Association between coronary artery vitamin D receptor expression and select systemic risks factors for coronary artery atherosclerosis.

OBJECTIVE: The aim of this study is to analyze the association between coronary artery vitamin D receptor (VDR) expression and systemic coronary artery atherosclerosis (CAA) risk factors. METHODS: Female cynomolgus monkeys (n = 39) consumed atherogenic diets containing the women's equivalent of 1000 IU/day of vitamin D3. After 32 months consuming the diets, each monkey underwent surgical menopause. After 32 postmenopausal months, CAA and VDR expression were quantified in the left anterior descending coronary artery. Plasma 25OHD3, lipid profiles and serum monocyte chemotactic protein-1 (MCP-1) were measured. RESULTS: In postmenopausal monkeys receiving atherogenic diets, serum MCP-1 was significantly elevated compared with baseline (482.2 ± 174.2 pg/ml vs. 349.1 ± 163.2 pg/ml, respectively; p < 0.001; d = 0.79) and at the start of menopause (363.4 ± 117.2 pg/ml; p < 0.001; d = 0.80). Coronary VDR expression was inversely correlated with serum MCP-1 (p = 0.042). Additionally, the change of postmenopausal MCP-1 (from baseline to necropsy) was significantly reduced in the group with higher, compared to below the median, VDR expression (p = 0.038). The combination of plasma 25OHD3 and total plasma cholesterol/high-density lipoprotein cholesterol was subsequently broken into low-risk, moderate-risk and high-risk groups; as the risk increased, the VDR quantity decreased (p = 0.04). CAA was not associated with various atherogenic diets. CONCLUSION: Coronary artery VDR expression was inversely correlated with markers of CAA risk and inflammation, including MCP-1, suggesting that systemic and perhaps local inflammation in the artery may be associated with reduced arterial VDR expression.

Impairment of ovarian function and associated health-related abnormalities are attributable to low social status in premenopausal monkeys and not mitigated by a high-isoflavone soy diet.

BACKGROUND: Psychological stress may impair premenopausal ovarian function and contribute to risk for chronic disease. Soy isoflavones may also influence ovarian function and affect health. Here, we report the effects of a psychological stressor (subordinate social status) and dietary soy on reproductive function and related health indices in female monkeys. We hypothesized that reproductive compromise and adverse health outcomes would be induced in subordinate when compared with dominant monkeys and be mitigated by exposure to soy. METHODS: Subjects were 95 adult cynomolgus monkeys (Macaca fascicularis) housed in social groups of five or six. Animals consumed a soy-free, animal protein-based diet during an 8-month Baseline phase and then, during a 32-month Treatment phase, consumed either the baseline diet or an identical diet that substituted high-isoflavone soy protein for animal protein. RESULTS: Across more than 1200 menstrual cycles, subordinate monkeys consistently exhibited ovarian impairment [increased cycle length (P < 0.02) and variability (P < 0.02) and reduced levels of progesterone (P < 0.04) and estradiol (P < 0.04)]. Subordinate status was confirmed behaviorally and was associated with elevated cortisol (P < 0.04) and relative osteopenia (P < 0.05). Consumption of the soy diet had no significant effects. CONCLUSIONS: (i) Psychological stress adversely affects ovarian function and related health indices in a well-accepted animal model of women's health; (ii) Similar effects may extend to women experiencing reproductive impairment of psychogenic origin; (iii) soy protein and isoflavones neither exacerbate nor mitigate the effects of an adverse psychosocial environment; and (iv) this study was limited by an inability to investigate the genetic and developmental determinants of social status.

Impact of soy supplementation on sex steroids and vascular inflammation markers in postmenopausal women using tibolone: role of equol production capability.

OBJECTIVES: Tibolone is often taken concurrently with soy. Tibolone, soy and equol-producing capacity each affect vascular health, whereas their concomitant effects are unknown. We studied the effects of soy on sex steroids and vascular inflammation markers in long-term tibolone users. METHODS: Postmenopausal women (n = 110) on tibolone were screened with a soy challenge to find 20 equol producers and 20 non-producers. All women were treated for 8 weeks in a cross-over trial with soy (52 g of soy protein containing 112 mg of isoflavones) or placebo. Serum estrone, 17beta-estradiol, testosterone, androstenedione, dehydroepiandrosterone sulfate (DHEAS), sex hormone binding globulin (SHBG), C-reactive protein (CRP), vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1), and platelet-selectin (P-selectin) were assessed. RESULTS: Soy decreased (7.1%) the estrone level, significantly (12.5%) only in equol producers (from 80.2 +/- 10.8 to 70.3 +/- 7.0 pmol/l; p = 0.04). Testosterone was reduced (15.5%; from 586 +/- 62.6 to 495 +/- 50.1 pmol/l, p = 0.02) during soy treatment, and more markedly in equol producers than non-producers (22.1% vs. 10.0%). No changes appeared in SHBG, CRP or ICAM-1, but VCAM-1 increased (9.2%) and P-selectin decreased (10.3%) during soy treatment. CONCLUSIONS: Soy modified the concentrations of estrone, testosterone and some endothelial markers. Equol production enforced these effects. Soy supplementation may be clinically significant in tibolone users.

Individual differences in equol production capability modulate blood pressure in tibolone-treated postmenopausal women: lack of effect of soy supplementation.

OBJECTIVES: Equol, a gut bacterial metabolite of the isoflavone daidzein, has been associated with beneficial health effects. Recent studies indicate that women with intestinal capacity to convert daidzein to equol also have the capacity to alter steroid metabolism and bioavailability of estrogens. METHODS: We evaluated whether individual equol production capability, while not consuming soy supplement, was associated with lower blood pressure in postmenopausal women using tibolone. In addition, in a randomized, placebo-controlled, cross-over trial we assessed the effect of soy supplementation on blood pressure in both equol-producing (n = 20) and non-equol-producing (n = 20) women using tibolone. Blood pressure was recorded with a validated oscillometric technique. RESULTS: The circulating equol levels rose 20-fold in the equol producers and 1.9-fold in the non-equol producers. At baseline, systolic blood pressure (129.9 +/- 2.6 vs. 138.5 +/- 3.1 mmHg, p = 0.02), diastolic blood pressure (72.2 +/- 1.5 vs. 76.6 +/- 1.3 mmHg, p = 0.01) and mean arterial blood pressure (93.5 +/- 1.7 vs. 99.9 +/- 1.8 mmHg, p = 0.007) were lower in equol producers compared to non-equol producers. Soy supplementation had no effect on blood pressure in either group, whereas the baseline differences persisted. CONCLUSIONS: Postmenopausal women using tibolone characterized as equol producers had lower blood pressure compared to non-equol producers. Soy supplementation for 2 months had no blood pressure-lowering effect.

Multisystem evaluations of the long-term effects of tibolone on postmenopausal monkeys.

This long-term study (2 years) was designed to compare the effects of tibolone (LoTib at 0.05 mg/kg and HiTib at 0.2 mg/kg) with those of conjugated equine oestrogens (CEE) alone (0.042 mg/kg) and CEE continuously combined with medroxyprogesterone acetate (MPA) (0.167 mg/kg) on coronary artery atherosclerosis, bone, mammary gland and uterus in ovariectomised cynomolgus monkeys fed a moderately atherogenic diet. Despite reductions in plasma concentrations of high density lipoprotein cholesterol in tibolone-treated monkeys, there was no exacerbation of coronary artery atherosclerosis. Tibolone was equivalent to, or slightly better than, CEE and CEE + MPA in protecting against postmenopausal bone loss and loss of bone strength. Tibolone also resulted in less stimulation of breast and endometrial tissue compared with CEE and CEE + MPA. In conclusion, the results suggest that tibolone is a cardiovascular-safe treatment that is effective for the prevention of osteoporosis and that may have advantages over CEE or CEE + MPA with regard to endometrial and breast safety.

Effects of long-term hormone replacement and of tibolone on choline acetyltransferase and acetylcholinesterase activities in the brains of ovariectomized, cynomologus monkeys.

We examined long-term effects of low and high doses of tibolone, conjugated equine estrogens, and conjugated equine estrogens plus medroxyprogesterone acetate on choline acetyltransferase and acetylcholinesterase activities within different regions of the brain in cynomologus monkeys. All treatments were administered for 2 years. None of the treatments produced significant increases in either choline acetyltransferase or acetylcholinesterase in any of eight brain regions analyzed. In contrast, treatment with conjugated equine estrogens plus medroxyprogesterone acetate, but not conjugated equine estrogens alone, produced significant reductions in both choline acetyltransferase and acetylcholinesterase in the medial septum/diagonal band of Broca compared with untreated controls. Treatment with tibolone also resulted in significant reductions in both choline acetyltransferase and acetylcholinesterase in the medial septum/diagonal band of Broca, and this effect was dose-related. These findings are the first to report the effects of long-term therapies used by postmenopausal women on cholinergic measures in the primate brain. The findings are consistent with recent reports in rats, and suggest that any positive effects of long-term estrogen or hormone replacement therapy on cognitive processes are probably not due to significant effects on choline acetyltransferase or acetylcholinesterase activities.

Premenopausal social status and hormone exposure predict postmenopausal atherosclerosis in female monkeys.

OBJECTIVE: To determine whether premenopausal social subordination in female monkeys predicts postmenopausal atherosclerosis, and whether any such effect is altered by chronic exposure to contraceptive steroids or postmenopausal hormone replacement. METHODS: One hundred seventy-seven (177) premenopausal cynomolgus monkeys (Macaca fascicularis) housed in social groups of five or six were fed an atherogenic diet that, for half of the animals, also contained an oral contraceptive (OC). Individuals were judged socially dominant or subordinate based on behavioral observations. After 26 months animals were oophorectomized, biopsied for iliac atherosclerosis, and for the next 36 months were fed one of three atherogenic diets containing soy protein: 1) phytoestrogen-free; 2) phytoestrogens intact; and 3) phytoestrogen-free plus conjugated equine estrogens. Plasma lipids and menstrual cyclicity were also assessed. Finally, all animals were necropsied and the extent of atherosclerosis measured in the coronary and iliac arteries. RESULTS: The interaction of premenopausal social status and OC exposure predicted postmenopausal coronary artery atherosclerosis (P =.02). Subordinate animals not receiving OCs developed twice the coronary atherosclerosis of similarly untreated dominants (P <.01), an outcome mitigated by premenopausal OC exposure (P <.01). These effects occurred across postmenopausal treatment groups and independent of variation in plasma lipids. The same associations were observed in the iliac arteries, and, to a similar extent, both pre- and post-menopausally. Hormone data suggest that untreated premenopausal subordinates may have been estrogen deficient. CONCLUSION: Premenopausal social subordination exacerbates postmenopausal atherosclerosis, an effect possibly mediated by estrogen deficiency and shown here to be prevented by premenopausal OC exposure. These results occur irrespective of postmenopausal treatment.

A comparison of tibolone and conjugated equine estrogens effects on coronary artery atherosclerosis and bone density of postmenopausal monkeys.

This study compared the effects of tibolone, a tissue-specific compound for the treatment of climacteric symptoms and the prevention of osteoporosis, with those of conjugated equine estrogens (CEE) with and without medroxyprogesterone (MPA) on bone mineral density and coronary atherosclerosis (CAA) of postmenopausal cynomolgus monkeys. The groups were tibolone [two doses were used, 0.05 mg/kg (LoTib) and 0.2 mg/kg (HiTib)], CEE (0.042 mg/kg), CEE (0.042 mg/kg) plus MPA (0.167 mg/kg given continuously), and a control group given no treatment for 2 yr. Compared with no treatment, bone mineral density was higher by 6.3% (P = 0.0004) in the LoTib group and by 9.5% (P = 0.02) in the HiTib group compared with 4.3% (P = 0.12) for CEE and 4.5% (P = 0.10) for CEE+MPA. Plasma high density lipoprotein cholesterol was reduced by 49% with HiTib and by 34% with LoTib. There were no differences in CAA between control and HiTib (P = 0.60) or LoTib (P = 0.58). CEE and CEE+MPA both reduced CAA by about 62% (CEE vs. control, P = 0.02; CEE+MPA vs. control, P = 0.01). Despite adverse effects of tibolone on plasma lipoprotein concentrations, there was no increase in CAA, suggesting that tibolone is a cardiovascular-safe treatment for climacteric symptoms and the prevention of osteoporosis.

A comparison of oral micronized estradiol with soy phytoestrogen effects on tail skin temperatures of ovariectomized rats.

OBJECTIVE: Whether phytoestrogen-containing soy supplements have beneficial effects on hot flashes of postmenopausal women and how those effects, if any, compare to estrogen replacement therapy has been uncertain. It is possible that the uncertainty is due to the low doses of soy isoflavones (30-60 mg per day) used in the studies. We used ovariectomized retired breeder rats and a higher dose of soy phytoestrogens to approach these uncertainties experimentally. DESIGN: The treatment groups were as follows: (1) Control group fed a casein/lactalbumin-based diet; (2) Soy(-) group fed alcohol-washed soy protein isolate with the phytoestrogens extracted; (3) Soy(+) group fed phytoestrogen-containing soy protein (equivalent to a woman's dose of 144 mg isoflavones per day)--a dose two to three times higher than that in most studies with women; and (4) E2 group fed oral micronized estradiol (E2) at a dose equivalent to a woman's dose of 1 mg per day. A temperature-transponder was taped to the surface of the tail to measure temperature. Tail skin temperature was significantly increased within a week after ovariectomy. The animals were pair-fed during the last 21 days of treatment for daily temperature measurement. RESULTS: Soy(-) had no effect on skin temperature. E2 had a large effect on skin temperature (about 1.4 degrees C reduction from Control). Soy(+) was intermediate between the E2 treatment and no treatment (about 0.8 degrees C reduction from Control). CONCLUSIONS: Soy phytoestrogens have a modest effect on average skin temperatures, being about half that of E2, even at high doses in the rat model.

SERMs and cardiovascular disease in women. How do these agents affect risk?

The beneficial effects of SERMs, specifically tamoxifen in the treatment and prevention of breast cancer and raloxifene in the prevention of osteoporosis, are well established. In addition, numerous groups of investigators have reported that these agents have a positive impact on cardiovascular health, possibly as a result of their cholesterol-lowering and anticoagulation actions. Although studies clearly showed that tamoxifen therapy improved the levels of some types of lipids, the changes did not appear to translate into a decreased risk of cardiovascular disease. However, the risk of thromboembolic events (as well as endometrial cancer) was increased with the use of tamoxifen, which is often prescribed for breast cancer prevention in healthy women. Similarly, raloxifene treatment had modest positive effects on cardiovascular risk factors but was associated with an increased risk of thromboembolism. When viewed as a whole, study results dictate that the benefits of SERM use for the prevention of cardiovascular disease be carefully weighed against the potential risks.

Inhibition of postmenopausal atherosclerosis progression: a comparison of the effects of conjugated equine estrogens and soy phytoestrogens.

Experimental evidence was sought concerning whether soy phytoestrogens (SPEs) inhibit postmenopausal atherosclerosis progression/extent and, if so, their effectiveness relative to traditional estrogen replacement therapy. Premenopausal cynomolgus monkeys were fed a moderately atherogenic diet (26 months) to induce atherosclerosis. After ovariectomy, the moderately atherogenic diet was continued, and they were treated (36 months) with a control diet (soy protein depleted of SPEs), a diet containing SPEs in soy protein isolate, or a diet containing SPE-depleted soy protein with conjugated equine estrogens (CEE; Premarin) added. SPE effects on plasma lipids were better than those of CEE (higher high density lipoprotein cholesterol and no increase in triglyceride). Relative to the control group, CEE treatment inhibited (P = 0.0001), and SPE treatment partially inhibited (P = 0.10) the progression of atherosclerosis (common iliac artery atherosclerosis before and after treatment). CEE-treated monkeys had much less coronary artery atherosclerosis than the controls (P = 0.0002), whereas SPE-treated monkeys were intermediate in lesion extent between the controls and the CEE-treated animals (P = 0.02). Both CEE and SPE significantly reduced the extent of common carotid and internal carotid artery atherosclerosis, and the two treatment groups were not significantly different.

Soy phytoestrogens improve radial arm maze performance in ovariectomized retired breeder rats and do not attenuate benefits of 17beta-estradiol treatment.

OBJECTIVE: Soy phytoestrogens (SPEs) seem to have beneficial effects on the cardiovascular system with no adverse effects on the breast and uterus. Our objective was to examine the effects of oral estradiol alone, soy protein with phytoestrogens alone, and combinations of estradiol and SPEs on working memory of ovariectomized retired breeder female rats using the radial arm maze test. DESIGN: Eighty-four bilaterally ovariectomized retired breeder female rats were randomized into 12 groups to examine the effects of chronic treatment (10 months) with oral micronized estradiol (0, 0.5, 1, and 2 mg/1,800 Cal), SPEs (0, 72, and 144 mg/1,800 Cal), and all combinations of these doses of estradiol and SPEs on working memory. RESULTS: Oral administration of estradiol or SPEs resulted in a dose-dependent improvement in the performance of the radial arm maze tests. In addition, at each of the three doses of oral micronized estradiol tested, the performance of the radial arm tests was not significantly different in the presence or absence of SPEs. CONCLUSIONS: Our data suggest that SPEs may function as estrogen agonists in improving working memory in the ovariectomized retired breeder female rats and that SPEs do not antagonize the beneficial effects of estradiol on the working memory of these rats. No additional benefits on the radial arm maze test performance were observed with the tested combinations of estradiol and SPEs.

Effect of estradiol and soy phytoestrogens on choline acetyltransferase and nerve growth factor mRNAs in the frontal cortex and hippocampus of female rats.

We report here the effects of oral micronized estradiol and soy phytoestrogens on uterine weight, choline acetyltransferase (ChAT) and nerve growth factor (NGF) mRNAs in the frontal cortex and hippocampus of ovariectomized young and retired breeder rats. Within each age category, 15 bilaterally ovariectomized rats were randomized equally into three groups: control (OVX), estradiol (E2), and soy phytoestrogens (SBE). The OVX rats were fed a casein/lactalbumin-based control diet; the E2 rats were fed with the control diet with added estradiol; and the SBE rats were fed with the control diet with added soy phytoestrogens. After 8 weeks of treatment, blood, uteri, frontal cortex, and hippocampus were collected at necropsy. Results showed that the uterine weights and serum estradiol concentrations were significantly higher in the E2 group compared with those in the OVX and SBE groups. In the hippocampus of young rats, E2 treatment resulted in significantly higher NGF mRNA levels than no treatment (OVX), and NGF mRNA levels in the SBE group were intermediate between the E2 and OVX groups. ChAT mRNA levels were significantly higher in the frontal cortex of E2 and SBE-treated retired breeder rats compared to OVX retired breeder rats. There were no differences among treatment groups for ChAT mRNA levels in the frontal cortex of young rats and in the hippocampus of both young and retired breeder rats. Our data suggest that soy phytoestrogens may function as estrogen agonists in regulating ChAT and NGF mRNAs in the brain of female rats.

Effects of hormone replacement therapy and social stress on body fat distribution in surgically postmenopausal monkeys.

OBJECTIVE: To investigate the effects of hormone replacement therapy (HRT) and social stress on body fat distribution in an animal model of women's health, the female cynomolgus macaque (Macaca fascicularis). DESIGN/SUBJECTS: Adult female cynomolgus monkeys were ovariectomized and fed an atherogenic diet for two years while housed in social groups of 3-8 monkeys each. Animals were then fed a lipid-lowering diet and randomized into four experimental groups: a baseline group which was necropsied immediately and not included in the study reported here, 26 females fed diet only (CONTROL), 22 females fed diet plus conjugated equine estrogens (CEE), and 21 females fed the diet plus CEE and medroxyprogesterone acetate (CEE + MPA). Treatment lasted 30 months. MEASUREMENTS: During the last nine months of treatment, social status was determined three times at three month intervals. At the end of the study, whole body obesity and fat distribution patterns were determined using anthropometry and computerized tomography (CT). RESULTS: The addition of a progestin to the estrogen replacement regimen administered to surgically postmenopausal monkeys, increased all anthropometric and CT measures of obesity except intra-abdominal fat. HRT had no effect on patterns of fat distribution. Socially-dominant, ovariectomized females were more obese than subordinates using both anthropometric and CT measurements of whole body obesity. Dominant females were more likely to have their fat deposited centrally as measured anthropometrically. However, CT measures revealed a trend for dominants to preferentially deposit fat in the subcutaneous abdominal depot in contrast to subordinates who deposited fat in the intra-abdominal depot. CONCLUSIONS: The results of this study suggest that progestins, when administered in combination with estrogens, may increase fat deposition, particularly in subcutaneous depots. In addition, the social stress experienced by subordinate monkeys, may have mild effects on fat deposition patterns, even after removal of ovarian function as a factor. These observations may have implications for treatment recommendations in postmenopausal women. Lastly, CT may measure different characteristics of fat distribution than skinfolds and circumferences.

Evidence for up-regulation of brain-derived neurotrophic factor mRNA by soy phytoestrogens in the frontal cortex of retired breeder female rats.

In this study, we examined the effects of oral micronized estradiol and soy phytoestrogens on the mRNA levels of brain-derived neurotrophic factor (BDNF) in the frontal cortex of ovariectomized retired breeder female rats. Fifteen ovariectomized rats were randomized into three groups and fed either a soy-free control diet (OVX), a control diet with added soybean estrogens (SBE), or a control diet with added estradiol (E2) for 8 weeks. Frontal cortex samples were collected for RNA isolation. Northern and Phosphorlmager analyses were used to determine the relative levels of BDNF mRNA. Both estradiol and soy phytoestrogens significantly increased the mRNA levels of BDNF compared to ovariectomized controls, suggesting that soy phytoestrogens may act as estrogen agonists in regulating BDNF mRNA in the frontal cortex of retired breeder female rats.

Progestogens and cardiovascular disease. A critical review.

Although progestational hormones are clearly beneficial in preventing estrogen-induced endometrial hyperplasia and cancer, their effect on other areas is far less clear. Of particular interest is the attenuating effect medroxyprogesterone acetate (MPA) has on the cardiovascular benefits of postmenopausal estrogen treatment. MPA reduces the dilatory effect of estrogens on coronary arteries, increases the progression of coronary artery atherosclerosis, accelerates low-density lipoprotein uptake in plaque, increases the thrombogenic potential of atherosclerotic plaques and promotes insulin resistance and its consequent hyperglycemia. These effects may be largely limited to MPA and not shared with other progestogens.

Effects of soy isoflavones on atherosclerosis: potential mechanisms.

It has long been recognized that coronary heart disease rates are lower in Japan, where soy consumption is common, than in Western countries. In experimental studies, atherosclerosis was reduced in animals fed diets containing soy protein compared with those fed diets with animal protein. Recently, several lines of evidence have suggested that the components of soy protein that lower lipid concentrations are extractable by alcohol (eg, the isoflavones genistein and daidzein). We recently evaluated the relative effect of the soy protein versus the alcohol-extractable components of soy on cardiovascular disease and its risk factors. Young male and female cynomolgus monkeys were fed diets that contained either 1) casein-lactalbumin as the source of protein (casein), 2) soy protein isolate from which the isoflavones were alcohol extracted (SPI-), or 3) isoflavone-intact soy protein (SPI+). The SPI+ group had significant improvements in LDL cholesterol and HDL cholesterol. Only HDL cholesterol was significantly improved in the SPI- group males compared with the casein group. The casein group had the most atherosclerosis, the SPI+ group had the least, and the SPI- group was intermediate but did not differ significantly from the casein group. Potential mechanisms by which soy isoflavones might prevent atherosclerosis include a beneficial effect on plasma lipid concentrations, antioxidant effects, antiproliferative and antimigratory effects on smooth muscle cells, effects on thrombus formation, and maintenance of normal vascular reactivity.