Intermittent fasting (IF) is a promising strategy for weight loss and improving metabolic health, but its effects on bone health are less clear. This review aims to summarise and critically evaluate the preclinical and clinical evidence on IF regimens (the 5:2 diet, alternate-day fasting (ADF) and time-restricted eating (TRE)/time-restricted feeding and bone health outcomes. Animal studies have utilised IF alongside other dietary practices known to elicit detrimental effects on bone health and/or in models mimicking specific conditions; thus, findings from these studies are difficult to apply to humans. While limited in scope, observational studies suggest a link between some IF practices (e.g. breakfast omission) and compromised bone health, although lack of control for confounding factors makes these data difficult to interpret. Interventional studies suggest that TRE regimens practised up to 6 months do not adversely affect bone outcomes and may even slightly protect against bone loss during modest weight loss (< 5 % of baseline body weight). Most studies on ADF have shown no adverse effects on bone outcomes, while no studies on the ‘5–2’ diet have reported bone outcomes. Available interventional studies are limited by their short duration, small and diverse population samples, assessment of total body bone mass exclusively (by dual-energy X-ray absorptiometry) and inadequate control of factors that may affect bone outcomes, making the interpretation of existing data challenging. Further research is required to better characterise bone responses to various IF approaches using well-controlled protocols of sufficient duration, adequately powered to assess changes in bone outcomes and designed to include clinically relevant bone assessments.

For an optimal performance, soccer players and referees need to consume a high-quality diet. The Healthy Eating Index (HEI) is a tool that can estimate diet quality and has been shown to be associated with body composition. The aims of the present study were first to determine the HEI-2015 score of the diets consumed by athletes and second its association with different body composition parameters of athletes. We conducted a cross-sectional study on 198 soccer players and referees. Dietary intakes were recorded using a validated food frequency questionnaire (FFQ), and HEI scores were calculated. Body composition parameters were measured using the bioelectrical impedance analysis. The mean score for the HEI-2015 was 65⋅04. A multiple linear regression model showed significant associations of the HEI-2015 score with percent body fat (PBF), percent muscle mass (PMM), waist-to-hip ratio (WHR) and waist-to-height ratio (WHtR) in male soccer players aged <18 years, body mass index (BMI) in male soccer players aged ≥18 years and BMI and waist-to-height ratio (WHtR) in male soccer referees after adjustment for covariates (P < 0⋅05). The mean overall score for the HEI-2015 shows that Iranian soccer players and referees have an acceptable quality of diet. We also found significant associations between the HEI-2015 score and different body composition parameters in male soccer players and referees but we did not find any significant association in female athletes (P > 0⋅05).

Few investigations have evaluated the validity of current body composition technology among racially and ethnically diverse populations. This study assessed the validity of common body composition methods in a multi-ethnic sample stratified by race and ethnicity. One hundred and ten individuals (55 % female, age: 26·5 (sd 6·9) years) identifying as Asian, African American/Black, Caucasian/White, Hispanic, Multi-racial and Native American were enrolled. Seven body composition models (dual-energy X-ray absorptiometry (DXA), air displacement plethysmography (ADP), two bioelectrical impedance devices (BIS, IB) and three multi-compartment models) were evaluated against a four-compartment criterion model by assessing total error (TE) and standard error of the estimate. For the total sample, measures of % fat and fat-free mass (FFM) from multi-compartment models were all excellent to ideal (% fat: TE = 0·94–2·37 %; FFM: TE = 0·72–1·78 kg) compared with the criterion. % fat measures were very good to excellent for DXA, ADP and IB (TE = 2·52–2·89 %) and fairly good for BIS (TE = 4·12 %). For FFM, single device estimates were good (BIS; TE = 3·12 kg) to ideal (DXA, ADP, IB; TE = 1·21–2·15 kg). Results did not vary meaningfully between each race and ethnicity, except BIS was not valid for African American/Black, Caucasian/White and Multi-racial participants for % fat (TE = 4·3–4·9 %). The multi-compartment models evaluated can be utilised in a multi-ethnic sample and in each individual race and ethnicity to obtain highly valid results for % fat and FFM. Estimates from DXA, ADP and IB were also valid. The BIS may demonstrate greater TE for all racial and ethnic cohorts and results should be interpreted cautiously.

Maintaining Mg status may be important for military recruits, a population that experiences high rates of stress fracture during initial military training (IMT). The objectives of this secondary analysis were to (1) compare dietary Mg intake and serum Mg in female and male recruits pre- and post-IMT, (2) determine whether serum Mg was related to parameters of bone health pre-IMT, and (3) whether Ca and vitamin D supplementation (Ca/vitamin D) during IMT modified serum Mg. Females (n 62) and males (n 51) consumed 2000 mg of Ca and 25 μg of vitamin D/d or placebo during IMT (12 weeks). Dietary Mg intakes were estimated using FFQ, serum Mg was assessed and peripheral quantitative computed tomography was performed on the tibia. Dietary Mg intakes for females and males pre-IMT were below the estimated average requirement and did not change with training. Serum Mg increased during IMT in females (0·06 ± 0·08 mmol/l) compared with males (–0·02 ± 0·10 mmol/l; P < 0·001) and in those consuming Ca/vitamin D (0·05 ± 0·09 mmol/l) compared with placebo (0·001 ± 0·11 mmol/l; P = 0·015). In females, serum Mg was associated with total bone mineral content (BMC, β = 0·367, P = 0·004) and robustness (β = 0·393, P = 0·006) at the distal 4 % site, stress–strain index of the polaris axis (β = 0·334, P = 0·009) and robustness (β = 0·420, P = 0·004) at the 14 % diaphyseal site, and BMC (β = 0·309, P = 0·009) and stress–strain index of the polaris axis (β = 0·314, P = 0·006) at the 66 % diaphyseal site pre-IMT. No significant relationships between serum Mg and bone measures were observed in males. Findings suggest that serum Mg may be modulated by Ca/vitamin D intake and may impact tibial bone health during training in female military recruits.

Understanding the effects of acute feeding on body composition and metabolic measures is essential to the translational component and practical application of measurement and clinical use. To investigate the influence of acute feeding on the validity of dual-energy X-ray absorptiometry (DXA), a four-compartment model (4C) and indirect calorimetry metabolic outcomes, thirty-nine healthy young adults (n 19 females; age: 21·8 (sd 3·1) years, weight; 71·5 (sd 10·0) kg) participated in a randomised cross-over study. Subjects were provided one of four randomised meals on separate occasions (high carbohydrate, high protein, ad libitum or fasted baseline) prior to body composition and metabolic assessments. Regardless of macronutrient content, acute feeding increased DXA percent body fat (%fat) for the total sample and females (average constant error (CE):–0·30 %; total error (TE): 2·34 %), although not significant (P = 0·062); the error in males was minimal (CE: 0·11 %; TE: 0·86 %). DXA fat mass (CE: 0·26 kg; TE: 0·75 kg) and lean mass (LM) (CE: 0·83 kg; TE: 1·23 kg) were not altered beyond measurement error for the total sample. 4C %fat was significantly impacted from all acute feedings (avg CE: 0·46 %; TE: 3·7 %). 4C fat mass (CE: 0·71 kg; TE: 3·38 kg) and fat-free mass (CE: 0·55 kg; TE: 3·05 kg) exceeded measurement error for the total sample. RMR was increased for each feeding condition (TE: 1666·9 kJ/d; 398 kcal/d). Standard pre-testing fasting guidelines may be important when evaluating DXA and 4C %fat, whereas additional DXA variables (fat mass and LM) may not be significantly impacted by an acute meal. Measuring body composition via DXA under less stringent pre-testing guidelines may be valid and increase feasibility of testing in clinical settings.

For many people, micronutrient requirement means the amount needed in the diet to ensure adequacy. Dietary reference values (DRV) provide guidance on the daily intake of vitamins and minerals required to ensure the needs of the majority in the population are covered. These are developed on estimates of the quantity of each micronutrient required by the average person, the bioavailability of the micronutrient from a typical diet and the interindividual variability in these amounts. Sex differences are inherent in the requirements for many micronutrients because they are influenced by body size or macronutrient intake. These are reflected in different DRV for males and females for some micronutrients, but not all, either when data from males and females are combined or when there is no evidence of sex differences. Pregnancy and lactation represent times when micronutrient requirements for females may differ from males, and separate DRV are provided. For some micronutrients, no additional requirement is indicated during pregnancy and lactation because of physiological adaptations. To date, little account has been taken of more subtle sex differences in growth and maturation rates, health vulnerabilities and in utero and other programming effects. Over the years, the MRC Nutrition and Bone Health Group has contributed data on micronutrient requirements across the lifecourse, particularly for calcium and vitamin D, and shown that supplementation can have unexpected sex-specific consequences that require further investigation. The present paper outlines the current issues and the need for future research on sex differences in micronutrient requirements.

High dietary phosphorus (P), particularly soluble salts, may contribute to chronic kidney disease development in cats. The aim of the present study was to assess the safety of P supplied at 1 g/1000 kcal (4184kJ) from a highly soluble P salt in P-rich dry format feline diets. Seventy-five healthy adult cats (n 25/group) were fed either a low P control (1·4 g/1000 kcal [4184kJ]; Ca:P ratio 0·97) or one of two test diets with 4 g/1000 kcal (4184 kJ); Ca:P 1·04 or 5 g/1000 kcal (4184kJ); Ca:P 1·27, both incorporating 1 g/1000 kcal (4184 kJ) sodium tripolyphosphate (STPP) – for a period of 30 weeks in a randomised parallel-group study. Health markers in blood and urine, glomerular filtration rate, renal ultrasound and bone density were assessed at baseline and at regular time points. At the end of the test period, responses following transition to a commercial diet (total P – 2·34 g/1000 kcal [4184kJ], Ca:P 1·3) for a 4-week washout period were also assessed. No adverse effects on general, kidney or bone (skeletal) function and health were observed. P and Ca balance, some serum biochemistry parameters and regulatory hormones were increased in cats fed test diets from week 2 onwards (P ≤ 0·05). Data from the washout period suggest that increased serum creatinine and urea values observed in the two test diet groups were influenced by dietary differences during the test period, and not indicative of changes in renal function. The present data suggest no observed adverse effect level for feline diets containing 1 g P/1000 kcal (4184 kJ) from STPP and total P level of up to 5 g/1000 kcal (4184 kJ) when fed for 30 weeks.

Vitamin D supplementation in infancy is recommended to prevent rickets. At the population level, its effects on bone mineralisation are largely unknown. We aimed to explore whether adherence to national vitamin D supplementation guidelines (10 µg/d up to the age of 2 years), supplementation at the ages of 5 and 7 years, and serum 25-hydroxyvitamin D (s-25(OH)D) at various time points associated with bone mineral density (BMD) at the age of 7 years in the Odense Child Cohort, Denmark (n 1194). High adherence was defined as supplementation with 10 µg of vitamin D 6–7 times per week during ≥80 % of the observation time. s-25(OH)D was analysed using LC-MS/MS. Total-body-less-head (TBLH) BMD was measured by dual-energy X-ray absorptiometry. At the median age of 18·1 months, 53·9 % (n 475/881) reported high adherence. The median s-25(OH)D was 64·7, 78·8, 46·0 and 71·8 nmol/l in early pregnancy, late pregnancy, cord blood and at 5 years, respectively. The mean TBLH BMD at the median age of 7·1 years was 0·613 (SD 0·049) g/cm2 (z-score +0·363 (SD 0·824)). In adjusted analyses, vitamin D supplementation up to 18 months, and at 5 and 7 years, was not associated with TBLH BMD. Similarly, no robust associations were found between TBLH BMD and s-25(OH)D at any time point. No associations were found for TBLH bone mineral concentration or bone area. In this population with relatively high s-25(OH)D concentrations, no consistent associations were found between adherence to vitamin D supplementation recommendations or vitamin D status in pregnancy or childhood, and bone mineralisation at the age of 7 years.

The present paper reviews published literature on the relationship between dietary protein and bone health. It will include arguments both for and against the anabolic and catabolic effects of dietary protein on bone health. Adequate protein intake provides the amino acids used in building and maintaining bone tissue, as well as stimulating the action of insulin-like growth factor 1, which in turn promotes bone growth and increases calcium absorption. However, the metabolism of dietary sulphur amino acids, mainly from animal protein, can lead to increased physiological acidity, which may be detrimental for bone health in the long term. Similarly, cereal foods contain dietary phytate, which in turn contains phosphate. It is known that phosphate consumption can also lead to increased physiological acidity. Therefore, cereal products may produce as much acid as do animal proteins that contain sulphur amino acids. The overall effect of dietary protein on physiological acidity, and its consequent impact on bone health, is extremely complex and somewhat controversial. The consensus is now moving towards a synthesised approach. Particularly, how anabolic and catabolic mechanisms interact; as well as how the context of the whole diet and the type of protein consumed is important.

The purpose of the present study was: (1) to develop a new dual-energy X-ray absorptiometry (DXA)-derived body volume (BV) equation with the GE-Lunar prodigy while utilising underwater weighing (UWW) as a criterion and (2) to cross-validate the novel DXA-derived BV equation (4C-DXANickerson), Wilson DXA-derived BV equation (4C-DXAWilson) and air displacement plethysmography (ADP)-derived BV (4C-ADP) in Hispanic adults. A total of 191 Hispanic adults (18–45 years) participated in the present study. The development sample consisted of 120 females and males (50 % females), whereas the cross-validation sample comprised of forty-one females and thirty males (n 71). Criterion body fat percentage (BF %) and fat-free mass (FFM) were determined using a four-compartment (4C) model with UWW as a criterion for BV (4C-UWW). 4C-DXANickerson, 4C-DXAWilson and 4C-ADP were compared against 4C-UWW in the cross-validation sample. 4C-DXANickerson, 4C-DXAWilson and 4C-ADP all produced similar validity statistics when compared with 4C-UWW in Hispanic males (all P > 0·05). 4C-DXANickerson also yielded similar BF % and FFM values as 4C-UWW when evaluating the mean differences (constant error (CE)) in Hispanic females (CE = –0·79 % and 0·38 kg; P = 0·060 and 0·174, respectively). However, 4C-DXAWilson produced significantly different BF % and FFM values (CE = 3·22 % and –2·20 kg, respectively; both P < 0·001). Additionally, 4C-DXAWilson yielded significant proportional bias when estimating BF % (P < 0·001), whereas 4C-ADP produced significant proportional bias for BF % and FFM (both P < 0·05) when evaluated in Hispanic females. The present study findings demonstrate that 4C-DXANickerson is a valid measure of BV in Hispanics and is recommended for use in clinics, where DXA is the main body composition assessment technique.

The present study reports the effects of a high-fat (HF) diet of over 8 weeks on the Fe status of growing rats. Tissue Fe levels were analysed by atomic absorption spectrophotometry, and whole-body adiposity was measured by dual-energy X-ray absorptiometry. Histopathology and morphometry of adipose tissue were performed. Liver homogenates were used for measuring ferroportin-1 protein levels by immunoblotting, and transcript levels were used for Fe genes measured by real-time PCR. Tissue Fe pools were fit to a compartmental biokinetic model in which Fe was assessed using fourteen compartments and twenty-seven transfer constants (kj,i from tissue ‘i’ to tissue ‘j’) adapted from the International Commission on Radiological Protection (ICRP) 69. Ten kj,i were calculated from the experimental data using non-linear regression, and seventeen were estimated by allometry according to the formula ${k_{i,j}} = a \times {M^b}$. Validation of the model was carried out by comparing predicted and analysed Fe pool sizes in erythrocytes, the liver and the spleen. Body adiposity was negatively associated with serum Fe levels and positively associated with liver Fe stores. An inferred increase in Fe transfer from bone marrow to the liver paralleled higher hepatic Fe concentrations and ferritin heavy-chain mRNA levels in the HF diet-fed animals, suggesting that liver Fe accumulation occurred at least in part due to a favoured liver erythrocyte uptake. If this feeding condition was to be prolonged, impaired Fe decompartmentalisation may occur, ultimately resulting in dysmetabolic Fe overload.

The consumption of high-Ca, high-protein dairy foods (i.e. milk, cheese, yogurt) is advocated for bone health across the lifespan to reduce the risk of low-trauma fractures. However, to date, the anti-fracture efficacy of dairy food consumption has not been demonstrated in randomised controlled trials but inferred from cross-sectional and prospective studies. The anti-fracture efficacy of dairy food consumption is plausible, but testing this requires a robust study design to ensure outcomes are suitably answering this important public health question. The evidence of skeletal benefits of dairy food consumption is equivocal, not because it may not be efficacious but because the study design and execution are often inadequate. The key issues are compliance with dietary intervention, dropouts, sample sizes and most importantly lack of deficiency before intervention. Without careful appraisal of the design and execution of available studies, precarious interpretations of outcomes may be made from these poorly designed or executed studies, without consideration of how study design may be improved. Dairy food interventions in children are further hampered by heterogeneity in growth: in particular sex and maturity-related differences in the magnitude, timing, location and surface-specific site of bone accrual. Outcomes of studies combining children of different sexes and maturity status may be masked or exaggerated by these differences in growth, so inaccurate conclusions are drawn from results. Until these critical issues in study design are considered in future dairy food interventions, the anti-fracture efficacy of dairy food consumption may remain unknown and continue to be based on conjecture.

Consumption of a high-fat diet, rich in SFA, causes deterioration of bone properties. Some studies suggest that feeding inulin to animals may increase mineral absorption and positively affect bone quality; however, these studies have been carried out only on rodents fed a standard diet. The primary objective of this study was to determine the effect of inulin on bone health of pigs (using it as an animal model for humans) fed a high-fat diet rich in SFA, having an unbalanced ratio of lysine:metabolisable energy. It was hypothesised that inulin reduces the negative effects of such a diet on bone health. At 50 d of age, twenty-one pigs were randomly allotted to three groups: the control (C) group fed a standard diet, and two experimental (T and TI) groups fed a high-fat diet rich in SFA. Moreover, TI pigs consumed an extra inulin supply (7 % of daily feed intake). After 10 weeks, whole-body bone mineral content (P=0·0054) and bone mineral density (P=0·0322) were higher in pigs of groups TI and C compared with those of group T. Femur bone mineral density was highest in pigs in group C, lower in group TI and lowest in group T (P=0·001). Femurs of pigs in groups TI and C had similar, but higher, maximum strength compared with femurs of pigs in group T (P=0·0082). In conclusion, consumption of a high-fat diet rich in SFA adversely affected bone health, but inulin supplementation in such a diet diminishes this negative effect.

The relevance of dairy produce for the diminishment of osteoporotic risk is still a matter of scientific debate due to the outcome of a few single observational studies. This review will address the most robust point estimate on the role of dairy products, as reported in systematic reviews and meta-analyses on randomised controlled trials in the case of bone mineralisation or prospective studies in the case of fracture risk. Plain dairy products or those fortified with Ca and/or vitamin D improve total body bone mineral content (BMC) by 45–50 g over 1 year when the daily baseline Ca intake is lower than 750 mg in Caucasians and Chinese girls. In Caucasian and Chinese women, Ca from (fortified) dairy products increases bone mineral density (BMD) by 0·7–1·8 % over 2 years dependent on the site of measurement. Despite the results on BMC, there are currently no studies that have investigated the potential of dairy products to reduce fracture risk in children. In adult Caucasian women, daily intake of 200–250 ml of milk is associated with a reduction in fracture risk of 5 % or higher. In conclusion, the role of dairy products for BMC or BMD has been sufficiently established in Chinese and Caucasian girls and women. In Caucasian women, drinking milk also reduces fracture risk. More research on the role of dairy products within the context of bone health-promoting diets is needed in specific ethnicities, other than Chinese and Caucasians, and in men.

Oestrogen and n-3 PUFA, especially EPA and DHA, have been reported to have beneficial effects on bone loss. Thus, the purpose of the present study was to investigate the synergistic bone-protective mechanism of combined treatments of EPA+DHA supplementation and oestrogen injection in ovariectomised rats. Rats were fed a modified American Institute of Nutrition-93G diet with 0 %, 1 % or 2 % n-3 PUFA (EPA+DHA) relative to the total energy intake for 12 weeks. Rats were surgically ovariectomised at week 8, and after a 1-week recovery period rats were injected with either 17β-oestradiol-3-benzoate (E2) or maize oil for the last 3 weeks. Combined use of n-3 PUFA and E2 synergistically increased femoral cortical bone volume, bone mineral content and the bone expression of runt-related transcription factor 2 (RUNX2), but decreased the bone expression of IL-1β. Both n-3 PUFA and E2 decreased the bone expressions of IL-7, TNF-α and PPAR-γ, and increased the bone expression of oestrogen receptor-α. n-3 PUFA in the presence of E2 and E2 alone significantly decreased the bone expressions of IL-1β and IL-6 and increased the bone expression of RUNX2. E2 significantly decreased the serum levels of bone turnover markers and the bone expression of receptor activator of NF-κB ligand, but decreased the bone expression of osteoprotegerin. The combined use of n-3 PUFA and E2 exerted synergistic bone-protective efficacy through up-regulation of RUNX2, an essential transcription factor for bone formation, as well as the suppression of bone-resorbing cytokine IL-1β.

Maternal dietary vitamin D carry-over effects were assessed in young pigs to characterise skeletal abnormalities in a diet-induced model of kyphosis. Bone abnormalities were previously induced and bone mineral density (BMD) reduced in offspring from sows fed diets with inadequate vitamin D3. In a nested design, pigs from sows (n 23) fed diets with 0 (−D), 8·125 (+D) or 43·750 (++D) µg D3/kg from breeding through lactation were weaned and, within litter, fed nursery diets arranged as a 2×2 factorial design with 0 (−D) or 7·0 (+D) µg D3/kg, each with 95 % (95P) or 120 % (120P) of P requirements. Selected pigs were euthanised before colostrum consumption at birth (0 weeks, n 23), weaning (3 weeks, n 22) and after a growth period (8 weeks, n 185) for BMD, bone mechanical tests and tissue mRNA analysis. Pigs produced by +D or ++D sows had increased gain at 3 weeks (P<0·05), and at 8 weeks had increased BMD and improved femur mechanical properties. However, responses to nursery diets depended on maternal diets (P<0·05). Relative mRNA expressions of genes revealed a maternal dietary influence at birth in bone osteocalcin and at weaning in kidney 24-hydroxylase (P<0·05). Nursery treatments affected mRNA expressions at 8 weeks. Detection of a maternal and nursery diet interaction (P<0·05) provided insights into the long-term effects of maternal nutritional inputs. Characterising early stages of bone abnormalities provided inferences for humans and animals about maternal dietary influence on offspring skeletal health.

Breast-feeding has been associated with later bone health, but results from previous studies are inconsistent. We examined the associations of breast-feeding patterns and timing of introduction of solids with bone mass at the age of 6 years in a prospective cohort study among 4919 children. We collected information about duration and exclusiveness of breast-feeding and timing of introduction of any solids with postnatal questionnaires. A total body dual-energy X-ray absorptiometry scan was performed at 6 years of age, and bone mineral density (BMD), bone mineral content (BMC), area-adjusted BMC (aBMC) and bone area (BA) were analysed. Compared with children who were ever breast-fed, those never breast-fed had lower BMD (−4·62 mg/cm2; 95 % CI −8·28, −0·97), BMC (−8·08 g; 95 % CI −12·45, −3·71) and BA (−7·03 cm2; 95 % CI −12·55, −1·52) at 6 years of age. Among all breast-fed children, those who were breast-fed non-exclusively in the first 4 months had higher BMD (2·91 mg/cm2; 95 % CI 0·41, 5·41) and aBMC (3·97 g; 95 % CI 1·30, 6·64) and lower BA (−4·45 cm2; 95 % CI −8·28, −0·61) compared with children breast-fed exclusively for at least 4 months. Compared with introduction of solids between 4 and 5 months, introduction <4 months was associated with higher BMD and aBMC, whereas introduction between 5 and 6 months was associated with lower aBMC and higher BA. Additional adjustment for infant vitamin D supplementation did not change the results. In conclusion, results from the present study suggest that ever breast-feeding compared with never breast-feeding is associated with higher bone mass in 6-year-old children, but exclusive breast-feeding for 4 months or longer was not positively associated with bone outcomes.

The loss of estrogen during menopause causes changes in the female body, with wide-ranging effects on health. Estrogen-containing hormone replacement therapy (HRT) leads to a relief of typical menopausal symptoms, benefits bone and muscle health, and is associated with tissue-specific gene expression profiles. As gene expression is controlled by epigenetic factors (including DNA methylation), many of which are environmentally sensitive, it is plausible that at least part of the HRT-associated gene expression is due to changes in DNA methylation profile. We investigated genome-wide DNA methylation and gene expression patterns of white blood cells (WBCs) and their associations with body composition, including muscle and bone measures of monozygotic (MZ) female twin pairs discordant for HRT. We identified 7,855 nominally significant differentially methylated regions (DMRs) associated with 4,044 genes. Of the genes with DMRs, five (ACBA1, CCL5, FASLG, PPP2R2B, and UHRF1) were also differentially expressed. All have been previously associated with HRT or estrogenic regulation, but not with HRT-associated DNA methylation. All five genes were associated with bone mineral content (BMC), and ABCA1, FASLG, and UHRF1 were also associated with body adiposity. Our study is the first to show that HRT associates with genome-wide DNA methylation alterations in WBCs. Moreover, we show that five differentially expressed genes with DMRs associate with clinical measures, including body fat percentage, lean body mass, bone mass, and blood lipids. Our results indicate that at least part of the known beneficial HRT effects on body composition and bone mass may be regulated by DNA methylation associated alterations in gene expression in circulating WBCs.

Increasing dietary Ca intake may prevent the excessive mobilisation of bone mineral in nursing mothers. We aimed to investigate whether higher Ca intake could positively modulate the bone mineral changes in Chinese postpartum lactating women. The study was a 12-month randomised, double-blinded, parallel group trial conducted over 12 months. A total of 150 postpartum women were randomly selected to receive either 40 g of milk powder containing 300 mg of Ca and 5 μg of vitamin D (Low-Ca group) or same milk powder additionally fortified with 300 mg of Ca (Mid-Ca group) or 600 mg of Ca (High-Ca group). Bone mineral density (BMD) for the whole body, the lumbar spine, the total left hip and its sub-regions was measured using dual-energy X-ray absorptiometry. A total of 102 subjects completed the whole trial. The duration of total lactating time was 7·9 (sd 2·8) months on average. The intention-to-treat analysis yielded the following mean percentage changes in BMD for the whole body, the lumbar spine and the total left hip, respectively: −0·93 (sd 1·97), 2·11 (sd 4·90) and −1·60 (sd 2·65) % for the Low-Ca group; −0·56 (sd 1·89), 2·21 (sd 3·77) and −1·43 (sd 2·30) % for the Mid-Ca group; and −0·44 (sd 1·67), 2·32 (sd 4·66) and −0·95 (sd 4·08) % for the High-Ca group. The differences between the groups were not statistically significant (P: 0·5–0·9). The results of the complete case analysis were similar. In sum, we found no significant differences in the bone mineral changes from baseline to 12 months in postpartum lactating women consuming milk powder fortified with different levels of Ca.