Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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 expression of parathyroid hormone-related protein (PTHrP) in abnormal human parathyroids was investigated. Northern blot analysis of RNA extracted from human benign parathyroid adenomata (n = 4) revealed multiple PTHrP mRNA species ranging in size from 1.8 to 4 kb. The relative abundance of PTHrP mRNA expressed in two of the adenomata was similar to that of a tumour (DAF) associated with humoral hypercalcaemia of malignancy, whereas PTHrP mRNA was of low abundance in a third and was undetectable in the fourth. PTHrP-like immunoreactivity was detected in extracts of abnormal parathyroid tissue (benign adenoma (n = 7), hyperplasia (n = 5) and parathyroid carcinoma (n = 2] using a sensitive specific two-site immunoradiometric assay for human (h) PTHrP(1-86) and a radioimmunoassay for hPTHrP(1-34). Ratios of hPTHrP(1-86)- and hPTHrP(1-34)-like immunoreactivities relative to hPTH(1-84)-like immunoreactivity in the parathyroid tissue extracts were, on average, less than 1%. PTHrP bioactivity in the extracts could not be distinguished from that of PTH, by an osteosarcoma cell bioassay. We conclude that, despite reports of over-expression of PTHrP mRNA in parathyroid adenomata, the potential contribution of PTHrP to the total PTH-like activity of adenomata and other abnormal parathyroid tissue may be insignificant relative to PTH.
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PMID:Expression of parathyroid hormone-related protein in abnormal human parathyroids. 206 98

[Nle8,18,Tyr34]bPTH-(1-34)amide (NlePTH) was biotinylated using sulfosuccinimidyl 6-(biotinamido)hexanoate, in dimethyl sulfoxide, and the multiple resulting peptides peaks were separated by reverse-phase high performance liquid chromatography. Their biological activities were compared with those of NlePTH, the parent compound, in radioreceptor and cAMP accumulation bioassays using rat osteosarcoma 17/2.8 cells; the earliest two eluting products, bioPTH 1 and 2, were equipotent, a third, bioPTH 3, was only 10% as potent, and the remaining, later eluting derivatives all were less than 0.1% as active. Competitive avidin binding assays using [3H]biotin suggested that bioPTH 1 and 2 had a single biotin congener per molecule, while bioPTH 3 contained two biotin residues. Upon Edman degradation, bioPTH 1 contained biotin on the lysine at position 13 of NlePTH; bioPTH 2's biotin was on the lysine at position 26 (or 27) and bioPTH 3 had biotins on lysines at both positions 13 and 26 (or 27). Avidin tagged with 125I, peroxidase, or fluorescein isothiocyanate was detected on bone-derived cells which had been incubated initially with bioPTH 2 (1, 10, and 100 nM) for 4 h, but not when NlePTH (1 microM) was added with bioPTH 2. A fluorescence-activated cell sorter detected a symmetrical shift in fluorescence of bone-derived cells incubated with 10 nM of bioPTH 2 and 10 micrograms/ml fluorescein isothiocyanate-avidin. Addition of a 30-fold molar excess of NlePTH, or omission of bioPTH 2, completely reversed this fluorescence shift, and no shift in fluorescence was seen with cells lacking PTH receptors. This fully active, high affinity biotinylated PTH-derivative should prove useful in the study of PTH receptor-bearing cells.
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PMID:Characterization of fully active biotinylated parathyroid hormone analogs. Application to fluorescence-activated cell sorting of parathyroid hormone receptor bearing cells. 215 27

We have used both biochemical and morphological techniques to characterize PTH receptors on the clonal osteosarcoma cell line UMR-106, a widely used model of the osteoblast phenotype. 125I-labeled rat (r) PTH-(1-34) bound to a single class of specific saturable receptors on both whole cells and membranes prepared from UMR-106 cells in a time- and temperature-dependent manner. A decrease in PTH receptor affinity seen in the presence of guanine nucleotides demonstrated that PTH receptors on the UMR-106 cells are coupled to guanyl nucleotide-binding proteins. Although PTH is a potent stimulator of adenylate cyclase in the UMR-106 cells, comparison of PTH-stimulated adenylate cyclase and PTH binding curves indicated the presence of receptors that are not linked to the adenylate cyclase system. Our studies also demonstrated that 125I-labeled rPTH-(1-34) bound to UMR-106 cells is rapidly internalized at 22 C, whereas PTH bound at 4 C remains intact and on the cell surface. Internalization of 125I-labeled rPTH-(1-34) was associated with degradation and release of the hormone at 22 C. Three morphologically distinct cell types were identified in subconfluent cultures of UMR-106 cells. Autoradiographic analysis of 125I-labeled rPTH-(1-34) binding demonstrated differential PTH receptor expression in these cell types. The most abundant PTH binding was observed over a cell type with long cytoplasmic extensions. This cell was reminiscent of the predominant PTH target cell previously identified in the rat metaphysis in vivo, suggesting that the UMR-106 cell line may represent neoplastic transformation of the PTH target cell.
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PMID:Biochemical and morphological characterization of parathyroid hormone receptor binding to the rat osteosarcoma cell line UMR-106. 215 23

We have examined the mechanisms of homologous and heterologous regulation of PTH receptor binding and receptor-mediated adenylate cyclase activity in the osteosarcoma cell line UMR-106. Pretreatment with PTH resulted in a time- and dose-dependent decrease in PTH-stimulated adenylate cyclase which was maximal after 2 h and at a concentration of 10(-8) M rat (r)PTH-(1-34). PTH pretreatment over the same dose range also diminished receptor binding of 125I-labeled rPTH-(1-34); however, maximal loss of binding required 14 h and was greater than the loss of maximal adenylate cyclase activity. After 24 h pretreatment with rPTH-(1-34), cell surface receptors were decreased from 21,000 sites per cell to 2,700 sites per cell, and these down-regulated PTH receptors could not be detected in either vesicular or cytosolic subcellular fractions. Recovery from such homologous down-regulation appeared to require new receptor synthesis. Heterologous down-regulation of PTH receptors was demonstrated when UMR-106 cells were preincubated with prostaglandin E2 or (Bu)2cAMP. Heterologous desensitization was shown to be the result of a reversible modification of the PTH receptor which decreased binding affinity and decreased PTH-stimulated adenylate cyclase. Postreceptor components were also examined, and PTH but not prostaglandin E2 pretreatment was shown to decrease guanyl nucleotide binding (G) protein-mediated adenylate cyclase stimulation. This decrease in G protein function was associated with a loss of cholera toxin-catalyzed ADP ribosylation and was also detected by immunoblotting. These results indicate that PTH responses in osteoblastic cells are modulated by diverse mechanisms involving modifications both to the receptor and to postreceptor components of adenylate cyclase.
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PMID:Mechanisms of homologous and heterologous regulation of parathyroid hormone receptors in the rat osteosarcoma cell line UMR-106. 215 32

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

Previous studies examining the interaction of PTH and PTH-related protein (PTHrP) with target tissue have for the most part emphasized the similarity between the two hormones in binding to and activating receptors. This observation that two peptides with limited homology have equal affinities for the same receptor is unusual. In this report we investigated two aspects of PTH/PTHrP-receptor interactions. First, the nonhomologous 14-34 regions of PTH and PTHrP were synthesized and evaluated. Second, hybrid peptides containing the 7-18 fragment of one hormone combined with the 19-34 region of the other hormone were studied to determine whether interactions between these two regions are required for receptor recognition. All four peptides were examined in bovine renal cortical membrane and rat osteosarcoma (ROS 17/2.8) cell PTH-binding and PTH-stimulated adenylate cyclase assays. The results indicate that the receptor-binding domains of PTH and PTHrP lie outside of the 1-13 region, the region containing sequence homology shared by the two hormones, and that two peptides of different amino acid sequence bind with equal affinity to the bovine renal PTH receptor. However, in the absence of the N-terminal region, the rat bone PTH receptor displays a preference for the C-terminal (19-34 sequence) region of PTHrP.
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PMID:The bovine renal parathyroid hormone (PTH) receptor has equal affinity for two different amino acid sequences: the receptor binding domains of PTH and PTH-related protein are located within the 14-34 region. 216 27

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

The human osteosarcoma cell line MG-63 has been used to study the production of the bone-specific protein, osteocalcin. In the absence of any stimuli, MG-63 cells secreted very low levels of osteocalcin. The secretion of osteocalcin started after a lag time of 10-12 h upon 1,25-(OH)2D3 treatment. Osteocalcin secretion was measured at doses as low as 0.03 nM (fourfold increase, p less than 0.05), and this activity increased further with higher doses of 1,25-(OH)2D3 to reach a plateau at 50 nM. The secretion increased transiently from very low levels in sparse cell cultures to peak values in subconfluent cultures (+/- 40%), two- to threefold above values obtained for confluent cells. Values for confluent cells average 55.9 +/- 2.0 ng/ml protein per 48 h. A similar behavior is observed for 1,25-(OH)2D3 receptor concentration under similar experimental conditions. Bmax increased transiently from sparse to subconfluent cell cultures (40-60% confluent) and reached values 50% lower in confluent cells. However, the receptor affinity was not affected by cell density. MG-63 cells also possessed an alkaline phosphatase isoenzyme of the bone-liver-kidney type that was stimulated by 1,25-(OH)2D3 treatment (two- to threefold) and inhibited by parathyroid hormone (40 nM, -25%, p less than 0.025). PTH and PGE2 increased cAMP production in a dose-dependent manner, but the cells were irresponsive to salmon calcitonin. Basal and PTH-responsive cyclic AMP production were also modulated by cell density. Dexamethasone pretreatment (100 nM, 48 h) stimulated the PTH-dependent cAMP production but failed to influence the response to PGE2.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Osteocalcin secretion by the human osteosarcoma cell line MG-63. 217 53

Recent studies have indicated that neutral collagenase can be produced in bones of rats. In addition, it has been demonstrated by in vitro studies that the enzyme is likely secreted by osteoblasts. Cells of the osteoblastic tumor cell line UMR-106 can be stimulated to produce not only collagenase, but also collagenase inhibitor and plasminogen activator. However, it is conceivable that not all osteoblasts produce all of these proteins. In this study, in which UMR cells were maximally stimulated with PTH, only a subpopulation of cells was observed to produce enhanced levels of collagenase but all cells had the ability to synthesize plasminogen activator. Cells of the rat osteosarcoma line UMR-106-01 were stained for the presence of collagenase and tissue plasminogen activator using an immunohistochemical procedure. In many cases, the cells were exposed to monensin for the final 3 h of incubation as well as to the inducing agent PTH. Monensin prevented export of the enzymes, enabling them to be visualized within their cell or origin. Maximal stimulation of collagenase was demonstrated to occur 8 h after exposure to 10(-8) -10(-7) M PTH. Under these conditions, 14-17% of the cells appeared to synthesize elevated amounts of collagenase (as determined by intense staining). Without PTH stimulation, there was a low level of collagenase in all cells, but less than 1% of the cells stained heavily for the enzyme. In contrast, strong staining for plasminogen activator was observed in all cells with or without PTH treatment.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Stimulation of collagenase production by rat osteosarcoma cells can occur in a subpopulation of cells. 217 54

The recent demonstration of estrogen receptors in bone derived cells has stimulated the study of direct effects of sex steroids on bone. We have shown direct stimulation of proliferation by 17 beta-estradiol (E2) of ROS 17/2.8 rat osteogenic osteosarcoma cells, and other bone-derived cells in culture, as well as sex-specific stimulation of diaphyseal bone in vivo by estrogen and testosterone, using [3H]thymidine incorporation into DNA and stimulation of the specific activity of creatine kinase as markers. ROS 17/2.8 cells were used as models of osteoblast-like cells to study the reciprocal modulation of stimulation of bone cell proliferation by sequential treatment by sex steroid and calciotrophic hormones. Pretreatment with 1,25(OH)2D3 and PTH augmented stimulation by E2, while pretreatment with PGE2 followed by E2 resulted in no additional stimulation. Reciprocally, pretreatment with E2 significantly reduced the response to PGE2 while showing an insignificant effect on the response to the other hormones. Gonadectomized Wistar-derived rats provided a useful model system for study of postmenopausal osteoporosis. In diaphyseal bone, [3H]thymidine incorporation and creatine kinase activity decreased 4 weeks after gonadectomy. At that time, a single i.p. injection of E2 in females, and testosterone in males, resulted in a highly significant increase in both these parameters within 24 h.
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PMID:Hormonal stimulation of bone cell proliferation. 225 46


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