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European Journal of (2011) 165 293–300 ISSN 0804-4643

CLINICAL STUDY Long-term DHEA substitution in female adrenocortical failure, body composition, muscle function, and : a randomized trial Jens J Christiansen1, Jens M Bruun2, Jens S Christiansen1, Jens Otto Jørgensen1 and Claus H Gravholt1 Department of Endocrinology and Internal Medicine MEA, 1Aarhus Sygehus NBG and 2Aarhus Sygehus THG, Aarhus University Hospital, DK-8000 Aarhus C, Denmark (Correspondence should be addressed to C H Gravholt; Email: [email protected])

Abstract Context: Adrenal derived androgens are low in women with adrenal failure. The physiological consequences of substitution therapy are uncertain. Objective: To investigate the effects of DHEA substitution in women with adrenal failure on body composition, fuel metabolism, and inflammatory markers. Design, participants and intervention: In this study, ten female patients (median age 38.5 years, range 28–52) with adrenal failure were treated with DHEA 50 mg for 6 months in a double-blind, randomized, placebo-controlled, and crossover study. The participants underwent dual-energy X-ray absorptiometry (DXA) scan, computed tomography scan of abdominal fat, indirect calorimetry, bicycle ergometry, muscle and fat biopsies, and blood samples. Results: Baseline androgens were normalized to fertile range during active treatment. Anthropo- metric data were unaffected, but lean body mass (LBM) slightly increased compared with placebo (delta LBM (kg) placebo versus DHEA: K0.48G6.1 vs 1.6G3.4, PZ0.02) with no alterations in total or abdominal fat mass. PTH increased with DHEA, but no significant changes were observed in other bone markers or in bone mineral content. The mRNA levels of markers of tissue inflammation (adiponectin, interleukin 6 (IL6), IL10, monocyte chemoattractant protein 1, and tumor necrosis factor a) in fat and muscle tissue were unaffected by DHEA treatment, as was indirect calori- metry and maximal oxygen uptake. A high proportion of self-reported seborrheic side effects were recorded (60%). Conclusion: In female adrenal failure, normalization of androgens with DHEA 50 mg for 6 months had no effects on muscle, fat, and bone tissue and on fuel metabolism in this small study. A small increase in LBM was observed. Treatment was associated with a high frequency of side effects.

European Journal of Endocrinology 165 293–300

Introduction including (7–9). Questionnaires in con- trolled clinical trials have shown some (9–12) or no The , being a member of the nuclear (13–15) effects on quality of life, and a high proportion receptor superfamily, is responsible for mediating of self-reported skin related side effects (9–11, 13, 14). development of primary and secondary sex charac- Substantial physiological alterations upon androgen teristics during fetal life, childhood, and and for substitution with DHEA in female adrenocortical failure maintaining these features during adult life. The are still controversial and probably obscured by the beneficial impact of endogenous testosterone as well associated deficiencies (i.e. hypocortisolism as testosterone replacement in hypogonadal males on with or without other aspects of hypopituitarism), and it maintenance of bone (1, 2), muscle mass (2, 3), body fat remains to be determined whether DHEA has any effect distribution, and quality of life (4) is well documented. on muscle and fat distribution. Cessation of androgen secretion in the adult female Therefore, we evaluated the influence of DHEA is observed in adrenal insufficiency where adrenal substitution in female adrenal failure on muscle, fat, androgen precursor production is blunted or almost and bone tissue as well as on indicators offuel metabolism. absent (5, 6). Substitution therapy with an oral dose of We undertook biopsies on both fat and muscle, which 50 mg of the adrenal androgen DHEA is followed by were analyzed for expression of enzymes and proteins normalization of the spectrum of circulating androgens known to be involved in hormonal and inflammatory

q 2011 European Society of Endocrinology DOI: 10.1530/EJE-11-0289 Online version via www.eje-online.org

Downloaded from Bioscientifica.com at 09/25/2021 11:41:52AM via free access 294 J J Christiansen and others EUROPEAN JOURNAL OF ENDOCRINOLOGY (2011) 165 pathways, and determined muscle and fat distribution for capsules in the European Pharmacopoeia. That by dual-energy X-ray absorptiometry (DXA) scan and means uniformity of capsule content, uniformity of abdominal computed tomography (CT) scan. A daily dose capsule mass and disintegration of capsules. According of 50 mg DHEA was administered for 6 months in a to the analysis certificate the results are assay 101.5% crossover, randomized and placebo-controlled trial. (95–105%), mean weight variation 410.4G3.6 mg (401–419 mg), and disintegration 3 min (!15 min). Figures in parenthesis indicate the requirements of the Subjects and methods law. Physical examination consisted of anthropometrics, muscle strength, ergometry, indirect calorimetry, and CT In this study, ten females with adrenocortical failure scan of abdominal fat distribution. Blood samples were included. Exclusion criteria were lactation and included routine biochemistry, bone markers, sex pregnancy, neoplastic, thromboembolic, and cardiovas- , and regulatory hormones. This program cular diseases and diabetes mellitus. Two participants was performed in similar sequence before and at the end discontinued the study because of seborrheic side effects of each treatment period. Muscle and fat biopsy and and anxiety. The examined study group (Addison’s DEXA scan were done at the end of each study period. At disease, nZ6; monotrope ACTH deficiency, nZ2) had a baseline (tZ120–150 min), percutaneous muscle biop- median age of 38.5 years (range 28–52). All were on sies from the quadriceps muscle were performed after stable hydrocortisone (20–40 mg/day) and fludrocorti- injecting local anesthesia (10% lidocaine) in the skin sone (0–0.2 mg/day) treatment. One of two postmeno- and percutaneous regions. A small incision was made at pausal patients was treated with vaginal 17b- the lateral aspect of the thigh in the mid-belly of the (E2; vaginal capsules, E2 25 mg). Naturally menstruat- vastus lateralis of the quadriceps muscle. Using a ing patients used contraceptives (four intra uterine modified Bergstrom needle (5 mm), muscle tissue was devices and two combined oral contraceptives). Other obtained by suction. The biopsy was immediately put in concomitant included thyroid hormones liquid nitrogen and kept at K80 8C. Fat biopsies were (one patient) and and (two patients). taken from the subcutaneous abdominal adipose tissue None of the participants were previously treated with depot (periumbilically). Using local anesthesia (5 mg/ml DHEA or androgens. Studies were performed during the lidocaine), the biopsies were taken by needle aspiration early (days 5–10) of regular cycle or in (liposuction). The adipose tissue was washed thoroughly the corresponding phase of a tablet cycle. The protocol with isotonic saline and then frozen in liquid nitrogen for was approved by the Aarhus County ethics scientific later RNA extraction. committee (no. 2001 0130). All participants gave informed oral and written consent. The clinical trial was registered at ClinicalTrials.gov (NCT00471900). Determination of adipokine mRNA levels Cardiovascular data from this study have previously In the fat tissue biopsies, mRNA expression of adiponectin, been published (16). interleukin 6 (IL6), IL10, tumor necrosis factor a (TNFa), monocyte chemoattractant protein 1 (MCP1 or CCL2), and 11b-HSD (HSD11B1) were assessed. In accordance Study protocol with tissue specificity only mRNA expression of IL6, TNFa, The patients were treated for 6 months with a daily and MCP1 were assessed in the muscle tissue biopsies. morning dose of DHEA (50 mg) or placebo in a double- For mRNA determination the following oligonucleotide blind, randomized, placebo-controlled, and crossover primer pairs were used: adiponectin, 50-CATGACCAGGA- design study. Blinding and randomization was per- AACCACGACT-30 and 50-TGAATGCTGAGCGGTAT-30; formed by the Hospital Pharmacy in blocks of 5. IL6,50-AAATGCCAGCCTGCTGACGAAG-30 and 50-AAC- Everybody else was blinded to this procedure until the AACAATCTGAGGTGCCCATGCTAC-30; IL10,50-ATGGA- GCP unit had finalized monitoring the study and the GCGAAGGT-TAGTGGTCA-30 and 50-CTCGCTTTAA- randomization code was broken. A washout period of at TTGTCATGTATGCT-30; TNFa,50-CGAGTGACAAGCCTG- least 2 months was inserted before the next exami- TAGC-30 and 50-GGTGTGGGTGAGGAGCACAT-30; MCP1, nation period. During the treatment period, participants 50-CAATCAATGCCCCAGTCAC-30 and 50-GATTCTTGGG- continued normal daily living. The project was TTGTGGGAGTG-30; 11b-HSD,50-GACCATGACCTTCG- conducted and monitored according to the Inter- CAGAGCAATTTGT-30 and 50-GACGCCAAGAACACA- national Conference on Harmonization-Good Clinical GAGAGTGATTGA-30; b-actin, 50-ACGGGGTCACCCA- Practice guidelines (ICH-GCP; The GCP-unit, Aarhus CACTGTGC-30 and 50-CTAGAAGCATTTGCGGTGGAC- University Hospital). This study drug was produced GATG-30. RNA was isolated from w250 mg of adipose according to Good Manufacturing Practice (GMP) in a tissue using the TriZol Reagent (Invitrogen). RNA was licensed laboratory (Terapharm, Katwijk, Holland). Raw quantified by measuring absorbance at 260 and 280 nm. materials fulfilled the requirements of the European The integrity of the RNA was examined by visual Pharmacopoeia (Ph. Eur. 3rd ed.). Final analyses inspection of the two ribosomal RNAs, 18S and 28S, on document the tests corresponding to the requirements an agarose gel. RNA was reversely transcribed with RT

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Downloaded from Bioscientifica.com at 09/25/2021 11:41:52AM via free access EUROPEAN JOURNAL OF ENDOCRINOLOGY (2011) 165 DHEA and body composition 295 and random hexamer primers at 23 8C for 10 min, 42 8C Similarly, rates of protein, and glucose oxidation for 60 min, and 95 8C for 10 min according to the were estimated after correction for urinary excretion of manufacturer’s instructions (GeneAmp RNA PCR kit, urea (17). Maximal oxygen consumption (VO2max) test PerkinElmer Life and Analytical Sciences, Boston, MA, was performed on a bicycle ergometer by a standardized USA). Then 2 ml of each RT reaction was amplified in PCR protocol. The initial workload was increased with 10 W mastermix containing the specific primers, Hot Star Taq every 30 s until exhaustion. Breath-by-breath gas DNA polymerase, and SYBR-Green PCR buffer. All exchange analysis was performed and VO2maxwas samples were determined as duplicates. Real-time determined as the highest O2 consumption achieved quantification of target gene (adiponectin, IL6, IL10, during exercise with a calorimeter (Jaeger Oxycon Delta, TNFa, MCP1,and11b-HSD) relative to housekeeping Erich Jaeger, Hoechberg, Germany), and VO2maxwas gene (b-actin) was performed by a SYBR-Green real-time calculated. The isometric strength of the right biceps PCRassayand an iCycler PCR machine (Bio-Rad). In brief, brachialis and quadriceps muscles was assessed by samples were incubated in separate tubes for initial means of a dynamometer (Good strength, Metitur Ltd, denaturation at 95 8C for 10 min, followed by 40 PCR Jyva¨skyla¨, Finland), which electronically measures amplification cycles. Each cycle consisted of 30 s at 95 8C, the isometric muscle functions in the upper and lower 30 s at 57 8C, and extension for 60 s at 74 8C. During final limbs. The strength was calculated as the mean of three extension increasing fluorescence was measured in real voluntary maximum isometric contractions separated time. Data were obtained as threshold cycle values (Ct,the by 1 min intervals. cycle number at which fluorescence reached ten times the S.D. of the background fluorescence). Relative gene expression of target gene to b-actin was calculated as Assays described in the User Bulletin No. 2, 1997 from We measured DHEAS, a-4-androstendione, testoster- PerkinElmer Life and Analytical Sciences (PerkinElmer one, (DHT), and E2 by an in-house Cetus, Norwalk, CT, USA). RIA after extraction and subsequent celite chromatog- raphy (18). We estimated free testosterone (fT) by a DXA and CT scans method described by Bartsch (19), based on measure- ment of -binding globulin (SHBG), total Bone mineral content (BMC) (g) and area-adjusted bone testosterone, and DHT, using the law of mass action, the mineral density (aBMD) (g/cm2) were measured at the binding constant of testosterone and DHT to SHBG, and lumbar spine (L2–L4), the hip (femoral neck and including a calculation of testosterone binding to trochanteric region), and the nondominant forearm albumin (assuming a constant association constant to (ultradistal and proximal part of distal third) by DXA on albumin). In this system binding to -binding Hologic 1000/w or 2000/w osteodensitometers globulin is thought to be negligible. We analyzed SHBG (Hologic, Inc., Waltham, MA, USA). Cross-calibration by double-monoclonal immunofluorometric assay was ensured through the use of double measurements (AutoDelfia, Wallac, OY, Finland); intra- and inter- and a phantom. Precision for BMD was 1.5% for the assay coefficient of variation (CV) was 7.5 and 5.2% lumbar spine, 2.1% for the femoral neck, and 1.9% for respectively. Our inter- and intra-assay CV were the ultradistal forearm. These quantities included as follows: SHBG, 7.5 and 5.2%; testosterone, 13.8 crossover calibration, change in hardware, change in and 8.2%; fT, 6.4 and 4.7%; DHT, 11.0 and 9.1%; technicians, and long-term stability (!0.2%/year). The a-4-androstendione, 11.4 and 9.4%; DHEAS, 11.5 amounts of intra abdominal (visceral) and subcu- and 8.5%; E2, 10.5 and 7.4%. Plasma intact PTH was taneousfat,aswellastotalabdominalareawas measured by a chemiluminescence assay using an determined by CT with a Somatom Plus-S scanner. automated instrument (Immulite, DPC, Los Angeles, The subjects were studied in the supine position. The CA, USA). N-terminal cross-linking telopeptide of area scanned comprised of a 10 mm cross-sectional slice type I collagen (NTX) was measured by an immuno- at the umbilicus, using 120 kV and 330 mA. The same metric assay using a Vitros ECI analyzer (Ortho Clinical technician performed all the scans, which subsequently Products, Amersham Pharmacia Biotech). This assay were analyzed blind by the same radiologist. uses MAbs against human NTX (20). Plasma osteo- calcin (total OC) was measured using the N-Mid-OC Indirect calorimetry, muscle strength, assay on an automated analyzer (Elecsys 2010 and maximal oxygen uptake analyzer, Roche Diagnostics) with antibodies that determine both intact OC and the large N-Mid-terminal Indirect calorimetry (Deltatrac metabolic monitor; fragment. The CVtotal was 4–7% for the various Datex, Helsinki, Finland) was performed for 30 min in automated assays. Plasma bone-specific alkaline phos- basal state. The initial 5 min were used for acclimatiz- phatase (bone ALP) was measured after lectin precipi- ation, and calculations of respiratory exchange ratio tation using a Hitachi 917 automated analyzer (Roche and resting energy expenditure (EE) were based on Diagnostics) (21). This assay was performed with a mean values of 25 measurements of 1 min each. CVtotal of 8%. Plasma concentrations of procollagen I

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Table 1 Serum values (median, range) of androgens and androgen precursor metabolites in female adrenocortical failure before and after 6 months treatment with 50 mg DHEA or placebo.

Placebo DHEA Normal range (fertile) Start End Start End P value

DHEA (nmol/l) 3.8–15.9 1.74 (1.57–2.59) 2.25 (1–3.92) 2.11 (0.47–3.97) 10.28 (5.13–17.17) 0.012 DHEAS (nmol/l) 1200–9500 99 (99–240) 99 (99–210) 99 (99–230) 8450 (1000–19 000) 0.012 Androstendione (nmol/l) 2.4–8.9 1.9 (0.27–3.91) 2.1 (0.21–3.75) 2.0 (0.18–5.15) 5.1 (1.89–9.77) 0.017 DHT (nmol/l) 0.25–1.2 0.31 (0.09–0.45) 0.27 (0.09–0.35) 0.28 (0.09–0.45) 0.83 (0.38–2.02) 0.012 Testosterone (nmol/l) 0.55–1.8 0.55 (0.2–0.71) 0.52 (0.18–0.96) 0.49 (0.16–1.5) 1.30 (0.69–2.65) 0.012 Free testosterone (nmol/l) 0.006–0.034 0.01 (0–0.01) 0.01 (0–0.01) 0.00 (0–0.02) 0.02 (0.01–0.03) 0.018 E2 (pmol/l) !40–400 155 (39–540) 195 (39–1100) 290 (39–630) 220 (39–630) 0.3 SHBG (nmol/l) 41–170 88 (27–114) 96 (37–156) 78 (36–163) 79 (26–144) 0.2

N-terminal propeptide (PINP) (22), procollagen III carryover effects, which did not affect the level of N-terminal propeptide (PIIINP) (23), and C-terminal significance. Analysis was done on delta values (‘end cross-linking telopeptide of type I collagen (ICTP) (24) period value’ minus ‘start period value’). P!0.05 was were determined by commercial RIA kits (Orion considered statistically significant. Diagnostica, Espoo, Finland). Intra- and inter-assay CV of 5 and 7% respectively, were observed. As indices of the IGF axis, serum IGF1 and IGF2 were measured by noncompetitive time-resolved immunofluorometric Results assays (TR-IFMA) (25), serum IGFBP2 was measured Levels of DHEA, DHEAS, a-4-androstendione, testoster- by RIA, and IGFBP3 was measured by an immuno- one, and fT rose significantly to medium normal fertile radiometric assay (Diagnostics Systems Laboratories, range during active treatment compared with baseline. Inc., Webster, TX, USA). was determined by No significant alterations were detected in E and SHBG a commercial ELISA (Dako, Glostrup, Denmark). Leptin 2 values (Table 1). was determined by a commercial RIA (Linco, St Louis, Anthropometric data (weight (kg) 71.3G4.3; BMI MO, USA). Cortisol was measured by an automated (kg/m2) 25.7G1.7; waist–hip ratio 0.81G0.01) did not chemiluminescence system (Chiron Diagnostics, change significantly during the treatment period. Lean Fernwald, Germany). Free fatty acids (FFAs) were body mass (LBM) increased during active treatment determined by a colorimetric method using a commer- (delta LBM (kg) placebo versus DHEA: K0.48G6.1 vs cial kit (Wako Chemicals, Neuss, Germany). Total 1.6G3.4, PZ0.02). There was no change in total fat concentrations of thyroxine (T ) and tri-iodothyronine 4 mass (delta TFM (kg), placebo versus DHEA: K1.2G0.6 (T ) were measured in serum as described previously 3 vs 0.6G0.9, PZ0.1). We observed no change in (26), whereas TSH was measured in a solid-phase, two- abdominal fat distribution measured by CT scan in site chemiluminescent enzyme immunometric assay total abdominal area, subcutaneous fat area or intra- (Immulite; Diagnostic Products Corp., Los Angeles, CA, abdominal fat area (Fig. 1). USA). We measured serum total adiponectin by a EE and estimated oxidation rates of glucose (PZ0.4), novel in-house TR-IFMA. Urea excretion in urine was lipid (PZ0.6), and protein (PZ0.6), as well as the determined by an indophenol method and serum urea respiratory quotient did not change during the study by a commercial kit (COBAS INTEGRA; Roche). (Fig. 2). No changes were observed in thyroid hormones,

Safety parameters CT abdominal area, transversal section view Total abdominal area 800 Standard blood count, enzymes, electrolytes, and P = 0.5 renal parameters were measured four times during each 600 Subcutaneous fat treatment period and analyzed with routine methods in 2 400 P = 0.6 hospital central laboratory. There were no significant Cm Intra-abdominal fat changes and no remarkable alterations. 200 P = 1.0 0 Statistical analysis Start End Start End Start End Start End Start End Start End All statistical calculations were carried out using SPSS Placebo DHEA Placebo DHEA Placebo DHEA for Windows version 13.0 (SPSS, Inc., Chicago, IL, Figure 1 Intra abdominal and subcutaneous fat tissue area USA). Data were examined by Student’s two-tailed estimated by computed tomography at umbilical level in transversal unpaired t-test or the Mann–Whitney U test, where section view, in female adrenocortical failure before and after appropriate. All data were tested for period as well as 6 months treatment with 50 mg DHEA/placebo.

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Indirect calorimetry PZ0.6) and right leg knee extension (362G26 to 375G26 vs 346G26 to 385G20 Nm, PZ0.5) and 1600 P = 0.5 VO2 max ((ml/kg per min): 31.1G2.5 to 30.9G3.0 vs 1400 32.7G2.0 to 31.3G2.2, PZ0.5) did not change during 1200 the study. Six patients experienced side effects during active 1000 treatment only and one patient reported mild facial 800 greasiness after 4 days of placebo treatment. Side effects

kcal/24 h 600 were self reported and were not evaluated by physical examination. Two patients reported symptoms from the 400 scalp region after 64 and 15 days of treatment. 200 Symptoms were described as ‘painful spots’ and ‘an unpleasant crawling sensation’ respectively. Both 0 Start End Start End patients left the study. Three patients reported facial Placebo DHEA greasiness after 29, 36, and 46 days of active treatment and one patient experienced regain of axils and pubic Figure 2 Resting energy expenditure and estimated fractions of hair after 161 days of active treatment. glucose (black), lipid (bright), and protein (gray) oxidation from indirect calorimetry in female adrenocortical failure before and after 6 months treatment with 50 mg DHEA/placebo. Discussion IGF1, IGF2, IGFBP3, GHPB, FFAs, leptin or adiponectin (Table 2). Insulin levels did not respond differently to In this randomized placebo-controlled trial, 6 months of DHEA treatment and substituted cortisol levels were DHEA substitution in women with adrenocortical stabile throughout the study (data not shown). insufficiency was associated with a complete bio- chemical normalization of circulating androgen levels. Total and regional BMC did not increase in response The only significant effect was a small gain in LBM of to DHEA treatment (Fig. 3). A slight, but significant 1.6 kg. This occurred without concomitant changes in increase in PTH was detected during DHEA compared body weight or fat mass. All other examined variables with placebo treatment (Fig. 4); however, this change were essentially unchanged including both subcu- was not followed by any changes in levels of calcium or taneous and intra-abdominal fat mass determined by ALP or indicators of resorptive and formative bone cell CT scan, and we found no alterations in fuel metabolism activity (Fig. 4). or in the expression of mRNA of a host of inflammatory The mRNA expression of the investigated inflam- markers. matory markers (adiponectin, IL6, IL10, MCP1, and DHEA is regarded an androgen precursor with no TNFa) in subcutaneous abdominal fat and in skeletal affinity for the androgen receptor but with in vitro effects muscle did not change significantly in response to on several membrane-associated receptors (27), includ- DHEA treatment, nor did the expression of 11b-HSD ing direct binding to receptors in the growth mRNA (Table 3). plate (28). The downstream conversion of DHEA to Muscle strength of right arm elbow flexion (placebo testosterone and DHT, as well as aromatization to start to end versus DHEA start to end; 182G5 to 189 estrogen, may take place inside target cells. A local G9 vs 188G8 to 189G6 Newton meters (Nm), degradation of the receptor-bound hormone would then

Table 2 Serum values (mean GS.E.M.) of hormones, fat tissue markers and free fatty acids in female adrenocortical failure before and after 6 months treatment with 50 mg DHEA or placebo.

Placebo DHEA

Start End Start End P value

Cortisol (nmol/l) 640G77 463G142 475G89 492G142 0.3 Insulin (pmol/l) 97G45 81G16 47G754G6 0.7 Leptin (ng/ml) 17.0G2.2 13.5G2.1 14.5G2.8 15.0G2.2 0.2 T3 (nmol/l) 1.7G0.1 1.5G0.1 1.6G0.1 1.7G0.1 0.1 T4 (nmol/l) 93G689G789G791G9 0.6 TSH (mU/ml) 2.0G0.6 2.3G0.6 2.7G1.0 2.6G0.5 0.9 GHBP (nmol/l) 2.4G0.2 2.7G0.3 2.5G0.3 2.2G0.3 0.2 IGF1 (mg/l) 212G26 201G17 212G27 213G15 0.5 IGF2 (mg/l) 796G50 800G46 751G40 798G56 0.1 IGFBP3 (mg/l) 4853G261 4986G320 4823G370 5039G402 0.7 FFA (mmol/l) 0.38G0.05 0.40G0.10 0.48G0.07 0.35G0.07 0.2 Adiponectin (mg/l) 14.9G2.3 14.0G2.4 15.8G2.9 14.9G2.6 0.9

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∆BMC spine ∆BMC hip increase in LBM, but without concomitant changes in 2 any other parameters as mentioned above. However, P = 0.2 P = 0.4 LBM and other DXA-derived measures are determined 1 with a precision of about 2%. The observed difference in LBM could not be replicated in relation to body weight 0 (determined with a precision (or imprecision) of up to 3 kg (about 5%)), or supported by any other measures of Gram –1 body composition, anthropometrics, serum samples or fat and muscle samples, why we rather consider it a –2 chance finding. Placebo DHEA Placebo DHEA –3 We detected a high proportion (60%) of self-reported side effects during active treatment. All reports related Figure 3 Changes in bone mineral content (BMC) in spine and hip in female patients with adrenocortical failure before and after 6 months ABAlk Phos (U/l) µ treatment with 50 mg DHEA/placebo (expressed as delta changes). ( g/l) 30 40 25 obscure a sex hormone action from serological 30 detection, hence the term ‘intracrinology’ has been 20 introduced (29). Effects of DHEA treatment in adreno- 15 20 P = 0.6 10 P = 0.3 cortical failure beyond 12 months have not been 10 evaluated, and in agreement with our findings 1 year 5 of DHEA treatment increased total LBM by 1.8%, with 0 0 the increase being observed in females only (14). In the Placebo DHEA Placebo DHEA same study an increase in BMD at the femoral neck was observed only in DHEA treated patients. On the CDPTH (pmol/l) Calcium (mmol/l) contrary, no significant alterations were observed on 8 DXA scans of body composition and BMD in 39 male 7 1.2 and female patients after 3 months (10) and in 38 6 P = 0.035 1.0 P = 0.2 female hypopituitary women after 6 months (11). Like 5 in this study, no changes in circulating bone markers 4 0.8 have been reported in either mixed female and male 3 0.6 populations (10) or in female patients only (11, 13, 30). 2 In this study, we also evaluated muscle function, Placebo DHEA Placebo DHEA determined as muscle strength, and found absolutely no change in this measure, and likewise we did not EFITCP (µg/l) PIIINP (µg/l) detect any change in maximal oxygen uptake, another 6 4 measure which reflects the size of the muscle mass. 5 3 In spite of some evidence of minor LBM accumulation 4 in response to DHEA substitution, we (31) and others 3 P = 0.2 2 P = 0.2 (32) have not found any alterations in forearm and 2 1 whole body amino acid turnover in short-term trials 1 (1 and 2 weeks). In accordance with our previous short- 0 0 term study on GH secretory patterns (8), we did not find Placebo DHEA Placebo DHEA any augmentation of GH derived growth factors or their binding proteins. Slightly stimulated IGF1 levels are GHPICP (µg/l) PINP (µg/l) reported in some (9, 14, 15) but not all studies (8, 12, 160 13, 33). In a 3 months study with DHEA or placebo 60 treatment (50 mg) to a group of middle aged female 120 40 Addison patients insulin sensitivity increased (33), 80 P = 0.4 P = 0.8 while others found unchanged insulin sensitivity (11, 20 31). In this study, we did not find any evidence of altered 40 fuel metabolism by indirect calorimetry or any changes 0 0 in pertinent hormones and metabolites. As expected, Placebo DHEA Placebo DHEA the surrogate markers of adipose tissue mass, adipo- nectin and leptin, were unaffected by DHEA treatment. Figure 4 Serum levels of markers of bone metabolism (PTH and calcium), bone resorption (PINP, PIIINP, ITCP, and PICP), and In line with these findings, mRNA expression of specific bone formation (alkaline phosphatase and osteocalcin) in female inflammatory markers in both fat tissue and skeletal patients with adrenocortical failure before and after 6 months muscle remained unchanged. We did find a slight treatment with 50 mg DHEA or placebo.

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Table 3 mRNA expression (relative gene expression of target gene Funding to b-actin) of adipokines in subcutaneous abdominal fat and in muscle tissue in female adrenocortical failure after 6 months J J Christiansen was supported by a PhD research fellowship by the treatment with placebo or DHEA. University of Aarhus. This study was supported by a grant from the Danish Health Research Council, grant number 9600822. Placebo DHEA P value

Fat Acknowledgements G G TNFa 0.007 0.002 0.008 0.003 0.8 We thank Elsebeth Hornemann, Hanne Pedersen, Lone Svendsen, and G G MCP1 0.002 0.001 0.006 0.003 0.2 Bente Mortensen for expert technical help. IL6 0.023G0.010 0.032G0.018 0.5 IL10 0.008G0.002 0.019G0.009 0.2 Adiponectin 10.0G3.6 10.0G2.5 1.0 11b-HSD 0.308G0.128 0.497G0.209 0.2 Muscle References TNFa 0.028G0.023 0.008G0.002 0.4 MCP1 0.006G0.005 0.001G0.000 0.3 1 Finkelstein JS, Klibanski A, Neer RM, Greenspan SL, Rosenthal DI IL6 0.130G0.105 0.028G0.006 0.4 & Crowley WF Jr. Osteoporosis in men with idiopathic hypogona- dotropic . Annals of Internal Medicine 1987 106 354–361. 2 Katznelson L, Finkelstein JS, Schoenfeld DA, Rosenthal DI, to skin problems and probably reflect stimulation of the Anderson EJ & Klibanski A. Increase in bone density and lean body mass during testosterone administration in men with pilosebaceous unit. Our findings are consistent with the acquired hypogonadism. Journal of Clinical Endocrinology and frequency reported in previous trials (9–14) and it may Metabolism 1996 81 4358–4365. (doi:10.1210/jc.81.12.4358) be concluded that the skin is the most DHEA sensitive 3 Sheffield-Moore M. Androgens and the control of organ. However, attempts to evaluate this phenomenon protein synthesis. Annals of Medicine 2000 32 181–186. (doi:10. 3109/07853890008998825) with sebutape (14) and score (11) have failed. 4 Davidson JM, Camargo CA & Smith ER. Effects of androgen The design of the present pilot study, however, has on sexual behavior in hypogonadal men. Journal of Clinical limitations. Only eight patients completed the trial, Endocrinology and Metabolism 1979 48 955–958. (doi:10.1210/ which introduces a risk for a type 2 error. It is also jcem-48-6-955) possible that longer term duration of treatment would 5 Christiansen JJ, Fisker S, Gravholt CH, Bennett P, Svenstrup B, Andersen M, Feldt-Rasmussen U, Christiansen JS & Jorgensen JO. show (positive) changes in body composition, although Discontinuation of estrogen replacement therapy in GH-treated we doubt this, due to the almost complete absence of hypopituitary women alters androgen status and IGF-I. European positive effects after 6 months treatment. In addition, Journal of Endocrinology 2005 152 719–726. (doi:10.1530/eje.1. the expected effect size of the measured variables may 01898) 6 Miller KK, Sesmilo G, Schiller A, Schoenfeld D, Burton S & also preclude definite conclusions. However, under Klibanski A. Androgen deficiency in women with hypopituitarism. similar conditions using small study samples, significant Journal of Clinical Endocrinology and Metabolism 2001 86 561–567. effects on body composition has been found. For (doi:10.1210/jc.86.2.561) instance, using the same methodology in our own 7 Young J, Couzinet B, Nahoul K, Brailly S, Chanson P, Baulieu EE & laboratory, we observed profound changes in body Schaison G. Panhypopituitarism as a model to study the Z metabolism of (DHEA) in humans. composition in females with (n 12) Journal of Clinical Endocrinology and Metabolism 1997 82 treated with GH or placebo after only 2 months of 2578–2585. (doi:10.1210/jc.82.8.2578) treatment (34). In addition, a study of postmenopausal 8 Christiansen JJ, Gravholt CH, Fisker S, Svenstrup B, Bennett P, women treated with a weak androgen () ten Veldhuis J, Andersen M, Christiansen JS & Jorgensen JO. Dehydroepiandrosterone supplementation in women with adrenal females increased their muscle mass and decreased fat failure: impact on twenty-four hour GH secretion and IGF-related mass in comparison with ten females receiving placebo parameters. Clinical Endocrinology 2004 60 461–469. (doi:10. and ten females receiving an (spirono- 1111/j.1365-2265.2004.01998.x) lactone) (35). Thus, exclusion of a physiological effect of 9 Arlt W, Callies F, van Vlijmen JC, Koehler I, Reincke M, DHEA may not be possible based on the present data, Bidlingmaier M, Huebler D, Oettel M, Ernst M, Schulte HM & Allolio B. Dehydroepiandrosterone replacement in women with but we may be able to conclude that we could not find adrenal insufficiency [see comments]. New England Journal of any physiologically relevant effect of DHEA treatment in Medicine 1999 341 1013–1020. (doi:10.1056/NEJM19990930 hypoadrenal women. 3411401) In conclusion, the present evaluation of DHEA 10 Hunt PJ, Gurnell EM, Huppert FA, Richards C, Prevost AT, Wass JAH, Herbert J & Chatterjee VK. Improvement in mood and treatment in female adrenocortical failure does not fatigue after dehydroepiandrosterone replacement in Addison’s document convincing anabolic effects and does not disease in a randomized, double blind trial. Journal of Clinical contribute to a scientific justification for this treatment Endocrinology and Metabolism 2000 85 4650–4656. (doi:10. modality. 1210/jc.85.12.4650) 11 Johannsson G, Burman P, Wiren L, Engstrom BE, Nilsson AG, Ottosson M, Jonsson B, Bengtsson BA & Karlsson FA. Low dose Declaration of interest dehydroepiandrosterone affects behavior in hypopituitary andro- gen-deficient women: a placebo-controlled trial. Journal of Clinical The authors declare that there is no conflict of interest that could be Endocrinology and Metabolism 2002 87 2046–2052. (doi:10. perceived as prejudicing the impartiality of the research reported. 1210/jc.87.5.2046)

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12 Brooke AM, Kalingag LA, Miraki-Moud F, Camacho-Hubner C, 24 Risteli J, Elomaa I, Niemi S, Novamo A & Risteli L. Radioimmuno- Maher KT, Walker DM, Hinson JP & Monson JP. Dehydroepian- assay for the pyridinoline cross-linked carboxy-terminal telopep- drosterone improves psychological well-being in male and female tide of type I collagen: a new serum marker of bone collagen hypopituitary patients on maintenance replace- degradation. Clinical Chemistry 1993 39 635–640. ment. Journal of Clinical Endocrinology and Metabolism 2006 91 25 Frystyk J, Dinesen B & Orskov H. Non-competitive time-resolved 3773–3779. (doi:10.1210/jc.2006-0316) immunofluorometric assays for determination of human insulin- 13 Lovas K, Gebre-Medhin G, Trovik TS, Fougner KJ, Uhlving S, like growth factor I and II. Growth Regulation 1995 5 169–176. Nedrebo BG, Myking OL, Kampe O & Husebye ES. Replacement of 26 Weeke J, Boye N & Orskov H. Ultrafiltration method for direct dehydroepiandrosterone in adrenal failure: no benefit for sub- radioimmunoassay measurement of free thyroxine and free jective health status and sexuality in a 9-month, randomized, tri-iodothyronine in serum. Scandinavian Journal of Clinical and parallel group clinical trial. Journal of Clinical Endocrinology and Laboratory Investigation 1986 46 381–389. (doi:10.3109/ Metabolism 2003 88 1112–1118. (doi:10.1210/jc.2002- 00365518609083686) 020769) 27 Webb SJ, Geoghegan TE, Prough RA & Michael Miller KK. The 14 Gurnell EM, Hunt PJ, Curran SE, Conway CL, Pullenayegum EM, biological actions of dehydroepiandrosterone involves multiple Huppert FA, Compston JE, Herbert J & Chatterjee VK. Long-term receptors. Drug Metabolism Reviews 2006 38 89–116. (doi:10. DHEA replacement in primary adrenal insufficiency: a random- ized, controlled trial. Journal of Clinical Endocrinology and 1080/03602530600569877) Metabolism 2008 93 400–409. (doi:10.1210/jc.2007-1134) 28 Sun H, Zang W, Zhou B, Xu L & Wu S. DHEA suppresses 15 van Thiel SW, Romijn JA, Pereira AM, Biermasz NR, Roelfsema F, longitudinal bone growth by acting directly at growth plate van Hemert A, Ballieux B & Smit JW. Effects of dehydroepian- through estrogen receptors. Endocrinology 2011 152 1423. drostenedione, superimposed on growth hormone substitution, (doi:10.1210/en.2010-0920) on quality of life and insulin-like growth factor I in patients 29 Labrie F. Intracrinology. Molecular and Cellular Endocrinology 1991 with secondary adrenal insufficiency: a randomized, placebo- 78 C113–C118. (doi:10.1016/0303-7207(91)90116-A) controlled, cross-over trial. Journal of Clinical Endocrinology and 30 Callies F, Fassnacht M, van Vlijmen JC, Koehler I, Huebler D, Metabolism 2005 90 3295–3303. (doi:10.1210/jc.2004-1802) Seibel MJ, Arlt W & Allolio B. Dehydroepiandrosterone replace- 16 Christiansen JJ, Andersen NH, Sorensen KE, Pedersen EM, ment in women with adrenal insufficiency: effects on body Bennett P, Andersen M, Christiansen JS, Jorgensen JO & composition, serum leptin, bone turnover, and exercise capacity. Gravholt CH. Dehydroepiandrosterone substitution in female Journal of Clinical Endocrinology and Metabolism 2001 86 adrenal failure: no impact on endothelial function and cardio- 1968–1972. (doi:10.1210/jc.86.5.1968) vascular parameters despite normalization of androgen status. 31 Christiansen JJ, Gravholt CH, Fisker S, Moller N, Andersen M, Clinical Endocrinology 2007 66 426–433. (doi:10.1111/j.1365- Svenstrup B, Bennett P, Ivarsen P, Christiansen JS & Jorgensen JO. 2265.2007.02750.x) Very short term dehydroepiandrosterone treatment in female 17 Frayn KN. Calculation of substrate oxidation rates in vivo adrenal failure: impact on carbohydrate, lipid and protein from gaseous exchange. Journal of Applied Physiology 1983 55 metabolism. European Journal of Endocrinology 2005 152 77–85. 628–634. (doi:10.1530/eje.1.01810) 18 Lykkesfeldt G, Bennett P, Lykkesfeldt AE, Micic S, Moller S & 32 Burt MG, Johannsson G, Umpleby AM, Chisholm DJ & Ho KK. Svenstrup B. Abnormal androgen and oestrogen metabolism in Impact of growth hormone and dehydroepiandrosterone on men with sulphatase deficiency and recessive X-linked protein metabolism in -treated patients. Journal of ichthyosis. Clinical Endocrinology 1985 23 385–393. (doi:10. Clinical Endocrinology and Metabolism 2008 93 688–695. (doi:10. 1111/j.1365-2265.1985.tb01096.x) 1210/jc.2007-2333) 19 Bartsch W. Interrelationships between sex hormone-binding 33 Dhatariya K, Bigelow ML & Nair KS. Effect of dehydroepiandros- globulin and testosterone, 5 alpha-dihydrotestosterone and terone replacement on insulin sensitivity and in hypoadrenal oestradiol-17 beta in blood of normal men. Maturitas 1980 2 women. Diabetes 2005 54 765–769. (doi:10.2337/diabetes.54.3. 109–118. (doi:10.1016/0378-5122(80)90044-4) 20 Hanson DA, Weis MA, Bollen AM, Maslan SL, Singer FR & 765) Eyre DR. A specific immunoassay for monitoring human 34 Gravholt CH, Naeraa RW, Brixen K, Kastrup KW, Mosekilde L, bone resorption: quantitation of type I collagen cross-linked Jørgensen JO & Christiansen JS. Short term growth hormone N-telopeptides in urine. Journal of Bone and Mineral Research 1992 treatment in girls with Turner syndrome decreases fat mass and 7 1251–1258. (doi:10.1002/jbmr.5650071119) insulin sensitivity. A randomized double-blind, placebo-controlled 21 Rosalki SB, Foo AY, Burlina A, Prellwitz W, Stieber P, Neumeier D, cross-over study. Pediatrics 2002 110 889–896. (doi:10.1542/ Klein G, Poppe WA & Bodenmuller H. Multicenter evaluation of peds.110.5.889) Iso-ALP test kit for measurement of bone alkaline phosphatase 35 Lovejoy JC, Bray GA, Bourgeois MO, Macchiavelli R, Rood JC, activity in serum and plasma. Clinical Chemistry 1993 39 Greeson C & Partington C. Exogenous androgens influence body 648–652. composition and regional body fat distribution in obese post- 22 Melkko J, Kauppila S, Niemi S, Risteli L, Haukipuro K, Jukkola A & menopausal women – a clinical research center study. Journal of Risteli J. Immunoassay for intact amino-terminal propeptide of Clinical Endocrinology and Metabolism 1996 81 2198–2203. human type I procollagen. Clinical Chemistry 1996 42 947–954. (doi:10.1210/jc.81.6.2198) 23 Risteli J, Sogaard H, Oikarinen A, Risteli L, Karvonen J & Zachariae H. Aminoterminal propeptide of type III procollagen in methotrexate-induced liver fibrosis and cirrhosis. British Journal of Dermatology 1988 119 321–325. (doi:10.1111/ Received 20 May 2011 j.1365-2133.1988.tb03224.x) Accepted 23 May 2011

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