Lipolysis in Adipocytes Isolated from Deep and Superficial Subcutaneous Adipose Tissue

Objective: Abdominal subcutaneous adipose tissue (SAT) occurs in two depots separated by a fascial plane: deep SAT and superficial SAT. In a recent study it was demonstrated that the amount of deep SAT has a much stronger relationship to insulin resistance than does superficial SAT. Because insulin resistance may be related to fatty acid release from adipose tissue, we hypothesized that the two SAT depots may have a different lipolytic activity. Research Methods and Procedures: To test this hypothesis, we obtained samples of deep and superficial SAT from patients undergoing elective abdominal surgery. The rate of lipolysis was determined in the collagenase‐digested adipocytes obtained from the two fat depots by measuring glycerol release in the presence and absence of isoproterenol. In addition, the relative concentration of hormone‐sensitive lipase was determined in both SAT depots by Western blot analysis. Results: Our results showed that the rate of isoproterenol‐stimulated lipolysis was ～20% higher in cells from deep SAT compared with those from superficial SAT, indicating that the deep SAT is more lipolytically active. The concentration of hormone‐sensitive lipase did not differ between the two adipose tissue depots. Discussion: These findings suggest that the higher lipolytic activity of deep SAT may account for its stronger association with insulin resistance. The mechanism seems to be independent of differences in hormone‐sensitive lipase concentration.     

Translocation of hormone-sensitive lipase and perilipin upon lipolytic stimulation of rat adipocytes

Adipocyte lipolysis was compared with hormone-sensitive lipase (HSL)/perilipin subcellular distribution and perilipin phosphorylation using Western blot analysis. Under basal conditions, HSL resided predominantly in the cytosol and unphosphorylated perilipin upon the lipid droplet. Upon lipolytic stimulation of adipocytes isolated from young rats with the beta-adrenergic agonist, isoproterenol, HSL translocated from the cytosol to the lipid droplet, but there was no movement of perilipin from the droplet to the cytosol; however, perilipin phosphorylation was observed. By contrast, upon lipolytic stimulation and perilipin phosphorylation in cells from more mature rats, there was no HSL translocation but a significant movement of perilipin away from the lipid droplet. Adipocytes from younger rats had markedly greater rates of lipolysis than those from the older rats. Thus high rates of lipolysis require translocation of HSL to the lipid droplet and translocation of HSL and perilipin can occur independently of each other. A loss of the ability to translocate HSL to the lipid droplet probably contributes to the diminished lipolytic response to catecholamines with age.     

Reactive oxygen species facilitate translocation of hormone sensitive lipase to the lipid droplet during lipolysis in human differentiated adipocytes

In obesity, there is an increase in reactive oxygen species (ROS) within adipose tissue caused by increases in inflammation and overnutrition. Hormone sensitive lipase (HSL) is part of the canonical lipolytic pathway and critical for complete lipolysis. This study hypothesizes that ROS is a signal that integrates regulation of lipolysis by targeting HSL. Experiments were performed with human differentiated adipocytes from the subcutaneous depot. Antioxidants were employed as a tool to decrease ROS, and it was found that scavenging ROS with diphenyliodonium, N-acetyl cysteine, or resveratrol decreased lipolysis in adipocytes. HSL phosphorylation of a key serine residue, Ser552, as well as translocation of this enzyme from the cytosol to the lipid droplet upon lipolytic stimulation were both abrogated by scavenging ROS. The phosphorylation status of other serine residues on HSL were not affected. These findings are significant because they document that ROS contributes to the physiological regulation of lipolysis via an effect on translocation. Such regulation could be useful in developing new obesity therapies.     

Several agents and pathways regulate lipolysis in adipocytes

Adipose tissue is the only tissue capable of hydrolyzing its stores of triacylglycerol (TAG) and of mobilizing fatty acids and glycerol in the bloodstream so that they can be used by other tissues. The full hydrolysis of TAG depends on the activity of three enzymes, adipose triglyceride lipase (ATGL), hormone-sensitive lipase (HSL) and monoacylglycerol lipase, each of which possesses a distinct regulatory mechanism. Although more is known about HSL than about the other two enzymes, it has recently been shown that HLS and ATGL can be activated simultaneously, such that the mechanism that enables HSL to access the surface of lipid droplets also permits the stimulation of ATGL. The classical pathway of lipolysis activation in adipocytes is cAMP-dependent. The production of cAMP is modulated by G-protein-coupled receptors of the Gs/Gi family and cAMP degradation is regulated by phosphodiesterase. However, other pathways that activate TAG hydrolysis are currently under investigation. Lipolysis can also be started by G-protein-coupled receptors of the Gq family, through molecular mechanisms that involve phospholipase C, calmodulin and protein kinase C. There is also evidence that increased lipolytic activity in adipocytes occurs after stimulation of the mitogen-activated protein kinase pathway or after cGMP accumulation and activation of protein kinase G. Several agents contribute to the control of lipolysis in adipocytes by modulating the activity of HSL and ATGL. In this review, we have summarized the signalling pathways activated by several agents involved in the regulation of TAG hydrolysis in adipocytes.     

Comparative studies of the role of hormone-sensitive lipase and adipose triglyceride lipase in human fat cell lipolysis

Hormone-sensitive lipase (HSL) and adipose triglyceride lipase (ATGL) regulate adipocyte lipolysis in rodents. The purpose of this study was to compare the roles of these lipases for lipolysis in human adipocytes. Subcutaneous adipose tissue was investigated. HSL and ATGL protein expression were related to lipolysis in isolated mature fat cells. ATGL or HSL were knocked down by RNA interference (RNAi) or selectively inhibited, and effects on lipolysis were studied in differentiated preadipocytes or adipocytes derived from human mesenchymal stem cells (hMSC). Subjects were all women. There were 12 lean controls, 8 lean with polycystic ovary syndrome (PCOS), and 27 otherwise healthy obese subjects. We found that norepinephrine-induced lipolysis was positively correlated with HSL protein levels (P < 0.0001) but not with ATGL protein. Women with PCOS or obesity had significantly decreased norepinephrine-induced lipolysis and HSL protein expression but no change in ATGL protein expression. HSL knock down by RNAi reduced basal and catecholamine-induced lipolysis. Knock down of ATGL decreased basal lipolysis but did not change catecholamine-stimulated lipolysis. Treatment of hMSC with a selective HSL inhibitor during and/or after differentiation in adipocytes reduced basal lipolysis by 50%, but stimulated lipolysis was inhibited completely. In contrast to findings in rodents, ATGL is of less importance than HSL in regulating catecholamine-induced lipolysis and cannot replace HSL when this enzyme is continuously inhibited. However, both lipases regulate basal lipolysis in human adipocytes. ATGL expression, unlike HSL, is not influenced by obesity or PCOS.     

Regulation of lipolytic activity by long-chain acyl-coenzyme A in islets and adipocytes

Intracellular lipolysis is a major pathway of lipid metabolism that has roles, not only in the provision of free fatty acids as energy substrate, but also in intracellular signal transduction. The latter is likely to be particularly important in the regulation of insulin secretion from islet beta-cells. The mechanisms by which lipolysis is regulated in different tissues is, therefore, of considerable interest. Here, the effects of long-chain acyl-CoA esters (LC-CoA) on lipase activity in islets and adipocytes were compared. Palmitoyl-CoA (Pal-CoA, 1-10 microM) stimulated lipase activity in islets from both normal and hormone-sensitive lipase (HSL)-null mice and in phosphatase-treated islets, indicating that the stimulatory effect was neither on HSL nor phosphorylation dependent. In contrast, we reproduced the previously published observations showing inhibition of HSL activity by LC-CoA in adipocytes. The inhibitory effect of LC-CoA on adipocyte HSL was dependent on phosphorylation and enhanced by acyl-CoA-binding protein (ACBP). In contrast, the stimulatory effect on islet lipase activity was blocked by ACBP, presumably due to binding and sequestration of LC-CoA. These data suggest the following intertissue relationship between islets and adipocytes with respect to fatty acid metabolism, LC-CoA signaling, and lipolysis. Elevated LC-CoA in islets stimulates lipolysis to generate a signal to increase insulin secretion, whereas elevated LC-CoA in adipocytes inhibits lipolysis. Together, these opposite actions of LC-CoA lower circulating fat by inhibiting its release from adipocytes and promoting fat storage via insulin action.     

Region‐specific effects of oestradiol on adipose‐derived stem cell differentiation in post‐menopausal women

The goal of this study was to determine the effect of acute transdermal 17β‐oestradiol (E₂) on the adipogenic potential of subcutaneous adipose‐derived stem cells (ASC) in post‐menopausal women. Post‐menopausal women (n = 11; mean age 57 ± 4.5 years) were treated for 2 weeks, in a randomized, cross‐over design, with transdermal E₂ (0.15 mg) or placebo patches. Biopsies of abdominal (AB) and femoral (FEM) subcutaneous adipose tissue (SAT) were obtained after each treatment and mature adipocytes were analysed for cell size and ASC for their capacity for proliferation (growth rate), differentiation (triglyceride accumulation) and susceptibility to tumour necrosis factor alpha‐induced apoptosis. Gene expression of oestrogen receptors α and β (ESR1 and ESR2), perilipin 1 and hormone‐sensitive lipase (HSL), was also assessed. In FEM SAT, but not AB SAT, 2 weeks of E₂ significantly (P = 0.03) increased ASC differentiation and whole SAT HSL mRNA expression (P = 0.03) compared to placebo. These changes were not associated with mRNA expression of oestrogen receptors α and β, but HSL expression was significantly increased in FEM SAT with transdermal E₂ treatment. Adipose‐derived stem cells proliferation and apoptosis did not change in either SAT depot after E₂ compared with placebo. Short‐term E₂ appeared to increase the adipogenic potential of FEM, but not AB, SAT in post‐menopausal women with possible implications for metabolic disease. Future studies are needed to determine longer term impact of E₂ on regional SAT accumulation in the context of positive energy imbalance.     

Adipogenic and lipolytic effects of chronic glucocorticoid exposure

Glucocorticoids have been proposed to be both adipogenic and lipolytic in action within adipose tissue, although it is unknown whether these actions can occur simultaneously. Here we investigate both the in vitro and in vivo effects of corticosterone (Cort) on adipose tissue metabolism. Cort increased 3T3-L1 preadipocyte differentiation in a concentration-dependent manner, but did not increase lipogenesis in adipocytes. Cort increased lipolysis within adipocytes in a concentration-dependent manner (maximum effect at 1-10 μM). Surprisingly, removal of Cort further increased lipolytic rates (～320% above control, P < 0.05), indicating a residual effect on basal lipolysis. mRNA and protein expression of adipose triglyceride lipase and phosphorylated status of hormone sensitive lipase (Ser563/Ser660) were increased with 48 h of Cort treatment. To test these responses in vivo, Sprague-Dawley rats were subcutaneously implanted with wax pellets with/without Cort (300 mg). After 10 days, adipose depots were removed and cultured ex vivo. Both free fatty acids and glycerol concentrations were elevated in fed and fasting conditions in Cort-treated rats. Despite increased lipolysis, Cort rats had more visceral adiposity than sham rats (10.2 vs. 6.9 g/kg body wt, P < 0.05). Visceral adipocytes from Cort rats were smaller and more numerous than those in sham rats, suggesting that adipogenesis occurred through preadipocyte differentiation rather than adipocyte hypertrophy. Visceral, but not subcutaneous, adipocyte cultures from Cort-treated rats displayed a 1.5-fold increase in basal lipolytic rates compared with sham rats (P < 0.05). Taken together, our findings demonstrate that chronic glucocorticoid exposure stimulates both lipolysis and adipogenesis in visceral adipose tissue but favors adipogenesis primarily through preadipocyte differentiation.     

Control of adipose triglyceride lipase action by serine 517 of perilipin A globally regulates protein kinase A-stimulated lipolysis in adipocytes

Phosphorylation of the lipid droplet-associated protein perilipin A (Peri A) mediates the actions of cyclic AMP-dependent protein kinase A (PKA) to stimulate triglyceride hydrolysis (lipolysis) in adipocytes. Studies addressing how Peri A PKA sites regulate adipocyte lipolysis have relied on non-adipocyte cell models, which express neither adipose triglyceride lipase (ATGL), the rate-limiting enzyme for triglyceride catabolism in mice, nor the "downstream" lipase, hormone-sensitive lipase (HSL). ATGL and HSL are robustly expressed by adipocytes that we generated from murine embryonic fibroblasts of perilipin knock-out mice. Adenoviral expression of Peri A PKA site mutants in these cells reveals that mutation of serine 517 alone is sufficient to abrogate 95% of PKA (forskolin)-stimulated fatty acid (FA) and glycerol release. Moreover, a "phosphomimetic" (aspartic acid) substitution at serine 517 enhances PKA-stimulated FA release over levels obtained with wild type Peri A. Studies with ATGL-and HSL-directed small hairpin RNAs demonstrate that 1) ATGL activity is required for all PKA-stimulated FA and glycerol release in murine embryonic fibroblast adipocytes and 2) all PKA-stimulated FA release in the absence of HSL activity requires serine 517 phosphorylation. These results provide the first demonstration that Peri A regulates ATGL-dependent lipolysis and identify serine 517 as the Peri A PKA site essential for this regulation. The contributions of other PKA sites to PKA-stimulated lipolysis are manifested only in the presence of phosphorylated or phosphomimetic serine 517. Thus, serine 517 is a novel "master regulator" of PKA-stimulated adipocyte lipolysis.     

Perilipin promotes hormone-sensitive lipase-mediated adipocyte lipolysis via phosphorylation-dependent and -independent mechanisms

Hormone-sensitive lipase (HSL) is the predominant lipase effector of catecholamine-stimulated lipolysis in adipocytes. HSL-dependent lipolysis in response to catecholamines is mediated by protein kinase A (PKA)-dependent phosphorylation of perilipin A (Peri A), an essential lipid droplet (LD)-associated protein. It is believed that perilipin phosphorylation is essential for the translocation of HSL from the cytosol to the LD, a key event in stimulated lipolysis. Using adipocytes retrovirally engineered from murine embryonic fibroblasts of perilipin null mice (Peri-/- MEF), we demonstrate by cell fractionation and confocal microscopy that up to 50% of cellular HSL is LD-associated in the basal state and that PKA-stimulated HSL translocation is fully supported by adenoviral expression of a mutant perilipin lacking all six PKA sites (Peri Adelta1-6). PKA-stimulated HSL translocation was confirmed in differentiated brown adipocytes from perilipin null mice expressing an adipose-specific Peri Adelta1-6 transgene. Thus, PKA-induced HSL translocation was independent of perilipin phosphorylation. However, Peri Adelta1-6 failed to enhance PKA-stimulated lipolysis in either MEF adipocytes or differentiated brown adipocytes. Thus, the lipolytic action(s) of HSL at the LD surface requires PKA-dependent perilipin phosphorylation. In Peri-/- MEF adipocytes, PKA activation significantly enhanced the amount of HSL that could be cross-linked to and co-immunoprecipitated with ectopic Peri A. Notably, this enhanced cross-linking was blunted in Peri-/- MEF adipocytes expressing Peri Adelta1-6. This suggests that PKA-dependent perilipin phosphorylation facilitates (either direct or indirect) perilipin interaction with LD-associated HSL. These results redefine and expand our understanding of how perilipin regulates HSL-mediated lipolysis in adipocytes.     

Shrinking and development of lipid droplets in adipocytes during catecholamine-induced lipolysis

Time-lapse observation of adipocytes during catecholamine-induced lipolysis clearly shows that shrinking of existing lipid droplets (LDs) occurs in some adipocytes and that small LDs are newly developed in almost all cells. Immunofluorescence imaging reveals that activation and localization of hormone-sensitive lipase (HSL) on the surface of LDs, which are required for conferring maximal lipolysis, are necessary for the shrinking of the LDs. However, not all adipocytes in which phosphorylated HSL is localized on LDs exhibit shrinking of LDs. The simultaneous shrinking and development of LDs yield apparent fragmentation and dispersion of LDs in adipocytes stimulated with catecholamine.     

The influence of preadipocyte differentiation capacity on lipolysis in human mature adipocytes.

The ability of catecholamines to maximally stimulate adipocyte lipolysis (lipolytic capacity) is decreased in obesity. It is not known whether the lipolytic capacity is determined by the ability of adipocytes to differentiate. The aim of the study was to investigate if lipolytic capacity is related to preadipocyte differentiation and if the latter can predict lipolysis in mature adipocytes. IN VITRO experiments were performed on differentiating preadipocytes and isolated mature adipocytes from human subcutaneous adipose tissue. In preadipocytes, noradrenaline-induced lipolysis increased significantly until terminal differentiation (day 12). However, changes in the expression of genes involved in lipolysis (hormone sensitive lipase, adipocyte triglyceride lipase, the alpha2-and beta1-adrenoceptors, perilipin, and fatty acid binding protein) reached a plateau much earlier during differentiation (day 8). A significant positive correlation between lipolysis in differentiated preadipocytes and mature adipocytes was observed for noradrenaline (r=0.5, p<0.01). The late differentiation capacity of preadipocytes measured as glycerol-3-phosphate dehydrogenase activity was positively correlated with noradrenaline-induced lipolysis in preadipocytes (r=0.51, p<0.005) and mature fat cells (r=0.35, p<0.05). In conclusion, intrinsic properties related to terminal differentiation determine the ability of catecholamines to maximally stimulate lipolysis in fat cells. The inability to undergo full differentiation might in part explain the low lipolytic capacity of fat cells among the obese.     

Adipose triglyceride lipase regulates basal lipolysis and lipid droplet size in adipocytes

In adipocytes, lipid droplet (LD) size reflects a balance of triglyceride synthesis (lipogenesis) and hydrolysis (lipolysis). Perilipin A (Peri A) is the most abundant phosphoprotein on the surface of adipocyte LDs and has a crucial role in lipid storage and lipolysis. Adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL) are the major rate-determining enzymes for lipolysis in adipocytes. Each of these proteins (Peri A, ATGL, and HSL) has been demonstrated to regulate lipid storage and release in the adipocyte. However, in the absence of protein kinase A (PKA) stimulation (basal state), the lipases (ATGL and HSL) are located mainly in the cytoplasm, and their contribution to basal rates of lipolysis and influence on LD size are poorly understood. In this study, we utilize an adenoviral system to knockdown or overexpress ATGL and HSL in an engineered model system of adipocytes in the presence or absence of Peri A. We are able to demonstrate in our experimental model system that in the basal state, LD size, triglyceride storage, and fatty acid release are mainly influenced by the expression of ATGL. These results demonstrate for the first time the relative contributions of ATGL, HSL, and Peri A on determination of LD size in the absence of PKA stimulation.     

Calyculin and okadaic acid promote perilipin phosphorylation and increase lipolysis in primary rat adipocytes

Lipolysis is primarily regulated by protein kinase A (PKA), which phosphorylates perilipin and hormone-sensitive lipase (HSL), and causes translocation of HSL from cytosol to lipid droplets in adipocytes. Perilipin coats lipid droplet surface and assumes to prevent lipase access to triacylglycerols, thus inhibiting basal lipolysis; phosphorylated perilipin facilitates lipolysis on PKA activation. Here, we induced lipolysis in primary rat adipocytes by inhibiting protein serine/threonine phosphatase with specific inhibitors, okadaic acid and calyculin. The incubation with calyculin promotes incorporation of 32Pi into perilipins, thus, confirming that perilipin is hyperphosphorylated. The lipolysis response to calyculin is gradually accompanied by increased accumulation of phosphorylated perilipin A in a concentration- and time-responsive manner. When perilipin phosphorylation is abrogated by the addition of N-ethylmaleimide, lipolysis ceases. Different from a considerable translocation of HSL upon PKA activation with isoproterenol, calyculin does not alter HSL redistribution in primary or differentiated adipocytes, as confirmed by both immunostaining and immunoblotting. Thus, we suggest that inhibition of the phosphatase by calyculin activates lipolysis via promoting perilipin phosphorylation rather than eliciting HSL translocation in adipocytes. Further, we show that when the endogenous phosphatase is inhibited by calyculin, simultaneous PKA activation with isoproterenol converts most of the perilipin to the hyperphosphorylated species, and induces enhanced lipolysis. Apparently, as PKA phosphorylates perilipin and stimulates lipolysis, the phosphatase acts to dephosphorylate perilipin and attenuate lipolysis. This suggests a two-step strategy governed by a kinase and a phosphatase to modulate the steady state of perilipin phosphorylation and hence the lipolysis response to hormonal stimulation.     

Berberine attenuates cAMP-induced lipolysis via reducing the inhibition of phosphodiesterase in 3T3-L1 adipocytes

Berberine, a hypoglycemic agent, has been shown to decrease plasma free fatty acids (FFAs) level in insulin-resistant rats. In the present study, we explored the mechanism responsible for the antilipolytic effect of berberine in 3T3-L1 adipocytes. It was shown that berberine attenuated lipolysis induced by catecholamines, cAMP-raising agents, and a hydrolyzable cAMP analog, but not by tumor necrosis factor α and a nonhydrolyzable cAMP analog. Unlike insulin, the inhibitory effect of berberine on lipolysis in response to isoproterenol was not abrogated by wortmannin, an inhibitor of phosphatidylinositol 3-kinase, but additive to that of PD98059, an extracellular signal-regulated kinase kinase inhibitor. Prior exposure of adipocytes to berberine decreased the intracellular cAMP production induced by isoproterenol, forskolin, and 3-isobutyl-1-methylxanthine (IBMX), along with hormone-sensitive lipase (HSL) Ser-563 and Ser-660 dephosphorylation, but had no effect on perilipin phosphorylation. Berberine stimulated HSL Ser-565 as well as adenosine monophosphate-activated protein kinase (AMPK) phosphorylation. However, compound C, an AMPK inhibitor, did not reverse the regulatory effect of berberine on HSL Ser-563, Ser-660, and Ser-565 phosphorylation, nor the antilipolytic effect of berberine. Knockdown of AMPK using RNA interference also failed to restore berberine-suppressed lipolysis. cAMP-raising agents increased AMPK activity, which was not additive to that of berberine. Stimulation of adipocytes with berberine increased phosphodiesterase (PDE) 3B and PDE4 activity measured by hydrolysis of ³[H]cAMP. These results suggest that berberine exerts an antilipolytic effect mainly by reducing the inhibition of PDE, leading to a decrease in cAMP and HSL phosphorylation independent of AMPK pathway.     

Analysis of lipolysis in adipocytes using a fluorescent fatty acid derivative

For facilitation of the experimental analysis of the mechanism and regulation of mobilization of fatty acids from adipose triacylglycerol (TAG) stores, which also represents important targets for pharmacological intervention with the pathogenesis of diabetes and obesity, we developed a convenient and reliable non-radioactive cell-based assay. Isolated rat adipocytes are incubated with the fluorescent fatty acid derivative, 12-((7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)dodecanoic acid (NBD-FA), in the presence of insulin. The resulting NBD-FA-labeled TAG is efficiently cleaved by hormone-sensitive lipase (HSL) in vitro. After removal of insulin and excess of free NBD-FA, lipolysis is initiated by addition of isoproterenol and/or adenosine deaminase. The amount of NBD-FA generated in total or released into the incubation medium in the presence of modulatory hormones or compounds is then monitored by thin layer chromatography and fluorescence imaging. Release of NBD-FA, glycerol and [3H]oleic acid from TAG follows similar kinetics and concentration dependence in response to various lipolytic and anti-lipolytic stimuli as well as inhibitors of HSL. Release of NBD-FA from adipocytes correlates well to translocation of HSL from the cytosol to TAG droplets. In addition, we found that a cell-free system consisting of NBD-FA-labeled TAG droplets with endogenous associated HSL closely reflects the lipolytic state of the adipocytes used for its preparation. In conclusion, release of NBD-FA from TAG in vivo and in vitro can be used as accurate index for (regulation of) lipolysis in primary and cultured adipocytes.     

Subdivisions of subcutaneous abdominal adipose tissue and insulin resistance

Whereas truncal (central) adiposity is strongly associated with the insulin resistant metabolic syndrome, it is uncertain whether this is accounted for principally by visceral adiposity (VAT). Several recent studies find as strong or stronger association between subcutaneous abdominal adiposity (SAT) and insulin resistance. To reexamine the issue of truncal adipose tissue depots, we performed cross-sectional abdominal computed tomography, and we undertook the novel approach of partitioning SAT into the plane superficial to the fascia within subcutaneous adipose tissue (superficial SAT) and that below this fascia (deep SAT), as well as measurement of VAT. Among 47 lean and obese glucose-tolerant men and women, insulin-stimulated glucose utilization, measured by euglycemic clamp, was strongly correlated with both VAT and deep SAT (r = -0.61 and -0.64, respectively; both P < 0.001), but not with superficial SAT (r = -0.29, not significant). Also, VAT and deep SAT followed a highly congruent pattern of associations with glucose and insulin area under the curve (75-g oral glucose tolerance test), mean arterial blood pressure, apoprotein-B, high-density lipoprotein cholesterol, and triglyceride. Superficial SAT had markedly weaker association with all these parameters and instead followed the pattern observed for thigh subcutaneous adiposity. We conclude that there are two functionally distinct compartments of adipose tissue within abdominal subcutaneous fat and that the deep SAT has a strong relation to insulin resistance.