Influence of vitamin D supplementation on bone mineral content, bone turnover markers, and fracture risk in South African schoolchildren: multicenter double-blind randomized placebo-controlled trial (ViDiKids).

Randomized controlled trials (RCTs) to determine the influence of vitamin D on BMC and fracture risk in children of Black African ancestry are lacking. We conducted a sub-study (n = 450) nested within a phase 3 RCT of weekly oral supplementation with 10 000 IU vitamin D3 vs placebo for 3 yr in HIV-uninfected Cape Town schoolchildren aged 6-11 yr. Outcomes were BMC at the whole body less head (WBLH) and LS and serum 25-hydroxyvitamin D3 (25(OH)D3), PTH, alkaline phosphatase, C-terminal telopeptide, and PINP. Incidence of fractures was a secondary outcome of the main trial (n = 1682). At baseline, mean serum 25(OH)D3 concentration was 70.0 nmol/L (SD 13.5), and 5.8% of participants had serum 25(OH)D3 concentrations <50 nmol/L. Among sub-study participants, end-trial serum 25(OH)D3 concentrations were higher for participants allocated to vitamin D vs placebo (adjusted mean difference [aMD] 39.9 nmol/L, 95% CI, 36.1 to 43.6) and serum PTH concentrations were lower (aMD -0.55 pmol/L, 95% CI, -0.94 to -0.17). However, no interarm differences were seen for WBLH BMC (aMD -8.0 g, 95% CI, -30.7 to 14.7) or LS BMC (aMD -0.3 g, 95% CI, -1.3 to 0.8) or serum concentrations of bone turnover markers. Fractures were rare among participants in the main trial randomized to vitamin D vs placebo (7/755 vs 10/758 attending at least 1 follow-up; adjusted odds ratio 0.70, 95% CI, 0.27 to 1.85). In conclusion, a 3-yr course of weekly oral vitamin D supplementation elevated serum 25(OH)D3 concentrations and suppressed serum PTH concentrations in HIV-uninfected South African schoolchildren of Black African ancestry but did not influence BMC or serum concentrations of bone turnover markers. Fracture incidence was low, limiting power to detect an effect of vitamin D on this outcome.

Vitamin D­the "sunshine vitamin"­is essential for helping the body to absorb calcium from the diet, which is laid down in bone to improve its strength. There is a lack of clinical trials testing whether vitamin D supplements can improve bone content of calcium and other minerals, or reduce risk of bone fractures (broken bones) in children of Black African ancestry. We therefore conducted such a study, recruiting 1682 schoolchildren aged 6­11 yr living in Cape Town, South Africa. We found that a weekly dose of 10 000 international units (250 micrograms) of vitamin D3, given by mouth for 3 yr, was effective in boosting vitamin D levels in trial participants who received it. However, this did not have any effect on bone content of calcium and other minerals. Relatively few children experienced a broken bone during the study, so we were unable to say with confidence whether or not vitamin D supplements might affect this outcome.

Effects of 25-hydroxycholecalciferol on performance, gut health, and bone quality of broilers fed with reduced calcium and phosphorus diet during Eimeria challenge.

This study evaluated the effects of 25-hydroxycholecalciferol (25-OHD) on performance, gut health, and bone quality of broilers fed with reduced calcium (Ca) and phosphorus (P) diet during Eimeria spp. challenge. A total of 576 fourteen-day-old Cobb 500 male chicks were randomly distributed in a 2 × 2 × 2 factorial arrangement, with 6 replicates of 12 birds each. The main factors were 25-OHD level (0 or 3,000 IU/kg of feed), mineral level (0.84% of Ca/0.42% of P, the levels recommended for the grower phase (NOR) or 0.64% of Ca/0.22% of P (RED), and mid-high mixed Eimeria challenge or nonchallenge. 25-OHD improved phosphorus retention (P = 0.019), bone ash weight (P = 0.04), cortical bone trabecular connectivity (P = 0.043) during coccidiosis. For birds fed with reduced mineral levels, 25-OHD supplementation increased bone ash weight (P = 0.04). However, 25-OHD did not improve bone ash weight when birds were challenged and fed with reduced mineral levels. The dietary 3,000 IU of 25-OHD supplementation did not improve performance or gut morphology but support bone health during coccidiosis. Future investigations are needed for better understand 25-OHD role on bone microarchitecture and oxidative metabolism during coccidiosis.

Dietary 25-hydroxy-cholecalciferol and additional vitamin E improve bone development and antioxidant capacity in high-density stocking broilers.

This study aimed to investigate the effects of diets supplemented with 25-hydroxycholecalciferol [25-(OH)D3] and additional vitamin E on growth performance, antioxidant capacity, bone development, and carcass characteristics at different stocking densities on commercial broiler farms. A total of 118,800 one-day-old Arbor Acres broilers were assigned to a 2 × 2 factorial treatment consisting of two dietary vitamin levels (5,500 IU vitamin D3 and 60 IU vitamin E: normal diet, using half 25-(OH)D3 as a source of vitamin D3 and an additional 60 IU of vitamin E: 25-(OH)D3+VE diet) and two stocking densities (high density of 20 chickens/m2: HD and 16 chickens/m2: LD). The experiment lasted for 42 d. The results showed that high-density stocking negatively affected the growth performance of broilers during the first four weeks, whereas the vitamin diet treatment significantly improved the feed conversion ratios (FCR) during the last 2 wk. Vitamin diets increased catalase at 14 and 42 d, and the glutathione peroxidase (GSH-px) levels at 42 d in high-density-stocked broilers. The interaction showed that serum vitamin E levels were significantly improved at 28 d of age in high-density-stocked broilers as a result of the vitamin diets. Stocking density and dietary treatments were found to significantly affect bone development, with the vitamin diet significantly increasing metatarsal length and femoral bone strength in broilers from high-density stocking density at 28 d of age. High stocking density increased the proportion of leg muscles and meat yield per square meter. In general, 25-(OH)D3 and additional vitamin E suppressed oxidative stress and ameliorated the negative effects of high-density stocking on bone development in a commercial chicken farm setting. Vitamin diets improved the FCR of broilers, while high-density stocking resulted in better economic outcomes.

High-density stocking is often associated with animal welfare risks in broilers, mainly in terms of oxidative stress and bone development. Nevertheless, farming at too low a density remains for the most part economically unviable. Modulation of antioxidant capacity and bone development by nutritional strategies in high-density-farmed broilers has proven an effective tool in developing countries. Therefore, the present study investigated the effects of applying diets with a higher biological potency of vitamin D3 25-hydroxycholecalciferol [25-(OH)D3] and a higher concentration of vitamin E on broiler production performance, antioxidant capacity and meat production performance at different densities of stocking under commercial farming conditions. The results indicated that the vitamin dietary treatments suppressed oxidative stress and ameliorated the negative effects of high-density farming on bone development.

Effects of 25-hydroxycholecalciferol (25-OH-D3) and 1-hydroxycholecalciferol (1-OH-D3) on serum bone biomarkers and calcium and phosphorus balance and concentrations of energy in diets without or with microbial phytase fed to sows in late gestation.

The objective was to test the hypothesis that supplementation of diets for gestating sows with 25-hydroxycholecalciferol (25-OH-D3) or 1-hydroxycholecalciferol (1-OH-D3) affects serum biomarkers for bone and increases Ca and P balance and the apparent total tract digestibility (ATTD) of gross energy (GE), and the concentrations of digestible energy (DE) and metabolizable energy (ME) in diets without or with microbial phytase. Sixty multiparous sows were allotted to 1 of 6 diets. Diets were formulated using a 3 × 2 factorial with 3 inclusions of supplemental vitamin D metabolite (no metabolite, 25-OH-D3, or 1-OH-D3) and 2 inclusion levels of microbial phytase (0 or 1,000 units). Sows were housed individually in metabolism crates and feces and urine were collected quantitatively. Results indicated that there was no difference in the ATTD of dry matter (DM) and GE and concentration of DE among the 3 diets containing microbial phytase, but the ATTD of DM and GE and concentration of DE was greater (P < 0.05) in diets containing 1-OH-D3 compared with the diet without a vitamin D metabolite if phytase was not used (interaction; P < 0.05). In diets without microbial phytase, ME was greater in diets containing either one of the 2 vitamin D metabolites than in the diet without a vitamin D metabolite, but among diets with microbial phytase, the ME of the 1-OH-D3 diet was less than of the 25-OH-D3 diet (interaction; P < 0.05). No effect of microbial phytase on concentrations of DE and ME was observed. There was no interaction between supplementation of microbial phytase and vitamin D metabolites for Ca and P balances, and regardless of metabolite supplementation, use of microbial phytase increased (P < 0.05) the ATTD and retention of Ca and P. Regardless of dietary phytase, the ATTD and retention of Ca and P increased (P < 0.05) for sows fed a diet containing one of the vitamin D metabolites compared with sows fed the diet without a vitamin D metabolite. Serum biomarkers for bone resorption or bone tissue synthesis were not affected by experimental diets. In conclusion, the ATTD of DM and GE, concentrations of DE and ME, and Ca and P balance in phytase-free diets fed to sows in late gestation were increased by supplementation with 1-OH-D3 or 25-OH-D3, but no differences between the 2 vitamin D metabolites were observed. Supplementation of diets with microbial phytase increased Ca and P balance, but did not affect DE and ME of diets.

The role of vitamin D is to increase absorption of calcium and phosphorus in the gastrointestinal tract and maintain serum concentrations of calcium, but dietary vitamin D needs to be converted to an active form by 2-hydroxylation steps that take place in the liver and the kidneys. The conversion efficiency to active vitamin D may be increased if pre-hydroxylated metabolites rather than vitamin D are provided, which also increases calcium and phosphorus utilization. In a previous experiment it was also demonstrated that a vitamin D metabolite increases energy absorption in gestating sows. It is possible that use of a vitamin D metabolite and phytase have additive effects and the hypothesis, therefore, was that supplementation of a vitamin D metabolite increases calcium and phosphorus balance and energy digestibility in diets fed to gestating sows without or with microbial phytase. Results indicated that in diets without phytase, the 2 vitamin D metabolites increased energy concentration in diets by increasing apparent energy digestibility. There was no interaction between supplementation of phytase and vitamin D metabolites for calcium and phosphorus balances. Use of phytase and vitamin D metabolites increased calcium and phosphorus digestibility and retention.

Effect of calcifediol supplementation on renin-angiotensin-aldosterone system mediators in dogs with chronic kidney disease.

BACKGROUND: Chronic kidney disease (CKD) leads to low serum concentrations of vitamin D metabolites. Thus, hypovitaminosis D associated with CKD might contribute to disease progression via increased concentration of renin angiotensin aldosterone system (RAAS) mediators. OBJECTIVES: To evaluate whether supplementation with calcifediol affects equilibrium concentrations of selected mediators of the RAAS. We hypothesized that vitamin D supplementation will decrease concentration of circulating RAAS mediators in dogs with CKD. ANIMALS: Six client-owned adult dogs with IRIS Stage 2 and 3 CKD. METHODS: Prospective study. Serum 25-hydroxyvitamin D (25[OH]D), 1,25-dihydroxyvitamin D (1,25[OH]2 D), 24,25-dihydroxyvitamin D (24,25[OH]2 D), RAAS mediators (angiotensin I/II/III/IV/1-5/1-7, and aldosterone), and surrogate angiotensin converting enzyme (ACE) activity (calculated by the ratio of angiotensin II to angiotensin I) were evaluated at baseline, after 3 months of calcifediol supplementation, and 2 months after discontinuing administration of supplement. RESULTS: All serum vitamin D metabolite concentrations increased significantly by month 3 (P < .001): 25(OH)D (median 250 ng/mL; range, 204-310), compared to baseline (median 43.2 ng/mL; range, 33.8-58.3 ng/mL); 1,25(OH)2 D (median 66.1 pg/mL; range, 57.3-88.1 pg/mL) compared to baseline (median 35.2 pg/mL; range, 29.3-56.7 pg/mL); 24,25(OH)2 D (median 68.4 ng/mL; range, 22.1-142.0 ng/mL) compared to baseline (median 14.4 ng/mL; range, 9.0-21.3 ng/mL). Calculated ACE activity was significantly lower at month 3 (median 0.5; range, 0.4-1.0) compared to baseline (median 0.7; range, 0.6-1.3; P = .01). There were no significant differences in any of the evaluated RAAS variables at any other time-point. CONCLUSIONS AND CLINICAL IMPORTANCE: Short-term calcifediol supplementation in this small group of CKD dogs appeared to decrease ACE activity.

Effect of combined maternal and post-hatch dietary 25-hydroxycholecalciferol supplementation on broiler chicken Pectoralis major muscle growth characteristics and satellite cell mitotic activity.

Skeletal muscle growth is largely dependent on the proliferation and differentiation of muscle-specific stem cells known as satellite cells (SC). Previous work has shown that dietary inclusion of the vitamin D3 metabolite, 25-hydroxycholecalciferol (25OHD3), also called calcidiol, can promote skeletal muscle growth in post-hatch broiler chickens. Improving vitamin D status of broiler breeder hens by feeding 25OHD3 in addition to vitamin D3 has also been shown to positively impact progeny. Yet, whether combined pre- and post-hatch supplementation with 25OHD3 produces an additive or synergistic SC-mediated, skeletal muscle growth response remains unanswered. To evaluate the effect of combined maternal and post-hatch dietary 25OHD3 supplementation on the growth and SC mitotic activity of the Pectoralis major (PM) muscles in broiler chickens, a randomized complete block design experiment with the main effects of maternal diet (MDIET) and post-hatch diet (PDIET) arranged in a 2 × 2 factorial treatment structure was conducted. From 25 to 36 wk of age, broiler breeder hens were fed 1 of 2 MDIET formulated to provide 5,000 IU D3 (MCTL) or 2,240 IU of D3 + 2,760 IU of 25OHD3 per kg of feed (M25OHD3). Their male broiler chick offspring (n = 400) hatched from eggs collected from 35 to 36 wk of age were reared in raised floor pens. Broilers were fed 1 of 2 PDIET formulated to provide 5,000 IU of D3 per kg of feed (PCTL) or 2,240 IU of D3 + 2,760 IU of 25OHD3 per kg of feed (P25OHD3). Muscle was collected at days 4, 8, 15, 22, and 29 and stored until immunofluorescence analysis. Data were analyzed as a 2-way ANOVA with SAS GLIMMIX. Dietary 25OHD3 was effectively transferred from hen plasma to egg yolks (P = 0.002) and to broiler progeny plasma (days 4 to 22; P ≤ 0.044). Including 25OHD3 in either MDIET or PDIET altered PM hypertrophic growth prior to day 29 (P ≥ 0.001) and tended to reduce Wooden Breast severity (P ≤ 0.089). Mitotic SC populations were increased in PM of MCTL:P25OHD3 and M25OHD:PCTL-fed broilers at d 4 (P = 0.037). At d 8, the PM mitotic SC populations were increased 33% by P25OHD3 (P = 0.054). The results of this study reveal that combined maternal and post-hatch 25OHD3 supplementation does not produce additive or synergistic effects on SC-mediated broiler muscle growth. However, vitamin D status improvement through dietary 25OHD3 inclusion in either the maternal or post-hatch diet stimulated broiler breast muscle growth by increasing proliferating SC populations.

Skeletal muscle growth is largely dependent on the proliferation and differentiation of muscle-specific stem cells known as satellite cells (SC). Previous work has shown that dietary inclusion of the vitamin D3 metabolite, 25-hydroxycholecalciferol (25OHD3), also called calcidiol, can promote skeletal muscle growth in post-hatch broiler chickens. Improving vitamin D status of broiler breeder hens by feeding 25OHD3 in addition to vitamin D3 has also been shown to positively impact progeny. Yet, whether combined pre- and post-hatch supplementation with 25OHD3 produces an additive or synergistic SC-mediated, skeletal muscle growth response remains unanswered. The results of this study reveal that combined maternal and post-hatch 25OHD3 supplementation does not produce additive or synergistic effects on SC-mediated broiler muscle growth. However, vitamin D status improvement through dietary 25OHD3 inclusion in either the maternal or post-hatch diet stimulated broiler breast muscle growth by increasing proliferating SC populations. Overall, this work answers not only practical questions for the broiler industry regarding the possible benefits of combining maternal and post-hatch dietary 25OHD3 supplementation but also improves our understanding of vitamin D's role in pre- and post-hatch broiler skeletal muscle growth.

Dietary supplementation with canthaxanthin and 25-hydroxycholecalciferol on the incubation performance and fertility of European quail breeders.

This study assessed the effects of combined supplementation with canthaxanthin (Cx) and 25-hydroxycholecalciferol (25-OH-D3) on incubation performance, fertility, and chick quality in European quail breeders. A total of 240 birds were distributed in a completely randomized design with 5 diets and 8 replicates. The animals were fed a basal diet containing 50 µg of vitamin D3 or the basal diet supplemented with 3 ppm Cx and 34.5 µg 25-OH-D3, 6 ppm Cx, and 69 µg 25-OH-D3, 9 ppm Cx and 103.5 µg 25-OH-D3, or 12 ppm Cx and 138 µg 25-OH-D3. Incubation performance was analyzed in 2 periods (32 and 38 wk). Breeders aged 32 wk produced eggs with higher hatchability (P = 0.024), hatchability of fertile eggs (P = 0.026) and lower initial plus mid embryonic mortality (P = 0.021), whereas 38-week-old breeders generated chicks with a higher length at hatching (P < 0.001) and lower final plus pipped embryonic mortality (P = 0.021). In both age groups, Cx + 25-OH-D3 levels had a quadratic effect on egg fertility (P < 0.001), hatchability of total (P < 0.001), and fertile eggs (P < 0.001). The fertility and the number of sperm cells in the perivitelline membrane was analyzed in two periods (26 and 40 wk). A quadratic effect of diet and days after mating on both parameters (P < 0.05) was observed. Eggs from supplementing breeders showed a high fertility (P < 0.001) and sperm cell counts (P < 0.001) for up to 7 and 3 d after mating, respectively, then the control group. Moreover, the supplementation of quail breeder diets with 6 ppm Cx + 69 µg 25-OH-D3 enhances sperm cell longevity in sperm storage tubules, hatchability of total and fertile eggs, fertility, and chick quality, especially in older quail's breeders and reduces embryonic mortality.

Differential Effects of 25-Hydroxyvitamin D3 versus 1α 25-Dihydroxyvitamin D3 on Adipose Tissue Browning in CKD-Associated Cachexia.

Patients with chronic kidney disease (CKD) often have low serum concentrations of 25(OH)D3 and 1,25(OH)2D3. We investigated the differential effects of 25(OH)D3 versus 1,25(OH)2D3 repletion in mice with surgically induced CKD. Intraperitoneal supplementation of 25(OH)D3 (75 µg/kg/day) or 1,25(OH)2D3 (60 ng/kg/day) for 6 weeks normalized serum 25(OH)D3 or 1,25(OH)2D3 concentrations in CKD mice, respectively. Repletion of 25(OH)D3 normalized appetite, significantly improved weight gain, increased fat and lean mass content and in vivo muscle function, as well as attenuated elevated resting metabolic rate relative to repletion of 1,25(OH)2D3 in CKD mice. Repletion of 25(OH)D3 in CKD mice attenuated adipose tissue browning as well as ameliorated perturbations of energy homeostasis in adipose tissue and skeletal muscle, whereas repletion of 1,25(OH)2D3 did not. Significant improvement of muscle fiber size and normalization of fat infiltration of gastrocnemius was apparent with repletion of 25(OH)D3 but not with 1,25(OH)2D3 in CKD mice. This was accompanied by attenuation of the aberrant gene expression of muscle mass regulatory signaling, molecular pathways related to muscle fibrosis as well as muscle expression profile associated with skeletal muscle wasting in CKD mice. Our findings provide evidence that repletion of 25(OH)D3 exerts metabolic advantages over repletion of 1,25(OH)2D3 by attenuating adipose tissue browning and muscle wasting in CKD mice.

1,25-Dihydroxyvitamin D3 and 20-Hydroxyvitamin D3 Upregulate LAIR-1 and Attenuate Collagen Induced Arthritis.

Vitamin D plays a crucial role in regulation of the immune response. However, treatment of autoimmune diseases with 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] doses sufficient to be effective is prohibitive due to its calcemic and toxic effects. We use the collagen-induced arthritis (CIA) model to analyze the efficacy of the noncalcemic analog of vitamin D, 20S-hydroxyvitamin D3 [20S(OH)D3], as well as 1,25(OH)2D3, to attenuate arthritis and explore a potential mechanism of action. Mice fed a diet deficient in vitamin D developed a more severe arthritis characterized by enhanced secretion of T cell inflammatory cytokines, compared to mice fed a normal diet. The T cell inflammatory cytokines were effectively suppressed, however, by culture of the cells with 20S(OH)D3. Interestingly, one of the consequences of culture with 1,25(OH)2D3 or 20S(OH)D3, was upregulation of the natural inhibitory receptor leukocyte associated immunoglobulin-like receptor-1 (LAIR-1 or CD305). Polyclonal antibodies which activate LAIR-1 were also capable of attenuating arthritis. Moreover, oral therapy with active forms of vitamin D suppressed arthritis in LAIR-1 sufficient DR1 mice, but were ineffective in LAIR-1-/- deficient mice. Taken together, these data show that the effect of vitamin D on inflammation is at least, in part, mediated by LAIR-1 and that non-calcemic 20S(OH)D3 may be a promising therapeutic agent for the treatment of autoimmune diseases such as Rheumatoid Arthritis.

Organic zinc and manganese and 25-hydroxycholecalciferol improves eggshell thickness in late-phase laying hens.

The aim of this study was to evaluate the effects of organic micromineral zinc (Zn) and manganese (Mn) and 25-hydroxycholecalciferol supplementation in late-phase laying hens' diets on performance, egg quality, lipid stability of fresh and stored eggs, and bone quality. The treatments were a basal diet and diets supplemented with 32 mg Zn-Met/kg diet; 26 mg Mn-Met/kg diet; 32 mg Zn-Met/kg diet and 26 mg Mn-Met/kg diet; 1500 IU 25-hydroxycholecalciferol/kg diet; 32 mg Zn-Met/kg diet; 26 mg Mn-Met/kg diet; and 1500 IU 25-hydroxycholecalciferol/kg diet. On performance, the birds supplemented with organic manganese had the lowest feed intake. Regarding egg quality, the birds supplemented with Zn-Met and Mn-Met, with 25-hydroxycholecalciferol alone, and with Zn-Met, Mn-Met and 25-hydroxycholecalciferol presented a greater eggshell thickness than those receiving the basal diet. Lipid stability of the yolk varied only according to storage time. No effect of supplementation was observed on bone quality. Supplementation with Zn-Met and Mn-Met, or associated with 25-hydroxycholecalciferol, or 25-hydroxycholecalciferol alone, improved eggshell thickness in aged white layers. However, the associated or isolated supplementation with these nutrients did not influence performance, lipid stability of fresh and stored egg yolk or bone quality.

FDA-Approved Drug Screening for Compounds That Facilitate Hematopoietic Stem and Progenitor Cells (HSPCs) Expansion in Zebrafish.

Hematopoietic stem cells (HSCs) are a specialized subset of cells with self-renewal and multilineage differentiation potency, which are essential for their function in bone marrow or umbilical cord blood transplantation to treat blood disorders. Expanding the hematopoietic stem and progenitor cells (HSPCs) ex vivo is essential to understand the HSPCs-based therapies potency. Here, we established a screening system in zebrafish by adopting an FDA-approved drug library to identify candidates that could facilitate HSPC expansion. To date, we have screened 171 drugs of 7 categories, including antibacterial, antineoplastic, glucocorticoid, NSAIDS, vitamins, antidepressant, and antipsychotic drugs. We found 21 drugs that contributed to HSPCs expansion, 32 drugs' administration caused HSPCs diminishment and 118 drugs' treatment elicited no effect on HSPCs amplification. Among these drugs, we further investigated the vitamin drugs ergocalciferol and panthenol, taking advantage of their acceptability, limited side-effects, and easy delivery. These two drugs, in particular, efficiently expanded the HSPCs pool in a dose-dependent manner. Their application even mitigated the compromised hematopoiesis in an ikzf1-/- mutant. Taken together, our study implied that the larval zebrafish is a suitable model for drug repurposing of effective molecules (especially those already approved for clinical use) that can facilitate HSPCs expansion.

20S-Hydroxyvitamin D3, a Secosteroid Produced in Humans, Is Anti-Inflammatory and Inhibits Murine Autoimmune Arthritis.

The ability to use large doses of vitamin D3 (D3) to chronically treat autoimmune diseases such as rheumatoid arthritis (RA) is prohibitive due to its calcemic effect which can damage vital organs. Cytochrome P450scc (CYP11A1) is able to convert D3 into the noncalcemic analog 20S-hydroxyvitamin D3 [20S(OH)D3]. We demonstrate that 20S(OH)D3 markedly suppresses clinical signs of arthritis and joint damage in a mouse model of RA. Furthermore, treatment with 20S(OH)D3 reduces lymphocyte subsets such as CD4+ T cells and CD19+ B cells leading to a significant reduction in inflammatory cytokines. The ratio of T reg cells (CD4+CD25+Foxp3+ T cells) to CD3+CD4+ T cells is increased while there is a decrease in critical complement-fixing anti-CII antibodies. Since pro-inflammatory cytokines and antibodies against type II collagen ordinarily lead to destruction of cartilage and bone, their decline explains why arthritis is attenuated by 20(OH) D3. These results provide a basis for further consideration of 20S(OH)D3 as a potential treatment for RA and other autoimmune disorders.

Role of long-term supplementation of 25-hydroxyvitamin D3 on egg production and egg quality of laying hen.

A study was conducted to evaluate the effect of dietary 25-hydroxyvitamin D3 (25OHD) on pullet and egg-laying hen growth performance, egg production, and egg quality. Three hundred and ninety 1-day-old Hy-Line W36 pullets were randomly allocated to 3 treatments with 10 replicated cages and 13 birds per cage. Dietary treatments were vitamin D3 at 2,760 IU/kg (D); vitamin D3 at 5,520 IU/kg (DD), and vitamin D3 at 2,760 IU/kg plus 25OHD at 2,760 IU (69 µg)/kg (25D). Body weight and feed intake were recorded at the end of each stage: starter 1 (0-3 wk), starter 2 (4-6 wk), grower (7-12 wk), developer (13-15 wk), prelay (15-17 wk), peaking (18-38 wk), layer 2 (39-48 wk), layer 3 (49-60 wk), layer 4 (61-75 wk), and layer 5 (76-95 wk). Egg production was recorded daily. Egg quality was evaluated every 8 wk starting from 25 wk. There was no difference in growth performance during the rearing period (0-17 wk). In the laying period (18-95 wk), DD showed lower feed intake at layer 2, but higher intake at layer 3 along with lower hen day production (HDP) from 22 to 48 wk compared to the other treatments. During the same period, the DD group laid smaller eggs with higher specific gravity and shell thickness compared with the other treatments or D alone at 40 wk, which may be partly due to the lower body weight. In contrast, 25D had better feed conversion ratio (feed intake per dozen of eggs) at layer 2, and higher overall (22-60 wk) HDP compared with DD. For the egg quality analysis, at 25 and 33 wk, both DD and 25D had higher Haugh unit compared with D. However, 25OHD has no effects on eggshell quality during the entire production period and no beneficial effects on egg production during the later laying period (after 60 wk). In summary, long-term and early supplementation of 25OHD has positive effects on egg production and egg quality, and the beneficial effects were mainly observed during the early laying stage.

Effects of normal and low calcium and phosphorus levels and 25-hydroxycholecalciferol supplementation on performance, serum antioxidant status, meat quality, and bone properties of broilers.

To determine the effects of normal and low dietary calcium (Ca) and phosphorus (P) levels and 25-hydroxycholecalciferol (25-OH-D3) supplementation on performance, serum antioxidant status, meat quality, and bone properties of broilers, 224 1-day-old Arbor Acre male broilers were used in this study. Broilers were allotted randomly to 1 of 4 treatments in a 2 × 2 factorial arrangement that included normal or low Ca and P diet with or without 69 µg/kg 25-OH-D3. The trial consists of a starter phase from day 1 to 21 and a grower phase from day 22 to 42. Dietary 25-OH-D3 supplementation increased (P < 0.05) average daily weight gain from day 22 to 42 and decreased feed conversation ratio from day 22 to 42 and day 0 to 42. On day 21, 25-OH-D3 increased serum concentrations of total antioxidant capacity (T-AOC), catalase (CAT), and glutathione peroxidase in broilers fed low Ca and P diet (Interaction, P < 0.05). 25-hydroxycholecalciferol significantly decreased serum malondialdehyde concentration. Dietary Ca and P deficiencies significantly decreased serum Ca and P concentrations and increased serum parathyroid hormone (PTH) concentration, and serum Ca and 25-OH-D3 concentrations were significantly increased by 25-OH-D3 supplementation. On day 42, serum T-AOC and CAT concentrations were decreased by dietary Ca and P deficiencies without 25-OH-D3 (Interaction, P < 0.05) and unaffected by dietary Ca and P deficiencies with 25-OH-D3. Dietary Ca and P deficiencies significantly decreased Ca, P, and alkaline phosphatase concentrations and increased PTH concentration in serum. Dietary 25-OH-D3 increased (P < 0.05) serum Ca and 25-OH-D3 concentrations and decreased (P < 0.05) serum tartrate-resistant acid phosphatase concentration. The interaction between CaP level and 25-OH-D3 was observed (P < 0.05) for tibial Ca content and femoral bone density. 25-hydroxycholecalciferol significantly increased tibial breaking strength. These data indicated that 25-OH-D3 supplementation at 69 µg/kg increased growth performance in some periods, enhanced serum antioxidant capacity, and improved bone mineralization and breaking strength of broilers.

Role of long-term supplementation of 25-hydroxyvitamin D3 on laying hen bone 3-dimensional structural development.

Egg-laying hens have a unique bone development pattern due to the medullary bone formation and high bone turnover rate. The role of long-term supplementation of an intermediate form of vitamin D3, 25-hydroxyvitamin D3 (25OHD), on skeletal development of pullets and laying hens is not well established. Exploring its effects on layer bone development will help develop a strategy for preventing laying hen osteoporosis. The purpose of this study was to investigate the role of long-term supplementation of 25OHD in layer diets on bone 3-dimensional structural development. A total of 390 1-day-old Hy-Line W36 pullets were randomly allocated to 3 treatments with 10 replicate cages and 13 birds/cage. Dietary treatments were 1) vitamin D3 at 2,760 IU/kg, 2) vitamin D3 at 5,520 IU/kg, and 3) vitamin D3 at 2,760 IU/kg plus 25OHD at 2,760 IU (69 µg)/kg. The level of 25OHD in the serum was tested throughout the whole experimental period (0-95 wk). Bone growth rate (BGR) was measured at 10 wk using a calcein injection technique. Femurs were scanned using Micro-CT for 3D structural analysis, and the whole-body composition analysis was performed using a dual-energy x-ray absorptiometry method at 17, 60, and 95 wk. Dietary supplementation of 25OHD significantly increased 25OHD level in the serum from 0 to 95 wk. During the rearing period (0-17 wk), 25OHD increased BGR, cortical tissue volume, and bone marrow area at 17 wk, simultaneously. 25OHD created more pores in cortical bone, which resulted in a lower cortical bone mineral density (BMD) but without alerting bone mineral content (BMC). This effect allowed more space for mineral deposition in bones during the later egg-laying period. At 60 wk, the 25OHD group had significantly greater BMD, which led to the highest total BMC, cortical volume, and trabecular bone connectivity. At 95 wk, the birds fed 25OHD had significantly higher cortical bone volume and lower porosity. The 25OHD group also had higher total BMD and medullary bone volume but a lower BMC and volume of trabecular bone than vitamin D3 or double dosage vitamin D3 treatment. This indicated that the bone resorption rate was lower in cortical bone than that in trabecular bone in the late laying period. In conclusion, supplementation with dietary 25OHD could stimulate bone growth and increase bone volume in pullets to provide more space for mineral deposition during the laying period with positive effects on laying hen bone quality.

Dietary supplementation with canthaxanthin and 25-hydroxycholecalciferol has beneficial effects on bone and oxidative metabolism in European quail breeders.

This study aimed to investigate the effect of supplementation with canthaxanthin (Cx) and 25-hydroxycholecalciferol (25-OH-D3) on the production performance, egg quality, bone mineral content, blood biochemical parameters, and antioxidant status of European quail breeders. Two hundred and forty quail breeders were distributed in a completely randomized design with 5 diets and 8 replicates of 4 females and 2 males were used. All quail breeders received one of 5 diets: basal diet (containing 2,000 IU vitamin D3) or the same diet supplemented with 3 ppm Cx and 34.5 µg 25-OH-D3, 6 ppm Cx and 69 µg 25-OH-D3, 9 ppm Cx and 103.5 µg 25-OH-D3, or 12 ppm Cx and 138 µg 25-OH-D3. Production performance and internal and external egg quality parameters were not influenced by diet. Eggshell dry weight decreased linearly with increasing supplementation levels, and eggshell ash and calcium content increased quadratically. Plasma phosphorus, calcium, and ionic calcium levels in females and plasma ionic calcium levels in males showed a positive quadratic response to dietary supplementation. Femoral and tibiotarsal dry weight and calcium content were influenced by diet. The 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity in the liver of males and females and in the serum of females showed a positive quadratic relationship with Cx and 25-OH-D3 levels, whereas the malonaldehyde concentration showed a negative quadratic relationship. DPPH scavenging activity in the serum of male quail increased linearly with supplementation. There was a positive quadratic effect on superoxide dismutase gene expression and a positive linear effect on glutathione peroxidase 7 gene expression, suggesting that dietary enrichment with Cx and 25-OH-D3 might help protect spermatozoa against oxidative damage. The dietary supplement was pro-oxidative at high concentrations (above 9 ppm Cx). The results indicate that diets with adequate levels of Cx and 25-OH-D3 have a beneficial effect on calcium and phosphorus metabolism as well as on the antioxidant defense system. We recommend supplementing European quail breeders in the laying period with 6 ppm Cx and 69 µg 25-OH-D3.

Vitamin D ameliorates adipose browning in chronic kidney disease cachexia.

Patients with chronic kidney disease (CKD) are often 25(OH)D3 and 1,25(OH)2D3 insufficient. We studied whether vitamin D repletion could correct aberrant adipose tissue and muscle metabolism in a mouse model of CKD-associated cachexia. Intraperitoneal administration of 25(OH)D3 and 1,25(OH)2D3 (75 µg/kg/day and 60 ng/kg/day respectively for 6 weeks) normalized serum concentrations of 25(OH)D3 and 1,25(OH)2D3 in CKD mice. Vitamin D repletion stimulated appetite, normalized weight gain, and improved fat and lean mass content in CKD mice. Vitamin D supplementation attenuated expression of key molecules involved in adipose tissue browning and ameliorated expression of thermogenic genes in adipose tissue and skeletal muscle in CKD mice. Furthermore, repletion of vitamin D improved skeletal muscle fiber size and in vivo muscle function, normalized muscle collagen content and attenuated muscle fat infiltration as well as pathogenetic molecular pathways related to muscle mass regulation in CKD mice. RNAseq analysis was performed on the gastrocnemius muscle. Ingenuity Pathway Analysis revealed that the top 12 differentially expressed genes in CKD were correlated with impaired muscle and neuron regeneration, enhanced muscle thermogenesis and fibrosis. Importantly, vitamin D repletion normalized the expression of those 12 genes in CKD mice. Vitamin D repletion may be an effective therapeutic strategy for adipose tissue browning and muscle wasting in CKD patients.

Effects of additional dosage of vitamin D3, vitamin D2, and 25-hydroxyvitamin D3 on calcium and phosphorus utilization, egg quality and bone mineralization in laying hens.

Vitamin D is essential for the metabolism of calcium (Ca) and phosphorus (P) in birds. The objective of the study was to evaluate the effect of different isoforms of dietary vitamin D on Ca and P utilization, egg quality, and bone mineralization of laying hens. A total of 42 Lohmann white laying hens at 57 wk of age were randomly assigned to 7 dietary treatments for 6 wk. Dietary treatments were: 3,000 IU/kg Vit D3 as control, and control with additional 3,000 IU/kg 25-hydroxyvitamin D3 (T1), 9,000 IU/kg 25-hydroxyvitamin D3 (T2), 3,000 IU/kg vitamin D3 (T3), 9,000 IU/kg vitamin D3 (T4), 3,000 IU/kg of vitamin D2 (T5), or 9,000 IU/kg of vitamin D2 (T6). Egg production and egg quality were measured weekly. Fecal samples were collected at weeks 2 and 6 to measure Ca and P utilization. After 6 wk, the left tibia and femurs were collected to measure bone mineral density (BMD) and bone mineral content (BMC). A 1-way ANOVA with Tukey HSD means separation test was used for statistical analysis. There were no significant differences in egg production, egg quality, BMD, or BMC of tibia and femurs among the treatments (P > 0.05). T6 significantly reduced feed intake (P < 0.05). The apparent total tract digestibility (ATTD) of Ca was higher (P < 0.012) in treatments supplemented with additional vitamin D, irrespective of forms. The ATTD of P was higher (P < 0.0001) in T5 compared to the other treatments at both time points. The utilization of Ca and P by laying hens can be improved through the addition of different isoforms of vitamin D in diets. However, additional vitamin D supplementation to laying hens, regardless of forms, had no effect on either bone mineralization or measures of egg quality.

Feeding supplemental 25-hydroxyvitamin D3 increases serum mineral concentrations and alters mammary immunity of lactating dairy cows.

Objectives were to determine the effects of feeding supplemental 25-hydroxyvitamin D3 [25(OH)D3] on concentrations of vitamin D metabolites and minerals in serum, mammary immune status, and responses to intramammary bacterial infection in dairy cows. Sixty multiparous, pregnant lactating Holstein cows with somatic cell count <200,000/mL were blocked by days in milk and milk yield and randomly assigned to receive a daily top-dressed dietary supplement containing 1 or 3 mg of vitamin D3 (1mgD or 3mgD), or 1 or 3 mg 25(OH)D3 (1mg25D or 3mg25D) for 28 d (n = 15/treatment). Cows were kept in a freestall barn and fed a total mixed ration in individual feeding gates. Individual dry matter intake (DMI) and milk yield were recorded daily, and milk and blood samples were collected at 0, 7, 14, and 21 d relative to the start of treatment. At 21 d, cows fed 1mgD and 3mg25D received an intramammary challenge with Streptococcus uberis. Cows were observed for severity of mastitis, and blood and milk samples were collected every 12 h to measure inflammation. The 1mg25D and 3mg25D cows had greater serum 25(OH)D3 concentrations at 21 d compared with 1mgD and 3mgD cows (62 ± 7, 66 ± 8, 135 ± 15, and 232 ± 26 ng/mL for 1mgD, 3mgD, 1mg25D, and 3mg25D, respectively). The 3mg25D cows had greater concentrations of Ca and P in serum at 21 d compared with other treatments (Ca = 2.38, 2.4, 2.37, and 2.48 ± 0.02 mM, 1.87, 1.88, and 2.10 ± 0.08 mM for 1mgD, 3mgD, 1mg25D, and 3mg25D, respectively). Yields of milk and milk components, DMI, body weight, and concentrations of 1,25-dihydroxyvitamin D and Mg in serum did not differ among treatments. Abundance of mRNA transcripts for interleukin-1ß (IL1B) and inducible nitric oxide synthase (iNOS) in milk somatic cells before S. uberis challenge were increased in cows fed 25(OH)D3 compared with cows fed vitamin D3. Furthermore, IL1B, iNOS, ß-defensin 7, and ß-defensin 10 in milk somatic cells increased as concentrations of 25(OH)D3 increased in serum. Cows fed 3mg25D had less severe mastitis at 60 and 72 h after challenge with S. uberis compared with cows fed 1mgD. Concentrations of bacteria, somatic cells, and serum albumin in milk after challenge did not differ between treatments; however, an interaction between treatment and day was detected for lactate dehydrogenase in milk. Expression of adhesion protein CD11b on milk neutrophils after the S. uberis challenge was greater among 3mg25D cows compared with 1mgD cows. Transcripts of CYP24A1 and iNOS in milk somatic cells during mastitis also were greater in 3mg25D cows compared with 1mgD cows. Feeding 25(OH)D3 increased serum 25(OH)D3 more effectively than supplemental vitamin D3, resulting in increased serum mineral concentrations, increased expression of vitamin D-responsive genes, and altered immune responses to intramammary bacterial challenge.

The dietary supplementation of canthaxanthin in combination with 25OHD3 results in reproductive, performance, and progeny quality gains in broiler breeders.

The objective of this study was to evaluate the effects of the dietary supplementation of canthaxanthin and cholecalciferol (25OHD3) to broiler breeders and their progenies on their performance. Eighty 25-wk-old Cobb 500 broiler breeders were distributed according to a completely randomized design in 2 experimental treatments, with 10 replicates of 4 birds each. The treatments consisted of the supplementation or not of a corn-soybean meal diet with canthaxanthin + 25OHD3 to broiler breeders fed from 25 to 62 wk of age. Egg production, fertility, hatchability, hatchability of fertile eggs, embryonic mortality, and egg specific gravity were evaluated. Breeders were inseminated at 35, 45, and 62 wk of age, the eggs were incubated, and the performance of the progenies was evaluated. From the progeny of each breeder age, 300 male chicks were distributed according to a completely randomized design in a 2 × 2 factorial arrangement (chicks from breeders supplemented or not with canthaxanthin + 25OHD3 and chicks supplemented or not with canthaxanthin + 25OHD3), totaling 4 treatments with 5 replicates of 15 birds each. Canthaxanthin + 25OHD3 were supplied to the chicks until 21 D of age. The combination of canthaxanthin and 25OHD3, containing 69 mg of 25OHD3 and 6 g of canthaxanthin, was supplemented at 1 kg/t of feed. Breeders supplemented with canthaxanthin + 25OHD3 showed higher egg production, total hatchability, hatchability of fertile eggs, and lower early embryo mortality compared with those fed the control diet. Broilers from breeders fed canthaxanthin + 25OHD3 and supplemented with this additive up to 21 D of age presented a better feed conversion ratio and higher carcass and breast yields than those derived from nonsupplemented breeders.