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Endocrinology Vol. 139, No. 9 3849-3854
Copyright © 1998 by The Endocrine Society

ARTICLES

Furosemide Inhibits 11ß-Hydroxysteroid Dehydrogenase Type 21

Daniel Fuster, Geneviève Escher, Bruno Vogt, Daniel Ackermann, Bernhard Dick, Brigitte M. Frey and Felix J. Frey

Division of Nephrology and Hypertension, Department of Medicine, University Hospital of Berne, Inselspital, 3010 Berne, Switzerland

Address all correspondence and requests for reprints to: Felix J. Frey, M.D., Division of Nephrology, Freiburgstrasse 3, Inselspital, 3010 Berne, Switzerland. E-mail: felix.frey{at}insel.ch


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
11ß-Hydroxsteroid dehydrogenase 2 (11ß-OHSD2) protects the nonselective renal mineralocorticoid receptor from the endogenous glucocorticoid cortisol. Thus, drugs inhibiting 11ß-OHSD2 might enhance urinary loss of potassium. As diuretics influence the renal handling of potassium, we analyzed the impact of 13 commonly used diuretics on 11ß-OHSD2.

Furosemide was the only inhibitor. Its inhibition constant (Ki) was 30 µmol when extracts from COS-1 cells transfected with human 11ß-OHSD2 were used as an enzyme source. The type of inhibition was competitive. To establish whether furosemide inhibits 11ß-OHSD2 and 11ß-OHSD1 in the renal target tissue, isolated tubular segments from rats were analyzed. Furosemide decreased the oxidative activity of 11ß-OHSD2 in intact distal tubules and 11ß-OHSD1 in proximal convoluted tubules. For the assessment of furosemide on the excretion of corticosterone metabolites in vivo, rats were given furosemide ip, and the ratio of tetrahydrocorticosterone plus 5{alpha}-tetrahydrocorticosterone to 11-dehydrotetrahydrocorticosterone was determined in urine. This ratio increased after the administration of furosemide in all animals, indicating inhibition of the oxidative activity of 11ß-OHSD. Thus, furosemide inhibits the 11ß-OHSD2 enzyme in the target tissue and might by that mechanism enhance the mineralocorticoid effect of 11ß-hydroxyglucocorticoids.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
11ß-HYDROXYSTEROID dehydrogenase (11ß-OHSD) enzymes catalyze the interconversion of endogenous or exogenous biologically active 11ß-hydroxy-glucocorticosteroids (cortisol, corticosterone, or prednisolone) and inactive keto-glucocorticosteroids (cortisone, 11-dehydrocorticosterone, or prednisone) (1, 2, 3, 4). Two isoenzymes accounting for 11ß-OHSD activity have been cloned and characterized: 11ß-OHSD1 is NADP(H) dependent and catalyzes both the oxidation and the reduction reactions (5), whereas 11ß-OHSD2 requires NAD as a cofactor and exhibits only oxidative activity (6). 11ß-OHSD1 is expressed in a wide variety of tissues (5, 7, 8). Its biological function is only partly defined. There is evidence that the activity of 11ß-OHSD1 determines the antiinflammatory effect of 11ß-hydroxyglucocorticosteroids and the access of these agents to glucocorticoid receptors (1, 9, 10, 11). The biological role of 11ß-OHSD2 is most likely to provide selective access of aldosterone to the mineralocorticoid receptor by inactivating cortisol (1, 2, 12, 13). The absence of 11ß-OHSD2 results in apparent mineralocorticoid excess, with hypertension and hypokalemia. Because specific inactivation of cortisol is relevant only in distal tubular cells of the kidney, salivary glands, and colon (the target cells of aldosterone), 11ß-OHSD2 is almost exclusively expressed in this subset of cells (1, 14).

Several endo- and xenobiotics have been found to modulate the activity of 11ß-OHSD (15, 16, 17, 18, 19, 20, 21). We have previously shown that furosemide inhibits 11ß-OHSD activity, as assessed by using kidney and liver microsomal enzyme preparations or extracts from COS-1 cells transfected with 11ß-OHSD1 (22). Furthermore, the effect of furosemide was assessed by determining the concentration ratio of prednisolone to its 11-ketometabolite prednisone in vivo (22). Based on these results, it was speculated that the urinary loss of potassium in patients treated with furosemide might at least partly be attributable to the activation of mineralocorticoid receptors by glucocorticoids. In the meantime, it became clear that 11ß-OHSD1 did not fulfill the prerequisites for the protection of the mineralocorticoid receptor, and subsequently, the relevant candidate 11ß-OHSD isozyme, 11ß-OHSD2, was cloned (1, 6). Therefore, the main purpose of the present investigation was to establish whether furosemide inhibits 11ß-OHSD2. Furthermore, we attempted to determine whether furosemide inhibits 11ß-OHSD activity at the site of action, i.e. in the distal tubule, using isolated segments to assess the activity of 11ß-OHSD.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Supplies
Corticosterone, 11-dehydrocorticosterone, furosemide, and the tests substances mentioned below were purchased from Sigma Chemical Co. (Buchs, Switzerland). NADP+/NADPH, NAD+/NADH, and restriction enzymes were obtained from Boehringer Mannheim (Mannheim, Germany). Collagenase was a product of Serva (Heidelberg, Germany). Bicinchoninic acid protein assay reagent (BCA) was received from Pierce Chemical Co. (Rockford, IL). TLC plates (60 F 254) coated with silica gel were obtained from Merck (Rahway, NJ). DMEM, penicillin G (100,000 U/liter), and streptomycin sulfate (10,000 µg/liter) were obtained from Life Technologies (Basel, Switzerland). FCS was obtained from Biological Industries (Beit Haemek, Israel). Tissue culture plates (100 mm) and 24-well plates were purchased from Becton Dickinson Labware (Basel, Switzerland). [1,2,6,7-3H]Corticosterone (SA, 83 Ci/mmol) was purchased from Amersham (Aylesbury, UK). 5{alpha}-Androstane-3{alpha},17{alpha}-diol, stigmasterol, and cholesteryl butyrate were purchased from Steraloids (Wilton, NH).

Test substances
The following diuretics were assessed with respect to their inhibitory capacity on 11ß-OHSD: acetazolamide (10/100 µg/ml; 45/450 µM), amiloride hydrochloride (10/100 ng/ml; 0.04/0.4 µM), bendroflumethiazide (0.1/1 µg/ml; 0.24/2.4 µM), buthiazide (12/120 ng/ml; 0.035/0.35 µM), chlorthalidone (8/80 µg/ml; 23.6/236 µM), furosemide (6/60 µg/ml; 18.2/180 µM), hydrochlorothiazide (0.26/2.6 µg/ml; 0.87/8.7 µM), indapamide (0.14/1.4 µg/ml; 0.38/3.8 µM), piroxicam (2/20 µg/ml; 6.04/60.4 µM), spironolactone (0.2/2 µg/ml; 0.48/4.8 µM), torasemide (2.7/27 µg/ml; 7.75/77.5 µM), and triamterene (4/40 µg/ml; 15.8/158 µM).

Transfection
COS-1 cells were cultured in DMEM supplemented with 10% FCS at 37 C in a CO2 incubator. Transfection of cells with the complementary DNA (cDNA) of rat 11ß-OHSD1 and human 11ß-OHSD2 in pcDNA3 was performed using the diethylaminoethyl-dextran method (22). Forty-eight hours after transfection, cells were either harvested and extracted with the appropriate buffer (see below) or incubated in situ for 1 h with the indicated drug concentrations in the presence of 10 µM (for 11ß-OHSD1) or 10 nM (for 11ß-OHSD2) corticosterone and 100 respectively, 50 nCi [3H]corticosterone. After sonication, protein determination was performed using the BCA reagent. For measuring the activity of 11ß-OHSD, approximately 10 µg total protein from transfected cells was used to obtain a conversion rate of 30–50%.

Assay for 11ß-OHSD1
The assay was performed as previously described by Monder et al. (24). Oxidation or reduction at C-11 was determined by measuring the rate of conversion of corticosterone to 11-dehydrocorticosterone in the presence of NADP or the rate of conversion of dehydrocorticosterone to corticosterone in the presence of NADPH. Transfected COS-1 cells with the cDNA of 11ß-OHSD1 were extracted with 10 mM Tris-HCl (pH 7.5), 5 mM EDTA (pH 8), 1% Triton X-100, 2 mM phenylmethylsulfonylfluroide, and 10 µg total protein were used for the reaction. The assay was performed in 0.25 mM NADP, 100 mM Tris (pH 8.3), 100 nCi [3H]corticosterone, and 5 µM corticosterone in the presence or absence of the mentioned drug. Samples were incubated for 1 h at 37 C, the reaction was stopped on ice, and steroids were extracted with 1 vol ethyl acetate. The organic layer was separated by centrifugation at 13,000 rpm and evaporated under a stream of nitrogen. The steroid residue was dissolved in 20 µl methanol containing a mixture of 20 µg each of unlabeled corticosterone and dehydrocorticosterone. This was quantitatively transferred to thin layer plates and developed in chloroform-methanol (90:10, vol/vol). The spots corresponding to the steroids were located under a UV lamp, cut out, transferred to scintillation vials, and counted in scintillation fluid in a Tricarb 2000 CA (Canberra Packard, Zurich, Switzerland) fluid scintillation counter. Specific activity was expressed as micromoles of product formed per µg of protein/h.

For activity measurements in single nephron segments, the assay was performed in a final volume of 10 µl, incubated for 1 h at 37 C. To the supernatant, 20 µg each of unlabeled corticosterone and dehydrocorticosterone were added, and the mixture was transferred directly onto the TLC plate without prior ethyl acetate extraction.

Assay for 11ß-OHSD2
The assay was performed as previously described by Albiston et al. (6). COS-1 cells transfected with the cDNA of 11ß-OHSD2 were homogenized in a buffer containing 250 mM sucrose and 10 mM Tris-HCl, pH 7.5. Ten micrograms of protein extract were incubated for 1 h at 37 C with 1 mM NAD, 10 nM corticosterone, and 50 nCi [3H]corticosterone in 500 µl homogenization buffer in the presence or absence of the above-mentioned drugs. The subsequent steps were the same as those described for 11ß-OHSD1.

For activity measurements in single nephron segments, the assay was performed with the same buffers and incubation conditions, but in a volume of 10 µl.

Experimental animals
The protocol was approved by the ethics committee at our institution. Male Wistar rats, weighing 200–230 g, were kept in a temperature-, humidity-, and light (12-h light, 12-h dark cycle)-controlled room and maintained on a normal chow diet without fluid restriction. On the morning of the experiment, rats were separated into four groups of four animals each. The animals received ip furosemide (400 mg/kg), torasemide (20 mg/kg), glycyrrhetinic acid (400 mg/kg), or the solvent dimethylsulfoxide.

Preparation of tubular segments
One hour after administration of furosemide, rats were anesthetized with 10 mg/kg pentobarbital and perfused via the aorta with an ice-cold perfusion solution (containing 137 mM NaCl, 5 mM KCl, 0.8 mM MgSO4, 0.33 mM Na2HPO4, 0.44 mM KH2PO4, 1 mM MgCl2, 1 mM CaCl2, 5 mM glucose, and 10 mM Tris-HCl, pH 7.4) followed by perfusion with a collagenase solution (similar solution containing, in addition, 10 mg/ml collagenase) (23, 24, 25). At the end of the perfusion, the kidney was removed, cut into thin pyramid pieces, and incubated at 30 C for 15 min in a perfusion solution containing 5 mg/ml collagenase and 0.1% BSA. The corresponding solutions without collagenase were used during microdissection under a stereoscope as previously described (26, 27). Tubular length was measured using a millimeter scale placed under the microdissection dish. The same concentrations of furosemide were added into the solutions for the dissection of the kidneys from the animals treated with furosemide to minimize the loss of furosemide from the tubule during manipulation procedures. Tubular integrity was assessed using the dye exclusion method (28).

For activity measurements of 11ß-OHSD1 and 11ß-OHSD2 in single nephron segments, the assay was performed with the same buffers and incubation conditions as those mentioned above, but in a volume of 10 µl. Briefly, 2.5 mm of proximal convoluted tubule (PCT) were incubated for 2 h for measurement of 11ß-OHSD1 activity, and 1 mm of cortical collecting duct (CCT) was incubated for 30 min for measurement of 11ß-OHSD2 activity. In preliminary experiments lengths of tubules and incubation time were chosen to obtain a conversion rate of the steroidal substrates of about 25%.

Effect of furosemide on urinary steroid metabolites in rats
Animals were kept in metabolic cages for urine collection. On the morning of the experiment, rats were separated and administered ip furosemide, torasemide, glycyrrhetinic acid, or dimethylsulfoxide. For the assessment of steroid metabolite excretion in urine, gas chromatography was performed according to the method of Shackelton (29, 30). Briefly, the analytical procedure consisted of hydrolysis, solid phase extraction, derivatization, and purification by gel filtration. For each measurement, 2.5 ml urine mixed with 0.5 ml acetate buffer (0.5 M) were hydrolyzed at 55 C for 3 h with Sigma type I powdered Helix promatia enzyme (12 mg) and 0.0125 ml ß-glucuronidase/arylsulfatase (Boehringer Mannheim) liquid enzyme. After extraction on a Sep-Pak cartridge (Waters Corp., Milford, MA) and adding the internal standards (5{alpha}-androstane-3{alpha},17{alpha}-diol, stigmasterol, and cholesteryl butyrate), the samples were derivatized to form the methyloxime-trimethylsilyl ethers. After purification by gel filtration on Lipidex-5000 columns, the samples were analyzed on a Shimadzu GC-9A with an integrator Chromatopac C-R2A (Shimadzu Corp., Kyoto, Japan). A temperature programed run of 210–270 C was used over a 40-min period. Using this method, the following metabolites were determined in urine: tetrahydrocorticosterone (THB), 5{alpha}-tetrahydrocorticosterone (5{alpha}-THB), and 11-dehydrotetrahydrocorticosterone (THA).

Analysis of data
The effect of furosemide on 11ß-OHSD2 was assessed by analyzing the concentration-response curves and Lineweaver-Burk linear transformations of the Michaelis-Menten equation. The Ki was calculated by unweighted linear regression analysis with the mean values of at least three experiments. Results are given as the mean ± SEM.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
11ß-OHSD2 was subcloned into the expression vector pcDNA3 and transfected into COS-1 cells, a cell line that does not express 11ß-OHSD enzymes. Enzyme preparations from these transfected cells were incubated with the following 13 diuretics at therapeutic and 10-fold higher concentrations: acetazolamide, amiloride bendroflumethiazide, butizide, chlorthalidone, furosemide, hydrochlorothiazide, indapamide, piretanide, piroxicam, spironolactone, torasemide, and triamterene. Only furosemide had an inhibitory effect. A concentration-response curve for furosemide is given in Fig. 1Go. The concentrations of furosemide required to exhibit half-maximal inhibition of 11ß-OHSD2 were 30–40 µM. A Ki value of similar magnitude was recently described for 11ß-OHSD1 (22). Double reciprocal plots of the concentration-dependent oxidation of corticosterone in the presence of increasing concentrations of furosemide are depicted in Fig. 2Go. The data converged on the ordinate in a pattern consistent with competitive inhibition.



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  		Figure 1. Concentration-response curve of furosemide vs. percent inhibition of 11ß-OHSD2. Human 11ß-OHSD2 from transfected COS cells was used as an enzyme source.

 


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  		Figure 2. Lineweaver-Burk plot showing competitive inhibition of 11ß-OHSD2 by furosemide. The rate (V) was defined as percentage of substrate oxidized per µg protein/h. Linearized curves are given for the following furosemide concentrations: open circles, no furosemide; closed circles, 15 µM/liter; and closed squares, 120 µM/liter. r2 > 0.95.

 
Furosemide was incubated with intact COS cells transfected with 11ß-OHSD2 (Fig. 3Go, upper panel). Whereas a strong inhibition of the activity was observed when enzyme extracts were used, the inhibition was much less pronounced in the intact cells. A similar difference was observed between the effect of furosemide in intact cells and enzyme extracts when 11ß-OHSD1 was considered (Fig. 4Go, upper panel). To exclude that this difference between the inhibitory effect of furosemide on the two types of enzyme source was restricted to furosemide, glycyrrhetinic acid was added to intact cells and enzyme extracts (Figs. 3Go and 4Go, lower part). A similar difference between the dose-response curve in intact cells and cell extracts was found with this model compound for the inhibition for the 11ß-OHSD enzymes.



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  		Figure 3. Inhibition of human 11ß-OHSD2 by furosemide and glycyrrhetinic acid in transfected COS-1 cells. Furosemide and glycyrrhetinic acid were incubated in situ (black columns) and with cell extracts (hatched columns). Each column represents the mean (±SE) of three experiments. The inhibition was more pronounced in cell extracts than in intact cells.

 


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  		Figure 4. Inhibition of human 11ß-OHSD1 dehydrogenase activity by furosemide and glycyrrhetinic acid in transfected COS-1 cells. Furosemide and glycyrrhetinic acid were incubated in situ (black columns) and with cell extracts (hatched columns). Each column represents the mean (±SE) of three experiments. The inhibition was more pronounced in cell extracts than in intact cells.

 
The impact of furosemide on the activity of 11ß-OHSD was assessed in rat tubular segments. 11ß-OHSD1 oxidation activity is present in PCT (23). When PCT segments of rats with or without pretreatment with furosemide were incubated with furosemide, a dose-dependent inhibition of 11ß-OHSD1 dehydrogenase activity was observed (Fig. 5Go). CCT segments express a high activity of 11ß-OHSD2 (23). In these segments furosemide at high concentrations inhibited the dehydrogenase activity (Fig. 5Go). When CCT were incubated with high concentrations of piretanide or torasemide, two structurally different loop diuretics, no inhibition of 11ß-OHSD2 was observed (results not shown).



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  		Figure 5. Effect of furosemide on 11ß-OHSD1 dehydrogenase activity in isolated PCT and on 11ß-OHSD2 dehydrogenase activity in isolated CCT from rats. Rats pretreated or not with furosemide were used for the isolation of tubules. The tubules were incubated without (black columns) or with 0.12 mM (11ß-OHSD1) and 0.012 mM (11ß-OHSD2; hatched columns) or 1.2 mM (11ß-OHSD1) and 0.12 mM (11ß-OHSD2; open columns) furosemide. Each column represents the mean (±SE) of three experiments.

 
The influence of a dose of furosemide on the urinary ratio of (THB plus 5{alpha}-THB)/THA was analyzed. Furosemide increased this ratio in all four rats investigated (Fig. 6Go). When another loop diuretic, torasemide (20 mg/kg), was given ip, no changes in the urinary ratio of (THB+5{alpha}-THB)/THA were observed, indicating that the furosemide dose-associated changes in the pattern of steroid metabolites excreted in urine were not due to the enhanced diuresis induced by the diuretic. As a positive control, four rats given 400 mg/kg glycyrrhetinic acid, ip, showed a significant increase in the ratio of (THB plus 5{alpha}-THB)/THA in all animals (mean ±SD values at baseline, 1.22 ± 0.11; 24 h after treatement, 1.75 ± 0.13).



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  		Figure 6. Effect of ip furosemide on the urinary ratio of corticosterone metabolites (THB plus 5{alpha}-THB)/THA in rats. Each symbol represents one rat. Urine was collected before the administration of furosemide (BASELINE) and during two collecting periods thereafter.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
Recently, we have shown that the inhibition of the 11ß-OHSD by glycyrrhetinic acid increases the concentration ratio of 11ß-hydroxy- to 11-keto-glucocorticosteroids in renal tissue (4). A similar effect was observed for furosemide, indicating that furosemide inhibits 11ß-OHSD activity in vivo (22). An inhibitory effect of furosemide, but not of other diuretics, was recently shown when rat microsomal preparations from testis or kidneys were used as a source of enzymes (31). In the present investigation we defined the isoforms of 11ß-OHSD inhibited by furosemide in the mineralocorticoid target tissue. Furosemide is a potent inhibitor of 11ß-OHSD2 when cell extracts from transfected COS-1 cells or from constitutively 11ß-OHSD2 expressing LLC-PK1 cells (results not shown) are used as an enzyme source. The concentrations of glycyrrhetinic acid required to induce half-maximal inhibition of the dehydrogenase reaction of 11ß-OHSD2 (~10 nM) in cell extracts from transfected COS cells was in the same range as that found by Albiston et al. (6) using CHOP-transfected cells as a source of enzymes (~50 nM). When furosemide was added to intact transfected cells, the inhibition of 11ß-OHSD2 was less pronounced. A similar discrepancy between inhibition of 11ß-OHSD obtained from microsomal preparations and intact cells was previously described by Monder et al. (32). These researchers observed a 75% inhibition in the presence of 20 nM glycyrrhetinic acid when microsomal preparations were used and a 60% inhibition in the presence of 100 µM of glycyrrhetinic acid when intact cells were used. (32). On the other hand, Bujalska et al. (33) observed a significant inhibition of 11ß-OHSD2 at much lower concentrations of glycyrrhetinic acid (100–1000 nM) in intact cells when fetal kidney 293 cells transfected with 11ß-OHSD2 were analyzed. These researchers did not analyze the impact of glycyrrhetinic acid on extracts of 11ß-OHSD enzymes from the transfected 293 cells.

With respect to inhibition of 11ß-OHSD isoenzymes, we have screened a number of diuretics other than furosemide, including acetazolamide, amiloride hydrochloride, bendroflumethiazide, butizide, chlorthalidone, indapamide, piroxicam, spironolactone, torasemide, piretanide, thiazides, and triamterene. None of these diuretics inhibited 11ß-OHSD enzymes (results not shown), indicating that the inhibitory effect of furosemide is a particular feature of this loop diuretic. The inhibition of 11ß-OHSD1 in PCT and that of 11ß-OHSD2 in CCT by furosemide are consistent with a direct effect of this diuretic in the kidney tubule. The renal effect of furosemide is furthermore supported by the changes in the urinary ratio of (THB plus 5{alpha}-THB)/THA, an observation consistent with inhibition of 11ß-OHSD dehydrogenase activity. The impact of furosemide on the urinary excretion of glucocorticoid metabolites is not a nonspecific consequence of the enhanced diuresis, because rats given another loop diuretic (torasemide) did not exhibit a similar change in the urinary excretion of the steroid metabolites.

The role of 11ß-OHSD2 inhibition in cortical collecting tubules by furosemide remains speculative. The kaliuretic effect of furosemide is attributed to the inhibition of the Na+/K+/2Cl cotransporter in the thick ascending limb of Henle (34), resulting in an increased delivery of potassium to the distal nephron. Microperfusion studies showed no conclusive direct effect of furosemide in the distal nephron (35). Interestingly, these researchers observed an increasing potassium concentration in the furosemide perfusions not completely explained by the slightly higher potassium concentration in the furosemide perfusion solution, suggesting tubular secretion of potassium. Distal tubular secretion of potassium is enhanced by mineralocorticoids. Glucocorticosteroids such as cortisol act as mineralocorticoids whenever the enzyme 11ß-OHSD2 is inhibited. As the concentrations of furosemide required to show a significant inhibition of 11ß-OHSD2 were of the same magnitude as those found after a moderate dose of iv or oral furosemide in humans and the concentrations of furosemide increase along the kidney tubule, furosemide might enhance the urinary excretion of potassium (36, 37, 38).

Hypokalemia from furosemide is traditionally attributed to 1) increased delivery of sodium to the distal tubule, so that a larger fraction of sodium is available for exchange with potassium; 2) a shift of potassium into the cells because of alkalosis; and 3) secondary hyperaldosteronism due to volume depletion (39, 40). Inhibition of 11ß-OHSD might be a fourth mechanism to account for the increased urinary loss of potassium in subjects treated with furosemide. Indeed, the apparent absence of the activity of 11ß-OHSD or its inhibition by glycyrrhetinic acid leads to the activation of mineralocorticoid receptors by cortisol with urinary loss of potassium (1, 2, 41, 42). Such an activation of the mineralocorticoid receptors, however, should cause sodium retention and hypertension, an effect apparently overcome by the potent inhibition of the Na+/K+/2Cl cotransporter in the thick ascending limb of the loop of Henle. Alternatively, glucocorticoids might induce kaliuresis without concomitant sodium retention, as recently shown in patients with Addison’s disease (43). The latter effect might not be mediated through mineralocorticoid receptors.

One might speculate that some of the hitherto poorly understood side-effects of furosemide, such as insulin resistance or increased lipid levels, are attributable to increased access of cortisol to glucocorticoid receptors in the presence of furosemide (44). This mechanism could, however, account only for part of these furosemide-related unwanted effects, as other diuretics, such as thiazides, do not inhibit 11ß-OHSD, but also induce insulin resistance and high lipid concentrations. 11ß-OHSD2 is expressed in placental tissue (1). Evidence is growing that it plays a pivotal role in fetal physiology by excluding maternal glucocorticoids from the fetal circulation. Thus, inhibition of the 11ß-OHSD in hypertensive pregnant women by furosemide might be hazardous, a hypothesis in line with the fact that many authorities discourage the use of furosemide during pregnancy (45).


   Acknowledgments
 
We acknowledge Rita Häberli for secretarial work.


   Footnotes
 
1 This work was supported by Grant 3200-50820.97 from the Swiss National Foundation for Scientific Research. Back

Received March 4, 1998.


   References
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 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

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