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)

The influence of dexamethasone on expression of the osteocalcin gene which encodes the most abundant non-collagenous and only reported bone-specific protein was examined in ROS 17/2.8 osteosarcoma cells which express a broad spectrum of genes related to bone formation. Consistent with previous reports, quantitation of cellular osteocalcin mRNA levels by Northern blot analysis, osteocalcin gene transcription by activity of the osteocalcin gene promoter fused to a chloramphenicol acetyl-transferase (CAT) mRNA coding sequence following transfection into ROS 17/2.8 cells, and osteocalcin biosynthesis by radioimmunoassay indicate that dexamethasone in a concentration range of 10(-6) to 10(-9) M only modestly modifies basal levels of osteocalcin gene expression. However, dexamethasone significantly inhibits these parameters of the vitamin D-induced upregulation of osteocalcin gene expression in both proliferating and in confluent ROS 17/2.8 cells. In this study, we observed that the extent to which abrogation of the vitamin D response occurs is dependent on basal levels of osteocalcin gene expression as reflected by a complete inhibition of the vitamin D-induced upregulation in a ROS 17/2.8K subline with low basal expression and only a partial reduction of the vitamin D stimulation in a ROS 17/2.8C subline with eightfold higher levels of basal expression. This effect of glucocorticoid appears to be at the transcriptional and post-transcriptional levels as demonstrated by a parallel decline in the cellular representation of osteocalcin mRNA, osteocalcin gene promoter activity, and osteocalcin biosynthesis. The complexity of the glucocorticoid effect on vitamin D-mediated transcriptional properties of the osteocalcin gene is indicated by persistence of sequence-specific protein-DNA interactions at two principal osteocalcin gene promoter regulatory elements, the osteocalcin (CCAAT) box which modulates basal level of transcription, and the vitamin D responsive element, where vitamin D-mediated enhancement of osteocalcin gene transcription is controlled.
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PMID:Influence of dexamethasone on the vitamin D-mediated regulation of osteocalcin gene expression. 175 81

1,25-Dihydroxyvitamin D3 [1,25-(OH)2D3] is the active hormonal form of vitamin D3 and has potent effects on bone and calcium regulation. Over the past decade it has become apparent that 1,25-(OH)2D3 has other effects on cellular proliferation that potentially could be developed for therapy in human malignancy. Since the hypercalcemic effects of 1,25-(OH)2D3 have limited that use in the human, novel nonhypercalcemic analogs of 1,25-(OH)2D3 have been synthesized. The molecular mechanism of this divergence in these antiproliferative and calcium-regulating actions is unexplained. We have previously examined the human bone-specific gene osteocalcin as a model of the molecular mechanisms of vitamin D action in bone and have shown that induction of the osteocalcin gene by 1,25-(OH)2D3 is mediated through an unique and complex palindromic region of the promoter similar to but distinct from those of other steroid hormone-responsive elements. Using an osteosarcoma cell line permanently transfected with the vitamin D-responsive promoter of the human osteocalcin gene linked to a "reporter" gene, we have shown that there is a dose-dependent induction of CAT activity by 1,25-(OH)2D3 and that the potencies of vitamin D metabolites and analogs are comparable to those found in other vitamin D bioassays. Furthermore, vitamin D analogs, including MC-903, 22-oxa-1,25-(OH)2D3, and delta 22-1,25S,26-trihydroxyvitamin D3, which effect cellular differentiation but lack hypercalcemic activity in vivo, exhibit osteocalcin promoter inductive actions virtually identical to those of 1,25-(OH)2D3. Consideration of these and other data support the hypothesis that the divergent effects of such analogs on differentiation and calcium homeostasis reflect pharmacokinetic differences in vivo rather than distinct 1,25-(OH)2D3-sensitive pathways.
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PMID:Nonhypercalcemic 1,25-(OH)2D3 analogs potently induce the human osteocalcin gene promoter stably transfected into rat osteosarcoma cells (ROSCO-2). 178 78

The heterologous regulation of hormone receptors is well described in the hormone receptor literature. We were interested in determining whether human 1,25-dihydroxyvitamin D-3 receptor (hVDR) and glucocorticoid receptor (GR), members of the steroid/thyroid hormone receptor family, are heterologously regulated by other steroids and related hormones. We used human osteosarcoma cells (MG-63) and measured hVDR and GR mRNA levels after androgen, estrogen, glucocorticoid, progesterone, thyroid hormone, vitamin A and vitamin D treatments. Each hormone, except androgen and progesterone, was capable of increasing hVDR mRNA levels like the natural ligand in human osteosarcoma cells. On the other hand, GR gene expression was not affected by these hormones. To study whether the cells responded to the 1,25(OH)2D3-treatment with changes in differentiation and proliferation, we also studied c-myc and c-fos gene expression. Both genes were only regulated by 1,25(OH)2D3. 1,25(OH)2D3 slightly increased the accumulation of c-fos mRNA within 4-12 h from the hormone addition, while the increase in c-myc mRNA appeared at 24 h.
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PMID:Homologous and heterologous regulation of 1,25-dihydroxyvitamin D-3 receptor mRNA levels in human osteosarcoma cells. 184 64

The metabolism of 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] by a human osteoblastic sarcoma cell line, U-2 OS, and by primary cultures of human bone-derived cells was examined at physiologic (5 x 10(-11) M) and pharmacologic (3.5 x 10(-7) M) substrate concentrations. For metabolite identification purposes, cells nearing confluency were incubated for 18 h with 3.5 x 10(-7) M 1,25-(OH)2D3 in serum-free medium. The putative vitamin D metabolites produced during this incubation were isolated from a total lipid extract of cells and medium. Identification of the metabolites was achieved by comigration with authentic standards on three high-performance liquid chromatography systems, UV spectral analysis, mass spectrometry, and chemical modification by sodium borohydride and sodium metaperiodate. The identified metabolites produced from 1,25-(OH)2D3 by the human osteosarcoma cells include 1,24,25-trihydroxyvitamin D3; 24-oxo-1,25-dihydroxyvitamin D3; 24-oxo-1,23,25-trihydroxyvitamin D3; and 24,25,26,27-tetranor-1,23-dihydroxyvitamin D3. Evidence is presented that (1) 1,25-(OH)2D3 metabolism occurs constitutively in U-2 OS osteosarcoma cells at a physiologic substrate concentration (5 x 11(-11) M), (2) the pathway can be further induced by pharmacologic 1,25-(OH)2D3 concentrations (10(-7) M), and (3) this pathway is present in primary cultures of normal human bone-derived cells.
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PMID:1,25-Dihydroxyvitamin D3 metabolism in a human osteosarcoma cell line and human bone cells. 216 23

In UMR 106 rat osteosarcoma cells, parathormone (1-34hPTH) and calcitonin (sCT) stimulated adenylate cyclase (AC) activity 5.5-and 2.8-fold, respectively. AC in osteoblasts (OB) from collagenase-treated calvaria of 3-day-old rats responded similarly to 1-34hPTH. In contrast, fibroblasts (mouse fibroblastomas) displayed a marginal 1-34hPTH sensitive AC. Osteoclasts (OC) of collagenase-treated rat calvariae, rat monocytes and mouse macrophages did not demonstrate 1-34hPTH inducable AC activity. Physiological concentrations of 24,25-dihydroxyvitamin D-3 attenuated PTH-sensitive AC in OB and UMR 106 cells within 20 min, while 1,25-dihydroxyvitamin D-3 showed no such immediate effect. In contrast, the AC response to Gpp(NH)p was unaffected by 24,25-(OH)2D3, indicating that 24,25-(OH)2D3 interrupts the coupling of the PTH receptor to the GTP binding protein Gs. OB and UMR 106 cells were also subjected to long-term (48 h) incubation with vitamin D-3 metabolites, 1-34hPTH or 20% serum from patients with secondary hyperparathyroidism (sHBT-serum), respectively. PTH-sensitive AC was markedly attenuated by pre-exposure to both 1-34hPTH and 1,25-(OH)2D3, while minimally affected by corresponding 24,25-(OH)2D3 and 20% sHPT-serum treatment. The secretion of alkaline phosphatase (Alphos) from the two cell types was strongly increased by 1-34hPTH, the effect being abolished by the presence of 24,25-(OH)2D3. Iliac crest biopsies of normal individuals exhibited a clear negative correlation between PTH-sensitive AC and corresponding serum 24,25-(OH)2D3 levels. Basal AC activity was, however, negatively correlated to serum 1,25-(OH)2D3 concentrations. In summary, the results show that 24,25-(OH)2D3 reduces PTH-stimulated AC activity in and Alphos secretion from osteoblastic bone cells by rapidly and directly interfering with the plasma membrane. These data reinforce the probable in vivo significance of 24,25-(OH)2D3. Moreover, the negative correlation between basal AC activity and serum 1,25-(OH)2D3 levels indicates a possible role for 1,25-(OH)2D3 in regulating bone cell synthesis of AC components in vivo.
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PMID:1,25-dihydroxyvitamin D-3 and 24,25-dihydroxyvitamin D-3 affect parathormone (PTH) -sensitive adenylate cyclase activity and alkaline phosphatase secretion of osteoblastic cells through different mechanisms of action. 216 95

1,25-dihydroxycholecalciferol (1,25(OH)2D3) rapidly affects calcium (Ca2+) transport in several cell systems, suggesting physiological actions independent of genomic activation. To test this hypothesis, we studied immediate to early effects (0.5-300 sec) of 1,25(OH)2D3 on cytosolic Ca2+ [Ca2+]i in single osteogenic sarcoma ROS 17/2.8 cells loaded with fura-2. An acute rise in [Ca2+]i was observed in 40% of the cells following addition of 1,25(OH)2D3, with a threshold concentration of 10(-11) M. In most cases, the [Ca2+]i rise was transient, with return to baseline within 1 min; less frequently a more prolonged effect was observed, with variable recovery times. 25-hydroxycholecalciferol (25(OH)D3) reproduced the effect of 1,25(OH)2D3 on [Ca2+]i, with equal potency and similar responses, whereas 24,25-dihydroxycholecalciferol, 1 alpha-hydroxycholecalciferol, and 22 oxa-1,25(OH)2D3 were not effective. 1,25(OH)2D3 also increased [Ca2+]i in ROS 24/1 cells, which are defective of receptors for the vitamin D metabolites. At high doses (10(-8)-10(-7) M) of 1,25(OH)2D3 the [Ca2+]i rise in ROS 17/2.8 cells was due to both influx of extracellular Ca2+ and release of Ca2+ from intracellular stores, as the effect was only partially inhibited by Ca2(+)-channel blockade by nifedipine. At low doses (10(-9)-10(-10) M), the effect was entirely dependent on extracellular Ca2+. 1,25(OH)2D3 also increased the production of inositol 1,4,5 trisphosphate (Ins(1, 4, 5)P3) and diacylglycerol, at a threshold dose of 10(-9) M, indicating activation of phospholipase C (PLC). In two thirds of the cells studied, a second addition of 1,25(OH)2D3 within 5 min to cells prestimulated with equimolar doses of the vitamin D metabolite resulted in a [Ca2+]i transient of higher amplitude than the first, a phenomenon occurring at all doses of the hormone, and associated with production of Ins(1, 4, 5)P3. This response amplification was not produced by 25(OH)D3, and pretreatment with 1 alpha(OH)D3 did not significantly enhance 1,25(OH)2D3-induced production of Ins(1, 4, 5)P3. In conclusion, activation of the Ca2+ message system by vitamin D metabolites is a rapid, nongenomic effect; 1,25(OH)2D3 specifically activates both PLC and dihydropyridine-sensitive Ca2+ channels, and "primes" the cells to respond with an enhanced [Ca2+]i rise to a subsequent homologous stimulation; the presence of both the 1 alpha and 25 hydroxyl groups is necessary to express the full hormonal action of vitamin D on [Ca2+]i.
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PMID:Nongenomic activation of the calcium message system by vitamin D metabolites in osteoblast-like cells. 222 14

Expression of genes encoding transferrin and the vitamin D-binding protein is described in a cell line, U-2 OS, derived from a human osteogenic sarcoma. The mRNA transcripts of transferrin and vitamin D-binding protein were shown to be the lengths of those found in normal human liver. The cells synthesize and secrete the transferrin and vitamin D-binding proteins, in addition to human albumin and ceruloplasmin. The U-2 OS cells were successfully transfected with chimeric genes carrying 670 bp of the 5' regulatory sequence of the human transferrin gene fused to a reporter chloramphenicol acetyltransferase gene. These data indicate that the appropriate transcriptional factors required for expression of four plasma proteins are produced by U-2 OS nuclei and that the U-2 OS cell line will be useful for studies analyzing regulation of these genes.
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PMID:Expression of transferrin and vitamin D-binding protein genes in an osteogenic sarcoma cell line. 229 48

Monoclonal antibodies were elicited to membrane constituents of the osteoblastic human osteosarcoma cell line Saos-2. Two types of antibody reactivities were characterized: one group of antibodies identified fibroblastic and osteoblastic cultured cells, whereas the other group was specific for the parent cell line, Saos-2. Primary endothelial cells and hepatoma cells were not recognized by either group of antibodies. Through indirect immunofluorescent microscopy, the Saos-2-specific antigen was demonstrated to reside on the surface of these osteosarcoma cells. Metabolic radiolabeling of cultured Saos-2 cells and subsequent immunoprecipitation, electrophoretic separation, and autoradiography revealed this protein to have a Mr of 80,000. Similar experiments in the presence of hormones showed that the expression of this cell surface protein was influenced in an opposing fashion by the bone-regulating hormones parathyroid hormone and vitamin D. Vitamin D stimulated expression by 300%, whereas parathyroid hormone depressed expression by 50%. Thus, Saos-2 human osteoblastic cells demonstrate hormonal regulation through an apparently specific membrane protein.
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PMID:Identification of a vitamin D-responsive protein on the surface of human osteosarcoma cells. 266 84

The antifungal imidazole, ketoconazole, was tested for effects on 1,25-dihydroxyvitamin D-3 (1,25-(OH)2D3) metabolism and binding in intact osteoblast-like osteogenic sarcoma cells (UMR-106). Ketoconazole inhibited the C-24 oxidation of 1,25-(OH)2D3 in a dose-dependent manner. Furthermore, inhibition of 1,25-(OH)2D3 metabolism by ketoconazole resulted, after a lag time of 2 h, in a sharp increase of receptor-bound 1,25-(OH)2D3. The data suggest that the self-induced 1,25-(OH)2D3 metabolism may play an important role in controlling the intracellular levels of and, consequently, receptor occupancy by the active form of vitamin D. Furthermore the results are compatible with the existence of a homologous up-regulation of the 1,25-(OH)2D3-receptor.
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PMID:Effect of ketoconazole on metabolism and binding of 1,25-dihydroxyvitamin D-3 by intact rat osteogenic sarcoma cells. 282 40

We have used a monoclonal antibody (9A7) against the purified avian 1,25-dihydroxyvitamin D3 receptor to develop an immunocytochemical technique for visualization of the protein in fixed tissues and cultured cells. In Bouin's-fixed, chick intestine, 1,25-dihydroxyvitamin D3 receptor-like immunoreactivity was localized mainly in nuclei of epithelial cells and was more abundant in the crypt than in the villar cells. Receptor staining was low or undetectable in liver hepatocytes but was present in nuclei of cells lining the hepatic sinusoids. In rat brain, receptor-like immunoreactivity was abundant and widely distributed, but did not always coincide with the presence of vitamin D-dependent calcium-binding protein; 1,25-dihydroxyvitamin D3 receptor was absent from cerebellar Purkinje cells that contained abundant calcium-binding protein. In disaggregated rat bone cells, receptor immunoreactivity was present in mononuclear cells including osteoblasts and fibroblasts but was absent from osteoclasts. Two separate clones of osteoblast-like, rat osteosarcoma cells, shown in previous studies to be either receptor positive (17/2.8) or negative (24.1), demonstrated nuclear immunoreactivity in exact concordance with receptor levels as determined by ligand binding. The phenomenon of hormone-induced up-regulation of receptor was visualized in receptor-positive 3T6 fibroblasts by demonstration of markedly enhanced nuclear reactivity in cells treated with 10(-7) M 1,25-dihydroxyvitamin D3 for 48 h. Our studies demonstrate the feasibility of the immunocytochemical approach to visualize the 1,25-dihydroxyvitamin D3 receptor in target tissues and show that it is predominantly a nuclear protein in the relatively unoccupied and fully activated states. Moreover, the vitamin D-dependent calcium binding is not a universal marker for 1,25-dihydroxyvitamin D3 action. Rather, our observations suggest that the expression of the 1,25-dihydroxyvitamin D3 receptor may be connected with the state of cellular differentiation.
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PMID:Immunocytochemical localization of the 1,25-dihydroxyvitamin D3 receptor in target cells. 283 Oct 24


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