Vitamin D status of 3-year-old children in Denmark: determinants and associations with bone mineralisation and blood lipids.

PURPOSE: Low vitamin D status is a global problem and has been associated with reduced skeletal and cardiometabolic health. However, evidence in young children is lacking. We, therefore, aimed to characterise vitamin D status in toddlers, identify its determinants, and explore if vitamin D status was associated with bone mineralisation and lipid profile. METHODS: We used cross-sectional data from 3-year-old children (n = 323) living in Denmark (latitude: 55°N). Bone mineralisation (n = 108) was measured by DXA. Blood samples were analysed for serum 25-hydroxyvitamin D (s-25(OH)D) by LC-MS/MS, triacylglycerol, and total, low- and high density lipoprotein cholesterol. RESULTS: Mean ± SD s-25(OH)D was 69 ± 23 nmol/L, but varied with season. During winter, 38% had inadequate s-25(OH)D (< 50 nmol), whereof 15% had deficiency (< 30 nmol/L); these numbers were only 7 and 1% during summer. In terms of status determinants, supplement use (66% were users) was associated with s-25(OH)D (P < 0.001), whereas dietary vitamin D intake (median [25-75th percentile] of 1.3 [0.9-1.9] µg/d), sex, parental education, BMI, and physical activity were not. There were no associations between s-25(OH)D and blood lipids or bone measurements, using either unadjusted or adjusted regression models. CONCLUSION: More than 1/3 of Danish toddlers had inadequate vitamin D intake during winter, but acceptable mean vitamin D status. In addition to season, supplement use was the main determinant of vitamin D status, which was, however, not associated with bone mineralisation or lipid profile. The results support recommendations of vitamin D supplements during winter at northern latitudes, but potential health effects need further investigation.

Vitamin D supplementation and increased dairy protein intake do not affect muscle strength or physical function in healthy 6-8-year-old children: the D-pro randomized trial.

PURPOSE: To investigate separate and combined effects of vitamin D supplementation during the extended winter and increased dairy protein intake on muscle strength and physical function in children, and furthermore to explore potential sex differences. METHODS: In a 2 × 2-factorial, randomized winter trial, 183 healthy, 6-8-year-old children received blinded tablets with 20 µg/day vitamin D3 or placebo, and substituted 260 g/day dairy with yogurts with high (HP, 10 g protein/100 g) or normal protein content (NP, 3.5 g protein/100 g) for 24 weeks during winter at 55° N. We measured maximal isometric handgrip and leg press strength, and physical function by jump tests and a 30 s sit-to-stand test. Physical activity was measured by 7-day accelerometry. RESULTS: Baseline (mean ± SD) serum 25-hydroxyvitamin D was 80.8 ± 17.2 nmol/L, which increased to 88.7 ± 17.6 nmol/L with vitamin D supplementation and decreased to 48.4 ± 19.2 nmol/L with placebo. Baseline protein intake was 15.5 ± 2.4 E%, which increased to 18.4 ± 3.4 E% with HP and was unchanged with NP. We found no separate or combined effects of vitamin D supplementation and/or increased dairy protein intake on muscle strength or physical function (all P > 0.20). There was an interaction on the sit-to-stand test (Pvitamin×yogurt = 0.02), which however disappeared after adjusting for physical activity (P = 0.16). Further, vitamin D supplementation increased leg press strength relatively more in girls compared to boys (mean [95% CI] 158 [17, 299] N; Pvitamin×sex = 0.047). CONCLUSION: Overall, vitamin D and dairy protein supplementation during the extended winter did not affect muscle strength or physical function in healthy children. Potential sex differences of vitamin D supplementation should be investigated further. REGISTERED AT CLINICALTRIALS.GOV: NCT0395673.

Wholegrain intake, growth and metabolic markers in Danish infants and toddlers: a longitudinal study.

PURPOSE: Wholegrain intake is linked to lower risk of lifestyle diseases, but little is known about its role in growth and metabolic health during the first years of life. We characterized wholegrain and dietary fibre intake in 439 Danish children at 9 and 36 months of age and explored associations with height z-scores (HAZ), body mass index z-scores (BMIZ) and metabolic markers. METHODS: We used pooled data from two infant cohorts and estimated intakes of total wholegrain, dietary fibre and wholegrain subtypes from 7-day dietary records. Associations with HAZ, BMIZ and non-fasting plasma low-density (LDLC) and high-density-lipoprotein cholesterol, triacylglycerol, insulin and glucose were analysed in mixed models, adjusted for potential confounders. RESULTS: Median (25th, 75th percentile) wholegrain intake was 7.5 (4.9, 10.5) and 6.5 (4.6, 9.0) g/MJ at 9 and 36 months. Neither wholegrain nor dietary fibre intake were associated with HAZ (P ≥ 0.09). At 36 months, wholegrain intake was inversely associated with LDLC (P = 0.05) and directly with glucose (P < 0.001). In secondary analyses, wholegrain rye was inversely associated with glucose at 9 months and insulin at 36 months (both P ≤ 0.03). Oat and wheat wholegrain were directly associated with glucose (both P ≤ 0.01) and wheat with BMIZ (P = 0.02) at 36 months. CONCLUSION: Danish infants and toddlers have high intakes of wholegrain and dietary fibre, with no indication of compromised growth. In line with studies in adults, wholegrain intake was inversely associated with LDLC. The observed direct association between wholegrain intake and plasma glucose and associations with wholegrain subtypes should be investigated further.

Effects of high dairy protein intake and vitamin D supplementation on body composition and cardiometabolic markers in 6-8-y-old children-the D-pro trial.

BACKGROUND: Increasing evidence suggests that prevention of lifestyle diseases should begin early. Dairy protein and vitamin D can affect body composition and cardiometabolic markers, yet evidence among well-nourished children is sparse. OBJECTIVES: We investigated combined and separate effects of high dairy protein intake and vitamin D on body composition and cardiometabolic markers in children. METHODS: In a 2 × 2-factorial, randomized trial, 200 white, Danish, 6-8-y-old children substituted 260 g/d dairy in their diet with high-protein (HP; 10 g protein/100 g) or normal-protein (NP; 3.5 g protein/100 g) yogurt and received blinded tablets with 20 µg/d vitamin D3 or placebo for 24 wk during winter. We measured body composition (by DXA), blood pressure, and fasting blood glucose, insulin, C-peptide, and lipids. RESULTS: In total, 184 children (92%) completed the study. Baseline median (25th-75th percentile) dairy protein intake was median: 3.7 (25th-75th percentile: 2.5-5.1) energy percentage (E%) and increased to median: 7.2 (25th-75th percentile: 4.7-8.8) E% and median: 4.2 (25th-75th percentile: 3.1-5.3) E% with HP and NP. Mean ± SD serum 25-hydroxyvitamin D concentration changed from 81 ± 17 to 89 ± 18 nmol/L and 48 ± 13 nmol/L with vitamin D and placebo, respectively. There were no combined effects of dairy protein and vitamin D, except for plasma glucose, with the largest increase in the NP-vitamin D group (Pinteraction = 0.005). There were smaller increases in fat mass index (P = 0.04) with HP than with NP, and the same pattern was seen for insulin, HOMA-IR, and C-peptide (all P = 0.06). LDL cholesterol was reduced with vitamin D compared with placebo (P < 0.05). Fat-free mass and blood pressure were unaffected. CONCLUSIONS: High compared with normal dairy protein intake hampered an increase in fat mass index. Vitamin D supplementation counteracted the winter decline in 25-hydroxyvitamin D and the increase in LDL cholesterol observed with placebo. This study adds to the sparse evidence on dairy protein in well-nourished children and supports a vitamin D intake of ∼20 µg/d during winter. This trial was registered at clinicaltrials.gov as NCT03956732.

Effects of vitamin D and high dairy protein intake on bone mineralization and linear growth in 6- to 8-year-old children: the D-pro randomized trial.

BACKGROUND: Vitamin D and dairy protein may stimulate bone mineralization and linear growth in children, but previous studies show inconsistent results and have not examined their combined effects. OBJECTIVES: To investigate combined and separate effects of vitamin D supplementation and high-protein (HP) compared with normal-protein (NP) yogurt intake on children's bone mineralization and linear growth. METHODS: In a 2 × 2-factorial trial, 200 healthy, 6- to 8-year-old, Danish, children with light skin (55°N) were randomized to 20 µg/d vitamin D3 or placebo and to substitute 260 g/d dairy with HP (10 g protein/100 g) or NP (3.5 g protein/100 g) yogurt for 24 weeks during an extended winter. Outcomes were total body less head (TBLH) and lumbar spine bone mineral density (BMD), bone mineral content (BMC), and bone area (BA) by dual-energy X-ray absorptiometry, height, and biomarkers of bone turnover and growth. The primary outcome was TBLH BMD. RESULTS: In total, 184 children (92%) completed the study. The baseline serum 25-hydroxyvitamin D was 80.8 ± 17.2 nmol/L, which increased by 7.2 ± 14.1 nmol/L and decreased by 32.3 ± 17.5 nmol/L with vitamin D and placebo, respectively. The baseline protein intake was 15.4 ± 2.4 energy percentage (E%), which increased to 18.3 ± 3.4 E% with HP. There were no vitamin D-yogurt interactions and no main effects of either intervention on TBLH BMD. However, vitamin D supplementation increased lumbar spine BMD and TBLH BMC compared to placebo, whereas HP groups showed lower increments in lumbar spine BMD, TBLH BMC and BA, and plasma osteocalcin compared to NP groups. Height, growth factors, and parathyroid hormone levels were unaffected. CONCLUSIONS: Although there were no effects on whole-body BMD, vitamin D increased bone mass and spinal BMD, whereas high compared with normal dairy protein intake had smaller incremental effects on these outcomes. This supports a recommended vitamin D intake of around 20 µg/d during winter but not use of HP dairy products for improved bone mineralization among healthy, well-nourished children. This trial was registered at clinicaltrials.gov as NCT03956732.

The effect of milk and rapeseed protein on growth factors in 7-8 year-old healthy children - A randomized controlled trial.

OBJECTIVE: Milk protein may stimulate linear growth through insulin-like growth factor-1 (IGF-1). However, the effect of plant proteins on growth factors is largely unknown. This study assesses the effect of combinations of milk and rapeseed protein versus milk protein alone on growth factors in children. DESIGN: An exploratory 3-armed randomized, double-blind, controlled trial was conducted in 129 healthy 7-8 year-old Danish children. Children received 35 g milk and rapeseed protein (ratio 54:46 or 30:70) or 35 g milk protein per day for 4 weeks. The primary outcome was difference in IGF-1 changes between intervention groups after 4 weeks. Secondary outcomes included changes in IGF-1 after 1 week and changes in insulin-like growth factor binding protein-3 (IGFBP-3), IGF-1/IGFBP-3, insulin, height, weight and body composition after 1 and 4 weeks. Results were analysed by multiple linear mixed-effect models. RESULTS: There were no differences in changes of plasma IGF-1, insulin-like growth factor binding protein-3 (IGFBP-3), IGF-1/IGFBP-3 ratio or insulin between groups after 1 or 4 weeks based on 89 complete cases (P > 0.10). IGF-1 increased by 13.7 (95% CI 9.7;17.7) ng/mL and 18.0 (14.0;22.0) ng/mL from baseline to week 1 and 4, respectively, a 16% increase during the intervention. Similarly, insulin increased by 31% (14; 50) and 33% (16; 53) from baseline to week 1 and 4. Fat-free mass index (FFMI) increments were higher with milk alone than rapeseed blends (P < 0.05), coinciding with a trend towards a lower height increment. Body mass index increased within all groups (P < 0.05), mainly due to an increase in FFMI (P < 0.01). CONCLUSION: There were no differences in changes of growth factors between the combinations of milk and rapeseed protein and milk protein alone in healthy, well-nourished children with a habitual intake of milk. Within groups, growth factors increased considerably. Future studies are needed to investigate how intakes of plant and animal proteins affect childhood growth.

Associations between body mass index and height during childhood and adolescence and the risk of coronary heart disease in adulthood: A systematic review and meta-analysis.

Body mass index (BMI) at child and adolescent ages is positively associated with adult coronary heart disease (CHD) whereas height at these ages may be inversely associated with CHD. However, potential effects of age, sex, and socioeconomic status on associations between BMI and CHD are less investigated. We conducted a systematic review and meta-analysis of BMI and height at ages 2-19 years in relation to adult CHD and examined effects of age, sex, socioeconomic status, and other factors. Twenty-two studies on BMI and five on height were included, comprising 5,538,319 individuals and 69,830 CHD events. Random effects meta-analyses were conducted. Child and adolescent BMI were positively associated with CHD (hazard ratio = 1.12; 95% confidence interval [CI] [1.01, 1.25] per standard deviation [SD]), and categorical analyses supported these findings. The associations did not significantly differ by age, sex, or by adjustment for socioeconomic status. Child and adolescent height were inversely associated with CHD (hazard ratio = 0.87; 95% CI [0.81, 0.93] per SD), and categorical analyses agreed. Insufficient studies on height precluded subgroup analyses. Heterogeneity was generally high in all analyses. We found that BMI in youth is positively associated with adult CHD regardless of sex or adjustment for socioeconomic status whereas height is inversely associated with later risk of CHD.

Vitamin D supplementation to prevent acute respiratory infections: a systematic review and meta-analysis of aggregate data from randomised controlled trials.

BACKGROUND: A 2017 meta-analysis of data from 25 randomised controlled trials (RCTs) of vitamin D supplementation for the prevention of acute respiratory infections (ARIs) revealed a protective effect of this intervention. We aimed to examine the link between vitamin D supplementation and prevention of ARIs in an updated meta-analysis. METHODS: For this systematic review and meta-analysis, we searched MEDLINE, Embase, the Cochrane Central Register of Controlled Trials, Web of Science, and the ClinicalTrials.gov registry for studies listed from database inception to May 1, 2020. Double-blind RCTs of vitamin D3, vitamin D2, or 25-hydroxyvitamin D (25[OH]D) supplementation for any duration, with a placebo or low-dose vitamin D control, were eligible if they had been approved by a research ethics committee, and if ARI incidence was collected prospectively and prespecified as an efficacy outcome. Studies reporting results of long-term follow-up of primary RCTs were excluded. Aggregated study-level data, stratified by baseline 25(OH)D concentration and age, were obtained from study authors. Using the proportion of participants in each trial who had one or more ARIs, we did a random-effects meta-analysis to obtain pooled odds ratios (ORs) and 95% CIs to estimate the effect of vitamin D supplementation on the risk of having one or more ARIs (primary outcome) compared with placebo. Subgroup analyses were done to estimate whether the effects of vitamin D supplementation on the risk of ARI varied according to baseline 25(OH)D concentration (<25 nmol/L vs 25·0-49·9 nmol/L vs 50·0-74·9 nmol/L vs >75·0 nmol/L), vitamin D dose (daily equivalent of <400 international units [IU] vs 400-1000 IU vs 1001-2000 IU vs >2000 IU), dosing frequency (daily vs weekly vs once per month to once every 3 months), trial duration (≤12 months vs >12 months), age at enrolment (<1·00 years vs 1·00-15·99 years vs 16·00-64·99 years vs ≥65·00 years), and presence versus absence of airway disease (ie, asthma only, COPD only, or unrestricted). Risk of bias was assessed with the Cochrane Collaboration Risk of Bias Tool. The study was registered with PROSPERO, CRD42020190633. FINDINGS: We identified 1528 articles, of which 46 RCTs (75 541 participants) were eligible. Data for the primary outcome were obtained for 48 488 (98·1%) of 49 419 participants (aged 0-95 years) in 43 studies. A significantly lower proportion of participants in the vitamin D supplementation group had one or more ARIs (14 332 [61·3%] of 23 364 participants) than in the placebo group (14 217 [62·3%] of 22 802 participants), with an OR of 0·92 (95% CI 0·86-0·99; 37 studies; I2=35·6%, pheterogeneity=0·018). No significant effect of vitamin D supplementation on the risk of having one or more ARIs was observed for any of the subgroups defined by baseline 25(OH)D concentration. However, protective effects of supplementation were observed in trials in which vitamin D was given in a daily dosing regimen (OR 0·78 [95% CI 0·65-0·94]; 19 studies; I2=53·5%, pheterogeneity=0·003), at daily dose equivalents of 400-1000 IU (0·70 [0·55-0·89]; ten studies; I2=31·2%, pheterogeneity=0·16), for a duration of 12 months or less (0·82 [0·72-0·93]; 29 studies; I2=38·1%, pheterogeneity=0·021), and to participants aged 1·00-15·99 years at enrolment (0·71 [0·57-0·90]; 15 studies; I2=46·0%, pheterogeneity=0·027). No significant interaction between allocation to the vitamin D supplementation group versus the placebo group and dose, dose frequency, study duration, or age was observed. In addition, no significant difference in the proportion of participants who had at least one serious adverse event in the vitamin supplementation group compared with the placebo group was observed (0·97 [0·86-1·07]; 36 studies; I2=0·0%, pheterogeneity=0·99). Risk of bias within individual studies was assessed as being low for all but three trials. INTERPRETATION: Despite evidence of significant heterogeneity across trials, vitamin D supplementation was safe and overall reduced the risk of ARI compared with placebo, although the risk reduction was small. Protection was associated with administration of daily doses of 400-1000 IU for up to 12 months, and age at enrolment of 1·00-15·99 years. The relevance of these findings to COVID-19 is not known and requires further investigation. FUNDING: None.

Exploring the effects of oily fish consumption on measures of acute and long-term stress in healthy 8-9-year-old children: the FiSK Junior randomised trial.

Long-chain n-3 PUFA (n-3 LCPUFA) are known to reduce blood pressure (BP), heart rate and vagal tone, but potential stress-mitigating effects of n-3 LCPUFA are not well investigated. We explored the effects of oily fish consumption on long-term stress and the stress response in schoolchildren. Healthy 8-9-year-old children were randomised to receive about 300 g/week of oily fish or poultry for 12 weeks (199 randomised, 197 completing). At baseline and endpoint, we measured erythrocyte n-3 LCPUFA, hair cortisol and the response to a 1-min cold pressor test (CPT) on saliva cortisol, BP and continuous electrocardiogram recordings. Post-intervention hair cortisol did not differ between the groups, but sex-specificity was indicated (Psex × group = 0·074, boys: -0·9 (95 % CI -2·9, 1·0) ng/g, girls: 0·7 (95 % CI -0·2, 1·6) ng/g). Children in the fish group tended to be less prone to terminate CPT prematurely (OR 0·20 (95 % CI 0·02, 1·04)). Mean heart beat interval during CPT was 18·2 (95 % CI 0·3, 36·6) ms longer and high frequency power increased (159 (95 % CI 29, 289) ms2) in the fish v. poultry group. The cardiac autonomic response in the 10 min following CPT was characterised by a sympathetic peak followed by a parasympathetic peak, which was most pronounced in the fish group. This exploratory study does not support a strong effect of oily fish consumption on stress but indicates that oily fish consumption may increase vagal cardiac tone during the physiological response to CPT. These results warrant further investigation.

Sleep and physical activity in healthy 8-9-year-old children are affected by oily fish consumption in the FiSK Junior randomized trial.

PURPOSE: Studies indicate that long-chain n-3 PUFA (n-3LCPUFA) affect sleep and physical activity (PA) in childhood. However, few studies used objective tools and none studies examined the effect of fish per se. We aimed to explore if fish consumption affected sleep and PA assessed by accelerometry in children, and if effects were modified by sex. METHODS: In a randomized 12-week trial, 199 healthy 8-9-year-old children received ~ 300 g/week of oily fish or poultry. Sleep and PA were pre-specified explorative outcomes examined by accelerometers that the children wore on their hip for 7 days at baseline and endpoint, while parents registered sleep. Compliance was verified by erythrocyte n-3LCPUFA. RESULTS: The children slept 9.4 ± 0.5 h/night but the sleep duration variability across the week was 6.0 (95%CI: 0.8, 11.1) min lower in the fish vs poultry group. Furthermore, children in the fish group exhibited increased spare time sedentary activity [9.4 (95%CI: 1.8, 16.9) min/day] at the expense of light PA [- 8.2 (95%CI: - 14.4, - 2.0) min/day]. These effects were supported by dose-dependency with n-3LCPUFA. Additionally, latency to sleep onset was reduced by 3.6 (95%CI: 1.0, 6.3) min on weekends and moderate-vigorous PA during school hours was 3.5 (95%CI: 0.1, 6.8) min longer in fish vs poultry. P values for sex interactions were all > 0.05 but the effects tended to be most pronounced on sleep in girls and PA in boys. CONCLUSION: Oily fish intake altered sleep and PA patterns among healthy schoolchildren, with some slight indications of sex differences. These findings warrant further investigation. CLINICAL TRIAL REGISTRY: At clinicaltrials.gov (NCT02809508) and a published protocol in Trials [Damsgaard et al. in Trials, 2016].

Does polymorphisms in PPAR and APOE genes modify associations between fatty acid desaturase (FADS), n-3 long-chain PUFA and cardiometabolic markers in 8-11-year-old Danish children?

n-3 Long-chain PUFA (LCPUFA) can improve cardiometabolic blood markers, but studies in children are limited. SNP in the FADS genes, which encode fatty acid desaturases, influence endogenous LCPUFA production. Moreover, SNP in genes that encode PPAR and apoE may modulate the effects of n-3 LCPUFA. We explored whether FADS polymorphisms were associated with blood cholesterol and TAG, insulin and glucose and whether polymorphisms in PPAR and APOE modified associations between FADS or n-3 LCPUFA status and the cardiometabolic blood markers. We measured fasting cholesterol and TAG, insulin, glucose and n-3 LCPUFA in 757 Danish 8-11-year-old children and genotyped SNP in FADS (rs1535 and rs174448), PPARG2 (rs1801282), PPARA (rs1800206) and APOE (rs7412+rs429358). Carriage of two FADS rs174448 major alleles was associated with lower TAG (P = 0·027) and higher HDL-cholesterol (P = 0·047). Blood n-3 LCPUFA was inversely associated with TAG and insulin in PPARG2 minor allele carriers and positively with LDL-cholesterol in major allele homozygotes (Pn-3 LCPUFA × rs180182 < 0·01). Associations between n-3 LCPUFA and cardiometabolic markers were not modified by APOE genotype (Pn-3 LCPUFA × APOE > 0·11), but interaction between FADS rs1535 and APOE showed that rs1535 major allele homozygotes who also carried APOE2 had higher HDL-cholesterol than all other genotype combinations (Prs1535 × APOE = 0·019, pairwise-P < 0·05). This indicates that FADS genotypes, which increase endogenous LCPUFA production, may beneficially affect children's cardiometabolic profile in a partly APOE-dependent manner. Also, the degree to which children benefit from higher n-3 LCPUFA intake may depend on their PPARG2 genotype.

Vitamin D supplementation to prevent acute respiratory infections: systematic review and meta-analysis of aggregate data from randomised controlled trials.

BACKGROUND: A 2017 meta-analysis of data from 25 randomised controlled trials of vitamin D supplementation for the prevention of acute respiratory infections revealed a protective effect of the intervention. Since then, 20 new RCTs have been completed. METHODS: Systematic review and meta-analysis of data from randomised controlled trials (RCTs) of vitamin D for ARI prevention using a random effects model. Pre-specified sub-group analyses were done to determine whether effects of vitamin D on risk of ARI varied according to baseline 25-hydroxyvitamin D (25[OH]D) concentration or dosing regimen. We searched MEDLINE, EMBASE, the Cochrane Central Register of Controlled Trials (CENTRAL), Web of Science and the ClinicalTrials.gov registry from inception to 1st May 2020. Double-blind RCTs of supplementation with vitamin D or calcidiol, of any duration, were eligible if they were approved by a Research Ethics Committee and if ARI incidence was collected prospectively and pre-specified as an efficacy outcome. Aggregate data, stratified by baseline 25(OH)D concentration, were obtained from study authors. The study was registered with PROSPERO (no. CRD42020190633). FINDINGS: We identified 45 eligible RCTs (total 73,384 participants). Data were obtained for 46,331 (98.0%) of 47,262 participants in 42 studies, aged 0 to 95 years. For the primary comparison of vitamin D supplementation vs. placebo, the intervention reduced risk of ARI overall (Odds Ratio [OR] 0.91, 95% CI 0.84 to 0.99; P for heterogeneity 0.01). No statistically significant effect of vitamin D was seen for any of the sub-groups defined by baseline 25(OH)D concentration. However, protective effects were seen for trials in which vitamin D was given using a daily dosing regimen (OR 0.75, 95% CI 0.61 to 0.93); at daily dose equivalents of 400-1000 IU (OR 0.70, 95% CI 0.55 to 0.89); and for a duration of ≤12 months (OR 0.82, 95% CI 0.72 to 0.93). No significant interaction was seen between allocation to vitamin D vs. placebo and dose frequency, dose size, or study duration. Vitamin D did not influence the proportion of participants experiencing at least one serious adverse event (OR 0.97, 95% CI 0.86 to 1.09). Risk of bias within individual studies was assessed as being low for all but three trials. A funnel plot showed left-sided asymmetry (P=0.008, Egger's test). INTERPRETATION: Vitamin D supplementation was safe and reduced risk of ARI, despite evidence of significant heterogeneity across trials. Protection was associated with administration of daily doses of 400-1000 IU vitamin D for up to 12 months. The relevance of these findings to COVID-19 is not known and requires investigation. FUNDING: None.

Effects of oily fish intake on cognitive and socioemotional function in healthy 8-9-year-old children: the FiSK Junior randomized trial.

BACKGROUND: Long-chain n-3 PUFAs (n-3 LCPUFAs) accrete in the brain during childhood and affect brain development. Randomized trials in children show inconsistent effects of n-3 LCPUFAs on cognitive and socioemotional function, and few have investigated effects of fish per se. OBJECTIVES: We aimed to investigate the effects of oily fish consumption on overall and domain-specific cognitive and socioemotional scores and explore sex differences. METHODS: Healthy 8-9-y-old children (n = 199) were randomly allocated to receive ∼300 g/wk oily fish or poultry (control) for 12 ± 2 wk. At baseline and endpoint, we assessed attention, processing speed, executive functions, memory, emotions, and behavior with a large battery of tests and questionnaires and analyzed erythrocyte fatty acid composition. RESULTS: One hundred and ninety-seven (99%) children completed the trial. Children in the fish group consumed 375 (25th-75th percentile: 325-426) g/wk oily fish resulting in 2.3 (95% CI: 1.9, 2.6) fatty acid percentage points higher erythrocyte n-3 LCPUFA than in the poultry group. The overall cognitive performance score tended to improve by 0.17 (95% CI: -0.01, 0.35) points in children who received fish compared with poultry, supported by n-3 LCPUFA dose dependency. This was driven mainly by fewer errors [-1.9 (95% CI: -3.4, -0.3)] in an attention task and improved cognitive flexibility measured as faster reaction time [-51 ms (95% CI: -94, -7 ms)] in a complex relative to a simple task ("mixing cost"). The fish intervention furthermore reduced parent-rated Strength and Difficulties Questionnaire total difficulties by -0.89 (95% CI: -1.60, -0.18) points mainly due to a -0.63 (95% CI: -1.11, -0.16) points reduction in internalizing problems that was reflected in tendency to a decrease in the overall socioemotional problems score of -0.13 (95% CI: -0.26, 0.01) points. The overall effects were similar in boys and girls. CONCLUSIONS: Oily fish dose-dependently improved cognitive function, especially attention and cognitive flexibility, and reduced socioemotional problems. The results support the importance of n-3 LCPUFAs for optimal brain function and fish intake recommendations in children.The trial was registered at www.clinicaltrials.gov as NCT02809508.

Vitamin D-related genes and cardiometabolic markers in healthy children: a Mendelian randomisation study.

Observational studies show associations between low serum 25-hydroxyvitamin D (25(OH)D) and cardiometabolic risk markers. This Mendelian randomisation study examined associations between cardiometabolic markers in children and SNP in genes related to vitamin D metabolism (DHCR7; group-specific complement (GC); cytochrome P450 subfamily IIR1 (CYP2R1); and CYP24A1) and action (CYP27B1 and VDR). In 699 healthy 8-11-year-old children, we genotyped eleven SNP. We generated a genetic risk score based on SNP associated with low 25(OH)D and investigated associations between this and blood pressure, plasma lipids and insulin. Furthermore, we examined whether SNP related to vitamin D actions modified associations between 25(OH)D and the cardiometabolic markers. All GC and CYP2R1 SNP influenced serum 25(OH)D. A risk score based on four of the six SNP was associated with 3·4 (95 % CI 2·6, 4·2) mmol/l lower 25(OH)D per risk allele (P < 0·001), but was not associated with the cardiometabolic markers. However, interactions were indicated for the three VDR SNP (Pinteraction < 0·081) on associations between 25(OH)D and TAG, systolic blood pressure and insulin, which all decreased with increasing 25(OH)D only in major allele homozygotes (ß -0·02 (95 % CI -0·04, -0·01) mmol/l; ß -0·5 (95 % CI -0·9, -0·1) mmHg; and ß -0·5 (95 % CI -1·4, 0·3) pmol/l, respectively). In conclusion, genetic variation affected 25(OH)D substantially, but the genetic score was not associated with cardiometabolic markers in children. However, VDR polymorphisms modified associations with vitamin D, which warrants further investigation of VDR's role in the relationship between vitamin D and cardiometabolic risk.

Is high oily fish intake achievable and how does it affect nutrient status in 8-9-year-old children?: the FiSK Junior trial.

PURPOSE: Most children do not meet dietary guidelines for fish intake. Fish is the main source of EPA (20:5n-3), DHA (22:6n-3) and vitamin D, but may replace better iron sources such as meat. We investigated if intake of 300 g/week oily fish was achievable in children and how it affected their nutrient status. Additionally, we validated a fish food frequency questionnaire (FFQ) by correlations against EPA + DHA in red blood cells (RBC). METHODS: In a randomised 12-week trial, 199 children (8-9 years) received oily fish or poultry (control) to be eaten five times/week. We measured dietary intake and analysed fasting RBC EPA + DHA, serum 25-hydroxyvitamin D (25(OH)D), blood haemoglobin and plasma ferritin. RESULTS: 197 (99%) children completed the study. The median (25th-75th percentile) intake was 375 (325-426) and 400 (359-452) g/week oily fish and poultry, respectively. The fish group increased their intake of EPA + DHA by 749 (593-891) mg/day and vitamin D by 3.1 (1.6-3.8) µg/day. Endpoint RBC EPA + DHA was 2.3 (95% CI 1.9; 2.6) fatty acid %-point higher than the poultry group (P < 0.001). The fish group avoided the expected 25(OH)D winter decline (P < 0.001) and had 23%-point less vitamin D insufficiency (winter subgroup, n = 82). Haemoglobin and ferritin decreased slightly in both groups (P < 0.05), but the number of children with low values did not change (P > 0.14). FFQ estimates moderately reflected habitual intake (r = 0.28-0.35) and sufficiently captured intervention-introduced changes in intake (r > 0.65). CONCLUSION: Oily fish intake of 300 g/week was achievable and improved children's EPA + DHA and 25(OH)D status, without markedly compromising iron status. These results justify public health initiatives focusing on children's fish intake.

Effects of oily fish intake on cardiometabolic markers in healthy 8- to 9-y-old children: the FiSK Junior randomized trial.

BACKGROUND: Fish oil improves cardiometabolic markers in adults, but results in children are inconsistent. Few children meet the recommended fish intake and no randomized trials have investigated how fish intake per se affects children's cardiometabolic profile. OBJECTIVE: We investigated whether oily fish consumption modulated serum triacylglycerol and diastolic blood pressure (coprimary outcomes) and other cardiometabolic markers in healthy Danish children and whether effects were sex-specific. METHODS: In a randomized controlled 12-wk trial, 199 children (aged 8-9 y) received ∼300 g/wk of oily fish or poultry (control). We measured blood pressure, heart rate, and heart rate variability (HRV) via 3-h continuous electrocardiograms and collected fasting blood samples for analysis of erythrocyte EPA [20:5n-3 (ω-3)] + DHA (22:6n-3) and serum triacylglycerol, LDL and HDL cholesterol, glucose, and insulin. RESULTS: One hundred and ninety-seven children (99%) completed the trial. The fish group consumed a median (IQR) of 375 (325-426) g oily fish/wk and the poultry group consumed 400 (359-452) g poultry/wk, which resulted in 2.25 (95% CI: 1.88, 2.62) fatty acid percentage-point higher erythrocyte EPA + DHA in the fish group (P < 0.001). In the fish group, serum triacylglycerol decreased by 0.05 mmol/L (95% CI: 0.00, 0.11 mmol/L) (P = 0.04) and HDL cholesterol increased by 0.07 mmol/L (95% CI: 0.01, 0.13 mmol/L) (P = 0.02); the triacylglycerol effect showed dose-dependency with erythrocyte EPA + DHA (r = -0.15, P = 0.04), whereas HDL showed a tendency for such an association(r = 0.13, P = 0.08). Additional analyses indicated sex-specificity (Pdiet*sex < 0.10), because triacylglycerol was reduced by 0.09 mmol/L (95% CI: 0.02, 0.16 mmol/L) in boys only (girls: -0.00; 95% CI: -0.07, 0.07 mmol/L) and heart rate was reduced by 3.4 bpm (95% CI: 0.2, 6.6 bpm) in girls only (boys: 0.6; 95% CI: -2.6, 3.8 bpm). Blood pressure, HRV, and glucose homeostasis were unaffected. CONCLUSIONS: Oily fish intake improved serum triacylglycerol and HDL cholesterol in a dose-dependent manner in 8- to 9-y-old children, but had no effect on blood pressure, HRV, or glucose homeostasis. This supports recommendations for fish intake in children and underlines the importance of initiatives to increase children's intake of oily fish. This trial was registered at clinicaltrials.gov as NCT02809508.

Exploring correlations between neuropsychological measures and domain-specific consistency in associations with n-3 LCPUFA status in 8-9 year-old boys and girls.

Long-chain n-3 polyunsaturated fatty acids (n-3 LCPUFA) have in some studies been associated with cognitive and socioemotional outcomes in children, but results are inconsistent possibly due to the use of different tests and potential gender-specific effects. The objective of this cross-sectional study was to explore overall patterns in neuropsychological scores as well as correlations between scores within specific domains, and to examine potential gender differences and consistency in associations with n-3 LCPUFA status. In 199 Danish 8-9 year-old children, we performed a large battery of tests and questionnaires on attention, processing speed, executive functions, memory, and socioemotional traits, and measured erythrocyte fatty acid composition. Principal component analyses (PCA) showed that most of the variation in both cognitive performance and socioemotional traits was explained by overall performance, followed by speed-accuracy trade off and externalizing vs. internalizing problems, respectively. Boys had higher speed, lower attention and higher externalizing problem scores than girls. Measures of performance within both processing speed and attention domains correlated moderately, whereas no correlations were found for measures of executive functions apart from some weak correlations for impulsivity. Parent-rated scores for both externalizing and internalizing problems correlated strongly, whereas correlations with child-rated scores were weak. Scores within specific domains did not consistently associate with n-3 LCPUFA, except for processing speed measures which all pointed to faster processing with increased n-3 LCPUFA status. Gender differences in the associations were observed for attention and impulsivity. Child- but not parent-rated internalizing and social problems tended to associate directly with n-3 LCPUFA, supported by increased internalizing problems measured by the PCA component. In conclusion, measures of speed and attention seem to represent these domains in general, whereas single measures of more complex cognitive functions should be interpreted with caution. One approach could be to use multiple tests and create multivariate scores to guide interpretations. Furthermore, the results indicate a need to consider both parent- and child-rated socioemotional scores and gender differences in neuropsychological functions e.g. in investigations of n-3 LCPUFA effects.

FADS and PPARG2 Single Nucleotide Polymorphisms are Associated with Plasma Lipids in 9-Mo-Old Infants.

BACKGROUND: Dietary intake of polyunsaturated fatty acids (PUFAs), e.g., linoleic acid and n-3 (ω-3) long-chain PUFAs, has been shown in adults to affect plasma cholesterol and triglycerides (TGs), respectively. Little is known about the effects of PUFAs on plasma lipids in early life. OBJECTIVE: The aim of this study was to explore the associations between plasma concentrations of total, LDL, and HDL cholesterol and TGs in infants and 2 single nucleotide polymorphisms (SNPs) in the fatty acid desaturase genes (FADS) oppositely associated with docosahexaenoic acid (rs1535 and rs174448) and potential effect modification by a functional peroxisome proliferator-activated receptor-γ2 gene variant (PPARG2 Pro12Ala). METHODS: In 9-mo-old infants (n = 561) from 3 Danish cohorts, we analyzed associations between plasma lipids, erythrocyte PUFAs, and FADS SNPs, and interactions with PPARG2 Pro12Ala genotype, by multiple linear regression. We also examined potential effect modification by breastfeeding, as 46% of the infants were still being breastfed. RESULTS: Minor allele carriage of rs174448 was associated with lower total cholesterol (difference: -0.22 mmol/L; 95% CI: -0.37, -0.06 mmol/L; P = 0.006) and LDL cholesterol (difference: -0.15 mmol/L; 95% CI: -0.29, -0.01 mmol/L; P = 0.035), but no associations were observed with TGs or for rs1535. Minor allele carriage of both FADS SNPs was associated with 1 SD lower HDL cholesterol, but only in currently breastfed infants (rs174448 × breastfeeding, P = 0.080; rs1535 × breastfeeding, P = 0.030) and PPARG2 minor allele carriers (rs174448 × PPARG2, P = 0.001; rs1535 × PPARG2, P = 0.004). Erythrocyte arachidonic acid and eicosapentaenoic acid were inversely associated with LDL cholesterol [estimated effect (ß): -0.3 mmol/L; 95% CI: -0.06, -0.00 mmol/L per percentage of fatty acids (FA%); P = 0.035] and TGs (ß: -0.23 mmol/L; 95% CI: -0.41, -0.05 mmol/L per FA%; P = 0.015), respectively. CONCLUSIONS: The observed associations with FADS variants indicate that PUFAs are involved in plasma lipid regulation in 9-mo-old infants. Observed FADS SNP differences and interactions with breastfeeding and PPARG2 warrant additional studies to explore the effects of individual FADS SNPs on PUFA status and potential genetic modification of dietary PUFA effects.

Winter vitamin D3 supplementation does not increase muscle strength, but modulates the IGF-axis in young children.

PURPOSE: To explore whether muscle strength, the insulin-like growth factor axis (IGF-axis), height, and body composition were associated with serum 25-hydroxyvitamin D [25(OH)D] and affected by winter vitamin D supplementation in healthy children, and furthermore to explore potential sex differences. METHODS: We performed a double-blind, placebo-controlled, dose-response winter trial at 55ºN. A total of 117 children aged 4-8 years were randomly assigned to either placebo, 10, or 20 µg/day of vitamin D3 for 20 weeks. At baseline and endpoint, we measured muscle strength with handgrip dynamometer, fat mass index (FMI), fat free mass index (FFMI), height, plasma IGF-1, IGF-binding protein 3 (IGFBP-3), and serum 25(OH)D. RESULTS: At baseline, serum 25(OH)D was positively associated with muscle strength, FFMI, and IGFBP-3 in girls only (all p < 0.01). At endpoint, baseline-adjusted muscle strength, FMI and FFMI did not differ between intervention groups. However, baseline-adjusted IGF-1 and IGFBP-3 were higher after 20 µg/day compared to placebo (p = 0.043 and p = 0.006, respectively) and IGFBP-3 was also higher after 20 µg/day compared to 10 µg/day (p = 0.011). Children tended to be taller after 20 µg/day compared to placebo (p = 0.064). No sex interactions were seen at endpoint. CONCLUSIONS: Avoiding the winter-related decline in serum 25(OH)D may influence IGF-1 and IGFBP-3 in children. Larger trials are required to confirm these effects, and the long-term implication for linear growth.