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

---

Query: EC:3.1.3.1 (alkaline phosphatase)
47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A tumor-derived factor believed to cause hypercalcemia by acting on the parathyroid hormone (PTH) receptor was recently purified, cloned, and found to have NH2-terminal sequence homology with PTH. The 1-34 region of this protein was synthesized, evaluated for its postreceptor effects on the ROS 17/2.8 cell line, and its properties were compared to 1-34 PTH. Both 1-34 human humoral hypercalcemia factor (HCF) and 1-34 PTH stimulated adenylate cyclase with an effective concentration (EC)50 of approximately 1 nM. The extent of stimulation by both peptides was equally enhanced by dexamethasone. They both had a pronounced inhibitory effect on growth in the presence of dexamethasone, with an EC50 of approximately 0.1 nM, reduced alkaline phosphatase (AP) activity by approximately 70% in the absence of dexamethasone and by approximately 80% in the presence of dexamethasone with an EC50 of 0.03 nM, and when present at a concentration of 10 nM, reduced AP mRNA levels (estimated by Northern analysis) by approximately 80% in the presence or absence of dexamethasone. Thus, in addition to similar dose-response curves for adenylate cyclase stimulation, both HCF and PTH produced identical postreceptor effects in ROS 17/2.8 cells. These effects of HCF are probably mediated by the interaction of the tumor-derived factor with the PTH receptor.
...
PMID:Comparison of postreceptor effects of 1-34 human hypercalcemia factor and 1-34 human parathyroid hormone in rat osteosarcoma cells. 283 Mar 17

A photoreactive derivative of a sulfur-free bovine parathyroid hormone (PTH) analogue, [Nle8,N-epsilon-(4-azido-2-nitrophenyl)Lys13,Nle18,Tyr34]bovine PTH-(1-34)-NH2 (NAP-NlePTH), was purified from the products of the reaction of [Nle8,Nle18,Tyr34]bovine PTH-(1-34)-NH2 (NlePTH) with 4-fluoro-3-nitro-phenylazide and was used to identify binding components of the PTH receptor in clonal rat osteosarcoma cells (ROS 17/2.8). The purified analogue, NAP-NlePTH, is a fully active agonist in three different ROS 17/2.8 cell bioassays: 1) specific binding to saturable PTH receptors; 2) stimulation of cyclic AMP accumulation; and 3) inhibition of cellular alkaline phosphatase activity; this analogue gave dose response curves parallel to and 25-33% as potent as its parent molecule, NlePTH. Radioiodinated NAP-NlePTH (125I-labeled NAP-NlePTH) retained maximal receptor-binding potency. Radioligand saturation studies in intact cells showed that the Kd of PTH receptors for the photoligand was slightly less than that for 125I-labeled NlePTH (2.8 and 0.8 nM, respectively), but that the Bmax was essentially identical for both radioligands (8 fmol/10(5) cells). Photoaffinity labeling of ROS 17/2.8 cells revealed several 125I-labeled macromolecular components by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. One predominant 125I-labeled band, having an apparent Mr of 80,000 daltons (including Mr = 4,347 ligand; hereafter referred to as the Mr = 80,000 protein), was consistently demonstrated in both reducing and nonreducing conditions. Its labeling was completely inhibited by coincubation with NlePTH (10 nM) at 26-fold molar excess to the photoligand, but not by biologically inactive PTH fragments or unrelated hormone. Labeling of several other macromolecular components persisted in the presence of NlePTH (1 microM). Only the labeling of the Mr = 80,000 protein showed saturation kinetics for photoaffinity labeling; the dose of 125I-labeled NAP-NlePTH (0.8 nM) to half-saturate labeling of the Mr = 80,000 protein was close to the Kd (2.8 nM) of specific binding of the photoligand to receptors in intact ROS 17/2.8 cells. Pretreatment of the cells with NlePTH and dexamethasone led to the predicted proportional decrease or increase, respectively, in labeling of the Mr = 80,000 protein. Our data, using a highly purified photoactive derivative of PTH, having carefully defined chemical and biological properties, show a plasma membrane component of Mr = 80,000 in ROS 17/2.8 cells that possesses the affinity, binding capacity, and physiological characteristics of the PTH receptor.
...
PMID:Photoaffinity labeling of parathyroid hormone receptors in clonal rat osteosarcoma cells. 283 Dec 8

The lateral mobility of alkaline phosphatase (AP) in the plasma membrane of osteoblastic and nonosteoblastic cells was estimated by fluorescence redistribution after photobleaching in embryonic and in tumor cells, in cells that express AP naturally, and in cells transfected with an expression vector containing AP cDNA. The diffusion coefficient (D) and the mobile fraction, estimated from the percent recovery (%R), were found to be cell-type dependent ranging from (0.58 +/- 0.16) X 10(-9) cm2s-1 and 73.3 +/- 10.5 in rat osteosarcoma cells ROS 17/2.8 to (1.77 +/- 0.51) X 10(-9) cm2s-1 and 82.8 +/- 2.5 in rat osteosarcoma cells UMR106. Similar values of D greater than or equal to 10(-9) cm2s-1 with approximately 80% recovery were also found in fetal rat calvaria cells, transfected skin fibroblasts, and transfected AP-negative osteosarcoma cells ROS 25/1. These values of D are many times greater than "typical" values for membrane proteins, coming close to those of membrane lipid in fetal rat calvaria and ROS 17/2.8 cells (D = [4(-5)] X 10(-9) cm2s-1 with 75-80% recovery), estimated with the hexadecanoyl aminofluorescein probe. In all cell types, phosphatidylinositol (PI)-specific phospholipase C released 60-90% of native and transfection-expressed AP, demonstrating that, as in other tissue types, AP in these cells is anchored in the membrane via a linkage to PI. These results indicate that the transfected cells used in this study possess the machinery for AP insertion into the membrane and its binding to PI. The fast AP mobility appears to be an intrinsic property of the way the protein is anchored in the membrane, a conclusion with general implications for the understanding of the slow diffusion of other membrane proteins.
...
PMID:High lateral mobility of endogenous and transfected alkaline phosphatase: a phosphatidylinositol-anchored membrane protein. 288 41

Activation of the 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] receptor-hormone complex was studied in-vitro using cytosolic preparations of rat osteogenic sarcoma cell ROS 17/2-8 and a DNA-cellulose assay. We found that salt was required for extraction of the unoccupied receptor indicating its possible nuclear localization. The 1,25(OH)2D3 receptor underwent an activation process similar to other steroid hormones which could be stimulated by heat and salt. At physiological ionic strength 100% of the complexes were, however, activated at 2 degrees C, indicating that the activation process is not absolutely temperature dependent. In contrast to other steroid hormones, 30-50% of the complexes were in an activated state in the absence of heat and salt moreover, alkaline phosphatase and ammonium sulphate had no effect on activation. Activation was also stimulated by ATP and ATP plus 8BrcAMP indicating the possible role of phosphorylation in the activation process; however, further work is required to clarify this point.
...
PMID:Activation of the 1,25-dihydroxyvitamin D3 receptor in cultured rat osteogenic sarcoma cells. 299 3

To analyze the phenotypic diversity of a clonal rat osteosarcoma cell line (ROS 17/2) we have subcloned the cell line and characterized four subclones, ROS 17/2-A.II, A.III, A.V, and A.XIV. The subclones retained many of the characteristics of the parent clone that are considered typical of normal osteoblast-like cells; they responded to parathyroid hormone and isoproterenol, and had a negligible response to prostaglandin E2 as measured by their respective changes in cyclic AMP concentration. In addition up to a 75% decrease in 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) binding was observed over a four-fold increase in cell density. The morphologies of the subclones varied from spindle-shaped, fibroblast-like to cuboidal. Doubling times varied from 24 to 48 hours, and basal alkaline phosphatase (AP) levels differed by as much as 10 times over the initial 3 months in culture. After 6 months (approximately 100 PDL), the population doubling time of subclone A.XIV decreased from approximately 48 to approximately 20 hrs and there was a 2.5 to 3-fold increase in saturation density. This cell line was designated A.XIV.1 and was compared to a thawed sample from frozen stock of the original A.XIV isolate, designated A.XIV.2. These two populations, the parent cell line (ROS 17/2) and subclone A.V had similar growth properties, but differed with respect to changes in their alkaline phosphatase activity (AP) with time in culture: that is, all clones increased AP with time but there was a three to five-fold difference in their respective AP levels at various times in culture. All clones except A.V exhibited decreased AP activity upon reaching their saturation densities.(ABSTRACT TRUNCATED AT 250 WORDS)
...
PMID:Subclone heterogeneity in a clonally-derived osteoblast-like cell line. 299 73

Based on the finding that retinoic acid (RA) increases 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] receptor number in ROS 17/2 cells, we investigated the effects of RA on the ability of 1,25-(OH)2D3 to regulate alkaline phosphatase activity and PTH-responsive adenylate cyclase in these cells. A maximally effective dose of 1,25-(OH)2D3 (10(-8) M) caused a 75-80% increase in alkaline phosphatase activity and an approximately 70-75% attenuation of the cAMP response to PTH, while RA (10(-6) M) decreased alkaline phosphatase activity by 30-45% and decreased PTH-stimulated cAMP levels by approximately 20%. Preincubation with RA did not enhance the 1,25-(OH)2D3-induced increases in alkaline phosphatase activity. The ED50 values for control and RA-treated cultures were approximately 8 X 10(-10) M and 6 X 10(-10) M, respectively. With regard to PTH responsiveness, the effects of RA preincubation on the 1,25-(OH)2D3 attenuation of cAMP response varied with the concentration of 1,25-(OH)2D3. At low doses (less than 10(-9) M), the effects of 1,25-(OH)2D3 and RA were additive. At higher doses of 1,25-(OH)2D3, the effects of RA and 1,25-(OH)2D3 were not additive, and there were no differences between control- and RA-treated cultures. The ED50 values for control- and RA-treated cultures were 10(-10) M and 3 X 10(-11) M, respectively. None of the above effects were observed using equimolar doses of the vitamin D3 metabolites 24,25-dihydroxyvitamin D3 and 25-hydroxyvitamin D3. The data show that pretreating ROS 17/2A cells with RA to increase 1,25-(OH)2D3 receptors does not correspond with a concomitant increase in the cellular responsiveness to 1,25-(OH)2D3, as measured by increases in alkaline phosphatase activity and decreases in PTH-responsive adenylate cyclase.
...
PMID:Modulation by retinoic acid of 1,25-dihydroxyvitamin D3 effects on alkaline phosphatase activity and parathyroid hormone responsiveness in an osteoblast-like osteosarcoma cell line. 301 60

Cells of the dental papilla are capable of odontoblastic, fibroblastic, and endothelial differentiation and formation of dentin and the dental pulp. In the present study dental papilla cells, obtained from human tooth buds (HDP cells), were cultured in vitro through 3 to 7 passages. After exposure to prostaglandin E2 there was a marked decrease in intracellular cyclic AMP (cAMP) levels as compared to hormone-free controls. Parathyroid hormone and calcitonin had stimulatory effects with 1 and 2 log increases in cAMP, respectively. The HDP cells showed moderate activity of alkaline phosphatase, 1 log higher than that of hamster kidney fibroblasts (BHK 13) and 1 log lower than that of osteoblastic osteosarcoma cells (ROS 17/2). When cultured for 4 or 8 wk in diffusion chambers (DC) implanted in athymic mice, many of the HDP cells underwent odontoblastic morphodifferentiation with very long, single processes extending into the matrix. This matrix contained banded and unbanded collagen fibers. Neither light nor electron microscopy of the DC content revealed mineral deposits. These results suggest that HDP cells have an intrinsic potential for partial odontoblastic differentiation; inductive signals like those originating from odontogenic epithelium are probably essential for the completion of hard tissue formation.
...
PMID:Hormone-responsive cells derived from human dental papilla: characterization in vitro and in vivo in diffusion chambers. 302 24

Ros 17/2 clonal rat osteosarcoma cells calcify when cultured in the presence of 10 micrograms/ml beta-glycerol phosphate in an agarose gel. Culture in 1% agarose inhibited cell division while allowing cells to remain metabolically active and viable for over 21 days. Serial photography of the same microscopic field shows a progressive deposition of calcium phosphate during the course of the experiment. The deposition of calcium around cells was confirmed by calcium-specific stains, and by energy dispersive X-ray analysis (EDX) during scanning electron microscopy. Cells with high calcium content analyzed by EDX had Ca:P ratios similar to hydroxyapatite. Total calcium progressively increased in beta-glycerol phosphate-treated cultures whereas the control plates maintained a constant calcium content over 16 days. Alkaline phosphatase activity increased with time in culture whereas cells with beta-glycerol phosphate maintained the alkaline phosphatase values achieved at the time of initial calcification. Alkaline phosphatase staining revealed no correlation between the presence of the enzyme activity and calcification. Radioimmunoassay for the bone-specific vitamin K-dependent protein bone Gla protein showed that beta-glycerol phosphate-treated cells accumulate over sixfold greater amounts of this protein. Our studies show that ROS cells can calcify and accumulate bone-specific matrix components when cultured in a 3-dimensional agarose matrix.
...
PMID:Calcification of osteoblastlike rat osteosarcoma cells in agarose suspension cultures. 312 Nov 51

In order to elucidate the mechanism of increased alkaline phosphatase (AI-P) activity of bone origin in serum of patients with hyperthyroidism, the effects of thyroid hormone on mouse osteoblast-like cells (MC3T3-E1) were studied in vitro. Triiodo-L-thyronine (T3) and thyroxine (T4) produced a dose-dependent increase in AI-P activity in the cells at minimum concentrations of 10(-10)M T3 (free T3, 5 x 10(-12) M) and 10(-8) M T4 (free T4, 8 x 10(-11) M), respectively. Scatchard analysis revealed that MC3T3-E1 cells contained nuclear binding sites specific for T3 with an apparent Kd of 120 pM (maximum number of binding sites, approximately 2500 per cell). When cells were cultured with T3 in alpha-minimal essential medium (alpha-MEM) for a prolonged period, AI-P activity also became detectable in the conditioned medium. In contrast to rat osteosarcoma cells (ROS 17/2.8), MC3T3-E1 cell growth was inhibited by T4 in a concentration-dependent manner. These findings suggest that thyroid hormone inhibits proliferation and stimulates differentiation of mouse osteoblast-like cells. Since T3 and T4 stimulate AI-P activity not only in the cells but also in the medium, we speculate that the hyper-alkaline phosphatasia frequently seen in patients with hyperthyroid Graves' disease is partly due to a direct effect of thyroid hormone on osteoblasts or osteoblast-like cells.
...
PMID:Stimulation of alkaline phosphatase activity by thyroid hormone in mouse osteoblast-like cells (MC3T3-E1): a possible mechanism of hyperalkaline phosphatasia in hyperthyroidism. 319 Dec 90

To examine the role of lipid metabolism in the growth and function of osteoblast-like cells, we studied ROS 17/2.8 osteosarcoma cells and primary cultures of rat calvarial osteoblasts during growth in a serum-free medium supplemented by purified human lipoproteins or by liposomes. Increase in ROS cell number was measured in sparse (1-5 X 10(3)/cm2) cultures over 6-8 days. Liposomes (0-300 micrograms/ml) and high (HDL), low (LDL), and very low density (VLDL) lipoprotein fractions (0-300 micrograms apoprotein) markedly stimulated cell growth. Cells plated at 5 X 10(3)/cm2 achieved growth rates in the presence of LDL or HDL comparable to 10% fetal bovine serum. Serum-free culture with exogenous lipid maintained the response of cell cyclic AMP accumulation to parathyroid hormone. Cyclic AMP response to parathyroid hormone was enhanced by glucocorticosteroid, and was attenuated by 1,25-dihydroxyvitamin D (1,25(OH)2D) with an EC50 (10(-10) M) comparable to that previously observed in serum-cultured cells (J. Biol. Chem. 258:736, 1985). 1,25(OH)2D also increased the alkaline phosphatase activity in ROS cells cultured in lipid-supplemented serum-free culture. Lipoproteins or liposomes also markedly enhanced the proliferative response of sparse cultures of normal rat osteoblasts to polypeptide mitogens.
...
PMID:Growth of rat osteoblast-like cells in a lipid-enriched culture medium and regulation of function by parathyroid hormone and 1,25-dihydroxyvitamin D. 322 57


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>

---