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1.
In addition to classical roles in calcium homeostasis and bone development, 1,25 dihydroxyvitamin D3 [1,25(OH)2D3] inhibits the growth of several cancer types, including breast cancer. Although cellular effects of 1,25(OH)2D3 traditionally have been attributed to activation of a nuclear vitamin D receptor (VDR), a novel receptor for 1,25(OH)2D3 called 1,25D3-MARRS (membrane-associated, rapid response steroid-binding) protein was identified recently. The purpose of this study was to determine if the level of 1,25D3-MARRS expression modulates 1,25(OH)2D3 activity in breast cancer cells.Relative levels of 1,25D3-MARRS protein in MCF-7, MDA MB 231, and MCF-10A cells were estimated by real-time RT-PCR and Western blotting. To determine if 1,25D3-MARRS receptor was involved in the growth inhibitory effects of 1,25(OH)2D3 in MCF-7 cells, a ribozyme construct designed to knock down 1,25D3-MARRS mRNA was stably transfected into MCF-7 cells. MCF-7 clones in which 1,25D3-MARRS receptor expression was reduced showed increased sensitivity to 1,25(OH)2D3 ( IC50 56 ± 24 nM) compared to controls (319 ± 181 nM; P < 0.05). Reduction in 1,25D3-MARRS receptor lengthened the doubling time in transfectants treated with 1,25(OH)2D3. Knockdown of 1,25D3-MARRS receptor also increased the sensitivity of MCF-7 cells to the vitamin D analogs KH1060 and MC903, but not to unrelated agents (all-trans retinoic acid, paclitaxel, serum/glucose starvation, or the isoflavone, pomiferin). These results suggest that 1,25D3-MARRS receptor expression interferes with the growth inhibitory activity of 1,25(OH)2D3 in breast cancer cells, possibly through the nuclear VDR. Further research should examine the potential for pharmacological or natural agents that modify 1,25D3-MARRS expression or activity as anticancer agents. 相似文献
2.
Yasuko Koshihara Mieko Kawamura Sachiko Endo Chiharu Tsutsumi Hiroaki Kodama Hiromi Oda Shohzo Higaki 《In vitro cellular & developmental biology. Plant》1989,25(1):37-43
Summary We isolated osteoblastic cells derived from human periosteum and established them in culture. Their growth depended on the
presence of ascorbic acid, and the doubling time was 40 to 60 h. The requirement for ascorbic acid was used to high production
of collagen. These cells produced mainly type I collagen and only small amounts of type III collagen determined by reducing
sodium dodecyl sulfate SDS-polyacrylamide gel electrophoresis. The total collagen yield was about 10 mg from 2×107 cells. The cells could be continuously cultured in α-minimum essential medium supplemented with 10% fetal bovine serum for
18 to 40 population doubling levels, depending on the age of the donated periosteum. These cells have the ability to calcify
when incubated with 2 mM α-glycerophosphate-Na2. Calcification as viewed by the naked eye appeared from Day 15 after treatment. Treatment with the active formed vitamin
D3, 1, 25 dihydroxyvitamin D3 enhanced calcification significantly and stimulated osteocalcin production. By electron microscopy, cells with many projections
on their surfaces showed well-developed rough endoplasmic reticulum and actinlike fibers, and large numbers of lysosomes,
mitochondria, and secretion granules. Many matrix vesicles, in which minerals were initially localized, and well-banded collagen
fibrils were seen in the intercellular spaces. These observations demonstrate typical osteoblastic morphology. The above results
indicate that cultured cells from human periostem are osteoblastic cells that have the capacity to differentiate into osteocytes
and to deposit calcified minerals in response to 1,25 dihydroxyvitamin D3. 相似文献
3.
J.L. Taft M. French J.A. Danks R.G. Larkins 《Biochemical and biophysical research communications》1984,121(1):355-363
In contrast to dibuturyl cyclic AMP, the methylxanthine phosphodiesterase inhibitors theophylline and caffeine were found to inhibit the conversion of 25 hydroxyvitamin D3 to 1,25 dihydroxyvitamin D3 in isolated renal tubules from vitamin D deficient chicks. This inhibition occurred at concentrations of methylxanthines which were shown to increase renal tubule cyclic AMP levels. No effect of theophylline or caffeine on 25 hydroxyvitamin D3 metabolism in isolated chick renal mitochondria was detected. Because of a demonstrated inhibitory action of calcium (10 and 20 μmol/l) on renal mitochondrial conversion of 25 hydroxyvitamin D3 to 1,25 dihydroxyvitamin D3, the effect of theophylline and dibutyryl cyclic AMP on cellular calcium-45 efflux and total renal tubule calcium content was estimated. Theophylline 10 mmol/l was found to inhibit renal tubular calcium efflux and to increase total cellular calcium content, while dibutyryl cyclic AMP 1 mmol/l had the reverse effect on both parameters. Divergent actions of the methylxanthines and dibutyryl cyclic AMP on the formation of 1,25 dihydroxyvitamin D3 and renal tubule calcium efflux and content support the hypothesis that intracellular calcium is an important regulator of renal vitamin D metabolism. The results indicate that observed actions of methylxanthines cannot always be ascribed to cyclic AMP accumulation. 相似文献
4.
Prema B. Rapuri J.C. Gallagher Zafar Nawaz 《The Journal of steroid biochemistry and molecular biology》2007,103(3-5):368
Of the various risk factors contributing to osteoporosis, dietary/lifestyle factors are important. In a clinical study we reported that women with caffeine intakes >300 mg/day had higher bone loss and women with vitamin D receptor (VDR) variant, tt were at a greater risk for this deleterious effect of caffeine. However, the mechanism of how caffeine effects bone metabolism is not clear. 1,25-Dihydroxy vitamin D3 (1,25(OH)2D3) plays a critical role in regulating bone metabolism. The receptor for 1,25(OH)2D3, VDR has been demonstrated in osteoblast cells and it belongs to the superfamily of nuclear hormone receptors. To understand the molecular mechanism of the role of caffeine in relation to bone, we tested the effect of caffeine on VDR expression and 1,25(OH)2D3 mediated actions in bone. We therefore examined the effect of different doses of caffeine (0.2, 0.5, 1.0 and 10 mM) on 1,25(OH)2D3 induced VDR protein expression in human osteoblast cells. We also tested the effect of different doses of caffeine on 1,25(OH)2D3 induced alkaline phosphatase (ALP) activity, a widely used marker of osteoblastic activity. Caffeine dose dependently decreased the 1,25(OH)2D3 induced VDR expression and at concentrations of 1 and 10 mM, VDR expression was decreased by about 50–70%, respectively. In addition, the 1,25(OH)2D3 induced alkaline phosphatase activity was also reduced at similar doses thus affecting the osteoblastic function. The basal ALP activity was not affected with increasing doses of caffeine. Overall, our results suggest that caffeine affects 1,25(OH)2D3 stimulated VDR protein expression and 1,25(OH)2D3 mediated actions in human osteoblast cells. 相似文献
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WEHI-3B D− cells differentiate in response to 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) but not to all-trans-retinoic acid (RA) or other inducing agents. Combinations of RA with 1,25-(OH)2D3 interact to produce synergistic differentiation of WEHI-3B D− cells. To determine factors involved in the synergistic interaction, expression of the 1,25-(OH)2D3 receptor (VDR) and retinoid receptors, RARα and RXRα, was measured. No VDR was detected in untreated WEHI-3B D− cells; however, RA and 1,25-(OH)2D3 when used as single agents caused a slight induction of the VDR and in combination produced a marked increase in the VDR. In contrast, no changes in RARα and RXRα were initiated by these compounds. An RAR-selective agonist combined with 1,25-(OH)2D3 produced synergistic differentiation of WEHI-3B D− cells, whereas an RXR-selective agonist did not. To gain information on the role of the VDR in the synergistic interaction, the VDR gene was transferred into WEHI-3B D+ cells, in which no VDR was detected and no synergism was produced. Expression of the VDR conferred differentiation responsiveness to 1,25-(OH)2D3 in WEHI-3B D+ cells. These findings suggest that (a) induction of VDR expression is a key component in the synergistic differentiation induced by 1,25-(OH)2D3 and RA and (b) RAR and not RXR must be activated for enhanced induction of the VDR and for the synergistic differentiation produced by RA and 1,25-(OH)2D3. 相似文献
8.
W. E. Stumpf M. Sar F. A. Reid S. Huang R. Narbaitz H. F. DeLuca 《Cell and tissue research》1981,221(2):333-338
Summary After injection of radiolabeled 1,25 (OH)2 vitamin D3, nuclear concentration of radioactivity is observed in parenchymal cells of the parathyroid gland in pregnant, adult male, and 10-day male neonatal rats. In competition studies with unlabeled 1,25 (OH)2 vitamin D3, but not with 25 (OH) vitamin D3, nuclear uptake is prevented. Experiments with 3H 25 (OH) vitamin D3, in contrast to 3H 1,25 (OH)2 vitamin D3, do not show nuclear concentration in cells of the parathyroid. The results of the autoradiographic studies suggest the presence of receptors for a direct effect of 1,25 (OH)2 vitamin D3 on the parathyroid gland for modulation of parathyroid hormone secretion. 相似文献
9.
《Autophagy》2013,9(5):739-753
In MCF-7 breast tumor cells, ionizing radiation promoted autophagy that was cytoprotective; pharmacological or genetic interference with autophagy induced by radiation resulted in growth suppression and/or cell killing (primarily by apoptosis). The hormonally active form of vitamin D, 1,25D3, also promoted autophagy in irradiated MCF-7 cells, sensitized the cells to radiation and suppressed the proliferative recovery that occurs after radiation alone. 1,25D3 enhanced radiosensitivity and promoted autophagy in MCF-7 cells that overexpress Her-2/neu as well as in p53 mutant Hs578t breast tumor cells. In contrast, 1,25D3 failed to alter radiosensitivity or promote autophagy in the BT474 breast tumor cell line with low-level expression of the vitamin D receptor. Enhancement of MCF-7 cell sensitivity to radiation by 1,25D3 was not attenuated by a genetic block to autophagy due largely to the promotion of apoptosis via the collateral suppression of protective autophagy. However, MCF-7 cells were protected from the combination of 1,25D3 with radiation using a concentration of chloroquine that produced minimal sensitization to radiation alone. The current studies are consistent with the premise that while autophagy mediates a cytoprotective function in irradiated breast tumor cells, promotion of autophagy can also confer radiosensitivity by vitamin D (1,25D3). As both cytoprotective and cytotoxic autophagy can apparently be expressed in the same experimental system in response to radiation, this type of model could be utilized to distinguish biochemical, molecular and/or functional differences in these dual functions of autophagy. 相似文献
10.
Hirokazu Hara 《Free radical research》2013,47(3):279-285
Cytochrome P450 (CYP)-dependent drug metabolism decreases in vivo and in cultured hepatocytes under various immunostimulatory conditions. Nitric oxide (NO) released during inflammation is presumed to be involved in this phenomenon. CYP3A4, which is abundant in the liver and small intestine and participates in the metabolism of various drugs, is known to be induced by 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) in the colon carcinoma cell line Caco-2. In this study we examined whether NO affected CYP3A4 gene expression induced by 1,25(OH)2D3 in Caco-2 cells. Induction of CYP3A4 mRNA by 1,25(OH)2D3 was suppressed in a dose-dependent manner by treatment with the NO donors NOR-4 (15–500 μM) or S-nitroso-N-acetyl-penicillamine (30 μM-1 mM), which spontaneously release NO. These results indicated that NO has an inhibitory effect on the induction of CYP3A4 mRNA by 1,25(OH)2D3 in Caco-2 cells. Treatment with the guanylate cyclase inhibitor ODQ failed to prevent the inhibition of induction of CYP3A4 mRNA by 1,25(OH)2D3. 8-Bromo cGMP had no effect on 1,25-(OH)2D3-induced CYP3A4 gene expression. Therefore, the suppression of CYP3A4 mRNA by NO might be mediated through a guanylate cyclase-independent pathway. 相似文献
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Dafu Chen Yang Li Xuejun Dai Xinhua Zhou Wei Tian Yixin Zhou Xuenong Zou Chi Zhang 《International journal of biological sciences》2013,9(6):649-655
Osteoarthritis (OA) is the most prevalent degenerative joint disease. The highly regulated balance of matrix synthesis and degradation is disrupted in OA, leading to progressive breakdown of articular cartilage. The molecular events and pathways involved in chondrocyte disfunction of cartilage in OA are not fully understood. It is known that 1,25-dihydroxyvitamin D₃ (1,25-(OH)2D3) is synthesized by macrophages derived from synovial fluid of patients with inflammatory arthritis. Vitmain D receptor is expressed in chondrocytes within osteoarthritic cartilage, suggesting a contributory role of 1,25-(OH)2D3 in the aberrant behavior of chondrocytes in OA. However, the physiological function of 1,25-(OH)2D3 on chondrocytes in OA remains obscure. Effect of 1,25-(OH)2D3 on gene expression in chondrocytes was investigated in this study. We found that 1,25-(OH)2D3 activated MMP13 expression in a dose-dependent and time-dependent manner, a major enzyme that targets cartilage for degradation. Interestingly, a specific mitogen-activated protein kinase p38 inhibitor SB203580, but not JNK kinase inhibitor SP600125, abrogated 1,25-(OH)2D3 activation of MMP13 expression. 1,25-(OH)2D3-induced increase in MMP13 protein level was in parallel with the phosphorylation of p38 in chondrocytes. To further address the effect of 1,25-(OH)2D3 on MMP13 expression, transfection assays were used to show that 1,25-(OH)2D3 activated the MMP13 promoter reporter expression. MMP13 is known to target type II collagen and aggrecan for degradation, two major components of cartilage matrix. We observed that the treatment of 1,25-(OH)2D3 in chondrocytes results in downregulation of both type II collagen and aggrecan while MMP13 was upregulated. Taken together, we provide the first evidence to demonstrate that 1,25-(OH)2D3 activates MMP13 expression through p38 pathway in chondrocytes. Since MMP13 plays a major role in cartilage degradation in OA, we speculate that the ability of 1,25-(OH)2D3 to potentiate MMP13 expression might facilitate cartilage erosion at the site of inflammatory arthritis. 相似文献
14.
B. Shannan M. Seifert D.A. Boothman W. Tilgen J. Reichrath 《The Journal of steroid biochemistry and molecular biology》2007,103(3-5):721
Prostate cancer is the most commonly diagnosed cancer in the majority of western countries. Due to their antiproliferative and proapoptotic activity, vitamin D analogues have been introduced recently as an experimental therapy for prostate cancer. Clusterin (CLU) is a glycoprotein that has two known isoforms generated in human cells. A nuclear form of CLU protein (nCLU) is pro-apoptotic, and a secretory form (sCLU) is pro-survival. In this study, we analyzed whether proapoptotic and antiproliferative effects of 1,25(OH)2D3 on LNCaP prostate cancer cells are modulated by expression of sCLU. Using colony forming assay, we studied the effect of treatment with different doses of 1,25(OH)2D3 (10−6, 10−7, 10−10 M) on proliferation of LNCaP cells that were stable transfected and over-express sCLU (LNT-1) as compared to empty vector-transfected cells (LN/C). We also measured apoptosis using TUNEL assay. sCLU over-expression protected against both antiproliferative (30%) and proapoptotic (15%) effects of 1,25(OH)2D3, although this effect was statistically not significant. In conclusion, our findings demonstrate that expression of sCLU modulates growth regulatory effects of 1,25(OH)2D3 in prostate cancer indicating that CLU interferes with vitamin D signalling pathways. 相似文献
15.
Structure-functional characterization of vitamin D receptor (VDR) requires identification of structurally distinct areas of VDR-ligand-binding domain (VDR-LBD) important for biological properties of 1α,25-dihydroxyvitamin D3 (1,25(OH)2D3). We hypothesized that covalent attachment of the ligand into VDR-LBD might alter ‘surface structure’ of that area influencing biological activity of the ligand. We compared anti-proliferative activity of three affinity alkylating derivatives of 1,25(OH)2D3 containing an alkylating probe at 1,3 and 11 positions. These compounds possessed high-affinity binding for VDR; and affinity labeled VDR-LBD. But, only the analog with probe at 3-position significantly altered growth in keratinocytes, compared with 1,25(OH)2D3. Molecular models of these analogs, docked inside VDR-LBD tentatively identified Ser237 (helix-3: 1,25(OH)2D3-1-BE), Cys288 (β-hairpin region: 1,25(OH)2D3-3-BE,) and Tyr295 (helix-6: 1,25(OH)2D3-11-BE,) as amino acids that are potentially modified by these reagents. Therefore, we conclude that the β-hairpin region (modified by 1,25(OH)2D3-3-BE) is most important for growth inhibition by 1,25(OH)2D3, while helices 3 and 6 are less important for such activity. 相似文献
16.
《Bioscience, biotechnology, and biochemistry》2013,77(1):312-315
1α,25-dihydroxyvitamin D3 [1,25-(OH)2D3] phosphorylates the extracellular signal-regulated kinase (ERK), a member of the mitogen-activated protein kinase (MAPK) family, within 30 sec in primary cultured chick skeletal muscle cells. MAPK of HeLa cell lines, which had been stably transfected with a cDNA library derived from mRNA of chick skeletal muscle cells, was also rapidly phosphorylated by 1,25-(OH)2D3. These cell lines have the potential to be a good tool for further investigation of rapid non-genomic mechanism activated by 1,25-(OH)2D3. 相似文献
17.
Dr. Madhabananda Sar Walter E. Stumpf Hector F. DeLuca 《Cell and tissue research》1980,209(1):161-166
Summary In the anterior pituitary of the rat, target cells of 1,25 (OH)2 vitamin D3 are identified as those that secrete thyroid stimulating hormone by means of a combined technique of thaw-mount autoradiography and immunohistochemistry. The results for the first time provide evidence that suggests a central effect of 1,25 (OH)2 vitamin D3 on the modulation of thyrotropin secretion in a manner similar to that of other steroid hormones at the level of the pituitary.Supported by PHS grant NS09914 相似文献
18.
1,25D3 is critical for the maintenance of normal reproduction since reduced fertility is observed in male rats on a vitamin D-deficient diet. Vitamin D-deficient male rats have incomplete spermatogenesis and degenerative testicular changes. In the present study we have examined the ionic involvement and intracellular messengers of the stimulatory effect of 1,25D3 on amino acid accumulation in immature rat testis. 1,25D3 stimulates amino acid accumulation from 10−12 to 10−6 M by increasing the slope to reach a maximum value at 10−10 M, as compared to the control group. No effect was observed at a lower dose (10−13 M). Time-course showed an increase on amino acid accumulation after 15, 30, and 60 min of incubation with 1,25D3 (10−10 M). 1,25D3 stimulated amino acid accumulation in 11-day-old rat testis but not in testis that were 20 days old. Cycloheximide totally blocked the 1,25D3 action on amino acid accumulation. Furthermore, a localized elevation of cAMP increased the stimulatory effect of 1,25D3 and the blockage of PKA nullified the action of the hormone. In addition, 1,25D3 action on amino acid accumulation was also mediated by ionic pathways, since verapamil and apamine diminished the hormone effect. The stimulatory effect of 1,25D3 on amino acid accumulation is age-dependent and specific to this steroidal hormone since testosterone was not able to change amino acid accumulation in both ages studied. This study provides evidence for a dual effect for 1,25D3, pointing to a genomic effect that can be triggered by PKA, as well as to a rapid response involving Ca2+/K+ channels on the plasma membrane. 相似文献
19.
Quantitation of vitamin D and its metabolites and their plasma concentrations in five species of animals 总被引:5,自引:0,他引:5
Chromatographic methods suitable for the resolution of 24,25-dihydroxyvitamin D3, 24,25-dihydroxyvitamin D2, 25-hydroxyvitamin D3-26,23 lactone, and 25,26-dihydroxyvitamin D2 are described. These four metabolites comigrated in high-pressure liquid chromatography on silicic acid columns developed in 11:89 isopropanol:hexane. Adequate resolution was achieved by subjecting the four-metabolite complex to high-pressure liquid chromatography column developed in 2:98 isopropanol:methylene chloride. This additional chromatographic step, coupled with modifications of assay procedures previously described, allowed for the estimation of plasma concentrations of vitamin D2, vitamin D3, 25-hydroxyvitamin D2, 25-hydroxyvitamin D3, 24,25-dihydroxyvitamin D2, 24,25-dihydroxyvitamin D3, 25,26 dihydroxyvitamin D2, 25,26-dihydroxyvitamin D3, 25-hydroxyvitamin D3-26,23 lactone, and 1,25-dihydroxyvitamin D (1,25-dihydroxyvitamin D2 plus 1,25-dihydroxyvitamin D3). The samples automatically were introduced onto the high-pressure liquid chromatography columns with a Waters 710A “intelligent” processor. The metabolites were automatically collected with the aid of a programmable timer that advanced a fraction collector at predetermined intervals. The assays were used to determine the plasma vitamin D and vitamin D metabolite concentrations in five species of adult farm animals. 相似文献
20.
Susumi Hatakeyama Satoshi Nagashima Naoko Imai Keisuke Takahashi Jun Ishihara Atsuko Sugita Takeshi Nihei Hitoshi Saito Fumiaki Takahashi Noboru Kubodera 《The Journal of steroid biochemistry and molecular biology》2007,103(3-5):222
1α,25-Dihydroxy-2β-(3-hydroxypropoxy)vitamin D3 (ED-71), an analog of active vitamin D3, 1α,25-dihydroxyvitamin D3 [1,25(OH)2D3], possesses a hydroxypropoxy substituent at the 2β-position of 1,25(OH)2D3. ED-71 has potent biological effects on bone and is currently under phase III clinical studies for bone fracture prevention. It is well-known that the synthesis and secretion of parathyroid hormone (PTH) is regulated by 1,25(OH)2D3. Interestingly, during clinical development of ED-71, serum intact PTH in osteoporotic patients did not change significantly upon treatment with ED-71. The reason remains unclear, however. Brown et al. reported that 3-epi-1,25(OH)2D3, an epimer of 1,25(OH)2D3 at the 3-position, shows equipotent and prolonged activity compared to 1,25(OH)2D3 at suppressing PTH secretion. Since ED-71 has a bulky hydroxypropoxy substituent at the 2-position, epimerization at the adjacent and sterically hindered 3-position might be prevented, which may account for its weak potency in PTH suppression observed in clinical studies. We have significant interest in ED-71 epimerization at the 3-position and the biological potency of 3-epi-ED-71 in suppressing PTH secretion. In the present studies, synthesis of 3-epi-ED-71 and investigations of in vitro suppression of PTH using bovine parathyroid cells are described. The inhibitory potency of vitamin D3 analogs were found to be 1,25(OH)2D3 > ED-71 ≥ 3-epi-1,25(OH)2D3 3-epi-ED-71. ED-71 and 3-epi-ED-71 showed weak activity towards PTH suppression in our assays. 相似文献