Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0029463 (osteosarcoma)
 16,637 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A quantitative method for measuring 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) was developed utilizing a luciferase reporter gene under the control of the highly inducible 25-hydroxyvitamin D3 24-hydroxylase promoter in a stably transfected cell line. Transient transfections with constructs containing the 24-hydroxylase gene promoter 5' to a luciferase reporter were first performed in cell lines with high levels of vitamin D receptor, i.e., the rat osteosarcoma (ROS 17/2.8) and human breast cancer (T-47D) cell lines. ROS 17/2.8 cells, stably transfected with the plasmid, gave a 60-fold stimulation with 10(-10) M 1,25-(OH)2D3. A standard curve was constructed showing a large range of response to 1,25-(OH)2D3 (1 pg to 1 ng). The assay was adapted to microtiter plates, which permits a large number of samples to be assayed simultaneously. Other metabolites of vitamin D and analogs such as 25-hydroxyvitamin D3, 24,25-dihydroxyvitamin D3, and 1 alpha-hydroxyvitamin D3 have negligible effects on the detection of 1,25-(OH)2D3, thus eliminating the need for purification of sample. The sensitivity of the method permitted the use of 100 microliters of serum with excellent results. Comparison of this method with a commercially available assay demonstrates that it gives higher sensitivity, simpler manipulations, and comparable results.
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PMID:A highly sensitive method for large-scale measurements of 1,25-dihydroxyvitamin D. 944 54

The molecular mechanisms that couple osteoblast structure and gene expression are emerging from recent studies on the bone extracellular matrix, integrins, the cytoskeleton, and the nucleoskeleton (nuclear matrix). These proteins form a dynamic structural network, the tissue matrix, that physically links the genes with the substructure of the cell and its substrate. The molecular analog of cell structure is the geometry of the promoter. The degree of supercoiling and bending of promoter DNA can regulate transcriptional activity. Nuclear matrix proteins may render a change in cytoskeletal organization into a bend or twist in the promoter of target genes. We review the role of nuclear matrix proteins in the regulation of gene expression with special emphasis on osseous tissue. Nuclear matrix proteins bind to the osteocalcin and type I collagen promoters in osteoblasts. One such protein is Cbfa1, a recently described transcriptional activator of osteoblast differentiation. Although their mechanisms of action are unknown, some nuclear matrix proteins may act as "architectural" transcription factors, regulating gene expression by bending the promoter and altering the interactions between other trans-acting proteins. The osteoblast nuclear matrix is comprised of cell- and phenotype-specific proteins including proteins common to all cells. Nuclear matrix proteins specific to the osteoblast developmental stage and proteins that distinguish osteosarcoma from the osteoblast have been identified. Recent studies indicating that nuclear matrix proteins mediate bone cell response to parathyroid hormone and vitamin D are discussed.
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PMID:Nuclear matrix proteins and osteoblast gene expression. 949 8

1,25-Dihydroxyvitamin D3 (1,25(OH)2D3), the active metabolite of vitamin D, induces nerve growth factor (NGF) synthesis in a variety of different cell lines. The mechanism by which 1,25(OH)2D3 induces NGF, however, is poorly understood. We used a series of full-length and truncated NGF promoter-human growth hormone (hGH) reporter gene plasmids to investigate the mechanism of 1,25(OH)2D3-induced NGF expression in osteoblasts. Untransfected rat osteosarcoma cells (ROS 17/2.8) treated with 1,25(OH)2D3 showed a 2-fold increase in NGF expression compared to control cells. ROS 17/2.8 osteosarcoma cells were transfected with the NGF-hGH reporter plasmids and treated with 10(-)8 M 1,25(OH)2D3. The full-length NGF promoter (-1800 to +120)-hGH reporter construct showed an approximately 2-fold increase in hGH release. Plasmids with successive 5'-deletions showed enhanced hGH expression in treated cells and control cells. A similar series of NGF promoter-hGH reporter gene constructs, lacking the AP-1 site located within the first intron of the NGF gene, were also transiently transfected into ROS 17/2.8 cells. When these cells were treated with the same dose of 1,25(OH)2D3, no increase in hGH expression was seen compared to control cells, demonstrating that this AP-1 site is essential for 1,25(OH)2D3-mediated NGF up-regulation. Since 1,25(OH)2D3 is known to activate the transcription of several genes through its interaction with the vitamin D receptor (VDR), we performed a series of gel electrophoretic mobility shift assays to determine if the VDR binds directly to the AP-1 sequence. No evidence of VDR binding, either as a homodimer or as a heterodimer, to the AP-1 sequence was observed. Treatment of ROS 17/2.8 cells with 1,25(OH)2D3, however, resulted in an increase in AP-1 binding activity; however, no significant changes in c-jun and c-fos levels were observed. Our data show that in osteoblasts, 1,25(OH)2D3 induces NGF expression indirectly by increasing AP-1 binding activity.
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PMID:An AP-1 site in the nerve growth factor promoter is essential for 1, 25-dihydroxyvitamin D3-mediated nerve growth factor expression in osteoblasts. 955 35

The physiologically active metabolite of the vitamin D seco-steroid hormone, 1,25-dihydroxyvitamin D3 (1,25(OH)2D3), is a major regulator of mineral homeostasis. Recent evidence also suggests its role in regulating proliferation and differentiation of cells, including cancer cells. Therapeutic application of 1,25(OH)2D3 to hyperproliferative disease, such as cancer, is thwarted by its hypercalcemic activity. To overcome this problem, analogs of 1,25(OH)2D3 have been produced which retain growth regulating properties and exhibit decreased hypercalcemic activity. In the present study, the efficacy of the vitamin D2 analog, 1,24(S)-dihydroxyvitamin D2 (1,24(S)-(OH)2D2) in the inhibition of cancer cell proliferation and in inducing differentiation of cancer cells was compared to that of 1,25(OH)2D3. By the [3H]-thymidine incorporation procedure, 1,24(S)-(OH)2D2 is as equipotent as 1,25(OH)2D3 in inhibiting the proliferation of five different cell lines, ROS 17/2.8, the rat osteosarcoma cell line, MCF-7, the human breast cancer cell line, HD-11, the chick bone marrow v myc transformed cell line, HT-29, the human colon cancer cell line and HL-60, the human leukemia cell line. The inhibitory action was dose and time-dependent. The NBT reduction method indicated that 1,24(S)-(OH)2D2 induces the differentiation of the human leukemia cell (HL-60) to the same extent as 1,25(OH)2D3. Notwithstanding the vast similarity between 1,24(S)-(OH)2D2 and 1,25(OH)2D3 with regard to the above activities, they differ in their effects on calcium regulation. In conclusion, the present results encourage the use of 1,24(S)-(OH)2D2 for the treatment of cancer disease in vivo.
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PMID:The novel analog 1,24(S)-dihydroxyvitamin D2 is as equipotent as 1,25-dihydroxyvitamin D3 in growth regulation of cancer cell lines. 967 3

The 9,000 Mr calcium-binding protein calbindin-D9k (CaBP9k) is markedly induced by 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] in mammalian intestine. However, although a vitamin D response element (VDRE) has been reported in the promoter of the rat CaBP9k gene (at -490/-472), the CaBP9k promoter is weakly transactivated by 1,25-(OH)2D3. Previous studies indicated that when MCF-7 cells are transfected with the rat CaBP9k VDRE ligated to the thymidine kinase promoter and treated with both 1,25-(OH)2D3 and T3 there is an enhancement of the response observed with 1,25-(OH)2D3 alone, suggesting direct cross-talk between thyroid hormone and the vitamin D endocrine system and activation via the formation of vitamin D receptor (VDR)-thyroid hormone receptor (TR) heterodimers. To determine whether the weak response of the rat CaBP9k natural promoter to 1,25-(OH)2D3 could be enhanced by T3, CaBP9k promoter/reporter chloramphenicol acetyltransferase constructs were transfected in MCF-7 cells, and the cells were treated with the two hormones alone or in combination. No induction with T3 alone and no enhancement of reporter activity in the presence of both hormones was observed. To determine whether a lack of effect by T3 was specific for the CaBP9k promoter and to further examine the possibility of cross-talk between the TR- and VDR-signaling pathways, the 1,25-(OH)2D3-responsive rat 24 hydroxylase [24(OH)ase] promoter and the rat osteocalcin VDRE (-457/-430), both fused to reporter genes were similarly examined in MCF-7 cells. Again, no enhancement of the response to 1,25-(OH)2D3 was observed in the presence of T3. In addition, a similar lack of response to T3 but responsiveness to 1,25-(OH)2D3 was observed when UMR106-01 osteosarcoma cells [which, like MCF-7 cells, express VDR, TR, and the retinoid X receptor (RXR) endogenously] were transfected with a 1,25-(OH)2D3 responsive mouse osteopontin promoter reporter. In vitro DNA binding assays were carried out using purified human VDR, human RXRalpha, and chick T3Ralpha and 24(OH)ase, osteocalcin, osteopontin, and CaBP9k VDRE oligonucleotide probes. No VDR-TR heterodimer binding on any of these VDREs was observed, although, as expected, there was binding by the VDR-RXR complex and strong TR-RXR binding to a consensus thyroid hormone response element. Simultaneous gel retardation assays using similar and lower concentrations of TR with RXR showed strong binding of TR-RXR on a 32P-labeled thyroid response element. Studies using the yeast two-hybrid system also did not provide evidence for the formation of a VDR-TR protein-protein interaction. In addition, in vivo data showed that transfection of TR, in fact, repressed VDR-mediated transcription and that the repression could be reversed by the addition of RXR. Thus, in vitro and in vivo experiments do not support ligand-sensitive transactivation mediated by VDR-TR heterodimer formation but rather suggest that TR expression can repress 1,25-(OH)2D3-induced transcription predominantly by sequestering RXR.
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PMID:Thyroid hormone receptor does not heterodimerize with the vitamin D receptor but represses vitamin D receptor-mediated transactivation. 973 5

We have synthesized several 1alpha,25-dihydroxyvitamin D [1alpha,25(OH)2D] derivatives and evaluated their biological activity in terms of their binding affinity for the vitamin D receptor (VDR) and vitamin D-binding protein (DBP), antiproliferative or differentiation-inducing effects on human promyelocytic leukemic HL-60 cells, and transcriptional activity on a rat 25-hydroxyvitamin D3-24-hydroxylase gene promoter, including two vitamin D-responsive elements (VDREs), and human osteocalcin gene promoter, including a VDRE in transfected human osteosarcoma MG-63 cells. Furthermore, human VDR- or retinoic acid X receptor alpha (RXR alpha)-mediated luciferase activities of the derivatives were also measured by a one-hybrid system in human epitheloid carcinoma, cervix HeLa cells and African green monkey kidney CV-1 cells. Binding affinity for VDR, bone-resorbing activity, antiproliferative and cell-differentiating effects, transactivation potencies on target genes and VDR- or RXR alpha-mediated gene regulations of 1alpha,25(OH)2D2 and 1alpha,25(OH)2D4 were almost comparable to the effects of 1alpha,25(OH)2D3 while 24-epi-1alpha,25(OH)2D2 and 1alpha,25(OH)2D7 were much less active than 1alpha,25(OH)2D3 in these respects. This is the first report concerning biological assessment of 1alpha,25(OH)2D2, 1alpha,25(OH)2D3, 1alpha,25(OH)2D4, 24-epi-1alpha,25(OH)2D2 and 1alpha,25(OH)2D7 at the molecular level, especially with regards to the structural differences at the 24R- or 24S-methyl group and a double bond between carbons 22 and 23 in the side chain of 1alpha,25(OH)2D derivatives.
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PMID:Biological activity profiles of 1alpha,25-dihydroxyvitamin D2, D3, D4, D7, and 24-epi-1alpha,25-dihydroxyvitamin D2. 1032 56

Although canine osteosarcoma is one of the most malignant, aggressive and lethal neoplasms originating from undifferentiated bone cells, it may retain some capacity for normal differentiation. The purpose of this study was to ascertain if the residual capacity for differentiation could be used to suppress its malignant properties. We tested the efficacy of vitamin D and retinoids in inducing differentiation and inhibiting growth of the POS canine osteosarcoma and four of its clonal cell lines, POS 14A (fibroblast type), POS 53B (chondroblast type), POS 53C (undifferentiated type) and POS 53D (osteoblastic type). Treatment with 10(-10)to 10(-8)M concentrations of calcitriol, OCT, cholecalciferol, all-trans retinoic acid (ATRA) and 9-cis retinoic acid for 48-120 hours changed the morphology of POS, POS 53B, POS 53C and POS 53D cells to cells that were elongated and spindle-shaped. Increased number of cytoplasmic organelles and pronounced nuclear activities were induced by concentrations of 10(-8)M and 10(-7)M for 120 hours. All drugs at concentrations of 10(-10)to 10(-8)M for 72 hours inhibited POS growth dose-dependently. OCT significantly reduced the cell number in all cell lines when used at concentrations between 10(-9)and 10(-8)M for 72 hours and exerted significant anti-proliferative effects for eight days culture. This study demonstrated that changed morphology and inhibition of growth was induced by treatment of the cells with these vitamins, that the loss of control of differentiation in the neoplasia was not irreversible and that these drugs may be useful in the clinic.
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PMID:Effects of vitamin D and retinoids on the differentiation and growth in vitro of canine osteosarcoma and its clonal cell lines. 1033 64

The active hormonal form of vitamin D3, 1alpha,25-dihydroxyvitamin D3 (1alpha,25(OH)2D3), has been described as a principal mediator of skeletal homeostasis. Treatment of rat osteosarcoma (ROS)17/2.8, an osteoblast-like cell line, with 1alpha,25(OH)2D3 results in a ligand-dependent increase in transcription of the bone-specific osteocalcin gene. We isolated permanent cell lines that were established by transfecting ROS 17/2.8 cells with plasmids consisting of the human osteocalcin gene promoter containing the vitamin D responsive element linked to a bacterial beta-galactosidase gene. In one of many cell lines, especially in clone NK-31, 1alpha,25(OH)2D3 strongly stimulated beta-galactosidase activity. Reverse transcription-polymerase chain reaction analysis also showed endogenous osteocalcin gene expression and beta-galactosidase gene expression in clone NK-31 cells, which paralleled the increase in beta-galactosidase activity. Using a synthetic analogue of 1alpha,25(OH)2D3, 24,24-difluoro-1alpha,25-dihydroxyvitamin D3, we found that the levels of this activity and these gene expressions were nearly parallel to those of 1alpha,25(OH)2D3. 24R,25-dihydroxyvitamin D3 and 25-hydroxyvitamin D3 at high doses (concentration: 10(-7) M) also induced beta-galactosidase activity in clone NK-31. These cell lines, harboring the plasmid-carrying beta-galactosidase gene under the control of the osteocalcin gene promoter, may contribute to studies on the regulation by 1alpha,25(OH)2D3 or to the development of synthetic analogues of 1alpha,25(OH)2D3.
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PMID:Establishment of permanent cell lines exhibiting vitamin D-dependent expression of beta-galactosidase activity. 1042 66

Tissue-type plasminogen activator (t-PA) is a positive modulator of the plasminogen-plasmin system, which is involved in bone remodeling. In the present study, 1,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] was found to stimulate t-PA gene expression in ROS17/2.8 osteosarcoma cells. Transient transfection analysis and in vitro DNA binding studies identified two vitamin D-responsive elements (VDRE) in the human t-PA enhancer. The first VDRE (bp -7175 to -7146) comprised an inverted palindrome separated by 9 bp (IP9) overlapping a palindrome separated by 3 bp. The second VDRE (bp -7315 to -7302) is an IP2 element overlapping the previously identified retinoic acid-responsive element. 1,25(OH)(2)D(3) treatment of primary osteoblasts derived from t-PAlacZ transgenic mice containing 9 kb of 5' sequence of the human t-PA gene increased the number of lacZ-positive cells, fitting with the probability model of enhancer function.
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PMID:1,25-Dihydroxyvitamin D(3) induction of the tissue-type plasminogen activator gene is mediated through its multihormone-responsive enhancer. 1054 52

1alpha,25-Dihydroxyvitamin D3 [1alpha,25(OH)2D3] has been shown to exert both its nuclear vitamin D receptor (nVDR)-mediated genomic actions and membrane vitamin D receptor (mVDR)-mediated nongenomic actions. In this study, the effects of 1alpha,25(OH)2D3 and its analogues on transmembrane Ca2+ influx were examined in the growth phase of rat osteosarcoma ROS17/2.8 cells. Like BAYK8644 (2 x 10(-5)M), a well-known L-type Ca2+ channel agonist, 1alpha,25(OH)2D3 (10(-8)M) increased transmembrane influx of Ca2+ through voltage-dependent Ca2+ channels and increased intracellular Ca2+ concentration within 2 min of addition to the medium. The 1alpha,25(OH)2D3-induced Ca2+ influx was completely blocked by pre-treatment with nifedipine (2 x 10(-5)M), an L-type Ca2+ channel antagonist. Two vitamin D analogues, 22-oxa-1alpha,25(OH)2D3 (OCT, 10(-8) M) and 20-epi-22-oxa-24a, 26a,27a-trihomo-1alpha,25(OH)2D3 (KH1060, 10(-8)M), which were 3.8 and 3600-fold more active than 1alpha,25(OH)2D3 in stimulating differentiation on human promyelocytic leukemic HL-60 cells, respectively, also increased intracellular Ca2+ concentration, while their Ca2+ channeling activities were similar to or significantly weaker than that of 1alpha,25(OH)2D3. Furthermore, the enhanced transmembrane Ca2+ influx induced by 1alpha,25(OH)2D3 (10(-8)M) or OCT (10(-8)M) was completely blocked by pre-treatment with the respective 1beta epimer [1beta,25(OH)2D3 and 1beta-OCT] at equal concentration. These findings suggest that 1alpha,25(OH)2D3 and its analogues modulate transmembrane Ca2+ influx in osteoblast-like cells by opening L-type Ca2+ channels which can recognize 1alpha-hydroxy analogues as agonists and 1beta-hydroxy analogues as antagonists.
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PMID:Rapid control of transmembrane calcium influx by 1alpha,25-dihydroxyvitamin D3 and its analogues in rat osteoblast-like cells. 1054 55


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