Plant-derived polyphenols have been shown to influence bone turnover and bone properties in the estrogen-depleted state. We used a crossover design in ovariectomized rats (n = 16 rats for each diet) to investigate the effect of supplementation of two doses each of blueberry, plum, grape, grape seed extract, and resveratrol on bone. We tested the aglycon and glucoside forms of genistein to quantify differences in efficacy on bone calcium retention. Rats were given an intravenous dose of 45Ca to prelabel bone, and bone calcium retention was assessed by urinary excretion of 45Ca:Ca ratio during an intervention period compared with nonintervention. Genistein aglycon increased bone calcium retention significantly (p<0.05) more than the glucoside (22% vs 13%, respectively). Plum extract (0.45% w/w total dietary polyphenols) and resveratrol (0.2% w/w total dietary polyphenols) were also effective, increasing bone calcium retention by 20% (p=0.0153) and 14% (p=0.0012), respectively. Several polyphenolic-rich diets improved bone calcium retention.
Mexican Americans are an understudied ethnic group for determinants of bone health, although the risk of age-related osteoporosis is high in this rapidly growing sector of the U.S. population. Thus, the objective of the present study was to establish the dietary calcium requirements for bone health in Mexican-American adolescents by measuring calcium retention calculated from balance in response to a range of dietary calcium intakes and to determine predictors of skeletal calcium retention. Adolescents aged 12-15 y were studied twice on paired calcium intakes ranging from 600 to 2300 mg/d using randomized-order, crossover 3-wk balance studies. Skeletal calcium retention was calculated as dietary calcium intake minus calcium excreted in feces and urine over the last 2 wk of balance. A linear model was developed to explain the variation in calcium retention. Boys (n = 20) were taller and had higher lean mass, usual dietary calcium intake, bone mineral content, and serum alkaline phosphatase compared with girls, whereas girls (n = 20) had higher Tanner scores and greater fat mass. Calcium retention increased with calcium intake (P < 0.0001) and did not differ by sex (P = 0.66). In boys and girls considered together, calcium intake explained 33% of the variation in calcium retention. Serum alkaline phosphatase explained an additional 11% of the variation in calcium retention. Other variables measured, including the urine N-telopeptide of type I collagen/creatinine ratio, Tanner score, serum parathyroid hormone and 25-hydroxyvitamin D, weight, height, and body mass index, did not contribute to the variance in calcium retention. In adolescence, calcium retention in both Mexican-American boys and girls was higher than determined previously in adolescent nonHispanic white girls. This trial was registered at clinicaltrials.gov as NCT01277185.
Soluble maize fibre (SCF) has been found to significantly improve bone mineral density and strength in growing rats compared with several other novel prebiotic fibres. The objective of the present study was to investigate the effect of SCF on Ca absorption and retention in pubertal children by studying the potential absorption mechanisms of the intestinal microbiota. A total of twenty-four adolescent boys and girls (12-15 years) participated in two 3-week metabolic balance studies testing 0 g/d SCF (control (CON) treatment) and 12 g/d SCF (SCF treatment) in a random order by inclusion in a low-Ca diet (600 mg/d). Fractional Ca absorption was measured at the end of the two intervention periods using a dual-stable isotope method. Diet composites and faecal and urine samples were collected daily and analysed for Ca content. Ca retention was calculated as dietary Ca intake minus Ca excretion in faeces and urine over the last 2 weeks. Microbial community composition in the faecal samples collected at the beginning and end of each session was determined by 454 pyrosequencing of the PCR-amplified 16S ribosomal RNA gene. Fractional Ca absorption was 12 % higher (41 mg/d) after the SCF treatment compared with that after the CON treatment (0·664 (sd 0·129) and 0·595 (sd 0·142), respectively; P= 0·02), but Ca retention was unaffected. The average proportion of bacteria of the phylum Bacteroidetes was significantly greater in the participants after the SCF treatment than after the CON treatment. These results suggest that moderate daily intake of SCF, a well-tolerated prebiotic fibre, increases short-term Ca absorption in adolescents consuming less than the recommended amounts of Ca.
Soy isoflavones and their metabolism by intestinal microbiota have gained attention because of potential health benefits, such as the alleviation of estrogen/hormone-related conditions in postmenopausal women, associated with some of these compounds. However, overall changes in gut bacterial community structure and composition in response to addition of soy isoflavones to diets and their association with excreted isoflavone metabolites in postmenopausal women has not been studied. The aim of this study was to determine fecal bacterial community changes in 17 postmenopausal women after a week of diet supplementation with soy bars containing isoflavones, and to determine correlations between microbial community changes and excreted isoflavone metabolites. Using DGGE profiles of PCR amplified 16S rRNA genes (V3 region) to compare microbial communities in fecal samples collected one week before and one week during soy supplementation revealed significant differences (ANOSIM p<0.03) before and after soy supplementation in all subjects. However, between subjects comparisons showed high inter-individual variation that resulted in clustering of profiles by subjects. Urinary excretion of isoflavone (daidzein) metabolites indicated four subjects were equol producers and all subjects produced O-desmethylangolensin (ODMA). Comparison of relative proportions of 16S rRNA genes from 454 pyrosequencing of the last fecal samples of each treatment session revealed significant increases in average proportions of Bifidobacterium after soy consumption, and Bifidobacterium and Eubacterium were significantly greater in equol vs non-S-(-)equol producers. This is the first in vivo study using pyrosequencing to characterize significant differences in fecal community structure and composition in postmenopausal women after a week of soy diet-supplementation, and relate these changes to differences in soy isoflavones and isoflavone metabolites.
Previously, we showed that black girls retained more calcium than white girls did and that salt loading negatively affected calcium retention. Racial differences likely exist in other bone minerals also, such as magnesium, in response to salt loading during growth.
Adolescence is a time for rapid growth that represents an opportunity to influence peak bone mass. Prebiotic agents, such as galacto-oligosaccharides (GOS), increase Ca absorption in animal models and postmenopausal women. The objectives of the present study were to investigate the dose-response relationship of GOS supplementation on Ca absorption during growth and to assess changes in colonic microbiota to better understand the mechanism by which GOS is acting. A total of thirty-one healthy adolescent girls aged 10-13 years consumed smoothie drinks twice daily with 0, 2·5 or 5 g GOS for three 3-week periods in a random order. Fractional Ca absorption was determined from urinary Ca excretion over 48 h at the end of each 3-week period using a dual stable isotope method. Faecal microbiota and bifidobacteria were assessed by PCR-denaturing gradient gel electrophoresis and quantitative PCR. Fractional Ca absorption after the 48 h treatment with control, 5 and 10 g GOS/d was 0·393 (SD 0·092), 0·444 (SD 0·086) and 0·419 (SD 0·099), respectively. Significant improvements in Ca absorption were seen with both low and high doses of GOS compared with the control (P,0·02), but itwas not a dose-response relationship. The increase in absorption was greatest in the urine collected after 24 h, which is consistent with lower gut absorption. Faecal bifidobacteria increased (control 10·89 (SD 13·86), 5 g GOS 22·80 (SD 15·74) and 10 g GOS 11·54 (SD 14·20)) with the GOS treatment (P,0·03). The results suggest that daily consumption of 5 g GOS increases Ca absorption, which may be mediated by the gut microbiota, specifically bifidobacteria.
Increasing calcium bioavailability by decreasing calcium salt particle size in the supplement may be one way to increase calcium absorption. The aim of the study was to compare (1) large versus small particle size CaCO(3) supplements and (2) small particle size CaCO(3) supplement versus placebo on calcium absorption and retention in adolescent girls.
Prebiotics and phytoestrogens have sparked great interest because evidence indicates that the consumption of these dietary constituents leads to lower cholesterol levels and inhibition of postmenopausal bone loss. The aim of this study was to determine the effect of both a prebiotic (Synergy) and a phytoestrogen (genistein) on bone and blood lipid levels in an animal model of postmenopausal women.
Galactooligosaccharides (GOS), prebiotic nondigestible oligosaccharides derived from lactose, have the potential for improving mineral balance and bone properties. This study examined the dose-response effect of GOS supplementation on calcium and magnesium absorption, mineral retention, bone properties, and gut microbiota in growing rats. Seventy-five 4-week-old male Sprague-Dawley rats were randomized into one of five treatment groups (n = 15/group) and fed a diet containing 0, 2, 4, 6, or 8% GOS by weight for 8 weeks. Dietary GOS significantly decreased cecal pH and increased cecal wall weight and content weight in a dose-dependent manner (p < 0.0001). Fingerprint patterns of the 16S rRNA gene PCR-DGGE from fecal DNA indicated the variance of bacterial community structure, which was primarily explained by GOS treatments (p = 0.0001). Quantitative PCR of the samples revealed an increase in the relative proportion of bifidobacteria with GOS (p = 0.0001). Net calcium absorption was increased in a dose-response manner (p < 0.01) with GOS supplementation. Dietary GOS also increased (p < 0.02) net magnesium absorption, femur ??Ca uptake, calcium and magnesium retention, and femur and tibia breaking strength. Distal femur total and trabecular volumetric bone mineral density (vBMD) and area and proximal tibia vBMD increased (p < 0.02) with GOS supplementation. Trabecular-rich bones, that is, those that rapidly turn over, were most benefited. Regression modeling showed that GOS benefited calcium and magnesium utilization and vBMD through decreased cecal pH, increased cecal wall and content weight, and increased proportion of bifidobacteria.
New food sources are needed to bridge the gap between vitamin D intake and recommended intake. We assessed the bioavailability and efficacy of vitamin D in an 8 week dose-response study of bread made with vitamin D2-rich yeast compared to vitamin D3 in growing, vitamin D-deficient rats. Plasma 25-hydroxyvitamin D (25OHD) levels increased in a curvilinear, dose-dependent manner with both forms of vitamin D, but rats fed vitamin D2-rich yeast achieved lower levels than rats fed vitamin D3. Rats fed the highest doses of vitamin D had significantly greater (p<0.05) trabecular BMC, BMD, bone volume, and connectivity density, and greater midshaft total cross-sectional area, compared to rats on the vitamin D-deficient diets, with no significant difference due to vitamin D source. Vitamin D2-rich yeast baked into bread is bioavailable and improves bone quality in vitamin D-deficient animals.
Several studies have shown a positive effect of fructo-oligosaccharides on calcium absorption and retention in animals and humans. Effects of levels of these pre-biotics that can be functionally incorporated into manufactured foods, have not been studied in controlled feeding studies.
In healthy adolescents, cross-sectional studies show either no or negative relationships between serum 25-hydroxyvitamin D [25(OH)D] and calcium (Ca) absorption. Using a 2-period metabolic balance study, the effect of vitamin D supplementation on Ca absorption and retention in adolescent girls was investigated. Eleven girls aged 12-14 y with a mean entry serum 25(OH)D of 35.1 nmol/L consumed a controlled intake (providing 5 ?g vitamin D and 1117 mg Ca/d) for two 3-wk metabolic balance periods separated by a 1-wk washout period. Sunlight exposure was minimized by sunscreen with a sun protection factor ? 15. After the first metabolic balance period, participants received 25 ?g/d cholecalciferol supplementation for 4 wk. Fractional Ca absorption was measured in each metabolic balance period using a stable Ca isotope method. All urine and fecal samples were collected and analyzed to measure net Ca absorption and Ca retention. Paired t tests and correlations were used to analyze the data. Daily supplementation with 25 ?g vitamin D resulted in a mean increase in serum 25(OH)D of 13.3 nmol/L (P < 0.01) but a decrease in fractional Ca absorption of 8.3% (P < 0.05) and no significant change in fasting serum 1,25-dihydroxyvitamin D, parathyroid hormone, net Ca absorption, or Ca skeletal retention. In pubertal girls with vitamin D status considered insufficient in adults, vitamin D supplementation of 25 ?g/d for 4 wk did not improve fractional Ca absorption, net Ca absorption, or Ca retention.
Adequate intake (AI) standards for water in adolescents range between 2.4-3.3 l/day for males and 2.1-2.3 l/day for females, independent of obesity status. Water intakes and excretions of this population are not well documented. The purposes of this study were to assess water turnover, inputs, and outputs in overweight adolescents, compare these parameters between males and females, and evaluate the reproducibility of water turnover. Eighteen girls (BMI 31.7 ± 4 kg/m(2); mean ± s.d.) and nine boys (BMI 26.3 ± 3 kg/m(2)) aged 12-15 years completed two 3-week metabolic balance trials. Rate of water turnover (rH(2)O) was measured by tracking the decline of deuterated water from the body over 14 days. Water inputs (diet*, ad libitum(#), metabolic(#)) and outputs (urine*, feces*, insensible(#)) were assessed (*measured, #estimated). rH(2)O was lower (P = 0.002) in girls vs. boys (3,742 ± 536 vs. 4,537 ± 623 g/day). Per kg body weight, rH(2)O was 28% lower in girls vs. boys (46 ± 7 vs. 64 ± 9 g·kg(-1)·day(-1)). Water input from food and beverages provided and metabolic production were 44 and 28% lower, respectively, in girls vs. boys. Urine and insensible water losses were 21 and 17% lower in girls vs. boys. BMI was positively associated with water turnover in both sexes (girls P = 0.037; boys P = 0.014). The intraclass correlation of rH(2)O between trials was 0.981 (P < 0.001). In conclusion, these overweight adolescents consumed water well in excess of sex-specific AI standards. The lower rH(2)O in girls compared to boys is consistent with adult females and males.
Calcium-41 (t(1/2) = 10(5) years) can be used after a single dose to follow calcium metabolism over a subjects lifetime. The aims of this study were to expand a (41)Ca kinetic model and estimate bone resorption in women with stable bone loss, compare the rates with those calculated with classical isotope studies, and to use the model to simulate dynamic changes in urinary (41)Ca:Ca ratios and bone balance for the design and interpretation of (41)Ca studies. Forty-two women >5 years post-menopause were given (41)Ca intravenously. Bone mineral content and bone mineral density of total body were measured by dual-energy X-ray absorptiometry at the beginning of the study. Urine collections were made periodically for up to ~5 years while subjects were free living. Urinary (41)Ca:Ca ratios were measured using accelerator mass spectrometry. The isotope data were analyzed by compartmental modeling. Four compartments were necessary to fit the urinary tracer data and total bone calcium. The final model included pathways for absorption, distribution, urinary excretion, and endogenous excretion and was used to calculate rates of bone turnover. Estimates of bone resorption in a subset of the women (n = 13), studied previously in a 3-week balance and full kinetic study with (45)Ca, agreed with those using (41)Ca methodology. Thus, rates of bone resorption can be estimated from (41)Ca urinary data in stable post-menopausal women. The model was used to simulate dynamic changes in urinary (41)Ca:Ca ratios and bone balance, as a result of interventions that perturb calcium metabolism to aid in study design and interpretation.
Dietary fibers are thought to benefit bone health through increasing mineral absorption and retention following fermentation in the lower gut and solubilization of minerals. This study compared eight fibers to cellulose following a 12 week intervention for production of short-chain fatty acids (SCFA), calcium absorption, mineral retention and bone content, and bone density and strength in a weanling rat model. Benefits to bone were poorly to modestly related to SCFA production, calcium absorption, or mineral retention, but some parameters were better predicted by cecal content weight, suggesting other mechanisms may be important. Nevertheless, two resistant starches, a soluble fiber dextrin and Polydextrose, increased bone calcium content. Soluble corn fiber and soluble fiber dextrin had the greatest benefit to bone properties including whole body bone mineral content and density and greater volumetric bone mineral density, cortical thickness and area, and peak breaking strength of the distal femur.
(3)H-tetracycline ((3)H-TC) is thought to be superior to calcium (Ca) isotopic tracers for estimating bone resorption rates due to the less redeposition upon release in animal models. However, these 2 tracers have not been compared directly using complete kinetic studies with sampling of blood, urine, feces, and bone. Our goal was to compare the 2 isotopes for evaluating bone turnover. We firstly developed a model for (3)H-TC kinetics in 4-mo-old female rats (n = 3) by measuring the tracer in serum, urine, and feces over 4 d. Then, 9-mo-old ovariectomized (OVX) rats (n = 6) were given both (45)Ca and (3)H-TC subcutaneously. Urine was collected in 24-h pools and assayed for both tracers. Rats were killed 7 and 46 d after the dose and whole skeleton was harvested. We calculated bone resorption rates by modeling the (45)Ca and (3)H-TC data in urine and bone. (3)H-TC kinetics revealed that, like Ca, there are 2 exchangeable compartments between serum and bone. An additional pool was required to account for bone mass of Ca. Bone resorption rates determined from urinary (45)Ca and (3)H-TC did not differ significantly. The tracers (45)Ca and (3)H-TC can be used interchangeably to determine bone resorption rates in OVX rats. Thus, both labels can be used to screen dietary and other interventions for beneficial effects on bone.
Simultaneous and accurate measurement of vitamin D and 25-hydroxyvitamin D in biological samples is a barrier limiting our ability to define "optimal" vitamin D status. Thus, our goal was to optimize conditions and evaluate an LC-MS method for simultaneous detection and quantification of vitamin D(2) , vitamin D(3) , 25-hydroxyvitamin D(2) and 25-hydroxyvitamin D(3) in serum. Extraction and separation of vitamin D forms were achieved using acetone liquid-liquid extraction and by a reversed phase C8 column, respectively. Detection was performed on a triple quadrupole tandem mass spectrometer (QQQ-MS/MS) equipped with atmospheric pressure photo ionization source. The LOQs for all analytes tested were 1 ng/mL for hydroxylated molecules and 2 ng/mL for the parent vitamin Ds. RSD at lower LOQ (2 ng/mL) and in medium (80 ng/mL) and high (200 ng/mL) quality control samples did not exceed 20 and 15% CV, respectively. Accuracy of the method for determination of hydroxylated molecules was also validated using National Institutes of Standards and Technology standard samples and found to be in the range of 90.9-111.2%. In summary, a sensitive and reproducible method is reported for simultaneous quantification of vitamin D(2) , vitamin D(3) , 25-hydroxyvitamin D(2) and 25-hydroxyvitamin D(3) molecules in biological samples.
ECTO-NOX proteins are growth-related cell surface proteins that catalyze both hydroquinone or NADH oxidation and protein disulfide interchange and exhibit time-keeping and prion-like properties. A bacterially expressed truncated recombinant 46 kDa ENOX2 with full ENOX2 activity bound ca 2 moles copper and 2 moles of zinc per mole of protein. Unfolding of the protein in trifluoroacetic acid in the presence of the copper chelator bathocuproine resulted in reversible loss of both enzymatic activities and of a characteristic pattern in the Amide I to Amide II ratios determined by FTIR with restoration by added copper. The H546-V-H together with His 562 form one copper binding site and H582 represents a second copper site as determined from site-directed mutagenesis. Bound copper emerges as having an essential role in ENOX2 both for enzymatic activity and for the structural changes that underly the periodic alternations in activity that define the time-keeping cycle of the protein.
Calcium requirements of North American adolescents were set at 1300 mg/day based on data from white girls. Calcium requirements for Asian-American adolescents have not been studied. Using metabolic balance protocols and a range in calcium intakes, skeletal calcium retention was determined in Chinese-American adolescents. A sample of 29 adolescents, 15 boys aged 12 to 15 years and 14 girls aged 11 to 15 years, was studied twice on paired calcium intakes ranging between 629 to 1835 mg/day using a randomized-order crossover design. Calcium absorption and bone turnover rates using double-stable calcium isotope kinetic analysis on two calcium intakes per subject were measured and compared in boys and girls. Girls and boys had low habitual mean calcium intakes of 648 and 666 mg/day, respectively, and low mean serum 25-hydroxyvitamin D concentrations of 19.1 and 22.2 ng/mL, respectively. True fractional calcium absorption varied inversely with calcium load. Boys had significantly higher bone turnover rate than girls at the same calcium intake. Calcium retention increased with calcium intake; calcium intakes to achieve maximal calcium retention were 1100 mg/day in boys and 970 mg/day in girls. Recommendations for calcium requirements should be lowered for Chinese-American adolescents.
Adolescence is a critical time to achieve peak bone mass. However, results of national surveys suggest that young people are not consuming adequate amounts of calcium. A simple calcium self-assessment tool with low respondent burden could provide immediate feedback that may encourage calcium consumption. This study used data reduction methods to shorten a previously evaluated calcium-specific food frequency questionnaire to the 15-item Brief Calcium Assessment Tool (BCAT). Sample 1 was a cross-sectional sample of 1,426 students in sixth and 11th grades (April 2002 to February 2003) who completed the calcium food frequency questionnaire. Sample 1 was divided into two matching samples (Group 1 and Group 2). Sample 2 was a cross-sectional sample of 745 girls in sixth grade that completed the calcium food frequency questionnaire (July 2002 to February 2005). The development of the BCAT and its calcium score used two unique samples (Group 1 of Sample 1 and Sample 2) followed by confirmation of the score in a second sample (Group 2 of Sample 1). Bone mineral content was measured among the girls in Sample 2. The BCAT calcium score was associated with total hip bone mineral content (beta=.014, P=0.003) and femoral neck bone mineral content (beta=.002, P=0.006) while adjusting for covariates. Sample 3 (n=265), a subsample of Sample 2, completed bone measures 12 months later. The BCAT calcium score at the baseline was a predictor of total body bone mineral content (beta=.279, P=0.041) at 12 months. The correlation of BCAT scores 1 week apart was .76 (P<0.001) among a convenience sample of 41 boys and girls aged 9 to 17 years (October 2005 to December 2005). The BCAT took about 5 minutes to complete. The significant relationship between bone mass and the BCAT calcium score supports the use of BCAT as a tool to assess food calcium intake among adolescents, especially Asian, Hispanic, and non-Hispanic white girls. The reliability and low respondent burden of the BCAT make the tool useful for large-scale studies and community-based programs.
Techniques for assessing bone dynamic are in high demand. Calcium (Ca) kinetic studies are currently being used in our clinical studies of bone turnover in adolescents and elderly. The technique has rarely been compared to the standard method of bone dynamic histomorphometry. We perturbed bone turnover through ovariectomy and sub-optimal dietary Ca in a female rat model to cross-calibrate Ca kinetics against dynamic histomorphometry. Kinetic studies involved oral and intravenous administration of (45)Ca and monitoring the tracer in blood, urine, feces, and bone over a 3-day period as part of a metabolic Ca balance study. Histomorphometric indices of mineral apposition rate, mineralizing surface, and bone formation rate were obtained from proximal metaphysis and mid-diaphysis region of tibial bone. Bone mineralization and resorption rates at the whole skeletal level as evaluated by kinetic studies were significantly correlated with the volume-based bone formation rate (BFR/BV) evaluated by dynamic histomorphometry in metaphyseal trabecular bone (r=0.72 and r=0.61, respectively, p<0.001) and surface-based bone formation rate (BFR/BS) in tibial cortex (r=0.63, p<0.001 and r=0.59, p<0.01, respectively). Significant correlations were also demonstrated between bone resorption and mineralization rates at the whole skeletal level (r=0.91, p<0.001) using (45)Ca kinetic data. Ca kinetic modeling showed an increase (p<0.001) in skeletal resorption and formation rates in response to ovariectomy (27.6 vs. 13.8 mg/d for bone resorption and 42.7 vs. 28 mg/d for bone formation in ovariectomized vs. their Sham-operated control animals, respectively). Ca kinetic data also showed that bone formation decreased by 30% and whole bone balance by 50%, when dietary Ca level was reduced from 0.4% to 0.2% (34.2 vs. 23.8 mg/d and 10.4 vs. 5.1 mg/d, respectively, p<0.001). Our data suggest that Ca kinetic studies can be used reliably to rapidly detect changes in bone turnover at the whole skeletal level in response to interventions.
Soy isoflavones and their metabolites, with estrogenic activity, have been considered candidates for reducing postmenopausal bone loss. In this study, we examined the effect of dietary equol, a bioactive metabolite of the soy isoflavone daidzein, on equol tissue distribution, bone parameters, and reproductive tissue activity using an adult ovariectomized (OVX) rat model. An 8-wk feeding study was conducted to compare 4 dietary treatments of equol (0, 50, 100, 200 mg/kg diet) in 6-mo-old OVX female Sprague-Dawley rats. A dose response increase in tissue equol concentrations was observed for serum, liver, kidney, and heart, and a plateau occurred at 100 mg equol/kg diet for intestine. In OVX rats receiving 200 mg equol/kg diet, femoral calcium concentration was greater than those receiving lower doses but was still less than SHAM (P < 0.05), and other bone measures were not improved. Tibia calcium concentrations were lower in OVX rats receiving 100 and 200 mg equol/kg diet compared with the OVX control rats. Trabecular bone mineral density of tibia was also lower in equol-fed OVX rats. At this dietary equol intake, uterine weight was higher (P < 0.05) than in other OVX groups but lower than the SHAM-operated intact rats. The 200 mg/kg diet dose of dietary equol significantly increased proliferative index in the uterine epithelium. Dietary equol had no stimulatory effect on mammary gland epithelium. We conclude that in OVX rats, a dietary equol dose that had modest effect on bone also exerts mild uterotropic effects.
Calcium (Ca) supplements, especially Ca carbonate (CaCO3), are the main alternative sources of dietary Ca and an important part of a treatment regimen for osteoporosis, the most common metabolic bone disorder of aging and menopause. In a female ovariectomized (OVX) rat model for studying postmenopausal osteoporosis, we tested the hypothesis that a small compared with a large particle size of CaCO3 (13.0- vs. 18.5-mum geometric diameter) would result in increased Ca balance and subsequently bone mass and that this would be affected by dietary Ca level. We used 6-mo-old rats that were OVX either at 6 or 3 mo of age as models of early or stable menopausal status, respectively. The rats received semipurified diets that contained either 0.4 or 0.2% dietary Ca provided from CaCO3 of 2 particle sizes. A group of Sham-operated rats with intact ovaries served as control and were fed 0.4% dietary Ca from large particles. Estrogen deficiency as a result of ovariectomy had an adverse effect on bone density, mineral content, and bone mechanical properties (P < 0.001). Reducing dietary Ca from 0.4 to 0.2% resulted in significant adverse effects on bone density and mechanical properties (P < 0.001). The particle size of CaCO3 did not affect total Ca balance, bone dual energy X-ray absorptiometry and peripheral quantitative computed tomography indices, bone ash and Ca content, or the mechanical determinants of bone strength. We conclude that a decrease in particle size of CaCO3 to 70% of that typically found in Ca supplements does not provide a benefit to overall Ca metabolism or bone characteristics and that the amount of Ca consumed is of greater influence in enhancing Ca nutrition and skeletal strength.
Dairy products provide most of the calcium in the diet, but consumption of milk has declined steadily over the last six decades. We determined (1) benefits to bone formation in growing female Sprague-Dawley rats when calcium is derived from dairy versus CaCO(3) and (2) residual benefits of calcium provided by dairy to rats subsequently receiving a low-calcium diet. During growth, femurs from rats fed nonfat dry milk solids (NFDM) had 8.4% higher peak breaking force, 6.4% greater Ca content, 4.8% greater weight, 4% greater width, 1.2% greater density, 13.1% greater midshaft cortical thickness, and 16.7% greater midshaft cortical area than from rats fed CaCO(3). These effects were unrelated to differences in calcium absorption or serum IGF-1, but the NFDM group had higher rates of bone formation. If maintained on an adequate calcium diet, many of these advantages disappeared. However, rats fed adequate Ca as NFDM versus CaCO(3) during growth and subsequently switched to deficient Ca as CaCO(3) had significantly (p < 0.0001) higher femoral BMD (1.3%), total bone Ca (7.2%), Ca concentration (4.6%), and cortical thickness (9.4%) and a trend (p = 0.02) toward greater peak breaking force (17%). Thus, NFDMs improved bone measures during growth and protected bone against a subsequent period of calcium depletion compared with CaCO(3).
Patients with chronic kidney disease (CKD) are given calcium carbonate to bind dietary phosphorus, reduce phosphorus retention, and prevent negative calcium balance; however, data are limited on calcium and phosphorus balance during CKD to support this. Here, we studied eight patients with stage 3 or 4 CKD (mean estimated glomerular filtration rate 36 ml/min) who received a controlled diet with or without a calcium carbonate supplement (1500 mg/day calcium) during two 3-week balance periods in a randomized placebo-controlled cross-over design. All feces and urine were collected during weeks 2 and 3 of each balance period and fasting blood, and urine was collected at baseline and at the end of each week. Calcium kinetics were determined using oral and intravenous (45)calcium. Patients were found to be in neutral calcium and phosphorus balance while on the placebo. Calcium carbonate supplementation produced positive calcium balance, did not affect phosphorus balance, and produced only a modest reduction in urine phosphorus excretion compared with placebo. Calcium kinetics demonstrated positive net bone balance but less than overall calcium balance, suggesting soft-tissue deposition. Fasting blood and urine biochemistries of calcium and phosphate homeostasis were unaffected by calcium carbonate. Thus, the positive calcium balance produced by calcium carbonate treatment within 3 weeks cautions against its use as a phosphate binder in patients with stage 3 or 4 CKD, if these findings can be extrapolated to long-term therapy.
Low serum 25-hydroxyvitamin D [25 (OH) D] is common in healthy children particularly in blacks. However, serum 25 (OH) D concentrations for optimal bone turnover in children is unknown and few data exist that describe effects of increasing serum 25 (OH) D on bone turnover markers during puberty. The purpose of this study was to determine the relationships between serum 25 (OH) D and changes in serum 25 (OH) D and bone turnover in white and black pubertal adolescents. Bone turnover markers were measured in 318 healthy boys and girls from Georgia (34°N) and Indiana (40°N) who participated in a study of oral vitamin D(3) supplementation (0 to 4000 IU/d). Serum 25 (OH) D, osteocalcin, bone alkaline phosphatase, and urine N-telopeptide cross-links were measured at baseline and 12 weeks. Relationships among baseline 25 (OH) D and bone biomarkers, and between changes over 12 weeks were determined and tested for effects of race, sex, latitude, and baseline 25 (OH) D. Median 25 (OH) D was 27.6 ng/mL (n=318, range 10.1-46.0 ng/mL) at baseline and 34.5 ng/mL (n=302, range 9.7-95.1 ng/mL) at 12 weeks. Neither baseline nor change in 25 (OH) D over 12 weeks was associated with bone turnover. The lack of association was not affected by race, sex, latitude, or baseline serum 25 (OH) D. Serum 25 (OH) D in the range of 10-46 ng/mL appears to be sufficient for normal bone turnover in healthy black and white pubertal adolescents.
Age-related changes in calcium metabolism play a role in the development of osteoporosis. A 4-wk feeding study was conducted in 5-mo-old ovariectomized (OVX) Sprague-Dawley rats to assess the effect of various dietary fibers on mineral metabolism and bone health parameters. There were 6 treatment groups: sham-Control, OVX-Control, OVX rats receiving daily estradiol (E?) injections, and OVX rats receiving an AIN-93M diet supplement with either an inulin-based fiber (Synergy1® or Fruitafit HD®) or a novel fiber (polydextrose) at 5% wt. of diet. Calcium and magnesium metabolic balances were performed after early (3 d) and late exposure (4 wk) to dietary treatments. Rats receiving polydextrose had significantly higher net calcium absorption efficiency and retention than all control groups and a trend (P? 0.10) for higher calcium absorption when compared to inulin-based fibers after early exposure but the advantage did not persist over long-term exposure. The inulin-based fibers had positive chronic effects on calcium metabolism that were related to changes in the gut, that is, production of short chain fatty acids and higher cecal wall weights. All fibers improved magnesium absorption and retention in early and late metabolic balances; effects on magnesium metabolism were more pronounced than for calcium.
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