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425 , , and sulphate inhibit in female but not in male samples of omental adipose : lack of effect of

V Pin˜ eiro1, X Casabiell1, R Peino´ 1, M Lage1, J P Camin˜a1, C Menendez1, J Baltar2, C Dieguez3 and F F Casanueva1 1Endocrine Section, Department of , Santiago de Compostela University, Complejo Hospitalano´ de Santiago, Santiago de Compostela, Spain 2Division of General Surgery, Hospital de Conxo, Complejo Hospitalario de Santiago, Santiago de Compostela, Spain 3Department of Physiology, Santiago de Compostela University, Santiago de Compostela, Spain (Requests for offprints should be addressed to F F Casanueva, PO Box 563, E-15780, Santiago de Compostela, Spain)

Abstract Leptin, the product of the Ob , is a polypeptide secretion in either gender (4856&366 in women, expressed in adipocytes which acts as a signalling 3322&505 in men). Coincubation of factor from the adipose tissue to the central nervous with dihydrotestosterone (DHT) induced a significant system, regulating food intake and energy expenditure. It (P<0·05) leptin decrease in samples taken from women has been reported that circulating leptin levels are higher (3119&322) but not in those taken from men in women than in men, even after correction for body fat. (2042&430). Stanozolol, a non-aromatizable , This gender-based difference may be conditioned by decreased (P<0·05) leptin secretion in female samples differences in the levels of androgenic . (2809&383) but not in male (1553&671). Dehydro- To explore this possibility, a systematic in vitro study sulphate (DHEA-S) induced a significant with organ cultures from human omental adipose tissue, (P<0·01) leptin decrease in female samples (2996&473), either stimulated or not with (1 µM), was with no modifications in samples derived from males undertaken in samples obtained from surgery on 44 (1596&528). Exposure to androstenedione also resulted non-obese donors (21 women and 23 men). The assay was in a significant reduction (P<0·01) of leptin secretion in standardized in periods of 24 h, ending at 96 h, with no samples taken from women (2231&264), with no effect apparent tissue damage. Leptin results are expressed as on male adipose tissue (1605&544). the mean&... of the integrated secretion into the In conclusion, DHT, stanozolol, DHEA-S and andro- medium, expressed as ng leptin/g tissue per 48 h. stenedione induced a significant inhibition of in vitro leptin Spontaneous leptin secretion in samples from female secretion in samples from female donors, without affecting donors (4149&301) was significantly higher (P<0·01) the secretion in samples from men. Testosterone was than that from male donors (2456&428). Testosterone devoid of activity in either gender. did not exert any significant effect on in vitro leptin Journal of Endocrinology (1999) 160, 425–432

Introduction of secretion (Carro et al. 1997), gonadal function and gestation (Ahima et al. 1997, Butte et al. Leptin, the 167-amino acid product of the Ob 1997, Garcia-Mayor et al. 1997, Mantzoros et al. 1997, gene (Zhang et al. 1994) is produced in adipose tissue Pombo et al. 1997) and possibly also placental function (Lonnqvist et al. 1995, Masuzaki et al. 1995). After its (Sen˜aris et al. 1997). Furthermore, the presence of leptin in release into the bloodstream, leptin acts by signalling both placental tissue (Sen˜aris et al. 1997) and human milk the amount of body fat stores to hypothalamic centres (Casabiell et al. 1997) would suggest further unexpected regulating appetite and energy expenditure (Ahima functions for this hormone. et al. 1996, Caro et al. 1996). Besides its main role in It is widely assumed that circulating leptin levels are and in the neuroendocrine adaptation to determined by the size of the body fat stores (Maffei et al. fasting (Ahima et al. 1996), the participation of leptin in 1995, Havel et al. 1996). However, the large variations in new functions has been described, such as in the regulation leptin values for any given proportion of body fat suggest

Journal of Endocrinology (1999) 160, 425–432  1999 Society for Endocrinology Printed in Great Britain 0022–0795/99/0160–425 Online version via http://www.endocrinology.org

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that other hormonal and nutritional factors may contribute under a humidified atmosphere of 95% air–5% CO2. After to its regulation (Caro et al. 1996). In this sense, serum a preincubation period of 1 h the media were discarded and leptin concentrations have been shown to be lower in men 2·5 ml fresh medium (with or without stimuli) were than in women with similar amounts of body fat (Maffei dispensed into each well. Culture media were then et al. 1995, Caprio et al. 1996, Considine et al. 1996, collected completely every 24 h and replaced with fresh Hassink et al. 1996, Havel et al. 1996, Ostlund et al. 1996, medium, again with or without stimuli. The drugs tested Rosenbaum et al. 1996), a fact that has already been were testosterone, dihydrotestosterone (DHT), stanozolol, described at birth, when no differences in the adipose dehydroepiandrosterone sulphate (DHEA-S) and andro- tissue reserves are expected (Matsuda et al. 1997, Tome stenedione, all at 1 µM, with the appropriate vehicle added et al. 1997). to control samples; this dose was selected as the most When considering the gender-based differences in appropriate for the organ culture system used (Casabiell leptin secretion, testosterone and other androgen et al. 1998). Unless specifically indicated, all drugs and have been postulated as the putative mediators. Several reagents were from Sigma Chemical Co. (St Louis, MO, indirect pieces of evidence obtained in vivo suggest that USA). testosterone normalized the abnormally elevated leptin For each tested variable, either untreated or treated levels in hypogonadal men ( Jockenhovel et al. 1997, Sih sample, the adipose tissue from a given subject was et al. 1997), while other studies failed to find any sig- independently incubated in triplicate, and the medium nificant relationship between leptin concentrations and was collected and analysed every 24 h to obtain: the 24 h androgenic hormones (Haffner et al. 1997). In peripubertal secretion, the cumulative secretion till 96 h, and the boys, leptin rises in parallel with body fat until the time integrated secretion (area under the curve, AUC), in such of the initial testosterone rise, when a sudden leptin a way that each subject acted as her/his own control. reduction occurs (Garcia-Mayor et al. 1997). As the Samples were stored at "20 )C until assayed for leptin. majority of these reports are based on indirect and infer- Serum leptin levels were measured in duplicate by ential evidence, the exact role of androgens in leptin RIA using commercial kits (Human Leptin RIA, Linco secretion is still an open question. The aims of the present Research Inc., St Charles, MO, USA). The limit of work were twofold: to study the effect of androgens on sensitivity was 0·5 µg/l, the intraassay coefficient of vari- leptin secretion by using an in vitro system of human ation was 8·3%, and the interassay coefficient of variation omental adipose tissue organ culture; and to assess any was 6·2%. gender-based difference in the action of androgens. This work was presented at the 80th Annual Meeting of Statistical analysis The Endocrine Society, New Orleans, LA, USA, 1998. The mean BMI, defined as the weight in kilograms divided by the square of the height in metres was Materials and Methods calculated. Data are presented as the mean&...ofthe group. Leptin secretion was expressed as the total amount Omental adipose tissue was obtained from 44 non-obese of leptin secreted into the well by a given sample (in patients during elective abdominal surgery; malignancy nanograms) divided by the amount of fat tissue of the was an exclusion criterion. The tissue donor group was sample and the time considered, i.e. ng leptin/g tissue composed of 21 women (age 56·8&4·0 years, body mass per 48 h (Barr et al. 1997, Casabiell et al. 1998). The index (BMI) 27·2&1·09) and 23 men (57·0&3·6 years, integrated AUC was calculated by the trapezoidal method. BMI 26·8&0·76). Patients were taking no drugs or Comparisons between groups were made by the paired antibiotics. The study was approved by the hospital and unpaired t-tests when appropriate. Values of P<0·05 ethical committee, and each participating subject provided were considered significant. informed consent. Excised adipose tissue was immediately transported to Results the laboratory in ice-cold Krebs–Ringer–Hepes buffer (NaCl, 125 mM; KCl, 5 mM; MgSO4, 1·2 mM; CaCl2, Spontaneous (non-stimulated) leptin secretion into the 2 mM; KH2PO4, 1·2 mM; , 6 mM; Hepes, incubation medium was well maintained in samples from 25 mM; pH 7·4). After removing blood vessels and con- both men and women, from 0 to 96 h (Fig. 1). No tissue nective tissue, adipose tissue was washed with sterile damage was observed even in longer incubation periods Krebs–Ringer–Hepes and cut into small pieces with sharp (Casabiell et al. 1998). As there was a progressive decrease scissors. Tissue fragments were placed in six-well dishes in the spontaneous leptin secretion after 96 h, no longer (300–400 mg adipose tissue/well) containing 2·5 ml times were studied. The leptin secretion in periods of 24 h Dulbecco’s modified Eagle’s medium plus 0·5% fetal calf reached its maximum net increment at the 48 h point, serum, supplemented with (100 U/ml) and afterwards the leptin secretion reached a plateau state; for streptomycin sulphate (100 µg/ml), and incubated at 37 )C this reason the integrated secretion was analysed in 48 h

Journal of Endocrinology (1999) 160, 425–432

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Figure 1 Mean&S.E.M. of leptin secretion into the incubation medium from non-stimulated omental adipose tissue samples of 21 women and 23 men. Tissue samples from each subject were incubated in triplicate and values are presented either as the secretion for each 24 h incubation period (lower panel), the cumulative release into the medium (upper panel, line graph), or the integrated leptin secretion expressed as the AUC at 48 h (upper panel, bar graph). ††P<0·01 vs secretion from female samples in the same period. •, women; ●, men. periods as previously described (Hardie et al. 1996, Barr et al. 1997, Casabiell et al. 1998, Halleux et al. 1998). Analysis of leptin secretion over longer periods (96 h), did Figure 2 Mean&S.E.M. leptin secretion into the incubation medium not qualitatively change the results obtained by the after treatment with 1 ìM testosterone, expressed as the standard 48 h periods (data not shown). Spontaneous leptin increment over the respective control values for cumulative secretion at both 24 and 48 h was significantly higher secretion (lines) or for each 24 h incubation period (bars); women (P<0·01) in women than in men (Fig. 1, lower panel). n=15, men n=16. Analysed as the integrated secretion (AUC as ng leptin/g tissue per 48 h, Fig. 1, bars in upper panel), leptin secretion in female samples (n=21, 4149&301) was sig- was observed in female, but not in male samples (Fig. 3). nificantly higher (P<0·01) than that of male samples The time-period at which maximal reduction in leptin (n=23, 2456&428). occurred was in women at the 24 h and in men at the 72 h The addition of testosterone (1 µM) in the incubation period. Examined as ng leptin/g tissue per 48 h (Fig. 7), medium throughout the 96 h of incubation, induced a the spontaneous leptin release from tissue fragments moderate non-significant increase in the rate of leptin from female donors (n=16, 4253&320) was significantly secretion into the medium of samples taken from both decreased by DHT (3119&322, P<0·05), while it was men and women (Fig. 2). This non-significant enhance- not affected in samples from male donors (n=15, ment was quite consistent at any moment studied and 2188&405 in controls vs 2042&430 in samples treated when analysed as integrated secretion in 48 h (see later with DHT). Fig. 7), the secretion of leptin in samples treated with When the adipose tissue fragments were incubated testosterone was higher (P<0·05) in female (n=15, throughout the 96 h period in the presence of the non- 4856&366) than in male samples (n=16, 3322&505), aromatizable androgen stanozolol (1 µM), the spontaneous but not different from the respective control values. leptin secretion in samples from male donors was un- On the contrary, when the adipose tissue was incubated affected, while a clear decrease was observed in female with DHT (1 µM), a clear-cut decrease in leptin secretion samples (Fig. 4). The inhibitory action of stanozolol on

Journal of Endocrinology (1999) 160, 425–432

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Figure 4 Mean&S.E.M. leptin secretion into the incubation medium Figure 3 Mean&S.E.M. leptin secretion into the incubation medium after treatment with 1 ìM stanozolol, expressed as the increment after treatment with 1 µM DHT, expressed as the increment over over the respective control values for cumulative secretion (lines) the respective control values for cumulative secretion (lines) or for or for each 24 h incubation periods (bars); women n=10, men each 24 h incubation period (bars); women n=16, men n=15. n=7. **P<0·01 vs control secretion in the same period. *P<0·05 vs control secretion in the same period. 24 h incubation periods were considered, a significant female samples was evident at the 24 h period (P<0·01). difference vs control samples was observed for all periods As integrated secretion (Fig. 7), in samples taken from (P<0·01 at 24 and 48 h, P<0·05 at 72 and 96 h) in adipose women (n=10), the spontaneous secretion (3906&511) tissue from female donors, while no action was observed was inhibited by stanozolol (2809&383, P<0·05), but was in male samples. The integrated leptin secretion after the not affected in those taken from men (n=7, 1254&425 administration of 1 µM androstenedione (Fig. 7) was control vs 1553&671 stanozolol). significantly lower (P<0·01, 2231&264) than secretion in As Fig. 5 shows, DHEA-S induced a decrease in leptin the control incubations (4321&420) in women’s samples release on tissue samples taken from women (P<0·05 at (n=10). In male samples androstenedione was devoid of the 24 h period and P<0·01 at 48 h), while in samples action, the spontaneous secretion (n=10; 1670&510) not from men there was a weak inhibition which was only being altered by the (1605&544). statistically significant at the 72 h point (P<0·05). The It appears that adrenal androgens, such as DHEA-S and spontaneous release of leptin in female samples androstenedione, were more potent inhibitors of leptin (4321&420) (Fig. 7) was significantly (P<0·01) inhibited secretion than either DHT or stanozolol. by DHEA-S (n=10, 2996&473), leaving again unaffected the secretion in male samples (n=10, 1670&510 in controls vs 1596&528 in samples with DHEA-S). Discussion Incubation of adipose tissue fragments with 1 µM androstenedione (Fig. 6) induced a clear-cut inhibition of It is undisputed that the net amount of body fat is the leptin secretion in samples from females. In fact, when the main determinant of leptin concentrations in serum (Caro

Journal of Endocrinology (1999) 160, 425–432

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Figure 5 Mean&S.E.M. leptin secretion into the incubation medium after treatment with 1 ìM DHEA-S, expressed as the increment over the respective control values for cumulative secretion (lines) Figure 6 Mean&S.E.M. leptin secretion into the incubation medium or for each 24 h incubation period (bars); women n=10, men after treatment with 1 ìM androstenedione, expressed as the n=10. **P<0·01 or *P<0·05 vs control secretion in the same increment over the respective control values for cumulative period. secretion (lines) or for each 24 h incubation period (bars); women n=10, men n=10. **P<0·01 or *P<0·05 vs control secretion in the same period. et al. 1996, Ferron et al. 1997, Leal-Cerro et al. 1998). However, the wide variability in leptin values from evidence (Haffner et al. 1997, Jockenhovel et al. 1997, individuals with similar amounts of fat mass, and the lack Matsuda et al. 1997, Sih et al. 1997, Tome et al. 1997). of stoichiometry between fat mass and leptin values, However controversial, the main support for a negative suggest that in addition to the number and size of role of testosterone on leptin comes from the studies in adipocytes, other factors could operate by modulating the hypogonadal men who present inappropriately high leptin release of leptin. As it is now accepted that women have levels that are normalized after testosterone replacement higher serum leptin levels than men, even after adjusting therapy ( Jockenhovel et al. 1997, Sih et al. 1997). Never- for adiposity (Hassink et al. 1996, Ostlund et al. 1996, theless, in most of these studies the steroid treatment Rosenbaum et al. 1996, Tome et al. 1997), sex steroids induced changes in body composition that may per se have been considered as potential modulators of leptin explain the leptin changes. Interestingly, when shorter release. In fact, oestradiol is an in vitro releaser of leptin treatment schedules were used to prevent variations in with a similar to that of (Casabiell body composition, testosterone was devoid of relevant et al. 1998), and this may well be the background for the action over circulating leptin levels in both hypogonadal gender-based differences in serum leptin concentrations. and eugonadal men (V Popovic and F F Casanueva, It is not known whether, besides oestrogens, other sex unpublished observations). steroids may contribute to leptin regulation in humans; To further clarify this topic, in the present work the testosterone, for example, has been suggested to exert a action of different androgenic compounds has been sys- negative regulation over leptin secretion based on indirect tematically studied in an adipose tissue organ culture

Journal of Endocrinology (1999) 160, 425–432

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Although subcutaneous fat is the main contributor to circulating leptin levels, the participation of the abdominal omental adipose tissue to the amount of circulating leptin is qualitatively similar to that of the subcutaneous fat (Masuzaki et al. 1995, Hardie et al. 1996, Halleux et al. 1998), the former being easier to prepare and manipulate due to its relative lack of fibrous and non-adipose com- ponents. Furthermore, in vitro leptin secretion has been followed for more than 96 h with no morphological or biochemical signs of tissue damage, and well-preserved leptin secretion. The good response to glucocorticoids and oestrogens plus the inhibition by forskolin (Casabiell et al. 1998), and the here reported inhibition by androgenic compounds, further support the functionality of the assay. On the other hand, no histological evidence of gender- based differences in adipocyte size or number between women and men’s samples was found (Casabiell et al. 1998), ruling out differences in secretion rate as an explanation for the findings observed. In this study, steroids were employed at doses slightly over their habitual use in dispersed cells in order to compensate for the reduced accessibility to the inner cells of organ culture (Wabitsch et al. 1997). The first observation of the present work was the confirmation, with different samples from different patients, of previous work (Casabiell et al. 1998) reporting that in vivo female adipose tissue samples secreted more leptin than those from male donors, a finding that may be the biological basis of the gender differences reported in vivo for leptin secretion. Interestingly enough, testoster- one was devoid of inhibitory action on leptin secretion both in female and in male samples, showing only non-significant mild stimulatory activity. In female samples of human omental adipose tissue, both DHT, the active of testosterone, and the non-aromatizable androgen stanozolol were clear inhibi- tors of leptin secretion. It is possible that the tendency of testosterone to increase leptin levels may be the final result of two antagonistic actions, i.e. an inhibitory one mediated by its conversion to DHT, plus the stimulatory one exerted by its previous to oestradiol (Gray et al. 1979). In fact, oestradiol is as potent as gluco- Figure 7 Mean&S.E.M. values of the integrated leptin secretion (ng leptin/g tissue per 48 h) in samples from human omental adipose corticoids for stimulating leptin secretion in vivo and in vitro tissue. Upper panel: results for adipose tissue obtained from (Casabiell et al. 1998). It is worthy of note that the two women. Lower panel: results for samples taken from men. adrenal androgens tested, i.e. androstenedione and *P<0·05 or **P<0·01 vs control values. DHEA-S, were the strongest inhibitors of leptin secretion in samples from female donors. In fact, while DHT and stanozolol have inhibitory actions only in the first 48 h system previously validated (Casabiell et al. 1998). This after their addition to the culture medium, the inhibitory method allows for the precise evaluation of hormonal action of the adrenal androgens was evident at all periods action directly on the adipocytes without the confusing of incubation analysed until the 96 h point. This suggests variables of an in vivo system. This organ culture has the a long-lasting activity of these compounds, or the fact that advantage of convenience, lack of enzymatic damage to no receptor desensitization to their activity occurs. There cellular structures, and most relevant, that small adipose is no clear explanation for such differences in potency cells are not lost, as is the case when adipose cells are among compounds that are acting through the same isolated after enzymatic dispersion ( Julien et al. 1989). cytoplasmic receptor.

Journal of Endocrinology (1999) 160, 425–432

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By far the most surprising finding was the weak activity, Acknowledgements or lack of activity, of the androgens studied on leptin secretion by male samples in vitro. These results are This work was supported by research grants from: consistent with our previous report in which both Fondo de Investigacion Sanitaria, Spanish Ministry of oestradiol and , which act as potent leptin Health, and Conselleria de Educacion, Xunta de Galicia. secretagogues in female adipose tissue, were devoid of action in male samples (Casabiell et al. 1998). This then results in a generalized absence of response to steroid References hormones (glucocorticoids, oestrogens and androgens) in Ahima RS, Prabakaran D, Mantzoros C, Qu D, Lowell B, Maratos adipose tissue coming from male donors. Considering that Flier E & Flier JS 1996 Role of leptin in the neuroendocrine leptin secretion in male samples is inhibited in vitro by response to fasting. Nature 382 250–252. forskolin (Casabiell et al. 1998), a general lack of modu- Ahima RS, Dushay J, Flier SN, Prabakaran D & Flier JS 1997 Leptin lation may be ruled out. As the existence of androgen accelerates the onset of in normal female mice. Journal of Clinical Investigation 99 391–395. receptors in human adipose tissue is well established, and Barr V, Malide D, Zarnowski M, TaylorS&Cushman S 1997 no known sex-based differences in the amount of steroids stimulates both leptin secretion and production by rat white receptors have been reported in peripheral tissues, the adipose tissue. Endocrinology 138 4463–4472. biological basis for the observed gender-based differences Butte NF, Hopkinson JM & Nicolson MA 1997 Leptin in human reproduction: serum leptin levels in pregnant and lactating women. in leptin secretion in vitro has made future molecular Journal of Clinical Endocrinology and Metabolism 82 585–589. analysis mandatory. Whether these differences in leptin Caprio S, Tamborlane WV, Silver D, Robinson C, Leibel R, release reflect gender-based different regulatory mech- McCarthy S, Grozman A, Belous A, MaggsD&Sherwin RS 1996 anisms leading to altered transcription or stabilization of Hyperleptinemia: an early sign of juvenile obesity. Relations to the Ob gene mRNA in adipocytes merits further study. body fat depots and insulin concentrations. American Journal of Physiology 271 E626–E630. Alternatively, it cannot be excluded that the androgenic Caro JF, Sinha MK, Kolaczynski JW, Zhang PL & Considine RV action on leptin release may be mediated by indirect 1996 Leptin: the tale of an obesity gene. Diabetes 45 1455–1462. mechanisms of action such as regulation of lipolysis. In Carro E, Sen˜aris R, Considine RV, Casanueva FF & Dieguez C 1997 fact, androgens do regulate lipolysis (Xu et al. 1990), and Regulation of in vivo growth hormone secretion by leptin. Endocrinology 138 2203–2206. fatty acids seem to modulate the expression of the Ob Casabiell X, Pin˜eiro V, Tome´ MA, Peino´R,Die´guezC& gene (Rentsch & Chiesi 1996), although, to date, a Casanueva FF 1997 Presence of leptin in colostrum and/or breast sex-based difference in lipolysis regulation seems an milk from lactating mothers: a potential role in the regulation of unlikely possibility. neonatal food intake. Journal of Clinical Endocrinology and Metabolism In any case, the results here reported make it unlikely 82 4270–4273. Casabiell X, Pin˜eiro V, Peino´ R, Lage M, Camin˜a J, Gallego R, that the reported gender-based differences in circulating Garcia-Vallejo L, DieguezC&Casanueva FF 1998 Gender leptin levels in humans can be due to any androgenic differences in both spontaneous and stimulated leptin secretion by action. First of all, the inhibitory role of androgens, if any, omental adipose tissue in vitro: dexamethasone and is little in comparison with the clear-cut stimulatory action stimulate leptin release in women but not in men samples. Journal of Clinical Endocrinology and Metabolism 83 2149–2155. of oestrogens. Secondly, both compounds, androgens and Considine RV, Sinha MK, Heiman ML, Kriauciunas A, Stephens oestrogens, act exclusively on women’s samples, being TW, Nyce MR, Ohannesian JP, Marco CC, McKee LJ, Bauer TL devoid of effect on male-derived adipose tissue. These & Caro UK 1996 Serum immunoreactive-leptin concentrations in considerations, together with the well-known fact that normal-weight and obese humans. New England Journal of Medicine women have higher levels of oestrogens than androgens, 334 292–295. Ferron F, Considine RV, Peino´ R, Lado IG, DieguezC&Casanueva make minimal any role of circulating androgens in the FF 1997 Serum leptin concentrations in patients with anorexia regulation of plasma leptin levels in humans. nervosa, bulimia nervosa and non-specific eating disorders correlate In conclusion: (a) it is confirmed that spontaneous leptin with the body mass index but are independent of the respective secretion by human omental adipose tissue in vitro is larger disease. Clinical Endocrinology 46 289–293. Garcia-Mayor R, Andrade M, Rios M, Lage M, Dieguez C & in female than in male samples; (b) testosterone is devoid Casanueva FF 1997 Serum leptin levels in normal children: of action on leptin secretion; and (c) androgenic steroids relationship to age, gender, body mass index, pituitary–gonadal are potent inhibitors of leptin secretion in the adipose hormones and pubertal stage. Journal of Clinical Endocrinology and tissue of female donors, while they are scarcely or not Metabolism 82 2849–2855. effective in samples from men. The lack of effect of steroid Gray JM, Nunez AA, Siegel LI & Wade GN 1979 Effects of testosterone on body weight and adipose tissue: role of compounds on male adipose tissue, together with the aromatization. Physiology and Behavior 23 465–469. stimulatory role previously reported for oestrogens on Haffner SM, Miettinen H, Karhapaa P, MykkanenL&Laakso M female adipocytes, strongly suggests that the gender-based 1997 Leptin concentrations, sex hormones, and in differences in leptin levels observed in vivo are due to the nondiabetic men. Journal of Clinical Endocrinology and Metabolism 82 1807–1809. hormonally modulated leptin secretion in women rather Halleux CM, Servais I, Reul BA, DetryR&Brichard SM 1998 than to an inhibitory effect of androgens on men, as Multihormonal control of ob gene expression and leptin secretion previously postulated. from cultured human visceral adipose tissue: increased

Journal of Endocrinology (1999) 160, 425–432

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Journal of Endocrinology (1999) 160, 425–432

Downloaded from Bioscientifica.com at 09/27/2021 08:42:39PM via free access