Genistein accumulates in body depots and is mobilized during fasting, reaching estrogenic levels in serum that counter the hormonal actions of estradiol and organochlorines.

Isoflavones are important dietary compounds that are consumed with the daily diet and elicit important biological actions. Here we report on the ability of genistein to partially accumulate in body depots of male mice, be released following fasting, and modulate the actions of estradiol and environmental estrogens in reproductive and nonreproductive target organs of estrogen-reporter mice (ERE-tK-luciferase). After the consumption of 50 mg/kg/day for 3 days, genistein accumulates in body compartments where it remains at functionally active levels for at least 15 days. Following 48 h of fasting, its concentration increased in serum from 99 +/- 13 to 163 +/- 17 nM. These levels are sufficient to exert an estrogenic effect in the testis and liver, as revealed by a twofold increase in luciferase gene expression. beta-Benzene-hexachloride (betaBHC) given at the concentration of 100 mg/kg/day for 3 days also accumulates in the body and is released by fasting, reaching serum levels of 176 +/- 33 nM, upregulating the luciferase gene in the liver and inhibiting its expression in the testis. When genistein was given in combination with betaBHC at doses sufficient to induce accumulation of both in body depots, the genistein mobilized by fasting reversed the action of the mobilized betaBHC in the testis. Acute administration of nutritional doses of genistein inhibited the action of estradiol and reversed the antiestrogenic action of o,p'-DDT: 1,1,1,-trichloro-2(p-chlorophenyl)-2-(o-chlorophenyl)ethane in the liver and the antiestrogenic action of betaBHC in the testis. Genistein had an additive effect with the ER agonist p,p'-DDT: 1,1,1,-trichloro-2,2-bis(p-chlorophenyl)ethane in the liver. The observed effects may be relevant to a protective action of phytoestrogens against estrogen receptor-interacting pollutants as well as the dietary modulation of estradiol action.

Evidence of a lack of effect of a phytoestrogen regimen on the levels of C-reactive protein, E-selectin, and nitrate in postmenopausal women.

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.

Urinary and serum concentrations of seven phytoestrogens in a human reference population subset.

Diets rich in naturally occurring plant estrogens (phytoestrogens) are strongly associated with a decreased risk for cancer and heart disease in humans. Phytoestrogens have estrogenic and, in some cases, antiestrogenic and antiandrogenic properties, and may contribute to the protective effect of some diets. However, little information is available about the levels of these phytoestrogens in the general US population. Therefore, levels of phytoestrogens were determined in urine (N=199) and serum (N=208) samples taken from a nonrepresentative subset of adults who participated in NHANES III, 1988-1994. The phytoestrogens quantified were the lignans (enterolactone, enterodiol, matairesinol); the isoflavones (genistein, daidzein, equol, O-desmethylangolensin); and coumestrol (urine only). Phytoestrogens with the highest mean urinary levels were enterolactone (512 ng/ml), daidzein (317 ng/ml), and genistein (129 ng/ml). In serum, the concentrations were much less and the relative order was reversed, with genistein having the highest mean level (4.7 ng/ml), followed by daidzein (3.9 ng/ml) and enterolactone (3.6 ng/ml). Highly significant correlations of phytoestrogen levels in urine and serum samples from the same persons were observed for enterolactone, enterodiol, genistein, and daidzein. Determination of phytoestrogen concentrations in large study populations will give a better insight into the actual dietary exposure to these biologically active compounds in the US population.

Quantification of isoflavones and lignans in serum using isotope dilution liquid chromatography/tandem mass spectrometry.

Phytoestrogens (isoflavones and lignans) are receiving increasing attention due to a potential protective effect against a number of complex diseases. However, in order to investigate these associations, it is necessary to accurately quantify the levels of phytoestrogens in foods and biological fluids. We report an assay for three isoflavones (daidzein, genistein, and glycitein), two metabolites of daidzein (O-desmethylangolensin and equol), and two lignans (enterodiol and enterolactone) in human serum using electrospray ionisation liquid chromatography/mass spectrometry (LC/MS) with selective reaction monitoring. A simple, highly automated sample preparation procedure requires only 200 microL of sample and utilises one solid-phase extraction stage. Limits of detection are in the region of 10 pg/mL for all analytes except equol, which had a limit of detection of approximately 100 pg/mL. The method developed is suitable for measuring the concentrations of phytoestrogens in blood samples collected from large epidemiological studies. The results of the analysis of serum samples from 300 men and women living in the UK are reported.

A randomized placebo-controlled crossover trial with phytoestrogens in treatment of menopause in breast cancer patients.

OBJECTIVE: Phytoestrogens are popular in treatment of menopause, although scientific evidence is insufficient as to their efficacy. We studied the effects of daily use of isoflavonoids on climacteric symptoms and quality of life in patients with a history of breast cancer. METHODS: Sixty-two postmenopausal symptomatic women were randomized to use either phytoestrogen (tablets containing 114 mg of isoflavonoids) or a placebo for 3 months; the treatment regimens were reversed after a 2-month washout period. Fifty-six women completed the study. Menopausal symptoms were recorded on the Kupperman index and the visual analogue scale, and working capacity and mood changes were assessed via validated questionnaires. In addition, we followed the levels of phytoestrogens, follicle-stimulating hormone (FSH), luteinizing hormone (LH), estradiol, and sex hormone-binding globulin. Liver enzymes and creatinine were also assessed at each visit. RESULTS: The phytoestrogen regimen raised the circulating levels of phytoestrogens (daidzein, genistein, equol) 19- to 106-fold. The Kupperman index was reduced by 4.2 +/- 9.6 (mean +/- standard deviation) (15.5%) during phytoestrogen use and similarly by 4.0 +/- 8.1 (14.7%) during placebo use (P nonsignificant). The quality of life parameters (working capacity, mood changes) were unaffected by phytoestrogen. In addition, the phytoestrogen regimen caused no changes in FSH, LH, estradiol, or sex hormone-binding globulin. Phytoestrogen treatment was well tolerated and caused no changes in liver enzymes, creatinine, body mass index, or blood pressure. Of the 56 women, 25 (44.6%) preferred the phytoestrogen regimen, 15 preferred the placebo (26.8%), and 16 (28.6%) reported no preference (nonsignificant). CONCLUSION: Pure isoflavonoids did not alleviate subjective menopausal symptoms in breast cancer patients.

The soya isoflavone content of rat diet can increase anxiety and stress hormone release in the male rat.

RATIONALE: Most commercial rodent diets are formulated with soya protein and therefore contain soya isoflavones. Isoflavones form one of the main classes of phytoestrogens and have been found to exert both oestrogenic and anti-oestrogenic effects on the central nervous system. The effects have not been limited to reproductive behaviour, but include effects on learning and anxiety and actions on the hypothalamo-pituitary axis. It is therefore possible that the soya content of diet could have significant effects on brain and behaviour and be an important source of between-laboratory variability. OBJECTIVES: To determine whether behaviour in two animal tests of anxiety, and stress hormone production, would differ between rats that were fed a diet which was free of soya isoflavones and other phytoestrogens (iso-free) and those that were fed a diet which contained 150 microg/g of the isoflavones genistein and daidzein (iso-150). This controlled diet has an isoflavone concentration similar to that in the maintenance diet routinely used in our institution. METHODS: Male rats were randomly allocated to the iso-free and iso-150 diets and their body weights and food and water consumption were recorded for 14 days. They were then maintained on the same diets, but housed singly for 4 days, before testing in the social interaction and elevated plus-maze tests of anxiety. Corticosterone concentrations in both dietary groups were determined under basal conditions and after the stress of the two tests of anxiety. Vasopressin and oxytocin concentrations were determined after brief handling stress. RESULTS: The groups did not differ in food or water intake, body weight or oxytocin concentrations. Compared with the rats fed the iso-free diet, the rats fed the iso-150 diet spent significantly less time in active social interaction and made a significantly lower percentage of entries onto the open arms of the plus-maze, indicating anxiogenic effects in both animal tests. The groups did not differ in their basal corticosterone concentrations, but the iso-150 group had significantly elevated stress-induced corticosterone concentrations. Stress-induced plasma vasopressin concentrations were also significantly elevated in the iso-150 diet group compared with the iso-free rats. CONCLUSIONS: Major changes in behavioural measures of anxiety and in stress hormones can result from the soya isoflavone content of rat diet. These changes are as striking as those seen following drug administration and could form an important source of variation between laboratories.

Quantitation of soy-derived phytoestrogens in human breast tissue and biological fluids by high-performance liquid chromatography.

A new and reliable HPLC method for the quantitation of daidzein, equol, and genistein in human breast tissue has been developed. The method was applied to biopsies from women undergoing breast reductions, who, prior to surgery, had ingested either a soy isoflavone preparation or a placebo tablet. The results were compared with data collected for urine and serum of the same subjects using standard methods. The limits of detection in the breast tissue homogenate were 24.7 nmol/l for daidzein, 148.0 nmol/l for equol, and 28.4 nmol/l for genistein (S/N of 3). The chromatographic limits of quantitation were 62.5 nmol/l for daidzein and genistein, and 125.0 nmol/l for equol, for which the accuracies were 86.0%, 83.6%, and 81.8%, respectively. The coefficients of variation of these measurements were all below 20% (11.1% for daidzein, 16.4% for genistein, and 13.2% for equol). The sample preparation comprised a concentration step and the absolute limits of quantitation were, therefore, 4.7 nmol/l, 18.8 nmol/l, and 0.94 nmol/l for daidzein and genistein, and 9.4 nmol/l, 37.5 nmol/l, and 1.9 nmol/l for equol in urine, serum, and breast tissue homogenate, respectively. Recoveries were between 70% (+/-5.6%) in breast tissue homogenate and 100% (+/-14.1%) in urine and serum for all three compounds. Equol (less than 1 micromol/l homogenate) was found to be the predominant phytoestrogen in breast tissue and its concentrations exceeded those in serum. The concentrations of phytoestrogens were at least 100-fold higher in urine than in serum and breast tissue.

Enterolactone inhibits the growth of 7,12-dimethylbenz(a)anthracene-induced mammary carcinomas in the rat.

The inverse association between a high enterolactone (ENL) concentration in both urine and serum, and the risk of breast cancer found in epidemiological studies suggests a chemopreventive action for ENL. However, no causal relationship has been established in clinical studies or in experimental models for breast cancer. In the present study, the potential chemopreventive action of p.o. administered ENL (1 or 10 mg/kg of body weight) was tested in 7,12-dimethylbenz(a)anthracene-induced mammary cancers of the rat. Rats were maintained on a standard open-formula chow diet. Daily p.o. administration of ENL at a dose of 10 mg/kg of body weight for 7 weeks significantly inhibited tumor growth. The growth-inhibitory effect of ENL was more pronounced on the new tumors, which developed during the treatment period, but ENL also inhibited the growth of those tumors established before the start of the lignan administration. The rat serum concentration of ENL, which illustrated a permanent positive effect on breast cancer growth, was 0.4 microM, which is >10-fold as compared with the serum concentrations found in the general human population. The effect of ENL was not restricted to any specific histological tumor type. ENL was demonstrated to act as a weak aromatase inhibitor in vitro and to reduce the relative uterine weight of the 7,12-dimethylbenz(a)anthracene-treated nonovariectomized rats. However, in a short-term assay ENL had no effect on the uterine growth of the intact or androstenedione-treated immature rats. Thus, the mechanism of the ENL action and its minimum or optimal daily dose remains to be clarified.

Determination of trace isoflavone phytoestrogens in biological materials by capillary electrochromatography.

Capillary electrochromatography using a specialty monolithic matrix was utilized in developing a rapid and highly efficient separation of isoflavones in biological materials. Without a preconcentration technique, it is relatively easy to reach ppm-ppb concentrations of these compounds in soy-based foods and verify them structurally using a photodiode array detector. With on-column preconcentration, we were able to measure low-ppb levels in human serum. Using blood samples from human volunteers, whose diet was supplemented by a soy-based product, the method has been validated for high-throughput screening of isoflavones in clinical studies.

Phytoestrogen concentrations in serum from Japanese men and women over forty years of age.

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.

Liquid chromatographic-photodiode array mass spectrometric analysis of dietary phytoestrogens from human urine and blood.

Dietary phytoestrogens have been implicated in the prevention of chronic diseases. However, it is uncertain whether the phytoestrogens or the foods associated with phytoestrogens account for the observed effects. We report here a new liquid chromatography photodiode array mass spectrometry (LC-PDA-MS) assay for the determination of nanomolar amounts of the most prominent dietary phytoestrogens (genistein, dihydrogenistein, daidzein, dihydrodaidzein, glycitein, O-desmethylangolensin, hesperetin, naringenin, quercetin, enterodiol, enterolactone) in human plasma or serum and urine. This assay was found to be suitable for the assessment of quercetin exposure in an onion intervention study by measuring urinary quercetin levels. Other successful applications of this assay in clinical and epidemiologic studies validated the developed method and confirmed previous results on the negative association between urinary isoflavone excretion and breast cancer risk.

Human 17beta-hydroxysteroid dehydrogenase type 5 is inhibited by dietary flavonoids.

Phytoestrogens contained in a vegetarian diet are supposed to have beneficial effects on the development and progression of a variety of endocrine-related cancers. We have tested the effect of a variety of dietary phytoestrogens, especially flavonoids, on the activity of human 17beta-hydroxysteroid dehydrogenase type 5 (17beta-HSD 5), a key enzyme in the metabolism of estrogens and androgens. Our studies show that reductive and oxidative activity of the enzyme are inhibited by many compounds, especially zearalenone, coumestrol, quercetin and biochanin A. Among flavones, inhibitor potency is enhanced with increased degree of hydroxylation. The most effective inhibitors seem to bind to the hydrophilic cofactor binding pocket of the enzyme.

Genistein affects ER beta- but not ER alpha-dependent gene expression in the hypothalamus.

Isoflavone phytoestrogens are growing increasingly popular because of their reported cardiovascular and anticarcinogenic properties, but the effects of these compounds in the brain are largely unknown. In a previous study, we found that an isoflavone supplement, containing a mixture of soy phytoestrogens, inhibited estrogen-dependent female sexual behavior and was antiestrogenic for both ER alpha- and ER beta-dependent gene expression in the hypothalamus. Here we examined the impact of the soy isoflavone genistein, a major component of the supplement, on estrogen-dependent female sexual behavior and ER alpha- and ER beta-dependent gene expression in the rat brain. Genistein, at a dietary concentration of 100 or 500 ppm had no effect on lordosis behavior in rats. However, at 500 ppm genistein had differential activity through ER alpha and ER beta in the hypothalamus. Genistein had no effect, in either the presence or absence of 17 beta-E2, on oxytocin receptor density in the ventromedial nucleus of the hypothalamus, an estrogen-dependent action thought to be regulated via ER alpha. However, genistein increased ER beta mRNA expression in the paraventricular nucleus of the hypothalamus by 24%, whereas 17 beta-E2 decreased ER beta mRNA expression by 26%, a process likely mediated by ER beta itself. These results suggest that at this dose, genistein has antiestrogenic action through ER beta in the paraventricular nucleus but negligible activity through ER alpha in the brain.

Estrogenic isoflavones in rodent diets.

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.

The correlation between dietary soy phytoestrogens and neuropathic pain behavior in rats after partial denervation.

UNLABELLED: Soy diets suppress the development of neuropathic pain behavior in rats undergoing partial sciatic nerve ligation (PSL) injury. Phytoestrogens, plant isoflavones and lignans, abundantly found in soy products, have powerful estrogenic properties. Because, in some preparations, steroid estrogens were found to exert antinociception, we examined whether the analgesic effect of dietary soy is mediated by phytoestrogens. Male Wistar rats were fed five different diets containing 8-180 microg of phytoestrogens per gram. These diets were administered 2 wk before and 2 wk after PSL injury. Levels of tactile allodynia and mechanical and heat hyperalgesia of these rats were determined on Days 3, 8, and 14 after PSL injury. Plasma levels of two major phytoestrogens (genistein and daidzein) and two daidzein metabolites (equol and dihydrodaidzein) were assessed on Day 14 postoperatively. We found that the plasma concentration of these phytoestrogens and the levels of allodynia and hyperalgesia varied highly among dietary groups. Average plasma concentrations of phytoestrogens were associated with reduced levels of tactile allodynia and mechanical hyperalgesia, but not with reduced heat allodynia. Low and high plasma phytoestrogen levels were not analgesic in these tests. This report is the first to show that, at certain plasma concentrations, phytoestrogens reduce neuropathic pain in rats. IMPLICATIONS: Dietary soy suppresses neuropathic pain in rats after partial sciatic nerve ligation. Some of the pain-suppression properties of soy can be attributed to phytoestrogens, isoflavones abundantly found in soy products. Average, but not low or high, plasma levels of phytoestrogens are associated with analgesia.

Altered sexually dimorphic nucleus of the preoptic area (SDN-POA) volume in adult Long-Evans rats by dietary soy phytoestrogens.

Naturally occurring estrogen-like molecules in plants (phytoestrogens), present via soy, in animal diets can alter morphology and physiology in rodents. Phytoestrogens have the ability to bind estrogen receptors and exert many of the biological responses evoked by physiological estrogens. This study characterized the effects of dietary phytoestrogens on the expression of body and prostate weight, circulating testosterone and estradiol levels, puberty onset, vaginal cyclicity, and volume of the sexually dimorphic nucleus of the preoptic area (SDN-POA) in Long-Evans rats. Using different experimental protocols, animals were fed either a phytoestrogen-rich (Phyto-600) or a phytoestrogen-free (Phyto-free) diet. Animals fed the Phyto-600 diet displayed significantly decreased body weights (in males and females), prostate weights and delayed puberty in females compared to that of animals fed the Phyto-free diet. Circulating testosterone or estradiol levels in males or estrous cyclicity were not altered by the diets. The volume of the SDN-POA was significantly altered by a change in diet at 80 days of age where one-half of the males or females fed the Phyto-600 diet (from birth) were switched to the Phyto-free diet until 120 days of age. Males initially fed a Phyto-600 diet but changed to a Phyto-free diet had significantly smaller SDN-POA volumes compared to males fed the Phyto-600 diet (long-term). These data suggest that consumption of phytoestrogens via a soy diet, significantly: (1) decreases body and prostate weight, (2) delays puberty onset, and (3) alters SDN-POA volumes during adulthood.

Dietary soy-phytoestrogens decrease testosterone levels and prostate weight without altering LH, prostate 5alpha-reductase or testicular steroidogenic acute regulatory peptide levels in adult male Sprague-Dawley rats.

Nutritional factors, especially phytoestrogens, have been extensively studied for their potential beneficial effects against hormone-dependent and age-related diseases. The present study describes the short-term effects of dietary phytoestrogens on regulatory behaviors (food/water intake, locomotor activity and body weight), prostate weight, prostate 5alpha-reductase enzyme activity, reproductive hormone levels, and testicular steroidogenic acute regulatory peptide (StAR) levels in adult Sprague-Dawley rats. Animals were fed either a phytoestrogen-rich diet containing approximately 600 microg/g isoflavones (as determined by HPLC) or a phytoestrogen-free diet. After 5 weeks of consuming these diets, plasma phytoestrogen levels were 35 times higher in animals fed the phytoestrogen-rich vs phytoestrogen-free diets. Body and prostate weights were significantly decreased in animals fed the phytoestrogen-rich diet vs the phytoestrogen-free fed animals; however, no significant change in prostate 5alpha-reductase enzyme activity was observed between the treatment groups. Locomotor activity levels were higher in the phytoestrogen-rich vs the phytoestrogen-free animals during the course of the treatment interval. Plasma testosterone and androstenedione levels were significantly lower in the animals fed the phytoestrogen-rich diet compared with animals fed the phytoestrogen-free diet. However, there were no significant differences in plasma LH or estradiol levels between the diet groups. Testicular StAR levels were not significantly different between the phytoestrogen-rich vs the phytoestrogen-free fed animals. These results indicated that consumption of dietary phytoestrogens resulting in very high plasma isoflavone levels over a relatively short period can significantly alter body and prostate weight and plasma androgen hormone levels without affecting gonadotropin or testicular StAR levels. The findings of this study identify the biological actions of phytoestrogens on male reproductive endocrinology and provide insights into the protective effects these estrogen mimics exert in male reproductive disorders such as benign prostatic hyperplasia and prostate cancer.

Effects of dietary phytoestrogen on global myocardial ischemia-reperfusion injury in isolated female rat hearts.

We investigated the effects of phytoestrogen on global myocardial ischemia-reperfusion injury in five groups of female rats. A high-phytoestrogen group (HPE) was ovariectomized (Ovx) and fed a diet containing soybean protein and a high-isoflavone soy extract. Another Ovx group of rats was fed the same diet as the HPE group but treated with the estrogen receptor blocker ICI-182,780 (HPE + ICI). A third group of Ovx rats was fed a diet containing soybean protein alone (low-phytoestrogen content; LPE). A fourth Ovx group was fed a diet free of phytoestrogen (Ovx). The fifth group of rats was sham ovariectomized (sham). Hearts from all rats were subjected to 30 min of global, hypothermic (4 degrees C), cardioplegic ischemia and 120 min of normothermic (37 degrees C) reperfusion with oxygenated Krebs-Henseleit buffer. Compared with either the sham or the HPE group, the Ovx and HPE + ICI groups had significantly decreased first derivative of left ventricular pressure (dP/dt), coronary flow rate (CFR), nitrite production and mitochondrial respiratory function and significantly increased Ca2+ accumulation and myocardial histological and ultrastructural injury. The CFR of the LPE group was significantly different from that of either Ovx or HPE + ICI group but the dP/dt, nitrite production, Ca2+ accumulation, and mitochondrial function were not. Our results indicate that diets containing phytoestrogen extract play a cardioprotective role in global myocardial ischemia-reperfusion in female rats.

Animal models impacted by phytoestrogens in commercial chow: implications for pathways influenced by hormones.

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.

Maternal and perinatal brain aromatase: effects of dietary soy phytoestrogens.

Phytoestrogens are extensively investigated for their potential to prevent many hormone-dependent cancers and age-related diseases, however little is known about their effects in brain. Brain aromatase and plasma phytoestrogen levels were determined in Sprague-Dawley rats fed a phytoestrogen-rich diet during pregnancy/lactation. Ingested phytoestrogens cross the placenta and become concentrated in maternal milk as evident from high infantile plasma concentrations. Dietary phytoestrogens, however, do not alter brain aromatase during pregnancy/lactation or perinatal development.