Ob Gene Expression Research Articles

Effect of clofibrate on malic enzyme and leptin mRNAs level in rat brown and white adipose tissue.

Two previous studies have reported contradictory results regarding the effect of fibrates treatment on obese (ob) gene expression in rodents. The purpose of the present study was to reinvestigate this issue. We examined the effect of clofibrate (fibrate derivative) administration for 14 days to rats on malic enzyme (as an adequate control of fibrates action) and leptin mRNAs level in the white and brown adipose tissues (WAT and BAT, respectively). The malic enzyme activity and malic enzyme mRNA level in white adipose tissue increased significantly after clofibrate feeding. In brown adipose tissue, the drug treatment resulted in depression of malic enzyme activity and malic enzyme mRNA level. Under the same conditions, leptin mRNA level did not change in these tissues. The results presented in this paper provide further evidence that the clofibrate (activator of peroxisome proliferator activated receptor alpha), feeding is without effect on ob gene expression in rat white and brown adipose tissue. Furthermore, the present study demonstrates that clofibrate causes opposite effects on malic enzyme gene expression in WAT (up-regulation) and BAT (down-regulation).

Regulation of ob gene expression: evidence for epinephrine-induced suppression in human obesity.

Leptin acts as satiety factor and increases energy expenditure. Studies conducted on animals and in vitro on adipocytes culture have shown that infusion of catecholamines leads to a significant reduction of ob gene expression; it appears of interest to evaluate the in vivo effects of adrenergic activation on the expression of the ob gene in humans. We studied ob gene expression in adipose tissue samples from 13 obese subjects before and after epinephrine (25 ng/min x kg ideal body weight for 3 h) and 6 obese patients during saline infusion. Hormonal infusion led to a significant increase in epinephrine plasma levels (from 27 +/- 4 to 339 +/- 75 pg/mL; P < 0.001), plasma free fatty acids (from 0.73 +/- 0.05 to 0.98 +/- 0.07; P < 0.05), heart rate (13.5 +/- 3.1 beats/min; F = 2.9; P < 0.03), and systolic blood pressure (F = 2.7; P < 0.05), whereas diastolic blood pressure did not show significant variation. Plasma leptin levels decreased by the end of the infusion (from 63 +/- 13 to 49 +/- 11 ng/mL; P < 0.05), and ob messenger ribonucleic acid levels were significantly reduced (decrease amounting to 47 +/- 5% of basal values). Our study shows that adrenergic activation contributes to regulate ob messenger ribonucleic acid levels in humans. The interaction between epinephrine and leptin may operate during metabolic and psychological stress to regulate energy expenditure and food intake.

Regulation of ob Gene Expression: Evidence for Epinephrine-Induced Suppression in Human Obesity

Leptin acts as satiety factor and increases energy expenditure. Studies conducted on animals and in vitro on adipocytes culture have shown that infusion of catecholamines leads to a significant reduction of ob gene expression; it appears of interest to evaluate the in vivo effects of adrenergic activation on the expression of the ob gene in humans. We studied ob gene expression in adipose tissue samples from 13 obese subjects before and after epinephrine (25 ng/min·kg ideal body weight for 3 h) and 6 obese patients during saline infusion. Hormonal infusion led to a significant increase in epinephrine plasma levels (from 27 ± 4 to 339 ± 75 pg/mL; P < 0.001), plasma free fatty acids (from 0.73 ± 0.05 to 0.98 ± 0.07; P < 0.05), heart rate (13.5 ± 3.1 beats/min; F = 2.9; P < 0.03), and systolic blood pressure (F = 2.7; P < 0.05), whereas diastolic blood pressure did not show significant variation. Plasma leptin levels decreased by the end of the infusion (from 63 ± 13 to 49 ± 11 ng/mL; P < 0.05), and ob messenger ribonucleic acid levels were significantly reduced (decrease amounting to 47 ± 5% of basal values). Our study shows that adrenergic activation contributes to regulate ob messenger ribonucleic acid levels in humans. The interaction between epinephrine and leptin may operate during metabolic and psychological stress to regulate energy expenditure and food intake.

15-deoxy-Δ 12,14 prostaglandin J2: A putative endogenous promoter of adipogenesis suppresses the ob gene

Leptin is considered a key factor in the regulation of appetite and energy expenditure, but little is known about the control of its synthesis and release. Thiazolidinediones (TZDs) have recently been shown to downregulate leptin expression, and it has been speculated that downregulation of the ob gene occurs through activation of the transcription factor, peroxisome proliferator—activated receptor γ (PPARγ). However, there are no studies using an endogenous PPARγ ligand. We examined the effect of 15-deoxy-Δ 12,14 prostaglandin J 2 (15d-PGJ 2), a putative natural ligand of PPARγ, on ob gene expression in fully differentiated 3T3-L1 adipocytes and compared its effect with that of two other PPARγ activators, the TZD troglitazone (Trog) and indomethacin (indo). 15d-PGJ 2, Trog, and Indo all inhibited leptin expression at concentrations at which they activate PPARγ. The inhibition of leptin expression of PPARγ activators was surprising, since PPARγ, is known to induce adipogenesis during which the ob gene is expressed. To address the possibility that PPARγ plays different roles before and after the induction of adipogenesis, we examined the effects of the three PPARγ ligands on the expression of leptin and the glucose transporter protein GLUT4, both of which are expressed during differentiation of 3T3-L1 preadipocytes to adipocytes. In the absence of PPARγ ligands, leptin and GLUT4 synthesis increased from day 3 to day 9 or 10 during differentiation. However, in the presence of any of the three PPARγ ligands, GLUT4 expression was unaffected, while ob gene expression was inhibited. We hypothesize that PPARγ may be essential for induction of adipocyte differentiation but then needs to be inactivated to allow expression of the ob gene.

In Vivo and in Vitro ob Gene Expression and Leptin Secretion in Rat Adipocytes: Evidence for a Regional Specific Regulation by Sex Steroid Hormones

As a sexual dimorphism appears in plasma leptin levels, the aim of the present study was to investigate, in vivo and in vitro, the influence of sex steroid hormones on ob messenger RNA (mRNA) and leptin expressions in rat fat cells from various anatomical localizations. In male rats, castration resulted in a modulation of ob gene mRNA expression which was increased by 2-fold in perirenal and half-reduced in sc adipocytes. Moreover, in isolated fat cells from both perirenal and sc fat depots, ob gene mRNA expression was reduced by 20% after a 24-h in vitro exposure to dihydrotestosterone (10−8m). This effect of dihydrotestosterone on ob mRNA was prevented by exposure to the antiandrogen cyproterone acetate and also by actinomycin D. In contrast, leptin secretion from both perirenal and sc adipocytes was unchanged after 24 h exposure to dihydrotestosterone. In female rats, ovariectomy induced a 25% decrease in ob gene mRNA expression in perirenal fat cells. In vitro studies revealed that a 24-h exposure to 17-β estradiol (10−8m) induced a 1.4-, 1.2-, and 1.75-fold increase in ob mRNA expression and a 3.8-, 1.65- and 2-fold increase in leptin secretion in sc, perirenal and parametrial adipocytes, respectively. Moreover, these effects were prevented by the antiestrogen ICI182780 and also by actinomycin D. Altogether, these results demonstrate that in rat adipocytes, estrogens, and androgens modulate ob gene expression at the mRNA level through sex steroid receptor-dependent transcriptional mechanisms.

In vivo and in vitro ob gene expression and leptin secretion in rat adipocytes: evidence for a regional specific regulation by sex steroid hormones.

As a sexual dimorphism appears in plasma leptin levels, the aim of the present study was to investigate, in vivo and in vitro, the influence of sex steroid hormones on ob messenger RNA (mRNA) and leptin expressions in rat fat cells from various anatomical localizations. In male rats, castration resulted in a modulation of ob gene mRNA expression which was increased by 2-fold in perirenal and half-reduced in sc adipocytes. Moreover, in isolated fat cells from both perirenal and s.c. fat depots, ob gene mRNA expression was reduced by 20% after a 24-h in vitro exposure to dihydrotestosterone (10(-8) M). This effect of dihydrotestosterone on ob mRNA was prevented by exposure to the antiandrogen cyproterone acetate and also by actinomycin D. In contrast, leptin secretion from both perirenal and sc adipocytes was unchanged after 24 h exposure to dihydrotestosterone. In female rats, ovariectomy induced a 25% decrease in ob gene mRNA expression in perirenal fat cells. In vitro studies revealed that a 24-h exposure to 17-beta estradiol (10(-8) M) induced a 1.4-, 1.2-, and 1.75-fold increase in ob mRNA expression and a 3.8-, 1.65- and 2-fold increase in leptin secretion in sc, perirenal and parametrial adipocytes, respectively. Moreover, these effects were prevented by the antiestrogen ICI182780 and also by actinomycin D. Altogether, these results demonstrate that in rat adipocytes, estrogens, and androgens modulate ob gene expression at the mRNA level through sex steroid receptor-dependent transcriptional mechanisms.

Oleoyl-estrone treatment affects the ponderostat setting differently in lean and obese Zucker rats.

To determine whether the slimming effects of treatment with oleoyl-estrone (OE) in liposomes of normal and obese rats are permanent, or disappear as soon as the treatment with the drug ceased. This study was devised to gain further knowledge on the postulated role of OE as a ponderostat signal, evaluating whether (in addition) it can lower the ponderostat setting of the rat. The rats were infused for 14d (using osmotic minipumps) with oleoyl-estrone in liposomes at a dose of 3.5 micromol/kg x d, and were studied up to one month after the treatment ceased. Young adult lean controls (CL) or treated (TL) and obese controls (CO) or treated (TO) Zucker rats. Energy balance, blood glucose, liver glycogen, plasma insulin, leptin corticosterone, ACTH and estrone (free and total) concentrations, and expression of the OB gene in white adipose tissue (WAT). The loss of body weight caused by OE was recovered quickly in the TO, which gained weight at the same rate as the CO. TL rats, however remained at the low weight attained for one month after the treatment ceased. However, no differences were observed in calculated energy expenditure (EE) between the TL and TC rats once treatment had stopped. In TL and TO rats, liver glycogen concentrations decreased to normal shortly after treatment ceased, and leptin expression and concentrations remained normal and unchanged after the end of OE treatment. In TO rats, plasma glucose, insulin and leptin were lower than in the CO. Total estrone concentrations decreased rapidly in TL rats and more slowly in the TO, and free estrone followed a similar pattern. Continuous infusion of liposomes loaded with OE resulted in a decreased energy intake (EI), maintenance of EE and the utilization of body fat reserves in lean and obese rats alike. This process ended in obese rats as soon as the infusion ceased, so that even when the levels of free and total estrone in plasma remained high, there was a marked (and relatively fast) shift toward the basal situation, which translated into an increase in EI, maintenance of estimated EE and a marked buildup of energy stores. In lean rats, the effects of OE on leptin concentrations and OB gene expression persisted after infusion ended.

Acute Effects of Exercise on Circulating Leptin in Lean and Genetically Obesefa/faRats

Mechanisms of regulation of plasma leptin in lean and genetically obese animals are not completely understood. In particular a relation has been proposed between energy metabolism and leptin. However, it is not clear how energy expenditure and leptin are related under exercise in lean and obese animals. To clarify these aspects we investigated lean and genetically obese (fa/fa) Zucker rats undergoing a single bout (30 min) of swimming and measured several biochemical and hormonal parameters of energy metabolism and leptin changes throughout the study. Moreover ob-gene expression in adipose tissue was also measured. Our results showed that plasma leptin is decreased by 30% at the end of exercise in lean animals while resulting unaffected in obese animals. Leptin changes in lean rats are concomitant with the peak of NEFA and glycerol release from adipose tissue rather than with the reduction of plasma insulin. Ob-gene expression in adipose tissue was markedly increased in fa/fa compared to lean rats, but was not modified by exercise both in lean and obese animals. In conclusion our data show that leptin changes during exercise are related to lipolytic events in adipose tissue and support a link between leptin and energy expenditure.

Regulation of ob gene expression and leptin secretion by insulin and dexamethasone in rat adipocytes.

Leptin, the ob gene product, is produced by adipocytes, and it acts to decrease caloric intake and increase energy expenditure. To better understand the molecular mechanisms of hormone-regulated leptin synthesis and secretion, we assessed the ability of insulin and dexamethasone to acutely modulate ob gene expression and leptin secretion in rat adipocytes. Incubation of rat adipocytes with 100 nmol/l insulin for 2 h had no effect on ob mRNA levels, but it stimulated a twofold increase in leptin secretion. Dexamethasone (100 nmol/l) stimulated both a two- to fourfold increase in ob mRNA and a twofold increase in leptin secretion. Consonant with a posttranscriptional and transcriptional regulatory mechanism for insulin- and dexamethasone-stimulated leptin secretion, respectively, actinomycin D blocked dexamethasone-stimulated leptin secretion but did not affect insulin-stimulated leptin secretion. Cycloheximide treatment did not significantly affect ob mRNA accumulation, but it reduced total secreted leptin. Interestingly, however, insulin was still able to stimulate a twofold increase in leptin secretion. These data suggest that insulin, but not dexamethasone, is able to stimulate leptin secretion from a preexisting intracellular pool, although de novo protein synthesis is required for the full insulin-stimulated effect. Signaling pathways involved in leptin synthesis/secretion were also evaluated. The phosphatidylinositol 3-kinase inhibitor LY294002, the Map/Erk kinase inhibitor PD98059, and the immunosuppressant rapamycin had no effect on basal levels of leptin secretion. However, all three inhibitors markedly decreased both insulin- and dexamethasone-stimulated leptin secretion. These findings suggest a complex set of signaling pathways involved in mediating insulin- and dexamethasone-stimulated leptin synthesis and secretion.

Arginine infusion and/or acute changes of growth hormone levels do not acutely alter leptin serum levels.

Leptin, the ob gene product, is produced by differentiated adipocytes. It functions as an afferent signal to the central nervous system indicating satiety and fat mass status. It acts upon the hypothalamic-pituitary axis. Growth hormone (GH) secretion is thought to be stimulated by leptin. Conversely, leptin secretion and ob gene expression are regulated by classical neuroendocrine networks. Whether or not acute changes of GH concentrations directly alter leptin serum levels in vivo is still debated. We investigated whether or not acute changes in GH serum concentrations during arginine infusion (0.5 g/kg b. wt.) alter leptin serum levels in 45 children and adolescents (33 M, 12 F). GH and leptin serum levels were determined at -30, 0, 30, 60, 90, 120 min after arginine infusion using specific radioimmunoassays. Leptin serum concentrations remained unaltered throughout the arginine infusion in all children and adolescents whether or not GH secretion was normal. (1) Acute changes of GH levels do not alter leptin serum levels during acute arginine infusions over 120 min. (2) Arginine does not acutely modulate leptin secretion.

The regulation of obese (ob) gene expression by intracellular fatty acid concentration in adipocytes.

The regulation of obese (ob) gene expression by intracellular fatty acid concentration in adipocytes.

Short-term treatment with estrone oleate in liposomes (Merlin-2) does not affect the expression of the ob gene in Zucker obese rats.

Young female Zucker fa/fa rats of 370-430 g were implanted with osmotic minipumps releasing 3.5 micromol/day-kg of estrone oleate in liposomes (Merlin-2) into the bloodstream for up to 14 days. Merlin-2 induced a sustained loss of appetite, and a decrease in body weight of 3.5%, which contrasts with the 8.2% increase in controls during the period studied. Plasma insulin, glucose and urea decreased, and liver glycogen increased with Merlin-2 treatment. Plasma ACTH and corticosterone increased to a maximum at the end of the experiment. The expression of the ob gene in adipose tissue was unchanged, and plasma leptin levels were also unchanged by treatment. Estrone levels increased more than 1500-fold, and estrone oleate rose 100-fold during treatment. The fact that estrone oleate had no effect on the leptin levels or expression in obese rats, in contrast with the marked inhibition observed in the lean suggests that the functionality of the leptin receptor is essential for estrone oleate inhibition of the ob gene. This also suggests that leptin may control ob gene expression in white adipose tissue and that estrone oleate may activate this process. The slimming effect of estrone oleate is, thus, not directly dependent on leptin, since both normoleptinemic and hyperleptinemic animals lose fat following treatment nor are the effects on appetite and energy expenditure mediated by leptin. However, leptin levels and the expression of the ob gene are directly linked with estrone oleate function. A possible involvement of leptin in estrone oleate action is postulated. The results support the participation of estrone oleate in the control of body weight and hint at the complexity of its regulation by leptin and glucocorticoids.

Effects of intravenous neuropeptide Y on insulin secretion and insulin sensitivity in skeletal muscle in normal rats.

Intracerebroventricular administration of neuropeptide Y to normal rats induces a syndrome characterised by obesity, hyperinsulinaemia, insulin resistance and over expression of the adipose tissue ob gene. Little is known about the effect of circulating neuropeptide Y on glucose metabolism, insulin secretion and leptin. We therefore aimed to evaluate the effect of an intravenous infusion of neuropeptide Y on glucose disposal, endogenous glucose production, whole body glycolytic flux, and glucose storage as assessed during euglycaemic hyperinsulinaemic clamp. In addition, the insulin-stimulated glucose utilisation index in individual tissues was measured by the 2-deoxy-[1-3H]-glucose technique. The effect of neuropeptide Y on insulin secretion was evaluated by hyperglycaemic clamp. Infusion did not induce any change in endogenous glucose production during basal conditions or at the end of the clamp. Glucose disposal was significantly increased in the rats given neuropeptide Y compared with controls (27.8 +/- 1.3 vs 24.3 +/- 1.6 mg x min(-1) x kg(-1); p < 0.05) as was the glycolytic flux (18.9 +/- 1.6 vs 14.4 +/- 0.8 mg x min(-1) x kg(-1); p < 0.05), while glucose storage was comparable in the two groups. In skeletal muscle, the glucose utilisation index was increased significantly in rats given neuropeptide Y. The glucose utilisation index in subcutaneous and epididimal adipose tissue was not significantly different between the two groups. Plasma leptin was significantly increased by hyperinsulinaemia, but was not affected by neuropeptide Y infusion. Both the early and late phase of the insulin response to hyperglycaemia were significantly reduced by neuropeptide Y. In conclusion neuropeptide Y infusion may increase insulin-induced glucose disposal in normal rats, accelerating its utilisation through the glycolytic pathway. Neuropeptide Y reduces both phases of the insulin response to hyperglycaemia.

Low leptin gene expression and hyperleptinemia in chronic renal failure

The ob gene product leptin is thought to be a key regulator of food intake and body weight. Patients having advanced chronic renal failure (CRF) have markedly higher serum leptin levels. It is not known whether the increase in leptin levels in CRF is caused by a decreased plasma clearance and/or increased production. In the present study serum leptin levels and glomerular filtration rate (GFR) were measured in 219 patients having various degrees of renal failure. In addition, serum leptin levels, C-reactive protein (CRP), body composition (by dual-energy x-ray absorptiometry) and ob gene expression (by in situ hybridization histochemistry) were determined in 15 patients with advanced CRF. Seven of the patients were re-evaluated following 12 months of peritoneal dialysis (PD) treatment. Serum leptin levels correlated negatively to GFR (r = -0.26; P < 0.0001). The ob gene expression was significantly lower in patients with CRF than in healthy controls. A negative correlation between serum leptin levels and ob gene expression (r = -0.66; P < 0.05) was found in patients with CRP < 25 mg/liter. The ob gene expression (20.0 +/- 1.8 vs. 15.0 +/- 1.0 nCi/g; P < 0.05) was significantly higher in 5 patients with CRP > 25 mg/liter than in 10 patients with CRP < 25 mg/liter. Following 12 months of PD, the amount of body fat increased by 30% while the ob gene expression remained unchanged. The present study shows a correlation between serum leptin levels and GFR, and our results suggest that elevated leptin levels, due to a decreased plasma clearance, down-regulate the expression of the ob gene. We also found that an ongoing inflammation stimulates ob gene expression in patients with CRF. Therefore, it is suggested that the hyperleptinemia induced feedback inhibition of ob gene expression is overcome by inflammatory cytokines.

Increased OB gene expression leads to elevated plasma leptin concentrations in patients with chronic primary hyperinsulinemia.

Leptin, a hormone secreted by adipocytes, decreases food intake and increases energy expenditure. The role of insulin in the regulation of leptin secretion is poorly understood and is still a topic of debate. Insulin increases leptin mRNA synthesis in rodents, but in humans, the available data are discordant. To investigate the role of chronic hyperinsulinemia in the regulation of plasma leptin concentrations, we studied 13 patients with surgically confirmed insulinoma before and after tumor removal, along with 15 healthy control subjects matched for sex, age, and BMI. Immunoreactive plasma leptin levels were measured by radioimmunoassay; leptin mRNA levels were also determined by reverse transcription-competitive polymerase chain reaction in a subgroup of six patients with insulinoma and six control subjects. All determinations were made with subjects in the fasting state. Plasma leptin concentrations correlated positively with leptin mRNA levels (r = 0.880, P < 0.001). Leptin levels, both plasma protein and mRNA, were significantly higher in the insulinoma patients than in the control subjects (plasma protein: 17.5 +/- 3.6 vs. 2.9 +/- 0.4 ng/ml, respectively, P < 0.001; mRNA: 0.98 +/- 0.33 vs. 0.19 +/- 0.064 amol/microg RNA, respectively, P < 0.05), and they correlated positively with fasting plasma insulin levels in the patients with insulinoma (plasma protein: r = 0.686, P < 0.01; mRNA: 0.796, P < 0.05). Finally, removal of the insulin-secreting tumor was followed by the normalization of plasma leptin levels. In summary, in patients with insulinoma, 1) plasma leptin levels and leptin mRNA are elevated; 2) a direct relationship exists between leptin, both circulating protein and mRNA, and insulin concentrations; and 3) plasma leptin returns to normal levels after tumor removal. These data, therefore, support a role for insulin in the chronic regulation of leptin gene expression.

A nutrient-sensing pathway regulates leptin gene expression in muscle and fat.

Leptin, the protein encoded by the obese (ob) gene, is synthesized and released in response to increased energy storage in adipose tissue. However, it is still not known how incoming energy is sensed and transduced into increased expression of the ob gene. The hexosamine biosynthetic pathway is a cellular 'sensor' of energy availability and mediates the effects of glucose on the expression of several gene products. Here we provide evidence for rapid activation of ob gene expression in skeletal muscle by glucosamine. Increased tissue concentrations of the end product of the hexosamine biosynthetic pathway, UDP-N-acetylglucosamine (UDP-GlcNAc), result in rapid and marked increases in leptin messenger RNA and protein levels (although these levels were much lower than those in fat). Plasma leptin levels and leptin mRNA and protein levels in adipose tissue also increase. Most important, stimulation of leptin synthesis is reproduced by either hyperglycaemia or hyperlipidaemia, which also increase tissue levels of UDP-N-acetylglucosamine in conscious rodents. Finally, incubation of 3T3-L1 pre-adipocytes and L6 myocytes with glucosamine rapidly induces ob gene expression. Our findings are the first evidence of inducible leptin expression in skeletal muscle and unveil an important biochemical link between increased availability of nutrients and leptin expression.

The influence of ovariectomy on ob gene expression in rats.

Ovarian steroid hormones exert major influences on eating behaviour and body weight regulation of female rats. Ovariectomy (OVX) results in an increase in food intake and a concomitant increase in body weight, while estradiol (E2) replacement reverses these effects. In this study, we examined the influence of OVX on obese (ob) gene expression in rat adipose tissues and serum leptin concentration. Female Wistar rats, 10 weeks old, were divided into three groups: sham-operated control rats receiving corn oil (group 1, n = 4), ovariectomized rats receiving corn oil (group 2, n = 5), and ovariectomized rats receiving 17beta-E2 (10 microg/kg/day) replacement (group 3, n = 4). After 4 weeks, the rats and food consumption were weighed and serum E2 and leptin levels were measured by radioimmunoassays. Furthermore, the expression levels of ob mRNA obtained from the bilateral perimetric fat pads were estimated by Northern blot analysis. The mean weight and food consumption in group 2 were significantly (p < 0.01) heavier than those in group 1. But there were no significant differences between group 1 and group 3. The expression levels of ob mRNA in group 2 were lower than those in group 1, however, the levels of group 3 were restored to the level of group 1. On the other hand, no significant differences among the 3 groups as to serum levels of leptin were observed. The data herein clearly indicate that ovarian steroid hormones may be one of the factors involved in the regulation of ob gene.

Multihormonal control of ob gene expression and leptin secretion from cultured human visceral adipose tissue: increased responsiveness to glucocorticoids in obesity.

The direct role of hormones on leptin synthesis has not yet been studied in cultured adipose cells or tissue from lean and obese subjects. Moreover, this hormonal regulation has never been addressed in human visceral fat, although this site plays a determinant role in obesity-linked disorders. In this study, we investigated the hormonal control of ob expression and leptin production in cultured visceral adipose tissue from lean and obese subjects. We more particularly focused on the interactions between glucocorticoids and insulin. We also briefly tackled the role of cAMP, which is still unknown in man. Visceral (and subcutaneous) adipose tissues from eight obese (body mass index, 41 +/- 2 kg/m2) and nine nonobese (24 +/- 1 kg/m2) subjects were sampled during elective abdominal surgery, and explants were cultured for up to 48 h in MEM. The addition of dexamethasone to the medium increased ob gene expression and leptin secretion in a time-dependent manner. Forty-eight hours after dexamethasone (50 nmol/L) addition, the cumulative integrated ob messenger ribonucleic acid (mRNA) and leptin responses were, respectively, approximately 5- and 4-fold higher in obese than in lean subjects. These responses closely correlated with the body mass index. The stimulatory effect of the glucocorticoid was also concentration dependent (EC50 = approximately 10 nmol/L). Although the maximal response was higher in obese than in lean subjects, the EC50 values were roughly similar in both groups. Unlike dexamethasone, insulin had no direct stimulatory effect on ob gene expression and leptin secretion. Singularly, insulin even inhibited the dexamethasone-induced rise in ob mRNA and leptin release. This inhibition was observed in both lean and obese subjects, whereas the expected stimulation of insulin on glucose metabolism and the accumulation of mRNA species for the insulin-sensitive transporter GLUT4 and glyceraldehyde-3-phosphate dehydrogenase occurred in lean patients only. This inhibitory effect was already detectable at 10 nmol/L insulin and was also observed in subcutaneous fat. Although a lowering of intracellular cAMP concentrations is involved in some of the effects of insulin on adipose tissue, this cannot account for the present finding, because the addition of cAMP to the medium also decreased ob mRNA and leptin secretion (regardless of whether dexamethasone was present). In conclusion, glucocorticoids, at physiological concentrations, stimulated leptin secretion by enhancing the pretranslational machinery in human visceral fat. This effect was more pronounced in obese subjects due to a greater responsiveness of the ob gene and could contribute to the metabolic abnormalities associated with central obesity by para/endocrine actions of hyperleptinemia on adipocytes and liver. Unlike dexamethasone, insulin had no direct stimulatory effect on ob gene expression and leptin secretion, and even prevented the positive response to dexamethasone by a cAMP-independent mechanism that remained functional despite insulin resistance.

Multihormonal Control of ob Gene Expression and Leptin Secretion from Cultured Human Visceral Adipose Tissue: Increased Responsiveness to Glucocorticoids in Obesity

Multihormonal Control of ob Gene Expression and Leptin Secretion from Cultured Human Visceral Adipose Tissue: Increased Responsiveness to Glucocorticoids in Obesity

A molecular investigation of the obese phenotype in the Aston University strain of ob/ob mice and the Japanese Kuo Kondo mice.

The aim of this work was to determine the role of the ob gene in the obese phenotype observed in the Aston University Strain of ob/ob mouse and the Japanese Kuo Kondo (KK) mouse. After RT-PCR amplification of the ob RNA, the transcript was cloned into the vector pCR3 and three individual clones from each strain were sequenced. It was confirmed that the Aston University strain of ob/ob mice shared the same C-T mutation found in the Jackson Laboratory C57BL/6J ob/ob strain whereas the Japanese KK mice showed wild-type ob gene expression. This study indicates that the ob mutation has survived unchanged following the separation of the two strains of ob/ob mice, and secondly, that the molecular basis of the obese phenotype in the KK mice is not due to mutations in the ob gene.