Helen M. Macdonald

Common genetic determinants of vitamin D insufficiency: a genome-wide association study

BACKGROUND Vitamin D is crucial for maintenance of musculoskeletal health, and might also have a role in extraskeletal tissues. Determinants of circulating 25-hydroxyvitamin D concentrations include sun exposure and diet, but high heritability suggests that genetic factors could also play a part. We aimed to identify common genetic variants affecting vitamin D concentrations and risk of insufficiency. METHODS We undertook a genome-wide association study of 25-hydroxyvitamin D concentrations in 33 996 individuals of European descent from 15 cohorts. Five epidemiological cohorts were designated as discovery cohorts (n=16 125), five as in-silico replication cohorts (n=9367), and five as de-novo replication cohorts (n=8504). 25-hydroxyvitamin D concentrations were measured by radioimmunoassay, chemiluminescent assay, ELISA, or mass spectrometry. Vitamin D insufficiency was defined as concentrations lower than 75 nmol/L or 50 nmol/L. We combined results of genome-wide analyses across cohorts using Z-score-weighted meta-analysis. Genotype scores were constructed for confirmed variants. FINDINGS Variants at three loci reached genome-wide significance in discovery cohorts for association with 25-hydroxyvitamin D concentrations, and were confirmed in replication cohorts: 4p12 (overall p=1.9x10(-109) for rs2282679, in GC); 11q12 (p=2.1x10(-27) for rs12785878, near DHCR7); and 11p15 (p=3.3x10(-20) for rs10741657, near CYP2R1). Variants at an additional locus (20q13, CYP24A1) were genome-wide significant in the pooled sample (p=6.0x10(-10) for rs6013897). Participants with a genotype score (combining the three confirmed variants) in the highest quartile were at increased risk of having 25-hydroxyvitamin D concentrations lower than 75 nmol/L (OR 2.47, 95% CI 2.20-2.78, p=2.3x10(-48)) or lower than 50 nmol/L (1.92, 1.70-2.16, p=1.0x10(-26)) compared with those in the lowest quartile. INTERPRETATION Variants near genes involved in cholesterol synthesis, hydroxylation, and vitamin D transport affect vitamin D status. Genetic variation at these loci identifies individuals who have substantially raised risk of vitamin D insufficiency. FUNDING Full funding sources listed at end of paper (see Acknowledgments).

Effect of Vitamin D Supplementation on Blood Pressure: A Systematic Review and Meta-analysis Incorporating Individual Patient Data

IMPORTANCE Low levels of vitamin D are associated with elevated blood pressure (BP) and future cardiovascular events. Whether vitamin D supplementation reduces BP and which patient characteristics predict a response remain unclear. OBJECTIVE To systematically review whether supplementation with vitamin D or its analogues reduce BP. DATA SOURCES We searched MEDLINE, CINAHL, EMBASE, Cochrane Central Register of Controlled Trials, and http://www.ClinicalTrials.com augmented by a hand search of references from the included articles and previous reviews. Google was searched for gray literature (ie, material not published in recognized scientific journals). No language restrictions were applied. The search period spanned January 1, 1966, through March 31, 2014. STUDY SELECTION We included randomized placebo-controlled clinical trials that used vitamin D supplementation for a minimum of 4 weeks for any indication and reported BP data. Studies were included if they used active or inactive forms of vitamin D or vitamin D analogues. Cointerventions were permitted if identical in all treatment arms. DATA EXTRACTION AND SYNTHESIS We extracted data on baseline demographics, 25-hydroxyvitamin D levels, systolic and diastolic BP (SBP and DBP), and change in BP from baseline to the final follow-up. Individual patient data on age, sex, medication use, diabetes mellitus, baseline and follow-up BP, and 25-hydroxyvitamin D levels were requested from the authors of the included studies. For trial-level data, between-group differences in BP change were combined in a random-effects model. For individual patient data, between-group differences in BP at the final follow up, adjusted for baseline BP, were calculated before combining in a random-effects model. MAIN OUTCOMES AND MEASURES Difference in SBP and DBP measured in an office setting. RESULTS We included 46 trials (4541 participants) in the trial-level meta-analysis. Individual patient data were obtained for 27 trials (3092 participants). At the trial level, no effect of vitamin D supplementation was seen on SBP (effect size, 0.0 [95% CI, -0.8 to 0.8] mm Hg; P=.97; I2=21%) or DBP (effect size, -0.1 [95% CI, -0.6 to 0.5] mm Hg; P=.84; I2=20%). Similar results were found analyzing individual patient data for SBP (effect size, -0.5 [95% CI, -1.3 to 0.4] mm Hg; P=.27; I2=0%) and DBP (effect size, 0.2 [95% CI, -0.3 to 0.7] mm Hg; P=.38; I2=0%). Subgroup analysis did not reveal any baseline factor predictive of a better response to therapy. CONCLUSIONS AND RELEVANCE Vitamin D supplementation is ineffective as an agent for lowering BP and thus should not be used as an antihypertensive agent.

Vitamin D3 supplementation has no effect on conventional cardiovascular risk factors: a parallel-group, double-blind, placebo-controlled RCT.

CONTEXT Observational studies show an association between low vitamin D status assessed by circulating 25-hydroxyvitamin D and cardiovascular events and mortality. Data from randomized controlled trials are limited. OBJECTIVE The aim of this study was to test whether daily doses of vitamin D(3) at 400 or 1000 IU/d for 1 yr affected conventional markers of cardiovascular disease (CVD) risk. DESIGN We conducted a parallel-group, double-blind, placebo-controlled randomized controlled trial. Randomization was computer generated. Participants and study investigators were blinded to intervention groupings throughout the trial. SETTING The study was conducted at the Clinical Research Facility, University of Aberdeen, United Kingdom. PARTICIPANTS A total of 305 healthy postmenopausal women aged 60-70 yr were recruited for the study. INTERVENTION Each woman received a daily capsule of 400 or 1000 IU vitamin D(3) or placebo randomly allocated. MAIN OUTCOME MEASURES Primary outcomes were serum lipid profile [total, high-density lipoprotein, and low-density lipoprotein cholesterol; triglycerides; and apolipoproteins A-1 and B100], insulin resistance (homeostatic model assessment), inflammatory biomarkers (high-sensitivity C-reactive protein, IL-6, soluble intracellular adhesion molecule-1), and blood pressure. RESULTS A total of 265 (87%) participants completed all study visits. Small differences between groups for serum apolipoprotein B100 change [repeated measures ANOVA, P=0.04; mean (sd), -1.0 (10.0) mg/dl (400 IU); -1.0 (10.0) mg/dl (1000 IU); and +0.02 (10.0) mg/dl (placebo)] were not considered clinically significant. Other systemic markers for CVD risk remained unchanged. There was significant seasonal variation in systolic and diastolic blood pressure independent of vitamin D dose (P<0.001, linear mixed model). Mean (sd) reduction in systolic blood pressure from winter to summer was -6.6 (10.8) mm Hg. CONCLUSIONS Improving vitamin D status through dietary supplementation is unlikely to reduce CVD risk factors. Confounding of seasonality should be recognized and addressed in future studies of vitamin D.

Vitamin D status in postmenopausal women living at higher latitudes in the UK in relation to bone health, overweight, sunlight exposure and dietary vitamin D

For 5 months a year the UK has insufficient sunlight for cutaneous synthesis of vitamin D and winter requirements are met from stores made the previous summer. Although there are few natural dietary sources, dietary intake may help maintain vitamin D status. We investigated the relationship between 25-hydroxyvitamin D (25(OH)D), bone health, overweight, sunlight exposure and dietary vitamin D in 3113 women (age 54.8 [SD 2.3] years) living at latitude 57 degrees N between 1998-2000. Serum 25(OH)D was measured by high performance liquid chromatography (HPLC), dietary intakes (food frequency questionnaire, n=2598), sunlight exposure (questionnaire, n=2402) and bone markers were assessed. Bone mineral density (BMD) was measured by dual x-ray absorptiometry in all women at the sampling visit and 6 years before. Seasonal variation in 25(OH)D was not substantial with a peak in the autumn (23.7 [9.9] ng/ml) and a nadir in spring (19.7 [7.6] ng/ml). Daily intake of vitamin D was 4.2 [2.5] mug from food only and 5.8 [4.0] mug including vitamin D from cod liver oil and multivitamins. The latter was associated with 25(OH)D at each season whereas vitamin D simply from food was associated with 25(OH)D in winter and spring only. Sunlight exposure was associated with 25(OH)D in summer and autumn. 25(OH)D was negatively associated with increased bone resorption and bone loss (P<0.05) remaining significant after adjustment for confounders (age, weight, height, menopausal status/HRT use, physical activity and socio-economic status). Using an insufficiency cut-off of <28 ng/ml 25(OH)D, showed lower concentrations of bone resorption markers in the upper category (fDPD/Cr 5.1 [1.7] nmol/mmol compared to 5.3 [2.1] nmol/mmol, P=0.03) and no difference in BMD or bone loss. 25(OH)D was lower (P<0.01) and parathyroid hormone higher (P<0.01) in the top quintile of body mass index. In conclusion, low vitamin D status is associated with greater bone turnover, bone loss and obesity. Diet appears to attenuate the seasonal variation of vitamin D status in early postmenopausal women at northerly latitude where quality of sunlight for production of vitamin D is diminished.

Large-Scale Population-Based Study Shows No Evidence of Association Between Common Polymorphism of the VDR Gene and BMD in British Women

The VDR is a candidate gene for osteoporosis. Here we studied five common polymorphisms of VDR in relation to calcium intake and vitamin D status in a population‐based cohort of 3100 British women, but found no significant association with bone mass, bone loss, or fracture.

Dietary patterns, bone resorption and bone mineral density in early post-menopausal Scottish women

Background/Objectives:Several nutrients affect bone turnover. Dietary patterns may provide insights into which foods are important and how nutrition affects bone health. The aim of this study was to investigate the associations between dietary patterns, bone turnover and bone mineral density (BMD).Subjects/Methods:This cross-sectional study examined 3236 Scottish women age 50–59 years, who were members of the Aberdeen Prospective Osteoporosis Screening Study. They had hip and spine BMD measurements (dual-energy X-ray absorptiometry) and provided samples for bone turnover markers. Diet was assessed by a validated food frequency questionnaire encompassing 98 foods, from which 35 food groups were systematically created. Dietary patterns were defined by principal components analysis. The bone measures were regressed onto the dietary pattern and adjusted for potential confounders.Results:Five dietary patterns were identified, three of which were associated with bone health. The ‘healthy’ pattern was associated with decreased bone resorption (r=0.081, P<0.001). Two other patterns (processed foods and snack food) were associated with lower BMD (femoral neck r=−0.056, r=−0.044, P<0.001, respectively).Conclusions:Dietary pattern may influence bone turnover and BMD. A healthy dietary pattern with high intakes of fruit and vegetables may lead to less bone resorption, and a poor dietary pattern rich in processed foods is associated with a decrease in BMD. This study confirms that a healthy diet is required for strong bones, and highlights that a nutrient-poor diet is a risk factor for osteoporosis.

UK Food Standards Agency Workshop Report: an investigation of the relative contributions of diet and sunlight to vitamin D status.

The UK Food Standards Agency (FSA) convened an international group of scientific experts to review three Agency-funded projects commissioned to provide evidence for the relative contributions of two sources, dietary vitamin D intake and skin exposure to UVB rays from sunlight, to vitamin D status. This review and other emerging evidence are intended to inform any future risk assessment undertaken by the Scientific Advisory Committee on Nutrition. Evidence was presented from randomised controlled trials to quantify the amount of vitamin D required to maintain a serum 25-hydroxy vitamin D (25OHD) concentration >25 nmol/l, a threshold that is regarded internationally as defining the risk of rickets and osteomalacia. Longitudinal evidence was also provided on summer sunlight exposure required to maintain 25OHD levels above this threshold in people living in the British Isles (latitude 51 degrees-57 degrees N). Data obtained from multi-level modelling of these longitudinal datasets showed that UVB exposure (i.e. season) was the major contributor to changes in 25OHD levels; this was a consistent finding in two Caucasian groups in the north and south of the UK, but was less apparent in the one group of British women of South Asian origin living in the south of the UK. The FSA-funded research suggested that the typical daily intake of vitamin D from food contributed less than UVB exposure to average year-round 25OHD levels in both Caucasian and Asian women. The low vitamin D status of Asian women has been acknowledged for some time, but the limited seasonal variation in Asian women is a novel finding. The Workshop also considered the dilemma of balancing the risks of vitamin D deficiency (from lack of skin exposure to sunlight in summer) and skin cancer (from excessive exposure to sunlight with concomitant sunburn and erythema). Cancer Research UK advises that individuals should stay below their personal sunburn threshold to minimise their skin cancer risk. The evidence suggests that vitamin D can be produced in summer at the latitude of the UK, with minimal risk of erythema and cell damage, by exposing the skin to sunlight for a short period at midday, when the intensity of UVB is at its daily peak. The implications of the new data were discussed in the context of dietary reference values for vitamin D for the general population aged 4-64 years. Future research suggestions included further analysis of the three FSA-funded studies as well as new research.

ASSOCIATION OF OESTROGEN RECEPTOR ALPHA GENE POLYMORPHISMS WITH POSTMENOPAUSAL BONE LOSS, BONE MASS, AND QUANTITATIVE ULTRASOUND PROPERTIES OF BONE

Background: The gene encoding oestrogen receptor α (ESR1) appears to regulate bone mineral density (BMD) and other determinants of osteoporotic fracture risk. Objective: To investigate the relation between common polymorphisms and haplotypes of the ESR1 gene and osteoporosis related phenotypes in a population based cohort of 3054 Scottish women. Results: There was a significant association between a common haplotype “px”, defined by the PvuII andXbaI restriction fragment length polymorphisms within intron 1 of the ESR1 gene, and femoral neck bone loss in postmenopausal women who had not received hormone replacement therapy (n = 945; p = 0.009). Annual rates of femoral neck bone loss were ∼14% higher in subjects who carried one copy of px and 22% higher in those who carried two copies, compared with those who did not carry the px haplotype. The px haplotype was associated with lower femoral neck BMD in the postmenopausal women (p = 0.02), and with reduced calcaneal broadband ultrasound attenuation (BUA) values in the whole study population (p = 0.005). There was no association between a TA repeat polymorphism in the ESR1 promoter and any phenotype studied, though on long range haplotype analysis subjects with a smaller number of TA repeats who also carried the px haplotype had reduced BUA values. Conclusions: The ESR1px haplotype is associated with reduced hip BMD values and increased rates of femoral neck bone loss in postmenopausal women. An association with BUA may explain the fact that ESR1 intron 1 alleles predict osteoporotic fractures by a mechanism partly independent of differences in BMD.

COL1A1 Sp1 Polymorphism Predicts Perimenopausal and Early Postmenopausal Spinal Bone Loss

Genetic factors play an important role in the pathogenesis of osteoporosis but the genes that determine susceptibility to poor bone health are defined incompletely. Previous work has shown that a polymorphism that affects an Sp1 binding site in the COL1A1 gene is associated with reduced bone mineral density (BMD) and an increased risk of osteoporotic fracture in several populations. Data from cross‐sectional studies have indicated that COL1A1 Sp1 alleles also may be associated with increased rates of bone loss with age, but longitudinal studies, which have examined bone loss in relation to COL1A1> genotype, have yielded conflicting results. In this study, we examined the relationship between COL1A1 Sp1 alleles and early postmenopausal bone loss measured by dual‐energy X‐ray absorptiometry (DXA) in a population‐based cohort of 734 Scottish women who were followed up over a 5‐ to 7‐year period. The distribution of genotypes was as expected in a white population with 484 “SS” homozygotes (65.9%); 225 “Ss” heterozygotes (30.7%), and 25 “ss” homozygotes (3.4%). Women taking hormone‐replacement therapy (HRT; n = 239) had considerably reduced rates of bone loss at the spine (−0.40 ± 0.06%/year) and hip (−0.56 ± 0.06%/year) when compared with non‐HRT users (n = 352; spine, −1.36 ± 0.06%/year; hip, −1.21 ± 0.05%/year; p < 0.001 for both sites). There was no significant difference in baseline BMD values at the lumbar spine (LS) or femoral neck (FN) between genotypes or in the rates of bone loss between genotypes in HRT users. However, in non‐HRT users (n = 352), we found that ss homozygotes (n = 12) lost significantly more bone at the lumbar site than the other genotype groups in which ss = −2.26 ± 0.31%/year compared with SS = −1.38 ± 0.07%/year and Ss = −1.22 ± 0.10%/year (p = 0.004; analysis of variance [ANOVA]) and a similar trend was observed at the FN in which ss = −1.78 ± 0.19%/year compared with SS = −1.21 ± 0.06%/year and Ss = −1.16 ± 0.08%/year (p = 0.06; ANOVA). The differences in spine BMD loss remained significant after correcting for confounding factors. Stepwise multiple regression analysis showed that COL1A1 genotype independently accounted for a further 3.0% of the variation in spine BMD change after age (4.0%), weight (5.0%), and baseline BMD (2.8%). We conclude that women homozygous for the Sp1 polymorphism are at significantly increased risk of excess rates of bone loss at the spine, but this effect may be nullified by the use of HRT.

Associations between dietary flavonoid intakes and bone health in a scottish population

Flavonoids are bioactive polyphenols found particularly in fruit and vegetables, but little is known about their role in bone health in humans. The aim of this observational study was to investigate whether dietary flavonoid intake was associated with bone mineral density (BMD) and bone resorption in a large group of perimenopausal Scottish women. Over 3000 women completed a food frequency questionnaire as part of an osteoporosis screening study. The diets were analyzed for flavonoid intake using a food composition database. BMD was measured at the femoral neck (FN) and lumbar spine (LS) by dual‐energy X‐ray absorptiometry (DXA). Free pyridinoline (PYD) and deoxypyridinoline (DPD) were measured by high‐performance liquid chromatography (HPLC) in second early morning fasted urine samples. The mean flavonoid intake of the diet was 307 ±199 mg/d. The catechin family contributed the most to flavonoid intakes (55%), and the flavones the least (<1%). Associations were found between energy‐adjusted total flavonoid intakes and BMD at the FN and LS (FN r = 0.054, LS r = 0.036, p ≤ .05). Annual percent change in BMD was associated with intakes of procyanidins and catechins (p ≤ .05), and flavanones were negatively associated with bone‐resorption markers (PYD r = −0.049, DPD r = –0.057, p ≤ .001). These associations were still seen after adjusting for confounders. It is concluded that dietary flavonoid intakes are associated with BMD, supporting the evidence from animal and cellular studies.