Glucocorticoid receptor in circulating mononuclear leukocytes

T. Murakami, D. Brandon, D. Rodbard, Donald (Lynn) Loriaux, M. B. Lipsett

Research output: Contribution to journalArticle

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Abstract

Binding of [ 3H]dexamethasone ([ 3H]Dex) and [ 3H]cortisol to normal female goat mononuclear leukocytes (MNL) was studied at 4 and 20 C. Dissociation studies of [ 3H]Dex and [ 3H]cortisol from MNL after 3 h of incubation at 20 C indicated the presence of at least two components: a small pool with a dissociation rate constant (k(off)) of 15 ± 8 x 10 -3 min -1 for [ 3H]Dex and 35 ± 11 x 10 -3 min -1 for [ 3H]cortisol, and a large capacity pool with a k(off) of 1.0 ± 0.1 x 10 -3 min -1 for [ 3H]Dex and 11 ± 2 x 10 -3 min -1 for [ 3H]cortisol. The calculated k(off) for the slow component was indistinguishable from the k(off) for nuclei isolated from cells that had been preincubated with [ 3H]Dex or [ 3H]cortisol. Thus, for the intact cells the slow component probably represents the dissociation of an activated steroid-receptor complex transformed by a temperature-dependent process at 20 C. The rapid phase may represent the dissociation of an inactive steroid-receptor complex in the cytosol at 20 C. The K(d) obtained from equilibrium Scatchard analysis (Dex, 0.27 ± 0.02 nM; cortisol, 4.5 ± 0.1 nM) was in satisfactory agreement with the ratio of k(off) to association rate from kinetic studies. The dissociation of [ 3H]Dex and [ 3H]cortisol from MNL at 4 C can be described by a single exponential decay. The ratio of k(off) to association rate at 4 C corresponds to the K(d)s obtained using equilibrium Scatchard analysis (at 15 hr) for either intact cells (Dex, 2.0 ± 0.3 nM; cortisol, 8.0 ± 0.9 nM) or cytosols (Dex, 1.4 ± 0.4 nM; cortisol, 8.5 ± 1.9 nM). The higher affinity of Dex to the intact cell at 20 C may explain the previously noted discrepancy between the increased biological potency of Dex compared to that of cortisol and the modest increase in binding affinity of Dex to cytosol receptors.

Original languageEnglish (US)
Pages (from-to)500-505
Number of pages6
JournalEndocrinology
Volume104
Issue number2
StatePublished - 1979
Externally publishedYes

Fingerprint

Mononuclear Leukocytes
Glucocorticoid Receptors
Hydrocortisone
Cytosol
Steroid Receptors
Cellular Structures
Cell Nucleus
Goats
Dexamethasone

ASJC Scopus subject areas

  • Endocrinology
  • Endocrinology, Diabetes and Metabolism

Cite this

Murakami, T., Brandon, D., Rodbard, D., Loriaux, D. L., & Lipsett, M. B. (1979). Glucocorticoid receptor in circulating mononuclear leukocytes. Endocrinology, 104(2), 500-505.

Glucocorticoid receptor in circulating mononuclear leukocytes. / Murakami, T.; Brandon, D.; Rodbard, D.; Loriaux, Donald (Lynn); Lipsett, M. B.

In: Endocrinology, Vol. 104, No. 2, 1979, p. 500-505.

Research output: Contribution to journalArticle

Murakami, T, Brandon, D, Rodbard, D, Loriaux, DL & Lipsett, MB 1979, 'Glucocorticoid receptor in circulating mononuclear leukocytes', Endocrinology, vol. 104, no. 2, pp. 500-505.
Murakami T, Brandon D, Rodbard D, Loriaux DL, Lipsett MB. Glucocorticoid receptor in circulating mononuclear leukocytes. Endocrinology. 1979;104(2):500-505.
Murakami, T. ; Brandon, D. ; Rodbard, D. ; Loriaux, Donald (Lynn) ; Lipsett, M. B. / Glucocorticoid receptor in circulating mononuclear leukocytes. In: Endocrinology. 1979 ; Vol. 104, No. 2. pp. 500-505.
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abstract = "Binding of [ 3H]dexamethasone ([ 3H]Dex) and [ 3H]cortisol to normal female goat mononuclear leukocytes (MNL) was studied at 4 and 20 C. Dissociation studies of [ 3H]Dex and [ 3H]cortisol from MNL after 3 h of incubation at 20 C indicated the presence of at least two components: a small pool with a dissociation rate constant (k(off)) of 15 ± 8 x 10 -3 min -1 for [ 3H]Dex and 35 ± 11 x 10 -3 min -1 for [ 3H]cortisol, and a large capacity pool with a k(off) of 1.0 ± 0.1 x 10 -3 min -1 for [ 3H]Dex and 11 ± 2 x 10 -3 min -1 for [ 3H]cortisol. The calculated k(off) for the slow component was indistinguishable from the k(off) for nuclei isolated from cells that had been preincubated with [ 3H]Dex or [ 3H]cortisol. Thus, for the intact cells the slow component probably represents the dissociation of an activated steroid-receptor complex transformed by a temperature-dependent process at 20 C. The rapid phase may represent the dissociation of an inactive steroid-receptor complex in the cytosol at 20 C. The K(d) obtained from equilibrium Scatchard analysis (Dex, 0.27 ± 0.02 nM; cortisol, 4.5 ± 0.1 nM) was in satisfactory agreement with the ratio of k(off) to association rate from kinetic studies. The dissociation of [ 3H]Dex and [ 3H]cortisol from MNL at 4 C can be described by a single exponential decay. The ratio of k(off) to association rate at 4 C corresponds to the K(d)s obtained using equilibrium Scatchard analysis (at 15 hr) for either intact cells (Dex, 2.0 ± 0.3 nM; cortisol, 8.0 ± 0.9 nM) or cytosols (Dex, 1.4 ± 0.4 nM; cortisol, 8.5 ± 1.9 nM). The higher affinity of Dex to the intact cell at 20 C may explain the previously noted discrepancy between the increased biological potency of Dex compared to that of cortisol and the modest increase in binding affinity of Dex to cytosol receptors.",
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N2 - Binding of [ 3H]dexamethasone ([ 3H]Dex) and [ 3H]cortisol to normal female goat mononuclear leukocytes (MNL) was studied at 4 and 20 C. Dissociation studies of [ 3H]Dex and [ 3H]cortisol from MNL after 3 h of incubation at 20 C indicated the presence of at least two components: a small pool with a dissociation rate constant (k(off)) of 15 ± 8 x 10 -3 min -1 for [ 3H]Dex and 35 ± 11 x 10 -3 min -1 for [ 3H]cortisol, and a large capacity pool with a k(off) of 1.0 ± 0.1 x 10 -3 min -1 for [ 3H]Dex and 11 ± 2 x 10 -3 min -1 for [ 3H]cortisol. The calculated k(off) for the slow component was indistinguishable from the k(off) for nuclei isolated from cells that had been preincubated with [ 3H]Dex or [ 3H]cortisol. Thus, for the intact cells the slow component probably represents the dissociation of an activated steroid-receptor complex transformed by a temperature-dependent process at 20 C. The rapid phase may represent the dissociation of an inactive steroid-receptor complex in the cytosol at 20 C. The K(d) obtained from equilibrium Scatchard analysis (Dex, 0.27 ± 0.02 nM; cortisol, 4.5 ± 0.1 nM) was in satisfactory agreement with the ratio of k(off) to association rate from kinetic studies. The dissociation of [ 3H]Dex and [ 3H]cortisol from MNL at 4 C can be described by a single exponential decay. The ratio of k(off) to association rate at 4 C corresponds to the K(d)s obtained using equilibrium Scatchard analysis (at 15 hr) for either intact cells (Dex, 2.0 ± 0.3 nM; cortisol, 8.0 ± 0.9 nM) or cytosols (Dex, 1.4 ± 0.4 nM; cortisol, 8.5 ± 1.9 nM). The higher affinity of Dex to the intact cell at 20 C may explain the previously noted discrepancy between the increased biological potency of Dex compared to that of cortisol and the modest increase in binding affinity of Dex to cytosol receptors.

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