Glucocorticoid binding during the differentiation of 3T3-F442A fibroblasts into adipocytes. A possible regulatory effect of insulin.

Until recently, few studies had been carried out on receptors for glucocorticoids in adipocytes, although the role of these steroids is considerable. In the present studies, we chose the pre-adipocyte line 3T3-F442A, which constitutes an excellent model for investigating the differentiation and function of adipocytes. Using a whole cell assay system, we showed the existence of a homogenous class of sites with the characteristics of glucocorticoid receptors, that is, high-affinity binding which is reversible, specific and saturable. Whatever the state of cellular differentiation, the affinity of the receptor for dexamethasone did not vary, although we observed an increase in the number of sites during differentiation. When cells were differentiated in the presence of insulin, there was a further increase in the binding capacity; moreover, insulin deprivation of such adipocytes caused a decrease in the number of sites. Our results therefore suggest that factors other than the glucocorticoids themselves influence dexamethasone binding. It is suggested that insulin plays a role in the regulation of the number of glucocorticoid receptors.

Development under the control of insulin of lipogenic enzymes, lipoprotein lipase, isoproterenol and glucagon sensitivity in differentiating rat preadipocytes in primary culture.

In preadipose cellular fractions (I, II and III) isolated by density gradient centrifugation from the inguinal tissue of young rats, we followed the activity of fatty acid synthetase, ATP citrate lyase and lipoprotein lipase during differentiation in culture. 1.5 nM insulin when added at confluence markedly induced the activity of ATP citrate lyase and fatty acid synthetase in the cells derived from the lighter fractions (I and II). The magnitude of this response was 25-50-fold the initial value 15 days after plating. In the cells of the heaviest fraction (III) both enzymes exhibited low activity which was slightly stimulated by the presence of insulin, VLDL and heparin. In contrast, the activity of lipoprotein lipase appeared before confluence in cells from all three fractions and peaked at day 6 after plating. This early emergence was independent of the addition of insulin to the medium. However, insulin slightly enhanced the peak activity in post-confluent cells. The development of cAMP production in response to isoproterenol (100 microM) and to glucagon (0.3 microM) was determined in the cells of fraction II in the same culture conditions. The responsiveness to isoproterenol was present very early in these cells and rose rapidly during the exponential growth phase, reaching a peak value at day 8 after plating. In contrast, the development of glucagon sensitivity occurred only during late differentiation. The stimulatory effect of glucagon was enhanced when VLDL and heparin were added with insulin to the medium.

Adipose angiotensinogen is involved in adipose tissue growth and blood pressure regulation.

White adipose tissue and liver are important angiotensinogen (AGT) production sites. Until now, plasma AGT was considered to be a reflection of hepatic production. Because plasma AGT concentration has been reported to correlate with blood pressure, and to be associated with body mass index, we investigated whether adipose AGT is released locally and into the blood stream. For this purpose, we have generated transgenic mice either in which adipose AGT is overexpressed or in which AGT expression is restricted to adipose tissue. This was achieved by the use of the aP2 adipocyte-specific promoter driving the expression of rat agt cDNA in both wild-type and hypotensive AGT-deficient mice. Our results show that in both genotypes, targeted expression of AGT in adipose tissue increases fat mass. Mice whose AGT expression is restricted to adipose tissue have AGT circulating in the blood stream, are normotensive, and exhibit restored renal function compared with AGT-deficient mice. Moreover, mice that overexpress adipose AGT have increased levels of circulating AGT, compared with wild-type mice, and are hypertensive. These animal models demonstrate that AGT produced by adipose tissue plays a role in both local adipose tissue development and in the endocrine system, which supports a role of adipose AGT in hypertensive obese patients.

Angiotensinogen-deficient mice exhibit impairment of diet-induced weight gain with alteration in adipose tissue development and increased locomotor activity.

White adipose tissue is known to contain the components of the renin-angiotensin system, which gives rise to angiotensin II from angiotensinogen (AGT). Recent evidence obtained in vitro and ex vivo is in favor of angiotensin II acting as a trophic factor of adipose tissue development. To determine whether AGT plays a role in vivo in this process, comparative studies were performed in AGT-deficient (agt(-/-)) mice and control wild-type mice. The results showed that agt(-/-) mice gain less weight than wild-type mice in response to a chow or high fat diet. Adipose tissue mass from weaning to adulthood appeared altered rather specifically, as both the size and the weight of other organs were almost unchanged. Food intake was similar for both genotypes, suggesting a decreased metabolic efficiency in agt(-/-) mice. Consistent with this hypothesis, cellularity measurement indicated hypotrophy of adipocytes in agt(-/-) mice with a parallel decrease in the fatty acid synthase activity. Moreover, AGT-deficient mice exhibited a significantly increased locomotor activity, whereas metabolic rate and mRNA levels of uncoupling proteins remained similar in both genotypes. Thus, AGT appears to be involved in the regulation of fat mass through a combination of decreased lipogenesis and increased locomotor activity that may be centrally mediated.

Sequence of rat lipoprotein lipase-encoding cDNA.

A rat lipoprotein lipase (LPL)-encoding cDNA (LPL) has been entirely sequenced and compared to the sequences of all the LPL cDNAs reported in other species. As expected, high homology was found between the coding exons. The putative catalytic triad, Ser132, Asp156, His241, according to human numbering, is conserved in rat. As is the case in mouse, an Asn444 present in human LPL is also missing. The major divergences between human, mouse and rat LPLs were observed in the untranslated exon 10, where (i) the rat cDNA exhibits a 157-bp insertion and an 81-bp deletion relative to human; (ii) neither the B1 repeat nor the homopurine stretch reported in mouse can be recognized, and (iii) the rat cDNA displays several A+T-rich stretches.

Alterations of lipid metabolism and gene expression in rat adipocytes during chronic olanzapine treatment.

Although antipsychotics are established drugs in schizophrenia treatment, they are admittedly known to induce side effects favoring the onset of obesity and worsening its complications. Despite potential involvement of histamine receptor antagonism, or of other neurotransmitter systems, the mechanism by which antipsychotic drugs increase body weight is not elucidated. The aim of the present study was to investigate whether chronic antipsychotic treatments can directly alter the regulation of two main functions of white adipose tissue: lipolysis and glucose utilization. The influence of a classical antipsychotic (haloperidol) was compared to that of two atypical antipsychotics, one known to favor weight gain (olanzapine), the other not (ziprasidone). Cell size, lipolytic capacity and glucose transport activity were determined in white adipocytes of rats subjected to 5-week oral treatment with these antipsychotics. Gene expression of adipocyte proteins involved in glucose transport or fat storage and mobilization, such as glucose transporters (GLUT1 and GLUT4), leptin, matrix metallo-proteinase-9 (MMP9), hormone-sensitive lipase (HSL) and fatty acid synthase (FAS) was also evaluated. Adipocytes from chronic olanzapine-treated rats exhibited decreased lipolytic activity, lowered HSL expression and increased FAS expression. These changes were concomitant to enlarged fat deposition and adipocyte size. Alterations were observed in adipocytes from olanzapine-treated rats whereas the other antipsychotics did not induce any notable disorder. Our results therefore show evidence of an effect of chronic antipsychotic treatment on rat adipocyte metabolism. Thus, impairment of fat cell lipolysis should be considered as a side effect of certain antipsychotics, leading, along with the already documented hyperphagia, to the excessive weight gain observed in patients under prolonged treatment..

Long-term treatment with interleukin-1beta induces insulin resistance in murine and human adipocytes.

AIMS/HYPOTHESIS: Adipose tissue inflammation has recently been implicated in the pathogenesis of insulin resistance and is probably linked to high local levels of cytokines. IL1B, a proinflammatory cytokine, may participate in this alteration. MATERIALS AND METHODS: We evaluated the chronic effect (1-10 days) of IL1B (0.1-20 ng/ml) on insulin signalling in differentiating 3T3-F442A and differentiated 3T3-L1 murine adipocytes and in human adipocytes. We also assessed expression of the gene encoding IL1B in adipose tissue of wild-type and insulin-resistant mice (diet-induced and genetically obese ob/ob mice). RESULTS: IL1B inhibited insulin-induced phosphorylation of the insulin receptor beta subunit, insulin receptor substrate 1, Akt/protein kinase B and extracellular regulated kinase 1/2 in murine and human adipocytes. Accordingly, IL1B suppressed insulin-induced glucose transport and lipogenesis. Long-term treatment of adipose cells with IL1B decreased cellular lipid content. This could result from enhanced lipolysis and/or decreased expression of genes involved in lipid metabolism (acetyl-CoA carboxylase, fatty acid synthase). Down-regulation of peroxisome proliferating-activated receptor gamma and CCAAT/enhancer-binding protein alpha in response to IL1B may have contributed to the altered phenotype of IL1B-treated adipocytes. Moreover, IL1B altered adipocyte differentiation status in long-term cultures. IL1B also decreased the production of adiponectin, an adipocyte-specific protein that plays a positive role in insulin sensitivity. Expression of the gene encoding IL1B was increased in epididymal adipose tissue of obese insulin-resistant mice. CONCLUSIONS/INTERPRETATION: IL1B is upregulated in adipose tissue of obese and insulin-resistant mouse models and may play an important role in the development of insulin resistance in murine and human adipose cells.

Differential effects of retinoic acid upon early and late events in adipose conversion of 3T3 preadipocytes.

When confluent 3T3-F442A cell cultures (Day 0) were grown for 3 days in fetal calf serum-supplemented medium containing isobutyl methyl xanthine and dexamethasone (induction phase) and then shifted to serum-free hormone-defined medium (expression phase), they spontaneously exhibited a sharp rise in lipoprotein lipase activity (LPL); at the peak (Day 7) the LPL activity was about 25 times higher than in control cultures and was further enhanced by insulin. Although this expression of LPL activity was spontaneous, the emergence of glycerophosphate dehydrogenase (G3PDH) activity was completely dependent upon insulin as well as upon the expression of the differentiated phenotype. In committed cells, insulin elicited sustained DNA synthesis associated with limited cell proliferation. The addition of retinoic acid during the phase of expression inhibited insulin-dependent terminal differentiation (i.e., the emergence of G3PDH activity and acquisition of the differentiated phenotype). In addition, retinoic acid counteracted the stimulating effect of insulin upon LPL activity, but affected neither the mitotic process nor the spontaneous emergence of LPL activity. When added during the phase of induction, it prevented the overall process of adipogenic differentiation. Thus, the use of retinoic acid can indicate independent control of the mitogenic and lipogenic effects of insulin following commitment to adipogenic differentiation.

[Role of adipocyte precursors in the initiation of nutritional obesity before weaning]. / Rôle des précurseurs d'adipocytes dans l'installation d'une obésité nutritionnelle avant le sevrage.

The stroma vascular fraction of adipose tissue partly consists of adipose precursor cells which can convert into adipocytes in vitro. The aim of this study was to investigate the possible contribution of cells from the stroma vascular compartment to the initiation of obesity induced by overfeeding during the early neonatal weeks in rats. Overfeeding during the suckling period was obtained by reducing the litter size. The inguinal adipose tissue of overfed rats raised in litters of 4 pups each was overdeveloped compared to that of controls raised in litters of 8 pups each, and the difference between the two groups became significant as early as 10 days of age. At this age, adipose tissue enlargement was only due to adipocyte hypertrophy; afterwards, hyperplasia of the mature fat cells contributed to the overdevelopment of adipose tissue in 15-day old overfed rats. The cell number in the stroma vascular fraction increased in the overfed group as early as 10 days of age and thus preceded the onset of mature fat cell hyperplasia. The developmental pattern of lipoprotein lipase, glycerol-3-phosphate dehydrogenase, glycerol-acyl-transferase and acyl-CoA ligase activities in stromal cells did not depend on litter size, but specific enzyme activities wee increased in 10-day old overfed rats compared to the controls. These results indicate that early overfeeding induced cell proliferation in the stroma vascular compartment and also induced the enzyme activities involved in adipose conversion to increase in these cells. This strongly suggests that precursor cell differentiation was greater in overfed rats than in control rats.

Transcriptional down-regulation by insulin of the beta 3-adrenergic receptor expression in 3T3-F442A adipocytes: a mechanism for repressing the cAMP signaling pathway.

Modulation of the three beta-adrenergic receptor subtypes (beta-ARs) by insulin was investigated in mouse 3T3-F442A adipocytes. Saturation and competition experiments measuring binding of 125I-labeled (-)-cyanopindolol to adipocyte membranes demonstrated that cell exposure to insulin for 4 days caused a 3.5-fold decrease in the density of the major beta-AR component of the adipocyte, the beta 3-AR, while beta 1-AR sites remained unchanged and beta 2-ARs were undetectable. This correlated with a lower potency of the beta 3-AR-selective agonists CGP12177, ICI201651, and BRL37344 in stimulating adenylate cyclase. Northern blotting analysis indicated that insulin induced a rapid and sharp decrease in beta 3-AR mRNA levels. This effect was detectable at low insulin concentrations (EC50 = 3 nM) and was not observed in the presence of insulin-like growth factor I, suggesting an insulin receptor-mediated phenomenon. Reverse transcriptase-PCR analysis showed that, in contrast to its dramatic down-regulatory effect on beta 3-AR mRNA, insulin did not modify the levels of beta 1- and beta 2-AR transcripts. As assessed by nuclear run-on assays, insulin inhibited the beta 3-AR gene transcription rate by 90% within 30 min. mRNA turnover experiments showed that the half-life of beta 3-AR mRNA was short (90 min) and remained unaffected by insulin. These findings demonstrate the genetic control of a beta-AR subtype expression by insulin and reveal a mechanism for the regulation by this hormone of cAMP-dependent biological processes in adipocytes.

Impaired fat storage capacity in adipocyte precursors of I versus C57BL mice.

I mouse strain displays adipocyte hypoplasia responsible for smaller fat pad size compared with C57BL mice. We investigated possible alterations in the proliferation and/or differentiation capacity of preadipocytes from the stroma-vascular fraction of adipose tissue in the I mouse strain. Control C57BL and I mice were studied at 8 weeks of age, and both adipose and stromal cells were isolated from epididymal and inguinal adipose tissue localizations. Results showed that the lower epididymal adipose mass in I mice was accompanied by a decrease in stromal cell number compared with C57BL mice. In inguinal fat pads, total cell number in the stroma-vascular fraction was unmodified; lipoprotein lipase activity significantly increased in stromal cells from I mice compared with control mice. In this depot, further characterization of cells from the stroma-vascular fraction by separation of cells according to density showed an increased number of preadipocytes in the I mouse whole stromal cell population. These preadipocytes seemed unable to undergo terminal maturation, thus leading to a decrease in the number of mature adipocytes. These results indicated that resistance to fat accumulation in I mice is characterized by site-dependent impairment of both the proliferative rate and the differentiation capacity of adipocyte precursors.

[Decrease of gene expression of glycerophosphate dehydrogenase by dexamethasone in differentiated 3T3-F442A cells: antagonism with insulin and antiglucocorticoid RU38486]. / Diminution de l'expression du gène de la glycérophosphate déshydrogénase par la dexaméthasone, dans les cellules 3T3-F442A différenciées: antagonisme avec l'insuline et l'antiglucocorticoïde RU38486.

Preadipocyte subclones derived from mouse 3T3 cells differentiate into adipocytes; this differentiation is characterized by an increased activity of numerous enzymes required for triglyceride synthesis and/or mobilization. Among these enzymes, the role of glycerophosphate dehydrogenase in the differentiation process has been previously reported. In the present work, we studied the hormonal regulation of glycerophosphate dehydrogenase gene expression (G3PDH) in differentiated 3T3-F442A adipocytes. Dexamethasone (DEX) elicited a 50% decrease in both mRNA content and specific activity of G3PDH. This effect was due to a posttranscriptional event since DEX shortened the half life of the mRNA, whereas it did not modify the transcription rate of this gene. The DEX effect is specific to G3PDH, since the expression of another adipose-specific gene, namely adipsin, is not modified by DEX treatment. Insulin counteracts the inhibitory effect of DEX, mainly by stabilizing the mRNA encoding for G3PDH. The antiglucocorticoid RU38486 is able to reverse DEX inhibition. Latter phenomenon suggests that DEX action on G3PDH gene expression could be mediated by glucocorticoid receptors.

Dexamethasone regulation of terminal differentiation in 3T3-F442A preadipocyte cell line.

The 3T3-F442A preadipocyte cell line was previously shown to possess specific glucocorticoid receptors whose number increased in the time course of differentiation. We have examined the effects of a three day dexamethasone treatment, added at confluence, on cells differentiated in the presence or absence of insulin. Triglyceride accumulation, polyamine content as well as glycerophosphate dehydrogenase and fatty acid synthetase activities were measured during the adipose conversion. We have also determined 2-deoxyglucose uptake in non-differentiated and differentiated cells. Dexamethasone was shown to decrease the adipose conversion by 3T3-F442A cells in the presence or absence of insulin. Intracellular spermidine content in differentiating cells was sensitive to dexamethasone and insulin in the same way as an enzymatic marker of terminal differentiation, glycerophosphate dehydrogenase. Dexamethasone decreases the 2 deoxyglucose uptake in non-differentiated and differentiated cells while insulin increases this uptake only in differentiated cells. This work shows that glucocorticoids inhibit adipocyte metabolism at distinct levels and suggests that these hormones might play an important role in the regulation of adipose tissue mass.

Role of adipocyte precursors in the onset of obesity induced by overfeeding in suckling rats.

Rats were overfed during the suckling period by litter size manipulation in order to investigate the possible contribution of preadipocytes from the stroma-vascular compartment of adipose tissue to the development of obesity. Rats raised in litters of four pups were overfed; for normal feeding we assigned eight pups per litter. As early as 10 d of age, overfed rats became fatter than controls, and showed an increase in both plasma insulin and triacylglycerol levels. At this age, adipose tissue overdevelopment arose only from adipocyte hypertrophy, since hyperplasia occurred only at 15 d of age. Concurrently, compared to normal feeding, overfeeding led to significantly higher activities of lipoprotein lipase (LPL), glycerophosphate dehydrogenase (GPDH) and glycerophosphate acyltransferase (GPAT) in mature fat cells; 10-d-old overfed pups exhibited a higher stromal cell number. Further separation of this heterogeneous fraction by density gradient centrifugation showed a higher preadipocyte number as compared to that of controls. In stromal cells, LPL, GPDH, GPAT and acyl CoA ligase activities were detected during the suckling period. As compared to controls, overfeeding induced an increase in both LPL and GPDH activities in 10-d-old pups. Results indicate that overfeeding in early life induced an excess of fat storage capacity through a simultaneous increase in proliferation and differentiation rates of adipocyte precursors.

[Development of esterification capacity of fatty acids in the adipocytes of the rat before weaning: the effect of milk overfeeding]. / Evolution de la capacité d'estérification des acides gras dans les adipocytes de rat avant le sevrage: effet de la suralimentation lactée.

During the suckling period, lipid storage in rat adipose tissue arises from exogenous triglycerides, as milk contains a large amount of lipid. In the present work, we studied the effect of overfeeding during the suckling period on the developmental pattern of the fatty acid esterification pathway in fat cells. Early overfeeding was induced by restricting litter size; at 2 days of age, the pups were randomly distributed to form litters of 4 (overfed group) or 8 (controls). During the suckling period the activity of lipoprotein lipase (LPL), glycerophosphate dehydrogenase (G3PDH), glycerophosphate acyl transferase (GPAT) and acyl CoA ligase were measured in isolated fat cells from the inguinal adipose tissue. As early as 10 days of age, plasma triglyceride levels were increased in overfed pups as compared to the controls and, in these animals, the adipose tissue was overdeveloped, mainly due to fat cell hypertrophy. At that age, overfeeding induced a significant increase in LPL and GPAT activities, but acyl CoA ligase and GPDH were not modified in overfed pups. Fat cell hypertrophy appeared to be responsible for the increased GPAT activity in the overfed pups. On the contrary, the effect of overfeeding on LPL activity was not only related to fat cell enlargement, as the enzyme activity continued to increase in the overfed rats when expressed per unit of cell surface area. These results show that overfeeding during the suckling period rapidly induced an increase in the fat storage capacity of the adipose tissue. This suggests that circulating triglyceride levels could regulate directly or through hormonal control the activity of the fatty acid esterification pathway in fat cells.

A method for separating cultured preadipocytes according to their density: application to stromal cells from overfed suckling rats.

The stroma vascular fraction of adipose tissue consists of a heterogeneous cell population; not all the cells in this compartment undergo adipose conversion in primary culture. A density gradient centrifugation procedure was used to separate cultured cells on the basis of their triglyceride content. This method was applied to both stroma vascular cells from rat adipose tissue and to a 3T3 F442A preadipose cell line as a reference. Comparison of the results obtained from these two cell types suggests that this separation procedure can lead to a quantification of adipose differentiation in the heterogeneous stromal cell population. Separation procedures were applied to cultured stromal cells derived from young rats during the onset of nutritional obesity induced by overfeeding in early life. Results show that early overfeeding induced an increase in the stromal cell differentiation capacity which is expressed in vitro.

Adipsin mRNA amounts are not decreased in the genetically obese Zucker rat.

Adipsin gene expression as assessed by mRNA amounts was examined in adipose tissue of genetically obese rats at the onset (16 days of age) or at later stages (30 and 60 days of age) of obesity. Amounts of mRNA were equivalent in obese and lean rats at 16 days of age. In adult rats, we observed a 2-fold decrease in adipsin mRNA in the obese rats compared with control lean rats, which was abolished by weaning the animals on a high-fat diet. Our data show that, in sharp contrast with genetically obese mice, adipsin mRNA is not suppressed in genetically obese Zucker rats.

Growth hormone binding to cultured preadipocytes from obese fa/fa rats increases during cell differentiation.

Preadipocytes from 7 day-old obese Zucker rats and from their lean littermates were grown in primary culture. Specific binding of human growth hormone (hGH) to the cells is demonstrated. There is no change in hGH binding to preadipocytes from lean animals before (day 5 of the culture) and after adipose conversion (day 12). Moreover, addition of insulin to the culture medium does not affect the specific binding of 125I-hGH to the cells. On the contrary, when 1 nM insulin is added to the culture medium of preadipocytes from obese rats, the specific binding of 125I-hGH is enhanced during the cell differentiation, with a 3-fold increase in the number of binding sites. The functional significance of these additional GH sites in differentiated preadipocytes from obese rats has to be examined.

Increased lipoprotein lipase content in the adipose tissue of suckling and weaning obese Zucker rats.

The aim of this study was to determine whether the increase in lipoprotein lipase activity displayed by the adipose tissue of obese (fa/fa) rats as compared with that of lean (Fa/fa) rats could be ascribed to a change in the content or in the catalytic properties of the enzyme. The question was addressed in rats of two ages: in 7-day-old suckling and in 30-day-old post-weaning pups. Inguinal fat-pads were removed surgically (7 days of age) or after killing (30 days of age), and acetone-extract powders were prepared. The relative quantity of enzyme was assessed by immunotitration using an antiserum raised in goat against purified lipoprotein lipase from rat adipose tissue. The results indicate that increases in enzyme activity in obese animals were strictly paralleled by increases in the amount of enzyme in suckling as well as in post-weaning pups. Moreover, the apparent Km values of lipoprotein lipase for its substrate triacylglycerol were identical in the two genotypes. In conclusion, the genotype-mediated increase in lipoprotein lipase activity in adipose tissue of obese Zucker rats was fully accounted for by an increase in the content of the enzyme. In addition, this work documents the mechanism of the increase in lipoprotein lipase activity during weaning, which is mediated mainly through changes in the adipose-tissue enzyme content.

Molecular and cellular mechanisms of adipose secretion: comparison of leptin and angiotensinogen.

Besides their function of lipid storage, the adipose cells secrete a number of proteins of physiopathological importance. To get further insights into this function, which remains poorly characterized, we sought to compare the mechanisms and regulation of secretion of two individual proteins in the same cells. Leptin and angiotensinogen were chosen and assessed by radioimmunoassay and quantitative immunoblotting, respectively, in primary culture of epididymal adipose cells from young obese Zucker rats. Leptin was secreted at a steady rate of 4 ng/10(6) cells/h over 2-6 h. Despite secretion, leptin cellular content remained stable at 3 ng/10(6) cells. In contrast, the rate of angiotensinogen secretion decreased regularly from 45 arbitrary units/10(6) cells/h at 2 h, to half this value at 6 h, although cell content remained constant at 100 arbitrary units/10(6) cells. Inhibition of protein synthesis by cycloheximide depleted the cells from leptin, but not from angiotensinogen for up to 6 h. Insulin increased leptin secretion (+75%) and cell content (+70 %), without affecting angiotensinogen. Secretion of both proteins was inhibited by Golgi-disturbing agents, brefeldin A and monensin. The presence of brefeldin A led to a specific rise in leptin cell content, an effect inhibited by cycloheximide and enhanced by insulin (+80%). These data show that leptin and angiotensinogen are both secreted through Golgi-dependent pathways and that their respective intracellular pool exhibit distinct turn-over rate and insulin sensitivity. These characteristics might account for the differential response of these adipose proteins to variations in the systemic environment.