Sherry Agellon

Effect of Different Dosages of Oral Vitamin D Supplementation on Vitamin D Status in Healthy, Breastfed Infants: A Randomized Trial

IMPORTANCE Vitamin D supplementation in infancy is required to support healthy bone mineral accretion. A supplement of 400 IU of vitamin D per day is thought to support plasma 25-hydroxyvitamin D (25[OH]D) concentrations between 40 and 50 nmol/L; some advocate 75 to 150 nmol/L for bone health. OBJECTIVE To investigate the efficacy of different dosages of vitamin D in supporting 25(OH)D concentrations in infants. DESIGN, SETTING, AND PARTICIPANTS Double-blind randomized clinical trial conducted among 132 one-month-old healthy, term, breastfed infants from Montréal, Québec, Canada, between March 2007 and August 2010. Infants were followed up for 11 months ending August 2011 (74% completed study). INTERVENTION Participants were randomly assigned to receive oral cholecalciferol (vitamin D3) supplements of 400 IU/d (n=39), 800 IU/d (n=39), 1200 IU/d (n=38), or 1600 IU/d (n=16). MAIN OUTCOMES AND MEASURES The primary outcome was a plasma 25(OH)D concentration of 75 nmol/L or greater in 97.5% of infants at 3 months. Secondary outcomes included 25(OH)D concentrations of 75 nmol/L or greater in 97.5% of infants at 6, 9, and 12 months; 25(OH)D concentrations of 50 nmol/L or greater across all times; growth; and whole body and regional bone mineral content. Data were analyzed by intention to treat using available data, logistic regression, and mixed-model analysis of variance. RESULTS By 3 months, 55% (95% CI, 38%-72%) of infants in the 400-IU/d group achieved a 25(OH)D concentration of 75 nmol/L or greater vs 81%(95% CI, 65%-91%) in the 800-IU/d group, 92% (95% CI, 77%-98%) in the 1200-IU/d group, and 100% in the 1600-IU/d group. This concentration was not sustained in 97.5% of infants at 12 months in any of the groups. The 1600-IU/d dosage was discontinued prematurely because of elevated plasma 25(OH)D concentrations. All dosages established 25(OH)D concentrations of 50 nmol/L or greater in 97% (95% CI, 94%-100%) of infants at 3 months and sustained this in 98% (95% CI, 94%-100%) to 12 months. Growth and bone mineral content did not differ by dosage. CONCLUSIONS AND RELEVANCE Among healthy, term, breastfed infants, only a vitamin D supplement dosage of 1600 IU/d (but not dosages of 400, 800, or 1200 IU/d) increased plasma 25(OH)D concentration to 75 nmol/L or greater in 97.5% of infants at 3 months. However, this dosage increased 25(OH)D concentrations to levels that have been associated with hypercalcemia. TRIAL REGISTRATION clinicaltrials.gov Identifier: NCT00381914.

Vitamin D Status in Montréal Preschoolers Is Satisfactory Despite Low Vitamin D Intake

The 2007 to 2009 Canadian Health Measures Survey reported vitamin D status in a representative sample of Canadians (6-79 y); however, children <6 y were not assessed. Our objective was to measure vitamin D intake from food and supplements, sun exposure, and biological vitamin D status of children ages 2 through 5 y in Montréal (latitude 45°N). Preschoolers (n = 508) were recruited between June 2010 and 2011 in a random sample of licensed daycares in the regions of greater Montréal, Canada in a cross-sectional study. The total plasma 25-hydroxyvitamin D [25(OH)D] concentration was measured using a chemiluminescence assay (Liaison, Diasorin). Dietary intake was assessed during one 24-h period plus a 30-d FFQ. Socioeconomic, demographic, anthropometry, and sun exposure data were collected. Plasma 25(OH)D was ≥50 nmol/L in 88% of children, whereas 49.4% had concentrations ≥75 nmol/L during the 1-y study. Almost 95% of preschoolers had vitamin D intakes less than the Estimated Average Requirement (EAR), and 4.8% of preschoolers ≤3.9 y and 25.9% of preschoolers ≥4 y had calcium intakes less than the EAR. Plasma 25(OH)D was different across age, income, sun index, milk intake, and dietary and supplemental vitamin D intake tertiles. Despite vitamin D intakes less than the EAR, the vitamin D status of Montréal preschoolers attending daycare is mostly satisfactory even in winter, suggesting that the EAR value is too high in the context of typical exogenous intakes of vitamin D in North America.

Methodological issues in assessing plasma 25-hydroxyvitamin D concentration in newborn infants.

BACKGROUND Although no gold standard exists, liquid chromatography tandem mass spectrometry (LC-MS/MS) is a precise and accurate method for the analysis of plasma 25-hydroxyvitamin D (25(OH)D). Immunoassays are more readily available and require small volume sampling, ideal for infant testing. The objective was to compare two commercially available immunoassays for measuring circulating 25(OH)D concentration in infant plasma against LC-MS/MS. METHODS Capillary blood samples from 103 infants were analyzed for plasma 25(OH)D using an enzyme immunoassay (EIA, Octeia, IDS Ltd.) and radioimmunoassay (RIA, DiaSorin). Plasma 25(OH)D(3), C-3 epimer of 25(OH)D(3) (3-epi-25(OH)D(3)) and 24,25-dihydroxyvitamin D (24,25(OH)(2)D(3)) were measured on the same samples using LC-MS/MS. To establish whether plasma 24,25(OH)(2)D(3) or 3-epi-25(OH)D(3) interferes with these immunoassay results, the zero 25(OH)D calibrator from each assay kit was spiked with increasing amounts of 24,25(OH)(2)D(3) or 3-epi-25(OH)D(3). RESULTS Classifying infants below the common vitamin D status targets of 50 nmol/L and 75 nmol/L respectively, 58% and 99% fell below using the RIA, 19% and 56% with the EIA and 31% and 76% with LC-MS/MS. Compared to LC-MS/MS, both immunoassays showed poor Bland-Altman limits of agreement for 25(OH)D concentrations (RIA: limits of agreement -27 to +13%; EIA: -12 to +41%), and mountain plots (folded cumulative distribution) depicted significant skew and bias. Spiked 24,25(OH)2D3 concentrations, but not 3-epi-25(OH)D3, appeared as >100% of known values on the EIA but not on the RIA thus, suggesting that the EIA may cross-react with 24,25(OH)(2)D(3) to a greater extent than 3-epi-25(OH)D(3). CONCLUSION Two common immunoassays resulted in very different classifications of vitamin D status possibly related to the interference of other vitamin D metabolites. Based on these data, LC-MS/MS assessment of vitamin D status is recommended in young infants (4-6 weeks of age).

Vitamin D supplementation trial in infancy: body composition effects at 3 years of age in a prospective follow-up study from Montréal.

The impact of vitamin D status on body composition is not well understood.

Vitamin D Status is Associated With Bone Mineral Density and Bone Mineral Content in Preschool-Aged Children

This study examined the associations between vitamin D status, bone mineral content (BMC), areal bone mineral density (aBMD), and markers of calcium homeostasis in preschool-aged children. Children (n=488; age range: 1.8-6.0 y) were randomly recruited from Montreal. The distal forearm was scanned using a peripheral dual-energy X-ray absorptiometry scanner (Lunar PIXI; GE Healthcare, Fairfield, CT). A subset (n=81) had clinical dual-energy X-ray absorptiometry (cDXA) scans (Hologic 4500A Discovery Series) of lumbar spine (LS) 1-4, whole body, and ultradistal forearm. All were assessed for plasma 25-hydroxyvitamin D [25(OH)D] and parathyroid hormone concentrations (Liaison; Diasorin), ionized calcium (ABL80 FLEX; Radiometer Medical A/S), and dietary vitamin D and calcium intakes by survey. Age (p<0.001) and weight-for-age Z-score (p<0.001) were positively associated with BMC and aBMD in all regression models, whereas male sex contributed positively to forearm BMC and aBMD. Having a 25(OH)D concentration of >75 nmol/L positively associated with forearm and whole body BMC and aBMD (p<0.036). Sun index related to (p<0.029) cDXA forearm and LS 1-4 BMC and whole-body aBMD. Nutrient intakes did not relate to BMC or aBMD. In conclusion, higher vitamin D status is linked to higher BMC and aBMD of forearm and whole body in preschool-aged children.

DHA and EPA in red blood cell membranes are associated with dietary intakes of omega-3-rich fish in healthy children

Omega-3 long-chain polyunsaturated fatty acids (n-3 LCPUFA) are important in child development. The primary objective of this study was to investigate the associations between dietary intakes of n-3 LCPUFA and red blood cell (RBC) n-3 LCPUFA in young children. Healthy children, (2-8y) underwent RBC fatty acid profiling. Dietary intakes were parent-reported over 6 mo using three 24h dietary intake assessments and three 30 d food frequency questionnaires (FFQ). Participants (n = 49, 5.6 ± 1.9y), were 59% male, and had a body mass index (BMI) z-score of 0.65 ± 0.84. Dietary n-3 LCPUFA intakes were not different over time. RBC docosahexaenoic acid (DHA) positively correlated with average DHA from the 24h recalls. RBC DHA and eicosapentaenoic acid (EPA) positively correlated with average n-3 LCPUFA-rich fish intake from the FFQ. RBC appear to reflect long-term stable intakes of n-3 LCPUFA during growth in healthy young children.

Orchidectomy-induced alterations in volumetric bone density, cortical porosity and strength of femur are attenuated by dietary conjugated linoleic acid in aged guinea pigs

Age-related osteoporosis and sarcopenia are ascribed in part to reductions in anabolic hormones. Dietary conjugated linoleic acid (CLA) improves lean and bone mass, but its impact during androgen deficiency is not known. This study tested if CLA would attenuate the effects of orchidectomy (ORX)-induced losses of bone and lean tissue. Male guinea pigs (n=40; 70-72 weeks), were randomized into four groups: (1) SHAM+Control diet, (2) SHAM+CLA diet, (3) ORX+Control diet, (4) ORX+CLA diet. Baseline blood sampling and dual-energy X-ray absorptiometry (DXA) scans were conducted, followed by surgery 4 days later with the test diets started 7 days after baseline sampling. Serial blood sampling and DXA scans were repeated 2, 4, 8 and 16 weeks on the test diets. Body composition and areal BMD (aBMD) of whole body, lumbar spine, femur and tibia were measured using DXA. At week 16, muscle protein fractional synthesis rate (FSR), volumetric BMD (vBMD), microarchitecture and bone strength were assessed. Body weight declined after SHAM and ORX surgery, with slower recovery in the ORX group. Dietary CLA did not affect weight or lean mass, but attenuated gains in fat mass. Lean mass was stable in SHAM and reduced in ORX by 2 weeks with whole body and femur bone mineral content (BMC) reduced by 4 weeks; CLA did not alter BMC. By week 16 ORX groups had lower free testosterone and myofibrillar FSR, yet higher cortisol, osteocalcin and ionized calcium with no alterations due to CLA. ORX+Control had higher prostaglandin E2 (PGE2) and total alkaline phosphatase compared to SHAM+Control whereas ORX+CLA were not different from SHAM groups. Femur metaphyseal vBMD was reduced in ORX+CTRL with the reduction attenuated by CLA. Femur cortical thickness (Ct.Th.) and biomechanical strength were reduced and cortical porosity (Ct.Po.) elevated by ORX and attenuated by CLA. This androgen deficient model with a sarcopenic-osteoporotic phenotype similar to aging men responded to dietary CLA with significant benefits to femur density and strength.

Depleted iron stores and iron deficiency anemia associated with reduced ferritin and hepcidin and elevated soluble transferrin receptors in a multiethnic group of preschool-age children

Iron deficiency anemia is prevalent in subgroups of the Canadian population. The objective of this study was to examine iron status and anemia in preschool-age children. Healthy children (n = 430, 2-5 years old, Montreal, Quebec, Canada) were sampled from randomly selected daycares. Anthropometry, demographics, and diet were assessed. Biochemistry included hemoglobin, ferritin, soluble transferrin receptors (sTfR), ferritin index, markers of inflammation (C-reactive protein, interleukin 6 (IL-6), and tumour necrosis factor alpha (TNFα)), and hepcidin. Iron deficiency and anemia cutoffs conformed to the World Health Organization criteria. Differences among categories were tested using mixed-model ANOVA or χ(2) tests. Children were 3.8 ± 1.0 years of age, with a body mass index z score of 0.48 ± 0.97, and 51% were white. Adjusted intakes of iron indicated <1% were at risk for deficiency. Hemoglobin was higher in white children, whereas ferritin was higher with greater age and female sex. Inflammatory markers and hepcidin did not vary with any demographic variable. The prevalence of iron deficiency was 16.5% (95% confidence interval (CI), 13.0-20.0). Three percent (95% CI, 1.4-4.6) of children had iron deficiency anemia and 12.8% (95% CI, 9.6-16.0) had unexplained anemia. Children with iron deficiency, with and without anemia, had lower plasma ferritin and hepcidin but higher sTfR, ferritin index, and IL-6, whereas those with unexplained anemia had elevated TNFα. We conclude that iron deficiency anemia is not very common in young children in Montreal. While iron deficiency without anemia is more common than iron deficiency with anemia, the correspondingly reduced circulating hepcidin would have enabled heightened absorption of dietary iron in support of erythropoiesis.

Are skeletally mature female rats a suitable model to study osteoporosis

OBJECTIVE To analyze if female Wistar rats at 56 weeks of age are a suitable model to study osteoporosis. MATERIALS AND METHODS Female rats with 6 and 36 weeks of age (n = 8 per group) were kept over a 20-week period and fed a diet for mature rodents complete in terms of Ca, phosphorous, and vitamin D. Excised femurs were measured for bone mass using dual-energy x-ray absorptiometry, morphometry, and biomechanical properties. The following serum markers of bone metabolism were analyzed: parathyroid hormone (PTH), osteocalcin (OC), osteoprotegerin (OPG), receptor activator of nuclear factor Κappa B ligand (RANKL), C-terminal peptides of type I collagen (CTX-I), total calcium, and alkaline phosphatase (ALP) activity. RESULTS Rats at 56 weeks of age showed important bone metabolism differences when compared with the younger group, such as, highest diaphysis energy to failure, lowest levels of OC, CTX-I, and ALP, and elevated PTH, even with adequate dietary Ca. CONCLUSION Rats at 26-week-old rats may be too young to study age-related bone loss, whereas the 56-week-old rats may be good models to represent the early stages of age-related changes in bone metabolism.

Dietary supplementation with long chain polyunsaturated fatty acids in pregnant guinea pigs has sex-dependent effects on growth and bone outcomes in offspring.

Long chain PUFA enhance bone mass in non-pregnant mammals. We examined the effects of arachidonic (AA; 20:4n-6) and docosahexaenoic (DHA; 22:6n-3) acid on bone mass of mothers and neonates. Guinea pig sows (n=15) were fed control, DHA or AA+DHA diets from mating to weaning. Measurements included: osteocalcin (OC), deoxypyridinoline (DPD), areal bone mineral density (aBMD) in sows and neonates; and volumetric density (vBMD) in neonates. Only vertebral aBMD and OC:DPD ratio declined during reproduction and only DHA reduced OC:DPD. Male pup weight was reduced by DHA and female weight elevated by AA+DHA. Whole body and femur aBMD were reduced by DHA and AA+DHA; whereas tibia vBMD was reduced by DHA in males. Female whole body, tibia and vertebrae aBMD plus tibia vBMD were elevated by AA+DHA; and DHA elevated whole body, tibia and vertebrae aBMD. Dietary AA+DHA and DHA elicit sex-dependent effects on neonatal bone, with minimal impact on mothers.