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1.
The effect of vitamin D3 metabolites on membrane proteins of chick duodenal brush borders. 总被引:1,自引:0,他引:1
Chick intestinal brush border proteins were examined by polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulfate. Following injection of 25-hydroxyvitamin D3 and 1,25-dihydroxyvitamin D3, a large molecular weight protein present in the vitamin D-deficient brush borders diminishes and a larger protein appears. This change occurs before calcium binding protein can be detected by Chelex assay and prior to the increase in total alkaline phosphatase but correlates closely with increased intestinal calcium absorption in response to the metabolites. The two brush border proteins have been solubilized with n-butanol and partially characterized. The vitamin D-deficient protein has a molecular weight of about 200,000 and has alkaline phosphatase activity but no detectable calcium binding activity. The protein which appears in response to metabolites has a molecular weight of 230,000, binds calcium, and also has alkaline phosphatase activity. 相似文献
2.
A crude aqueous extract of the leaves of when administered orally to vitamin D-deficient chicks produced significant increases in plasma phosphate but had little effect on plasma calcium. When chicks, fed a high strontium diet to inhibit endogenous 1,25(OH)2 vitamin D3 production and intestinal calcium transport, were dosed with the extract or synthetic 1,25(OH)2D345Ca absorption from the duodenum was stimulated, whereas vitamin D3 was ineffective. Partial purification of the crude extract on a Sephadex GH25 column yielded two factors, one of which mimicked 1,25 (OH)2D3 activity in chicks fed the high strontium diet while the other produced a significant increase in plasma phosphate. The presence of these substances, together with previously demonstrated organic solvent soluble vitamin D-type activity, may account for the calcinogenic nature of the plant. 相似文献
3.
Viviana Centeno Gabriela Picotto Adriana Pérez Arturo Alisio Nori Tolosa de Talamoni 《Archives of biochemistry and biophysics》2011,(2):191
The role of 1,25(OH)2D3 on the intestinal NCX activity was studied in vitamin D-deficient chicks (-D) as well as the hormone effect on NCX1 protein and gene expression and the potential molecular mechanisms underlying the responses. Normal, -D and -D chicks treated with cholecalciferol or 1,25(OH)2D3 were employed. In some experiments, -D chicks were injected with cycloheximide or with cycloheximide and 1,25(OH)2D3 simultaneously. NCX activity was decreased by -D diet, returning to normal values after 50 IU daily of cholecalciferol/10 days or a dose of 1 μg calcitriol/kg of b.w. for 15 h. Cycloheximide blocked NCX activity enhancement produced by 1,25(OH)2D3. NCX1 protein and gene expression were diminished by -D diet and enhanced by 1,25(OH)2D3. Vitamin D receptor expression was decreased by -D diet, effect that disappeared after 1,25(OH)2D3 treatment. Rapid effects of 1,25(OH)2D3 on intestinal NCX activity were also demonstrated. The abolition of the rapid effects through addition of Rp-cAMPS and staurosporine suggests that non genomic effects of 1,25(OH)2D3 on NCX activity are mediated by activation of PKA and PKC pathways. In conclusion, 1,25(OH)2D3 enhances the intestinal NCX activity in -D chicks through genomic and non genomic mechanisms. 相似文献
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5.
M Bachelet A Ulmann B Lacour 《Biochemical and biophysical research communications》1979,89(2):694-700
Alkaline phosphatase activity (APA) stimulation in response to 1,25-dihydroxycholecalciferol (1,25 (OH)2D3) has been studied in vitamin-D-deficient rat intestinal brush borders prepared from --perfused duodeno-jejunal segments. Basal APA in intestines perfused with ethanol remained constant throughout the experiments. APA was significantly increased when intestines were perfused with 1,25 (OH)2D3 (3 nM) for 30, 45 or 60 min. A dose-effect response was observed when 1,25 (OH)2D3 increased in the perfusion medium. The maximal alkaline phosphatase activity after a 45-min perfusion (2404 ± 379 mTU/mg prot.) was observed when 1,25 (OH)2D3 concentration was 6 nM. Cholecalciferol had no effect in this system. 相似文献
6.
C. Tony Liang Janice Barnes Richard A. Balakir Bertram Sacktor 《The Journal of membrane biology》1986,90(2):145-156
Summary Thein vivo andin vitro effects of 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) on calcium uptake by isolated chick duodenal cells were studied.In vivo, 1,25-(OH)2D3 given orally to vitamin D-deficient chicks increased the initial rate of calcium uptake by cells prepared 1 hr after administration of the hormone. The rate was stimulated approximately 100%, 17 to 24 hr after repletion.In vitro, pre-incubation of 1,25-(OH)2D3 with cells from D-deficient chicks increased the cellular rate of calcium uptake in a concentration-dependent relationship. Enhancement was found with 10–15
m, was maximal at 10–13
m, and was diminished at higher (10–11
m) concentrations. Stimulation was observed after a pre-incubation period as brief as 1 hr. The potency order for vitamin D3 analogs was 1,25-(OH)2D3=1-(OH)D3>25-(OH)D3>1,24,25-(OH)3D3>24,25-(OH)2D3>D3. The maximal enhancement in calcium uptake induced by the analogs was the same, only the concentration at which the cell responded was different. The effectiveness of 1,25-(OH)2D3 was five orders of magnitude greater than D3. Kinetically, 1,25-(OH)2D3 increased theV
max of calcium uptake; the affinity for calcium (K
m=0.54mm) was unchanged. The enhanced uptake found after the cells were pre-incubated for 2 hr with the hormone was completely blocked by inhibitors of protein synthesis. 1,25-(OH)2D3,in vitro, also increased calcium uptake in cells isolated from D-replete chicks. The maximal rates of uptake were the same in cells from D-deficient and D-replete animals. The hormone had no effect of calcium efflux from cells. Calcium uptake in microvillar brush-border membrane vesicles was increased by 1,25-(OH)2D3. These findings suggest that thein vitro cell system described in this paper represents an appropriate model to examine the temporal relationships between 1,25-(OH)2D3 induction of calcium transport and specific biochemical correlates. 相似文献
7.
Vitamin D-deficient chicks were injected intracardially with physiological doses of 1,25-dihydroxycholecalciferol (1,25-(OH)2D3) and the formation of intestinal brush-border proteins was followed in vitro. Within 4 h of receiving the hormone the incorporation of radioactive leucine into at least two proteins in the brush-borders was increased. The apparent molecular weights of these proteins were 45 000 and 84 000. The change in the synthesis of these proteins was followed with time and compared with the concomitant changes in intestinal calcium transport. The relationship of these changes is such that there is a strong possibility that the proteins are involved in calcium absorption. 相似文献
8.
Wayne R. Wecksler Helen L. Henry Anthony W. Norman 《Archives of biochemistry and biophysics》1977,183(1):168-175
As a further means of evaluating 1,25-dihydroxyvitamin D3-parathyroid gland interaction and its relation to calcium homeostasis, a comparative study of the subcellular localization of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3]in the parathyroid glands, intestinal mucosa, kidney, and liver of rachitic chickens has been carried out. Only in the chromatin fraction from parathyroids and intestinal mucosa could there be demonstrated selective and specific localization of the 1,25(OH)2D3. The chromatin-bound picomoles of 1,25(OH)2D3 (per gram of tissue) was in the ratio (mucosa:parathyroids:kidney:liver) of 1.0:0.23:0.11:0.17 2 h after an intracardial injection of 290 pmol of [3H]1,25(OH)2D3. This same ratio after a 30-min (23 °C) homogenate incubation with 1 × 10?8m [3H]1,25(OH)2D3 was 1.0:1.0:0.10:0.03. Analogous results were obtained when reconstituted chromatin and cytosol fractions from the different tissues were compared for chromatin localization efficiency. This chromatin localization of 1,25(OH)2D3 in the parathyroid glands was temperature dependent. In addition, parathyroid glands were found to contain 3.0–3.5 S cytoplasmic and KCl-extractable chromatin receptors specific for 1,25(OH)2D3. 相似文献
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10.
Nuclear receptors for 1,25(OH)2 vitamin D3 in thymus reticular cells studied by autoradiography 总被引:1,自引:0,他引:1
Summary After injection of 3H 1,25(OH)2 vitamin D3 to rats fed a vitamin D-deficient diet, nuclear concentration and retention of radioactivity exists in reticular cells of the thymus medulla and cortex, as well as outer cells of developing Hassal's corpuscles. Lymphocytes do not show nuclear concentration of radioactivity. Nuclear concentration in reticular cells is prevented by prior injection of excess 1,25(OH)2 vitamin D3. The results indicate that reticular-endothelial cells contain nuclear receptors for 1,25(OH)2 vitamin D3 and suggest that effects of 1,25(OH)2 vitamin D3 on immune response and lymphocyte differentiation are indirect and mediated through genomic modulation of reticular cell functions such as messenger secretion. 相似文献
11.
Y. Tanaka H.F. Deluca H.K. Schnoes N. Ikekawa Y. Kobayashi 《Archives of biochemistry and biophysics》1980,199(2):473-478
24,24-Difluoro-1,25-dihydroxyvitamin D3 has been synthesized by in vitro incubation of vitamin D-deficient chick kidney homogenates with 24,24-difluoro-25-dihydroxyvitamin D3. The compound produced was isolated and purified by successive high-performance liquid chromatographic steps and then identified by means of ultraviolet absorption spectrophotometry and mass spectrometry. The difluoro analog of 1,25-dihydroxyvitamin D3 is found to be highly active in stimulating intestinal calcium transport and bone calcium mobilization in vitamin D3-deficient rats. 相似文献
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13.
Marthe Rizk-Rabin Rougui Zineb Bouizar Zhor Garabedian Michele Pavlovitch Jana 《Journal of cellular physiology》1994,159(1):131-141
The epidermis is both a target tissue for and a source of 1,25 dihydroxycholecal-ciferol. The present study determines which of the epidermal cell populations synthesizes 1,25 dihydroxycholecalciferol and which responds to this hormone. Epidermal keratinocytes from new born rat epidermis were separated by unit gravity sedimentation into poorly differentiated cells, slow-cycling more differentiated cells, actively proliferating cells, and terminally differentiating subpopulations. The keratinocyte populations were characterized by cell size analysis, cell morphology, and DNA and RNA contents (acridine orange flow cytometry). 1,25(OH)2D3 synthesis was studied by measuring the conversion of [3H] 25(OH)D3 to [3H] 1,25(OH)2D3. The purified product was tested for its ability to compete with synthetic [3H] 1,25(OH)2D3 for binding to chick intestinal cytosol. The responses of the keratinocyte subpopulations to exogeneous 1,25(OH)2D3 were evaluated by the increase in 25(OH)D3-24 hydroxylase activity. Furthermore the expression of 1,25(OH)2D3 receptors (VDR) was examined in these cell populations. The results show that only the least differentiated cells produced 1,25(OH)2D3. In contrast, immunocytochemical detection of VDR, the VDR mRNA, and a 25(OH)D3-24 hydroxylase response to 1,25(OH)2D3 were mainly found in the more differentiated cells. Thus, the ability of epidermis to synthesize 1,25(OH)2D3 and be simultaneously sensitive to it depends on the state of cell differentiation. This suggests that the mammalian epidermis contains a paracrine system in which the more differentiated keratinocytes are sensitive to the 1,25(OH)2D3 produced locally by neighboring immature ones. © 1994 wiley-Liss, Inc. 相似文献
14.
The essential role of vitamin D throughout the life of most mammals and birds as a mediator of calcium homeostasis is well established. In view of the complex endocrine system existent for the regulated metabolism of vitamin D3 to both 1α,25-dihydroxyvitamin D3 [1,25(OH)2D3] and 24R,25-dihydroxyvitamin D3 [24R,25-(OH)2D3] (both produced by the kidney), an intriguing problem is to elucidate whether only one or both of these dihydroxyvitamin D3 metabolites is required for the generation of all the biological responses mediated by the parent vitamin D3. In contrast to the accumulated knowledge concerning the short term actions of 1,25(OH)2-D3 on stimulating intestinal calcium absorption and bone calcium reabsorption, relatively little is known of the biological function of 24,25(OH)2D3. We report now the results of a nine month study in which chicks were raised on a vitamin D-deficient diet from hatching to sexual maturity and received as their sole source of “vitamin D” either 24,25(OH)2D3 or 1,25(OH)2D3 singly or in combination. Specifically we are describing the integrated operation of the vitamin D endocrine system as quantitated by the individual measurement in all birds of 22 variables related to “vitamin D status” and as evaluated by the statistical procedure of multivariate discriminant analysis. Twelve of these variables involved detailed analysis of the bone including quantitative histology and the other 10 variables reflect various manifestations of vitamin D action, e.g. serum Ca2+ and Pi levels, vitamin D-dependent calcium binding protein (CaBP) in the intestine and kidney, egg productivity etc. As evaluated by the multivariate analysis, it is clear that 24,25(OH)2D3 and 1,25(OH)2D3 are simultaneously required for normalization of calcium homeostasis. 相似文献
15.
We have determined the dose-response of 1,25-dihydroxyvitamin D-3 (1,25-(OH)2D3) on the intracellular free calcium-ion concentration ([Ca2+1]i) in the osteoblastic osteosarcoma cells, ROS 17/2.8, using 19F-NMR and the intracellular divalent cation indicator, 1,2-bis(2-amino-5-fluorephenoxy)ethane-N, N, N′, N′-tetraacetic acid (5F-BAPTA). The dose-response demonstrated an inverted U-shaped relationship with maximal elevation of [Ca2+]i at doses of 1 to 10 nM 1,25-(OH)2D3. At 10 nM, 1,25-(OH)2D3 elevated the [Ca2+]i from a control level of 118±4 nM to a peak value of 237±8 nM within 40 min. 1,25-(OH)2D3 also increased the intial rate of Ca2+ influx into ROs 17/2.8 cells, measured by 45Ca uptake, with a dose-response relationship which paralleled its effects on [Ca2+]i. Treatment of ROS 17/2.8 cells with Pb2+ at 1 and 5 μM significantly increased [Ca2+]i but significantly reduced the 1,25-(OH)2D3-induced elevation of [Ca2+]i. Simultaneous treatment of naive cells with 1,25-(OH)2D3 and Pb2+ produce little reduction of 1,25-(OH)2D3 and Pb2+ produce little reduction of 1,25-(OH)2D3-induced 45Ca uptake while 40 min treatment with Pb2+ before addition of 1,25-(OH)2D3 significantly reduced the 1,25-(OH)2D3-induced increase in 45Ca influx. These findings suggest that Pb2+ acts by inhibiting 1,25-(OH)2D3-activation of Ca2+ channels and interferes with 1,25-(OH)2D3 regulation of Ca2+ metabolism in osteoblastic bone cells. 相似文献
16.
Eberhard Mayer Roger Bouillon Anthony W. Norman 《Archives of biochemistry and biophysics》1982,217(1):257-263
The structural requirements for the interaction of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] with an anti-1,25(OH)2D3 antiserum and with the natural cytosolic receptor for 1,25(OH)2D3 isolated from chick intestine have been evaluated quantitatively. The antiserum was raised in a rabbit against a 1,25(OH)2D3-hemisuccinate derivative which was linked to bovine serum albumin at the C-3 position of the steroid. For these cross-reaction studies structural analogs of 1,25(OH)2D3 were used in competitive protein binding assays; their ability to interact with the binding proteins was expressed as relative competitive index (RCI) values where the RCI of 1,25(OH)2D3 is defined to be 100. The results indicate that the 25-hydroxyl group is the most important hydroxyl for the interaction of 1,25(OH)2D3 with this antiserum. The absence of this hydroxyl group decreases the RCI value to 0.2. Lack of the hydroxyl at carbon-3 or carbon-1 decreases the RCI value to 33 or 25, respectively, indicating that the specificity of this antiserum for the A ring is much lower than for the side chain. The high specificity for the side chain is underlined by the fact that insertion of an additional hydroxyl group at C-24 or C-26 of 1,25(OH)2D3 decreases the binding affinity to the antiserum markedly. The chick intestinal mucosal receptor shows a comparable high specificity for the side chain of 1,25(OH)2D3, but an even higher specificity for the A ring in comparison to the antiserum. With the intestinal receptor, the 3-hydroxyl is only 1/ 10th as important as the 1-hydroxyl group and the 25-hydroxyl group for the binding process. Scatchard analysis showed a KD value of 1.7 × 10?10m for the antiserum and 2.3 × 10?10m for the chick intestinal mucosal receptor for the equilibrium binding of 1,25(OH)2D3 at 2 °C. The association rate constant at 2 °C was determined to be 5.8 × 107 M?1 min?1 for the antiserum and 0.55 × 107 M?1 min?1 for the receptor, indicating a 10-fold more rapid association of 1,25(OH)2D3 to the antiserum in comparison to the receptor. Furthermore, the dissociation process was found to be slower for the chick intestinal receptor (dissociation rate constant 3.6 × 10?5 min?1 versus 21.0 × 10?5 min?1). 相似文献
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18.
These studies investigated the initial stimulation of intestinal calcium absorption in the rat by 1,25-dihydroxyvitamin D3. To produce a functional vitamin D3-deficiency, rats were fed a diet containing 2.4% strontium. After 10 days on the diet, intestinal calcium uptake, as measured by everted gut sacs, was significantly depressed. Strontium-fed rats were dosed orally with 1,25-dihydroxyvitamin D3, and changes in intestinal calcium uptake, intestinal alkaline phosphatase activity, and intestinal calcium-binding protein were measured as a function of time after dose. Calcium uptake was significantly increased in the proximal 2.5 cm of the duodenum at 4 h and along the whole duodenum by 7 h. Intestinal alkaline phosphatase activity, measured in a Triton extract of the mucosal homogenate and in isolated brush border complexes, was also increased by 7 h. Using both gel electrophoresis and immunodiffusion against a specific antiserum, an increase in intestinal calcium-binding protein was detected in intestinal supernate at 4 h after dosing. Almost no calcium-binding protein was detectable in strontium-fed rats dosed with propylene glycol only. These time studies are consistent with a role for both alkaline phosphatase and calcium-binding protein in the 1,25-dihydroxyvitamin D3-stimulated uptake of calcium by the intestine. In addition, the usefulness of strontium feeding for producing a functional vitamin D3 deficiency in rats is demonstrated. 相似文献
19.
M. Peterlik K. Bursac M.R. Haussler M.R. Hughes R.H. Wasserman 《Biochemical and biophysical research communications》1976,70(3):797-804
Administration of an aqueous extract of the calcinogenic plant (S.m.) to vitamin D-deficient or strontium fed chicks produces significant plasma 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) activity within 6 hr. (via radioreceptor assay) and subsequently elicits the appearance of immunoreactive intestinal calcium binding protein. Studies of a purified aqueous extract of S.m. show that it does not compete effectively with radioactive 1,25-(OH)2D3 for binding to the sterol's intestinal receptor. However, treatment of the extract with β-glucosidase releases a biologically active substance which is soluble in organic solvents and efficiently competes with labeled sterol for the receptor. This factor migrates exactly with tritiated 1,25-(OH)2D3 on high resolution Celite liquid-liquid partition columns. Thus, S.m. contains a molecule very similar or identical to 1,25-(OH)2D3 which is combined with one or more carbohydrate moieties in the native plant. This glycoside is probably cleaved before biological activity is attained. 相似文献
20.
Y Tanaka H K Schnoes C M Smith H F DeLuca 《Archives of biochemistry and biophysics》1981,210(1):104-109
A polar metabolite of vitamin D3 has been produced in vitro from either 1,25-dihydroxyvitamin D3 incubated with kidney homogenate from vitamin D-supplemented chickens or from 25,26-dihydroxyvitamin D3 incubated with vitamin D-deficient chicken kidney homogenate. This compound was isolated in pure form and identified as 1,25,26-trihydroxyvitamin D3 by ultraviolet absorption spectrophotometry and mass spectrometry. Furthermore, its periodate cleavage product comigrates with synthetic 1α-hydroxy-25-keto-27-norvitamin D3 on high-performance liquid chromatography. The 1,25,26-trihydroxyvitamin D3 is 0.1-0.01 as active as 1,25-dihydroxyvitamin D3 in the stimulation of intestinal calcium transport and bone calcium mobilization. 相似文献