic Agents, Hormonal; Chemoprevention; Dietary Supplements; Drug Interactions; Female; Hydrolysis; Isoflavones; Mammary Neoplasms, Experimental; Phytoestrogens; Rats; Rats, Sprague-Dawley; Tamoxifen

Topics: Female; Fungi; Genistein; Glycine max; Hot Flashes; Humans; Isoflavones; Phytoestrogens; Pilot Projects; Postmenopause

In this study, we investigated the effects of long-term administration of quercetin with or without daidzein on glutathione and the enzymes involved in its metabolism in rat liver in vivo. Male Sprague-Dawley rats were divided randomly into four groups and given oral quercetin (20 mg/day) and daidzein (20 mg/day) alone or in combination, or vehicle alone for six weeks. The serum and liver alpha-tocopherol concentrations were significantly increased following administration of quercetin and daidzein alone or in combination. Glutathione concentration and glutathione reductase activity was significantly (p < 0.05) decreased with quercetin treatment, while no such effect was observed with daidzein treatment. Interestingly, decrease in glutathione concentration and glutathione reductase activity by quercetin treatment was inhibited by combined administration of daidzein and quercetin. The malondialdehyde concentration was significantly decreased following administration of quercetin and daidzein alone or in combination. These results suggest that quercetin, but not daidzein, acts as a pro-oxidant agent by decreasing glutathione concentration and glutathione reductase activity. Interestingly, this pro-oxidant effect of quercetin was inhibited by the combined administration of quercetin and daidzein.

Topics: Administration, Oral; alpha-Tocopherol; Animals; Drug Combinations; Glutathione; Glutathione Peroxidase; Glutathione Reductase; Isoflavones; Lipid Peroxidation; Liver; Male; Phytoestrogens; Quercetin; Random Allocation; Rats; Rats, Sprague-Dawley; Time Factors

In adult rats, elongation of cardiac myocytes (CMs) correlates with dilatation (and sometimes dysfunction) of cardiac ventricles. Although sex steroids may constitute one possible factor that affects the dimensions of CMs, studies on their effects in rodents is complicated by the fact that most commercial soy-based diets also contain abundant phytoestrogens. We report that feeding Wistar-Kyoto rat dams during gestation and lactation with a phytoestrogen-rich soy-based diet caused the CMs of their adult offspring to be shorter than in counterparts originating from mothers fed with a phytoestrogen-free casein-based diet. The soy-based diet had no such effects when given to rats after 6 wk of age, and its effects were replicated when supplementing the maternal casein-based diet with the isoflavones daidzein and genistein (the most abundant phytoestrogens in soy-based diets). In contrast to rats whose mothers had been fed with a soy-based diet, the hearts of adult rats raised with a casein-based diet only featured dilated eccentric hypertrophy and progressed toward congestive heart failure when further challenged. Thus the presence of isoflavones in the maternal diet provides cardioprotection to the hearts of their offspring during adulthood.

Topics: Animals; Cardiotonic Agents; Caseins; Cell Size; Diet; Dietary Supplements; Female; Genistein; Glycine max; Isoflavones; Lactation; Myocytes, Cardiac; Phytoestrogens; Pregnancy; Pregnancy, Animal; Prenatal Exposure Delayed Effects; Rats; Rats, Inbred WKY

Phenols are present in the environment and are prevalent in human populations, as environmental contaminants, dietary components, or their metabolites. Many are potential endocrine-altering agents. Currently available methods analyze single components or single families of chemicals as biomarkers of exposure. In order to assess multiple biologically relevant exposures to such substances, we evaluated the feasibility of determining several phenols simultaneously in urine, using an electrochemical detector (ECD) in combination with high performance liquid chromatography (LC). Based on reported analyses in the literature and the ECD response, we selected four xenobiotic residues, including three phytoestrogens (enterolactone, daidzein, and genistein) and bisphenolA [BPA]. These compounds had detection limits below 1 microg/L in urine using the cleanup procedure (glucuronidase hydrolysis and C18 column) and the urine volume (2 mL) we employed. As a pilot study to demonstrate the method's utility, we determined urinary enterolactone, daidzein, genistein and BPA in samples from nine children and 24 adults.

Topics: 4-Butyrolactone; Benzhydryl Compounds; Biomarkers; Child; Chromatography, High Pressure Liquid; Electrochemistry; Environmental Exposure; Female; Genistein; Humans; Isoflavones; Lignans; Middle Aged; Phenols; Phytoestrogens

There is growing evidence that soy isoflavones exert hormonal and antioxidant effects in postmenopausal women. In the present study, 12 postmenopausal Korean women with diabetic retinopathy consumed 2 g of genistein combined polysaccharides (GCP), containing 120 mg of genistein and 57 mg of daidzein, daily for 12 weeks. Blood was collected prior to and after 12 weeks of GCP supplementation for analysis of fasting blood glucose, insulin, lipid profiles, sex hormone-binding globulin (SHBG), estradiol, testosterone, free testosterone, and osteocalcin and activities of glutathione peroxidase (GSH-Px), catalase, and paraoxonase. After GCP supplementation, blood glucose, insulin, total cholesterol, triglyceride, high-density lipoprotein, and low-density lipoprotein cholesterols did not change significantly. However, there were significant increases in SHBG (P < .05) and testosterone (P < .05) and a decrease in estradiol (P < .01). Free testosterone levels were not significantly affected by GCP supplementation. After supplementation, osteocalcin decreased, but the difference was not statistically significant. Although activities of catalase and paraoxonase were unchanged, GSH-Px activity (P < .01) was increased significantly. These findings suggest that GCP supplementation may change the levels of some hormones and improve antioxidant status in postmenopausal Korean women with diabetic retinopathy.

Topics: Antioxidants; Blood Glucose; Diabetic Retinopathy; Dietary Supplements; Estrogens; Female; Genistein; Glutathione Peroxidase; Humans; Insulin; Isoflavones; Korea; Lipids; Middle Aged; Osteocalcin; Phytoestrogens; Postmenopause; Sex Hormone-Binding Globulin; Surveys and Questionnaires; Testosterone

Although many dietary studies have focused on breast cancer risk, few have examined dietary influence on tumor characteristics such as estrogen receptor (ER) status. Because phytoestrogens may modulate hormone levels and ER expression, we analyzed ER status and phytoestrogen intake in a case-case study of 124 premenopausal breast cancer patients. We assessed intake with a food-frequency questionnaire and obtained ER status from medical records. Rather than focusing on risk, we evaluated whether low intakes were more strongly associated with ER-negative tumors than with ER-positive disease. In logistic regression adjusting for potential confounders, threefold greater risks of ER-negative tumors relative to ER-positive tumors were associated with low intake of the isoflavones genistein (odds ratio, OR=3.50; 95% confidence interval, CI=1.43-8.58) and daidzein (OR=3.10; 95% CI=1.31-7.30). Low intake of the flavonoid kaempferol (OR=0.36; 95% CI=0.16-0.83), the trace element boron (OR=0.33; 95% CI=0.13-0.83), and the phytosterol beta-sitosterol (OR=0.42; 95% CI=0.18-0.98) were associated with decreased risk of ER-negative tumors relative to ER-positive disease. Other phytoestrogens were not significantly associated with ER status. Thus, in premenopausal patients, some phytoestrogens may affect breast carcinogenesis by influencing ER status. Such findings suggest new directions for mechanistic research on dietary factors in breast carcinogenesis that may have relevance for prevention and clinical treatment.

Topics: Adult; Anticarcinogenic Agents; Boron; Breast Neoplasms; Diet; Female; Genistein; Humans; Hypolipidemic Agents; Isoflavones; Kaempferols; Middle Aged; Nutritional Status; Odds Ratio; Phytoestrogens; Premenopause; Receptors, Estrogen; Risk Factors; Sitosterols; Surveys and Questionnaires

Phytoestrogens have been described to be weak estrogens, SERMs or exhibit antiestrogenic properties. However, information about their activity in presence of estrogens is limited. Therefore, we have analysed the dose dependent combinatory activity of the phytoestrogens genistein (Gen), daidzein (Dai) and coumestrol (Cou), and 17beta-estradiol (E2) on cell proliferation and apoptosis induction in human MCF-7 breast cancer cells. Neither additive nor antagonistic effects on proliferation could be observed, but in contrast all phytoestrogens possessed the ability to inhibit apoptosis in the presence of 17beta-estradiol. In summary, our in vitro results demonstrate that Gen does not exhibit any antiestrogenic properties. The additive growth stimulatory effects of Gen, Dai and Cou in the presence of E2 are not the result of a stimulation of proliferation; these phytoestrogens, at least in MCF-7 cells, could be characterised as inhibitors of apoptosis.

Topics: Apoptosis; Breast Neoplasms; Cell Proliferation; Coumestrol; Drug Interactions; Estradiol; Estrogen Antagonists; Female; Genistein; Humans; Isoflavones; Phytoestrogens

The phytoestrogen daidzein was metabolized by Nocardia species NRRL 5646 to give two metabolites obtained by hydroxylation and methylation. These metabolites were spectrally characterized as 7-methoxy-4'-hydroxyisoflavone (isoformononetin) and 7,8-dimethoxy-4'-hydroxyisoflavone. Mortierella isabellina ATCC 38063 was able to metabolize daidzein to the unusual metabolite daidzein-4'-rhamnopyranoside.

Topics: Hydroxylation; Isoflavones; Methylation; Models, Chemical; Molecular Structure; Mortierella; Nocardia; Phytoestrogens

Glutathione-S-transferases and glutathione play a key role in the detoxification of most toxic agents. In the present study, the protective effects, if any, of isoflavone phytoestrogens--genistein and daidzein on the carbon tetrachloride (CCl4) induced changes in the activity of alanine aminotransferase (ALT), aspartate aminotransferase (AST), glutathione S transferase (GSH) and levels of glutathione (GSH) and thiobarbituric acid reactive substances (TBARS)-were studied. The activities of ALT and AST were assayed in the serum, whereas the activity of GST and levels of GSH and TBARS were determined in the livers of rats. The current study involved the division of animals into two main groups: (i) rats pretreated with genistein and daidzein for three days; and (ii) non-pretreated rats. In the pretreated group, rats received oral doses of genistein (7.9 micromol/kg body weight) and daidzein (7.9 micromol/kg body weight) for three consecutive days (once daily) followed by oral dose of CCl4 on the 4th and the 5th day concurrently with the phytoestrogens-genistein or daidzein. In the non-pretreated group animals received oral dose of CCl4 (1 ml/kg body weight) for two consecutive days along with the phytoestrogens-genistein or daidzein. Treatment of male rats with CCl4 significantly elevated the activity of ALT and AST in serum and levels of TBARS in the liver. On the other hand, CCl4 resulted in decreased activity of GST and lowered the GSH levels. Coadministration of genistein and daidzein with CCl4 could not restore the alterations in the activity of ALT and AST caused by CCl4 to normal control levels. However, repeated dose treatments with genistein and daidzein for three days prior to the administration of CCl4 restored such alterations to normal levels. Our results indicate that genistein is more effective than daidzein in counteracting the inhibition of GST activity caused by CCl4 and restoring it to normal levels. Genistein was also more effective than daidzein restoring the induced TBARS levels caused by CCl4 to normal control levels when rats were pretreated with the isoflavone orally for three days. It has been observed that the tested isoflavonoids were able to antagonize the toxic effects of CCl4. Such counteracting effects were more pronounced for genistein and when the phytoestrogens were administered as repeated doses prior CCl4 administration.

Topics: Animals; Carbon Tetrachloride; Genistein; Glutathione Transferase; Isoflavones; Liver; Male; Oxidative Stress; Phytoestrogens; Rats; Rats, Wistar; Thiobarbituric Acid Reactive Substances

The isoflavones--genistein and daidzein -- compounds found in high concentrations in soy play an important role in prevention of many diseases and affect some metabolic pathways. In the performed experiment it was demonstrated that genistein (5mg/kg b.w.) administered intragastrically for three days to male Wistar rats substantially diminished blood leptin level. Studies with isolated rat adipocytes revealed that this phytoestrogen strongly restricted leptin secretion from these cells. These effects were not accompanied by any changes in leptin gene expression in adipocytes. Daidzein-- an analogue of genistein -- used at similar concentrations did not affect blood leptin concentration, leptin secretion and expression of its gene. To determine the influence of genistein and daidzein on leptin release, adipocytes isolated from the epididymal fat tissue were incubated for 2h in Krebs--Ringer buffer. Leptin secretion stimulated by glucose with insulin was significantly diminished by genistein (0.25--1mM). This effect of genistein may arise from several aspects of its action in adipocytes documented in the literature such as the inhibition of glucose transport and metabolism, the attenuation of insulin signalling, the inhibition of cAMP phosphodiesterase and the stimulation of lipolysis. However, the bypassing of the restrictive action of genistein on glucose transport and glycolysis (by the use of alanine instead of glucose) and on insulin action (by the use of nicotinic acid) was not sufficient to restore leptin secretion from isolated adipocytes. It was also demonstrated that the restriction of the stimulatory influence of genistein on cAMP/protein kinase A (PKA) pathway (by the inhibition of PKA activity) did not improve leptin release. Results obtained in our experiments point at the restriction of glucose metabolism following formation of pyruvate as the pivotal reason of the inhibitory action of genistein on leptin release.

Topics: Adipocytes; Animals; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Epididymis; Gene Expression; Genistein; Glucose; Insulin; Isoflavones; Leptin; Male; Phytoestrogens; Rats; Rats, Wistar

This study investigated a phytoestrogen-rich herb formula, Xianlinggubao (XLGB) (including genistein 510 microg/g and daidzein 2500 microg/g), concerning prevention of OVX-induced deterioration of musculoskeletal tissues in 11-month-old female Wistar rats, which were randomized into Sham, OVX, and XLGB groups. Daily oral administration of XLGB (250 mg/kg/day) started after OVX for 3 months. mRNA of MHC-I IIa IIb of abductor muscle was determined by RT-PCR. The proximal femoral BMD and geometry, microarchitecture, and mechanical strength were evaluated by pQCT, micro-CT, and compressive testing, respectively. The bone turnover biochemical markers serum osteocalcin (OC) and urinary deoxypyridinoline (DPD) were evaluated. The results showed that (1) XLGB-treated OVX rats showed no difference compared to the Sham group, whereas OVX induced significant deterioration in variables related to bone density, microarchitecture, and mechanical strength (P < 0.05); (2) biochemical markers showed no difference between sham and XLGB groups as compared with higher bone turnover in OVX rats (P < 0.05); (3) mRNA expression of MHC-I IIa IIb was downregulated in OVX rats but upregulated after XLGB treatment (P < 0.05); and (4) as compared with the OVX group, no uterine hypertrophy was found in XLGB-treated rats. In conclusion, findings of this study suggested that the herbal preparation XLGB was able to prevent OVX-induced deterioration of musculoskeletal tissues at the hip without causing uterine stimulation.

Topics: Animals; Biomarkers; Bone Density; Female; Femur; Genistein; Growth Inhibitors; Isoflavones; Muscle, Skeletal; Osteoporosis; Ovariectomy; Phytoestrogens; Plant Preparations; Rats; Rats, Wistar

The soy isoflavone daidzein (DAI) is known to undergo metabolism to equol (EQO) and to 3'-hydroxy-DAI (3'-HO-DAI) and 6-hydroxy-DAI (6-HO-DAI) in humans. In order to better understand the implications of soy diets for human health, the hormonal and genotoxic activities of these DAI metabolites were studied in cultured human endometrial carcinoma cells. When the estrogenicity was tested by cell-free binding to recombinant human estrogen receptor (ER) alpha and beta as well as by the induction of enzyme activity and gene expression of alkaline phosphatase (ALP) in Ishikawa cells, the ranking order was EQO>DAI>3'-HO-DAI>6-HO-DAI. All compounds had a higher affinity to ERbeta than to ERalpha. No significant anti-estrogenic effects of the DAI metabolites were observed in the cells at non-cytotoxic concentrations. The in vitro genotoxicity was assessed by analyzing effects on cell cycle distribution and cell morphology as well as the induction of micronuclei (MN). EQO caused a slight increase in G1 and decrease in S phase of the cell cycle, and slightly but significantly induced kinetochore-positive as well as kinetochore-negative MN and an elevated proportion of abnormal mitotic spindles. 3'-HO-DAI, but not 6-HO-DAI, induced kinetochore-negative MN. The observation that major human metabolites of DAI exhibit estrogenic and genotoxic potential may be of relevance for the safety evaluation of diets containing soy isoflavones.

Topics: Adenocarcinoma; Alkaline Phosphatase; Cell Cycle; Cell Line, Tumor; Drug Screening Assays, Antitumor; Endometrial Neoplasms; Enzyme Induction; Equol; Estrogen Receptor alpha; Estrogen Receptor beta; Female; Gene Expression; Humans; Isoflavones; Micronuclei, Chromosome-Defective; Micronucleus Tests; Mutagens; Phytoestrogens; RNA, Messenger; Spindle Apparatus

An amphiphilic, hyperbranched polymer suitable for use in controlled drug delivery is reported. This polymer was obtained by modification of the hyperbranched aliphatic polyester Boltorn H20 (H20) with succinic anhydride and then glycidyl methacrylate, and formed nanoparticles in aqueous solution. The critical association concentration was 7.4 x 10(-3) g . L(-1), as determined by fluorescence spectroscopy using pyrene as a molecular probe. A static/dynamic laser light scattering (LLS) study revealed that the average particle size was 39.4 nm with a low particle size distribution (PDI=0.04), and that each particle was composed of about 350 amphiphilic molecules. Daidzein, a hydrophobic traditional Chinese medicine, was encapsulated during particle formation and the release properties were determined. The optimal feeding concentration of daidzein to hyperbranched polyester was 4.9 x 10(-5) g . mL(-1) to 5.0 x 10(-3) g . mL(-1) with a loading efficiency of 76.1%. In the presence of the enzyme Lipase PS, the drug loaded nanoparticles degraded in a random one-by-one manner and released the drug over a few days. This system is therefore a novel controlled drug release system based on nanoparticles formed of hyperbranched polyester. Encapsulation of daidzein by hyperbranched polyester particles.

Topics: Capsules; Delayed-Action Preparations; Hydrophobic and Hydrophilic Interactions; Isoflavones; Nanostructures; Phytoestrogens; Polyesters; Solutions; Water

The molecular mechanisms of the vascular effects of phytoestrogens are poorly studied. Prostacyclin is a potent vasodilator synthesized by two isoforms of cyclooxygenase (COX) in endothelium. This study examine the effects of two phytoestrogens, the isoflavones genistein and daidzein, on prostacyclin production by cultured human umbilical vein endothelial cells (HUVECs) and the possible role of not only estrogen receptors but also both COX isoforms. The two phytoestrogens significantly increased prostacyclin release in a time- and dose-dependent (0.01-1 microM) manner, being higher than control after 24 h. Selective inhibitors of COX-1, SC-560 [5-(4-chlorophenyl)-1-(4-methoxypjenyl)-3-(trifluoromethyl)-1H-pyrazole], and COX-2, NS-398 (N-[2-(cyclohexyloxy)-4 nitrophenyl]-methanesulfonamide), were used to investigate the relative contribution of each enzyme. Both inhibitors decreased basal production of prostacyclin, but only COX-2 inhibition completely abolished the isoflavone-stimulated prostacyclin production. Phytoestrogens also increased COX-2 mRNA expression and protein content without affecting COX-1 levels. All these effects were mediated through estrogen receptor activation since treatment of cells with the estrogen receptor antagonist ICI 182780 [7alpha-[9[(4,4,5,5,5-pentafluoropentyl)sulfinyl]nonyl]-estra-1,3,5(10)-triene-3,17beta diol] completely abolished the isoflavone-induced increase in prostacyclin production, COX-2 mRNA expression, and COX-2 protein content. The results clearly support the hypothesis that genistein and daidzein increased HUVEC prostacyclin production through estrogen receptor-dependent mechanism, which involved the enhancement of COX-2 protein and activity.

Topics: Cell Survival; Cyclooxygenase 1; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; Endothelial Cells; Epoprostenol; Estrogens, Non-Steroidal; Female; Genistein; Humans; Immunoblotting; Infant, Newborn; Isoflavones; Male; Phytoestrogens; Protein-Tyrosine Kinases; Receptors, Estrogen; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger

Soy food has been associated with a reduced incidence of hormonal cancer in Asian countries, and the soy isoflavones daidzein and genistein are believed to protect against tumors induced by the endogenous hormone 17beta-estradiol (E2). In the present study, we have examined if daidzein and genistein as well as several structurally related isoflavones are able to modulate the in vitro glucuronidation of E2 in human hepatic microsomes. It is known that different isoforms of UDP-glucuronosyltransferase (UGT) are involved in E2 glucuronidation: UGT1A1 leads exclusively to the 3-glucuronide and is stimulated by E2 via homotropic kinetics, whereas UGT2B7 gives rise to the 17-glucuronide of E2 following Michaelis-Menten kinetics. In our study, daidzein markedly stimulated the 3-glucuronidation, thereby enhancing the metabolic clearance of E2. In contrast, genistein inhibited the 3-glucuronidation. The 17-glucuronidation of E2 was not affected by either compound. Formononetin and the daidzein metabolites equol, 3'-hydroxy-daidzein, 6-hydroxy-daidzein and glycitein behaved similar to daidzein, whereas biochanin A resembled genistein. The effect of daidzein on the 3-glucuronidation of E2 in human hepatic microsomes was also obtained with human recombinant UGT1A1. Since the only other compound known to stimulate E2 glucuronidation via allosteric kinetics is 17alpha-ethynylestradiol, our study is the first report of the heterotropic stimulation of a UGT by a non-steroidal and naturally occurring compound. An enhanced rate of glucuronidation of E2 by daidzein and its metabolites may contribute to the putative protection of soy against hormonal cancer.

Topics: Allosteric Regulation; Antineoplastic Agents; Chromatography, High Pressure Liquid; Estradiol; Genistein; Glucuronides; Glucuronosyltransferase; Humans; Isoflavones; Kinetics; Microsomes, Liver; Phytoestrogens; Recombinant Proteins; Substrate Specificity

The extent of conversion of daidzein to its metabolite, equol, by intestinal microflora may be a critical step that determines if a diet rich in daidzein protects against the deterioration of bone after estrogen withdrawal. The objective was to determine the extent that daidzein is converted to equol. In addition, bone mineral content (BMC), bone mineral density (BMD) and strength of femurs and lumbar vertebrae (LV) in four mouse strains were measured. Mice were ovariectomized and fed control diet (AIN93G) with or without daidzein (200 mg daidzein/kg diet) for 3 weeks, after which serum, femurs and LV were collected. Serum daidzein and equol were elevated in all mice fed daidzein. Among mice fed daidzein, the CD-1 and Swiss-Webster (SW) mice had higher (P<.001) serum equol than C57BL/6 (C57) and C3H mice. Differences due to mouse strain were observed for all bone outcomes. C57 mice had lower femur BMC (P<.001), BMD (P<.001) and peak load at femur midpoint (P<.001) and neck (P<.001) than other mouse strains. C57 mice also had a lower femur midpoint yield load (P<.001) and resilience (P<.001) than C3H mice. C57 mice had a lower LV1-4 BMC (P<.001) and BMD (P<.001) compared with all mouse strains and peak load of LV3 was lower than CD-1 and SW mice. Differences in serum equol, BMD and bone strength properties should be considered when selecting a mouse strain for investigating whether dietary strategies that include isoflavones preserve bone tissue after ovariectomy.

Topics: Animals; Biomechanical Phenomena; Body Weight; Bone and Bones; Bone Density; Eating; Equol; Female; Femur; Isoflavones; Lumbar Vertebrae; Mice; Mice, Inbred C3H; Mice, Inbred C57BL; Ovariectomy; Phytoestrogens; Species Specificity

Phyto-oestrogens are polyphenolic non-steroidal plant compounds with oestrogen-like biological activity. Phyto-oestrogens have many biological effects including oestrogen agonist/antagonist properties. However, the effect of phyto-oestrogens on allergic responses remains unclear. In this study we investigated whether formononetin, a phyto-oestrogen, and its metabolites, daidzein and equol, affect production of interleukin-4 (IL-4), a pro-inflammatory cytokine closely associated with allergic immune response, in primary CD4+ T cells and EL4 T lymphoma cells. Formononetin, daidzein and equol significantly enhanced IL-4 production from both CD4+ T cells and EL4 cells in a dose-dependent manner. Formononetin, daidzein and equol also enhanced IL-4 gene promoter activity in EL4 cells transiently transfected with IL-4 gene promoter constructs, but this effect was impaired in EL4 cells transfected with an IL-4 promoter construct deleted of P4 site carrying nuclear factor of activated T cells (NF-AT) and activator protein-1 (AP-1) binding sites. In addition, formononetin, daidzein and equol increased AP-1 DNA binding activities while did not affect NF-AT DNA binding activities. The enhancing effects on IL-4 production and AP-1 DNA binding activities were abrogated by specific inhibitors for phosphatidylinositol-3-kinase (PI3K), protein kinase C (PKC) and p38 mitogen-activated protein kinase (MAPK), indicating that formononetin, daidzein and equol might enhance IL-4 production by increased activation of AP-1 through the PI3-K/PKC/p38 MAPK signalling pathway. These results suggest that phyto-oestrogens and some of their metabolites may increase allergic responses via the enhancement of IL-4 production in T cells.

Topics: Animals; CD4-Positive T-Lymphocytes; Cells, Cultured; DNA-Binding Proteins; Equol; Humans; Interleukin-4; Isoflavones; Lymphocyte Activation; Lymphoma, T-Cell; Mice; p38 Mitogen-Activated Protein Kinases; Phosphatidylinositol 3-Kinases; Phytoestrogens; Protein Kinase C; T-Lymphocyte Subsets; Tetradecanoylphorbol Acetate; Transcription Factor AP-1; Tumor Cells, Cultured

Phytoestrogens may be associated with a reduced risk of hormone dependent neoplasms such as prostate and breast cancers. We tried to determine the validity of the association between serum phytoestrogen concentrations and dietary habits obtained from a food frequency questionnaire used in the Japan Collaborative Cohort Study (JACC Study) for Evaluation of Cancer Risk sponsored by the Ministry of Education, Science, Sports and Culture of Japan (Monbusho).. The subjects were 151 male controls who were selected for a nested case-control study for evaluating prostate cancer risk as part of the JACC Study. Dietary habits were determined using a food frequency questionnaire at baseline, and the concentrations of genistein, daidzein, and equol in frozenstored serum samples assayed in 2002 were compared.. Tofu intake showed a significant association with the serum concentrations of genistein and daidzein (Spearman's correlation coefficients (rs)=0.30 and 0.27, respectively), and miso soup showed a slight association with serum concentrations of these phytoestrogens. In contrast, serum concentrations of equol were not associated with dietary intake of tofu and miso soup. After adjustment for serum daidzein concentration, serum equol concentration was associated with the intake of foods containing fat, meat, and coffee, but not green tea.. Serum genistein and daidzein concentrations were significantly associated with dietary intake of tofu, and slightly with intake of miso soup. Consumption of fat, meat, and coffee may be associated with equol production by intestinal microflora in this sample set.

Topics: Adult; Aged; Case-Control Studies; Cohort Studies; Diet Surveys; Feeding Behavior; Genistein; Humans; Isoflavones; Japan; Male; Middle Aged; Nutritional Status; Phytoestrogens; Prostatic Neoplasms; Soy Foods; Surveys and Questionnaires

As the epidemiological and physiological investigation of isoflavones and lignans expands, the need for sensitive methods for analyzing large numbers of samples intensifies. We have developed a method using high-performance liquid chromatography (HPLC) equipped with a coulometric electrode array detector for separation and sensitive detection of daidzein (Da), equol (Eq), genistein (Ge) and enterolactone (Enl) in dried blood spots (DBS). Detection limits ranged from 4.5 pg or 0.09 ng/mL (Eq) to 19 pg or 0.38 ng/mL (Ge) on column. Signal linearities ranged from detection limits to 200 ng/mL (Eq, Enl) and 600 ng/mL (Da, Ge) sample concentration. Correlations between DBS and serum concentrations were 0.66 (Enl), 0.88 (Eq), 0.98 (Ge) and 0.99 (Da). Intra-assay coefficients of variation (CVs) were less than 8% and inter-assay CVs ranged from 2.4 to 20.2% for Da, Eq and Ge for three levels of controls. Enl intra-assay CV was 13.6% for the low pooled control. Analytic recovery ranged from 87% (inter-assay Ge) to 98% (inter-assay Enl). DBS concentrations of Da, Ge and Eq were stable for at least 8 weeks at 4 and 25 degrees C, and at 37 degrees C for at least 5 weeks, with Enl showing greater variability at all temperatures but relative stability for 7 weeks. Measurement of samples from 135 perimenopausal Japanese women consuming habitual diets in Kyoto and Fukushima prefectures showed the former to have the expected lower concentrations of Da and Eq (416 and 87 nM) as well as Enl (49 nM) compared to the latter locale (566, 145 and 72 nM, respectively). This method could be useful in large epidemiological research or detailed physiological studies.

Topics: Chromatography, High Pressure Liquid; Drug Stability; Electrochemistry; Electrodes; Equol; Female; Genistein; Humans; Isoflavones; Japan; Perimenopause; Phytoestrogens; Temperature

Soy isoflavones display estrogenic activity in humans and animals, and thus are referred to as phytoestrogens. This study was performed to observe the effects of the soy isoflavones genistein, daidzein, and glycitein on cell cultures of rat skeletal muscles. [3H]Thymidine incorporation was used to determine cell proliferation, while protein synthesis and degradation were determined by tracking radiolabeled leucine. For the proliferation studies, insulin, estradiol, genistein, daidzein, or glycitein was supplemented at 0, 0.04, 0.08, 0.16, 0.31, 0.63, 1.25, 2.5, 5, 10, or 20 microM, respectively, or in combinations with final concentrations of 0, 0.1, 1, or 10 microM. Genistein reacted most similarly to estradiol, inhibiting proliferation at > or = 1 microM (P < .001). A combination of phytoestrogens resulted in significant inhibition of cell proliferation, but not to the extent observed with genistein alone. For the protein synthesis and degradation experiments, treatments of 0.1 microM dexamethasone or 1 microM concentrations of insulin, genistein, daidzein, or glycitein were used. Phytoestrogens did not inhibit or stimulate protein degradation or synthesis (P > .05). A one-tailed univariate analysis of variance revealed a trend (P < or = .1) in protein stimulation with genistein and glycitein treatments. These results suggest that the tyrosine kinase inhibiting activity of genistein may be affecting phosphorylation of the mitosis-promoting factor, preventing the advancement of the mitotic cell cycle. In addition, at higher total combined concentrations, daidzein and glycitein may be able to outcompete genistein for receptor sites. These results suggest that soy isoflavones in the diet may potentially modulate normal growth and development in humans and animals that ingest soy-based products.

Topics: Animals; Cell Division; Cell Line; Dexamethasone; Estradiol; Genistein; Glycine max; Insulin; Isoflavones; Muscle Proteins; Muscle, Skeletal; Phosphorylation; Phytoestrogens; Rats

The aim of this study was to characterize carbonic anhydrase II (CA2), as novel estrogen responsive gene, towards its usefulness to elucidate the molecular mechanisms of phytoestrogen action. Effects of estradiol-17beta (E2), and the phytoestrogens genistein (Gen), daidzein (Dai), as well as 8-prenylnaringenin (8PN) on CA2 mRNA expression were investigated in vivo in the uterus and liver of Wistar rats, and in vitro in Fe33 hepatoma cells. Relative amounts of mRNA levels of CA2 were measured by real-time RT-PCR. In vivo CA2 expression in uterus and liver is down-regulated by estrogen in time dependent manner with the most pronounced effect detectable 72 h after treatment. Treatment with Gen results in a slight down-regulation of CA2 expression in the uterus. In liver a response to Gen is detectable only after 7 h, where the expression of the gene is down-regulated to 60%. Treatment with Dai and 8PN for 72 h results in a slight down-regulation of CA2 in both tissues. In contrast in Fe 33 cells CA2 gene expression was up-regulated in response to the treatment with E2 for 7 h. In summary, we could demonstrate that the modulation of CA2 gene expression following treatment with E2 and Gen in rat uterus is comparable to the uterotrophic response of these substances, but with an inverted pattern. Remarkably, of all phytoestrogens 8PN exhibited the strongest uterotrophic response but only induced a very faint decrease of CA2 expression. In addition, we provide the first pieces of evidence that 8PN, like Gen and Dai, cannot be considered as a pure agonist. In conclusion, CA2 shows estrogen sensitivity not only in both tissues studied, but also in many others. Further, it exhibits a differential sensitivity thereby being capable to discriminate between different molecular qualities of phytoestrogens, like demonstrated for Gen and 8PN.

Topics: Animals; Carbonic Anhydrase II; Down-Regulation; Estradiol; Female; Flavanones; Gene Expression; Genistein; Isoflavones; Liver; Phytoestrogens; Rats; Tumor Cells, Cultured; Up-Regulation; Uterus

Phytoestrogens have been hypothesized to protect against prostate cancer via modulation of circulating androgen concentrations. We conducted a cross-sectional study of 267 men in the Norfolk arm of the European Prospective Investigation into Cancer and Nutrition (EPIC) cohort with 2 aims: first, to investigate the association between phytoestrogen exposure (measured from diet, urine, and serum) and plasma concentrations of sex hormone-binding globulin (SHBG), androstanediol glucuronide, testosterone and Free Androgen Index (FAI); and second, whether the association may be modified by polymorphisms in CYP19 and SHBG genes. Dietary daidzein and genistein intakes were obtained from food diaries and computed using an in-house food composition database. Urinary and serum concentrations of 3 isoflavones (daidzein, genistein, glycitein), 2 daidzein metabolites O-desmethylangolensin (O-DMA) and 2 lignan metabolites (enterodiol and enterolactone) were measured using mass spectrometry. There was no association between dietary, urinary, and serum phytoestrogens and plasma SHBG concentrations. Enterolactone was positively associated with plasma androstanediol glucuronide concentrations (urinary enterolactone: r = 0.127, P = 0.043; serum enterolactone: r = 0.172, P = 0.006) and FAI (urinary enterolactone: r = 0.115, P = 0.067; serum enterolactone: r = 0.158, P = 0.011). Both urinary and serum equol were associated with plasma testosterone (urinary equol: r = 0.332, P = 0.013; serum equol: r = 0.318, P = 0.018) and FAI (urinary equol: r = 0.297, P = 0.027; serum equol: r = 0.380, P = 0.004) among men with the TT genotype but not the CC or CT genotypes (r = -0.029 to -0.134, P = 0.091-0.717) for the CYP19 3'untranslated region (UTR) T-C polymorphism. Urinary and serum enterolactone showed similar genotype-dependent associations with testosterone but not with FAI. In this first study on phytoestrogen-gene associations in men, we conclude that enterolactone and equol are positively associated with plasma androgen concentrations, and interactions with CYP19 gene may be involved.

Topics: 4-Butyrolactone; Aged; Androgens; Androstane-3,17-diol; Aromatase; Diet; Equol; Genistein; Genotype; Humans; Isoflavones; Lignans; Male; Middle Aged; Phytoestrogens; Polymorphism, Genetic; Sex Hormone-Binding Globulin; Testosterone

Phytoestrogens are a diverse group of non-steroidal compounds that occur naturally in many plants. Because they possess a ring system similar to estrogens they are able to bind to estrogen receptors in humans. In the present study we tested the effects of the phytoestrogens genistein and daidzein on the production of human chorionic gondaotropin (hCG) in isolated trophoblast cells of term placentas in vitro.. Genistein and daidzein were incubated at different concentrations with trophoblast cells. Untreated cells were used as controls. At designated times aliquots were removed and tested for hCG production.. Production of the protein hormone hCG was influenced by the phytoestrogens genistein and daidzein in trophoblast cells. We found a significant decrease of hCG production in genistein- and daidzein-treated trophoblast cells that was concentration-dependent. Compared with daidzein, genistein seems to be a more efficient inhibitor of the production of hCG.. The phytoestrogens genistein and daidzein can reduce hCG production in human term trophoblasts. Both phytoestrogens belong to the group of isoflavones, which are enriched in soy-containing foods and are widely consumed by humans for putative beneficial health effects. Because both phytoestrogens have inhibitory effects on hCG production during pregnancy, exposure to these estrogen-like compounds during sensitive periods of development may have the capacity to alter the function of the reproductive system and thereby influence fertility.

Topics: Adult; Cells, Cultured; Chorionic Gonadotropin; Dose-Response Relationship, Drug; Female; Genistein; Humans; In Vitro Techniques; Isoflavones; Phytoestrogens; Phytotherapy; Pregnancy; Soy Foods; Trophoblasts

Particular intestinal bacteria are capable of metabolizing the soya isoflavone daidzein to equol and/or O-desmethylangolensin (O-DMA), and the presence of these metabolites in urine after soya consumption are markers of particular intestinal bacteria profiles. Prevalences of equol producers and O-DMA producers are approximately 30-50 % and 80-90 %, respectively, and limited observations have suggested that these daidzein-metabolizing phenotypes are stable within individuals over time. Characterizing stability of these phenotypes is important to understand their potential as markers of long-term exposure to particular intestinal bacteria and their associations with disease risk. We evaluated concordance within an individual for the equol-producer and O-DMA-producer phenotypes measured at two time points (T1, T2), 1-3 years apart. Phenotypes were ascertained by analysing equol and O-DMA using GC-MS in a spot urine sample collected after 3 d soya (source of daidzein) supplementation. In ninety-two individuals without recent (within 3 months before phenotyping) or current antibiotics use, 41 % were equol producers at T1 and 45 % were equol producers at T2, and 90 % were O-DMA producers at T1 and 95 % were O-DMA producers at T2. The percentage agreement for the equol-producer phenotype was 82 and for the O-DMA-producer phenotype was 89. These results indicate that these phenotypes are stable in most individuals over time, suggesting that they provide a useful biomarker for evaluating disease risk associated with harbouring particular intestinal bacteria responsible for, or associated with, the metabolism of the soya isoflavone daidzein.

Topics: Anti-Bacterial Agents; Bacterial Infections; Biomarkers; Dietary Supplements; Equol; Female; Humans; Intestines; Isoflavones; Male; Phenotype; Phytoestrogens; Time Factors

Cytosolic sulfotransferases (STs) are generally thought to be involved in detoxification of xenobiotics, as well as homeostasis of endogenous compounds such as thyroid/steroid hormones and catecholamine hormones/neurotransmitters. We report here the identification and characterization of a zebrafish estrogen-sulfating cytosolic ST. The zebrafish ST was bacterially expressed, purified, and examined for enzymatic activities using a variety of endogenous compounds as substrates. Results showed that the enzyme displayed much higher activities toward two endogenous estrogens, estrone (E(1)) and 17beta-estradiol (E(2)), in comparison with thyroid hormones, 3,3',5-triiodothyronine (T(3)) and thyroxine (T(4)), dopamine, dihydroxyphenylalanine (Dopa), and dehydroepiandrosterone (DHEA). The kinetic parameters, K(m), and V(max), with estrogens and thyroid hormones as substrates were determined. The calculated V(max)/K(m) for E(1), E(2), T(3), and T(4) were, respectively, 31.6, 16.7, 1.5, and 0.8 nmol min(-1) mg(-1) microM(-1), indicating clearly the estrogens being preferred physiological substrates for the enzyme. The inhibitory effects of isoflavone phytoestrogens on the sulfation of E(2) by this zebrafish ST were examined. The IC(50) determined for quercetin, genistein, and daidzein were 0.7, 2.5, and 8 microM, respectively. Kinetic analyses revealed that the mechanism underlying the inhibition by these isoflavones to be of the competitive type.

Topics: Animals; Cytosol; Dose-Response Relationship, Drug; Estrogens; Genistein; Glutathione Transferase; Humans; Isoflavones; Phytoestrogens; Plant Preparations; Quercetin; Recombinant Fusion Proteins; Substrate Specificity; Sulfotransferases; Thyroid Hormones; Zebrafish; Zebrafish Proteins

Ca absorption has been shown to be unaffected by high luminal concentrations of two commonly consumed soyabean phyto-oestrogens (PO) (genistein and daidzein) in Caco-2 cells grown under oestrogen-depleted conditions. However, these compounds exhibit dose-dependent biphasic effects in some tissues, such as reproductive tissue and bone. Thus, in light of this biphasic activity, the effect of lower concentrations of genistein and daidzein on Ca absorption requires further investigation. Therefore, the aim of the present study was to investigate the effect of a range of concentrations of genistein and daidzein on Ca absorption in the human Caco-2 intestinal-like cell model. Caco-2 cells were seeded onto permeable filter supports and allowed to differentiate into monolayers. On day 21, the Caco-2 monolayers (n 12 per treatment), grown in oestrogen-deplete media, were then exposed to 10 nm-1,25-dihydroxycholecalciferol (1,25 (OH)2D3), or 1, 10 and 50 microm-genistein or -daidzein for 24 h. After exposure, transepithelial and transcellular transport of (45)Ca and fluorescein transport were measured. As expected, 1,25 (OH)2D3 stimulated Ca absorption in Caco-2 cells, by up regulating transcellular transport. Ca absorption was unaffected by either PO at luminal concentrations of 1, 10 or 50 microm, typical of intakes by Western and Asian populations as well as supplemental levels, respectively. The results of this model suggest that the proposed beneficial effects of supplemental levels of these PO compounds on bone mass in postmenopausal women more probably arise from direct effects on bone cells, and not by an indirect effect of these compounds on Ca absorption.

Topics: Absorption; Biological Transport; Caco-2 Cells; Calcitriol; Calcium; Calcium Channel Agonists; Cell Survival; Culture Media; Diet; Dose-Response Relationship, Drug; Enzyme Inhibitors; Estrogens, Non-Steroidal; Genistein; Humans; Isoflavones; Phytoestrogens; Plant Preparations

To investigate the functional changes in rabbit penile corpus cavernosum (CC) secondary to experimental hyperestrogenism and attempt to identify sites of immunoexpression for estrogen receptor subtypes alpha and beta (ER-alpha and ER-beta) in the CC. Although the role of testosterone in sexual function has been extensively studied in clinical settings and experimental animal models, the effect of hormonal modulation/imbalance arising from estrogenic excess has not been characterized.. Eighteen New Zealand white male rabbits (2.5-3.0 kg) were divided into control and two treatment groups. The two treatment groups were given orally 0.1 mg of estradiol valerate (estradiol group) or phytoestrogen, daidzein (phytoestrogen group) daily for 12 weeks. Blood and tissue samples were collected for hormone levels and in vitro pharmacologic studies. CC samples from untreated rabbits (n = 4) were cryosectioned and incubated with appropriate mouse monoclonal antibody for identification of ER-alpha and ER-beta.. Through immunohistochemistry, color signals for nuclear ER-alpha and ER-beta receptors were localized within the CC. Chronic treatment with estradiol and phytoestrogen significantly reduced the systemic total testosterone levels. In organ bath experiments, relaxant responses to acetylcholine, nitroglycerin, and nitrergic transmission were significantly attenuated compared with the control response. With regard to the contractile effect, both types of estrogen treatments significantly potentiated norepinephrine-induced antierectile contraction of the CC.. These results indicate that estradiol treatment and chronic exposure of phytoestrogen may cause receptor-mediated pathophysiologic changes in erectile function, leading to erectile dysfunction.

Topics: Acetylcholine; Animals; Body Weight; Estradiol; Estrogen Receptor alpha; Estrogen Receptor beta; Isoflavones; Male; Muscle, Smooth; Nitric Oxide Donors; Nitroglycerin; Penile Erection; Phytoestrogens; Plant Preparations; Rabbits; Receptors, Estrogen; Testosterone; Vasodilator Agents

Estimating intake of phyto-oestrogens (PO) is difficult because there is inadequate information on the PO content of foods. Development of a biomarker of intake is therefore necessary for carrying out epidemiological studies. We aimed to validate a newly constructed PO database, containing more than 600 values assigned to foods by using duplicate diet analysis, and to investigate the relationships between measured PO intake, urinary excretion and plasma concentrations of PO. Fourteen subjects with estimated dietary intakes of PO ranging from 0 to 44 mg/d, measured by 7 d weighed intake, completed a duplicate diet collection over 24 h. Concurrently, a 24 h urine collection, validated using p-aminobenzoic acid, was obtained and one timed spot plasma sample taken. Duplicate diets, complete urine collections and plasma samples were analysed for total genistein and daidzein using liquid chromatography-MS to determine PO intake. The potential for 24 h urinary excretion and plasma PO concentrations to reflect dietary intake was investigated. Mean estimated and measured dietary PO intakes were 12.3 and 11.0 mg/d respectively. The correlation between estimated intake and measured intake of PO was highly significant (r 0.98, P<0.001). Urinary excretion (24 h) and plasma concentrations of PO were significantly related to measured dietary PO intake (r 0.97, P<0.001 and r 0.92, P<0.001 respectively). The relationship between 24 h urinary PO excretion and timed plasma concentrations was also significant (r 0.99, P<0.001). These findings validate the PO database and indicate that 24 h urinary excretion and timed plasma concentrations can be used as biomarkers of PO intake.

Topics: Adult; Aged; Biomarkers; Chromatography, Liquid; Databases, Factual; Diet; Diet Records; Female; Genistein; Humans; Isoflavones; Male; Mass Spectrometry; Middle Aged; Phytoestrogens; Plant Preparations; Reproducibility of Results

Equol is a metabolite produced in vivo from the soy phytoestrogen daidzein by the action of gut microflora. It is known to be estrogenic, so human exposure to equol could have significant biological effects. Equol is a chiral molecule that can exist as the enantiomers R-equol and S-equol. To study the biological activity of racemic (+/-)-equol, as well as that of its pure enantiomers, we developed an efficient and convenient method to prepare (+/-)-equol from available isoflavanoid precursors. Furthermore, we optimized a method to separate the enantiomers of equol by chiral HPLC, and we studied for the first time, the activities of the enantiomers on the two estrogen receptors, ERalpha and ERbeta. In binding assays, S-equol has a high binding affinity, preferential for ERbeta (K(i)[ERbeta]=16 nM; beta/alpha=13 fold), that is comparable to that of genistein (K(i)[ERbeta]=6.7 nM; beta/alpha=16), whereas R-equol binds more weakly and with a preference for ERalpha (K(i)[ERalpha]=50 nM; beta/alpha=0.29). All equol isomers have higher affinity for both ERs than does the biosynthetic precursor daidzein. The availability and the in vitro characterization of the equol enantiomers should enable their biological effects to be studied in detail.

Topics: Antineoplastic Agents; Binding Sites; Carcinoma, Endometrioid; Chromatography, High Pressure Liquid; Equol; Estrogen Receptor alpha; Estrogen Receptor beta; Estrogens, Non-Steroidal; Female; Genistein; Glycine max; Humans; Isoflavones; Phytoestrogens; Phytotherapy; Plant Preparations; Receptors, Estrogen; Tumor Cells, Cultured

The use of estrogen for protection against vascular dysfunction is limited due to its effects on the reproductive system, particularly in males. We postulated that daidzein, an isoflavone with estrogen-like effects on the systemic vasculature but not the reproductive system, might enhance nitric oxide (NO)-mediated cerebral vasodilatation. Male rats were administered vehicle, 17beta-estradiol (0.1 mg/kg s.c.), or daidzein (0.2 mg/kg s.c.) daily for 7 days. Basal and acetylcholine-stimulated NO release was assessed in vitro via carotid arterial rings or in vivo by measuring changes in basilar artery diameter. Levels of protein expression of endothelial NO synthase (eNOS), caveolin-1, and calmodulin were assessed in carotid arteries using Western analysis. Plasma NO levels were doubled by daidzein or 17beta-estradiol. NO production and endothelium-dependent contraction in response to the NOS inhibitor NG-nitro-L-arginine (L-NNA; 100 microM) was enhanced by 50 to 100% in carotid arteries from rats treated with daidzein or 17beta-estradiol. Acetylcholine-induced relaxation was selectively enhanced in carotid arteries from rats treated with daidzein. Similarly, constrictor responses of the basilar artery to L-NNA in vivo were selectively augmented by approximately 100% by 17beta-estradiol treatment and tended to be approximately 50% greater in daidzein-treated rats. Expression of caveolin-1 was decreased, and calmodulin was increased, in vessels from daidzein- or 17beta-estradiol-treated rats. eNOS expression was unaffected by the treatments. These data suggest that short-term administration of daidzein or 17beta-estradiol modulates cerebral artery reactivity in males by enhancing synthesis and release of endothelium-derived NO. Isoflavone therapy may therefore be a feasible approach to protect against cerebrovascular disease and stroke.

Topics: Animals; Carotid Arteries; Cerebral Arteries; Estradiol; Gene Expression; Isoflavones; Male; Nitric Oxide; Nitric Oxide Synthase; Phytoestrogens; Plant Preparations; Rats; Rats, Sprague-Dawley

A diet high in isoflavonoids (soy) is associated with lower breast cancer risk in Asian populations. Due to the low soy intake, dietary lignans may be the more important phytoestrogen class in Western populations. We used a population-based case-control study of breast cancer by age 50 in southern Germany to evaluate the association between dietary intake of different phytoestrogens and premenopausal breast cancer risk. Dietary information was collected from 278 premenopausal cases and 666 age-matched controls, using a validated FFQ. Using multivariate logistic regression, the highest vs. lowest intake quartiles of daidzein and genistein yielded significantly reduced ORs (95% CI) for breast cancer risk of 0.62 (0.40-0.95) and 0.47 (0.29-0.74), respectively. The protective effects of daidzein and genistein were found only for hormone receptor-positive tumors. High intake of other isoflavonoids, e.g., formononetin and biochanin A, as well as the sum of isoflavonoids were not associated with a decrease in risk. Breast cancer risk significantly decreased with a high intake of the plant lignan matairesinol (OR = 0.58, 95% CI 0.37-0.94) but not secoisolariciresinol or the sum of plant lignans. However, both estimated mammalian lignans, enterodiol and enterolactone, were inversely associated with breast cancer risk, with ORs (95% CI) of 0.61 (0.39-0.98) and 0.57 (0.35-0.92), respectively. No effect was found for total phytoestrogen intake. Our results suggest an important role of dietary intake of daidzein and genistein, despite low levels, as well as of matairesinol and mammalian lignans to reduce premenopausal breast cancer risk in this study population.

Topics: Adult; Breast Neoplasms; Case-Control Studies; Female; Genistein; Humans; Isoflavones; Lignans; Middle Aged; Phytoestrogens; Plant Preparations; Premenopause; Risk

Although deoxybenzoins are intermediates in the synthesis of isoflavones, their estrogenic activity has not been investigated. Eleven deoxybenzoins were synthesized and their estrogenicity was evaluated. While their affinities for estrogen receptors (ER) ERalpha and ERbeta were found grossly comparable to those of daidzein, some exhibited considerable selectivity and transcriptional bias toward ERbeta, which appeared to allow for enhancement of ER-mediated transcription via deoxybenzoin binding of ERbeta. Their activity to stimulate the proliferation of ER-positive breast cancer cells and regulate the expression of endogenous and stably transfected reporter genes differed considerably, with some inhibiting cell proliferation while effectively inducing gene expression at the same time. Molecular modeling confirmed that deoxybenzoins fit well in the ligand binding pocket of ERbeta, albeit with different orientations. Our data support the view that deoxybenzoins constitute a promising new class of ERbeta-biased phytoestrogens.

Topics: Alkaline Phosphatase; Benzoin; Cell Division; Cell Line; Enzyme Activation; Estradiol; Estrogen Receptor alpha; Estrogen Receptor beta; Genes, Reporter; Humans; Isoflavones; Models, Molecular; Molecular Conformation; Molecular Structure; Phytoestrogens; Plant Preparations; Receptors, Estrogen; Structure-Activity Relationship; Transcription, Genetic

To identify the metabolites produced from an isoflavonoid, daidzein, by colonic bacteria of rhesus monkeys.. The metabolism of daidzein by the fecal bacteria of nine monkeys was investigated. Daidzein was incubated anaerobically with fecal bacteria, and the metabolites were analyzed by use of liquid chromatography and mass spectrometry.. The fecal bacteria of all of the monkeys metabolized daidzein to various extents. Dihydrodaidzein was found in cultures of fecal bacteria from two monkeys; dihydrodaidzein and equol were found in cultures from four monkeys; dihydrodaidzein, equol, and an unknown metabolite (MW = 244) were found in cultures from one monkey; and dihydrodaidzein and the unknown metabolite were found in cultures from two monkeys.. Similar to that in humans, variation was evident in the metabolism of isoflavonoids by fecal bacteria from rhesus monkeys. Some metabolites produced by fecal bacteria from monkeys were the same as those produced by fecal bacteria from humans.

Topics: Age Factors; Animals; Bacteria; Feces; Female; Humans; Isoflavones; Macaca mulatta; Male; Phytoestrogens

We isolated from soybean miso 8-hydroxyglycitein and 6-hydroxydaidzein as DPPH-radical scavengers, and elucidated their chemical structures by mass spectrometric, and (1)H- and (13)C-NMR spectrosopic analyses. These compounds showed DPPH-radical scavenging activity as high as that of alpha-tocopherol, 8-hydroxygenistein and 8-hydroxydaidzein. This is the first report of the isolation of 8-hydroxyglycitein from a natural source.

Topics: Biphenyl Compounds; Cell Proliferation; Free Radical Scavengers; Glycine max; HL-60 Cells; Humans; Hydrazines; Isoflavones; Molecular Structure; Nuclear Magnetic Resonance, Biomolecular; Phytoestrogens; Picrates

In view of the possible health benefits of phytoestrogens, a pilot study was carried out to quantitate the phytoestrogen content of soy foods and tea commonly consumed in Hong Kong, and also of traditional Chinese medicinal (TCM) products that are prescribed for menopausal symptoms and diseases relating to the menopause. Assays of daidzein and genistein were carried out using high-performance liquid chromatography, after extraction procedures. The TCM products were found to contain phytoestrogen in quantities comparable with soy products. Moreover, certain types of Chinese tea contained large quantities of phytoestrogens in the leaves, but also yielded comparable quantities in the infusion for drinking. The phytoestrogen content of these TCM may provide a scientific basis for their actions. However, clinical efficacy can only be determined by clinical trials.

Topics: Chromatography, High Pressure Liquid; Female; Food Analysis; Genistein; Hong Kong; Humans; Isoflavones; Medicine, Chinese Traditional; Menopause; Phytoestrogens; Pilot Projects; Plant Preparations; Soy Foods; Tea; Women's Health

Circulating hormones are associated with mammographic density, an intermediate marker of breast cancer risk. Differences in circulating hormones, including estrone and testosterone, have been observed in premenopausal women based on their capacity to metabolize daidzein, an isoflavone found predominantly in soybeans. Equol and O-desmethylangolensin (O-DMA) are products of intestinal bacterial metabolism of daidzein. There is interindividual variability in the capacity to produce daidzein metabolites; individuals can be equol producers or non-producers and O-DMA producers or non-producers. We tested the hypothesis that daidzein-metabolizing phenotypes are associated with mammographic density. Participants were recruited from among 92 sedentary, postmenopausal women, ages 50 to 75 years, who participated in a 1-year physical activity intervention. Pre-intervention mammographic density was determined using a computer-assisted, gray-scale thresholding technique. Fifty-five of these women consumed supplemental soy protein (>10 mg daidzein/d) for 3 days and collected a first-void urine sample on the fourth day to determine daidzein-metabolizing phenotypes. Equol and O-DMA concentrations were measured using gas chromatography-mass spectrometry. Associations between daidzein-metabolizing phenotypes and percent mammographic density were adjusted for age, maximum adult weight, gravidity, family history of breast cancer, and serum follicle-stimulating hormone and free testosterone concentrations. Mammographic density was 39% lower in equol producers compared with non-producers (P = 0.04). O-DMA producers had mammographic density 69% greater than non-producers (P = 0.05). These results suggest that particular intestinal bacterial profiles are associated with postmenopausal mammographic density, and these associations are not entirely explained by differences in reproductive or anthropometric characteristics or circulating hormones.

Topics: Aged; Biomarkers, Tumor; Breast; Breast Neoplasms; Cross-Sectional Studies; Dietary Supplements; Equol; Female; Gas Chromatography-Mass Spectrometry; Humans; Intestines; Isoflavones; Mammography; Middle Aged; Obesity; Phenotype; Phytoestrogens; Postmenopause; Soybean Proteins

The effects of soy isoflavones, genistein and daidzein, which exhibit estrogenic, anti-estrogenic and/or tyrosine kinase inhibitory activity, on the dendritic morphology of B16 mouse melanoma cells were quantitatively evaluated and compared with those of 17 beta-estradiol (Est) and tyrphostin, a tyrosine kinase inhibitor. Dendricity was significantly stimulated in the order of Est >> genistein > daidzein = tyrphostin, but not by glycosides of genistein and daidzein. In competition experiments, Est counteracted the stimulatory activity of genistein and daidzein, but enhanced the activity of tyrphostin additively, suggesting that genistein and daidzein agonized Est. In addition, when the concentration ratios of genistein/Est and daidzein/Est were higher than 5000 and 50,000, respectively, genistein and daidzein agonized Est. In contrast, when the ratio of daidzein/Est was lower than 500, daidzein antagonized Est. Furthermore, genistein and daidzein competed with each other in stimulatory activity. These observations suggest that: 1) dendricity is stimulated by agonists (genistein and daidzein) of Est and tyrosine kinase inhibitors (genistein and tyrphostin), 2) the concentration ratio of isoflavone aglycone/Est is very important as one regulatory factor for estrogenic and/or anti-estrogenic activity, and 3) daidzein antagonizes not only Est but also genistein. It is concluded that a quantitative and simple dendricity assay using B16 mouse melanoma cells is available to evaluate estrogenic and anti-estrogenic activity in vitro.

Topics: Animals; Cell Line, Tumor; Drug Evaluation, Preclinical; Estradiol; Estrogen Receptor Modulators; Genistein; Isoflavones; Melanoma, Experimental; Mice; Phytoestrogens; Protein-Tyrosine Kinases; Skin Neoplasms; Tyrphostins

Dietary soy-isoflavones have recently been noted as phytoestrogens with potentially beneficial effects on human health, and they are biologically transformed in the intestinal tract into aglycones and further into several specific metabolites. Here we report that in laying hens daidzin, a soy isoflavone-glycoside, in the diet was transformed into equol, absorbed, transported in circulating peripheral blood, and preferentially accumulated into egg yolk in its conjugated form. Laying hens were fed experimental diets containing two levels of soy isoflavone-glycosides (177 or 528 mg per 100 g diet) for 21 or 42 days, and blood and eggs were collected at 1- to 9-day intervals. HPLC analyses revealed that most of the isoflavones (daidzein, glycitein, and genistein) and a metabolite, equol, were present in blood and egg yolk in conjugated form. The concentration of equol-conjugates in blood plasma and egg yolk was higher than any of the other three isoflavone-conjugates analyzed and, especially in egg yolk, the equol-conjugates comprised no less than 60% of the total isoflavone-conjugates. The isoflavones, including equol, distributed mostly (95%) in the high-density fraction of blood serum, and more (65%) in the granule fraction of egg yolk. These results raise the possibility that feeding domestic animals soy-based fodder produces animal-based foods rich in a more active form of phytoestrogens.

Topics: Animal Feed; Animals; Chickens; Diet; Egg Yolk; Equol; Estrogens, Non-Steroidal; Female; Glycine max; Humans; Intestinal Absorption; Isoflavones; Phytoestrogens; Plant Preparations

Epidemiological evidence suggests a reduction in the incidence of coronary heart disease, cancer and osteoporosis in populations with a high dietary intake of plant estrogen or phytoestrogen. The clinical benefit of phytoestrogens in cereals, vegetables and medicinal plants is attracting increasing attention for the general public. In the present study, we examined the effect of phytoestrogenic isoflavones daidzein and genistein on glucose toxicity-induced cardiac mechanical malfunction simulating diabetic cardiomyopathy. Adult rat ventricular myocytes were isolated and maintained for 24 hours in normal (NG, 5.5 mM) or high glucose (HG, 25.5 mM) medium in the absence or presence of isoflavones daidzein (50 microM) or genistein (20 microM). Cardiac contractile indices were evaluated using an IonOptix MyoCam system including peak shortening (PS), maximal velocity of shortening/relengthening (+/- dL/dt), time-to-PS (TPS) and time-to-90% relengthening (TR90). Myocytes maintained in HG medium displayed altered mechanical function simulating in vivo diabetes including reduced PS, +/- dL/dt and prolonged TR90 associated with normal TPS compared to those from NG myocytes. Interestingly, these HG-induced mechanical dysfunctions were abolished by co-incubation of daidzein or genistein. However, daidzein but not genistein itself depressed PS in NG myocytes. Neither daidzein nor genistein affected any other mechanical parameters tested in NG myocytes. Collectively, these data suggest that the phytoestrogenic isoflavones daidzein and genistein may reduce glucose toxicity-induced cardiac mechanical dysfunction and thus possess therapeutic potential against diabetes-associated cardiac defects.

Topics: Animals; Cells, Cultured; Genistein; Glucose; Heart Diseases; Heart Ventricles; Isoflavones; Male; Myocardial Contraction; Myocytes, Cardiac; Phytoestrogens; Rats; Rats, Sprague-Dawley

Regioselective sulfation of the phytoestrogens daidzein (DZ, 7,4'-dihydroxyisoflavone) and genistein (GS, 5,7,4'-trihydroxyisoflavone) was investigated using human liver cytosol and purified recombinant human sulfotransferase (SULT) isoforms, SULT1A1, SULT1A3, SULT2A1, and SULT1E1. 7-Position-preferential sulfation of DZ and GS was observed in human hepatic cytosols from 3 male and 3 female subjects. Average ratios for 7- to 4'-sulfate formation were 4.5:1 from DZ and 8.4:1 from GS in these human liver cytosols. Apparent K(m) values for the 7- and 4'-sulfation of DZ and GS by these cytosols were similar and in a range from 0.46 to 0.66 microM. All recombinant human SULTs had activity for 7- and 4'-sulfation of these phytoestrogens except for 7-sulfating activity of SULT1A3. SULT1A1 and SULT1E1 exhibited much higher catalytic efficiency, k(cat)/K(m), for 7- and 4'-sulfation of these substrates than did the other two, SULT1A3 and SULT2A1. SULT1A1 showed K(m) values of 0.47 and 0.52 microM for the mono-sulfation of DZ and GS, respectively, which were very similar to those of human cytosol. The observed k(cat)/K(m) indicated that SULT1A1 catalyzed 7-sulfation of DZ and GS at rates 4.4- and 8.8-fold higher, respectively, than such 4'-sulfation. However, with SULT1E1, catalytic efficiency was very similar for the sulfation of both positions. These data strongly suggest that SULT1A1 plays a major role in monosulfation of the phytoestrogens and determines the regioselectivity of sulfation in human hepatic cytosol. A kinetic study for 7,4'-disulfate formation of DZ and GS from their 7- and 4'-monosulfates indicated that SULT1E1 most efficiently catalyzed both reactions among human SULTs.

Topics: Adult; Aged; Arylsulfotransferase; Female; Genistein; Humans; Isoflavones; Liver; Male; Middle Aged; Phytoestrogens; Stereoisomerism; Sulfates; Sulfotransferases

The effects of Lactobacillus gasseri JCM 1131(T) on isoflavonoid levels within the caecum and plasma were assessed in adult mice. Male 5-week-old mice were fed an AIN 93M diet for 30 d. Two groups of mice were administered either L. gasseri JCM 1131(T) (the LGI group) or physiological saline solution (the control (CI) group) daily for 5 d before dissection. The plasma daidzein concentration was significantly higher in the LGI group, however, their plasma equol concentration was significantly less than in the CI group. The total amount of equol present as aglycone in the caecum was significantly greater in the CI group, but there was no significant difference in the total daidzein present as caecal aglycone. In an in vitro incubation of daidzein with the faecal flora of mice, the equol concentration was significantly higher in the CI group. The numbers of lactobacilli present were significantly higher in the LGI group. The present data suggest that the administration of L. gasseri is likely to influence the effect of isoflavonoids on the host via changes in the gastrointestinal environment.

Topics: Animals; Body Weight; Cecum; Colony Count, Microbial; Eating; Equol; Isoflavones; Lactobacillus; Male; Mice; Mice, Inbred ICR; Phytoestrogens

Reactive oxygen species (ROS) are produced by a wide variety of exogenous chemicals and metabolic processes and cause a broad spectrum of damage to biological systems. As a consequence, ROS react with DNA, among many other biological targets, disrupting its structure and functionality. Estrogen-like compounds mediate DNA damage by ROS generation, implying that their effects can be modulated by antioxidants such as catalase, superoxide dismutase, and vitamin C. We examined DNA damage in human lymphocytes and sperm after treatment with four estrogen-like compounds (beta-estradiol, diethylstilbestrol, daidzein, and genistein) and its modulation by flavonoids (quercetin and kaempferol) using the Comet assay. The results indicated that quercetin and kaempferol reduced the DNA damage produced in sperm and lymphocytes by the four estrogenic compounds. The flavonoids also reduced the DNA damage induced by hydrogen peroxide, which was used as a positive control. Our results demonstrate that the antioxidant properties of flavonoids can protect the integrity of human sperm and lymphocyte DNA from ROS induced by estrogenic compounds.

Topics: Antineoplastic Agents; Antioxidants; Carcinogens; Diethylstilbestrol; DNA Damage; Estradiol; Genistein; Humans; Isoflavones; Kaempferols; Lymphocytes; Male; Phytoestrogens; Quercetin; Reactive Oxygen Species; Spermatozoa

Soyfoods contain estrogenic isoflavones--namely, genistein (G) and daidzein (D)--that, like estrogens, display physiological effects in humans and animals. Previously we and others have demonstrated antioxidant and cardioprotective effects of orally ingested soy diets and soy isoflavones. The overall objective of this study was to test the effects of injected soy isoflavones, G and D, on liver lipids, liver free radical scavenger systems, and parameters of cardiovascular risk. Forty male rats were injected with G, D, estradiol (E), or a vehicle control (V) for 6 weeks. At the end of the study, body weight, food intake, feed efficiency ratio (FER), plasma glucose and cholesterol, abdominal fat pad weight, reproductive organ weight, liver weight, liver lipids, and liver free radical scavenger systems were compared. Food intake was significantly (P < .04) higher in the D-, G-, and E-treated animals compared with V-treated animals. FER was lower (P < .001) in D-, G-, and E-treated animals compared with the V- and the E-treated animals. Body weight, testis weight, and prostate weight were markedly (P < .001) lower in the E-treated animals compared with D-, G-, and V-treated animals. Intraabdominal fat pad weights were also significantly (P < .001) lower in the E group, although this effect was lost when corrected for body weight. Liver weights were considerably lower in the D-, G-, and E-treated animals versus the V group (P < .001). Total plasma cholesterol was reduced (P < .05) in D- and E-treated animals versus the V group. Liver lipids appeared to be unchanged by the isoflavones and slightly elevated by E treatment (P < .02). Liver catalase levels were numerically higher in the D- and E-treated animals compared with the V group (P < .1). Similarly, Cu/Zn superoxide dismutase (SOD) activity was significantly elevated in the D and E groups (P < .01), while G treatment (P < .03) elevated SOD to a lesser degree, versus the V group. These results suggest that subcutaneous injections of the naturally occurring soy isoflavone D and, to a lesser extent, G exert cardioprotective effects and stimulate antioxidant systems, while minimizing the undesirable effects elicited by E treatment.

Topics: Animals; Body Weight; Catalase; Cholesterol; Estradiol; Free Radical Scavengers; Genistein; Injections, Subcutaneous; Isoflavones; Lipid Metabolism; Lipids; Liver; Male; Organ Size; Phytoestrogens; Random Allocation; Rats; Rats, Sprague-Dawley; Superoxide Dismutase

Endocrine-disrupting chemicals (EDCs) are giving rise to serious concerns for humans and wildlife. Phytoestrogens, such as daidzein and genistein in plants, and organochlorine pesticides are suspected EDCs, because their chemical structure is similar to that of natural or synthetic estrogens and they have estrogenic activity in vitro and in vivo. We assessed estrogenic activity and dietary phytoestrogen and organochlorine pesticide contents of various fish diets made in the United Kingdom, and compared them with those features of diets made in Japan that were tested in a previous study. Genistein and daidzein were detected in all of the diets. Using an in vitro bioassay, many of these diets had higher activation of estrogen beta-receptors than estrogen alpha-receptors. Organochlorine pesticides such as hexachlorobenzene, beta-benzene hexachloride (BHC), and gamma-BHC were detected in all fish diets. On the basis of these data, we investigated the effect of differing dietary phytoestrogen content in Japanese fish diets on hepatic vitellogenin production and reproduction (fecundity and fertility) in medaka (Oryzias latipes). Assessment of the effects of a 28-day feeding period on reproduction of paired medaka did not indicate significant differences in the number of eggs produced and fertility among all feeding groups. However, hepatic vitellogenin values were significantly higher for male medaka fed diet C (genistein, 58.5 +/- 0.6 microg/g; daidzein, 37.3 +/- 0.2 microg/g) for 28 days compared with those fed diet A (genistein, < 0.8 microg/g; daidzein, < 0.8 microg/g) or diet B (genistein, 1.4 +/- 0.1 microg/g; daidzein, 2.0 +/- 0.1 microg/g). Our findings indicate that fish diets containing high amounts of phytoestrogens, such as diet C, have the potential to induce hepatic vitellogenin production in male medaka, even if reproductive parameters are unaffected. Therefore, some diets, by affecting vitellogenin production in males, may alter estrogenic activity of in vivo tests designed to determine activity of test compounds added to the diet.

Topics: Animal Feed; Animals; Diet; Endocrine Glands; Estradiol Congeners; Female; Genistein; Humans; Hydrocarbons, Chlorinated; Isoflavones; Liver; Male; Oryzias; Pesticides; Phytoestrogens; Reproduction; Vitellogenins

To explore the effect of soybean isoflavone on the cognitive function in ovariectomized mice and to study the cognitive function mechanism of soybean isoflavone.. Forty-five Kunming female mice were randomly assigned into 5 groups: A (sham operated); B (ovariectomized, OVX); C (OVX + low dose soybean isoflavone); D (OVX + moderate dose soybean isoflavone); and E (OVX + high dose soybean isoflavone). The experiment lasted 60 days.. Ovariectomy significantly elongated the destination time of water maze, shortened the latent time of step-down test, decreased SOD of serum and Na+ K+ -ATPase and Ca2+ Mg2+ -ATPase of brain, and increased malonaldehyde of serum and monoamine oxidase of brain, which-could be inhibited by soybean isoflavone consumption.. The continuous oral administration of soybean isoflavone can improve the cognitive function of ovariectomized mice.

Topics: Animals; Cognition; Female; Genistein; Glycine max; Isoflavones; Mice; Ovariectomy; Phytoestrogens

Phytoestrogens (isoflavones and lignans) are of increasing interest due to their potential to prevent certain types of complex diseases. However, epidemiological evidence is needed on the levels of phytoestrogens and their metabolites in foods and biological fluids in relation to risk of these diseases. We report an assay for phytoestrogens which is sensitive, accurate, and uses low volumes of sample. Suitable for epidemiological studies, the assay consists of a simple sample preparation procedure and has been developed for the analysis of five isoflavones (daidzein, O-desmethylangolensin, equol, genistein, and glycitein) and two lignans (enterodiol and enterolactone), which requires only 200 microl of urine and utilizes one solid-phase extraction stage for sample preparation prior to derivatization for GC/MS analysis. Limits of detection were in the region 1.2 ng/ml (enterodiol) to 5.3ng/ml (enterolactone) and the method performed well in the UK Government's Food Standards Agency-sponsored quality assurance scheme for phytoestrogens. For the first time, average levels of all the above phytoestrogens were measured in samples of urine collected from a free living population sample of women. Results show a large range in both the amount and the type of phytoestrogens excreted.

Topics: 4-Butyrolactone; Equol; Female; Gas Chromatography-Mass Spectrometry; Genistein; Humans; Isoflavones; Lignans; Molecular Structure; Phytoestrogens; Plant Preparations; Reproducibility of Results; Sensitivity and Specificity

For the simultaneous assessment of in vitro carcinogenicity and mutagenicity of phytoestrogens, the abilities of 5 phytoestrogens, daidzein, genistein, biochanin A, prunetin, and coumestrol, to induce cell transformation and genetic effects were examined using the Syrian hamster embryo (SHE) cell model. Cellular growth was inhibited by all phytoestrogens in a concentration-related manner. The growth inhibitory effect of the compounds was ranked: genistein, prunetin > coumestrol > biochanin A > daidzein, which did not correspond to their apoptosis-inducing abilities. Morphological transformation in SHE cells was elicited by all phytoestrogens, except, prunetin. The transforming activities were ranked as follows: genistein > coumestrol > daidzein > biochanin A. Somatic mutations in SHE cells at the Na(+)/K(+) ATPase and hprt loci were induced only by genistein, coumestrol, or daidzein. Chromosome aberrations were induced by genistein or coumestrol, and aneuploidy in the near diploid range was occurred by genistein or biochanin A. Genistein, biochanin A or daidzein induced DNA adduct formation in SHE cells with the abilities: genistein > biochanin A > daidzein. Prunetin was negative for any of these genetic endpoints. Our results provide evidence that genistein, coumestrol, daidzein and biochanin A induce cell transformation in SHE cells and that the transforming activities of these phytoestrogens correspond to at least 2 of the mutagenic effects by each phytoestrogen, i.e., gene mutations, chromosome aberrations, aneuploidy or DNA adduct formation, suggesting the possible involvement of mutagenicity in the initiation of phytoestrogen-induced carcinogenesis.

Topics: Animals; Anticarcinogenic Agents; Apoptosis; Cell Line; Cell Transformation, Neoplastic; Chromosome Aberrations; Coumestrol; Cricetinae; DNA Adducts; Dose-Response Relationship, Drug; Embryo, Mammalian; Estrogens, Non-Steroidal; Genistein; Isoflavones; Mesocricetus; Metaphase; Models, Chemical; Mutagens; Mutation; Phytoestrogens; Plant Preparations; Time Factors

Estrogen replacement therapy (ERT) is reported to lower the incidence of cardiovascular disease in postmenopausal women. ERT also lowers the levels of oxidatively modified low-density lipoprotein (LDL). Because modified LDL can mediate the development of atherosclerosis by inflammatory processes, ERT may exert its LDL protective effect through enhanced antioxidant activity in vascular tissues. Plant sources of estrogenic compounds have been used as alternatives for ERT because they avoid a number of negative health effects produced by estrogen. In this study, the antioxidant properties of the soy isoflavone metabolite, equol (an estrogenic metabolite of daidzein) were studied. Equol has a greater antioxidant activity than the parent isoflavone compounds genistein and daidzein, found in high concentration in soy. Equol inhibits LDL oxidation in vitro and LDL oxidative modification by J774 monocyte/macrophages to LDL(-), an electronegative modified LDL found in human plasma. An antioxidant effect of equol was found to be mediated by inhibition of superoxide radical (O(2)(-*)) production and manifested through enhanced levels of free nitric oxide (NO) that prevents LDL modification. Thus, when NO levels were increased by donor agents, generators, or compounds that facilitate nitric oxide synthase activity, LDL(-) formation by J774 cells was strongly inhibited. Conversely, inhibition of NO production enhanced LDL(-) formation, and the combination of reduced NO and increased O(2)(-*) production yielded maximum LDL(-) formation. Pretreatment of cells with equol inhibited production of O(2)(-*) by J774 cells apparently via the inactivation of the reduced nicotinamide adenine dinucleotide phosphate oxidase complex. Decreased O(2)(-*) production resulted in increased free NO levels (but not total NO production) indicating that decreased reactions between O(2)(-*) and NO are an outcome of equol's antioxidant activity in cell culture.

Topics: Animals; Antioxidants; Biological Availability; Cells, Cultured; Chromans; Copper; Equol; Estrogens, Non-Steroidal; Genistein; Iron; Isoflavones; Kinetics; Lipid Peroxidation; Lipoproteins, LDL; Macrophages; Mice; Monocytes; NADPH Oxidases; Nitric Oxide; Oxidation-Reduction; Phytoestrogens; Plant Preparations; Superoxides; Time Factors

In human prostate cancer cells, the availability of the steroid hormone 1,25-dihydroxyvitamin D(3) for antimitotic action is determined through the activity of the two enzymes CYP24 and CYP27B1, viz. 25-hydroxyvitamin D-24-hydroxylase and 25-hydroxyvitamin D-1alpha-hydroxylase. High performance liquid chromatography (HPLC) analysis of [(3)H]25(OH)D(3) metabolism in human prostate cancer DU-145 cells revealed that genistein and other isoflavonoids, such as dihydrogenistein and daidzein, as well as the antiestrogenic compound ICI 182,780, inhibited Vitamin D-metabolizing enzyme activities. Reverse transcriptase-polymerase chain reaction (RT-PCR) showed that only in case of genistein this was due to transcriptional inhibition of CYP24 and CYP27B1 gene expressions. In case of CYP27B1, reduction of gene activity involves histone deacetylation because genistein was inactive in the presence of the histone deactylase inhibitor trichostatin A. In contrast, under the same condition, CYP24 gene activity was largely suppressed. In summary, our results suggest that a combined effect of genistein and trichostatin A could increase the responsiveness of human prostate cancer cells to the antiproliferative action of 1,25-dihydroxyvitamin D(3).

Topics: 25-Hydroxyvitamin D3 1-alpha-Hydroxylase; Cell Division; Chromatography, High Pressure Liquid; Cytochrome P-450 Enzyme Inhibitors; Enzyme Inhibitors; Estradiol; Estrogen Antagonists; Estrogens, Non-Steroidal; Fulvestrant; Genistein; Histone Deacetylases; Humans; Hydroxamic Acids; Isoflavones; Male; Phytoestrogens; Plant Preparations; Prostatic Neoplasms; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Steroid Hydroxylases; Time Factors; Transcription, Genetic; Tumor Cells, Cultured; Vitamin D; Vitamin D3 24-Hydroxylase

Ovarian steroid hormones, estrogen and progestin, affect the function of the serotonin neural system by inhibiting serotonin re-uptake through allosteric interaction with the serotonin transporter (SERT) in a nongenomic mechanism. Blocking or reducing serotonin re-uptake at the synapse alleviates depression. The aim of this study was to test the effect of compounds of the isoflavan and isoflavene groups, subclasses of the flavonoids family, on serotonin re-uptake and to compare the results with the effect of other known phytoestrogens like genistein and daidzein to relate the activity of these compounds to their structure. The effect of these compounds on the re-uptake of radioactive serotonin was assayed in HEK-293 cells stably expressed the recombinant human serotonin transporter (hSERT). The results demonstrated that the isoflavans glabridin and 4'-O-methylglabridin (4'-OMeG) and the isoflavene glabrene inhibited serotonin re-uptake by 60, 53 and 47%, respectively, at 50 microM, whereas resorcinol, the isoflavan 2'-O-methylglabridin (2'-OMeG), and the isoflavones genistein and daidzein were inactive. The inhibition of serotonin re-uptake is dose dependant with glabridin and estradiol. These results emphasize the importance of the lipophilic part of the isoflavans, as well as the hydroxyl at position 2' on ring B. In conclusion, this study showed that several isoflavans are unique phytoestrogens, which like estradiol, affects the serotonergic system and inhibits serotonin re-uptake and, thus, potentially may be beneficial for mild to moderate depression in pre- and postmenopausal women.

Topics: Carrier Proteins; Cells, Cultured; Depressive Disorder; Dose-Response Relationship, Drug; Estradiol; Estrogens, Non-Steroidal; Female; Flavonoids; Genistein; Glycyrrhiza; Humans; Isoflavones; Membrane Glycoproteins; Membrane Transport Proteins; Menopause; Molecular Structure; Nerve Tissue Proteins; Phenols; Phytoestrogens; Plant Extracts; Plant Preparations; Resorcinols; Selective Serotonin Reuptake Inhibitors; Serotonin; Serotonin Plasma Membrane Transport Proteins

In an international context of promoting scientific research on food safety, the interest in molecules having potential hormonal disrupting effects is growing. While industrial endocrine disruptors (phthalates, alkylphenols, PCBs, etc.) have been studied for several years, natural compounds like phytoestrogens remain less investigated. Accordingly, a research project was initiated with its main objectives to develop efficient analytical methods for a wide range of phytoestrogens in various food matrices, and to evaluate their occurrence in food products. Electrospray ionization with tandem mass spectrometric (MS/MS) analysis of isoflavones (genistein, daidzein, equol, formononetin, biochanin A), lignans (enterolactone, enterodiol), and coumestans (coumestrol) was investigated. This study revealed the formation of a large number of fragment ions in both positive and negative modes, corresponding to specific cleavages of the hydroxyl, carbonyl, and/or methoxy groups, and to Retro-Diels-Alder reactions. An LC/ESI-MS/MS method was developed consistent with the 2002/657/EC European decision criteria. An extraction and clean-up method was developed for milk samples. The identification limit for the proposed method appears to be under 1 ng/mL. The developed methodology was applied to various milk samples, and the occurrence of isoflavones (particularly equol) was demonstrated in the concentration range 1-30 ng/mL. The efficiency of the proposed analytical method permitted evaluation of a new and promising approach to a global risk assessment of natural estrogenic active substances including phytoestrogens and their metabolites.

Topics: 4-Butyrolactone; Animals; Cattle; Chromans; Chromatography, Liquid; Coumestrol; Equol; Estrogens, Non-Steroidal; Genistein; Ions; Isoflavones; Lignans; Milk; Molecular Structure; Phytoestrogens; Plant Preparations; Spectrometry, Mass, Electrospray Ionization

Many commercial laboratory diets have detectable levels of isoflavones (e.g., phytoestrogens such as genistein [GN]) that have weak estrogenic activity both in vitro and in vivo. During validation studies of the uterotrophic bioassay, diet samples from 20 participating laboratories were collected and analyzed for three major phytoestrogens: GN, daidzein (DN), and coumestrol (CM). Soy phytoestrogens GN and DN were found at total phytoestrogen levels from 100 to 540 microg/g laboratory diet; a forage phytoestrogen, CM, ranged from nondetectable to 4 microg/g laboratory diet. The phytoestrogen levels were compared with both baseline uterine weights of the control groups and with the relative uterine weight increase of groups administered two weak estrogen agonists: bisphenol A (BPA) and nonylphenol (NP). The comparison uses a working assumption of additivity among the phytoestrogens, despite several significant qualifications to this assumption, to estimate total genistein equivalents (TGE). Some evidence was found that phytoestrogen levels in the diet > 325-350 microg/g TGE could diminish the responsiveness of the uterotrophic bioassay to weak agonists. This was especially true for the case of the intact, immature female version of the uterotrophic bioassay, where higher food consumption relative to body weight leads to higher intakes of dietary phytoestrogens versus ovariectomized adults. This dietary level is sufficient in the immature female to approach a biological lowest observable effect level for GN of 40-50 mg/kg/day. These same data, however, show that low to moderate levels of dietary phytoestrogens do not substantially affect the responsiveness of the assay with weak estrogen receptor agonists such as NP and BPA. Therefore, laboratories conducting the uterotrophic bioassay for either research or regulatory purposes may routinely use diets containing levels of phytoestrogens < 325-350 microg/g TGE without impairing the responsiveness of the bioassay.

Topics: Administration, Oral; Animal Feed; Animals; Biological Assay; Coumestrol; Diet; Dose-Response Relationship, Drug; Endocrine System; Environmental Pollutants; Enzyme Inhibitors; Estrogens, Non-Steroidal; Female; Genistein; Housing, Animal; Isoflavones; Laboratories; Observer Variation; Phytoestrogens; Plant Preparations; Rats; Reproducibility of Results; Research Design; Uterus

It is well known that neonatal exposure to estrogen induces masculinization or defeminization of the brain. In this study, the effects of neonatal treatment with two kinds of soybean isoflavone aglycone, genistein (GS) and daidzein (DZ), on the estrous cycle and lordosis behavior were investigated. Female rats were injected subcutaneously with 1 mg GS, 1 mg DZ, 100 microg estradiol (E2), or oil daily for 5 days from birth. As a result, vaginal opening was advanced in GS- or E2-treated females. A vaginal smear check indicated that oil- or DZ-treated females showed a constant 4- or 5-day estrous cycle, whereas GS- or E2-treated rats showed a persistent or prolonged estrus. Ovariectomy was performed in all females at 60 days of age. The ovaries in the GS- or E2-treated groups were smaller than those in the oil- and DZ-treated groups and contained no corpora lutea. In the DZ group, although corpora lutea were seen, ovaries were smaller than that of control females. Behavioral tests were carried out after implantation of E2-tubes. All of the oil- or DZ-treated females showed lordosis with a high lordosis quotient (LQ). On the other hand, as male rats, LQs were extremely low in the E2-treated group, when compared to the oil-treated group. In the GS-treated group, the mean LQ was lower than that in the oil-treated group, but higher than those in the E2-treated female or male groups. These results suggest that genistein acts as an estrogen in the sexual differentiation of the brain and causes defeminization of the brain in regulating lordosis and the estrous cycle in rats. In addition, neonatal daidzein also has some influence on ovarian function.

Topics: Animals; Animals, Newborn; Estradiol; Estrogens, Non-Steroidal; Estrous Cycle; Female; Genistein; Isoflavones; Ovary; Phytoestrogens; Plant Preparations; Plants; Posture; Rats; Rats, Wistar; Sex Characteristics; Sex Differentiation; Sexual Behavior, Animal

The effects of phytoestrogens on neuronal nicotinic acetylcholine receptor/channels were examined by expressing recombinant channels in Xenopus oocytes. When functional channels were expressed with human alpha4 and beta2 subunits, daidzein (10 and 100 microM) partially inhibited the ionic current activated by acetylcholine. The current inhibition was also observed when functional channels were expressed with human alpha3 and beta4 subunits or rat homologues. Genistin (100 microM) also inhibited the acetylcholine-activated current. Tamoxifen (100 microM), an antiestrogen did not antagonize the inhibition by daidzein. The results suggest that phytoestrogens, like estrogens and xenoestrogens, block human neuronal acetylcholine receptors through non-genomic mechanisms.

Topics: Animals; Female; Humans; Isoflavones; Neural Inhibition; Nicotinic Antagonists; Phytoestrogens; Plant Preparations; Rats; Receptors, Nicotinic; Recombinant Proteins; Xenopus

While estrogen-related receptors (ERRalpha, ERRbeta, and ERRgamma) share a high amino acid sequence homology with estrogen receptors (ERs), estrogens are not ligands of ERRs. Structure-function studies from this and other laboratories have revealed that ERRs have small ligand-binding pockets and have provided evidence to show that these receptors can activate gene transcription in a constitutive manner. To address the question as to whether there is any agonist for ERRs, our laboratory recently performed virtual ligand screening on ERRalpha that predicted flavone and isoflavone phytoestrogens to be ligands of this receptor. Our mammalian cell transfection and mammalian two-hybrid experiments revealed that three isoflavones (genistein, daidzein, and biochanin A) and one flavone (6,3',4'-trihydroxyflavone) behaved as agonists of ERRs. These phytoestrogens induced the activity of ERRalpha at concentrations that are comparable to those for the activation of ERalpha and ERbeta. In this study, we also used the results of ERRalpha ligand-binding site mutant, F232A, to verify our ERRalpha hypothetical computer model. Our recent ERR research has determined for the first time that flavone and isoflavone phytoestrogens are agonists of ERRs. In addition, our studies have demonstrated that an approach that combines structure-based virtual screening and receptor functional assays can identify novel ligands of orphan nuclear receptors.

Topics: Binding, Competitive; Computer Simulation; Databases, Factual; ERRalpha Estrogen-Related Receptor; Flavones; Flavonoids; Genistein; HeLa Cells; Humans; Isoflavones; Ligands; Luciferases; Models, Molecular; Molecular Structure; Phytoestrogens; Plant Preparations; Receptors, Cytoplasmic and Nuclear; Receptors, Estrogen; Recombinant Fusion Proteins; Structure-Activity Relationship; Transfection

Binding of estrogen receptor (ER) to estrogen response element (ERE) induces gene activation and is an important step in estrogen-induced biological effects. Here, we investigated the effects of some dietary phytoestrogens such as the isoflavones genistein and daidzein, its metabolite equol, and the coumestane coumestrol on the binding rate of ERalpha and ERbeta to ERE by a nonradioactive real-time method, the Biacore Technology. ERalpha and ERbeta were able to bind to ERE immobilized on the surface of a sensor chip even in the absence of estrogens. 17beta-Estradiol and phytoestrogens induced an increase in ER binding to ERE in a concentration-dependent manner. 17beta-Estradiol was a more potent activator of binding than the phytoestrogens studied. The concentrations of 17beta-estradiol inducing an increase in the binding response of ERalpha and ERbeta to ERE by 50% (EC(50)) as compared to unliganded ER were 0.03 and 0.01 microM, respectively. Regarding the efficacy of activation of ERalpha, from the most to the least effective compound, the sequence and the EC(50) were as follows: 17beta-estradiol (0.03 microM) > coumestrol (0.2 microM) > equol (3.5 microM) > genistein (15 microM) > daidzein (>300 microM) and for ERbeta 17beta-estradiol (0.01 microM) > coumestrol (0.025 microM) > genistein (0.03 microM) > daidzein (0.35 microM) > equol (0.4 microM). The ratios EC(50)alpha/EC(50)beta were calculated to be for 17beta-estradiol, 3; coumestrol, 8; equol, 8.8; genistein, 500; daidzein > 850. These ratios indicate that genistein and daidzein preferentially activate the binding of ERbeta to ERE. The endogenous hormone 17beta-estradiol as well as coumestrol and daidzein metabolite equol activate the binding of ERbeta to ERE only slightly more effectively than the binding of ERalpha to ERE. Thus, the effect of daidzein can be changed from a specific activator of ERbeta to an activator of both ER isotypes alpha and beta in humans who are able to convert daidzein to equol. While the results of the measurements with ERalpha were in line with the binding affinities of compounds tested for ER, there was a distinct difference between our results and the binding affinities of phytoestrogens for the ERbeta. This leads to the conclusion that phytoestrogens differ not only in their binding affinities for the ER, but also in their potential to increase the rate of receptor binding to the ERE.

Topics: Diet; DNA; Estrogen Receptor alpha; Estrogen Receptor beta; Genistein; Isoflavones; Phytoestrogens; Plant Preparations; Receptors, Estrogen; Response Elements

To evaluate the effects of daidzein on sperm quality of male mice.. Three different doses of daidzein were supplemented to pubertal male mice for 21 days(5 mg/kg, 100 mg/kg, 1,000 mg/kg ration). The viability of sperm was determined by eosin-Y, the acrosome was observed by Wright-Giemsa staining, and the testosterone was measured by radioimmunoassay.. Daidzein at the dose of 5 mg/kg ration significantly increased serum testosterone levels (P < 0.01), prompted testis gain (P < 0.05), and improved spermatozoa quality. Daidzein at dose of 1,000 mg/kg ration could inhibit the secretion of serum testosterone (P < 0.01), without significant variation in spermatozoa quality. Daidzein at dose of 100 mg/kg ration did not significantly affect sperm quality and other index.. Daidzein can affect sperm quality and in dosage-dependant ways.

Topics: Animals; Cell Survival; Dose-Response Relationship, Drug; Isoflavones; Male; Mice; Mice, Inbred Strains; Organ Size; Phytoestrogens; Spermatozoa; Testis; Testosterone

The purpose of the study reported here was to determine the effects of dietary phytoestrogens on the time of vaginal opening (VO) in immature CD-1 mice, and to correlate it with phytoestrogen and total metabolizable energy (ME) contents of the diet in an effort to determine the most appropriate diets(s) for comparing or evaluating the estrogenic or antiestrogenic activity of endocrine disruptor compounds (EDC). Mice were weaned at postnatal day (PND) 15 and fed the test diets from PND 15 to 30. Vaginal opening was recorded from PND 20 to 30. The phytoestrogen content of the diet was highly predictive (P < 0.0001) of the proportion of mice with VO at PND 24. Total ME content also was significantly (P < 0.01) correlated with time of VO, although this variable was somewhat less predictive than was phytoestrogen content. Time of VO in mice was significantly (P < 0.05) accelerated in mice fed diets high in phytoestrogens, compared with those containing low phytoestrogen content. It was concluded that: dietary daidzein and genistein can significantly (P < 0.01) accelerate the time of VO in CD-1 mice; the advancement in time of VO is more highly correlated with daidzein and genistein contents of the diets than with total ME content; advancement in the time of VO is a sensitive end point for evaluating the estrogenic activity of EDCs, and should be part of the standard protocol for evaluating EDCs. Phytoestrogen-free diet(s) containing the same amount of ME should be used in bioassays that compare the time of VO, or increases in uterine weight as end points for evaluating the estrogenic activity of an EDC.

Topics: Animal Feed; Animals; Biological Assay; Diet; Dose-Response Relationship, Drug; Energy Metabolism; Estrogens, Non-Steroidal; Female; Food, Formulated; Genistein; Isoflavones; Mice; Mice, Inbred Strains; Phytoestrogens; Plant Preparations; Sexual Maturation; Vagina

Daidzein, coumestrol and zearalenone - compounds called phytoestrogens, considered as active biological factors affecting many important physiological and biochemical processes appeared to be also significant regulators of adipocyte metabolism. In our experiments the influence of daidzein (0.01, 0.1 and 1 mM), coumestrol (0.001, 0.01 and 0.1 mM), zearalenone (0.01, 0.1 and 1 mM) and estradiol (0.01, 0.1 and 1 mM) on basal and insulin-stimulated (1 nM) lipogenesis from glucose and acetate was tested in adipocytes isolated from growing (160 +/- 5 g b.w) male Wistar rats. All tested compounds significantly attenuated glucose conversion to lipids. In the case of daidzein and coumestrol, this effect was probably due to inhibition of glycolysis. Daidzein (0.01, 0.1 and 1 mM), coumestrol (0.01 and 0.1 mM) and zearalenone (0.01, 0.1 and 1 mM) affected also basal and epinephrine-stimulated (1 microM) lipolysis. Daidzein (0.01 and 1 mM) augmented basal glycerides breakdown in adipocytes. The epinephrine-induced lipolysis was dependent on daidzein concentration and its stimulatory (0.1 mM) or inhibitory (1 mM) influence was observed. Zearalenone changed lipolysis only at the concentration of 1 mM and its effect was contradictory in the absence or presence of epinephrine (the stimulatory or inhibitory effect, respectively). Results obtained in experiments with inhibitors (insulin, 1 nM and H-89, 50 microM) and activators (dibutyryl-cAMP, 1 mM and forskolin, 1 microM) of lipolysis allowed us to assume that daidzein augmented basal lipolysis acting on PKA activity. The inhibitory effect of daidzein and zearalenone on epinephrine-induced lipolysis is probably due to restriction of HSL action. The influence of coumestrol on glycerides breakdown was less marked. Estradiol augmented only epinephrine-stimulated lipolysis.

Topics: Adipocytes; Animals; Coumestrol; Dose-Response Relationship, Drug; Epinephrine; Estradiol; Estrogens, Non-Steroidal; Isoflavones; Lipolysis; Male; Phytoestrogens; Phytotherapy; Plant Preparations; Rats; Rats, Wistar; Zearalenone

Phytoestrogens are plant substances that are similar to 17-beta-estradiol and produce estrogenic effects. A protective role in the development of breast and prostate cancer has been hypothesized. Estrogen receptors and their variant forms play a significant role in the pathogenesis of hepatocellular carcinoma (HCC); therefore weak estrogenic substances in the diet may play a role in its development. To investigate the role of phytoestrogens in HCC an investigation of dietary intake of these substances has been performed. Cases, patients at first diagnosis of cirrhosis or HCC were chosen. Questionnaire was built up using indications from previously published papers, extending the registration of details of the diet to reconstruct intake of nutrients for the last year. Interviews were always performed by the same dietician. Quantities determined with the help of photos of servings. Data were analyzed with Winfood database completed with data regarding content in phytoestrogens of food, beverages and seasonings. So far 92 cirrhotic patients and 32 HCCs have been interviewed. No significant difference was registered among the two groups regarding total caloric intake or single nutrients (lipids, carbohydrates, proteins). A significant lower intake of genistein was evidenced in patients at first diagnosis of HCC in comparison with cirrhotics; no significant difference was found in daidzein intake. Lignans intake was strictly related with wine intake; intake was significantly lower in cases only when wine was taken into account otherwise it was similar. Results can be summarized as follows: (1) there are no clear-cut differences (both qualitative or quantitative) between cirrhotics and HCC patients in the overall daily caloric intake while; (2) definite differences exist in the intake of some of the phytoestrogens (genistein, SEC, MAT); (3) differences between cases and controls in SEC and MAT are mainly attributable to lower alcohol intake in cases while; (4) significantly lower genistein intake in HCC only seems due to personal preferences of patients. In conclusion, these differences that we have evidenced in the diet in regard to estrogen-like substances may be relevant in modulating the risk of developing HCC in cirrhotic patients.

Topics: Adult; Aged; Carcinoma, Hepatocellular; Diet; Estrogens, Non-Steroidal; Female; Genistein; Humans; Isoflavones; Liver Cirrhosis; Male; Middle Aged; Phytoestrogens; Plant Preparations; Protective Agents; Surveys and Questionnaires

Animal and organ culture experiments have shown beneficial inhibitory estrogen effects on post injury neointima development. The purpose of this study was to investigate whether such estrogen effects are influenced by the estrogen receptor antagonist ICI 182,780. Different concentrations of 17beta-estradiol and the phytoestrogens genistein and daidzein were tested.. Female New Zealand White rabbits were benumbed. In situ vascular injury of the thoracic and abdominal aorta was performed by a 3F Fogarty catheter. Segments of 5 mm were randomised and held in culture for 21 days. Three test series were performed: 1) control group--20 microM ICI--30 microM ICI--40 microM ICI. 2) control group--20 microM ICI--40 microM 17beta-estradiol--40 microM 17beta-estradiol + 20 microM ICI. 3) control group--20 microM ICI--40 microM daidzein--40 microM daidzein + 20 microM ICI--20 microM genistein--20 microM genistein + 20 microM ICI. After 21 days the neointima-media-ratio was evaluated.. 1) Treatment with ICI 182,780 did not reduce neointima formation significantly (p = 0.05). 2) 40 microM 17beta-estradiol alone (p < 0.0001) and in combination with 20 microM ICI (p < 0.0001) reduced neointima formation significantly. 3) 20 microM genistein alone (p = 0.0083) and combined with 20 microM ICI (p = 0.0053) reduced neointima formation significantly. 40 microM daidzein did not have a significant (p = 0.0637) effect.. The estrogen receptor antagonist ICI 182,780 did not modulate the inhibitory estrogen effects on post injury neointima formation. These results do not support the idea that such effects are mediated by vascular estrogen receptors.

Topics: Animals; Aorta; Estradiol; Estrogen Antagonists; Estrogen Receptor Modulators; Female; Fulvestrant; Genistein; Isoflavones; Phytoestrogens; Plant Preparations; Pulmonary Artery; Rabbits; Random Allocation; Tunica Intima

Phytoestrogens have been shown to inhibit platelet activation by blocking platelet calcium channels. This study examined the effect of several synthetic derivatives of trans-resveratrol, genistein, and daidzein on platelet free intracellular calcium ([Ca2+]i) elevation in thrombin-activated platelets and the possible mechanisms of this inhibitory effect. Studies were conducted on fresh human platelets from healthy volunteers. The fluorescent dye fura-2 was used to monitor [Ca2+]i in platelets. At 10 microM-resveratrol, triacetyl-trans-resveratrol, and trimethoxy-trans-resveratrol produced, respectively, 57 +/- 4%, 40 +/- 4%, and 21 +/- 1% inhibition; genistein, acetylgenistein, and dihydrogenistein produced 51 +/- 10%, 26 +/- 7%, and 16 +/- 2% inhibition, respectively; daidzein and diacetyldaidzein produced 56 +/- 5% and 45 +/- 10% inhibition of thrombin-induced [Ca2+]i elevation. The inhibitory effect was immediate and appeared to directly affect the calcium influx channels. Phytoestrogen action on [Ca2+]i did not cause alteration in nitric oxide signaling. Tyrosine phosphorylation was not involved in the inhibition of [Ca2+]i elevation by phytoestrogens, because the percent inhibition produced by the tyrosine kinase inhibitor genistein and its inactive analogue daidzein on thrombin-induced and thapsigargin-induced [Ca2+]i elevation was not significantly different for either compound at any concentration tested. Structure-activity relationship studies on this limited set of compounds reveal the requirements for the stilbene pharmacophore for the calcium-blocking activity.

Topics: Adult; Blood Platelets; Calcium; Calcium Channel Blockers; Calcium Channels; Diet; Dose-Response Relationship, Drug; Estradiol Congeners; Estrogens, Non-Steroidal; Genistein; Humans; Isoflavones; Middle Aged; Phytoestrogens; Plant Preparations; Plants; Structure-Activity Relationship

We used the yeast estrogen screen (YES) containing a human estrogen receptor to evaluate the estrogenic activity of extracts obtained from Nigella damascena seeds. Alcohol extracts obtained by direct extraction of seeds showed a low estrogenic activity, while the alcohol extract obtained after extraction with solvents of increasing polarity showed a strong estrogenic activity. This suggests the presence in Nigella of polar components whose activity can be clearly demonstrated after previous elimination of interacting apolar components that may mask the activity of more polar components. The response of both alcohol fractions follow a bell-shaped curve indicating a concentration-dependent relationship.

Topics: Estrogens, Non-Steroidal; Gene Expression; Genes, Reporter; Genistein; Humans; Isoflavones; Nigella damascena; Phytoestrogens; Plant Extracts; Plant Preparations; Plants, Medicinal; Receptors, Estrogen; Recombinant Fusion Proteins; Saccharomyces cerevisiae; Seeds

Asian individuals have much lower incidences of prostate and breast cancer than populations from Western developed countries. They also consume a lower fat, higher fiber diet, with a large intake of phytoestrogens. These phytoestrogens may protect against hormone-dependent cancers and other diseases. Our study used established gas chromatography-mass spectrometry (GC-MS) methodologies to measure the concentrations of four phytoestrogens (daidzein, genistein, equol and enterolactone) in serum samples obtained from Japanese men (n = 102) and women (n = 125) > 40 y old. The results were compared with those obtained with samples from the UK. The Japanese men and women had higher (P < 0.001) concentrations of circulating daidzein, genistein and equol than individuals from the UK. The mean concentration of genistein in Japanese men, for example, was 492.7 nmol/L, compared with 33.2 nmol/L in men from the UK. The two populations, however, had similar serum concentrations of enterolactone. Furthermore, 58% of the Japanese men and 38% of the Japanese women had equol concentrations > 20 nmol/L, compared with none of the UK men and 2.2% of the UK women. These results support previously published GC-MS results from studies with low numbers of samples.

Topics: 4-Butyrolactone; Adult; Aged; Aged, 80 and over; Breast Neoplasms; Chromans; Diet; Equol; Estrogens, Non-Steroidal; Female; Gas Chromatography-Mass Spectrometry; Genistein; Humans; Isoflavones; Japan; Lignans; Male; Middle Aged; Phytoestrogens; Plant Preparations; Prostatic Neoplasms; United Kingdom

Flavonoids, such as daidzein and genistein, present in dietary plants like soybean, have unique chemical properties with biological activity relevant to cancer. Many flavonoids and polyphenols, including resveratrol in red wine and epigallocatechin gallate in green tea, are known antioxidants. Some of these compounds have estrogenic (and antiestrogenic) activity and are commonly referred to as phytoestrogens. A yeast-based estrogen receptor (ER) reporter assay has been used to measure the ability of flavonoids to bind to ER and activate estrogen responsive genes. Recently, estrogenic compounds were also shown to trigger rapid, nongenomic effects. The molecular mechanisms, however, have not been completely detailed and little information exists regarding their relevance to cancer progression. As a preliminary step toward elucidating rapid phytoestrogen action on breast cancer cells, we investigated the effect of 17-beta estradiol (E2), genistein, daidzein and resveratrol on the activation status of signaling proteins that regulate cell survival and invasion, the cell properties underlying breast cancer progression. The effect of these estrogenic compounds on the activation, via phosphorylation, of Akt/protein kinase B (Akt) and focal adhesion kinase (FAK) were analyzed in ER-positive and -negative human breast cancer cell lines. E2, genistein and daidzein increased whereas resveratrol decreased both Akt and FAK phosphorylation in nonmetastatic ER-positive T47D cells. In metastatic ER-negative MDA-MB-231 cells, all estrogenic compounds tested increased Akt and FAK phosphorylation. The inhibitory action of resveratrol on cell survival and proliferation is ER dependent. Therefore, all estrogenic compounds tested, including resveratrol, may exert supplementary ER-independent nongenomic effects on cell survival and migration in breast cancer cells.

Topics: Breast Neoplasms; Cell Division; Cell Survival; Enzyme Activation; Estradiol; Estrogen Receptor alpha; Estrogen Receptor beta; Estrogens, Non-Steroidal; Flavonoids; Focal Adhesion Kinase 1; Focal Adhesion Protein-Tyrosine Kinases; Genistein; Humans; Isoflavones; Phosphorylation; Phytoestrogens; Plant Preparations; Protein Serine-Threonine Kinases; Protein-Tyrosine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Receptors, Estrogen; Resveratrol; Stilbenes; Tumor Cells, Cultured

Topics: Anticarcinogenic Agents; Cohort Studies; Diet Records; Diet Surveys; England; Estrogens, Non-Steroidal; Female; Genistein; Glycine max; Humans; Isoflavones; Male; Middle Aged; Phytoestrogens; Plant Preparations; Prospective Studies; Sex Factors

Many rodent diets contain components such as soy isoflavones (daidzein and genistein) known to have estrogenic properties. The dietary background of phytoestrogens may modulate some responses to environmental estrogens when these compounds are tested in rodent bioassays. Thus, and since only few data were available on the phytoestrogen content of rodent diets commonly used in European laboratories, it was of interest to analyze the daidzein and genistein contents of our standard animal feeds. Isoflavone contents were determined in seven batches of rodent chow (from two suppliers in Germany, Altromin and Ssniff) by high-performance liquid chromatography, and also analyzed in six rodent diets from the United States. The soy-based rodent diets from Germany contained isoflavone (daidzein plus genistein) concentrations in the range of 0.3-0.55 mg/g feed. These isoflavone contents are similar to those analyzed in the US rodent diets, and similar to values reported by others, including one particular lot of feed (with 0.35 mg isoflavones per g) which produced a large uterotrophic response in immature ovariectomized rats [Environ. Health Perspect., 106 (1998) 369]. Coumestrol was found in a sample of commercial rabbit food at rather high levels (0.27 mg/g), but, this phytoestrogen was not detected (<1 microg/g feed) in any of the other samples we analyzed. The soy components in our rodent diet produce a measurable background of daidzein and genistein in blood of female DA/Han rats, total isoflavones (aglycone plus conjugates) ranging between 90 and 290 ng/ml plasma. The ovariectomized animals kept on this chow, showed no signs of estrogenization of the reproductive tract (uterus, vagina), and responded normally to (xeno-)estrogen administration in a uterotrophic assay [J. Steroid Biochem. Mol. Biol., 73 (2000) 1]. Moreover, ovariectomized Wistar rats on our standard rodent diet (Ssniff R/M H) had lower uterine weights than animals kept on the isoflavone-free (solvent extracted) chow; both groups of rats responded to genistein administration with an increase in uterine weights. These results suggest that--albeit the sensitivity of the rodent uterotrophic assay is not reduced by the use of a diet containing soy isoflavones at commonly encountered levels--attention should be given to a variable dietary phytoestrogen background.

Topics: Animal Feed; Animals; Estrogens, Non-Steroidal; Female; Genistein; Growth Inhibitors; Isoflavones; Organ Size; Ovariectomy; Phytoestrogens; Plant Preparations; Rats; Rats, Wistar; Statistics, Nonparametric; Uterus

There is widespread concern that fetal exposure to hormonally active chemicals may adversely affect development of the reproductive tract. Therefore, the present study was performed to develop the necessary analytical methods and test the hypothesis that dietary phytoestrogens can be quantified in second trimester human amniotic fluid. Amniotic fluid samples (n=59) from women (n=53) undergoing routine amniocentesis between 15 and 23 weeks of gestation were analyzed by gas chromatography/mass spectrometric (GC/MS). Analytes included the phytoestrogens daidzein, genistein, formononetin, biochanin A, and coumestrol. Dietary phytoestrogens were quantified in 96.2% of second trimester amniotic fluid samples tested. The mean (+/- standard deviation (S.D.)) concentration of daidzein and genistein in amniotic fluid was 1.44 +/- 1.34 and 1.69 +/- 1.48 ng/ml with maximum levels of 5.52 and 6.54 ng/ml, respectively. Second trimester amniotic fluid contains quantifiable levels of dietary phytoestrogens and thus is a marker of mid pregnancy fetal exposure.

Topics: Adult; Amniocentesis; Amniotic Fluid; Coumestrol; Diet; Estrogens, Non-Steroidal; Female; Gas Chromatography-Mass Spectrometry; Genistein; Humans; Isoflavones; Middle Aged; Phytoestrogens; Plant Preparations; Pregnancy; Pregnancy Trimester, Second

Oxidation of low-density lipoproteins (LDL) promotes the formation of atherosclerotic plaques. Estrogenic compounds (EC) from foods and other natural products, and synthetic estrogenic compounds (SECs) may prevent heart disease by inhibiting LDL oxidation. In the present study, we tested the antioxidant capacities of two phytoestrogens, daidzein (DAI) and genistein (GEN), and four SECs, (+)- and (-)-Z-bisdehydrodoisynolic acid (ZBDDA), and (+)- and (-)-hydroxy-allenoic acid (HAA), on isolated human LDL subjected to oxidation by cupric sulfate. The effects of these estrogenic compounds on the kinetics of conjugated diene formation in LDL undergoing oxidation were evaluated with a lag-time assay with continuous monitoring of absorbance at 234 nm. Lag-time data revealed that (+)-HAA, (-)-HAA, (+)-ZBDDA, and (-)-ZBDDA had similarly stronger antioxidant activities than either GEN or DAI. We also found that (+)-HAA, (-)-HAA, (+)-ZBDDA, and (-)-ZBDDA strongly inhibited the formation of Cu+-induced thiobarbituric acid reactive substances (TBARS) in LDL, and that GEN and DAI were less effective for inhibiting LDL lipid peroxidation. Finally, electrophoretic evaluation suggested that (+)-HAA, (-)-HAA, (+)-ZBDDA, and (-)-ZBDDA protected the apolipoprotein B-100 of LDL against oxidation better than did GEN or DAI. In summary, the four SECs, (+)-HAA, (-)-HAA, (+)-ZBDDA, and (-)-ZBDDA, were more potent antioxidants than the phytoestrogens, DAI and GEN.

Topics: Adult; Antioxidants; Copper; Estradiol Congeners; Estrogens, Non-Steroidal; Genistein; Humans; Isoflavones; Lipoproteins, LDL; Oxidation-Reduction; Phytoestrogens; Plant Preparations

Diets containing the soya-derived phytoestrogens, genistein and daidzein, decrease plasma cholesterol in humans and experimental animals. The mechanisms responsible for the hypocholesterolaemic effects of these isoflavones are unknown. The present study was conducted to determine if genistein and daidzein regulate hepatocyte cholesterol metabolism and apolipoprotein (apo) B secretion in cultured human hepatoma (HepG2) cells. ApoB secretion was decreased dose-dependently by up to 63% and 71% by genistein and daidzein (100 microM; P<0.0001) respectively. In contrast, no effect on apoAI secretion was observed. Cellular cholesterol synthesis was inhibited 41% by genistein (100 microM; P<0.005) and 18% by daidzein (100 microM; P<0.05), which was associated with significant increases in 3-hydroxy-3-methylglutaryl-CoA reductase mRNA. Cellular cholesterol esterification was decreased 56% by genistein (100 microM; P<0.04) and 29% by daidzein (100 microM; P<0.04); however, mRNA levels for acyl-CoA:cholesterol acyltransferase (ACAT) 1 and ACAT2 were unaffected. At 100 microM, both isoflavones equally inhibited the activities of both forms of ACAT in cells transfected with either ACAT1 or ACAT2. Genistein (100 microM) and daidzein (100 microM) significantly decreased the activity of microsomal triacylglycerol transfer protein (MTP) by 30% and 24% respectively, and significantly decreased MTP mRNA levels by 35% and 55%. Both isoflavones increased low-density lipoprotein (LDL)-receptor mRNA levels by 3- to 6-fold (100 microM; P<0.03) and significantly increased the binding, uptake and degradation of (125)I-labelled LDL, suggesting that enhanced reuptake of newly secreted apoB-containing lipoproteins contributed to the net decrease in apoB secretion. These results indicate that genistein and daidzein inhibit hepatocyte apoB secretion through several mechanisms, including inhibition of cholesterol synthesis and esterification, inhibition of MTP activity and expression and increased expression of the LDL-receptor.

Topics: Apolipoproteins B; Carcinoma, Hepatocellular; Cholesterol Esters; Dose-Response Relationship, Drug; Estrogens, Non-Steroidal; Genistein; Glycine max; Humans; Hydroxymethylglutaryl CoA Reductases; Isoflavones; Kinetics; Liver Neoplasms; Phytoestrogens; Plant Preparations; RNA, Messenger; Transcription, Genetic; Tumor Cells, Cultured

Hormonally active chemicals (HACs) that are capable of inducing adverse effects on wildlife as well as human beings are featured as "endocrine disruptors". Various animal studies conducted to clarify the characteristics of HACs, including the uterotrophic assay, are sufficiently sensitive to detect the effect of 17-beta-estradiol in micrograms per kilogram of body weight or lower. In such systems, a trace amount of HACs in the dietary pellets may interfere with the test results and thus can be a serious problem for the low-dose issue, which is now a major topic in the field of endocrine disruptor research. Here, the significance of the hormonal effects of phytoestrogen components in the NIH-07 diet is confirmed and a NIH-07-based open formula "phytoestrogen-low diet" (PLD) is proposed, which effectively reduces uterine weight as well as the uterine luminal epithelial labeling index in ovariectomized rats.

Topics: Animals; Diet; Estrogens, Non-Steroidal; Female; Genistein; Isoflavones; Organ Size; Ovariectomy; Phytoestrogens; Plant Preparations; Rats; Uterus

The objective of this study was to determine the effect of dietary phytoestrogens on the incidence of spontaneous vulvar carcinomas in 129/J mice using three natural ingredient diets and two purified diets containing predetermined levels of daidzein and genistein. Eighty weanling female mice without clinical evidence of vulvar carcinomas were randomly assigned 16 per diet to each of 5 test diets. Mice were clinically examined for vulvar masses weekly for 3 months and at monthly intervals thereafter. Vulvar carcinomas in representative groups of mice were confirmed using routine histological procedures. The incidence of vulvar carcinomas increased sharply in mice on all test diets during the first 2 months with minor changes during the remainder of the study. Within one month, the incidence of vulvar carcinomas in mice fed the AIN-76A modified soy protein diet was significantly (P < .05) increased over those of mice fed the AIN-76A modified casein diet, the #5K96, or the # 5058 diet. At three months, the incidence of vulvar carcinomas in mice fed the soy protein diet was significantly (P < .05) increased over those of mice fed the NIH-31 diet or the PMI #5K96 diet. There was a marginally significant (P < .10) correlation between the total daidzein and genistein levels in the five test diets and the incidence of vulvar carcinomas in mice as determined by clinical examination. We concluded that dietary levels of daidzein and genistein were associated with an increase in the incidence of vulvar carcinomas in mice and that the 129/J mouse may provide an animal model for studying the development of vulvar carcinomas.

Topics: Animals; Carcinoma, Squamous Cell; Diet; Estrogens, Non-Steroidal; Female; Genistein; Incidence; Isoflavones; Mice; Phytoestrogens; Plant Preparations; Vulvar Neoplasms

Five rodent diets have been evaluated for their possible effect on the sexual development of the rat. Groups of 12 pregnant Alpk rats were fed one of the following combinations of diets during pregnancy and postnatally: RM3/RM1, AIN-76A/AIN-76A, RM3/AIN-76A, Teklad Global 2016 (Global)/Global and Purina 5001/Purina 5001. AIN-76A is phytoestrogen-free while the other diets contained varying amounts of phytoestrogens. The phytoestrogens genistein and daidzein were determined in the diets studied, and the concentrations found agreed with earlier estimates. RM3/RM1 was selected as the control group, as this has been used routinely in this laboratory for the past decade. Determinations were made in offspring of the times of vaginal opening and first estrus among the females, and of prepuce separation and testes descent among the males. At postnatal day (PND) 26 the females from 6 of the 12 litters were terminated and tissue weights measured. Males from 6 of the 12 litters were similarly studied at PND 68. Animals from the remaining litters were transferred to RM1 diet at PND 70. Termination of the study was at PND 128 (males) and PND 140 (females) when body weights and tissue weights were determined. Marked differences in body weight, sexual development, and reproductive tissue weights were observed for rats maintained on AIN-76A or Purina 5001, with only minimal effects among rats maintained on the Global diet. These comparisons were against RM3/RM1 as the reference diet. However, using Purina 5001 as the reference diet reversed the direction of the differences seen when using RM3/RM1 as the reference diet. The differences observed when using RM3/RM1 as reference diet occurred mainly postnatally. In addition, the fact that similar differences were seen for the phytoestrogen-free diet, AIN-76A, and the phytoestrogen-rich diet, Purina 5001, indicate that these effects are more likely to be caused by nutritional differences between the diets that then have centrally mediated effects on rodent sexual development, rather than individual dietary components affecting peripheral estrogen receptors (ER). This proposal is supported by abolition of the uterotrophic activity of AIN-76A and Purina 5001 (relative to RM3/RM1) in the immature rat by coadministration of the gonadotrophin-releasing hormone (GnRH) antagonist ANTARELIX: The present data indicate that choice of diet may influence the timing of sexual development in the rat, and consequently, that when evaluating

Topics: Age Factors; Animal Feed; Animals; Animals, Newborn; Biological Assay; Body Weight; Diet; Estrogens, Non-Steroidal; Female; Genistein; Isoflavones; Male; Oligopeptides; Organ Size; Phytoestrogens; Plant Preparations; Pregnancy; Rats; Rats, Inbred Strains; Rats, Wistar; Sexual Maturation; Weaning

It is generally not known that most commercial rodent diets are formulated with soy protein and deliver large daily doses of isoflavones to animals throughout their lifespan, including the in utero period. Here, we demonstrate that isoflavones are bioavailable and show that commercial rodent diets universally used by animal facilities lead to very high steady-state serum isoflavone concentrations in adult rats (2613 +/- 873 ng/mL) and mice (2338 +/- 531 ng/mL), exceeding the animal's endogenous estrogen level by 30,000- to 60,000-fold. We demonstrate the maternal-fetal intrauterine transfer of isoflavones in animals fed a standard Purina 5001 soy-containing diet and show that newborn rat pups have high serum isoflavones levels (540 +/- 174 ng/mL) that are maintained throughout the suckling period by passage of isoflavones into maternal milk. These findings have profound implications for all animal experiments, including multigenerational studies and studies of transgenic animals, especially if biochemical or morphological end-points are influenced by the hormonal or nonhormonal properties of phytoestrogens. These compounds have the potential to modulate genotypic and phenotypic expression in general, and therefore, all investigators should be vigilant to the phytoestrogen composition of commercial rodent diets because there is a history of potent biological effects in larger animals and in humans from high circulating isoflavone concentrations.

Topics: Age Factors; Animal Feed; Animals; Animals, Newborn; Animals, Suckling; Chromans; Chromatography, High Pressure Liquid; Equol; Estrogens, Non-Steroidal; Female; Genistein; Hormones; Isoflavones; Mice; Models, Animal; Phytoestrogens; Plant Preparations; Rats

Phytoestrogens derived from soybeans reverse endothelial dysfunction in a number of animal models of systemic vascular disease. Based on these studies, we hypothesized that phytoestrogens would reverse chronic hypoxia-induced endothelial dysfunction in rat pulmonary arteries. To test this hypothesis we examined the effect of genistein, the major phytoestrogen found in soybeans, on carbachol-induced relaxation in phenylephrine-constricted pulmonary artery rings isolated from normoxic rats and rats exposed to 14 days of hypobaric hypoxia. Compared with that in normoxic rats, the response to carbachol was impaired in pulmonary arteries isolated from rats exposed to chronic hypoxia. In normoxic rat pulmonary arteries, genistein (30 microM) did not change the maximum relaxation to carbachol. In contrast, genistein significantly enhanced the relaxation response to carbachol in pulmonary arteries from hypoxic rats, restoring it to the levels seen in normoxic rats. 17beta-estradiol (10 microM) and daidzein (30 microM), a structural analog of genistein lacking inhibitory effects on tyrosine kinases, also restored the relaxation response to carbachol in hypoxic rat pulmonary arteries. The nitric-oxide synthase inhibitor N(omega)-nitro-L-arginine (100 microM) completely blocked the genistein, daidzein, and 17beta-estradiol-induced restoration of the relaxation response to carbachol, whereas the estrogen receptor antagonist ICI 182,780 (10 microM) had no effect on the relaxation responses. We conclude that the phytoestrogens genistein and daidzein act like estrogen in restoring nitric oxide-mediated relaxation in chronically hypoxic rat pulmonary arteries and that this effect does not appear to be mediated by inhibition of tyrosine kinases or by known estrogen receptors.

Topics: Animals; Carbachol; Cholinergic Agonists; Chronic Disease; Dose-Response Relationship, Drug; Estradiol; Estrogen Antagonists; Estrogens, Non-Steroidal; Fulvestrant; Genistein; Hypertension, Pulmonary; Hypoxia; In Vitro Techniques; Isoflavones; Male; Nitric Oxide; Nitric Oxide Synthase; Phytoestrogens; Plant Preparations; Pulmonary Artery; Rats; Rats, Sprague-Dawley; Vasoconstriction; Vasoconstrictor Agents; Vasodilation; Vasodilator Agents

The effect of phytoestrogen intake in combination with estrogen replacement therapy (ERT) on atherogenesis is largely unknown. The aim was thus to study the impact of phytoestrogens alone, or combined with oral estrogen, on experimental atherosclerosis in cholesterol-fed rabbits.. Two separate studies were performed in ovariectomized, cholesterol-fed female rabbits. In Study A, 45 rabbits were randomized to either a soy-free diet with or without oral 17 beta-estradiol (E2) 4 mg/day, or a soy-rich diet without any hormone for 14 weeks. In Study B, 100 rabbits were randomized into five groups (oral E2 0.5, 1, 2 or 4 mg/day, or no hormone) based on a soy-rich diet for 30 weeks.. By the end of treatment in Study A, aortic cholesterol content was twice the amount in the group treated with the soy-free diet compared with the soy-rich group and with the soy-free plus E2 group (p < 0.001). In Study B, aortic cholesterol content showed no significant difference between the groups (ANOVA, p = 0.49), but a tendency towards a lower aortic cholesterol content in the E2-treated animals compared with placebo was observed.. Dietary phytoestrogens significantly reduce aortic cholesterol content with a potency comparable to that of ERT, and seem to enhance (although mildly) the antiatherogenic effect of E2 in this model.

Topics: Animals; Aorta; Cholesterol; Chromatography, High Pressure Liquid; Coronary Artery Disease; Diet; Disease Models, Animal; Estradiol; Estrogens, Non-Steroidal; Female; Genistein; Glycine max; Isoflavones; Lipoproteins; Ovariectomy; Phytoestrogens; Plant Preparations; Rabbits; Random Allocation; Uterus

Numerous epidemiologic studies revealed that ethnic populations with higher dietary intake of phytoestrogens have the lowest incidence for breast cancer. The molecular mechanisms which may be responsible for this cancer protective action of phytoestrogens are so far only barely characterised. There are some hints that phytoestrogens may act like selective estrogen receptor modulators (SERMs) on the breast. For this reason we have investigated potential SERM-like properties of the phytoestrogens daidzein (Dai), coumestrol (Cou), and genistein (Gen) in the human breast cancer cell line MCF-7. Effects of these substances on progesterone (PR) and estrogen receptor alpha (ER) mRNA expression and estrogen receptor alpha protein levels were studied in comparison to estradiol (E2) and the synthetic SERMs raloxifene (Ral) and faslodex (ICI 182 780). PR mRNA expression was up-regulated after administration of Cou, whereas treatment with Dai and Gen induced only a faint increase. ER mRNA expression was down-regulated by Cou but not affected by Dai and Gen. The content of ER protein in the breast cancer cells was strongly decreased by Gen, only a faint reduction could be observed following administration of Cou, whereas administration of Dai slightly increases ER protein levels. In summary and in comparison to the effects observed after administration of E2, Ral, and ICI it turned out that Cou shows molecular properties which are very similar to an estrogen receptor agonist like E2, whereas the molecular properties of Gen are comparable to the SERMs ICI and Ral. These results clearly indicate that phytoestrogens differ significantly in regard to their molecular action on breast cancer cells and can be subdivided into distinct functional categories.

Topics: Blotting, Western; Breast; Coumestrol; Diet; Estradiol; Estrogen Antagonists; Estrogens, Non-Steroidal; Fulvestrant; Genistein; Humans; Isoflavones; Molecular Structure; Phytoestrogens; Plant Preparations; Raloxifene Hydrochloride; Receptors, Estrogen; Receptors, Progesterone; RNA, Messenger; Selective Estrogen Receptor Modulators; Tumor Cells, Cultured

The aim of this study was to assess the effect of phytoestrogens on the human vascular wall in vitro.. We compared the effects of E2 to those of genistein (G), daidzein (D), biochanin A (BA), equol (EQ), and quecertin (Qu) on 3[H] thymidine incorporation and creatine phosphokinase (CK) activity in human vascular smooth muscle cells (VSMC) and in a human endothelial cell line (E304).. In VSMC, E2, the estrogen antagonist raloxifene (RAL), G, and D stimulated DNA synthesis at low concentrations and suppressed 3[H] thymidine incorporation at higher concentrations. In contrast, BA and EQ had a monophasic stimulatory effect on 3[H] thymidine incorporation (87% +/- 9% and 54% +/- 17%, respectively) whereas Qu had only an inhibitory effect (-36 +/- 16% at 30 nmol/L). In E304 cells, all phytoestrogens stimulated DNA synthesis in a dose-related manner. In both cell types E2, RAL as well as all phytoestrogens increased CK-specific activity. The administration of phytoestrogens to immature female rats resulted in increased CK in the aorta (Ao) (60% to 220%) and in the left ventricle of the heart (Lv) (45% to 160%). Similar increases in Ao and Lv CK were also induced by E2 and all five phytoestrogens in ovariectomized (OVX) female rats. RAL antagonized phytoestrogen-induced CK activity in human vascular cells and in the rat Ao and Lv tissue but did not block phytoestrogen effects on DNA synthesis in human VSMC.. Although phytoestrogens have estrogen-mimetic effects on cell growth and CK in cultured human vascular cells and on CK in rat vascular tissues in vivo, the effects on human VSMC replication are highly dependent on the concentration and the particular phytoestrogen under investigation.

Topics: Animals; Aorta; Cells, Cultured; Chromans; Creatine Kinase; Creatine Kinase, MB Form; DNA; Endothelium, Vascular; Enzyme Activation; Enzyme Inhibitors; Equol; Estrogen Antagonists; Estrogens, Non-Steroidal; Female; Genistein; Humans; Isoenzymes; Isoflavones; Muscle, Smooth, Vascular; Phytoestrogens; Plant Preparations; Quercetin; Raloxifene Hydrochloride; Rats; Rats, Wistar; Thymidine; Tritium; Umbilical Arteries

Biological effects of dietary isoflavones, such as daidzein and genistein are of interest in preventive medicine. We estimated the dietary intake of isoflavones from dietary records and compared the values with the plasma concentrations and urinary excretions in Japanese middle-aged women. The dietary intake of daidzein and genistein was 64.6 and 111.6 mumol /day/capita (16.4 and 30.1 mg/day/capita), respectively. The isoflavones intake was mostly attributable to tofu, natto and miso. The median of plasma daidzein and genistein concentration was 72.46 and 206.09 nmol/L, respectively. The median of urinary excretion was 20.54 mumol /day for daidzein, 10.79 for genistein, 15.74 for equol and 1.64 for O-desmethylangolensin (O-DMA). Equol and O-DMA were excreted by 50% and 84% of all participants, respectively. Equol metabolizers were significantly lower the plasma and urinary daidzein and urinary O-DMA. The dietary intake of daidzein and genistein after the adjustment for total energy intake was significantly correlated with the urinary excretion (r = 0.365 for daidzein and r = 0.346 for genistein) and plasma concentration (r = 0.335 for daidzein and r = 0.429 for genistein). The plasma concentration of isoflavones was also significantly correlated with the urinary excretion. We conclude that in epidemiological studies measurements of plasma concentration or urinary excretion of these isoflavones are useful biomarkers of dietary intake and important for studies on their relation to human health.

Topics: Adult; Aged; Anticarcinogenic Agents; Biomarkers; Chromans; Diet; Diet Records; Energy Intake; Equol; Estrogens, Non-Steroidal; Female; Genistein; Glycine max; Humans; Isoflavones; Middle Aged; Monoamine Oxidase Inhibitors; Phytoestrogens; Plant Preparations; Plants; Regression Analysis

A time-resolved fluoroimmunoassay (TR-FIA), with europium labeled phytoestrogens as tracers, was developed for the quantitative determination of enterolactone, genistein and daidzein in human urine. The aim was to create a method for the screening of large populations in order to assess the possible correlations between the urinary levels and the risk of Western diseases. After the synthesis of the 5'-carboxymethoxy derivative of enterolactone and 4'-O-carboxymethyl derivatives of daidzein and genistein, the respective compound was coupled to bovine serum albumin and then used as an antigen in the immunization of rabbits. The same derivatives of the phytoestrogen were used in preparing the europium tracers. After the enzymatic hydrolysis, the TR-FIA was carried out using the Victor 1420 multilabel counter. The method has sufficient sensitivity to measure the phytoestrogens at concentrations even below 5 nmol/l. The intra- and inter-assay coefficients of variation, at three different concentrations, varied from 1.9 to 5.3 and from 2.4 to 9.7, respectively. We measured urinary enterolactone, genistein and daidzein in 215 samples from Finnish healthy women and found that more than 50% of the values ranged between 1 and 7, <0.1 and 0.6 and below 0.6 micromol/24 h, respectively. The TR-FIA method including only a hydrolysis step gave higher values than those measured by gas chromatography-mass spectrometry (GC-MS). However, the assay results by the present method showed strong correlation with those obtained by GC-MS. It is concluded that the TR-FIA is suitable for population screening of urinary phytoestrogens.

Topics: 4-Butyrolactone; Animals; Cross Reactions; Estrogens, Non-Steroidal; Evaluation Studies as Topic; Female; Fluoroimmunoassay; Gas Chromatography-Mass Spectrometry; Genistein; Humans; Isoflavones; Lignans; Phytoestrogens; Plant Preparations; Rabbits; Sensitivity and Specificity; Serum Albumin, Bovine

The function of the uterus is regulated by female sex steroids and it is, therefore, used as the classical target organ to detect estrogenic action. Uterine response to estrogens involves the activation of a large pattern of estrogen-sensitive genes. This fact offers the opportunity to analyze the estrogenic activity of xeno- and phytoestrogens, and the mechanisms of their molecular action by a correlation of the uterotropic activity and their ability to modulate the expression of estrogen-sensitive genes. We have analyzed the expression of androgen receptor (AR), progesterone receptor (PR), estrogen receptor (ER), clusterin (CLU), complement C3 (C3), and GAPDH mRNA in the rat uterus following oral administration of ethinylestradiol (EE), bisphenol A (BPA), o,p'-DDT (DDT), p-tert-octylphenol (OCT) and daidzein (DAI). A significant stimulation of the uterine wet weight could be observed after administration of all the substances. The activity of all analyzed compounds to stimulate uterine weight was low in comparison to EE. DDT has the highest activity to stimulate uterine weight whereas BPA and DAI turned out to be less potent. The analysis of gene expression revealed a very specific profile of molecular action in response to the different compounds which cannot be detected by judging the uterotropic response alone. A dose dependent analysis revealed that C3 mRNA is already modulated at doses where no uterotropic response was detectable. Although DAI and BPA were very weak stimulators of uterine growth, these substances were able to alter the expression of AR, ER and C3 very strongly. Based on these investigations the analyzed compounds can be subdivided into distinct classes: First, compounds which exhibit a similar gene expression fingerprint as EE (e.g. OCT); second, compounds exhibiting a significant uterotropic activity, but inducing a pattern of gene expression different from EE (e.g. DDT); and third, compounds like BPA and especially DAI which exhibit a very low uterotropic activity, but nevertheless modulate the expression of estrogen-sensitive genes. These findings strongly suggest that the fingerprint of uterine gene expression is a very sensitive tool to investigate estrogenicity of natural and synthetic compounds and offers the possibility to get information in regard to the molecular mechanisms involved in the action of the respective compounds.

Topics: Animals; Base Sequence; Benzhydryl Compounds; Complement C3; DDT; DNA Primers; Estrogens; Estrogens, Non-Steroidal; Ethinyl Estradiol; Female; Gene Expression; Isoflavones; Organ Size; Phenols; Phytoestrogens; Plant Preparations; Polymerase Chain Reaction; Rats; Receptors, Androgen; Receptors, Estrogen; RNA, Messenger; Uterus

Topics: Actins; Animals; Aorta, Abdominal; Endothelium, Vascular; Enzyme Inhibitors; Estradiol; Estrogens, Non-Steroidal; Female; Genistein; In Vitro Techniques; Isoflavones; Phytoestrogens; Plant Preparations; Rabbits; Receptors, Estrogen

We investigated the ability of genistein and daidzein, two soybean isoflavones, compared with that of 17 alpha-ethinylestradiol, to prevent bone loss in ovariectomized rats, a model for postmenopausal osteoporosis. Female Wistar rats (n = 65; 12 mo old) were either sham-operated (SH; n = 13) or ovariectomized (OVX; n = 52). On d 0, OVX rats were randomly assigned to groups as follows: 13 received genistein [G; 10 mcg/(g body weight. d)], 13 were treated with daidzein [D; 10 mcg/(g body weight. d)], 13 received 17 alpha-ethinylestradiol [E(2); 30 mcg/kg body weight. d)] and 13 were untreated (OVX). Compounds were mixed with a soy protein-free powdered semipurified diet and given orally for 3 mo. On d 90, the bone mineral density (BMD) in lumbar vertebrae, femur and its metaphyseal and diaphyseal zones (rich in cancellous and cortical bone, respectively) was lower in OVX than in SH (P < 0.01). In D or E(2), the four BMD were not different from SH, whereas in G, only the diaphyseal BMD was not different from SH. Image analysis performed in the distal femur metaphysis revealed that the cancellous bone area was lower in OVX than in SH (P < 0.01). Only the area in D was not different from that in SH. Finally, the bone turnover, which was higher in OVX than in SH (P < 0.005 and P < 0.05 for plasma osteocalcin concentration and urinary deoxypyridinoline excretion, respectively), was not different in G, D or E(2) compared with SH. Therefore, consumption of 17 alpha-ethinylestradiol or daidzein was more efficient than genistein in preventing ovariectomy-induced bone loss in rats.

Topics: Animals; Body Weight; Bone Density; Equidae; Estrogens, Non-Steroidal; Female; Genistein; Goats; Humans; Isoflavones; Osteocalcin; Osteoporosis, Postmenopausal; Ovariectomy; Phytoestrogens; Plant Preparations; Radioimmunoassay; Rats; Rats, Wistar

Increasing evidence indicates that oxidative modification of low-density lipoprotein (LDL) is an important determinant in atherogenesis, and following menopause, the incidence of coronary heart disease is as prevalent in women as it is in men. Estrogen has been demonstrated to inhibit the susceptibility of LDL to be oxidized, and more recently the use of phytoestrogens has been considered for estrogen replacement therapy. In this study the antioxidant activity of the three major phytoestrogens: genistein, daidzein, and equol were measured in terms of LDL oxidative susceptibility. Increasing levels of genistein, daidzein, and equol inhibited LDL oxidation, and this inhibitory effect was further enhanced in the presence of ascorbic acid. The synergism exhibited by these compounds is of clinical importance to phytoestrogen therapy since the efficacy of phytoestrogens as effective antioxidants is evident at concentration well within the range found in the plasma of subjects consuming soy products. However, this synergism, combined with the low reactivity of the phytoestrogens with peroxyl radicals, suggests that an antioxidant mechanism other then free radical scavenging reactions account for the phytoestrogen antioxidant effect. A structural basis for inhibition of LDL oxidation involving interaction of the phytoestrogens with apoB-100 is postulated.

Topics: Adult; Antioxidants; Ascorbic Acid; Chromans; Drug Synergism; Equol; Estrogens, Non-Steroidal; Female; Genistein; Humans; Isoflavones; Kinetics; Lipid Peroxidation; Lipid Peroxides; Lipoproteins, LDL; Male; Oxidation-Reduction; Phytoestrogens; Plant Preparations

An in vitro model, designated the Simulator of the Human Intestinal Microbial Ecosystem (SHIME), was used to study the effect of a soygerm powder rich in beta-glycosidic phytoestrogenic isoflavones on the fermentation pattern of the colon microbiota and to determine to what extent the latter metabolize the conjugated phytoestrogens. Initially, an inoculum prepared from human feces was introduced into the reactor vessels and stabilized over 3 wk using a culture medium. This stabilization period was followed by a 2-wk control period during which the microbiota were monitored. The microbiota were then subjected to a 2-wk treatment period by adding 2.5 g/d soygerm powder to the culture medium. The addition resulted into an overall increase of bacterial marker populations (Enterobacteriaceae:, coliforms, Lactobacillus: sp., Staphylococcus: sp. and Clostridium: sp.), with a significant increase of the Lactobacillus: sp. population. The short-chain fatty acid (SCFA) concentration increased approximately 30% during the supplementation period; this was due mainly to a significant increase of acetic and propionic acids. Gas analysis revealed that the methane concentration increased significantly. Ammonium and sulfide concentrations were not influenced by soygerm supplementation. Use of an electronic nose apparatus indicated that odor concentrations decreased significantly during the treatment period. The beta-glycosidic bonds of the phytoestrogenic isoflavones were cleaved under the conditions prevailing in the large intestine. The increased bacterial fermentation after addition of the soygerm powder was paralleled by substantial metabolism of the free isoflavones (genistein, daidzein and glycitein), resulting in recovery of only 12-17% of the supplemented isoflavones.

Topics: Bioreactors; Clostridium; Colon; Ecosystem; Electric Impedance; Enterobacteriaceae; Escherichia coli; Estrogens, Non-Steroidal; Feces; Fermentation; Glycine max; Humans; Intestines; Isoflavones; Lactobacillus; Models, Biological; Phytoestrogens; Plant Preparations; Staphylococcus

Animal experiments are widely accepted in arterosclerosis research. Estrogens have lipid lowering properties and beneficial effects on the vasomotion. They act antiproliferative on those cells of the vascular wall which play a major role in lumen narrowing after vascular injury and in atherogenesis. The aim of the present study was to establish an organ culture model (rabbit aorta) to investigate post injury estrogen effects in the vessel wall. We chose the rabbit abdominal aorta which is the target organ in various animal experiments on this matter. The endothelial mono-layer was manipulated in a way that caused a measurable and reproducible post injury reaction (neointima formation). Then the effect of different estrogens (17beta-Estradiol, the phytoestrogens Genistein and Daidzein) on neointima development was investigated in male and female rabbit aortae. In equivalent dosages of 50 microg/ml all three estrogens inhibited the neointima formation significantly in male and female vessels. By the use of this organ culture model it was possible to show post injury effects of different estrogens in the vasculature while the consumption of animals was significantly reduced. Because 10 aortic segments could be taken from one aortic vessel, the number of animals that would have been necessary for an in vivo experiment could be reduced by the factor 10.

Topics: Animal Testing Alternatives; Animals; Aorta, Abdominal; Estradiol; Estrogens, Non-Steroidal; Female; Genistein; Isoflavones; Male; Organ Culture Techniques; Phytoestrogens; Plant Preparations; Rabbits; Tunica Intima

A recent study demonstrated that reactive oxygen species (ROS) were involved in the maintenance of hypertension in stroke-prone spontaneously hypertensive rats (SHRSP). However, the role of oxidative stress in hypertension and its related diseases in SHRSP remains unknown. To determine whether phytoestrogens attenuate oxidative DNA damage in vascular smooth muscle cells (VSMC) from SHRSP and Wistar-Kyoto (WKY) rats, we investigated the effect of daidzein, genistein and resveratrol on oxidative DNA damage in VSMC, induced by advanced glycation end-products (AGEs).. VSMC were treated with AGEs in the presence or absence of phytoestrogens for the indicated time. Cellular degeneration induced by AGEs was characterized in terms of intracellular oxidant levels, intracellular total glutathione (GSH) levels, mRNA expression for gamma-glutamylcysteine synthetase (GCS), and a new marker of oxidative stress, 8-hydroxy-2'-deoxyguanosine (8-OHdG) contents.. AGEs stimulated 8-OHdG formation in VSMC in a time- and dose-dependent manner. We also confirmed that VSMC from SHRSP were more vulnerable to oxidative stress induced by AGEs, than VSMC from WKY rats. Daidzein, genistein or resveratrol reduced AGEs-induced 8-OHdG formation in a dose-dependent manner. The preventive effects of phytoestrogens on 8-OHdG formation remarkably paralleled changes in the intracellular oxidant levels in VSMC following AGEs treatment. We further demonstrated that phytoestrogens increase intracellular total GSH level in VSMC. Increased GSH synthesis was due to enhanced expression of the rate-limiting enzyme for GSH synthesis, GCS. Phytoestrogens-stimulated total GSH level in VSMC could lead to decreased intracellular oxidant levels, and thus prevent oxidative DNA damage, induced by AGEs. The phytoestrogens are powerful antioxidants able to interfere with AGEs-mediated oxidative DNA damage of VSMC, and are potentially useful against vascular diseases where ROS are involved in hypertension.

Topics: Animals; Base Sequence; Cells, Cultured; DNA Damage; DNA Primers; Estrogens, Non-Steroidal; Genistein; Glutathione; Glutathione Peroxidase; Glycation End Products, Advanced; Hypertension; Isoflavones; Muscle, Smooth, Vascular; Oxidation-Reduction; Oxidative Stress; Phytoestrogens; Plant Preparations; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Reactive Oxygen Species; Resveratrol; RNA, Messenger; Stilbenes; Stroke

The primary objective of this study was to assess the accuracy of a modified Block food frequency questionnaire (FFQ) with respect to its ability to assess usual dietary intakes of daidzein and genistein. Participants were a convenience sample of 51 Japanese and 18 Caucasian women. All interviews were conducted between February 1997 and October 1997. At each of the four study visits, participants provided a 24-hour urine specimen and a 48-hour dietary recall. At the first visit, participants also completed an interviewer-administered modified Block FFQ. The daidzein and genistein intakes estimated using the FFQ were moderately correlated with the mean estimates of daidzein and genistein intake calculated from four 48-hour dietary recalls (correlation for daidzein = 0.49-0.58 and correlation for genistein = 0.45-0.54) and estimates of urinary concentrations of these compounds calculated from four collections (correlations for daidzein and genistein = 0.49 and 0.30, respectively). The accuracy of the modified Block FFQ for assessment of usual daidzein and genistein intakes is supported by this study. These results support the use of this instrument in epidemiological studies as an easy and low-cost method to assess the usual dietary daidzein or genistein intake.

Topics: Anticarcinogenic Agents; Asian; Diet Records; Estrogens, Non-Steroidal; Female; Genistein; Humans; Interviews as Topic; Isoflavones; Mental Recall; Phytoestrogens; Plant Preparations; Reproducibility of Results; Sensitivity and Specificity; Surveys and Questionnaires; Urinalysis; White People

Time-resolved fluoroimmunoassays (TR-FIA), with europium labeled phytoestrogens as tracers, were developed for the quantitative measurement of genistein, daidzein and enterolactone in plasma and urine for the purpose of screening large populations and studies on possible correlation between the values in biological fluids and the risk of western diseases. The mean values of the three phytoestrogens in plasma as determined by TR-FIA were similar to those obtained by gas chromatography-mass spectrometry (GC-MS). The urinary excretion levels of total individual phytoestrogens were higher than those obtained by GC-MS, with the exception of the daidzein values. However, comparing the assay results obtained by the present method and those obtained by GC-MS, a strong correlation was evident (r = 0.87 - 0.99, p < 0.001). We measured plasma levels of genistein, daidzein and enterolactone in 111 healthy Japanese women The mean and median levels of genistein were 406.8 and 306.3 nmol/l, respectively, and those of daidzein were 118.4 and 76.8 nmol/l, respectively. These levels are higher than those reported for Americans and Western Europeans. Isoflavone intake as calculated from dietary records (genistein: mean, 86.5 mircomol/day and daidzein: mean, 57.4 micromol/day) was correlated with the plasma concentrations observed (genistein: r = 0.287, p < 0.01 and daidzein: r = 0.313, p < 0.01). Plasma enterolactone levels were low in Japanese women (mean, about 10 nmol/l). The levels of urinary excretions of genistein, daidzein were also measured and it was found that, in the majority, the levels ranged between 5-25 and 5-50 micromol/24 h, respectively. In contrast, healthy Finnish women showed very low values of isoflavones (below 10 nmol/l in plasma (n = 87) and below 0.6 micromol/24 h in urine (n = 126) for both compounds) and high levels of enterolactone in both plasma and urine (plasma: mean, 25 nmol/l and urine: majority range, 1-7 micromol/24 h).

Topics: 4-Butyrolactone; Adult; Aged; Diet; Estrogens, Non-Steroidal; Europium; Female; Finland; Fluoroimmunoassay; Gas Chromatography-Mass Spectrometry; Genistein; Humans; Isoflavones; Japan; Lignans; Middle Aged; Phytoestrogens; Plant Preparations; Reference Values; Sensitivity and Specificity

Prostate cancer is one of the most common male cancers in Western countries, yet the incidence of this fatal disease remains low in Asian populations. Environmental factors such as diet play an important role in hormone-dependent cancer etiology, and a high phytoestrogen intake may be one factor contributing to the low prostate cancer mortality in Eastern populations. In this study, we investigated the effects of the phytoestrogens genistein, daidzein, coumestrol, and equol on cell growth and DNA damage (strand breakage) in two human prostate tumor cell lines: androgen receptor-positive LNCaP and androgen receptor-negative PC-3. Each compound caused growth inhibition at physiologically relevant concentrations (<10 microM). Genistein induced DNA damage in both cell lines at <10 microM. Daidzein inhibited cell growth at 10-100 microM yet had no effect on DNA damage at up to 500 microM. Thus, despite their structural similarities, different phytoestrogens inhibit prostate tumor cell growth by independent mechanisms.

Topics: Antineoplastic Agents, Phytogenic; Cell Division; Cell Transformation, Neoplastic; Chromans; Coumestrol; DNA Damage; Equol; Estrogens, Non-Steroidal; Genistein; Humans; Isoflavones; Male; Phytoestrogens; Plant Preparations; Prostatic Neoplasms; Tumor Cells, Cultured

-Estrogens are known to induce cardioprotective effects by inhibiting smooth muscle cell (SMC) growth and neointima formation. However, the use of estrogens as cardioprotective agents is limited by carcinogenic effects in women and feminizing effects in men. If noncarcinogenic and nonfeminizing estrogenlike compounds, such as natural phytoestrogens, afford cardioprotection, this would provide a safe method for prevention of cardiovascular disease in both men and women. Therefore, we evaluated and compared in human aortic SMCs the effects of phytoestrogens (formononetin, genistein, biochanin A, daidzein, and equol) on 2.5% fetal calf serum-induced proliferation (3H-thymidine incorporation and cell number), collagen synthesis (3H-proline incorporation), and total protein synthesis (3H-leucine incorporation) and on PDGF-BB (25 ng/mL)-induced migration (modified Boydens chambers). Moreover, the effects of phytoestrogens on PDGF-BB (25 ng/mL)-induced mitogen-activated protein kinase (MAP kinase) activity in SMCs was also studied. Phytoestrogens inhibited proliferation, collagen and total protein synthesis, migration, and MAP kinase activity in a concentration-dependent manner and in the following order of potency: biochanin A>genistein>equol>daidzein>formononetin. In conclusion, our studies provide the first evidence that in human aortic SMCs phytoestrogens inhibit mitogen-induced proliferation, migration and extracellular matrix synthesis and inhibit/downregulate MAP kinase activity. Thus, phytoestrogens may confer protective effects on the cardiovascular system by inhibiting vascular remodeling and neointima formation and may be clinically useful as a safer substitute for feminizing estrogens in preventing cardiovascular disease in both women and men.

Topics: Adult; Animals; Aorta, Thoracic; Calcium-Calmodulin-Dependent Protein Kinases; Cattle; Cell Division; Cell Movement; Cells, Cultured; Chromans; Culture Media; DNA; Dose-Response Relationship, Drug; Equol; Estrogens, Non-Steroidal; Female; Genistein; Humans; Isoflavones; Kinetics; Male; Muscle, Smooth, Vascular; Phytoestrogens; Plant Preparations; Protein Biosynthesis

In previous investigations it was shown that the synthetic estrogen diethylstilbestrol (DES) induces a rise of the intracellular calcium level ([Ca2+]i) in C6 rat glioma cells [P. Tas, H. Stopper, K. Koschel, D. Schiffmann, Influence of the carcinogenic oestrogen diethylstilboestrol on the intracellular calcium level in C6 rat glioma cells. Toxic. In vitro 5 (1991) 463-465] which is accompanied by changes of the arrangement of the cytoskeleton. In the present study, we compared the induction of these effects in COS (monkey kidney cells) lacking estrogen receptors (ER) with those in RUCA-I (rat endometrial carcinoma) cells containing ER. The [Ca2+]i in RUCA-I and COS cells following 17beta-estradiol (ES), genistein (GEN), daidzein (DZ) and coumestrol (CES) treatment was analyzed. A significant increase of [Ca2+]i induced by all compounds was observed in RUCA-I cells. No effects were detected in COS cells after ES and GEN treatment. The anti-estrogen ICI 182780 completely blocked the ES-and GEN-induced rise of [Ca2+]i. Dose and time dependencies of changes of calcium levels were analyzed and a biphasic response could be observed. The actin staining showed disintegrated stress fibers in RUCA-I cells. The degree of the observed effects correlates with the known estrogenicity of the applied compounds (DES > ES > GEN). It remains to be elucidated whether or not the effects observed are mediated by the "classic" genomic estrogen receptor pathway or by alternate nongenomic or receptor-independent pathways.

Topics: Actins; Animals; Calcium; Chlorocebus aethiops; COS Cells; Coumestrol; Estradiol; Estrogen Antagonists; Estrogens, Non-Steroidal; Female; Fulvestrant; Genistein; Intracellular Fluid; Isoflavones; Phytoestrogens; Plant Preparations; Rats; Receptors, Estrogen; Tumor Cells, Cultured

We have recently reported that dietary intake of soybean isoflavone phytoestrogens resulted in increased oxidation resistance of isolated low density lipoprotein (LDL). In order to explore the underlying mechanisms we designed two types of in vitro experiments. First, we prepared several different isoflavone fatty acid esters to increase their lipid solubility and studied their incorporation into LDL. Second, the oxidation resistance of the isoflavone-containing LDLs was investigated with Esterbauer's 'conjugated diene' method using Cu2+ as prooxidant. Unesterified daidzein and genistein as well as genistein stearic acid esters were incorporated into LDL to a relatively small extent (0.33 molecules per LDL particle, or less) and they did not significantly influence oxidation resistance. The oleic acid esters of isoflavones were incorporated more effectively, reaching a level of 2.19 molecules per LDL particle or more, and the 4',7-O-dioleates of daidzein and genistein exhibited prolongations of lag times by 46% (P<0.05) and 202% (P<0.01), respectively. A smaller but significant increase in lag time (20.5%, P<0.01) was caused by daidzein 7-mono-oleate. In summary, esterification of soybean isoflavones daidzein and genistein with fatty acids at different hydroxyl groups provided lipophilicity needed for incorporation into LDL. Some isoflavone oleic acid esters increased oxidation resistance of LDL following their incorporation.

Topics: Antioxidants; Esters; Estrogens, Non-Steroidal; Fatty Acids; Genistein; Glycine max; Isoflavones; Lipoproteins, LDL; Molecular Structure; Oxidation-Reduction; Phytoestrogens; Plant Preparations

Phytoestrogens influence a variety of biological processes. As 17beta-estradiol alters adrenocortical cell function, we examined whether the dietary phytoestrogens, genistein and daidzein, have related effects. In cultured human fetal and postnatal adrenal cortical cells, genistein and daidzein (both 0.4-40 micromol/L) decreased ACTH-stimulated cortisol production to basal levels (ED50, 1-4 micromol/L). In the adult adrenocortical cell line, H295, genistein, daidzein, and 17beta-estradiol (10 micromol/L) decreased cAMP-stimulated cortisol synthesis in a similar fashion. Neither genistein nor daidzein altered basal or ACTH-stimulated dehydroepiandosterone sulfate (DHEA-S) production in fetal adrenocortical cells, whereas in postnatal adrenocortical cells, DHEA and DHEA-S were markedly increased (ED50, 1-4 micromol/L). In H295 cells, basal and cAMP-stimulated DHEA production were similarly increased by the phytoestrogens and 17beta-estradiol. Genistein and daidzein did not affect the expression of steroid-metabolizing enzymes. However, genistein and daidzein specifically inhibited the activity of 21-hydroxylase (P450c21); the activities of other steroidogenic enzymes were not affected. Thus, phytoestrogens may decrease cortisol synthesis by suppressing the activity of P450c21 and, as a consequence, increase DHEA/DHEA-S synthesis by shunting metabolites away from the glucocorticoid synthetic pathway. Therefore, consumption of foods containing phytoestrogens may alter adrenocortical function by decreasing cortisol and increasing androgen production.

Topics: Adrenal Cortex; Adrenocorticotropic Hormone; Adult; Androgens; Cell Line; Cells, Cultured; Cyclic AMP; Dehydroepiandrosterone; Dehydroepiandrosterone Sulfate; Enzyme Inhibitors; Estradiol; Estrogens, Non-Steroidal; Fetus; Genistein; Glucocorticoids; Humans; Hydrocortisone; Isoflavones; Phytoestrogens; Plant Preparations; Protein-Tyrosine Kinases; Steroid 21-Hydroxylase

1. The phytoestrogenic compound trans-resveratrol (trans-3,5, 4'-trihydroxystilbene) is found in appreciable quantities in grape skins and wine. It has been shown that both products rich in trans-resveratrol and pure trans-resveratrol inhibit platelet aggregation both in vivo and in vitro. However the mechanism of this action still remains unknown. 2. An essential component of the aggregation process in platelets is an increase in intracellular free Ca2+ ([Ca2+]i). Ca2+ must enter the cell from the external media through specific and tightly regulated Ca2+ channels in the plasma membrane. The objective of this study was to characterize what effect trans-resveratrol had on the Ca2+ channels in thrombin stimulated platelets. 3. In this study we showed that trans-resveratrol immediately inhibited Ca2+ influx in thrombin-stimulated platelets with an IC50 of 0.5 microM. trans-Resveratrol at 0.1, 1.0 and 10.0 microM produced 20+/-6, 37+/-6 and 57+/-4% inhibition respectively of the effect of thrombin (0.01 u ml(-1)) to increase [Ca2+]i. 4. trans-Resveratrol also inhibited spontaneous Ba2+ entry into Fura-2 loaded platelets, with 0.1, 1.0 and 10.0 microM trans-resveratrol producing 10+/-5, 30+/-5 and 50+/-7% inhibition respectively. This indicated that trans-resveratrol directly inhibited Ca2+ channel activity in the platelets in the absence of agonist stimulation. 5. trans-Resveratrol also inhibited thapsigargin-mediated Ca2+ influx into platelets. This suggests that the store-operated Ca2+ channels are one of the possible targets of trans-resveratrol. These channels rely on the emptying of the internal Ca2+ stores to initiate influx of Ca2+ into the cell. 6. The phytoestrogens genistein, daidzein, apigenin and genistein-glucoside (genistin) produced inhibitory effects against thrombin similar to those seen with trans-resveratrol. 7. We conclude that trans-resveratrol is an inhibitor of store-operated Ca2+ channels in human platelets. This accounts for the ability of trans-resveratrol to inhibit platelet aggregation induced by thrombin.

Topics: Adult; Barium; Blood Platelets; Calcium; Calcium Channel Blockers; Calcium Channels; Egtazic Acid; Estrogens, Non-Steroidal; Genistein; Humans; Inhibitory Concentration 50; Isoflavones; Phytoestrogens; Plant Preparations; Platelet Aggregation Inhibitors; Resveratrol; Stilbenes; Thapsigargin; Thrombin; Time Factors

Phytoestrogens exert estrogenic effects on the central nervous system, induce estrus, and stimulate growth of the genital tract of female animals. Over 300 plants and plant products, including some used in laboratory animal diets, contain phytoestrogens. Therefore, the source and concentration of phytoestrogens in rodent diets were determined.. Twelve rodent diets and six major dietary ingredients were assayed for phytoestrogens (daidzein, genistein, formononetin, biochanin A, and coumestrol), using high-performance liquid chromatography. Three rodent diets recently formulated to reduce phytoestrogen content also were assayed.. Formononetin, biochanin A, and coumestrol were not detected. Soybean meal was the major source of daidzein and genistein; their concentrations were directly correlated to the percentage of soybean meal in each diet.. High, variable concentrations of daidzein and genistein are present in some rodent diets, and dietary phytoestrogens have the potential to alter results of studies of estrogenicity. Careful attention should be given to diet phytoestrogen content, and their concentration should be reported. A standardized, open-formula diet in which estrogenic substances have been reduced to levels that do not alter results of studies that are influenced by exogenous estrogens is recommended.

Topics: Animal Feed; Animals; Animals, Laboratory; Coumestrol; Estrogens, Non-Steroidal; Female; Food, Formulated; Genistein; Glycine max; Isoflavones; Phytoestrogens; Plant Preparations; Rodentia

The oxidation of low density lipoproteins (LDLs) is thought to take place in the arterial intima when the particles have become isolated from circulating water-soluble antioxidants. We hypothesized that isoflavonoid antioxidants derived from soy could be incorporated into lipoproteins and possibly could protect them against oxidation, which is regarded as atherogenic. Six healthy volunteers received 3 soy bars [containing the isoflavonoid antioxidants genistein (12 mg) and daidzein (7 mg)] daily for 2 weeks. LDLs were isolated from blood drawn at the the end of a 2-week dietary baseline period, after 2 weeks on soy, and after discontinuation of soy. Large increases in plasma isoflavonoid levels occurred during soy feeding, but only minute amounts were stably associated with lipoproteins (less than 1% of plasma isoflavonoids in the LDL fraction). The LDLs were subjected to copper-mediated oxidation in vitro. Compared with off soy values, lag phases of LDL oxidation curves were prolonged by a mean of 20 min (P < 0.02) during soy intake, indicating a reduced susceptibility to oxidation. The results suggest that intake of soy-derived antioxidants, such as genistein and daidzein, may provide protection against oxidative modification of LDL. As only very small amounts of these substances were detected in purified LDL, modified LDL particles may have been produced in vivo by circulating isoflavonoids promoting resistance to oxidation ex vivo.

Topics: Adult; Antioxidants; Arteriosclerosis; Dietary Supplements; Estrogens, Non-Steroidal; Female; Genistein; Glycine max; Humans; Isoflavones; Lipoproteins, LDL; Male; Oxidation-Reduction; Phytoestrogens; Plant Preparations

We report a novel method for the measurement of urinary daidzein that is suitable for assessment of dietary soya exposure. The method incorporates the following features: (i) a highly specific monoclonal antibody to daidzein (clone 4E4) raised through the 7 position of daidzein and (ii) a europium labeled ovalbumin daidzein conjugate. In the present format, dilute urine samples of subjects who ingested soy milk are hydrolyzed with beta-glucuronidase for 30 min on rabbit anti-mouse coated plates. Afterwards, the specific monoclonal antibody to daidzein, clone 4E4, and europium labeled ovalbumin daidzein conjugate are added. After 1 h incubation, the wall bound fluorescence of europium is measured by time resolved fluorescence and is inversely proportional to the concentration of daidzein over the range 0.1-10 ng daidzein/well. The method demonstrates good sensitivity, precision and comparability with the chemical method GC-FID. Unlike the chemical method, the present immunoassay technique for daidzein is applicable for the measurement of large amounts of samples in epidemiological studies for the assessment and monitoring of human exposure to soya food.

Topics: Antibodies, Monoclonal; Calibration; Diet; Estrogen Antagonists; Estrogens, Non-Steroidal; Europium; Fluorescent Antibody Technique; Glucuronidase; Glycine max; Humans; Isoflavones; Ovalbumin; Phytoestrogens; Plant Preparations; Sensitivity and Specificity

Topics: Adenocarcinoma; BRCA1 Protein; Breast; Breast Neoplasms; Cells, Cultured; Epithelial Cells; Estrogens; Estrogens, Non-Steroidal; Female; Gene Expression Regulation; Genes, BRCA1; Genistein; Humans; Isoflavones; Neoplasm Proteins; Neoplasms, Hormone-Dependent; Phytoestrogens; Plant Preparations; Receptors, Estrogen; Signal Transduction; Tumor Cells, Cultured

Phytoestrogens can exhibit agonistic actions on estrogen-dependent gene expression in breast cancer cells. Since several different phytoestrogens may be found within a single dietary plant source, we sought to investigate whether estrogen-dependent gene expression may be further influenced by the collective treatment of breast cancer cells with multiple phytoestrogens. Accordingly, we transfected MCF-7 breast cancer cells with estrogen-responsive reporters followed by treatment with one of four phytoestrogens (genistein, daidzein, formononetin, and equol) or a combination of these in the absence of estradiol. Our results demonstrated clear-cut agonistic effects of phytoestrogens on estrogen-dependent gene expression. Moreover, combinatorial treatment consistently stimulated reporter activity above that observed for individual phytoestrogens. Inasmuch as the phytoestrogens tested are frequently found together in food sources, these combinatorial responses may more accurately reflect the consequences of in vivo exposure.

Topics: Breast Neoplasms; Chromans; Drug Interactions; Equol; Estrogens; Estrogens, Non-Steroidal; Gene Expression; Genistein; Humans; Isoflavones; Luciferases; Phytoestrogens; Plant Preparations; Transfection; Tumor Cells, Cultured

The hypothesis was tested that the rate of postmenopausal bone loss is inversely associated with long-term urinary excretion of phyto-oestrogens, as a marker of habitual dietary intake.. Secondary analysis of a 10-year follow-up study (1979 1989) among postmenopausal women in the Netherlands.. From the original population of 154 women, 32 women were selected with an annual rate of radial bone loss of < or = 0.5% over the first 5 years of the study and 35 women with a rate of > or = 2.5% per year.. The isoflavonoids genistein, daidzein and equol, and the lignan enterolactone were determined by gas chromatography mass spectrometry in aggregate samples from annually collected urine samples. Cortical bone density of the radius had previously been measured annually by single-photon absorptiometry.. Excretion of isoflavonoids did not differ between both groups, although in multivariate analysis equol excretion was weakly positively associated with rate of bone loss in the 5 years after the menopause. Enterolactone excretion was significantly higher in the group with high rate of bone loss. This positive association remained in multivariate linear regression analysis after adjustment for age, years since menopause, body mass index and intake of calcium, vegetable protein and dietary fibre.. Enterolactone excretion is likely to be an indicator of consumption of grains and legumes; it is not clear whether the observed positive association with rate of bone loss is a causal one. Our results do not support a preventive effect of low, unsupplemented dietary intake of phyto-oestrogens on postmenopausal cortical bone loss. However, no conclusions can be drawn about effects of higher doses of phyto-oestrogens.

Topics: 4-Butyrolactone; Aging; Bone Density; Chromans; Equol; Estrogens, Non-Steroidal; Female; Gas Chromatography-Mass Spectrometry; Genistein; Humans; Isoflavones; Lignans; Linear Models; Middle Aged; Osteoporosis, Postmenopausal; Phytoestrogens; Plant Preparations; Postmenopause; Prospective Studies; Time Factors

The aims of this study were to estimate the dietary intake of phytoestrogens and to measure urinary phytoestrogen excretion in postmenopausal Chinese women.. Postmenopausal Chinese women were recruited from the hormone replacement clinic of the Prince of Wales Hospital. Dietary intake of phytoestrogen-rich foods was estimated using a food frequency questionnaire. Urinary output of the isoflavonoids daidzein and genistein and the metabolite of daidzein, equol, was measured using high-performance liquid chromatography.. The mean daily excretion of daidzein, genistein and equol was 3.24 (+/- 3.63), 1.47 (+/- 1.75) and 0.64 (+/- 1.53) mumol, respectively. The total mean daily isoflavonoid excretion was 5.36 (+/- 5.27) mumol.. Urinary excretion of isoflavonoid phytoestrogens in this Chinese population was lower than that reported in Japanese subjects. This may be due to the higher consumption of legumes, especially soy products, in the Japanese compared to the Chinese diet. The intake of green vegetables was higher in the Chinese subjects, and this food group may be the main contributor to their total phytoestrogen intake.

Topics: Adult; Animals; China; Chromans; Diet; Diet Records; Equol; Estrogens, Non-Steroidal; Fabaceae; Female; Genistein; Glycine max; Humans; Isoflavones; Middle Aged; Milk; Phytoestrogens; Plant Preparations; Postmenopause; Vegetables

The aim of this work was to determine the antioxidant activities of a range of phytoestrogenic isoflavones. The antioxidant activity in the aqueous phase was determined by means of the ABTS.+ total antioxidant activity assay. The results show that the order of reactivity in scavenging the radical in the aqueous phase is genistein > daidzein = genistin approximately equal to biochanin A = daidzin > formononetin approximately equal to ononin, the latter displaying no antioxidant activity. The importance of the single 4'-hydroxyl group in the reactivity of the isoflavones, as scavengers of aqueous phase radicals, as well as the 5'7-dihydroxy structure is demonstrated. Examination of their abilities to enhance the resistance of low density lipoproteins to oxidation supports the observation that genistein is the most potent antioxidant among this family of compound studied, both in the aqueous and in the lipophilic phases.

Topics: Antioxidants; Copper; Estrogens, Non-Steroidal; Free Radical Scavengers; Genistein; Isoflavones; Lipoproteins, LDL; Oxidation-Reduction; Phytoestrogens; Plant Preparations; Plants

Topics: Estrogens, Non-Steroidal; Genistein; Glycine max; Humans; Infant; Infant Food; Isoflavones; Phytoestrogens; Plant Preparations; Plants

Thirteen isoflavonoids, flavonoids, and lignans, including some known phytoestrogens, were evaluated for their effects on DNA synthesis in estrogen-dependent (MCF-7) and -independent (MDA-MB-231) human breast cancer cells. Treatment for 24 hours with most of the compounds at 20-80 microM sharply inhibited DNA synthesis in MDA-MB-231 cells. In MCF-7 cells, on the other hand, biphasic effects were seen. At 0.1-10 microM, coumestrol, genistein, biochanin A, apigenin, luteolin, kaempferol, and enterolactone induced DNA synthesis 150-235% and, at 20-90 microM, inhibited DNA synthesis by 50%. Treatment of MCF-7 cells for 10 days with genistein or coumestrol showed continuous stimulation of DNA synthesis at low concentrations. Time-course experiments with genistein in MCF-7 cells showed effects to be reversed by 48-hour withdrawal of genistein at most concentrations. Induction of DNA synthesis in MCF-7 cells, but not in MDA-MB-231 cells, is consistent with an estrogenic effect of these compounds. Inhibition of estrogen-dependent and -independent breast cancer cells at high concentrations suggests additional mechanisms independent of the estrogen receptor. The current focus on the role of phytoestrogens in cancer prevention must take into account the biphasic effects observed in this study, showing inhibition of DNA synthesis at high concentrations but induction at concentrations close to probable levels in humans.

Topics: Anticarcinogenic Agents; Breast Neoplasms; Coumestrol; DNA; Estradiol; Estrogens, Non-Steroidal; Flavonoids; Genistein; Humans; Isoflavones; Kinetics; Lignans; Phytoestrogens; Plant Preparations; Tumor Cells, Cultured

The interactions of human Sex steroid binding protein (SBP) and the lignans [Nordihydrogaiaretic acid (NDGA) enterolactone (Ent), enterodiol (End)] and isoflavonoid phytoestrogens [Equol (Eq), diazein Dad), genistein (Gen)] were studied. The phytoestrogens had different dose-dependent inhibitory effects on steroid binding by SBP. Their relative efficiencies were: Ent> or = NDGA = Eq > Gen for displacing E2 and Eq > Ent > NDGA > Gen for displacing T. End and Dad were much less active. Scatchard analysis suggested that NDGA had similar non- competitive effects on T and E2 binding by reducing the number of binding sites without changing the association constants. But Eq seemed to inhibit E2 binding non-competitively and T binding competitively. NDGA binding to SBP reduced the immunorecognition of SBP by monospecific anti-SBP antibodies, suggesting that NDGA changed SBP immunoreactivity. Unlike NDGA, Eq binding to SBP caused no immunological changes in SBP, indicating qualitative differences in the effects of the lignan and isoflavonoid. Our results indicate that phytoestrogens may modulate the SBP activity and so influence the role of this protein in the delivery of hormonal information to sex steroid-dependent cells.

Topics: 4-Butyrolactone; Electrophoresis, Polyacrylamide Gel; Estradiol; Estrogens, Non-Steroidal; Genistein; Humans; Immunoelectrophoresis, Two-Dimensional; Isoflavones; Kinetics; Lignans; Masoprocol; Phytoestrogens; Plant Preparations; Plants; Sex Hormone-Binding Globulin; Structure-Activity Relationship; Testosterone

The metabolic fate of the dietary isoflavones daidzein and genistein was investigated in human volunteers challenged with soya. Urinary diphenols, isolated by partition chromatography on Sephadex LH-20, were characterized and identified by profile capillary gas chromatography (GC) and electron ionization mass spectrometry (GC-EIMS) analysis of the trimethylsilyl ether (TMS) derivatives. Novel isoflavonic phytoestrogens found in the urine of volunteers were those of tetrahydrodaidzein, dihydrogenistein, 6'-hydroxy-O-demethylangolesin and 2-dehydro-O-demethylangolensin. Other known diphenols identified were those of equal, dehydrodaidzein, O-demethylangolensin, daidzein, genistein, glycitein, and the lignan enterolactone. Two other urinary isomers with a fragmentation pattern closely resembling that of the persilylated TMS ethers of cis/trans-isomers of tetrahydrodaidzein, were characterized based on the elucidation of fragments associated with the loss of a non-phenolic-OTMS functional group in ring-C. These are fragments presented in the persilylated mass spectra of isoflavan-4-ols and isoflav-3-ene-4-ols, demonstrated here by a combination of simple and tandem mass spectrometry study of the deuterated persilylated TMS ethers of dihydrodaidzein. In a similar study we also present the data on the structural identification and fragment elucidation of the keto/enol tautomers of the TMS ether derivatives of the dihydro derivatives of daidzein and genistein, observed in the urine of volunteers and considered probable products of the derivatization process. Finally, the GC and GC-MS data of two unknown isoflavonoids and that of a lignan-like compound are presented together with those of dihydrodaidzein, dihydrogenistein, tetrahydrodaidzein and 2-dehydro-O-demethylangolensin. The latter four were obtained here as products of small scale chemical synthesis in a preliminary study on the tentative identification of urinary isoflavonoids in human volunteers challenged with soya.

Topics: Diet; Estrogens, Non-Steroidal; Fabaceae; Flavonoids; Genistein; Humans; Isoflavones; Mass Spectrometry; Phytoestrogens; Plant Preparations; Plants, Edible; Plants, Medicinal

The synthesis of the important diphenolic isoflavone type phytoestrogens starting from the corresponding unprotected phenols and arylacetic acids is discussed. The aryl rings may carry additional alkyl, methoxy, and/or halogeno groups. Intermediate polyhydroxy deoxybenzoins can also be isolated in good yield. Isotopically labeled isoflavone phytoestrogens were prepared for use as internal standards in ion exchange chromatography and GC-MS selected ion monitoring (SIM technique). Traditional methods rely on total synthesis using deuterated starting materials for the preparation of labeled isoflavonoid structures. We have used successfully an application where the H/D exchange is performed within the finished molecular framework, based on the exchange of aromatic protons that are ortho or para to a phenolic OH group. By this method the deuterated products are available in an isotopic purity of 90% or higher.

Topics: Chromatography, Ion Exchange; Deuterium; Estrogens, Non-Steroidal; Genistein; Isoflavones; Mass Spectrometry; Phytoestrogens; Plant Preparations; Trifluoroacetic Acid

Soybean consumption is associated with reduced rates of prostate and other cancers, possibly due in part to the presence of isoflavones. The metabolism and disposition of these soya-derived phytoestrogens after chronic soya exposure were studied on a metabolic unit in six healthy males (21-35 yrs of age) who consumed an unrestricted hospital diet and a 12-oz portion of soymilk with each meal for one month. The daily isoflavone intake was about 100 mg of daidzein (mostly as diadzin) and about 100 of mg of genistein (mostly as genistin). At two-week intervals, excretion of isoflavones in urine was studied, during which time the subjects consumed a constant basal diet for three to four days, ingested the full daily 36-oz portion of soymilk within 30 minutes each day for one to two days, and collected urine continuously. The urinary recovery of ingested diadzin plus daidzein (46.9 +/- 15.2%, mean +/- SD) and genistin plus genistein (14.6 +/- 9.2%) did not change with prolonged soya ingestion. The absorption half-lives (t1/2) for daidzein and genistein and the appearance t1/2 for equol (1 subject) were initially 1.5 +/- 0.4, 1.9 +/- 0.6, and 2.2 hours, respectively, and 2.5 +/- 1.1 (p = 0.06 compared with baseline) 1.4 +/- 0.9 (p = 0.03) compared with baseline), and 4.2 hours, respectively, during one month of soymilk ingestion. The excretion t1/2 for daidzein, genistein, and equol were initially 2.9 +/- 0.5, 3.8 +/- 0.7, and 5.2 hours, respectively, and 3.9 +/- 1.2 (p - 0.03), 5.5 +/- 1.6 (p = 0.02), and 9.7 hours, respectively, during one month of soymilk ingestion. These results indicate that chronic soya exposure did not induce significant changes in the metabolic pathways of isoflavones but altered the time courses of daidzein and genistein excretion. Thus chronic exposure to soya might prolong the tissue exposure to the presumed biologically active free and unconjugated forms of these isoflavones and thereby enhance their oncoprotective effects.

Topics: Absorption; Adult; Chromans; Equol; Estrogens, Non-Steroidal; Genistein; Glycine max; Humans; Isoflavones; Kinetics; Male; Phytoestrogens; Plant Preparations

Two estrogenic substances of plant origin have been identified in beer using gas chromatography/mass spectrometry. These phytoestrogens, daidzein and genistein, have previously been shown to be biologically active in animals. Confirming the presence of biologically active phytoestrogens in beer and their possible presence in other beverages, suggests that there may be clinically significant effects related to sustained exposure to phytoestrogens contained in alcoholic beverages.

Topics: Beer; Estrogens; Estrogens, Non-Steroidal; Gas Chromatography-Mass Spectrometry; Humans; Isoflavones; Phytoestrogens; Plant Preparations

Various phytoestrogens such as formononetin, daidzein, genistein and equol were synthesized. Their purity was assessed by various analytical techniques including melting point determination, thin-layer chromatography (TLC), infra-red spectra (i.r. spectra), nuclear magnetic resonance (1H- and 13C-NMR) and gas chromatography coupled with mass spectrometry (GC-MS). The estrogenic activity of these compounds, as well as biochanin A and coumestrol, was biologically tested by the induction of vitellogenin secretion in yearling sturgeon and compared to the activity of estradiol-17 beta. Pure daidzein, biochanin A, genistein, equol and coumestrol all had estrogenic activity as assessed by their induction of hepatic synthesis of vitellogenin when administrated intraperitoneally to yearling Siberian sturgeon. Coumestrol seemed to be the most potent compound, inducing the most vitellogenin secretion with the lowest dose administered. Formononetin was inactive when administered by the intraperitoneal route. All the phytoestrogens tested were considerably less potent than estradiol-17 beta.

Topics: Animals; Chromans; Chromatography, Thin Layer; Coumestrol; Equol; Estrogens; Estrogens, Non-Steroidal; Fishes; Gas Chromatography-Mass Spectrometry; Genistein; Isoflavones; Liver; Magnetic Resonance Spectroscopy; Phytoestrogens; Plant Preparations; Spectrophotometry, Infrared; Vitellogenins

The phytoestrogens daidzein, genistein, equol and coumestrol were found to stimulate microsomal prostaglandin H synthase (PHS) in vitro in a concentration-dependent manner when PHS-activity was measured by arachidonic acid-dependent oxygen uptake. These compounds were co-oxidized by PHS and the conversion of parent compounds was measured by HPLC analysis. The stimulation of PHS-cyclooxygenase by these compounds was partially reversed at high concentrations probably due to their antioxidant properties causing inhibition. In contrast, the monomethyl ethers of daidzein and genistein, formononetin and biochanin A, had little or weakly inhibitory effect on PHS, and appear to be no or poor co-substrates for PHS. Compared to the equine estrogen equilin, its metabolite d-equilenin was poorly metabolized by PHS and inhibited rather than stimulated PHS-cyclooxygenase activity in vitro. The resorcylic acid lactones zearalenone and zeranol, on the other hand, were surprisingly good inhibitors of PHS-cyclooxygenase. Furthermore, zeranol inhibited both the arachidonic acid and the hydrogen-peroxide-dependent oxidation of DES in contrast to indomethacin which inhibited only cyclooxygenase-dependent co-oxidation of DES. The results of this in vitro study are discussed in the context of data on synthetic and steroidal estrogens and support the idea that PHS-activity may be modulated by interaction with certain estrogenic compounds.

Topics: Cyclooxygenase Inhibitors; Equilin; Estrogens; Estrogens, Non-Steroidal; Indenes; Isoflavones; Phytoestrogens; Plant Preparations; Prostaglandin-Endoperoxide Synthases; Structure-Activity Relationship; Zearalenone

The thermospray mass spectra of the phytoestrogens have intense protonated molecular ions but contain few or no ions indicative of structure. Tandem mass spectrometry (MS/MS) was used to obtain daughter ion spectra containing ions unique to the different structural characteristics of each phytoestrogen subclass and was used both to confirm identification and propose structures for unknowns. In addition to unique daughter ion spectra, MS/MS was used as a class identifier to detect phytoestrogens through the neutral loss of 56 (due to consecutive losses of CO) that is common to all members of this family. Several sources of soy protein were investigated to confirm the presence or absence of phytoestrogens. In one preparation investigated, daidzein and genistein were detected as well as an unknown phytoestrogen of the Biochanin A subclass. This unknown has been tentatively identified as 6,7-dihydroxy-4'-methoxyisoflavone using its daughter ion spectrum.

Topics: Animal Feed; Chemical Phenomena; Chemistry; Chromatography, High Pressure Liquid; Chromatography, Liquid; Estrogens; Estrogens, Non-Steroidal; Genistein; Indicators and Reagents; Isoflavones; Mass Spectrometry; Phytoestrogens; Plant Preparations; Plant Proteins, Dietary; Soybean Proteins; Spectrophotometry, Ultraviolet

It is becoming increasingly apparent that dietary factors may play a role in the etiology of hormone dependent neoplasias. It has been hypothesized that estrogens play some role in the etiology of benign prostatic hyperplasia (BPH) in the canine. The presence of estrogen receptor binding activity in a fraction of canine urine purified by high performance liquid chromatography (HPLC) that did not correspond to estriol, estradiol, estrone or any of their primary metabolites was observed in the present study. We used thermospray-mass spectrometry and GC-MS to identify the phytoestrogens daidzein, equol, formononetin and genistein in HPLC purified fractions of urine obtained from male beagles. Using the same techniques we also confirmed the presence of daidzein and genistein in the commercial diet fed to these same dogs. Using the immature rat uterine cytosol estrogen receptor assay, relative binding affinities of 0.08, 1.1, less than 0.01 and 3.9% were obtained for daidzein, equol, formononetin and genistein, respectively when compared to estradiol (100%). In conclusion, phytoestrogens are present in urine of male beagles. Moreover, the commercial diet fed to these dogs contains isoflavones which can be converted to equol by intestinal microflora. These results suggest the need for investigations of phytoestrogens (e.g. equol) excreted into the urine daily and its relationship to the incidence and severity of BPH in the dog.

Topics: Animal Feed; Animals; Binding, Competitive; Chromans; Chromatography, High Pressure Liquid; Dogs; Equol; Estrogens; Estrogens, Non-Steroidal; Gas Chromatography-Mass Spectrometry; Genistein; Isoflavones; Male; Phytoestrogens; Plant Preparations; Prostatic Hyperplasia; Receptors, Estrogen; Reference Values

It was recently observed that the urinary excretion of animal lignans is low in postmenopausal breast cancer patients compared to normal omnivorous and vegetarian women. In addition, the mean excretion of the isoflavonic phytoestrogen equol tended to be lower. Because nonhuman primates appear to be remarkably resistant to the carcinogenic effect of estrogens, we investigated the possible occurrence of lignans and phytoestrogens in the urine of chimpanzees on their regular diet. Five major diphenols were isolated and identified by capillary gas chromatography and mass spectrometry by comparison with synthesized authentic reference compounds. Three of these compounds, the phytoestrogen equol and its precursor daidzein, the lignan enterolactone, were according to preliminary assays excreted in very large amounts. In addition, the lignan enterodiol and the daidzein metabolite O-desmethylangolensin were identified. It is concluded that the chimpanzee excretes both isoflavonic phytoestrogens and lignans in urine, apparently in high concentrations. It is suggested that these compounds may play a role in the maintenance of the resistance against carcinogenic effects of estrogens, which nonhuman primates possess, because both equol and enterolactone have been shown to have antiestrogenic properties in animals. However, much further work is necessary before the possible biological role of these compounds may be established.

Topics: 4-Butyrolactone; Animals; Butylene Glycols; Chromans; Diet; Equol; Estrogens; Estrogens, Non-Steroidal; Isoflavones; Lignans; Male; Pan troglodytes; Phytoestrogens; Plant Extracts; Plant Preparations