Effects of experimental diabetes on the vitamin D metabolism of pregnant rats and their fetuses

Summary The effects of streptozotocin-induced diabetes on the vitamin D metabolism of pregnant rats were investigated in mothers and their fetuses, 11 and 14 days after streptozotocin (SZ) injection, i.e., on days 18 and 21 of gestation. In the mothers' plasma, the levels of 25-hydroxycholecalciferol (25OHD) and 1,25-dihydroxycholecalciferol (1,25(OH)₂ D) were not different from control levels on day 18, but on day 21, 25OHD had increased, 1,25 (OH)₂ D had diminished, and significant hypercalcemia was noted (10.1±0.27 mg/dl vs. 9.47±0.19 mg/dl, mean ±SD). In hyperglycemic fetuses from the diabetic mothers, plasma insulin levels were reduced at day 18 but enhanced at day 21. 25OHD levels were not different from those of the controls at day 18, but were lower at day 21 (2.12±0.70 ng/g BW, n=13, vs. 3.75±1.40 ng/g BW n=29 controls, means ±SD). Fetal body levels of 1,25 (OH)₂ D were lower than that in the controls at day 18 (16.6±2.9 pg/g BW, n=9×2, vs. 28.7±6.3 pg/g BW, n=7×2, mean ±SDP <0.001), but identical to control levels on day 21. The role of fetal or placental enzymes in the regulation of vitamin D metabolism in fetuses is discussed.     

The effect of vitamin A deficiency in maternal rats on tumor formation in filial rats

Purpose

We established a vitamin A-deficient (VAD) model of pregnant rats to evaluate the effect of vitamin A deficiency in maternal rats on tumor formation in filial rats.

Methods

Ten pregnant Wistar rats were divided into 2 groups: (1) VAD group, 6 rats were given nonvitamin A diet from 2 weeks before mating till delivery and (2) normal diet (ND) group, 4 rats were given normal diet. Twenty random neonatal rats from each group were killed on the next day of delivery. The rest neonates were given normal diet for 1 year until killed. Serum levels of vitamin A, morphology of the kidney, incidence of tumor formation, and retinoid X receptor (RXR) α messenger RNA (mRNA) expression in renal tissue were assessed for the filial rats.

Results

Fifty-six and 49 neonatal rats were born for VAD group and ND group, respectively. The detection rate of nephrogenic rests (NRs) from neonates in VAD group (50%) was significantly higher than that in ND group (20%; P < .05). The incidence of nephroblastoma was 13.9% in filial rats of VAD group and 0% for ND group. The detection rate of NRs for filial rats of VAD group (30.6%) was significantly higher than that of ND group (6.9%; P < .01). The expression of RXRα mRNA in tumor tissue of the filial rats of VAD group (3.17 ± 0.15) was significantly lower than that in kidney tissue of ND group (3.58 ± 0.20; P < .01).

Conclusion

Deficiency in vitamin A for pregnant rats resulted in renal dysplasia, increased NRs, and higher incidence of nephroblastoma.     

Association of vitamin D and magnesium with insulin sensitivity and their influence on glycemic control

Insulin resistance increases the risk of developing diabetes, and the degree of resistance influences the glycemic control of patients with diabetes. Numerous researchers have focused on improving insulin sensitivity in order to prevent diabetes-related complications and other chronic diseases. Several studies have also linked vitamin D levels to insulin secretion and resistance, given that both vitamin D and its receptor complex play important roles in regulating pancreatic β-cells. It has been suggested that vitamin D supplementation improves vitamin D levels, but further research is needed to confirm this as neither insulin function nor glycemic control improves when vitamin D levels increase. Magnesium is a cofactor for many enzymes. Although the role of magnesium in the management of diabetes has long been evaluated, it has not yet been determined whether magnesium supplements improve insulin function. However, several researchers have found that patients with good glycemic control have high magnesium levels. Magnesium is closely related to vitamin D and is necessary for the transport and activation of vitamin D in humans. Combined supplementation with vitamin D and magnesium improves glycemic control in patients with diabetes.     

High-dose oral vitamin D3 supplementation in rheumatology patients with severe vitamin D3 deficiency

ObjectivesRecent large trials indicate that adherence associated with a daily regimen of vitamin D is low and limits anti-fracture efficacy with vitamin D supplementation. The aim of this report is to describe changes of 25-hydroxyvitamin D (25(OH)D) serum concentrations achieved with a single oral dose of 300 000 IU vitamin D3.MethodsOver a course of 4 months, we identified 33 elderly with severe vitamin D deficiency (25(OH)D < 25 nmol/l) on admission to acute care. Patients were admitted for musculoskeletal pain, bone disease, or gait abnormalities. The mean age was 80.5 years (SD ± 6.1). All patients were treated with a single oral dose of 300 000 IU D3 in combination with 500–1000 mg calcium supplements per day depending on their dietary calcium intake.ResultsBaseline mean 25(OH)D serum concentrations were 15 nmol/l (SD ± 5.5). Mean 25(OH)D serum concentrations increased to 81.4 nmol/l (SD ± 29.7) at 3 months (29 patients) and were still 69.0 nmol/l (SD ± 17.9) at 6 months (26 patients). Mean serum calcium levels were 2.24 mmol/l (SD ± 0.11) at baseline, 2.28 mmol/l (SD ± 0.18) at 3 months, and 2.28 mmol/l (SD ± 0.13) at 6 months. Two patients with mild hypercalcemia (2.69 mmol/l) at 3 months had normal values at 6 months.ConclusionBased on our observations, a single oral dose of 300 000 IU vitamin D3 raises mean 25(OH)D serum concentrations to the target mean of above 75 nmol/l at 3 months and a mean level of 69 nmol/l at 6 months. As calcium absorption is enhanced with higher 25(OH)D serum concentrations, calcium supplementation may need downward adjustment with this regimen to avoid mild hypercalcemia.     

Studies on the role of 24-hydroxylation of vitamin D in the mineralization of cartilage and bone of vitamin D-deficient rats

Summary To test the importance of 24-hydroxylation of vitamin D₃ on bone mineralization, rat pups born to vitamin D-deficient females were given either 25-hydroxyvitamin D₃ or 24,24-difluoro-25-hydroxyvitamin D₃ for 16 days beginning at the time of weaning. Following such treatment analysis of blood samples revealed no detectable 24R,25-(OH)₂D₃ and 1,25-(OH)₂D₃ in the rats given the difluoro compound while revealing the expected 24,24-difluoro-25-hydroxyvitamin D₃ and 24,24-difluoro-1,25-dihydroxyvitamin D₃. The rats given 25-hydroxyvitamin D₃ had the expected levels of 25-hydroxyvitamin D₃, 24,25-dihydroxyvitamin D₃, and 1,25-dihydroxyvitamin D₃. Following sacrifice at day 17, postweaning bone mineralization and modeling were studied in long bones using histological methods. Bones taken from vitamin D-deficient rats at the beginning and end of the experimental period had lesions typical of rickets. These included wide growth plates, excessive amounts of osteoid, and metaphyseal fibrosis. Following treatment with either 25-hydroxyvitamin D₃ or 24,24-difluoro-25-hydroxyvitamin D₃, bone mineralization returned to normal. Growth plate widths and the amount of osteoid on bone surfaces were both substantially reduced and to a similar degree in both treatment groups. Normal cartilage core formation and trabecularization of the metaphyseal primary spongiosa were also restored to a similar degree in both groups. In effect, no difference was observed in any bone parameter studied between the 25-hydroxyvitamin D₃- and the 24,24-difluoro-25-hydroxyvitamin D₃-treated animals. These results provide strong evidence that 24-hydroxylation of the vitamin D molecule plays little or no role in the modeling and mineralization of bone.     

Effect of vitamin D deficiency on urinary excretion of connective tissue derivatives (hydroxyproline and glycosaminoglycans) in rats

Summary The urinary excretion of two connective tissue metabolites was studied in both control and vitamin D deficient rats. Hydroxyproline (HyPRO) excretion was determined after 2, 13 and 22 months (experiment I). It decreased with aging in animals receiving the control diet. On the contrary, this excretion increased as a function of age in vitamin D deficient animals. At the age of 22 months, HyPRO excretion was respectively 31 and 1708 μg a day in control and deficient animals. HyPRO and glycosaminoglycans (GAG) excretion was measured on a group of both control and vitamin D deficient rats at the age of 21 months (experiment II). These results confirm the high excretion of HyPRO in deficient animals. On the contrary, the GAG excretion was higher in control animals than in deficient ones, the mean excretion being respectively 412 and 234 μg a day.     

Mechanical stress regulates bone regulatory gene expression independent of estrogen and vitamin D deficiency in rats

Mechanical stress determines bone mass and structure. It is not known whether mechanical loading affects expression of bone regulatory genes in a combined deficiency of estrogen and vitamin D. We studied the effect of mechanical loading on the messenger RNA (mRNA) expression of bone regulatory genes during vitamin D and/or estrogen deficiency. We performed a single bout in vivo axial loading with 14 N peak load, 2 Hz frequency and 360 cycles in right ulnae of nineteen weeks old female control Wistar rats with or without ovariectomy (OVX), vitamin D deficiency and the combination of OVX and vitamin D deficiency (N = 10/group). Total bone RNA was isolated 6 hours after loading, and mRNA expression was detected of Mepe, Fgf23, Dmp1, Phex, Sost, Col1a1, Cyp27b1, Vdr, and Esr1. Serum levels of 25(OH)D, 1,25(OH)₂D and estradiol were also measured at this time point. The effect of loading, vitamin D and estrogen deficiency and their interaction on bone gene expression was tested using a mixed effect model analysis. Mechanical loading significantly increased the mRNA expression of Mepe, and Sost, whereas it decreased the mRNA expression of Fgf23 and Esr1. Mechanical loading showed a significant interaction with vitamin D deficiency with regard to mRNA expression of Vdr and Esr1. Mechanical loading affected gene expression of Mepe, Fgf23, Sost, and Esr1 independently of vitamin D or estrogen, indicating that mechanical loading may affect bone turnover even during vitamin D deficiency and after menopause.     

孕妇维生素D缺乏及其对胎儿的影响

目的维生素D缺乏累及的人群非常广泛,而且与多种疾病的发生有关,特别是孕妇维生素D缺乏还可能影响到胎儿的健康。目前关于我国女性维生素D的资料报道很少。本研究的目的是了解健康孕妇(孕15~21w)以及同龄未孕对照组妇女维生素D水平以及孕妇维生素D水平对新生儿出生大小的影响。方法本文随机选取单胎妊娠孕妇63例和同龄对照妇女35例,用酶联免疫的方法测定血中25-羟维生素D水平。结果显示99%的检测病例25-羟维生素D水平低于正常值(≥75 nmol/L),有近93%的妇女为维生素D缺乏(50 nmol/L)。孕妇25-羟维生素D水平(28.40±9.19 noml/L)明显低于对照组妇女(38.46±10.77 noml/L;P0.001),两组维生素D缺乏的比率分别为孕妇96.8%,对照组85.7%,但重度维生素D缺乏的病例孕妇组接近半数,而对照组为零。新生儿身长与孕妇25-羟维生素D水平成显著正相关(r=0.323;P0.01。结论我国北方女性特别是孕妇是维生素D缺乏患病的高危人群,而且孕妇维生素D缺乏可能会影响到胎儿的生长发育。应该积极采用有效的措施防治维生素D的缺乏,从而减少维生素D缺乏对健康的影响对提高人口质量将具有重要的意义。     

Role of vitamin D in maternal skeletal changes during pregnancy and lactation: A histomorphometric study

Summary The effect of vitamin D on bone changes during the reproductive cycle in female rats has been investigated. One group of female rats was maintained on a vitamin D-deficient diet and another group on a vitamin D-replete diet from weaning. Both groups were mated with normal males and changes in their bones were determined histomorphometrically during pregnancy, lactation, and after weaning. All vitamin D-deficient rats had bone changes typical of rickets. Pregnancy caused significant reductions in mineralized tissue of trabecular and cortical bone in the vitamin D-deficient rats. Lactation caused further significant reductions in mineralized tissues of cortical and trabecular bone in both the vitamin D-deficient and vitamin D-replete animals, with the greatest changes seen at weaning. Some restoration of mineralized tissues occurred following weaning. There was an increase in tetracycline-labeled bone surface in the vitamin D-replete animals during lactation, likely due to an increase in bone formation rates. In the vitamin D-deficient animals during lactation, there was a decrease in tetracyclinelabeled bone surface, likely due to severely depressed bone mineralization. These results indicate that the mobilization of calcium from bone to maintain pregnancy and lactation occurs by a mechanism independent of vitamin D.     

维生素A缺乏对1,2-二甲基肼诱导大鼠肾母细胞瘤发生的影响

目的探讨维生素A缺乏对1,2二甲基肼(DMH)诱导大鼠肾母细胞瘤成瘤的影响。方法建立DMH诱导的Wistar幼鼠肾母细胞瘤动物模型,设立正常饮食组,维生素A缺乏饮食组和对照组,每组各30只。按Beckwith的诊断标准确定肾母细胞瘤和肾源性剩余。观察3组鼠肾母细胞瘤成瘤率和肾源性剩余发生率,流式细胞仪检测肿瘤标本的DNA指数(DI)和S期细胞分布,RTPCR方法检测肿瘤标本RARαmRNA基因表达。结果对照组无肿瘤发生,正常饮食组大鼠成瘤率13.3%,维生素A缺乏饮食组成瘤率36.7%,组间差异有统计学意义(P<0.05)。2组肾源性剩余发生率分别为3.3%和16.7%,差异有统计学意义(P<0.05)。维生素A缺乏饮食组肿瘤DI值1.49±0.22,S期细胞含量(24.1±5.1)%,明显高于正常饮食组的(1.32±0.13)和(15.2±3.6)%。维生素A缺乏饮食组RARαmRNA表达强度(0.18±0.03)明显低于正常饮食实验组(0.49±0.05)。结论维生素A缺乏影响后肾胚基分化、RARαmRNA表达和细胞增殖,导致DMH诱导的大鼠肾母细胞瘤成瘤率增高。     

Severe obesity and vitamin D deficiency treatment options before bariatric surgery: a randomized clinical trial

BackgroundObesity, which has various complications and co-morbidities, is an epidemic issue worldwide. Vitamin D deficiency (VDD) is a well-known metabolic disorder among patients with severe obesity. While they are good candidates for bariatric surgery, this deficiency can affect the outcome of surgery negatively.ObjectivesThe aim of this study was to compare 3 different VDD treatment strategies for use before bariatric surgery and compare serum vitamin D levels after 7 weeks.SettingsUniversity hospital, Isfahan, Iran.MethodsThis was a single-blinded, randomized clinical trial on 100 patients who were referred for bariatric surgery from 2016 to 2018. Vitamin D (VitD) level was checked before surgery for the patients included in the study, if their VitD level was <30 ng/mL. We rechecked their serum VitD in the 8^th week, after 7 weeks of treatment. The participants were randomly allocated into 3 groups: 33 patients were treated with 50,000 units VitD₃ capsules every week for 7 weeks; 33 patients were treated with a single dose of 300,000 units VitD₃ ampoule; and 34 patients were treated with a combination of a half of the injection dose, followed by the oral capsule for 4 weeks.ResultsNo case was lost during the follow-up time. No significant differences were found among the 3 groups in terms of their age (P = .654), body mass index (P = .434), sex (P = .799), initial 25(OH) VitD level (P = .273), and history of supplement use (P = .45). Mean serum VitD levels were 15.21, 13.16, and 13.37 ng/mL, respectively, before the surgery and reached 32.91, 24.74, and 29.49 ng/mL after 7 weeks of treatment in oral, injection, and combined groups, respectively. Finally, the 7-week oral treatment option had significantly higher levels of VitD (P value = .034).ConclusionVDD treatment with 50,000 units VitD₃ capsule every week for 7 weeks before bariatric surgery yields a higher level of VitD. Based on our findings, injectable supplements are not recommended for VDD treatment.     

Studies on the role of vitamin D in early skeletal development,mineralization, and growth in rats

Summary The role of vitamin D in early skeletal development was studied by measuring serum calcium and phosphorus, osseous tissue quantity and mineralization, and endochondral bone elongation in rat fetuses and pups from vitamin D-replete and vitamin D-deficient mothers. At the 20th day of pregnancy there was a slight, yet significant, increase in the amount of osteoid on trabecular bone surfaces in fetuses from vitamin D-deficient mothers. The fetal bones otherwise appeared normal in spite of severe skeletal changes in the vitamin D-deficient mothers. After parturition, the importance of vitamin D in skeletal development becomes progressively more obvious. Serum calcium levels were slightly, yet significantly, lower in vitamin D-deficient than in vitamin D-replete pups and these levels continued to fall in the vitamin D-deficient pups through lactation and after weaning. At 3 days postpartum, there was a small, yet significant, increase in the amount of osteoid on bone surfaces of the vitamin D-deficient pups. The relative amounts of osteoid in the vitamin D-deficient pups continued to increase through lactation and after weaning when compared with vitamin D-replete pups. By the 14th day of lactation and at later periods, there were significant reductions in metaphyseal mineralized tissues in the vitamin D-deficient pups when compared with the vitamin D-replete pups. At weaning and after weaning, there were substantial increases in growth plate thickness and decreases in longitudinal bone growth in the vitamin D-deficient pups when compared with the vitamin D-replete pups. The results from this study indicate that vitamin D does not appear to play a major role in fetal skeletal development. However, after birth, vitamin D becomes progressively more important with age for normal bone development, mineralization, and endochondral growth.     

High prevalence of vitamin D deficiency in women with breast cancer: The first Chilean study

BackgroundBreast cancer (BC) is the leading cause of female death from malignancy worldwide. One factor that has been associated to a higher incidence and poor prognosis is a Vitamin D deficiency (measured as 25-Hydroxi-Vitamin D (25OHD)). Our aim was to determine 25OHD levels in serum samples of Chilean BC patients before endocrine therapy and its association to clinical parameters at the time of diagnosis.MethodsWe analyzed clinical records of 105 women, evaluated at the Cancer Center of the Pontificia Universidad Católica de Chile. Serum levels of 25OHD were determined using a chemiluminescent microparticle immunoassay.ResultsThe prevalence of vitamin D deficiency before endocrine therapy was of 70.5%. Only 7% of our patients showed sufficient vitamin D levels at the beginning of the endocrine treatment. There was a significant correlation found between age and 25OHD levels, and also between body fat percentage and 25OHD levels (r² = 0.04; p = 0.021; r² = 0.028; p = 0.0432, respectively). Summer 25OHD levels were significantly higher than winter levels (p = 0.0322).ConclusionVitamin D deficiency is highly prevalent in Chilean BC women before endocrine therapy and 25OHD levels are inversely correlated to the age and body fat percentage of patients. Further studies are needed to determine causal relationship between vitamin D levels and BC development and outcome.     

Conundrum of vitamin D on glucose and fuel homeostasis

As an endocrine hormone, vitamin D plays an important role in bone health and calcium homeostasis. Over the past two decades, the non-calcemic effects of vitamin D were extensively examined. Although the effect of vitamin D on beta cell function were known for some time, the effect of vitamin D on glucose and fuel homeostasis has attracted new interest among researchers. Yet, to date,studies remain inconclusive and controversial, in part, due to a lack of understanding of the threshold effects of vitamin D. In this review, a critical examination of interventional trials of vitamin D in prevention of diabetes is provided. Like use of vitamin D for bone loss, the benefits of vitamin D supplementation in diabetes prevention were observed in vitamin D-deficient subjects with serum 25-hydroxyvitamin D 50 nmol/L(20 ng/mL). The beneficial effect from vitamin D supplementation was not apparent in subjects with serum 25-hydroxyvitamin D 75 nmol/L(30 ng/mL). Furthermore, no benefit was noted in subjects that achieved serum 25-hydroxyvitamin D 100 nmol/L(40 ng/mL).Further studies are required to confirm these observations.     

The relationship between serum vitamin D concentrations andin vivo tetracycline labeling of osteoid in crush fracture osteoporosis

Summary Twenty of 22 consecutive British patients with crush fracture osteoporosis had transiliac bone biopsies following doublein vivo tetracycline labeling synchronized with the collection of serum for the measurement of vitamin D metabolites. A significant but direct (rather than inverse) relationship was found between 25-hydroxyvitamin D (calcidiol) levels and the fraction of cancellous surfaces covered with osteoid not taking either tetracycline label (r=0.53,P<0.02). There was no correlation with 1,25-dihydroxyvitamin D levels. No patient had frankly thickneded osteoid seams although 3 had reduced but measurable calcidiol levels. These results make it unlikely that the majority of patients with osteoporosis who have osteoid of normal thickness but reduced uptake of tetracycline have a mineralization defect secondary to vitamin D deficiency. The pathophysiological significance of unlabeled osteoid in osteoporosis requires further investigation.     

Association of 1.25 vitamin D3 deficiency, disease activity and low bone mass in ankylosing spondylitis

Vertebral fractures due to osteoporosis are a common but frequently unrecognized complication in established ankylosing spondylitis (AS). It is known that inflammatory activity in rheumatic diseases (i.e., proinflammatory cytokines) itself plays a possible role in the pathophysiology of bone loss. The aim of this study was to analyze whether inflammatory activity and an alteration of the vitamin D metabolism play a substantial role in the loss of bone mass in AS. In this cross-sectional study, 58 patients with established AS and an age- and sex-matched control group were examined. The vitamin D status was investigated, as was, in parallel, the relationship to disease activity (erythrocyte sedimentation rate [ESR], C-reactive protein [CRP], Bath Ankylosing Spondylitis Disease Activity Index [BASDAI]), markers of bone metabolism (parathyroid hormone [PTH], 1.25 vitamin D₃, 25 vitamin D₃), calcium, bone alkaline phosphatase (bone-AP), urine cross-links, and plasma tumor necrosis factor (TNF). Bone mineral density was measured by quantitative computed tomography (QCT) of the lumbar spine. Osteoporosis was diagnosed in early as well as in progressive stages of AS (23/58=39.6%). Furthermore, serum levels of 1.25 vitamin D₃ and PTH were negatively correlated with disease activity and TNF. The excretion of cross-links showed a positive correlation with disease activity and TNF, and 1.25 vitamin D₃ and PTH were positively correlated with bone-AP. TNF also positively correlated with disease activity. AS patients with osteoporosis showed significantly increased CRP, ESR, cross-links and PTH and a significantly decreased 1.25 D₃. Osteoporosis is frequent in AS and high disease activity is associated with an alteration in vitamin D metabolites and increased levels of bone resorption in active AS. Our findings propose a close association of BMD, bone metabolism and inflammatory activity, possibly related to vitamin D inflammation interactions.     

Very high-dose ergocalciferol is effective for correcting vitamin D deficiency in children and young adults with cystic fibrosis

Approximately 10–80% of patients with Cystic Fibrosis (CF) have vitamin D deficiency. Obtaining therapeutic vitamin D levels has been a challenge for CF care providers using current recommended high-dose oral ergocalciferol (400,000 IU over 2 months). The objective of this study was to evaluate the safety and efficacy of a 2-week, very high dose ergocalciferol (700,000 IU over 14 days) repletion strategy in children and young adults with CF.As part of a quality improvement initiative, a prospective cohort study was performed from January through May 2007. Phase I included identifying individuals with CF who were subtherapeutic in 25-OH D. In phase II, 50,000 IU of ergocalciferol was prescribed for a 14 day term and administered daily. During phase III, a post treatment 25-OH D level was obtained to determine improvement. Baseline demographics and clinical characteristics were obtained at study entry. Stratification of the post 25-OHD levels was defined.Eighteen individuals with CF participated in the study. The mean age was 17 ± 5 years (range 6–25 years). One hundred percent were pancreatic insufficient and required pancreatic enzyme replacement. All 18 had 25-OHD levels less than 30 ng/mL pre-treatment.Seventeen of the 18 (94%) participants became therapeutic in the 2-week interval. No patients had values considered high abnormal (100–150 ng/mL) or toxic (> 150 ng/mL). Mean change was noted at an increase of 37.3 ± 22 ng/mL in the 2-week period (p < 0.001). Pre and peripubertal individuals had a significantly greater increase in 25-OH D levels.The results of this study demonstrate that very high dosing of vitamin D using oral ergocalciferol over a 14 day period is an effective strategy in achieving therapeutic levels of 25-OH vitamin D in children and young adults with CF. We believe this regimen deserves further study.     

Immobilization osteoporosis and active vitamin D: Effect of active vitamin D analogs on the development of immobilization osteoporosis in rats

Summary The therapeutic effects of vitamin D analogs, 1,24(R)-dihydroxycholecalciferol [1,24(R)(OH)₂D₃], 1,24(S)-dihydroxycholecalciferol [1,24(S)(OH)₂D₃], and 1,25-dihydroxycholecalciferol [1,25(OH)₂D₃] on immobilization osteoporosis were studied in rats. The right hind limb was immobilized through application of a plaster cast following the section of the sciatic nerve. The left hind limb was intact. Vitamin D analogs were orally administered for 6 weeks at dose levels of 0.02 and 0.10μg/kg/day, respectively. The mean lengths of the immobilized femurs were not significantly different from those of the intact femurs in all the experimental groups. In the immobilized femur of animals treated with 1,24(R)(OH)₂D₃, 0.10μg/kg, dry and ash weights were heavier and calcium and phosphorus contents greater than those in the nontreated group. Furthermore, the amount of calcified bone mass and the cortical thickness of the femurs of the immobilized limb in 1,24(R)(OH)₂D₃-treated animals were greater than those in the nontreated animals. Treatment with 1,25(OH)₂D₃ at 0.10μg/kg caused an increase of the bone mass in both immobilized and intact femurs when compared with those of the control group. It was concluded that the administration of 1,24(R)(OH)₂D₃ diminished the effect of immobilization in the development of osteoporosis without any side effects.     

Sequential changes in mineral metabolism and serum vitamin D metabolite concentrations produced by phenobarbital administration in the rat

Summary We examined the effect of daily phenobarbital administration on serum vitamin D metabolite levels and indices of vitamin D biologic activity in 7-week-old male rats maintained on parenteral vitamin D supplementation (125 ng/day). Treatment with phenobarbital (75 mg/kg/day) produced a biphasic response in parameters of vitamin D biologic effect, including serum calcium concentration, serum inorganic phosphate concentration, and intestinal⁴⁵Ca calcium absorption. An initial increase in these values, maximal after 3–5 days of treatment, was followed by a subsequent decline to subnormal levels by day 21. A parallel biphasic pattern was observed for serum 25-hydroxyvitamin D (25OHD) concentration. Serum 25OHD reached a peak increase of 87% above control levels (P<0.01), observed after 5 days of treatment, and subsequently declined to 62% of control animal values (P<0.01) by 21 days. Serum 24,25(OH)₂D concentration followed a similar course and exhibited a strong positive correlation with serum 25OHD concentrations (r=0.74,P<0.01). In contrast, serum 1,25(OH)₂D concentration was not significantly different from control values after 5 days but was increased 80% over control values (P<0.05) by day 21. Serum vitamin D concentration declined progressively in treated animals, falling to 50% of control levels (P<0.05) by day 5 and to 27% of control levels (P<0.001) by day 21. At the point of maximal increase in serum 25OHD concentration, hepatic microsomal vitamin D₃-25-hydroxylase activity was not increased in the treated animals whereas hepatic mitochondrial vitamin D₃-25-hydroxylase activity was increased by 2.4-fold. Increased hepatic mitochondrial vitamin D₃-25-hydroxylase activity persisted through 21 days of phenobarbital treatment. It is concluded that phenobarbital administration in the rat produces an initial increase in vitamin D biologic effect which correlates temporally with increased circulating levels of 25OHD, the latter possibly resulting from increased hepatic mitochondrial vitamin D-25-hydroxylase activity. A subsequent decline in serum 250HD concentration may be the result of decreased availability of vitamin D as substrate. This sequence of alterations in vitamin D metabolism bears potentially important implications for the timing of prophylactic vitamin D supplementation in patients treated with anticonvulsant drugs.     

Effect of vitamin D status on the activity of carbonic anhydrase in chicken epiphysis and kidney

Summary Chickens were raised for 6 weeks from the date of hatch under red light on a vitamin D-free diet; controls were given an oral vitamin D supplement. Vitamin D-deficient animals showed decreased total serum calcium concentration and decreased DNA content in epiphysis and kidney homogenates. In calcifying epiphysis, total carbonic anhydrase (CA) activity was decreased, but activity per μg DNA was slightly increased and specific activity was double that of the controls. Polyacrylamide gel isoelectric focusing after preparation of the enzyme showed a picture similar to that seen after parathyroid hormone (PTH) administration in chicks; therefore, this could be considered a secondary hyperparathyroidism. The CA activation was not seen in the kidney which can be explained by induction of an endogenous inhibitor protein of the cyclic AMP-dependent protein kinase exclusively in the kidney in vitamin D deficiency. In an additional experiment, chickens were raised for 3 weeks from the date of hatch under red light on a vitamin D-free diet. Daily oral substitution by different vitamin D metabolites (1,25 (OH)₂D₃, 25OHD₃, 24, 25(OH)₂D₃) over 7 days led to CA activation compared with controls probably by restoring protein kinase activity in the kidney. Our results show that CA activity is inversely correlated with serum calcium concentrations which is in agreement with a regulatory mechanism recently proposed by us.