Effect of Vitamin D Supplementation on Bone Mass in Infants With 25-Hydroxyvitamin D Concentrations Less Than 50 nmol/L: A Prespecified Secondary Analysis of a Randomized Clinical Trial.

Importance: The dose of supplemental vitamin D needed in infants born with serum 25-hydroxyvitamin D (25[OH]D) concentrations less than 50 nmol/L (ie, 20 ng/mL) is unclear. Objective: To determine whether a higher dose (1000 IU vs 400 IU per day) is required in infants born with 25(OH)D concentrations less than 50 nmol/L for bone mineral accretion across infancy. Design, Setting, and Participants: In this prespecified secondary analysis of a double-blinded randomized clinical trial, conducted from March 2016 to March 2019 in a single center in Greater Montreal, Quebec, Canada, a consecutive sample of 139 healthy term singletons were recruited from 866 infants screened for vitamin D status at birth. Data were analyzed from June 2021 to November 2022. Interventions: Capillary blood was collected 24 to 36 hours after birth to measure serum total 25(OH)D concentrations. Infants with 25(OH)D concentrations less than 50 nmol/L were randomized to receive either 1000 IU or 400 IU per day of oral vitamin D3 supplementation from age 1 to 12 months. Infants with 25(OH)D concentrations of 50 nmol/L or greater formed a reference group. Main Outcomes and Measures: Measures at age 1, 3, 6, and 12 months were preplanned and included whole-body bone mineral content, lumbar spine bone mineral content, and bone mineral density using dual-energy x-ray absorptiometry, and serum 25(OH)D3 using liquid chromatography tandem mass spectrometry. Results: Of 139 included infants, 81 (58.3%) were male, and the median (IQR) gestational age at birth was 39.6 (38.9-40.6) weeks. A total of 49 infants were included in the 1000 IU per day group, 49 infants in the 400 IU per day group, and 41 in the reference group. Mean (SD) whole-body bone mineral content was not different between trial groups over time (1000 IU per day, 173.09 [2.36] g; 400 IU per day, 165.94 [66.08] g). Similarly, no differences were observed in lumbar spine bone mineral content or density. Mean (SD) serum 25(OH)D3 concentrations were significantly higher in the 1000 IU per day group from age 3 to 12 months (3 months, 115.2 [35.3] nmol/L; 6 months, 121.6 [34.4] nmol/L; 12 months, 99.6 [28.8] nmol/L) compared with the 400 IU per day trial group (3 months, 77.4 [23.3] nmol/L; 6 months, 85.1 [18.6] nmol/L; 12 months, 82.3 [14.3] nmol/L). Conclusions and Relevance: In this study, a higher dose of vitamin D supplementation in infants born with 25(OH)D concentrations less than 50 nmol/L did not present advantages to bone mass in infancy. This study supports a standard dose of 400 IU per day of vitamin D supplementation for breastfed infants in Montreal. Trial Registration: ClinicalTrials.gov Identifier: NCT02563015.

Vitamin D Status of Infants of Mothers with Gestational Diabetes: Status at Birth and a Randomized Controlled Trial of Vitamin D Supplementation across Infancy.

BACKGROUND: Vitamin D status and requirements of infants of women with gestational diabetes mellitus (GDM) are unclear. OBJECTIVES: The objectives were to assess vitamin D status in infants of mothers with GDM and compare vitamin D status in response to 400 vs. 1000 IU/d vitamin D supplementation in infants born with serum 25-hydroxyvitamin D [25(OH)D] <50 nmol/L. METHODS: Women with GDM delivering full-term infants (n = 98; March 2017-2019, Montreal, Canada) were surveyed for demographic and lifestyle factors. Pregnancy history was obtained from medical records. Newborn serum 25(OH)D was measured (immunoassay) and categorized as <30 (deficient) or ≥40 nmol/L (adequate). Breastfed neonates (n = 16) with serum 25(OH)D <50 nmol/L at birth were randomly assigned to 400 or 1000 IU/d of supplemental cholecalciferol (vitamin D3), and serum 25(OH)D was measured at baseline (≤1 mo) and 3, 6, and 12 mo of age. Groups were compared using a linear mixed-effects model and Tukey-Kramer post hoc tests. RESULTS: Mean newborn serum 25(OH)D was 46.4 (95% CI: 43.9, 49.9) nmol/L, with 15.3% (95% CI: 8.2%, 22.4%) <30 nmol/L and 61.2% (95% CI: 51.6%, 70.9%) ≥40 nmol/L. During the trial, most infants were breastfed to 3 mo (400 IU/d: 87.5%; 1000 IU/d: 75.0%). Mean (± SEM) infant serum 25(OH)D was higher in the 1000-IU/d group at 3 mo (79.9 ± 5.9 vs. 111.5 ± 15.2 nmol/L; P = 0.0263), and although not different at 6-12 mo, was maintained at >50 nmol/L. CONCLUSIONS: Most infants of women with GDM had adequate vitamin D status in this study. In those born with serum 25(OH)D <50 nmol/L, vitamin D status was corrected by 3 mo of age in response to 400 or 1000 IU/d of supplemental vitamin D. Dietary guidance should continue to recommend that all women who could become pregnant take a multivitamin supplement and that breastfed infants receive 400 IU/d of supplemental vitamin D. This study and ancillary trial were registered at clinicaltrials.gov (https://www. CLINICALTRIALS: gov/ct2/show/NCT02563015) as NCT02563015.

Maternal and neonatal red blood cell n-3 polyunsaturated fatty acids inversely associate with infant whole-body fat mass assessed by dual-energy X-ray absorptiometry.

Research regarding polyunsaturated fatty acid (PUFA) status and body composition in neonates is limited. This study tested the relationship between newborn docosahexaenoic acid (DHA) status and body composition. Healthy mothers and their term-born infants (n = 100) were studied within 1 month postpartum for anthropometry and whole-body composition using dual-energy X-ray absorptiometry. Maternal and infant red blood cell (RBC) membrane PUFA profiles were measured using gas chromatography (expressed as percentage of total fatty acids). Data were grouped according to infant RBC DHA quartiles and tested for differences in n-3 status and infant body composition using mixed-model ANOVA, Spearman correlations, and regression analyses (P < 0.05). Mothers were 32.2 ± 4.6 years (mean ± SD) of age, infants (54% males) were 0.68 ± 0.23 month of age, and 80% exclusively breastfed. Infant RBC DHA (ranged 3.96% to 7.75% of total fatty acids) inversely associated with infant fat mass (r = -0.22, P = 0.03). Infant and maternal RBC n-6/n-3 PUFA ratio (r2 = 0.28, P = 0.043; r2 = 0.28, P = 0.041 respectively) were positively associated with fat mass. These results demonstrate that both maternal and infant long-chain PUFA status are associated with neonatal body composition. Novelty Our findings support an early window to further explore the relationship between infant n-3 PUFA status and body composition. Maternal and infant n-3 PUFA status is inversely related to neonatal whole-body fat mass. DHA appears to be the best candidate to test in the development of a lean body phenotype.

Dietary docosahexaenoic acid contributes to increased bone mineral accretion and strength in young female Sprague-Dawley rats.

Growing evidence suggests that docosahexaenoic acid (DHA: 22:6n-3) enhances bone mineral content (BMC) and bone mineral density (BMD) in adulthood and during aging, however the effects during and after sexual maturation are unclear. The aim of this study was to examine the dose-response of BMC, BMD and microarchitectural properties of bone to dietary DHA in healthy growing female rats during acquisition of peak bone mass (PBM). Female Sprague-Dawley rats (n = 12/diet) were randomized to receive a control diet (AIN-93 M, 60 g soybean oil/kg diet) or an experimental diet containing 0.1, 0.4, 0.8 and 1.2% DHA (w/w of total diet) for 10 weeks. Dietary DHA increased the whole body, lumbar spine and long bone BMC compared to the control, in addition to higher aBMD and also BMD. Additionally, an increase in cortical bone microarchitecture parameters of lumbar spine as well as peak force were observed in dietary DHA diet groups. Dietary DHA contributes to PBM when consumed during and after sexual maturation, however higher doses of DHA do not provide further benefits.

Docosahexaenoic Acid at 0.4% of Dietary Weight Enhances Lean Mass in Young Female Sprague-Dawley Rats.

BACKGROUND: Docosahexaenoic acid (DHA; 22:6n-3) is an n-3 (ω-3) fatty acid known for beneficial effects on body composition. OBJECTIVE: The objective of the study was to test the dose response of lean and fat mass to DHA in healthy growing female rats. METHODS: Female Sprague-Dawley rats (7 wk at baseline; n = 12/diet) were randomly assigned to receive a control (AIN-93M; 60 g soybean oil/kg diet) or experimental diet for 10 wk. Experimental diets contained 0.1%, 0.4%, 0.8%, or 1.2% DHA (wt:wt of total diet). Imaging for whole-body and abdominal composition was conducted using dual-energy X-ray absorptiometry and microcomputed tomography, respectively, at weeks 0, 5, and 10. Fatty acid profiles of several tissues were analyzed using gas chromatography. Serum leptin, C-reactive protein, and plasma insulin-like growth factor I concentrations were measured at each time point using immunoassays. Data were tested using Pearson's correlations and mixed-model ANOVA. RESULTS: No differences were observed in weight at baseline or food intake throughout the study. Overall, a 6% increase (P < 0.05) in whole-body and abdominal lean mass was observed in the 0.4%-DHA diet group compared with the control diet group. Moreover, the abdominal visceral fat mass was 31.4% lower in rats in the 0.4%-DHA than in the 1.2%-DHA diet group (P < 0.001). Rats in the 1.2%-DHA diet group showed greater percent differences in whole-body (32.5% and 40.6% higher) and in abdominal (33.9% and 49.4% higher) fat mass relative to the 0.1%- and 0.4%-DHA diet groups, respectively (P < 0.01). Accordingly, serum leptin concentration was lower in the 0.1%-DHA (38.2%) and 0.4%-DHA (43.8%) diet groups (P < 0.01) than in the 1.2%-DHA diet group and positively related to whole-body fat mass (r = 0.91, P < 0.0001). CONCLUSION: Dietary DHA at 0.4% of dietary weight effectively enhances lean mass and proportionally reduces fat mass in growing female rats.

Exercise training increased gene expression of LDL-R and PCSK9 in intestine: link to transintestinal cholesterol excretion.

Transintestinal cholesterol excretion (TICE) is known as an alternate non-biliary route of cholesterol excretion from the body. The aim of this study was to determine whether exercise training has effects on intestinal membrane receptors involved in TICE in intact and ovariectomized (Ovx) rats. Sprague-Dawley rats were first divided into 4 groups: Sham operated and Ovx rats fed a standard diet (Sham-SD; Ovx-SD), or a high cholesterol diet (Sham-Chol; Ovx-Chol). These 4 groups were subsequently subdivided into either sedentary or voluntary wheel running groups for 6 weeks. The cholesterol diet resulted in increased hepatic cholesterol accumulation (p< 0.001) in both Sham and Ovx rats. Exercise training increased (p < 0.01) transcripts of intestinal low density lipoprotein receptor (LDL-R) and proprotein convertase subtilisin/kexin type 9 (PCSK9), which are involved in trans-intestinal cholesterol uptake from circulation, in both Sham and Ovx rats compared to rats remaining sedentary in all diet conditions. The up-regulation of intestinal gene expression of LDL-R and PCSK9 following voluntary wheel running in intact and Ovx rats suggests that exercise training may contribute to elimination of cholesterol through the TICE pathway.

The effect of exercise training on upregulation of molecular markers of bile acid metabolism in the liver of ovariectomized rats fed a cholesterol-rich diet.

BACKGROUND: Small heterodimer partner (SHP) is an important transcriptional factor involved in the regulation of glucose, lipid, and bile acid metabolism in the liver. SHP has been reported to be down-regulated in ovariectomized (Ovx) mice and up-regulated by estrogens suggesting a link between estrogens and SHP. The aim of the present study was to determine the effects of exercise training on SHP and key molecular markers of cholesterol and bile acid homeostasis in Ovx rats under cholesterol feeding. METHODS: Our main experimental group was composed of Ovx rats fed a high-cholesterol diet (Ovx-Chol) that was compared to a group of Ovx rats fed a standard diet (Ovx-SD) and a group of sham operated rats fed the cholesterol diet (Sham-Chol). These three groups of Ovx and sham rats were subdivided into either voluntary wheel running (Tr) or sedentary (Sed) groups for 5 weeks. The mRNA expression of all genes was measured by quantitative real-time polymerase chain reaction. RESULTS: Liver total cholesterol levels were not affected by exercise training in any of the experimental conditions. Cholesterol feeding in both sham and Ovx rats resulted in significantly higher hepatic cholesterol accumulation than in Ovx-SD (P < 0.001). Hepatic low density lipoprotein receptor (LDL-R) involved in cholesterol uptake from circulation was not influenced by training. A main effect of training was, however, found for transcripts of SHP and cholesterol 7 alpha-hydroxylase (CYP7A1, P < 0.050). CYP7A1 is the main gene involved in bile acid biosynthesis from cholesterol. CONCLUSION: These results suggest that voluntary wheel running modulates cholesterol metabolism in Ovx animals through up-regulation of SHP and bile acid formation.

Lipid peroxidation and antioxidant enzymes activity in controlled and uncontrolled Type 2 diabetic patients.

BACKGROUND: This study was designed to compare lipid peroxidation and antioxidant enzymes activity in Type 2 diabetes patients with good or weak glycemic control. METHODS: In this case-control study, 62 Type 2 diabetic patients with glycated hemoglobin (HbA1c) between 6 and 8 were enrolled as the controlled group and 55 patients with HbA1c > 8 were selected as an uncontrolled group. Patients were all referred to Iranian Diabetes Association in Tehran, Iran, from 2010 onward. Groups were chosen by convenience sampling and were matched based on age, sex and duration of disease. Demographic questionnaire, two 24-hour food recall, HbA1c, insulin, malondialdehyde (MDA), superoxide dismutase (SOD), and catalase were measured in blood samples. Data were analyzed by Food Processor II and SPSS software. RESULTS: A mean daily consumption of energy, carbohydrate, protein, and fat was not significantly different between two groups. MDA in the uncontrolled group was significantly higher than controlled group (2.03 ± 0.88 vs. 1.65 ± 1.01 nmol/ml; P = 0.030). A mean SOD was slightly higher in the uncontrolled group comparing to the control group (843.3 ± 101.9 vs. 828.0 ± 127.3 U/g Hb; P = 0.400). CONCLUSION: These data suggest that MDA as a lipid peroxidation indicator is higher in uncontrolled diabetes probably due to chronic high blood sugar followed by higher oxidative stress.

GLUT2 proteins and PPARγ transcripts levels are increased in liver of ovariectomized rats: reversal effects of resistance training.

PURPOSE: This study investigated the effects of ovariectomy (Ovx) and 12 weeks of resistance training (RT) on gene expression of GLUT2, the main glucose transporter in the liver, and on PPARγ, a transcription factor known to target GLUT2 expression. METHODS: Forty Holtzman rats were divided into 5 groups: Sham-sedentary (Sed), Sham- RT, Ovx-Sed, Ovx-RT, and Ovx-Sed with hormone replacement (E2). The RT protocol consisted of sessions held every 72 h for 12 weeks, during which the animals performed 4 to 9 vertical climbs (1.1 m) at 2 min intervals with progressively heavier weights (30 g after the fourth climb) tied to the tail. The E2 silastic capsule was inserted into the rats' backs 48 hours before the first RT session. RESULTS: In addition to liver fat, GLUT2 protein levels and PPARγ transcripts were increased (P < 0.05) in Ovx compared to Sham-Sed animals, suggesting increased hepatic glucose uptake under estrogen deficient conditions. RT and E2 in Ovx rats decreased liver fat accumulation as well as GLUT2 and PPARγ gene expression to the level of Sham- Sed animals. CONCLUSION: The results of this study suggest that liver GLUT2 as well as PPARγ expression in Ovx rats are accompanied by increased fat accumulation and glucose uptake, thus providing a substrate for increased de novo lipogenesis. RT appears to be an appropriate exercise model to circumvent these effects.

High dietary cholesterol and ovariectomy in rats repress gene expression of key markers of VLDL and bile acid metabolism in liver.

BACKGROUND: The purpose of the study was to evaluate the effects of high dietary cholesterol in ovariectomized (Ovx) rats on several key markers of hepatic cholesterol and bile acid metabolism. METHOD: Ovx and sham operated (Sham) rats were given either a standard diet (SD), a SD diet supplemented with 0.25% cholesterol (SD + Chol), or a high fat diet supplemented with 0.25% cholesterol (HF + Chol) for 5 weeks. RESULTS: Ovx was associated with higher (P < 0.05) liver total cholesterol (TC) under the SD and the SD + Chol diet, while liver triglyceride (TG) content was higher in Ovx than in Sham rats in all 3 diet conditions. Surprisingly, the SD + Chol diet was associated with lower (P < 0.001) plasma TC and TG levels in Ovx than in Sham rats, suggesting a decrease in VLDL secretion. Accordingly, several transcripts of key markers of VLDL synthesis including microsomal TG transfer protein (Mttp) and Apob-100 were decreased (P < 0.05) in Ovx compared to Sham rats under the three dietary conditions and even more so for Mttp and Apob-100 when rats were fed the SD + Chol diet. Transcripts of bile acid transporters including bile salt export pump (Bsep) and Na + -taurocholate cotransporting polypeptide (Ntcp) were decreased by the addition of cholesterol to the SD diet in both Ovx and Sham rats. CONCLUSION: These results indicate that a high cholesterol feeding and ovariectomy combine to reduce the gene expression of key markers of VLDL synthesis suggesting a reduction in excretion of cholesterol from the liver.

Exercise training decreases gene expression of endo- and xeno-sensors in rat small intestine.

The purpose of the study was to test the hypothesis that gene expression of members of the nuclear receptor (NR) superfamily known to act as endo- and xeno-sensors is reduced in the ileum of exercise-trained (Tr) rats. Healthy female rats were either treadmill-trained for 8 weeks, 5 times/week, or remained sedentary (Sed). Training resulted in a significant (p < 0.05) decrease in plasma free fatty acid (0.18 ± 0.01 to 0.15 ± 0.01 mmol/L) and glycerol (24.8 ± 0.8 to 18.7 ± 0.8 mg/L) concentrations. Gene expressions of NRs farnesoid X receptor (FXR; p < 0.05), liver X receptor (LXR; p < 0.05), pregnane X receptor (PXR; p < 0.01), and retinoid X receptor (RXR; p < 0.06) were reduced in the ileum of Tr compared with Sed animals. Tr was also associated with a reduction (p < 0.05) in gene expression of FXR downstream heterodimeric organite solute transporters α (OSTα) and ß (OSTß) involved in the transport of bile acids, LXR downstream genes heterodimeric ATP-binding cassette transporters (ABCG5/G8) involved in transport of absorbed cholesterol back to the lumen, and Niemann-Pick C1-like 1 (NPC1L1) involved in cholesterol absorption. These data indicate that exercise training lowers the expression of molecules involved in the defense system of the ileum against endobiotic and xenobiotic insults under normal conditions, thus, suggesting that regular exercise contributes to the intestinal maintenance of cholesterol and bile acid homeostasis.