UMLS:C0029463 - FACTA Search

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Query: UMLS:C0029463 (osteosarcoma)
16,637 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The effects of the monokines tumor necrosis factor alpha (TNF) and interleukin 1 (IL 1) on parathyroid hormone (PTH)-responsive adenylate cyclase were examined in clonal rat osteosarcoma cells (UMR-106) with the osteoblast phenotype. Recombinant TNF and IL 1 incubated with UMR-106 cells for 48 hr each produced concentration-dependent inhibition of PTH-sensitive adenylate cyclase, with maximal inhibition of PTH response (40% for TNF, 24% for IL 1) occurring at 10(-8) M of either monokine. Both monokines also decreased adenylate cyclase stimulation by the tumor-derived PTH-related protein (PTHrP). In contrast, TNF and IL 1 had little or no inhibitory effect on receptor-mediated stimulation of adenylate cyclase by isoproterenol and nonreceptor-mediated enzyme activation by cholera toxin and forskolin; both monokines increased prostaglandin E2 stimulation of adenylate cyclase. Binding of the radioiodinated agonist mono-[125I]-[Nle8,18, Tyr34]bPTH-(1-34)NH2 to UMR-106 cells in the presence of increasing concentrations of unlabeled [Nle8,18, Tyr34]bPTH-(1-34)NH2 revealed a decline in PTH receptor density (Bmax) without change in receptor binding affinity (dissociation constant, Kd) after treatment with TNF or IL 1. Pertussis toxin increased PTH-sensitive adenylate cyclase activity but did not attenuate monokine-induced inhibition of PTH response. In time course studies, brief (1 hr) exposure of cells to TNF or IL 1 during early culture was sufficient to decrease PTH response but only after exposed cells were subsequently allowed to grow for prolonged periods. Inhibition of PTH response by monokines was blocked by cycloheximide. The results indicate that TNF and IL 1 impair responsiveness to PTH (and PTHrP) by a time- and protein synthesis-dependent down-regulation of PTH receptors linked to adenylate cyclase.
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PMID:Tumor necrosis factor and interleukin 1 inhibit parathyroid hormone-responsive adenylate cyclase in clonal osteoblast-like cells by down-regulating parathyroid hormone receptors. 132 78

In UMR-106 osteosarcoma cells we found that PTH activated both the cAMP/protein kinase A and the Ca(2+)-dependent phosphoinositide/protein kinase C (PKC) pathways, but prostaglandin E2 (PGE2) activated only the cAMP pathway. Activation of PKC by the phorbol ester PMA had no effect on cAMP production but enhanced PTH-stimulated cAMP production by 50% or more; the effect on PGE2-induced cAMP was negligible. Inhibition of the alpha-subunit of the inhibitory guanine nucleotide binding protein (Gi) by pertussis toxin pretreatment also enhanced PTH-mediated cAMP production but had no effect on PGE2-induced cAMP production. These results suggest that although PTH-mediated adenylate cyclase activity is regulated via both the stimulatory (Gs) and inhibitory (Gi) guanine nucleotide binding proteins, only Gs regulates PGE2-mediated adenylate cyclase activity in UMR-106 cells. Costimulation with pertussis toxin and PMA did not increase PTH-stimulated cAMP production above that obtained with PMA alone. This implies a similar target of action for pertussis toxin and PMA, that is, the alpha-subunit of Gi. The alpha-subunit of Gi was found to be a substrate for in vitro PKC phosphorylation of membrane fractions from UMR-106 cells, seen as a +/- 40 kD band on SDS-PAGE. Stimulation of in situ 32P-labeled cells with either PMA or PTH also enhanced incorporation of 32P into the 40 kD band. Using the peptide antisera AS/7 and EC/2, we showed that pertussis toxin-labeled subunits of both Gi1 alpha/Gi2 alpha and Gi3 alpha could be immunoprecipitated, respectively, but immunoprecipitation of membrane proteins after in situ phosphorylation and stimulation with PMA precipitated only Gi2 alpha.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Protein kinase C modulates parathyroid hormone- but not prostaglandin E2-mediated stimulation of cyclic AMP production via the inhibitory guanine nucleotide binding protein in UMR-106 osteosarcoma cells. 133

PTH activates multiple second messengers in its target cells, but the level at which the hormonal signal splits into different pathways is still unknown. To achieve insights on this issue, we have studied the structure-function relationship of PTH by analyzing the effects of bovine PTH-(1-34) [bPTH-(1-34)] and PTH fragments truncated at the N-terminus on the intracellular calcium concentration [( Ca2+]i) and cAMP production in the rat osteogenic sarcoma cell line UMR 106-01. [Ca2+]i was measured in single cells using fura-2. When exposed to 10(-7) M bPTH-(1-34), 20% of the cells responded with a transient increase in [Ca2+]i of variable amplitude. Equimolar doses of bPTH-(2-34), propionyl bPTH-(2-34) [(pbPTH-(2-34)], and bPTH-(3-34) also transiently increased [Ca2+]i, whereas both [tyrosine34]bPTH-(7-34) amide [bPTH-(7-34)] and bPTH-(30-34) were ineffective. The amplitude of the [Ca2+] i transients was dose-dependent, with threshold concentrations of 10(-10) M for bPTH-(1-34) and bPTH-(2-34), and 10(-9) M for bPTH-(3-34). The response rate to the active peptides ranged between 10-30%, without a clear dose-relatedness. A second addition of 10(-7) M bPTH-(1-34) to cells prestimulated with equimolar doses of bPTH-(2-34), pbPTH-(2-34), or bPTH-(3-34) produced another transient, whereas after exposure to 10(-7) M bPTH-(1-34), the cells were completely desensitized to a second homologous stimulation, suggesting that the binding affinity of the truncated peptides for the PTH receptor is lower than that of the intact bPTH-(1-34) fragment. In addition, both bPTH-(1-34) and bPTH-(2-34) dose-dependently stimulated cAMP production, but the former was more potent (ED50 = 10(-9) vs. 10(-7) M, respectively). On the contrary, pbPTH-(2-34), bPTH-(3-34), and bPTH-(7-34) had no effect on cAMP. Pretreating the cells with pertussis toxin to enhance cAMP responses via inhibition of Gi potentiated the effect of bPTH-(1-34) and bPTH-(2-34) and disclosed weak but detectable agonist action of pbPTH-(2-34). These results indicate that specific domains of the PTH molecule are linked to activation of different second messenger pathways; while the first two amino acids are indispensable for activating the cAMP system, generation of the [Ca2+]i signal appears to involve a longer domain, including the amino acid residue in position 3.
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PMID:Dissociation of second messenger activation by parathyroid hormone fragments in osteosarcoma cells. 164 59

Osteoblast-like osteosarcoma cells (ROS 17/2.8) display a rapid transmembrane influx of extracellular calcium after stimulation by 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] that is mediated largely by the opening of voltage-gated calcium channels. These cells also constitutively express high numbers (greater than 18,000/cell) of nuclear receptors for this seco-steroid hormone that are involved in the modulation of genomic activity in the osteoblast and in the up-regulation of transcript ion of osteoblast-specific genes such as osteocalcin. The objective of this study was to determine the structural hierarchy of vitamin D3 analogs with regard to their efficacy as molecular transducers of the genomic and nongenomic pathways that are activated upon treatment of osteoblasts with 1,25-(OH)2D3. To test the structural features of the agonist required for initiation of these distinct pathways, a series of ligand analogs and naturally occurring metabolites of 1,25-(OH)2D3 were used that contain A-ring, D-ring, and side-chain modifications. The abilities of these analogs/metabolites to 1) bind to nuclear receptors and 2) stimulate transmembrane calcium influx were measured. Several analogs (25-hydroxy-16-ene-23-yne-D3 and 25-hydroxy-23-yne D3) were found to stimulate Ca2+ channel opening, but bind only poorly to the 1,25-(OH)2D3 nuclear receptor. Conversely, other analogs (1,24-dihydroxy-22-ene-24-cyclopropyl D3 and 1,25-dihydroxy-16-ene-23-yne,26,27 F6-D3) were found to bind very well to the nuclear receptor, but displayed little or no activity in opening Ca2+ channels. Pertussis toxin, which interferes with coupling of certain ligand-gated receptors to ion channels, failed to block the activation of calcium channels by 1,25-(OH)2D3 or active agonist analogs. Our results indicate that there are likely to be distinct nuclear and plasma membrane-associated forms of the 1,25-(OH)2D3 receptor that are involved in genomic and nongenomic activation of osteoblast activity, respectively. The membrane-associated receptors do not appear to be coupled to pertussis toxin-sensitive G-proteins.
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PMID:Nongenomic actions of 1,25-dihydroxyvitamin D3 in rat osteosarcoma cells: structure-function studies using ligand analogs. 165 87

The effects of phorbol 12-myristate 13-acetate (PMA), a known activator of protein kinase C, on receptor-mediated stimulation of adenylate cyclase were evaluated in a rat osteosarcoma cell line (UMR-106) with the osteoblast phenotype. Pretreatment of UMR-106 cells with PMA increased parathyroid hormone (PTH)-stimulated adenylate cyclase activity and inhibited prostaglandin E2 (PGE2)-responsive enzyme activity. In addition, PMA enhanced enzyme activation by forskolin, which is thought to exert a direct stimulatory action on the catalytic subunit of adenylate cyclase. The regulatory effects of PMA were concentration dependent and of rapid onset (less than or equal to 1 min). Treatment with PMA also resulted in translocation of protein kinase C activity from the cytosol to the particulate cell fraction. Pertussis toxin, which attenuates inhibition of adenylate cyclase mediated by the inhibitory guanine nucleotide-binding regulatory protein (Gi), augmented PTH-sensitive adenylate cyclase activity and reduced the incremental increase in PTH response produced by PMA. The results suggest that activation of protein kinase C increases PTH-stimulated adenylate cyclase activity by actions on Gi and/or the catalytic subunit and decreases PGE2 responsiveness by a mechanism involving the PGE2 receptor.
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PMID:Protein kinase C differentially modulates PTH- and PGE2-sensitive adenylate cyclase in osteoblast-like cells. 173 55

Human alpha-thrombin is known to elicit bone resorption in vitro and has been proposed as a mediator of increased bone turnover in inflammatory diseases. We used UMR 106-H5 rat osteoblast-like osteosarcoma cells to explore the signal transduction mechanism utilized by thrombin in bone. Thrombin produced a dose-dependent increase in the accumulation of [3H]inositol phosphates (IPs) in UMR 106-H5 cells prelabeled with [3H]myo-inositol (EC50 15 U/ml). In saponin-permeabilized cells, GTP gamma S increased [3H]IP production, whereas GDP beta S inhibited the response to both GTP gamma S and thrombin, indicating involvement of a G-protein in thrombin action. Thrombin produced a dose-dependent increase in intracellular free calcium (Cai2+) in UMR 106-H5 cells (EC50 1 U/ml; maximal increase 4-fold), as well as a small (20%) increase in [3H]thymidine incorporation. Treatment of UMR 106-H5 membranes with pertussis toxin (PT) and [32P]NAD+ resulted in labeling of a 40-kDa protein. However, pretreatment of cells with a dose of PT sufficient to produce maximal endogenous labeling of this protein failed to influence thrombin action on IP accumulation, Cai2+, or [3H]thymidine incorporation. In contrast, PT treatment of CCL39 hamster lung fibroblasts significantly blunted thrombin-stimulated [3H]IP accumulation and [3H]thymidine incorporation. These results suggest that thrombin raises Cai2+ in UMR 106-H5 cells by activating polyphosphoinositide-specific phospholipase C. Whereas in fibroblasts and platelets, thrombin receptors appear to couple to both PT-sensitive and PT-insensitive G-proteins, only a PT-insensitive G-protein appears to mediate thrombin action in UMR 106-H5 cells. Either these cells lack the relevant PT-sensitive G-protein or they possess thrombin receptors that selectively couple to a pertussis toxin-insensitive G-protein.
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PMID:Thrombin stimulates inositol phosphate production and intracellular free calcium by a pertussis toxin-insensitive mechanism in osteosarcoma cells. 215 36

In the design and biological evaluation of PTH antagonists, certain analogs, although antagonists in vitro, possess partial agonist properties in vivo that preclude their utility as antagonists. In an effort to identify weak agonism of PTH analogs, an attempt was made to enhance the responsiveness of the widely employed rat osteosarcoma (ROS 17/2.8) cell adenylate cyclase assay. Because responsiveness to PTH in these cells is enhanced upon treatment with dexamethasone (dex) or pertussis toxin (PT), we have evaluated their use to aid in detection of partial agonism for PTH and PTH-related protein (PTHrP) antagonist analogs. Treatment of cells with dex alone (30 nM for 3 days) or with PT alone (40 ng/ml for 1 day) increased basal adenylate cyclase activity by 27%. However, combination of the dex and PT treatments increased basal cAMP production 70%. The in vivo partial agonist [Nle8,18,Tyr34]bPTH(3-34)NH2 increased cAMP production 3-fold over basal levels in untreated cells, nearly 5-fold in PT-treated cells, 8-fold in cells treated with dex, and 10-fold in cells treated with dex plus PT. Similar results were obtained with PTHrP(7-34)NH2: the 6-fold stimulation observed in control cells was converted to 14-fold in cells treated with dex plus PT. Agonist activity undetectable in the conventional assay was observed in the dex plus PT system: [Tyr34]- and [D-Trp12,Tyr34]bPTH(7-34)NH2, which exhibit no agonist activity under control conditions, stimulated cAMP production 2.6- and 2.1-fold, respectively, under dex plus PT treatment. In contrast, the antagonist analogs [Asn10,Leu11]- and [Leu11,D-Trp12]PTHrP(7-34)NH2, hybrid peptides of PTH and PTHrP, had no agonist activity under any conditions. Because of increased responsiveness, this assay should occupy an important step in the pathway for evaluation of PTH antagonists and permit identification of weak partial agonist activity before extensive in vivo testing.
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PMID:Treatment of bone-derived ROS 17/2.8 cells with dexamethasone and pertussis toxin enables detection of partial agonist activity for parathyroid hormone antagonists. 216 26

Pretreatment of the osteogenic sarcoma cell line UMR-106-01 with insulin results in sensitization to both parathyroid hormone (PTH) and isoproterenol. In insulin-pretreated cells, the two hormones cause a significantly greater cyclic AMP (cAMP) accumulation than in noninsulin-treated cells. In the presence of cholera toxin, which enhances cAMP production by these cells in both the basal and PTH-stimulated state, the effect of insulin is maintained. In the presence of pertussis toxin, which has no effect on basal cAMP accumulation but enhances both PTH and isoproterenol stimulation, insulin sensitization for both hormones is abolished. These data suggest that insulin sensitizes these cells to subsequent hormone stimulation by lessening the action of an inhibitory guanine nucleotide regulatory protein, possibly Gi.
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PMID:Insulin sensitizes a cultured rat osteogenic sarcoma cell line to hormones which activate adenylate cyclase. 216 43

The effects of transforming growth factor beta (TGF beta) on parathyroid hormone (PTH)-responsive adenylate cyclase were examined in clonal rat osteosarcoma cells (UMR-106) with the osteoblast phenotype. Purified TGF beta incubated with UMR-106 cells for 48 hr produced a concentration-dependent increase in PTH stimulation of adenylate cyclase, with maximal increase in PTH response (37%) occurring at 1 ng/ml TGF beta. TGF beta also enhanced receptor-mediated activation of adenylate cyclase by isoproterenol and prostaglandin E2 (PGE2) and nonreceptor-mediated enzyme activation by cholera toxin and forskolin. In cells in which PTH-stimulated adenylate cyclase activity was augmented by treatment with pertussis toxin, the incremental increase in PTH response produced by TGF beta was reduced by 33%. However, TGF beta neither mimicked nor altered the ability of pertussis toxin to catalyze the ADP-ribosylation of a 41,000-Da protein, presumably the alpha subunit of the inhibitory guanine nucleotide-binding regulatory component (Gi) of adenylate cyclase, in cholate-extracted UMR-106 cell membranes. TGF beta also had no effect on the levels of alpha or beta subunits of Gi, as assessed by immunotransfer blotting. In time course studies, brief (less than or equal to 30 min) exposure of cells to TGF beta during early culture was sufficient to increase PTH response but only after exposed cells were subsequently allowed to grow for prolonged periods. TGF beta enhancement of PTH and isoproterenol responses was blocked by prior treatment of cells with cycloheximide but not indomethacin. The results suggest that TGF beta enhances PTH response in osteoblast-like cells by action(s) exerted at nonreceptor components of adenylate cyclase. The effect of TGF beta may involve Gi, although in a manner unrelated to either pertussis toxin-catalyzed ADP-ribosylation of the alpha subunit of Gi or changes in levels of Gi subunits. The regulatory action of TGF beta on adenylate cyclase is likely to be mediated by the rapid generation of cellular signals excluding prostaglandins, followed by a prolonged sequence of events involving protein synthesis. These observations suggest a mechanism by which TGF beta may regulate osteoblast responses to systemic hormones.
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PMID:Transforming growth factor beta enhances parathyroid hormone stimulation of adenylate cyclase in clonal osteoblast-like cells. 239 78

1,25-Dihydroxyvitamin D3 [1,25-(OH)2D3] regulates the synthesis of bone gamma-carboxyglutamic acid (Gla) protein (BGP) by osteoblastic cells. In this study we examined the effect of cAMP, alone and in combination with 1,25-(OH)2D3, on the regulation of BGP mRNA levels in ROS 17/2 rat osteosarcoma cells. Elevation of intracellular cAMP levels by cAMP analogs or by isobutylmethylxanthine (IBMX), forskolin, or PTH, resulted in increased BGP mRNA levels and BGP secretion after 1 day of treatment. The effects of these agents were additive with 1,25-(OH)2D3 in stimulating BGP gene expression. After 4 days of treatment, pertussis toxin (PT) and 1,25-(OH)2D3 were synergistic in stimulating BGP mRNA, and the effect of PT could be mimicked by (Bu)2cAMP, IBMX, forskolin, cholera toxin, and to a lesser extent by PTH. The effect of 1-day treatment with cAMP alone and the synergistic effect with 1,25-(OH)2D3 on the stimulation of BGP mRNA were dependent on cell density, while basal and 1,25-(OH)2D3-stimulated synthesis were not. Cyclic AMP inhibited ROS 17/2 cell growth after 1 day of treatment, an effect that was also dependent on initial cell density. After 4 days of treatment, 1,25-(OH)2D3, cAMP, and PT all demonstrated inhibition of cell growth. When cells were treated with actinomycin D, both 1,25-(OH)2D3 and cAMP stimulation of BGP mRNA were blocked. In addition, neither agent was effective in enhancing BGP mRNA stability when prestimulated cells were exposed to actinomycin D.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Bone Gla protein messenger ribonucleic acid is regulated by both 1,25-dihydroxyvitamin D3 and 3',5'-cyclic adenosine monophosphate in rat osteosarcoma cells. 246 56

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