Isoelectric focusing analysis of detergent extracted renal 11β-hydroxysteroid dehydrogenase

Schulz, Wolfgang A.; Lichtenstein, I.; Siebe, H.; Hierholzer, Klaus

doi:10.1016/0022-4731(89)90393-2

Cited by 9 publications

(3 citation statements)

References 44 publications

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“…A separation of oxidase from reductase could not be achieved using the ELISA technique. This is in accordance with either a uniform bidirectional enzyme [27] or a dual enzyme system with an epitope common to an oxidase and a reductase.…”

Section: Discussionmentioning

confidence: 99%

Immunohistochemical Localization of 11-Hydroxysteroid Dehydrogenase in Rat Kidney with a Monoclonal Antibody

Castello¹,

Schwarting²,

Muller³

et al. 1989

Kidney Blood Press Res

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Monoclonal antibody (MAb) against 11-hydroxysteroid dehydrogenase (11-HSD) has been raised by immunization of female balb/c mice. 11-HSD from solubilized rat renal microsomal protein could be bound in a modified ELISA using antimouse IgG and MAb against 11-HSD. On Western blots of solubilized rat renal microsomes the MAb recognized a single protein band of an approximate molecular weight of 35 kD. Immunohistochemical staining of rat renal tissue with the above MAb and the APAAP staining technique displayed a heterogenous reginal and subcellular distribution: glomeruli and arterioles were practically devoid of specific staining, as were epithelial cells in inner and outer medulla. Intense immunostaining was observed in PCT and particularly in PST, appearing granular with highest density around the nuclei. Here the enzyme bound to intracellular membranes may exert an autocrine function such as signal inactivation. In contrast to cortex, staining of interstitial cells was observed in renal medulla. The latter localization is compatible with the concept of a paracrine function of 11-HSD which might prevent corticosterone from gaining access to collecting duct cells.

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Section: Discussionmentioning

confidence: 99%

Immunohistochemical Localization of 11-Hydroxysteroid Dehydrogenase in Rat Kidney with a Monoclonal Antibody

Castello¹,

Schwarting²,

Muller³

et al. 1989

Kidney Blood Press Res

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“…Incubation techniques in vitro were performed as previously described [9,10). Human kidney and liver samples were obtained from surgically removed mate rial (courtesy of the Departments of Urology + Sur gery).…”

Section: Biological Systems and General Techniquesmentioning

confidence: 99%

Metabolism of Dexamethasone: Sites and Activity in Mammalian Tissues

Siebe

Baude²,

Lichtenstein

et al. 1993

Kidney Blood Press Res

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We have used in vitro techniques to study the metabolism of dexamethasone. Tissue slices, homogenates and microsomal fractions of various mammalian organs from rats and humans have been used. We focused particularly on the question of whether or not dexamethasone (Dexa) is oxidized at the C11-OH group by 11β-hydroxysteroid-dehydrogenase. High activities of this enzyme system for Dexa were localized in renal cortex and rectum. Material from both human and murine liver was ineffective. The main metabolite formed from Dexa in renal and intestinal systems was identified by different mass-spectrometric techniques including on line HPLC mass spectrometry as 11-dehydro-dexamethasone. This finding was corroborated by the observation that both corticosterone and glycyrrhetinic acid block the metabolic transformation of Dexa.

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“…Sources o f compounds: [i-glycyrrhetinic acid, corti costerone and 11-dehydrocorticosterone were ob tained from Sigma (Dcisenhofcn, Germany), 1,2,6,7-'H-corticosterone (3.256-1015 Bq/mol) from New En gland Nuclear (Dreieich, Germany), 'H-l 1-dehydrocorticosterone was prepared and purified in our labo ratory as reported previously [26], NADVH, NADPV H, G-6-PDH and glucose-6-phosphate were purchased from Boehringer (Mannheim, Germany), CHAPS from Calbiochcm (Frankfurt, Germany).…”

Section: Methodsmentioning

confidence: 99%

Characterization of Human Placental 11β-Hydroxy steroid Dehydrogenase, a Key Enzyme of Corticosteroid Metabolism

Blum¹,

Bühler

Lichtenstein

et al. 1995

Cell Physiol Biochem

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Human placenta exhibits high levels of 11β-HSD oxidative activity. The enzymatic activity was tested in tissue slices, homogenates, microsomes and CHAPS-solubilized microsomal protein of spontaneously delivered fresh human placenta. Compared to liver and kidney the placenta exhibits the highest specific 11β-HSD activity in microsomal preparations. In contrast to liver and kidney, no strict cosubstrate dependence but some preference for NAD⁺ over NADP⁺ as cosubstrate was detectable. Conversion of corticosterone by microsomes with NAD⁺ as cofactor was 51.1 ± 3.7 ^. 10^–13 mol/(mg·min), withNADP+ 15.0 ± 0.8·10–13 mol/(mg·min). In contrast to tissue slices, subcellular fractions of placental preparations exhibited a small but significant reductive activity. A kinetic analysis of the conversion of corticosterone to 11-dehydrocorticosterone by Eadie-Hofstee plots revealed a biphasic course in the presence of NADP+ indicating the existence of isoenzymes. The k_M values were 6.3 ^. 10^–9 and 3.4 ^. 10^–8 mol/l respectively. In the presence of NAD⁺, a linear Eadie-Hofstee plot was obtained with a single k_M value of 4.4 ^. 10^–9 mol/l. Placental 11β-HSD was inhibited by β-glycyrrhetinic acid and by the steroidal detergent CHAPS. With NADP+ as cosubstrate, the IC50 values were 3.0 ^. 10^–7 and 1.2 ^. 10^-4 mol/l, with NAD⁺ 9.7 ^. 10^-8 and 5.6 ^. 10^–3 mol/l respectively. We conclude that the placenta constitutes an important barrier for 11-OH steroids during pregnancy between the maternal and fetal organism.

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Isoelectric focusing analysis of detergent extracted renal 11β-hydroxysteroid dehydrogenase

Cited by 9 publications

References 44 publications

Immunohistochemical Localization of 11-Hydroxysteroid Dehydrogenase in Rat Kidney with a Monoclonal Antibody

Immunohistochemical Localization of 11-Hydroxysteroid Dehydrogenase in Rat Kidney with a Monoclonal Antibody

Metabolism of Dexamethasone: Sites and Activity in Mammalian Tissues

Characterization of Human Placental 11β-Hydroxy steroid Dehydrogenase, a Key Enzyme of Corticosteroid Metabolism

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Isoelectric focusing analysis of detergent extracted renal 11β-hydroxysteroid dehydrogenase

Cited by 9 publications

References 44 publications

Immunohistochemical Localization of 11-Hydroxysteroid Dehydrogenase in Rat Kidney with a Monoclonal Antibody

Immunohistochemical Localization of 11-Hydroxysteroid Dehydrogenase in Rat Kidney with a Monoclonal Antibody

Metabolism of Dexamethasone: Sites and Activity in Mammalian Tissues

Characterization of Human Placental 11&beta;-Hydroxy steroid Dehydrogenase, a Key Enzyme of Corticosteroid Metabolism

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Characterization of Human Placental 11β-Hydroxy steroid Dehydrogenase, a Key Enzyme of Corticosteroid Metabolism