S-equol Exerts Estradiol-Like Anorectic Action with Minimal Stimulation of Estrogen Receptor-α in Ovariectomized Rats.

Chronic estrogen replacement in ovariectomized rats attenuates food intake and enhances c-Fos expression in the suprachiasmatic nucleus (SCN), specifically during the light phase. S-equol, a metabolite of daidzein, has a strong affinity for estrogen receptor (ER)-ß and exerts estrogenic activity. The purpose of the present study was to elucidate whether S-equol exerts an estrogen-like anorectic effect by modifying the regulation of the circadian feeding rhythm in ovariectomized rats. Ovariectomized female Wistar rats were divided into an estradiol (E2)-replaced group and cholesterol (vehicle; Veh)-treated group. These animals were fed either a standard diet or an S-equol-containing diet for 13 days. Then, the brain, uterus, and pituitary gland were collected along with blood samples. In the rats fed the standard diet, E2 replacement attenuated food intake (P < 0.001) and enhanced c-Fos expression in the SCN (P < 0.01) during the light phase. Dietary S-equol supplementation reduced food intake (P < 0.01) and increased c-Fos expression in the SCN (P < 0.01) in the Veh-treated rats but not in the E2-replaced rats during the light phase. Dietary S-equol did not alter ER-α expression in the medial preoptic area or the arcuate nucleus, nor did dietary S-equol affect pituitary gland weight or endometrial epithelial layer thickness. By contrast, E2 replacement not only markedly decreased ER-α expression in these brain areas (P < 0.001) but also increased both the pituitary gland weight (P < 0.001) and the endometrial epithelial layer thickness (P < 0.001). Thus, dietary S-equol acts as an anorectic by modifying the diurnal feeding pattern in a manner similar to E2 in ovariectomized rats; however, the mechanism of action is not likely to be mediated by ER-α. The data suggest a possibility that dietary S-equol could be an alternative to hormone replacement therapy for the prevention of hyperphagia and obesity with a lower risk of adverse effects induced by ER-α stimulation.

Relation between premenstrual syndrome and equol-production status.

AIM: Consumption of isoflavones, which are predominantly derived from soybeans, reduces the risk of estrogen-related diseases, such as menopausal symptoms, breast cancer, osteoporosis, and cardiovascular disease. Equol is more bioavailable than other soy isoflavones, and equol producers are believed to benefit to a greater extent. This study was conducted to evaluate the relation between premenstrual syndrome (PMS) and equol-production status in Japanese reproductive-age women. METHODS: This was a cross-sectional, observational study. The study included 144 Japanese women aged 20-45 years. PMS patients (n = 46) were recruited at three obstetrics and gynecology clinics. Control group women (n = 98) who were not receiving therapy for PMS were recruited from the local area by advertisement. The participants' equol-production status was determined using urine samples collected after a soy challenge test. RESULTS: The prevalence of equol producers was 41.8% in the control group and 23.9% in the patient group (P = 0.042). Using univariate analysis, significant risk factors for equol non-producers were being a PMS patient and being younger. In multivariate analysis with a step-wise model, being a PMS patient (odds ratio, 2.342; 95% confidence interval, 1.021-5.698) was shown to be a significant risk factor for being an equol non-producer. CONCLUSION: This study showed a relation between PMS and equol-production status in Japanese women.

A cross-sectional study of equol producer status and self-reported vasomotor symptoms.

OBJECTIVE: This study aims to evaluate the associations of vasomotor symptom (VMS) frequency, bother, and severity with equol producer status and dietary daidzein intake. METHODS: This is an observational study. This study included women aged 45 to 55 years, in postmenopause or in the menopausal transition, who had soy food intake of three or more servings per week. Exclusion criteria included severe concurrent disease, pregnancy or planned pregnancy, and current use of oral or transdermal hormones or selective estrogen receptor modulators. After screening, 375 participants completed a 3-day VMS diary and a 24-hour urine collection. Women with a urine daidzein or genistein concentration of 100 ng/mL or higher were included. We evaluated the association of VMS--dichotomized as lower than or equal to versus higher than the mean number of VMS per day (<2.33, ≥ 2.33)--with quartiles of daidzein intake. RESULTS: Overall, 129 (35%) of 365 women were equol producers. The mean (SD) urinary equol excretion was 0.67 (1.57) mg/day (50th percentile, 0 mg/d; 95th percentile, 4.12 mg/d). Among equol producers, the mean (SD) urinary equol excretion was 1.91 (2.15) mg/day (50th percentile, 1.09 mg/d; 95th percentile, 6.27 mg/d). Among equol producers, compared with those in the lowest quartile of dietary daidzein intake (mean, 4.9 mg/d), those in the highest quartile (mean, 28.5 mg/d) were 76% less likely to have VMS higher than the mean number of VMS (odds ratio, 0.24; 95% CI, 0.07-0.83; trend test across all daidzein levels, P = 0.06). Among equol nonproducers, there were no associations between daidzein intake and VMS frequency. There were no differences in VMS bother or severity among equol producers or nonproducers by dietary daidzein level. CONCLUSIONS: Among equol producers, higher equol availability attributable to higher soy consumption contributes to decreased VMS.

Estrogen stimuli promote osteoblastic differentiation via the subtilisin-like proprotein convertase PACE4 in MC3T3-E1 cells.

Estrogenic compounds include endogenous estrogens such as estradiol as well as soybean isoflavones, such as daidzein and its metabolite equol, which are known phytoestrogens that prevent osteoporosis in postmenopausal women. Indeed, mineralization of MC3T3-E1 cells, a murine osteoblastic cell line, was significantly decreased in medium containing fetal bovine serum treated with charcoal-dextran to deplete endogenous estrogens, but estradiol and these soybean isoflavones dose-dependently restored the differentiation of MC3T3-E1 cells; equol was tenfold more effective than daidzein. These differentiation-promoting effects were inhibited by the addition of fulvestrant, which is a selective downregulator of estrogen receptors. Analysis of the expression pattern of bone-related genes by reverse transcription PCR (RT-PCR)/quantitative real-time PCR (qRT-PCR), which focused on responsiveness to the estrogen stimuli, revealed that the transcription of PACE4, a subtilisin-like proprotein convertase, was tightly linked with the differentiation of MC3T3-E1 cells induced by estrogen stimuli. Moreover, treatment with RNAi of PACE4 in MC3T3-E1 cells resulted in a drastic decrease of mineralization in the presence of estrogen stimuli. These results strongly suggest that PACE4 participates in bone formation at least in osteoblast differentiation, and estrogen receptor-mediated stimuli induce osteoblast differentiation through the upregulation of PACE4 expression.

The effects of natural S-equol supplementation on skin aging in postmenopausal women: a pilot randomized placebo-controlled trial.

OBJECTIVE: The aim of this study was to investigate the effects of the natural S-equol supplement on skin aging in equol-nonproducing Japanese postmenopausal women. METHODS: A randomized, double-blind, placebo-controlled trial examined the use of the natural S-equol supplement for 12 weeks in 101 postmenopausal Japanese women who were equol nonproducers. They were randomly assigned to one of three groups: placebo (n = 34), 10 mg S-equol/day (EQL10; n = 34), or 30 mg S-equol/day (EQL30; n = 33). Skin parameters of crow's-feet wrinkles (area and depth), hydration, transepidermal water loss, and elasticity were measured at baseline and at monthly intervals during treatment. Vaginal cytology, endometrial thickness, and mammography were performed before and after treatment. Serum hormone concentrations were measured at the same time as skin parameters. RESULTS: The EQL10 and EQL30 groups showed significant reductions in wrinkle area compared with the placebo group (P < 0.05). There was a significant difference in wrinkle depth between the placebo group and the EQL30 group (P < 0.05). Other skin parameters did not show significant differences after the treatment in any group. There were no abnormal results in hormone status or gynecological examinations. CONCLUSIONS: Our data suggest that natural S-equol supplementation (EQL10 and EQL30) may have a beneficial effect on crow's-feet wrinkles in postmenopausal women without serious adverse events.

Effects of S-equol and natural S-equol supplement (SE5-OH) on the growth of MCF-7 in vitro and as tumors implanted into ovariectomized athymic mice.

Hormone replacement therapy (HRT) for treatment of menopausal symptoms is controversial because of reported breast cancer resulting from estrogen treatment and consequent estrogenic stimulation. S-equol, a natural metabolite of the soy isoflavone daidzein produced by intestinal bacteria, has been shown to ameliorate menopausal symptoms, with relatively low concomitant estrogenic receptor stimulation. Although synthesis of equol produces the racemate, the S-isomer may be produced in commercial amounts by bacterial fermentation of soy germ, during the production of the supplement SE5-OH. This study aims to investigate the effects of S-equol and SE5-OH on the growth of MCF-7 in vitro and in vivo. In vitro, purified S-equol, and the isoflavonoid mixture present in SE5-OH stimulated estrogenic transcriptional activity and proliferation of MCF-7-E10 cells, similar to that observed for genistein (another soy isoflavone), but at concentrations from 10(4)-fold to 10(6)-fold higher than seen with 17ß-estradiol (E2). Ovariectomized (OVX) mice implanted with MCF-7-E10 cells were fed diets containing 250 or 500 ppm of purified S-equol, isoflavonoid mixture, or genistein. There were no significant differences in tumor growth between the treatment groups and control group. These results suggest that S-equol and natural S-equol in the supplement (SE5-OH), do not promote the progression of breast cancer.

S-equol and the fermented soy product SE5-OH containing S-equol similarly decrease ovariectomy-induced increase in rat tail skin temperature in an animal model of hot flushes.

OBJECTIVE: The aim of this study was to compare the effect of SE5-OH, a fermented soy product containing S-equol, with purified S-equol on hot flushes in an ovariectomized rat model. METHODS: Eleven-week-old female Sprague-Dawley rats were assigned to either the sham group (vehicle; n = 30) or one of four ovariectomized groups: control (vehicle; n = 30), conjugated equine estrogens (CEE; 6.0 mg kg(-1) d(-1) CEE; n = 10), SE5-OH (2,000 mg kg(-1) d(-1) SE5-OH containing 11.7 mg kg(-1) d(-1) as S-equol; n = 30), and S-equol (11.7 mg kg(-1) d(_1) S-equol; n = 30). Three days after sham operation or ovariectomy, animals were treated once daily for 38 days. Tail skin temperature (TST) was assessed on days 21, 28, and 35 after surgery. Plasma estradiol and follicle-stimulating hormone levels and uterine weight and uteri histology were evaluated at the end of treatment. RESULTS: The rise in TST resulting from ovariectomy was inhibited by CEE, SE5-OH, and S-equol. Compared with the control, TST was decreased by 68.9% and 86.2% in SE5-OH group on days 21 and 28, respectively (P = 0.014, 0.020), and by 60.1% and 89.1% in S-equol group, respectively (P = 0.038, 0.016). Unlike in the CEE group, plasma estradiol and follicle-stimulating hormone levels, uterine weight, epithelial height, stromal expansion, and myometrial thickness were not affected in SE5-OH and S-equol groups. CONCLUSIONS: The results of this animal model of hot flushes suggest that S-equol is one of the primary components of SE5-OH and that both SE5-OH and S-equol represent promising alternatives for the treatment of menopausal symptoms. Clinical research is needed to confirm these findings.

[Prevalence of equol producer phenotype and its relations to lifestyle factors and dietary intakes among lifestyle factors and dietary intakes among healthy Chinese adults in Beijing].

OBJECTIVE: To investigate the prevalence of equol producers and the physiological range of urinary equol excretion, and also to evaluate relations between equol phenotype and lifestyle among Chinese adults in Beijing. METHODS: 100 male and 100 female adults participated in a cross-sectional study and provided twice 1d urine samples on regular diet and after 3d soy isoflavone challenge respectively. A health and demographics questionnaire, and 2d food record were completed before the urine collections. Isoflavones and their metabolites in urine were measured to determine equol phenotype by HPLC. RESULTS: The physiological range of 24h urinary equol excretion was 0-76.56 micromol/24h, and the percentage of the equol producer phenotype was 26.8% on regular diet and 60.4% after soy isofavone challenge, respectively. There was no indication that habitual consumption of soy foods is associated with the equol producer phenotype. The correlations of isoflavone intake from 2d food record with those from urinary isoflavone levels were 0.58 for total isoflavones, 0.49 for daidzein, 0.56 for genistein, and 0.50 for glycitein (P < 0.01). CONCLUSION: About one fourth of Chinese adults in Beijing were detected equol excretion in urine under the usually lifestyle. However, equol_producing potential was higher.

Dietary and lifestyle correlates of urinary excretion status of equol in Japanese women.

The isoflavone metabolite equol has been identified in urine or blood samples in some but not all humans. In this cross-sectional study, we examined the association between lifestyle, including diet, and the urinary excretion of equol. Study subjects were 419 Japanese women who were recruited from a breast cancer screening center. Each woman responded to a self-administered questionnaire seeking information about health and lifestyle factors. Diet was assessed by a validated semiquantitative food frequency questionnaire. Urinary isoflavones were measured using spot urine samples. Equol was detected in the urine of 84 (20.0%) women. After controlling for covariates, it was found that dairy product intake was significantly lower in those who excreted detectable equol levels in urine than in those who did not. Because equol is derived from daidzein, individuals with low intake of daidzein may produce undetectable levels of equol. To account for this, the study subjects were restricted to 163 women with urinary daidzein levels of 10 nmol/mg creatinine or higher. The association of equol excretion with dairy product intake remained significant. Demographic factors, smoking status, and menstrual and reproductive factors were unrelated to equol excretion. These data suggest that dairy product intake may be associated with the production of equol.

Associations among maternal soy intake, isoflavone levels in urine and blood samples, and maternal and umbilical hormone concentrations (Japan).

OBJECTIVES: In utero exposure to high levels of endogenous estrogens has been hypothesized to increase breast cancer risk in later life. A high intake of soy has been suggested to protect against breast cancer. We examined the hypothesis that maternal soy intake may be inversely associated with pregnancy hormone levels. METHODS: The concentrations of hormones (estradiol, estriol, and testosterone) and isoflavones (genistein, deidzein, and equol) were measured in the maternal urine and serum, and umbilical cord blood of 194 women during pregnancy and at delivery. Soy intake during pregnancy was assessed by 5-day diet records at approximately the 29th week of pregnancy. RESULTS: High correlations were observed for isoflavone levels between maternal samples and umbilical cord blood, indicating that isoflavone can be transferred from the maternal to the fetal compartment. None of the hormones measured in umbilical cord blood was significantly associated with any of the isoflavones measured. There were a few significant associations between maternal hormone levels and isoflavone measures during pregnancy, but their patterns of associations varied by gestational week and differed depending on whether isoflavone exposure was measured by diet records, urine or serum. CONCLUSION: Our data contain no strong evidence showing that soy intake affects hormone levels during pregnancy.