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: EC:3.1.4.1 (phosphodiesterase)
18,767 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The effect of the adenylate cyclase activator forskolin on bone resorption and cyclic AMP accumulation was studied in an organ-culture system by using calvarial bones from 6-7-day-old mice. Forskolin caused a rapid and fully reversible increase of cyclic AMP, which was maximal after 20-30 min. The phosphodiesterase inhibitor rolipram (30 mumol/l), enhanced the cyclic AMP response to forskolin (50 mumol/l) from a net cyclic AMP response of 1234 +/- 154 pmol/bone to 2854 +/- 193 pmol/bone (mean +/- S.E.M., n = 4). The cyclic AMP level in bones treated with forskolin (30 mumol/l) was significantly increased after 24 h of culture. Forskolin, at and above 0.3 mumol/l, in the absence and the presence of rolipram (30 mumol/l), caused a dose-dependent cyclic AMP accumulation with an calculated EC50 (concentration producing half-maximal stimulation) value at 8.3 mumol/l. In 24 h cultures forskolin inhibited spontaneous and PTH (parathyroid hormone)-stimulated 45Ca release with calculated IC50 (concentration producing half-maximal inhibition) values at 1.6 and 0.6 mumol/l respectively. Forskolin significantly inhibited the release of 3H from [3H]proline-labelled bones stimulated by PTH (10 nmol/l). The inhibitory effect by forskolin on PTH-stimulated 45Ca release was significant already after 3 h of culture. In 24 h cultures forskolin (3 mumol/l) significantly inhibited 45Ca release also from bones stimulated by prostaglandin E2 (1 mumol/l) and 1 alpha-hydroxycholecalciferol (0.1 mumol/l). The inhibitory effect of forskolin on spontaneous and PTH-stimulated 45Ca release was transient. A dose-dependent stimulation of basal 45Ca release was seen in 120 h cultures, at and above 3 nmol of forskolin/l, with a calculated EC50 value at 16 nmol/l. The stimulatory effect of forskolin (1 mumol/l) could be inhibited by calcitonin (0.1 unit/ml), but was insensitive to indomethacin (1 mumol/l). Forskolin increased the release of 3H from [3H]proline-labelled bones cultured for 120 h and decreased the amount of hydroxyproline in bones after culture. Forskolin inhibited PTH-stimulated release of Ca2+, Pi, beta-glucuronidase and beta-N-acetylglucosaminidase in 24 h cultures. In 120 h cultures forskolin stimulated the basal release of minerals and lysosomal enzymes.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Use of forskolin to study the relationship between cyclic AMP formation and bone resorption in vitro. 302 78

Although it is well known that aluminum (Al) plays a role in the development of osteomalacia in patients with chronic renal failure, the mechanisms are not fully understood. Since the osteoblasts are the cells responsible for the formation of osteoid tissue, which is greatly affected in patients with Al-induced osteomalacia, it is possible that Al could affect the number of osteoblasts or interfere with their function. To further characterize this potential mechanism, we performed studies in isolated perfused tibiae from normal and Al-treated dogs. In this system, when PTH is added to the perfusate, cAMP, a major marker of osteoblasts, is released. The dogs were divided into two groups: control, and Al-treated (0.75 mg/kg, iv, 5 days a week for 3 months). Thereafter, the dogs were killed, and the tibiae were perfused in vitro. PTH-(1-34) (3-4 ng/ml) and 3-isobutyl-1-methylxanthine (an inhibitor of phosphodiesterase) were added to the perfusate. Basal cAMP secretion was the same in both groups of dogs. After PTH was added to the perfusate, cAMP increased to a peak of 188.2 +/- 30.6 pmol/min in the normal dogs vs. 113 +/- 8.15 in Al-treated dogs (P less than 0.05). Cumulative cAMP secretion over a 30-min period was 766 +/- 127.9 pmol in the normal dogs vs. 455.6 +/- 38.2 pmol in the experimental animals (P less than 0.05). The histological appearance of bone biopsies taken before and after Al administration are consistent with a suppressive effect of the cation on osteoblast function. In particular, the number of osteoblasts had decreased 8-fold (P less than 0.01) under the influence of Al, and tetracycline-based measurements of mineralization kinetics show that osteoblast-mediated calcification was dysfunctional (P less than 0.01-0.025). On the other hand, the histological features of the post Al treatment biopsies suggest that at some time during its administration, the cation stimulates osteoblastic activity. For example, new (woven) bone formation was present in two dogs, and in another, lamellar bone, deposited under the influence of Al, covered the entire trabecular surface. Moreover, Al-associated osteoid was deposited independent of prior resorptive activity, indicating that the cation promotes bone formation in the absence of prior resorption. In keeping with its trophic effect on matrix deposition, Al also led to extensive marrow fibrosis in five dogs, indicating that Al also stimulates the activity of fibroblasts, cells closely related to osteoblasts.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Biological effects of aluminum on normal dogs: studies on the isolated perfused bone. 303 73

Treatment of ROS 17/2.8 osteosarcoma-derived cells with dexamethasone potentiates the PTH stimulation of adenylate cyclase in these cells, yielding a detectable response to as little as 10 pM PTH. Isoproterenol stimulation was also enhanced. The dexamethasone effect is first apparent at 12 h and increases with time of treatment. The apparent EC50 for dexamethasone is 3 nM. Hydrocortisone and corticosterone act similarly to dexamethasone, but require 30-fold higher concentrations. Dexamethasone treatment produces no change in high affinity phosphodiesterase activity. Glucocorticoid-potentiating effects are much more pronounced in whole cells than in broken cells and do not influence forskolin stimulation. Particulate fractions of dexamethasone-treated cells have higher adenylate cyclase specific activity, but are stimulated by guanyl-5'-yl imidodiphosphate to the same extent as control cells. These findings suggest that the glucocorticoids potentiate hormone responsiveness through promotion of hormone receptor-adenylate cyclase coupling by a mechanism dependent on cellular integrity.
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PMID:The effect of dexamethasone on parathyroid hormone stimulation of adenylate cyclase in ROS 17/2.8 cells. 608 91

Prior exposure to PTH markedly decreased the responsiveness of isolated, cultured bone cells to the stimulatory effect of the hormone on cyclic AMP formation. This process of desensitization developed within 30 min, persisted during prolonged incubation of the cells in PTH-free medium, and could not be attributed to enhanced excretion of cyclic AMP from the cells, nor to the extracellular accumulation of an inhibitor of PTH action. Adenylate cyclase activity in a subcellular fraction derived from PTH-treated cells was refractory to PTH and to sodium fluoride. These results indicate that PTH-mediated desensitization reflects, at least in part, impaired cyclic AMP formation. Adenosine and PGE2, known stimulators of bone cell cyclic AMP formation, elicited agonist-specific desensitization, and also desensitized bone cells to the effects of subsequently added PTH. PTH blunted the cellular response to adenosine, but not to PGE2. Modest refractoriness to PTH was evident in cells that had been treated previously with the cyclic AMP phosphodiesterase inhibitors IBMX, theophylline, and Bt2cAMP, whereas treatment with sodium butyrate had no effect. The actions of the inhibitors, like that of PTH, were rapid in onset and long-lasting. Desensitization caused by previous treatment with the phosphodiesterase inhibitors, and with PTH itself, was accompanied by enhanced phosphodiesterase activity in bone cell homogenates. Induction of phosphodiesterase activity may well contribute to desensitization in the bone cell system.
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PMID:Hormonal and nonhormonal desensitization in isolated bone cells. 615 60

The interaction between glucocorticoids (GC) and PTH has been suggested to play a role in the pathogenesis of GC-induced osteopenia. The present studies were designed to examine the effect of acute (5-h) or chronic (4-week) GC administration in vivo on 1) cAMP release by the isolated perfused dog tibia before (basal) and after the addition of synthetic bovine PTH-(1-34) [syn bPTH-(1-34)] (stimulated) to the perfusate in vitro, in the presence or absence of the phosphodiesterase inhibitor 3-isobutyl-1-methyl-xanthine (IBMX; 1 mM), and 2) the percent arteriovenous difference of immunoreactive PTH across bone. Acute administration of 6 mg/kg methylprednisolone (MP) did not affect the basal release of cAMP from bone (6.9 +/- 1.6 pmol/min in control vs. 6.1 +/- 1.2 pmol/min in MP-treated animals); however, syn bPTH-(1-34) stimulated release of cAMP was higher in the MP-treated animals (45 +/- 8.1 pmol/min) than in controls (26.8 +/- 3.0 pmol/min). When IBMX was added to the perfusate, basal cAMP release was not different in control and MP-treated bone (17.2 +/- 2.1 pmol/min in control vs. 19.1 +/- 1.9 pmol/min in MP-treated bone), and syn bPTH-(1-34)-stimulated release of cAMP was equivalent in both groups. In contrast, chronic prednisone therapy lead to a decrease in both basal and PTH-stimulated release of cAMP from bone (3.1 +/- 0.4 and 6.9 +/- 1.6 pmol/min for basal, and 13.1 +/- 1.7 and 26.8 +/- 3.0 pmol/min for stimulated values, respectively). However, the percent changes from the basal levels were not different in the two groups. These results were correlated with histological studies of rib biopsies obtained from these animals, which showed evidence of osteopenia and decreased bone turnover. Neither acute nor chronic GC administration had any effect on arterio-venous differences for PTH across the bone. Thus, these studies demonstrate that 1) acute administration of MP enhances the response of bone to PTH, an effect that is not apparent in the presence of the phosphodiesterase inhibitor IBMX; and 2) chronic prednisone therapy decreased basal and PTH-stimulated cAMP release, an effect that correlated with histological evidence of decreased bone turnover.
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PMID:Altered adenosine 3',5'-monophosphate release in response to parathyroid hormone by isolated perfused bone from glucocorticoid-treated dogs. 619 63

Synthetic bovine parathyroid hormone fragment containing the N-terminal 1-34 amino acids (bPTH-(1-34) ) relaxed the guinea-pig trachea constricted with histamine in vitro. Peptides with bovine and human sequences purchased from Peninsula Laboratories and Beckman Bioproducts produced similar effects. Substitution of methionine in positions 8 and 18 by norleucine did not affect this property of bPTH-(1-34). However, when the methionines were oxidized by treating the peptide with hydrogen peroxide, the peptide could no longer produce relaxation in the trachea. Oxidation of the methionine-replaced analog did not affect the action of the peptide on the trachea. It seems that the methionines per se are not necessary, but once oxidized the conformation of the molecule may be sufficiently altered to affect its ability to relax the trachea. While propranolol can block the relaxing action of isoproterenol, this blocking agent produces no inhibition of the bPTH-(1-34) effect. This action of PTH on the trachea may be related to cAMP because isobutyryl-methylxanthine, a phosphodiesterase inhibitor, potentiates and imidazole, a phosphodiesterase stimulator, inhibits the trachea relaxing action of bPTH-(1-34).
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PMID:Parathyroid hormone (PTH) fragments relax the guinea-pig trachea in vitro. 619 56

We used an in vivo infusion technique to assess the hypothesis that vitamin D metabolites and estrogens modulate tissue responsiveness to parathyroid hormone via effects on the adenylate cyclase-cAMP system. After treatment with these agents for 3-4 days, rats were thyroparathyroidectomized. Twenty-four hours later, parathyroid extract (PTE) was infused, and cAMP in calvaria was measured. The response to PTE was achieved by 2 min and represented a 4-fold increase in the tissue concentration of cAMP at the highest dose of hormone tested. Treatment with 1,25-dihydroxyvitamin D3 [1,25(OH)2D3], did not affect cAMP levels in bone. However, 24,25-dihydroxyvitamin D3 [24,25-(OH)2D3], either 0.25 or 1.25 micrograms daily, led to a major increase in PTE-stimulated cAMP formation, a result which persisted when carried out in chronically thyroparathyroidectomized animals. This effect did not reflect direct stimulation of adenylate cyclase or inhibition of cyclic nucleotide phosphodiesterase from bone by the vitamin metabolite, nor did it operate via the 1,25-(OH)2D3 receptor. 24,25-(OH)2D3 treatment also increased cAMP concentrations in renal cortical slices, but not in liver. Adenylate cyclase activity in kidneys from 24,25-(OH)2D3-treated rats was not different from that found in control tissue, but total cytosol phosphodiesterase activity was diminished. 17 beta-Estradiol, over a daily dose range of 2.5 micrograms to 5.0 mg, lowered basal cAMP levels but did not alter PTE-stimulated cAMP production. We conclude that modulation of PTH action in bone by estrogen does not involve modification of the acute cAMP response to PTH. Further, the results support the concept that there are unique actions of 24,25-(OH)2D3 on bone and kidney which are not duplicated by 1,25(OH)2D3.
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PMID:Effects in vivo of vitamin D metabolites and 17 beta-estradiol on parathyroid hormone-dependent formation of adenosine 3',5'-monophosphate in rat bone. 625 69

Circulating levels of calcium (Ca) and immunoreactive parathyroid hormone (IPTH), and the renal cyclic AMP responses to PTH, calcitonin (CT), and vasopressin (VP) were measured in fetal and neonatal rats. Serum Ca increased from a mean value of 9.1 mg/dl on the 19th day of gestation to 10.9 on day 20. Circulating IPTH decreased from 875 pg/ml to 213. Serum Ca declined rapidly after birth to a nadir of 7.6 by 3 h and IPTH increased to 2,006 pg/ml, indicating that fetal and newborn parathyroids are capable of responding appropriately to changes in circulating Ca. Renal responsiveness to hormones was assessed in vitro in the presence of methylisobutylxanthine, a phosphodiesterase inhibitor. The tissue cyclic AMP response to PTH and CT (15- to 18-fold over basal) was greatest at gestational days 18 and 19, progressively declined throughout the remainder of gestation, and remained low during the first 24 h after birth (6- to 7-fold). Renal cyclic AMP response to VP remained consistently low throughout this period. The depressed renal cyclic AMP response to PTH at the time of birth may contribute to the hypocalcemia found in newborn rats.
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PMID:Regulation of calcium homeostasis in the fetal and neonatal rat. 626 86

In the basal state cyclic nucleotide phosphodiesterase in cytosol of rat kidney cortex is characterized by at least two kinetically distinct activities, with Michaelis-Menten constant (Km) for cAMP of 3.3 X 10(-6) and 3.7 X 10(-5) M. Eight minutes after in vivo injection of parathyroid extract, and persisting over 4 h, phosphodiesterase showed only one apparent activity, Km = approximately 1 X 10(-5) M. This change in kinetic profile was mimicked by the ip injection of 10 mg/kg (Bu)2-cAMP, and was reversed by incubation of cytosol with trifluoperazine or EGTA. Incubation of control cytosol with purified calmodulin duplicated the effect of hormone injection. Chromatography of control cytosol over diethylaminoethyl-cellulose resolved two activity peaks. Peak I, corresponding to the high Km activity of whole cytosol, was stimulated by calmodulin, representing a change in maximum velocity but not in substrate affinity. In addition, calmodulin stimulated the hydrolysis by Peak I of 3',5'-cyclic guanosine monophosphate. Peak II activity was not altered by calmodulin. These studies demonstrate that PTH promotes a rapid and sustained alteration in phosphodiesterase kinetics which is mediated by calcium and calmodulin, and which reflects the activation of the high Km component of the enzyme.
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PMID:Parathyroid hormone induces a calmodulin-dependent alteration in phosphodiesterase activity of rat kidney in vivo. 629 44

Protein phosphatases regulate the activity of signal transduction mechanisms by dephosphorylating activated components. By utilizing selective inhibitors of these phosphatases, we investigated their role in regulating cAMP accumulation in the UMR 106 osteoblast-like tumor cell line. PTHrP, PTH and PGE2 stimulated cAMP accumulation up to 100-fold. Calyculin A, a potent inhibitor of protein phosphatase type 1 (PP1) and type 2A (PP2A), did not affect basal levels of cAMP, but concentrations of 10(-11) M to 10(-8) M increased PTHrP-, PTH-, and PGE2-stimulated cAMP accumulation up to 1.7-fold, and this increase was concentration-dependent. Similar results were obtained with tautomycin, another potent inhibitor of PP1 and PP2A. In contrast, okadaic acid, a potent inhibitor of PP2A which inhibited PP1 less potently, did not enhance PTHrP-, PTH-, or PGE2-stimulated cAMP accumulation. The effect of calyculin A on agonist-stimulated cAMP accumulation persisted in cells treated with isobutyl methylxanthine, a phosphodiesterase inhibitor. When the effect of calyculin A was compared with that of 4 beta-phorbol 12-myristate 13-acetate (PMA), it was found that while PMA enhanced both the receptor and forskolin-stimulated cAMP accumulation, calyculin A had no effect on the forskolin-stimulated cAMP accumulation. The effect of calyculin A on PTHrP- and PTH-stimulated cAMP accumulation persisted in cells treated with PMA. These results suggest that protein phosphatases play an important role in agonist-stimulated cAMP accumulation in osteoblast-like cells, and that PP1 but not PP2A may be the major phosphatase involved. In contrast to activation by protein kinase C, the site of action for the phosphatase appears to be predominantly at a step prior to the activation of adenylyl cyclase in the cAMP signal transduction pathway.
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PMID:Inhibition of serine/threonine protein phosphatases enhances agonist-stimulated cAMP accumulation in UMR 106 osteoblast-like cells. 754 25


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