Journal of Nutrition

Published by American Society for Nutrition
Online ISSN: 1541-6100
Publications
Article
Forty male weanling albino rats were divided into 4 groups and fed 20% casein diets of high fat (40% ) or low fat (5% ) content with adequate or exces sive quantities of niacin. The niacin content of both the high and low fat diets was increased by supplementing the diets with 0.1% of niacin. Two control groups, one fed high fat and the other low fat, containing adequate (0.5 mg/100 g) quantities of niacin, were fed the diet simultaneously. Blood samples were collected for pyridine nucleotide determinations on the twenty-first and the forty-second day of the experi ment. The animals were killed after 44 days and the livers analyzed for pyridine nucleotide concentration, fatty acid oxidase activity, and proximate composition. Re sults from this study indicate that excess niacin enters metabolic pathways to produce at least 2 unrelated effects, increased concentrations of pyridine nucleotides in blood and liver and increased levels of fat in the liver. The first effect occurs regardless of the level of fat in the diet, but the second occurs only in conjunction with a high level of dietary fat. The possibility that an increased consumption of niacin and fat increases the animals' requirement for choline is discussed.
 
Composition of the basal diet for gestating gilts 1 
Article
In this study, we determined the effects of L-arginine supplementation during early pregnancy on embryonic/fetal survival and growth in gilts. Gilts were housed individually in pens and fed twice daily 1 kg of a corn- and soybean meal-based diet supplemented with 0.0, 0.4, or 0.8% L-arginine (wt:wt) between d 0 and 25 of gestation (10 gilts/treatment). The diets were made isonitrogenous by addition of appropriate amounts of L-alanine. At d 25 of gestation, gilts were fed L-alanine or L-arginine and hysterectomized 30 min later to obtain uteri and conceptuses (embryos and associated fetal membranes and fluids). Dietary supplementation with 0.4 or 0.8% L-arginine enhanced (P < 0.05) its concentrations in maternal plasma (64 and 98%, respectively) as well as the vascularity of chorionic and allantoic membranes, compared with the control group. Reproductive performance [numbers of corpora lutea (CL) and fetuses, placental and fetal weights, and embryonic mortality] did not differ between the 0.4% Arg and control groups. However, supplementation with 0.8% L-arginine decreased (P < 0.05) uterine weight (-20%), total number of fetuses (-24%), CL number (-17%), total fetal weight (-34%), total volume of allantoic and amniotic fluids (-34 to 42%), concentrations of progesterone in maternal plasma (-33%), as well as total amounts of progesterone (-35%), estrone (-40%), and estrone sulfate (-37%) in allantoic fluid, compared with the control group. These results indicate that dietary supplementation with 0.8% L-arginine between d 0 and 25 of gestation, while increasing placental vascularity, adversely affects the reproductive performance of gilts.
 
Chemical structures of linoleic acid (18:2), 13-hy- droperoxylinoleic acid (13-HPODE) and 13-hydroxylinoleic acid (13- HODE).  
Nutrient composition of the animal diets fed to LDL r / mice in studies 1 and 2 1
Article
Studies suggest that heated oils contribute to the presence of oxidized components in the circulating lipoproteins and to the development of atherosclerosis in animals. We evaluated the effects of 11-13 wk of consumption of a well defined dietary oxidized fatty acid, 13-hydroxylinoleic acid (13-HODE) (8 mg), on atherosclerotic lesion development and plasma cholesterol concentrations in mice fed diets varying in fat and cholesterol contents. LDL receptor knockout mice were used in two feeding studies. In study 1, oxidized fatty acid consumption in association with a high fat diet increased aortic lesion areas by >100% (P < 0.05). Surprisingly, oxidized fatty acid intake also tended to increase plasma total cholesterol (P = 0.12) and LDL cholesterol (P < 0.05) as well as oxidative stress as measured by higher levels of autoantibodies to oxidatively modified proteins (P = 0.008). However, in mice fed a nonpurified diet, oxidized fatty acids were not atherogenic and may even have been beneficial, as indicated by a lower plasma triglyceride (TG) concentration (P < 0.05). In study 2, mice were fed either a high fat, medium fat or low fat diet to evaluate whether the increase in aortic lesions due to oxidized fatty acid consumption in study 1 was a result of the associated higher plasma total and LDL cholesterol concentrations. In study 2, 13-HODE-treated mice in the medium and low fat diet groups but not those fed the high fat diet had larger atherosclerotic lesions (P < 0.05). Additionally, plasma total and LDL cholesterol as well as TG were not affected by HODE treatment. However, the total cholesterol:HDL cholesterol ratio was higher in treated mice (P < 0.05) and HDL cholesterol was lower in HODE-treated mice that were fed the low fat diet (P < 0.05). Our results suggest that, in mice fed cholesterol, oxidized fatty acids may be atherogenic, both in terms of increased oxidative stress (as seen in study 1) and by increasing the atherogenicity of the plasma cholesterol profile.
 
Influence of fermented milk supplementation on rotavirus diarrhea in suckling rats. The control groups, CM (n 34) and CF (n 40), were supplemented daily from 2 d of age with milk or with milk fermented by L. casei-114 001; they were inoculated with modified Eagle's medium (MEM) at the age of 5 d. The infected groups, RM (n 39) and RF (n 55), were supplemented with milk or fermented milk from the age of 2 d and were inoculated at the age of 5 d with SA11 rotavirus. Results are expressed as the percentage of rats in each litter delivering feces immediately upon abdominal palpation. Values are means SD. For each time postinfection (p.i.), means with different letters are significantly different (ANOVA and subsequent Fisher's test, P 0.05).
Rotavirus shedding in infected rats supplemented with nonfermented milk (RM) or milk fermented by L. casei DN- 114 001(RF) 1
Sulfated mucin-containing cells in the jejunum of suckling rats infected with SA11 rotavirus, showing the influence of fermented milk supplementation. The control groups, CM (n 34) and CF (n 40), were supplemented daily from 2 d of age with milk or with milk fermented by Lactobacillus casei-114 001; they were inoculated with modified Eagle's medium (MEM) at the age of 5 d. The infected groups, RM (n 39) and RF (n 55), were supplemented with milk or fermented milk from the age of 2 d and were inoculated at the age of 5 d with SA11 rotavirus. Results are expressed as the number of sulfated mucin-containing cells/villi length (n/m). Values are means SD, n 3. *Significantly lower in the RM group compared with the others at the same time postinoculation (ANOVA and subsequent Scheffé F test, P 0.05). Abbreviation used: h p.i., hours postinfection.
Article
Group A rotavirus is the leading cause of diarrhea among children aged 3-36 mo worldwide. Introducing fermented milk products into the infant diet has been proposed for the prevention or treatment of rotavirus diarrhea. The preventive effect of milk fermented by the Lactobacillus casei strain DN-114 001 was studied in a model of germfree suckling rats supplemented daily from d 2 of life and infected with SA11 rotavirus at d 5 (RF group). One group was supplemented with nonfermented milk (RM) and two uninfected groups (CM and CF) received either nonfermented or fermented milk. Frequency and severity of diarrhea were observed. Rats were killed at various times from 0 to 120 h postinfection (p.i.). Bacteria were measured in the intestine, and rotavirus antigens were detected by ELISA in fecal samples and in different parts of the intestine. Histologic observations were made, including vacuolation, morphology of intestinal villi and number of mucin cells. RM rats had diarrhea for 6 d; compared with the CM group, they had alterations of the intestinal mucosa characterized by cellular vacuolation 48 and 72 h p.i. and a lower number of sulfated mucin cells 72 and 96 h p.i. (P: < 0.05). Early supplementation with fermented milk significantly decreased the clinical signs of diarrhea from 24 to 144 h p.i. (P: < 0.05) and prevented rotavirus infection in all sections of the intestine. Histologic lesions of the small intestine were greatly reduced (P: < 0.05) and the number of mucin cells remained unchanged. The data are discussed with respect to the possibility of reducing rotavirus diarrhea in young children by consumption of fermented milk.
 
Article
The effects of storage condition on the shelf life of the AIN-76 diet were investigated. Samples of the diet were stored at -70 degrees, 4 degrees, 20 degrees, and 23-30 degrees under atmospheric air and at 4 degrees and 20 degrees under argon. Levels of vitamin A, thiamine, rancidity, bacteria and mold were monitored during 168 days of storage. A 41.3% loss of vitamin A occurred in samples stored at 23-30 degrees. A marked decline in thiamine was observed in all samples stored at 20 degrees or above. Loss of thiamine was significantly greater in samples stored at 20 degrees than in those stored at 4 degrees, and greater in samples stored under air than those stored under argon. Rancidity reached a level previously shown to be associated with a disagreeable odor and taste (peroxide value greater than 140) with in 30, 50 and 90 days when the diet was stored at 23-30 degrees or 20 degrees under air and 20 degrees under argon, respectively. Peroxide values remained well below 140 in samples stored at 4 degrees or colder. None of the nutritional parameters tested reached unacceptable levels in any of the samples stored at 4 degrees or colder during the 168 days. These results indicate that to achieve maximum shelf life, the AIN-76 diet should be stored at 4 degrees or colder. Although less effective than low temperature, an argon atmosphere extended the shelf life and was additive with lower temperatures.
 
Diet composition Ingredients in basal diet1 Composition 
Effect of fluoride on the uptake of calcium and the release of calcium, hydroxyproline and alkaline phosphalase from embryonic chick bones1 
Article
White Leghorn cockerols were fed a semipurified diet (containing fluoride at 0 to 800 ppm) from the day of hatching. The birds were also injected daily with solutions containing ethane-1-hydroxy-1,1-diphosphonate (EHDP) at concentrations ranging from 5 to 20 mg as phosphorus/kg of body weight or isotonic saline. Changes in bone mineral composition and plasma Ca, Mg and F were determined as well as alterations in bone pyrophosphatase and the amounts of bone cyclic-3',5'-adenosine monophosphate (cAMP). Since dietary fluoride in high amounts is known to stimulate new bone formation, the purpose of the study was to assess whether a potent inhibitor of mineralization, such as EHDP, might alter the response to high dietary fluoride. Although bone fluoride and magnesium were increased in relationship to dietary fluoride, the administration of EHDP had little effect on these changes at low to moderate levels of fluoride. The levels of soluble bone pyrophosphatase were not greatly influenced by changes in dietary fluoride. Administration of EHDP at 20 mg P/kg, however, typically decreased the levels of bone pyrophosphatase. Bone cAMP did not appear to be influenced by either dietary fluoride or EHDP. Using tissue culture techniques, the effects of fluoride on calcium uptake and release from embryonic chick bone were also studied. The presence of 1 mM fluoride in the medium appeared to stimulate calcium uptake and reduced calcium release from chick bone. In general, the results were in keeping with previous suggestions that the major effect of fluoride on bone is the formation of fluoroapatite and subsequent effects this may have on other metabolic alterations.
 
Article
Chemical composition of liver chromatin was determined for rats fed a complete stock diet, or a diet lacking protein or fat. High carbohydrate, fat-free (diet 1) and low carbohydrate, protein-free (diet 2) diets were selected because they elicit structural alteration in chromatin as measured by incubation with micrococcal nuclease (E.C. 3.1.4.7). In the present study, either dietary treatment caused an increase in mass ratios of RNA:DNA and nonhistone:DNA, relative to control ratios. The nonhistone-DNA ratios in liver of rats fed diet 1 or diet 2 were 2.4-fold and 3.5-fold, respectively, larger than control ratios. The histone:DNA ratio remained relatively constant among all three dietary treatments. Liver nuclei were purified from rats fed each dietary treatment and were solubilized in 9 M urea. The nuclear proteins were analyzed by two-dimensional electrophoresis and visualized with a silver treatment that stains proteins in color. The electrophoretograms presented show preferentially proteins with low molecular weights and acidic pIs, two characteristics of nonhistones. The two-dimensional protein patterns are nearly identical for nuclear proteins from all three treatments. Analysis of the electrophoretograms indicates that the diet-induced increased nonhistone:DNA ratios are apparently not attributable to new species of protein, but rather to increased relative abundance of many proteins in the existing populations.
 
Article
Maltitol is fermented in the colon due to only partial hydrolysis in the small intestine. In the present study, we examined effects of dietary maltitol on dimethylhydrazine-induced intestinal tumor in rats. In experiment 1, rats were fed a fiber-free diet or diets supplemented with 1 or 5 g/100 g maltitol for 27 wk. Each group of rats was injected with dimethylhydrazine or vehicle alone for the first 14 wk of the experimental period. Maltitol supplementation at 1 g/100 g of the diet significantly reduced tumor incidence in the cecum and the 5% supplement reduced tumor incidence in both the cecum and proximal colon in dimethylhydrazine-treated rats. In experiment 2, we investigated the effect of the 1 g/100 g maltitol diet on the short chain fatty acid concentrations in cecal contents of placebo and dimethylhydrazine-treated rats. Intake of the 1 g/100 g maltitol diet doubled (P < 0.05) the concentration of butyrate but did not affect acetate or propionate in the cecal contents. These results suggest that dietary maltitol has a protective effect against dimethylhydrazine-induced tumors in rat cecum and proximal colon and that butyrate produced by bacterial fermentation of maltitol in the cecum may be involved in the protection.
 
Article
Torula yeast diets deficient (less than or equal to 0.02 ppm) in selenium (Se) and a control diet supplemented with sodium selenite (0.1 ppm Se), were fed to 52 male Sprague-Dawley rats per diet for 23 wk following weaning. 1,2-dimethylhydrazine (DMH) was administered (20 mg/kg body weight) in 20 weekly i.p. injections after 3 wk of feeding. After 23 wk, 67% of the rats fed the Se-deficient diet had intestinal adenocarcinomas versus 63% on the 0.1 ppm Se diets. Diet also had no effect on the number or size of tumors per tumor-bearing animal or on the location of the tumors within the colon. The effects of Se deficiency on the hematological variables of white blood cell count, hematocrit, serum urea nitrogen and cholesterol concentrations were examined. Serum urea nitrogen levels were lower in Se-deficient DMH-treated rats (9.6 +/- 0.7 vs. 13.7 +/- 0.9 mg/dl, P less than 0.01) and serum cholesterol was higher in Se-deficient DMH-treated animals (69.0 +/- 5.5 vs. 50.7 +/- 3.9 mg/dl, P less than 0.05). Body weights of the Se-depleted group were lower in the DMH-treated animals (P less than 0.01) although food consumption did not differ. Controls without DMH did not show differences due to Se status for any variable examined. Se deficiency appears to affect DMH toxicity but nutritional adequacy (0.1 ppm Se) does not inhibit tumor development. The results of this study do not support the belief that Se deficiency enhances colon carcinogenesis.
 
Article
Iron deficiency anemia in early life is related to altered behavioral and neural development. Studies in human infants suggest that this is an irreversible effect that may be related to changes in chemistry of neurotransmitters, organization and morphology of neuronal networks, and neurobiology of myelination. The acquisition of iron by the brain is an age-related and brain-region-dependent process with tightly controlled rates of movement of iron across the blood-brain barrier. Dopamine receptors and transporters are altered as are behaviors related to this neurotransmitter. The growing body of evidence suggests that brain iron deficiency in early life has multiple consequences in neurochemistry and neurobiology.
 
Effect of diet and 1,2-dimethylhydrazine on individual fecal bile acid metabolites1 Individual bile acids 
Article
The effects of varying colon bile acid concentrations on rat colon epithelial cell proliferation were studied. Bile acid concentrations were altered by intrarectally injecting either deoxycholic or lithocholic acid for 4 weeks or by increasing the dietary fat or fiber (wheat bran, agar, or carrageenan) intake for 4 weeks. 1,2-Dimethylhydrazine (DMH) was s.c. injected into half of the rats 1 week before treatments began. Colon epithelial cell proliferation was measured by [3H]thymidine autoradiography of colon crypts. Rats injected with DMH had more DNA-synthesizing cells per crypt. Neither bile acid injection nor any of the diets altered the number of DNA-synthesizing cells per crypt. DMH injections, deoxycholic and lithocholic acid intrarectal injections, and dietary agar and wheat bran all increased the total number of cells per crypt. High fat diets and dietary carrageenan did not affect cell number. All diets containing fiber lowered total fecal bile acid concentrations, but increasing the fat content of the diet did not affect them. These results indicate that the bile acid injections and dietary agar and wheat bran induce a slight hyperplasia in the colon.
 
Article
Dietary sphingomyelin (SM) inhibits early stages of colon cancer (appearance of aberrant crypt foci, ACF) and decreases the proportion of adenocarcinomas vs. adenomas in 1,2-dimethylhydrazine (DMH)-treated CF1 mice. To elucidate the structural specificity of this inhibition, the effects of the other major sphingolipids in milk (glycosphingolipids) were determined. Glucosylceramide (GluCer), lactosylceramide (LacCer) and ganglioside G(D3) were fed individually to DMH-treated (six doses of 30 mg/kg body weight) female CF1 mice at 0.025 or 0.1 g/100 g of the diet for 4 wk. All reduced the number of ACF by > 40% (P < 0.001), which is comparable to the reduction by SM in earlier studies. Immunohistochemical analysis of the colons revealed that sphingolipid feeding also reduced proliferation, with the most profound effect (up to 80%; P < 0.001) in the upper half of the crypts. Since the bioactive backbones of the glycosphingolipids (i.e., ceramide and other metabolites) are the likely mediators of these effects, the susceptibility of these complex sphingolipids to digestion in the colon was examined by incubating 500 microgram of each sphingolipid with colonic segments from mice and analysis of substrate disappearance and product formation by tandem mass spectrometry. All of the sphingolipids (including SM) disappeared over time with a substantial portion appearing as ceramide. Partially hydrolyzed intermediates (such as GluCer from LacCer or G(D3)) were not detected, which suggests that the cleavage involves colonic (or microflora) endoglycosidases. In summary, consumption of dairy SM and glycosphingolipids suppresses colonic cell proliferation and ACF formation in DMH-treated mice; hence, many categories of sphingolipids affect these key events in colon carcinogenesis.
 
Article
Measurement of micronutrient status in the presence of inflammation is difficult for several reasons. Changes in levels of acute phase proteins are associated with increased plasma levels of some indicators of micronutrient status, such as ferritin, and decrease of others, such as retinol. Alterations in the plasma levels of acute phase proteins can occur from hemodilution, sequestration and increased or decreased rates of synthesis and breakdown. How much these relate to functional deficiency is not known. Assays that are less perturbed by inflammation, such as the transferrin receptor assay, and adjustment of plasma micronutrient levels according to different cutoff levels for acute phase proteins are helpful but they do not enable precise assessment of micronutrient status among individuals who are infected. Improving assessment of micronutrient status is important if micronutrient interventions are to be targeted to those with the greatest need.
 
Article
The purpose of this study was to determine if high levels of dietary calcium could inhibit the induction of colon tumors in rats injected with a single dose of 1,2-dimethylhydrazine (DMH). Rats were given a single subcutaneous injection of DMH (200 mg/kg body weight) 2 wk before they were fed purified diets containing 5% fat and four different levels of calcium (as calcium gluconate). After 8 mo, the following incidences of colon tumors (total) were seen: 0.2% Ca, 56%; 0.5% Ca [National Academy of Sciences/National Research Council (NAS/NRC) recommended level], 75%; 1.0% Ca, 61%; 2.0% Ca, 41%. Thus, rats fed calcium at levels above or below the NAS/NRC recommendation had lower tumor incidences. The total tumor incidence and the incidence of adenocarcinomas (with or without invasion) were not significantly affected by calcium, but the incidences of benign adenomatous polyps and of distal colon tumors were significantly affected. Autoradiographic examination of [3H]thymidine-treated rats revealed that the level of calcium did not significantly alter the cell kinetic indices in the distal colon. In the proximal colon, however, the 0.2% Ca group had a significantly larger proliferative zone, with significantly more labeled cells present at the bottom of the colon crypt. Mineral analysis of tibias and serum samples revealed that rats fed higher levels of calcium had lower bone Fe and serum Mg contents, but no significant trends were seen for Ca, P, Zn or Cu. Therefore, increasing or decreasing the calcium content above or below the NAS/NRC recommendation (supplemented to low fat diets) during the promotional phase of colon carcinogenesis altered the tumor incidence, but the effect was confined to the distal colon and to benign adenomatous polyps.
 
Article
The hypothesis tested was that feeding rats sucrose rather than invert sugar (50:50 mixture of glucose and fructose) or cornstarch would result in a more rapid excretion of glucuronides and tritium from intravenously injected [1,2-3H]aldosterone. Thirty 56-d-old male rats of the Sprague-Dawley strain were fed for 8 wk one of three diets containing 45% of dietary energy from sucrose, invert sugar or cornstarch; 15% of energy was from protein and 40% from fat. Body weights and systolic blood pressures were measured weekly. After 60 d of feeding the diets ad libitum, all rats were injected intravenously with [1,2-3H]aldosterone and the percent recovery of tritium in both urine and feces was determined over the next 4 d. Urinary and fecal excretion of both free and conjugated glucuronic acid was determined over those 4 d. Urinary excretion of sodium and potassium (mg/d) was also determined. There were no differences between groups in food or water intakes, body weights, systolic blood pressures, daily fecal weights and daily urine volumes. The cornstarch-fed group excreted less sodium and potassium than did the other groups (P less than 0.05). The sucrose-fed group had a greater 4-d excretion of tritium (urinary + fecal) than did the invert sugar- or cornstarch-fed groups (P less than 0.01). The sucrose-fed group had a greater percentage of excreted glucuronic acid that was conjugated (urinary + fecal) than did the invert sugar- or cornstarch-fed groups (P less than 0.05). These results tended to confirm the hypothesis.
 
Plasma phosphorus as affected by dietary calcium (top) and vitamin D (bottom}. Values are means ± SEMfor 15 rats. Values at each time with differing super scripts are significantly different (P < 0.05). Calcium levels are (g/kg diet): low, 5; medium, 10; high, 15. Vitamin D levels are (mg/kg diet): low, 0.025; medium, 0.05; high, 0.1. 
Plasma 25-hydroxycholecalciferol as affected by dietary calcium (top) or vitamin D (bottom). Values are means ± SEMfor 15 rats. Values at each time with differing superscripts are significantly different (P < 0.05). Calcium levels are (g/kg diet): low, 5; medium, 10; high, 15. Vitamin D levels are (mg/kg diet): low, 0.025; medium, 0.05; high, 0.1. 
Article
We investigated whether increased levels of dietary calcium and vitamin D could inhibit colon carcinogenesis in rats injected with a single dose of 1,2-dimethylhydrazine. Rats were given a single subcutaneous injection (200 mg/kg body wt) 2 wk before they were fed purified diets containing 20% fat for 32 wk. Diets contained one of three levels of calcium (5, 10 or 15 g/kg diet) as calcium gluconate and one of three levels of vitamin D (0.025, 0.05 or 0.1 mg/kg diet) as cholecalciferol in a 3 x 3 factorial design. Rats receiving the highest level of vitamin D had greater plasma concentrations of 25-hydroxy-vitamin D. Autoradiographic examination of [3H]thymidine-treated rats demonstrated that a higher dietary level of calcium as well as higher levels of vitamin D significantly affected cellular kinetic indices. The total tumor incidence and tumor incidence in the distal colon was 45% lower in rats fed the highest level of both calcium and vitamin D compared with the other eight groups, although this decrease was not statistically significant (P = 0.12). The possible importance of these observations is discussed.
 
Article
Obesity is a state of chronic low-grade inflammation. Limiting white adipose tissue (WAT) expansion and therefore reducing inflammation could be effective in preventing the progression of obesity and the development of associated complications. We investigated the effects of 1,2-vinyldithiin (1,2-DT), a garlic-derived organosulfur, on the differentiation and inflammatory state of human preadipocytes. Preadipocytes were prepared from subcutaneous adipose tissue of nonobese young women and differentiated in the presence of 1,2-DT. Inflammatory preadipocytes were obtained following treatment with human macrophage-secreted factors. 1,2-DT (100 micromol/L) significantly reduced gene expression of PPARgamma2 (-40%), CCAAT/enhancer binding protein-alpha (-25%), lipoprotein lipase (-22%), leptin (-30%), and adiponectin (-15%). Lipid accumulation was also significantly diminished in preadipocytes differentiated in the presence of 100 micromol/L 1,2-DT (-37%) compared with controls. Furthermore, 100 micromol/L 1,2-DT treatment for 10 d significantly reduced PPARgamma activity (-27%). The protein expression of perilipin and the secretion levels for 2 adipokines, leptin and adiponectin, were significantly diminished in 1,2-DT-cultured preadipocytes (-37, -51, and -43%, respectively). Moreover, the secretion of inflammatory molecules (interleukin-6 and monocyte chemoattractant protein-1) induced by macrophage-secreted factors was partially abolished in 100 micromol/L 1,2-DT-treated preadipocytes (-28 and -25%, respectively). In conclusion, we demonstrated that 1,2-DT, a garlic-derived organosulfur, has antiadipogenic and antiinflammatory actions on human preadipocytes and may be a novel, antiobesity nutraceutical.
 
Composition of experimental diets Ingredient Casein BWP g/kg 
Article
This study was conducted to examine the effect of consumption of buckwheat protein product (BWP) on 1,2-dimethylhydrazine (DMH)-induced colon tumor in rats. Male growing Sprague-Dawley rats were fed diets containing either casein or BWP (net protein level, 200 g/kg; n = 20/group) for 124 d. The rats were gavaged weekly with DMH (20 mg/kg body) for the first 8 wk. Food intake and growth were unaffected by dietary manipulation. Dietary BWP caused a 47% reduction in the incidence of colonic adenocarcinoma (P < 0.05), but did not affect the incidence of colonic adenomas. BWP intake tended to reduce the number of colon adenocarcinomas (P = 0.16). Consumption of BWP significantly reduced cell proliferation and expression of c-myc and c-fos proteins in colonic epithelium. The results suggest that dietary BWP has a protective effect against DMH-induced colon carcinogenesis in rats by reducing cell proliferation.
 
Article
Dietary supplementation with milk sphingolipids inhibits colon tumorigenesis in CF1 mice treated with a colon carcinogen [1,2-dimethylhydrazine (DMH)] and in multiple intestinal neoplasia (Min) mice, which develop intestinal tumors spontaneously. Plant sphingolipids differ structurally from those of mammals [soy glucosylceramide (GlcCer) consists predominantly of a 4,8-sphingadiene backbone and alpha-hydroxy-palmitic acid], which might affect their bioactivity. Soy GlcCer was added to the AIN-76A diet (which contains <0.005% sphingolipid) to investigate whether it would also suppress tumorigenesis in these mouse models. Soy GlcCer reduced colonic cell proliferation in the upper half of the crypts in mice treated with DMH by 50 and 56% (P < 0.05) at 0.025 and 0.1% of the diet (wt/wt), respectively, and reduced the number of aberrant colonic crypt foci (an early marker of colon carcinogenesis) by 38 and 52% (P < 0.05). Min mice fed diets containing 0.025 and 0.1% (wt/wt) soy GlcCer developed 22 and 37% fewer adenomas (P < 0.05), respectively. The effects of dietary sphingolipids on gene expression in the intestinal mucosal cells of Min mice were analyzed using Affymetrix GeneChip microarrays. Soy GlcCer affected the expression of 96 genes by > or = 2-fold in a dose-dependent manner, increasing 32 and decreasing 64. Decreases in the mRNA expression of two transcription factors associated with cancer, hypoxia-induced factor 1 alpha (HIF1 alpha) and transcription factor 4 (TCF4), were confirmed by quantitative RT-PCR. In conclusion, soy GlcCer suppressed colon tumorigenesis in two mouse models; hence, plant sphingolipids warrant further investigation as inhibitors of colon cancer. Because soy contains relatively high amounts of GlcCer, sphingolipids may partially account for the anticancer benefits attributed to soy-based foods.
 
Article
Glycine plays several roles in human metabolism, e.g. as a 1-carbon donor, in purine synthesis, and as a component of glutathione. Glycine is decarboxylated via the glycine cleavage system (GCS) that yields concurrent generation of a 1-carbon unit as 5,10-methylenetetrahydrofolate (methyleneTHF). Serine hydroxymethyltransferase (SHMT) catalyzes the interconversion of glycine and serine, another 1-carbon donor. The quantitative role of glycine in human 1-carbon metabolism has received little attention. The aim of this protocol was to quantify whole body glycine flux, glycine to serine flux, and rate of glycine cleavage in humans. A primed, constant infusion with 9.26 micromol x kg(-1) x h(-1) [1,2-(13)C2]glycine and 1.87 micromol x kg(-1) x h(-1) [(2)H3]leucine was used to quantify the kinetic behavior of glycine in young, healthy volunteers (n = 5) in a fed state. The isotopic enrichment of infused tracers and metabolic products in plasma, as well as breath (13)CO2 enrichment, were determined for use in kinetic analysis. Serine synthesis by direct conversion from glycine via SHMT occurred at 193 +/- 28 micromol x kg(-1) x h(-1) (mean +/- SEM), which comprised 41% of the 463 +/- 55 micromol x kg(-1) x h(-1) total glycine flux. Nearly one-half (46%) of the glycine-to-serine conversion occurred using GCS-derived methyleneTHF 1-carbon units. Based on breath (13)CO2 measurement, glycine decarboxylation (190 +/- 41 micromol x kg(-1) x h(-1)) accounted for 39 +/- 6% of whole body glycine flux. This study is the first to our knowledge to quantify human glycine cleavage and glycine-to-serine SHMT kinetics. GCS is responsible for a substantial proportion of whole body glycine flux and constitutes a major route for the generation of 1-carbon units.
 
Article
Sphingolipids are in all eukaryotic cells and modulate cell growth, differentiation, and transformation; however, little is known about the physiological effects of their consumption. Mice were fed diets supplemented with milk sphingomyelin to determine effects on colon carcinogenesis. Cancer was initiated in CF1 mice by 1,2-dimethylhydrazine. Mice were then fed AIN76A diets supplemented with 0.025 to 0.1 g sphingomyelin/100 g for 28 wk until the supply of sphingomyelin was depleted and then fed unsupplemented diet for 24 wk. Sphingomyelin did not affect weight gain. Mice fed sphingomyelin had a 20% incidence of colon tumors compared with 47% in controls (P = 0.08 for all sphingomyelin-fed mice vs. controls). Tumors were adenomas or adenocarcinomas and located in the distal third of the colon. In shorter-term studies, colonic epithelial cell proliferation was significantly greater than controls in mice fed 0.025 g sphingomyelin/100 g diet, but not in those fed higher amounts of sphingomyelin. The number of aberrant crypts was significantly lower in 1,2-dimethylhydrazine-treated mice fed 0.05 g sphingomyelin/100 g diet than in controls. These results demonstrate that consumption of sphingomyelin affects the behavior of colonic cells. Because sphingolipids are present in food, the reduction in 1,2-dimethylhydrazine-induced premalignant lesions and the incidence of colon tumors in CF1 mice implies that these compounds may be another important class of nutritional modulators of carcinogenesis.
 
Article
The polyunsaturated fatty acid composition of dietary fat has been associated with increased oxidant stress in tissues. We previously showed that antioxidant mechanisms in colon mucosa are affected by dietary fat. In this study, the colonic antioxidant response to treatment with the colon-specific carcinogen 1,2-dimethylhydrazine (DMH) was determined. Sprague-Dawley rats were fed one of four AIN-76A-based diets with varied fat content. The basal diet contained 5% corn oil, the menhaden oil diet contained 19% menhaden oil and 1% corn oil, the corn oil diet contained 20% corn oil and the beef tallow diet contained 19% beef tallow and 1% corn oil. Animals were given one or two intraperitoneal injections of either DMH (20 mg/kg) or 1 mmol/L EDTA. Homogenates of colon mucosa were assayed to determine activity of catalase, glutathione peroxidase, total superoxide dismutase, manganese superoxide dismutase, glutathione reductase and glutathione-S-transferase, as well as total glutathione concentration, thiobarbituric acid-reacting substances and nuclear aberrations. Total glutathione concentration was greater in rats fed all diets following two injections of DMH, whereas glutathione reductase activity remained unchanged relative to controls. Lipid peroxidation was highest in the group fed menhaden oil; nuclear aberrations were greatest in the corn oil-fed group. In conclusion, both type and amount of dietary fat modified the response of colon tissue to genotoxic challenge, but none of the diets seemed more protective than the others.
 
Article
Recent evidence introduces the possibility that lutein and zeaxanthin may protect against the development of the two common eye diseases of aging, cataract and macular degeneration. This potential and the lack of other effective means to slow the progression of macular degeneration have fueled high public interest in the health benefits of lutein and zeaxanthin and the proliferation of supplements containing them on pharmacy shelves. An understanding of the biologic consequences of limiting or supplementing with these carotenoids is only beginning to emerge. Some epidemiologic evidence supports a role in eye disease and, to a lesser extent, cancer and cardiovascular disease. However, the overall body of evidence is insufficient to conclude that increasing levels of lutein and zeaxanthin, specifically, will confer an important health benefit. Future advances in scientific research are required to gain a better understanding of the biologic mechanisms of their possible role in preventing disease. Additional research is also required to understand the effect of their consumption, independent of other nutrients in fruits and vegetables, on human health. The newly advanced ability to measure levels of lutein and zeaxanthin in the retina in vivo creates a unique opportunity to contribute some of this needed evidence.
 
Article
The effects of two carcinogens benzo[a]pyrene (BP) and symmetrical 1,2-dimethylhydrazine dihydrochloride (DMH), on plasma amino acid concentrations and on excretion of lipids and nitrogenous metabolites were studied in 7- to 8-wk-old male and female B6C3F1 mice. BP and DMH were fed at concentrations of 0.3125 and 0.0225 g/kg, respectively, in purified diets containing 10 or 40% soybean protein. Nutritional balances were measured over a 7-d period after 7 d of acclimatization. Females excreted less urea and more NH3 than males. Urinary urea-nitrogen, NH3, allantoin, uric acid and total urinary nitrogen were consistently higher in mice fed 40% protein than in those fed 10% protein. The increases in total and NH3 nitrogen paralleled the increase in nitrogen intake. Nitrogen of urea rose more, while that of allantoin and uric acid rose less, than nitrogen intake. Fecal lipid excretion, as a percentage of intake, was consistently higher in mice fed the 40% protein diets than in mice fed 10% protein. Plasma glycine and branched-chain amino acids were higher, but citrulline was lower, when the 40% protein diet was fed. Body weight gain was higher when the 10% protein diet was fed with BP than without it, but BP made no apparent difference in weight gain when the 40% protein diet was fed. BP interacted with dietary protein to influence the excretion of nitrogenous metabolites. In addition, BP feeding produced numerous BP X sex and BP X protein interactions for plasma amino acid concentrations. Compared to controls, feed intake and weight gain were, respectively, 8 and 61% lower in DMH-fed animals during wk 1, but no differences in intake or weight gain were found during wk 2. In contrast to BP, DMH had no significant effects on urinary or fecal nitrogen metabolites, except that urinary uric acid (relative to nitrogen intake) was 9% higher in DMH-fed mice than in controls. DMH-fed mice had 43% higher serum glutamate and 6% lower glutamine than controls.
 
Article
The Grains for Health Foundation's Whole Grains Summit, held May 19-22, 2012 in Minneapolis, was the first meeting of its kind to convene >300 scientists, educators, food technologists, grain breeders, food manufacturers, marketers, health professionals, and regulators from around the world. Its goals were to identify potential avenues for collaborative efforts and formulate new approaches to whole-grains research and health communications that support global public health and business. This paper summarizes some of the challenges and opportunities that researchers and nutrition educators face in expanding the knowledge base on whole grains and health and in translating and disseminating that knowledge to consumers. The consensus of the summit was that effective, long-term, public-private partnerships are needed to reach across the globe and galvanize the whole-grains community to collaborate effectively in translating whole-grains science into strategies that increase the availability and affordability of more healthful, grain-based food products. A prerequisite of that is the need to build trust among diverse multidisciplinary professionals involved in the growing, producing, marketing, and regulating of whole-grain products and between the grain and public health communities.
 
Article
Weanling male Sprague-Dawley rats were used to determine whether the mechanism of the previously reported toxicity of 1,2-dimethylhydrazine (DMH) in selenium-deficient rats was related to a diminished capacity for detoxification of reactive oxygen species via glutathione peroxidase (GSH-Px) as well as by other known pathways of detoxification, including catalase (CAT), superoxide dismutase (SOD), glutathione-S-transferase (GST) and levels of glutathione (GSH). A 3 x 3 factorial experimental design was used to examine the acute effects of DMH treatment (0, 10 and 20 mg/kg body weight) interacting with dietary Se levels (less than 0.02, 0.1 and 0.5 mg/kg diet as sodium selenite). Animals were maintained on the test diets for 4 wk prior to challenge with DMH. Preliminary kinetics studies indicated the most appropriate time to examine antioxidant status was 3 h after DMH injection. At that time, livers and colons were analyzed for tissue levels of GSH-Px, CAT, SOD, GST and GSH. Data analysis demonstrated that Se deficiency impaired the ability of both liver and colon to mount an induced detoxification response to the acute oxidative stress generated by DMH challenge and may explain the toxicity of DMH in Se-deficient rats.
 
Article
Since propanediol is glucogenic and is extensively used in therapy of bovine ketosis, its metabolic fate was investigated. DL-1,2-propanediol-2-¹⁴C with 400 g of carrier propanediol was administered intraruminally to a lactating cow. During the next 24 hours, the percentage of the dose recovered was 43.7 in CO2, 12.4 in milk, 3.5 to 7 in urine, and less than 0.1 in feces. The propanediol was predominantly absorbed from the rumen without alteration, although some conversion to propionic acid in the rumen was detected. The maximal level of propanediol in milk was 0.04 mg/ml. Distribution of ¹⁴C among the carbons of lactose and glutamic acid indicated conversion of propanediol to glucose via carboxylation of pyruvate to oxalacetate. These results demonstrate that propanediol is glucogenic in the classical sense of that term; namely, that it is metabolized via intermediates, probably pyruvate and oxalacetate, which can lead to net synthesis of glucose. The small concentrations of propanediol in peripheral blood and the approximately 2-hour delay in attaining maximal specific activity in CO2 after attaining maximal specific activity of blood glucose indicated that glucogenesis was primarily hepatic with oxidation primarily occurring in other tissues.
 
Article
Garlic has been reported to have chemopreventive effects against a variety of cancers. However, different garlic preparations contain different constituents. We investigated the chemopreventive effect of aged garlic extract (AGE), an odorless product from prolonged extraction of fresh garlic, on colon carcinogenesis and cell proliferation in 1,2-dimethylhydrazine (DMH)-induced colon neoplastic rats. Rats were given weekly subcutaneous injections of DMH (20 mg/kg) for 20 wk, and fed either a basal diet or one containing 4% AGE. Serum from AGE-treated rats contained detectable S-allylcysteine. The AGE diet significantly reduced the number of colon tumors and aberrant crypt foci compared to the basal diet. Cell proliferation of normal-appearing colonic mucosa was assessed by MIB-5 immunohistochemistry. AGE treatment significantly decreased the mean MIB-5-labeling index. These findings suggest AGE has a chemopreventive effect on colon carcinogenesis through suppression of cell proliferation.
 
Article
C57BL/6J (B/6J) mice are genetically predisposed to become overweight and develop hyperglycemia if raised on a high fat diet. The purpose of the present study was to explore the effect of dietary supplementation of L-glutamine (Gln), an inhibitor of fatty acid oxidation, on the development of hyperglycemia and excessive weight gain. Groups of 10 age- and weight-matched male B/6J mice were raised on one of four diets: 1) a low fat, low sucrose (LL), studied separately, 2) a high fat, low sucrose (HL) diet alone, 3) high fat, low sucrose supplemented with L-glutamine (HL+Gln) and 4) high fat, low sucrose supplemented with L-alanine (HL+Ala). Energy intake, body weight, plasma glucose and insulin concentrations were monitored over time. We found no difference in energy intake per unit body weight between any groups after the first 2 wk of feeding. However, the mean +/- SEM for body weight (27.1 +/- 0.6 g) of the LL group measured at 16 wk was lower (P < 0.05) than that of the HL group at 37.9 +/- 1.9 g. Also, after 5.5 mo, the mean +/- SEM for plasma glucose and insulin concentrations in the LL group of mice were 6.9 +/- 0.4 mmol/l and 146 +/- 30 pmol/l, which were lower (P < 0.05) than those in the HL group at 10.1 +/- 0.9 mmol/l and 438 +/- 84 pmol/l, respectively. Although both amino acids caused a 10% reduction (P < 0.05) in body weight compared with HL feeding at wk 16, only Gln supplementation resulted in persistent reductions in both plasma glucose and insulin concentrations over 5.5 mo. In another experiment, when Gln was added to the high fat (HL) diet of heavy hyperglycemic animals for 2 mo, body weight gain, hyperglycemia and hyperinsulinemia were attenuated. In conclusion, supplementing glutamine to a high fat diet reduces body weight and attenuated hyperglycemia and hyperinsulinemia in B/6J mice.
 
Article
A number of model systems now exist for studying the nonnuclear actions of the seco-steroid hormone 1,25(OH)2D3. The perfused duodenal loop of vitamin D-replete chicks has provided the best correlation between nonnuclear actions and a physiological end point, namely enhanced calcium transport. Recent progress has been made in identifying and purifying an integral protein of the basal lateral membrane that may be a receptor for 1,25(OH)2D3. Studies with analogues (particularly 1,25(OH)2-7-dehydrocholesterol and 1,25(OH)2-lumisterol3) have provided definite correlations between binding to the solubilized membrane receptor and the ability to initiate transcaltachia (the rapid hormonal stimulation of calcium transport).
 
Duodenal calcium absorption and gene expression in 2-mo-old VDR KO and WT mice. ( A ) Ca absorption determined by in situ ligated loops (2 mmol/L Ca, 10-min test). ( B ) Calbindin D 9k mRNA. ( C ) TRPV6 mRNA levels. Calbindin D 9k and TRPV6 mRNA are ex- 
Effect of gender on the relation between duodenal gene expression and plasma 1,25(OH) 2 D levels in 3-mo-old mice fed diets with varying calcium concentrations. Mice were adapted to diets with different calcium concentrations (20, 5, or 0.2 g Ca/kg diet) for 7 d after which gene expression was studied (n 10 per gender and diet). TRPV6 and calbindin D 9k mRNA data were normalized for the expression of GAPDH within a sample and are expressed relative to the mean for male mice fed the high-calcium diet. (A) TRPV6 mRNA. Female: TRPV6 mRNA 0.042[plasma 1,25(OH) 2 D] 3.491, r 2 0.55; Male: TRPV6 mRNA 0.025[plasma 1,25(OH) 2 D] 1.799 r 2 0.51. (B) Calbindin D 9k mRNA. Female: calbindin D 9k 9.21 10 3 [plasma 1,25(OH) 2 D] 0.778, r 2 0.5; Male: calbindin D 9k 6.67 10 3 [plasma 1,25(OH) 2 D] 1.435, r 2 0.48.
Article
Recent advances in bone and calcium (Ca) metabolism have relied upon genetically modified mice. However, although human studies have identified gender as an important modulator of Ca metabolism, its effect on Ca metabolism has not been examined in mice. Here we examined basal and vitamin D-regulated Ca absorption (in situ ligated loops) and mRNA levels for the apical membrane calcium channel, TRPV6, and the calcium binding protein, calbindin D(9k) (CaBP) mRNA levels (real-time PCR) in duodenum of female and male mice. At 2 mo of age, females fed a 5 g Ca/kg diet had higher Ca absorption (62.3 +/- 4.8 vs. 47 +/- 3.6%) and TRPV6 mRNA levels than males even though plasma 1,25 dihydroxyvitamin D [1,25(OH)(2) D] was not different. In mice fed high (20 g/kg), normal (5 g/kg), or low (0.2 g/kg) Ca diets for 7 d to alter plasma 1,25(OH)(2) D (91 +/- 12, 322 +/- 25, and 587 +/- 43 pmol/L, respectively), the relation between Ca absorption (slope = 0.116 vs. 0.084, P = 0.021) or duodenum TRPV6 mRNA (slope = 0.042 vs. 0.025, P = 0.034) and circulating 1,25(OH)(2) D was steeper in females. After a single 1,25(OH)(2) D injection (200 ng/100 g body weight), peak induction of TRPV6 mRNA was 2-fold greater (at 6 h) and CaBP mRNA was 20% higher in females (at 16 h). Duodenal vitamin D receptor mRNA levels did not differ between genders. Our data indicate that female mice are more sensitive to changes in serum 1,25(OH)(2) D levels than males and that this must be considered when using mice to study calcium and bone biology.
 
Article
1,25-Dihydroxycholecalciferol [1,25-(OH)2D3] has been shown to inhibit the progression of experimental autoimmune encephalomyelitis (EAE). Here we tested the possibility that 1, 25-dihydroxycholecalciferol might be therapeutic for another autoimmune disease, arthritis. Two different animal models of arthritis were tested, namely, murine Lyme arthritis and collagen-induced arthritis. Infection of mice with Borrelia burgdorferi (the causative agent of human Lyme arthritis) produced acute arthritic lesions including footpad and ankle swelling. Supplementation with 1,25-dihydroxycholecalciferol of an adequate diet fed to mice infected with B. burgdorferi minimized or prevented these symptoms. Mice immunized with type II collagen also developed arthritis. The symptoms of this disease were also prevented by dietary supplementation with 1,25-dihydroxycholecalciferol. 1, 25-Dihydroxycholecalciferol given to mice with early symptoms of collagen-induced arthritis prevented the progression to severe arthritis compared with untreated controls. These results suggest that 1,25-dihydroxycholecalciferol and/or its analogs may be a valuable treatment approach to this disease.
 
Article
These studies were conducted to determine if supplementation of a corn-soybean meal diet with 1,25-dihydroxycholecalciferol [1,25-(OH)2D3] would increase the utilization of natural phytate phosphorus by broiler chickens. Two experiments were conducted to evaluate the effect of dietary 1,25-(OH)2D3 in the presence and absence of supplemental phytase and at several dietary levels of inorganic phosphorus supplementation. The criteria measured in these studies were weight gain, gain:feed ratio, bone ash, rickets due to phosphorus deficiency, plasma calcium and phosphorus and retention of calcium, phosphorus and phytate phosphorus. In the first experiment, the types and amounts of fecal inositol phosphates were determined by HPLC, and the total fecal phytate was determined by the classic FeCl3 precipitation technique. In the first experiment, the addition of 1,25-(OH)2D3 to the diet in the presence of dietary phytase resulted in greater 9-d weight and bone ash and lower incidence of rickets; the retention of total fecal phytate and phytate phosphorus was greater than in controls. The second experiment was a complete 2 x 2 x 2 factorial design [phosphorus levels x phytase x 1,25-(OH)2D3]. The addition of 1,25-(OH)2D3 alone to the diet resulted in greater 9-d weight and bone ash, lower incidence of rickets, and greater retention of total calcium and phosphorus and phytate phosphorus. The highest retention of phytate phosphorus (79.4%) was obtained when both phytase and 1,25-(OH)2D3 were present in the diet. The possible mode of action and importance of these results in many areas of nutrition and environmental science are discussed.
 
Article
1,25-Dihydroxyvitamin D3 [1,25-(OH)2-D3] is known to be an immunosuppressive hormone. This review primarily deals with in vitro and in vivo effects of 1,25-(OH)2-D3 and analogue, 1,25-dihydroxy-16ene-vitamin D3 [1,25-(OH)2-16ene-D3], on T helper subsets type 1 (Th1) or type 2 (Th2) that have distinctive functional characteristics in humans. Th1 secrete interferon (IFN-gamma), interleukin (IL-2) and induce B cells to produce immunoglobulin IgG2a while Th2 secrete IL-4, IL-10 and induce the production of IgG1 and IgE by B cells. The sterol inhibits the secretion of IL-12, a cytokine produced by monocytes and B cells, which leads to the activation and differentiation of Th1. In addition, 1,25-(OH)2-D3 directly inhibits IFN-gamma secretion by Th1 clones while it has little effect on IL-4 secretion by Th2 clones. The analogue, 1,25-(OH)2-16ene-D3, is 100-fold more potent than 1,25-(OH)2-D3 in inhibiting IFN-gamma secretion but also has little effect on IL-4 secretion. In mice, when given in vivo, the sterol prevents the induction of spontaneous and induced autoimmune diseases and inhibits Th1 induce IgG2a responses. These actions of the vitamin D3 compounds suggest that it may have potential therapeutic applications in Th1-mediated clinical situations such as autoimmunity and transplantation.
 
Article
1,25-Dihydroxycholecalciferol, the apparent active form of vitamin D3 (cholecalciferol) which mediates calcium translocation in bone and intestine, has been tested for its nutritional efficacy in preventing rickets. Chicks were fed a vitamin D-deficient diet and received oral supplements of 1,25-dihydroxycholecalciferol, 25-hydroxycholecalciferol or cholecalciferol for 3 weeks. Growth, plasma calcium concentration, calcium absorption and percentage bone ash were determined; sections of tibia were examined microscopically for evidence of rickets, 1,25-Dihydroxycholecalciferol displayed an effectiveness similar to 25-hydroxycholecalciferol, with both metabolities being between 1.5 and 2.2 times as active as cholecalciferol with respect to stimulation of weight gain and maintenance of plasma calcium levels. The antirachitic potency of 1,25-dihydroxycholecalciferol in chicks is estimated to be 1.3 times that of cholecalciferol. Thus, 1,25-dihydroxycholecalciferol was found to be more potent than the parent sterol in terms of supporting normal calcium and bone metabolism, and the metabolite alleviated all signs of rickets. These data substantiate the conclusion that 1,25-dihydroxycholecalciferol is the hormonal form of vitamin D and indicate that other metabolites of the vitamin are not required for the regulation of calcium metabolism and prevention of bone disease.
 
Article
The influence of triiodothyronine (T3) on the induction of intestinal calcium and inorganic phosphate (Pi) transport by 1,25-dihydroxycholecalciferol (1,25-(OH)2D3) was studied in 48 h cultures of embryonic chick jejunum. While T3 alone had no effect on calcium uptake by gut segments cultured on d 20 of embryonic development, the thyroid hormone amplified the effect of 1,25-(OH)2D3 on calcium transport and effectively shifted the dose-response curve to lower 1,25-(OH)2D3 concentrations. T3 had a dual effect on Pi uptake by cultured jejunum: It induced transport activity even in the absence of the steroid hormone, and, in addition, synergistically raised 1,25-(OH)2D3-related Pi uptake. In d 17 embryonic small intestine, which does not respond to 1,25-(OH)2D3 by a significant increase in Pi transport, T3 permitted the induction of Pi transport by the sterol. In general, the thyroid hormone enhanced the responsiveness of cultured embryonic intestine toward 1,25-(OH)2D3 by two orders of magnitude, resulting in facilitated induction of calcium and Pi transport by the sterol and, in particular, modulated the stage-specific expression of 1,25-(OH)2D3 action on intestinal Pi transport.
 
Article
Acute regulation of the vitamin D receptor in kidney by 1,25-dihydroxycholecalciferol and dietary calcium was investigated using vitamin D-deficient rats. 1,25-Dihydroxycholecalciferol administered to normocalcemic, vitamin D-deficient rats increased the renal receptor level, whereas serum calcium and phosphorus concentrations remained nearly constant. In hypocalcemic, vitamin D-deficient rats, 1,25-dihydroxycholecalciferol caused a sharp response at 4 h, which remained elevated for the remaining 20 h. Serum calcium rose gradually over 24 h, whereas serum phosphorus remained fairly constant. When hypocalcemic, vitamin D-deficient rats were fed a high calcium diet following 14 h without food, the serum calcium concentration increased and serum phosphorus concentration decreased, whereas renal receptor levels were unchanged. These data indicate that 1,25-dihydroxycholecalciferol rapidly regulates the renal vitamin D receptor, independent of serum calcium and phosphorus, but requires an adequate serum calcium concentration to sustain a prolonged effect.
 
Relationship between kidney tissue ratio of vitamin D receptor (VDR) mRNA/GAPDH mRNA, and serum 1,25-dihydroxycholecalciferol [1,25(OH) 2 D] in rats fed Purina nonpurified diet without or with additional vitamin D. The regression line is shown, along with its 95% confidence limits. The data show a significant negative relationship (r 0.714, P 0.006), with rats given a moderately higher vitamin D intake having lower 1,25(OH) 2 D and higher renal VDR mRNA by Northern blot, relative to expression of the GAPDH housekeeping gene. 
Effect of changes in diet calcium on urine pyridinoline/ creatinine ratio in groups of rats maintained on the indicated daily vitamin D intakes. Since there were no differences among groups, representative SEM are indicated, based on only the first group of rats.
Dynamic responses of urine calcium/creatinine ratio and serum 1,25-dihydroxycholecalciferol [1,25(OH) 2 D] to changes in dietary calcium content. Up to d 0, the diet contained 0.5 g of elemental calcium/ 100-g diet. Starting on d 0 the amount of calcium in the diet fed the rats was adjusted every 7 d to the amounts shown by the numbers across the top of the figure, in units of g/100-g diet. Groups of rats differed according to the amount and route of vitamin D nutrition, as indicated in the figures. The error whiskers indicate SEM The graphs show that urine calcium excretion paralleled diet calcium content, and that on the log scales of the bottom panels the serum 1,25(OH) 2 D levels of groups given 20 nmol/d of vitamin D were shifted lower by a constant amount at all calcium intakes, which indicates greater responsiveness to circulating 1,25(OH) 2 D. 
Article
We examined how cholecalciferol (vitamin D) nutrition affected serum 25-hydroxycholecalciferol (25(OH)D) and 1, 25-dihydroxycholecalciferol (1,25(OH)(2)D). Rats were fed conventional diet (vitamin D, 4.5 IU/g, or 7 nmol/d) or the same diet plus 18 nmol/d of extra vitamin D for 3 wk. The extra vitamin D resulted in greater serum 25(OH)D (51 +/- 3, vs. control of 21 +/- 2 nmol/L), and kidney mRNA for vitamin D receptor [VDR mRNA] (P = 0. 026) and lower serum 1,25(OH)(2)D (72 +/- 16 vs. control of 161 +/- 10 pmol/L, P = 0.001), and parathyroid hormone (PTH) (89 +/- 4 vs. control of 160 +/- 15 ng/L, P = 0.001). Kidney VDR mRNA relative to GAPDH mRNA correlated inversely with serum 1,25(OH)(2)D (r = -0.714, P = 0.006). There were no differences in serum calcium, phosphate, alkaline phosphatase, or weight gain. Experiment 2 compared groups supplemented with 0.2, 2 or 20 nmol/d of vitamin D orally, or 20 nmol/d dermally to see how vitamin D nutrition influenced the response of 1,25(OH)(2)D to changes in diet calcium. Vitamin D did not affect urinary calcium or pyridinoline excretion, serum calcium, phosphate, vitamin D binding protein or alkaline phosphatase. In groups given 20 nmol/d of vitamin D, renal mitochondrial 25(OH)D-1alpha-hydroxylase was lower (P < 0.01) and 25(OH)D-24-hydroxylase was higher (P < 0.05). Higher 25(OH)D concentration was related to proportionally lower 1,25(OH)(2)D at every calcium intake, indicating greater tissue sensitivity to 1, 25(OH)(2)D. We conclude suppression of 1,25(OH)(2)D and PTH, and higher renal VDR mRNA and 24-hydroxylase did not involve higher free 1,25(OH)(2)D concentration or a first pass effect at the gut. Thus, 25(OH)D or a metabolite other than 1,25(OH)(2)D is a physiological, transcriptionally and biochemically active, noncalcemic vitamin D metabolite.
 
Article
The effect of cholecalciferol on the intestinal absorption of 65Zn was assessed in zinc-deficient and zinc-replete rachitic chicks, using the in situ ligated loop techniques. Cholecalciferol did not significantly affect 65Zn absorption in either group, although the synthesis of the intestinal calcium-binding protein (CaBP) in both groups was similar. In an analogous study, 1,25-dihydroxycholecalciferol increased 47Ca absorption and induced the synthesis of CaBP but exerted on effect on 65Zn absorption in zinc-deficient rachitic chicks. When fed a diet adequate in cholecalciferol, more CaBP was present in the intestine of the zinc-adequate group than in the zinc-deficient group, possibly due to the greater rate of growth and therefore the greater need for calcium by the former group. These results suggest that cholecalciferol and its most active metabolite do not directly affect zinc absorption and, by inference, that the vitamin D-dependent transport mechanism is not involved in zinc homeostasis, or in the interaction between calcium and zinc.
 
First evaluation of the biological response in the chick to daily administration of 1£5-dihydroxycholecalciferol 
Article
The biological activity of the vitamin D2 metabolite 1,25-dihydroxycholecalciferol was assessed in three experiments in the chick and two experiments in the rachitic rat. In the official rat line test 1,25-dihydroxycholecalciferol was found to be equally as active as an equivalent dose of cholecalciferol. Doses of 20 to 250 pmoles (0.01 to 0.10 µg) of 1,25-dihydroxy-[26,27-²H] cholecalciferol or standard amounts of cholecalciferol were fed daily to White Leghorn cockerels for 3 weeks after hatching. The following vitamin D-related responses were measured: a) the rate of growth; b) intestinal transport of test doses of ⁴⁵Ca²⁺ in vivo and c) in vitro; d) percentage bone ash; e) serum calcium elevation. For each of these assays the 1,25-dihydroxycholecalciferol was judged to be “much more active” a) “more active” (b,d), and “as active” (c,e), as the parent cholecalciferol, where “much more active” is a response 2 to 4 times and “more active” is a response 1 to 2 times that obtained with an equivalent dose of cholecalciferol. In some instances (d,e), the relative increment of response mediated by 1,25-dihydroxycholecalciferol was greater at the low dose level (20 pmoles/day) than at a higher dose level (100 pmoles/day). The relative activity determination of 1,25-dihydroxycholecalciferol, obtained after daily administration, in comparison with cholecalciferol was markedly less than that previously obtained when the activity of single doses of these steroids were analyzed. A single dose of 1,25-dihydroxycholecalciferol was 4.4 ± 1.1 (se) [Science 171: 79 (1971)] times as effective as cholecalciferol in stimulating the intestinal absorption of Ca²⁺, and 5.5 ± 1.4 (se) [J. Biol. Chem. 247: 5728 (1972)] times as effective as cholecalciferol in stimulating serum Ca²⁺ elevation. These results collectively support the concept that 1,25-dihydroxycholecalciferol is the most highly potent form of Vitamin D3 yet known.
 
Article
The purpose of this study was to determine the effect of food restriction on age-related changes in serum 1,25-dihydroxyvitamin D and PTH, two important regulators of Ca metabolism. Starting at 6 wk, male F344 rats were fed a purified diet either ad libitum (non-restricted) or 60% of ad libitum (restricted). Rats from each group were killed at 5, 13, 22 and 28 mo of age. Dietary restriction increased the median lifespan from 24 to 31 mo. It delayed the rapid decrease in serum 1,25-dihydroxyvitamin D from 1.5-5.0 mo in the non-restricted group to 5-13 mo in the restricted group. It also completely suppressed the marked rise in serum PTH which occurred at 22 and 28 mo in the non-restricted group. Dietary restriction had these effects even though both groups of animals consumed the same amount of Ca per gram body weight. Diet had no effect on serum Ca and P, except at 28 mo. These effects of dietary restriction on serum 1,25-dihydroxyvitamin D and PTH may result in altered Ca metabolism in dietary restricted F344 rats.
 
NFB transcriptional activity in C3H10T1/2 murine fibroblast cells treated with 1,25(OH) 2 D 3 . C3H10T1/2 cells cotransfected with the NFB luciferase and Renilla luciferase plasmids were treated for indicated time points and luciferase activity was assayed and normalized for transfection efficiency via Renilla luciferase activity. Data are expressed as relative luciferase units as a ratio of treated samples to vehicle controls. Value represents the means of 4 independent experiments SE. Asterisks indicate a difference from the vehicle control: *P 0.05, **P 0.001.  
PI3K involvement in 1,25(OH) 2 D 3 regulation of NFB in C3H10T1/2 murine fibroblast cells. (A) A representative Western blot showing AKT activity at indicated time points after 1,25(OH) 2 D 3 treatment . (B) Histogram representative of the quantification of AKT activity. Data are expressed as a ratio of treated samples to vehicle controls. An asterisk indicates significant (P 0.05) difference compared with time zero. C3H10T1/2 cells were treated with PI3K inhibitors either LY294002 (C) or wortmannin (D). Luciferase activity was assayed after 4 h and was normalized for transfection efficiency via Renilla luciferase activity. Data are expressed as relative luciferase units as a ratio of treated samples to vehicle controls. Values represent the means of 3 independent experiments SE. An asterisk indicates a difference from the vehicle control: *P 0.05.  
Article
1,25-dihydroxycholecalciferol [1,25(OH)(2)D(3)] is important in the regulation of cell growth, differentiation, and apoptosis. Previous results from our laboratory demonstrate that 1,25(OH)(2)D(3) inhibits vitamin E succinate (VES) mediated apoptosis in untransformed C3H10T1/2 mouse fibroblast cells. The current work investigated cell survival signaling pathways that may be activated by 1,25(OH)(2)D(3), leading to protection from apoptosis. Results showed that nuclear factor kappaB (NFkappaB) transcriptional activity was significantly increased 1.8-fold over vehicle controls by 1,25(OH)(2)D(3) after 4 h of treatment. Protein kinase B/AKT, a downstream effector of phosphoinositide 3-kinase (PI3K), was activated 4-fold and 8-fold at 2 and 4 h, respectively, after treatment with 1,25(OH)(2)D(3). Pretreatment with two PI3K inhibitors, LY294002 and wortmannin, abolished the activation of NFkappaB by 1,25(OH)(2)D(3), suggesting that this pathway is essential for NFkappaB transcriptional activation. Additionally, the use of a p-21 activated kinase (PAK1) inhibitory construct (PAK(R299)) demonstrated that PAK1 was also required for NFkappaB transcriptional activation by 1,25(OH)(2)D(3). Inhibition of NFkappaB activity with transfection of the NFkappaB inhibitory construct (IkappaB(Ala32)) abolished the protective effect of 1,25(OH)(2)D(3) on VES-mediated apoptosis. In summary, NFkappaB transcriptional activation was essential to 1,25(OH)(2)D(3) protection from VES-mediated apoptosis and 1,25(OH)(2)D(3) regulated NFkappaB activity through PI3K and PAK pathways.
 
Article
Vitamin D is a conditionally required nutrient traditionally thought to influence physiology as the metabolite 1,25-dihydroxyvitamin D [1,25(OH)(2) D] by binding to the vitamin D receptor (VDR) and stimulating the transcription of genes through direct VDR-DNA interactions. However, over the past 15 y research has demonstrated that 1,25(OH)(2) D, as well as other steroid hormones, can rapidly stimulate ion fluxes and activate protein kinases by transcription-independent mechanisms. This review summarizes recent research on the rapid actions of 1,25(OH)(2) D and identifies questions that remain to be answered in this area.
 
Article
Anecdotal data suggest that the amount of vitamin D available in the environment either from sunshine exposure or diet may be an important factor affecting the development of inflammatory bowel disease (IBD) in humans. We tested the vitamin D hypothesis in an experimental animal model of IBD. Interleukin (IL)-10 knockout (KO) mice, which spontaneously develop symptoms resembling human IBD, were made vitamin D deficient, vitamin D sufficient or supplemented with active vitamin D (1,25-dihydroxycholecalciferol). Vitamin D-deficient IL-10 KO mice rapidly developed diarrhea and a wasting disease, which induced mortality. In contrast, vitamin D-sufficient IL-10 KO mice did not develop diarrhea, waste or die. Supplementation with 50 IU of cholecalciferol (5.0 microgram/d) or 1, 25-dihydroxycholecalciferol (0.005 microgram/d) significantly (P < 0. 05) ameliorated symptoms of IBD in IL-10 KO mice. 1, 25-Dihydroxycholecalciferol treatment (0.2 microgram/d) for as little as 2 wk blocked the progression and ameliorated (P < 0.05) symptoms in IL-10 KO mice with already established IBD.
 
Article
A possible role of vitamin D in the growth and development of rats was investigated. Impaired development was observed in normocalcemic, vitamin D-deficient male and female rats, as revealed by low intestinal calcium transport, low renal vitamin D receptor levels and poor bone mineralization. Analogs of 1,25-dihydroxycholecalciferol, possessing reduced calcium-mobilizing activity in intestine and bone but retaining differentiation activity in cultured cells, were unable to support normal development of normocalcemic, vitamin D-deficient male rats. These results suggest that either the calcium-mobilizing activity alone or both the calcium-mobilizing activity and differentiating activity of vitamin D are required for normal development or that the analogs are inactive in vivo. We also demonstrated sex-related differences in intestinal calcium transport, renal vitamin D receptor regulation and bone mineralization that were independent of vitamin D status.
 
Article
The adaptive increase in renal proximal tubule 25-hydroxyvitamin D-alpha-hydroxylase activity (1-OHase) during dietary calcium restriction is mediated by an increase in parathyroid hormone (PTH) and is inhibited by aging. Recent studies in mature (3-4 mo) rats demonstrated that insulin-like growth factor-I (IGF-I) restored stimulation of renal 1,25-dihydroxycholecalciferol [1,25(OH)(2)D(3)] production by low phosphorus diet (LPD), another major stimulus of 1-OHase. These studies were designed to determine whether IGF-I stimulates 1-OHase during low calcium intake in old rats. Male rats were fed a normal calcium diet (NCD, 6 g Ca/kg diet) or low calcium diet (LCD, 0.2 g Ca/kg diet) for 14 d, and recombinant human IGF-I [rhIGF-I, 1.4 mg/(24h 160 kg body wt)] or vehicle was administrated via miniosmotic pump for 72 h before killing. In 4-mo-old male Sprague-Dawley rats, LCD increased in vitro renal 1-OHase activity in the presence but not in the absence of rhIGF-I. LCD increased in vitro1-OHase activity in young (1-mo-old) but not old (24-mo-old) male Fischer 344 rats. RhIGF-I increased 1-OHase activity in 24 mo-old rats fed LCD to levels that were not different from those in 1-mo-old rats fed LCD. The results indicate that the adaptive increase in 1-OHase activity due to a LCD is lost by 4 mo in rats and can be restored by pharmacologic doses of rhIGF-I.
 
Article
Twenty-one-day-old rats placed on a vitamin D-deficient diet showed no decrease in serum, 1,25-dihydroxy vitamin D levels after 13 days on this diet. Between 13 and 20 days on this D-deficient diet there was a 50% decrease in serum 1,25-dihydroxy vitamin D. After 34 days, the level of 1,25-dihydroxy vitamin D in serum had dropped to near zero. With a vitamin D-deficient diet lacking calcium, there was an apparent stimulation of 1-hydroxylase, resulting in higher 1,25-dihydroxy vitamin D serum levels after 6--20 days on the diet. After 27 days there were no differences in 1,25-dihydroxy-vitamin D levels in animals fed a calcium-replete or calcium-deficient rachitogenic diet. When 1-day-old chickens were maintained on a rachitogenic diet for 1 week, 1,25-dihydroxy vitamin D levels were higher in animals fed the calcium-deficient diet compared with the calcium-replete diet. After 2 weeks on either rachitogenic diet, 1,25-dihydroxy vitamin D levels decreased to near zero. Measurement of 1,25-dihydroxy vitamin D levels has provided a biochemical indicator of vitamin D deficiency in chicks and rats which will complement other established biological criteria for vitamin D deficiency.
 
Top-cited authors
Guoyao Wu
  • China Agricultural University
Johanna T Dwyer
  • Tufts Medical Center
Sharon I Kirkpatrick
  • University of Waterloo
Regan L Bailey
  • Purdue University
Patricia M. Guenther
  • University of Utah