Influence of phosphatase inhibitors and nucleotides on [3H]dexamethasone binding in cytosol of human placenta

The purpose of this investigation was to establish the properties of [3H]dexamethasone binding sites in cytosol of human placenta at term. Cytosol containing 20 mM sodium molybdate (MoO4Na2) was incubated for 120min at 20°C with 40 nM [3H]dexamethasone. The following properties were observed: (a) a...

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Publicado: 1984
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Acceso en línea:https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_00224731_v21_n4_p381_Heller
http://hdl.handle.net/20.500.12110/paper_00224731_v21_n4_p381_Heller
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spelling paper:paper_00224731_v21_n4_p381_Heller2023-06-08T14:51:01Z Influence of phosphatase inhibitors and nucleotides on [3H]dexamethasone binding in cytosol of human placenta 11beta hydroxysteroid dehydrogenase 4 nitrophenyl phosphate acid phosphatase adenosine triphosphate alkaline phosphatase cortisone cyclic amp cyclic gmp dexamethasone estradiol fluoride sodium glucocorticoid receptor guanosine triphosphate hydrocortisone isobutylmethylxanthine molybdate sodium nucleotide phosphatase progesterone radioisotope testosterone dexamethasone h 3 drug analysis drug antagonism drug comparison drug identification drug interaction drug isolation drug protein binding female genital system human human cell hydrocortisone h 3 inhibitor normal human placenta pregnancy priority journal Acid Phosphatase Alkaline Phosphatase Binding, Competitive Cytosol Dexamethasone Female Human Kinetics Molybdenum Placenta Pregnancy Receptors, Glucocorticoid Receptors, Steroid Ribonucleotides Sodium Fluoride Support, Non-U.S. Gov't Tungsten The purpose of this investigation was to establish the properties of [3H]dexamethasone binding sites in cytosol of human placenta at term. Cytosol containing 20 mM sodium molybdate (MoO4Na2) was incubated for 120min at 20°C with 40 nM [3H]dexamethasone. The following properties were observed: (a) a single population of binding sites of high affinity and low capacity was measured by Scatchard analysis; (b) potent glucocorticoids such as dexamethasone and cortisol displaced the tritiated ligand, progesterone showed an intermediate activity, whereas cortisone, testosterone and 17β-estradiol were ineffective competitors; (c) ultracentrifugation on 16-41% glycerol gradients containing 20 mM MoO4Na2 yielded sedimentation values of 10.25 ± 0.35 S (n = 4 placentas); (d) the binding sites could be differentiated from the enzyme 11β-hydroxysteroid dehydrogenase, as the activity of the former, but not that of the latter, was greatly dependent on the presence of MoO4Na2 in the incubation medium. Inactivation of binding sites labelled with [3H]dexamethasone by incubation at 20°C was prevented by phosphatase inhibitors such as 20 mM MoO4Na2 (P < 0.01), 20 mM sodium tungstate (WO4Na2) (P < 0.01) and to a lower extent by 5 mM ATP and cAMP (P < 0.05). 50 mM NaF, 5 mM GTP or cGMP had no effect. The protection afforded by MoO4Na2 and WO4Na2 was correlated with a significant inhibition of the activity of acid phosphatase, but not alkaline phosphatase. Neither ATP nor cAMP modified phosphatase activity. It is suggested that binding sites for [3H]dexamethasone in cytosol of human placenta showed properties similar to those described for glucocorticoid receptors in target cells, and that these binding sites are regulated by phosphorylation and dephosphorylation mechanisms. © 1984. 1984 https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_00224731_v21_n4_p381_Heller http://hdl.handle.net/20.500.12110/paper_00224731_v21_n4_p381_Heller
institution Universidad de Buenos Aires
institution_str I-28
repository_str R-134
collection Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA)
topic 11beta hydroxysteroid dehydrogenase
4 nitrophenyl phosphate
acid phosphatase
adenosine triphosphate
alkaline phosphatase
cortisone
cyclic amp
cyclic gmp
dexamethasone
estradiol
fluoride sodium
glucocorticoid receptor
guanosine triphosphate
hydrocortisone
isobutylmethylxanthine
molybdate sodium
nucleotide
phosphatase
progesterone
radioisotope
testosterone
dexamethasone h 3
drug analysis
drug antagonism
drug comparison
drug identification
drug interaction
drug isolation
drug protein binding
female genital system
human
human cell
hydrocortisone h 3
inhibitor
normal human
placenta
pregnancy
priority journal
Acid Phosphatase
Alkaline Phosphatase
Binding, Competitive
Cytosol
Dexamethasone
Female
Human
Kinetics
Molybdenum
Placenta
Pregnancy
Receptors, Glucocorticoid
Receptors, Steroid
Ribonucleotides
Sodium Fluoride
Support, Non-U.S. Gov't
Tungsten
spellingShingle 11beta hydroxysteroid dehydrogenase
4 nitrophenyl phosphate
acid phosphatase
adenosine triphosphate
alkaline phosphatase
cortisone
cyclic amp
cyclic gmp
dexamethasone
estradiol
fluoride sodium
glucocorticoid receptor
guanosine triphosphate
hydrocortisone
isobutylmethylxanthine
molybdate sodium
nucleotide
phosphatase
progesterone
radioisotope
testosterone
dexamethasone h 3
drug analysis
drug antagonism
drug comparison
drug identification
drug interaction
drug isolation
drug protein binding
female genital system
human
human cell
hydrocortisone h 3
inhibitor
normal human
placenta
pregnancy
priority journal
Acid Phosphatase
Alkaline Phosphatase
Binding, Competitive
Cytosol
Dexamethasone
Female
Human
Kinetics
Molybdenum
Placenta
Pregnancy
Receptors, Glucocorticoid
Receptors, Steroid
Ribonucleotides
Sodium Fluoride
Support, Non-U.S. Gov't
Tungsten
Influence of phosphatase inhibitors and nucleotides on [3H]dexamethasone binding in cytosol of human placenta
topic_facet 11beta hydroxysteroid dehydrogenase
4 nitrophenyl phosphate
acid phosphatase
adenosine triphosphate
alkaline phosphatase
cortisone
cyclic amp
cyclic gmp
dexamethasone
estradiol
fluoride sodium
glucocorticoid receptor
guanosine triphosphate
hydrocortisone
isobutylmethylxanthine
molybdate sodium
nucleotide
phosphatase
progesterone
radioisotope
testosterone
dexamethasone h 3
drug analysis
drug antagonism
drug comparison
drug identification
drug interaction
drug isolation
drug protein binding
female genital system
human
human cell
hydrocortisone h 3
inhibitor
normal human
placenta
pregnancy
priority journal
Acid Phosphatase
Alkaline Phosphatase
Binding, Competitive
Cytosol
Dexamethasone
Female
Human
Kinetics
Molybdenum
Placenta
Pregnancy
Receptors, Glucocorticoid
Receptors, Steroid
Ribonucleotides
Sodium Fluoride
Support, Non-U.S. Gov't
Tungsten
description The purpose of this investigation was to establish the properties of [3H]dexamethasone binding sites in cytosol of human placenta at term. Cytosol containing 20 mM sodium molybdate (MoO4Na2) was incubated for 120min at 20°C with 40 nM [3H]dexamethasone. The following properties were observed: (a) a single population of binding sites of high affinity and low capacity was measured by Scatchard analysis; (b) potent glucocorticoids such as dexamethasone and cortisol displaced the tritiated ligand, progesterone showed an intermediate activity, whereas cortisone, testosterone and 17β-estradiol were ineffective competitors; (c) ultracentrifugation on 16-41% glycerol gradients containing 20 mM MoO4Na2 yielded sedimentation values of 10.25 ± 0.35 S (n = 4 placentas); (d) the binding sites could be differentiated from the enzyme 11β-hydroxysteroid dehydrogenase, as the activity of the former, but not that of the latter, was greatly dependent on the presence of MoO4Na2 in the incubation medium. Inactivation of binding sites labelled with [3H]dexamethasone by incubation at 20°C was prevented by phosphatase inhibitors such as 20 mM MoO4Na2 (P < 0.01), 20 mM sodium tungstate (WO4Na2) (P < 0.01) and to a lower extent by 5 mM ATP and cAMP (P < 0.05). 50 mM NaF, 5 mM GTP or cGMP had no effect. The protection afforded by MoO4Na2 and WO4Na2 was correlated with a significant inhibition of the activity of acid phosphatase, but not alkaline phosphatase. Neither ATP nor cAMP modified phosphatase activity. It is suggested that binding sites for [3H]dexamethasone in cytosol of human placenta showed properties similar to those described for glucocorticoid receptors in target cells, and that these binding sites are regulated by phosphorylation and dephosphorylation mechanisms. © 1984.
title Influence of phosphatase inhibitors and nucleotides on [3H]dexamethasone binding in cytosol of human placenta
title_short Influence of phosphatase inhibitors and nucleotides on [3H]dexamethasone binding in cytosol of human placenta
title_full Influence of phosphatase inhibitors and nucleotides on [3H]dexamethasone binding in cytosol of human placenta
title_fullStr Influence of phosphatase inhibitors and nucleotides on [3H]dexamethasone binding in cytosol of human placenta
title_full_unstemmed Influence of phosphatase inhibitors and nucleotides on [3H]dexamethasone binding in cytosol of human placenta
title_sort influence of phosphatase inhibitors and nucleotides on [3h]dexamethasone binding in cytosol of human placenta
publishDate 1984
url https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_00224731_v21_n4_p381_Heller
http://hdl.handle.net/20.500.12110/paper_00224731_v21_n4_p381_Heller
_version_ 1768546478893039616