Major involvement of mTOR in the PPARγ-induced stimulation of adipose tissue lipid uptake and fat accretion

Evidence points to a role of the mammalian target of rapamycin (mTOR) signaling pathway as a regulator of adiposity, yet its involvement as a mediator of the positive actions of peroxisome proliferator-activated receptor (PPAR)γ agonism on lipemia, fat accretion, lipid uptake, and its major determinant lipoprotein lipase (LPL) remains to be elucidated. Herein we evaluated the plasma lipid profile, triacylglycerol (TAG) secretion rates, and adipose tissue LPL-dependent lipid uptake, LPL expression/activity, and expression profile of other lipid metabolism genes in rats treated with the PPARγ agonist rosiglitazone (15 mg/kg/day) in combination or not with the mTOR inhibitor rapamycin (2 mg/kg/day) for 15 days. Rosiglitazone stimulated adipose tissue mTOR complex 1 and AMPK and induced TAG-derived lipid uptake (136%), LPL mRNA/activity (2- to 6-fold), and fat accretion in subcutaneous (but not visceral) white adipose tissue (WAT; 50%) and in brown adipose tissue (BAT; 266%). Chronic mTOR inhibition attenuated the upregulation of lipid uptake, LPL expression/activity, and fat accretion induced by PPARγ activation in both subcutaneous WAT and BAT, which resulted in hyperlipidemia. In contrast, rapamycin did not affect most of the other WAT lipogenic genes upregulated by rosiglitazone. Together these findings demonstrate that mTOR is a major regulator of adipose tissue LPL-mediated lipid uptake and a critical mediator of the hypolipidemic and lipogenic actions of PPARγ activation. Evidence points to a role of the mammalian target of rapamycin (mTOR) signaling pathway as a regulator of adiposity, yet its involvement as a mediator of the positive actions of peroxisome proliferator-activated receptor (PPAR)γ agonism on lipemia, fat accretion, lipid uptake, and its major determinant lipoprotein lipase (LPL) remains to be elucidated. Herein we evaluated the plasma lipid profile, triacylglycerol (TAG) secretion rates, and adipose tissue LPL-dependent lipid uptake, LPL expression/activity, and expression profile of other lipid metabolism genes in rats treated with the PPARγ agonist rosiglitazone (15 mg/kg/day) in combination or not with the mTOR inhibitor rapamycin (2 mg/kg/day) for 15 days. Rosiglitazone stimulated adipose tissue mTOR complex 1 and AMPK and induced TAG-derived lipid uptake (136%), LPL mRNA/activity (2- to 6-fold), and fat accretion in subcutaneous (but not visceral) white adipose tissue (WAT; 50%) and in brown adipose tissue (BAT; 266%). Chronic mTOR inhibition attenuated the upregulation of lipid uptake, LPL expression/activity, and fat accretion induced by PPARγ activation in both subcutaneous WAT and BAT, which resulted in hyperlipidemia. In contrast, rapamycin did not affect most of the other WAT lipogenic genes upregulated by rosiglitazone. Together these findings demonstrate that mTOR is a major regulator of adipose tissue LPL-mediated lipid uptake and a critical mediator of the hypolipidemic and lipogenic actions of PPARγ activation. Activation of the nuclear receptor peroxisome proliferator-activated receptor (PPAR)γ, a master regulator of adipogenesis and adipocyte lipid metabolism (1.Rosen E.D. MacDougald O.A. Adipocyte differentiation from the inside out.Nat. Rev. Mol. Cell Biol. 2006; 7: 885-896Crossref PubMed Scopus (1941) Google Scholar, 2.Festuccia W.T. Deshaies Y. Depot specificities of PPARγ ligand actions on lipid and glucose metabolism and their implication in PPARγ-mediated body fat redistribution.Clinical Lipidology. 2009; 4: 633-642Crossref Scopus (8) Google Scholar), is associated with marked fat accretion in subcutaneous white adipose (WAT) and brown adipose (BAT) tissues. We have shown in rodent models that such fat accretion is mainly attributable to the vastly enhanced uptake and storage of circulating lipids due to increased expression of genes involved in lipid uptake and esterification (2.Festuccia W.T. Deshaies Y. Depot specificities of PPARγ ligand actions on lipid and glucose metabolism and their implication in PPARγ-mediated body fat redistribution.Clinical Lipidology. 2009; 4: 633-642Crossref Scopus (8) Google Scholar–6.Festuccia W.T. Blanchard P.G. Turcotte V. Laplante M. Sariahmetoglu M. Brindley D.N. Deshaies Y. Depot-specific effects of the PPARγ agonist rosiglitazone on adipose tissue glucose uptake and metabolism.J. Lipid Res. 2009; 50: 1185-1194Abstract Full Text Full Text PDF PubMed Scopus (65) Google Scholar). This in turn is thought to be largely responsible for the marked postprandial hypolipidemic action of PPARγ activation in these models, with some contribution from reduced liver VLDL secretion (7.Laplante M. Festuccia W.T. Soucy G. Blanchard P.G. Renaud A. Berger J.P. Olivecrona G. Deshaies Y. Tissue-specific postprandial clearance is the major determinant of PPARγ-induced triglyceride lowering in the rat.Am. J. Physiol. Regul. Integr. Comp. Physiol. 2009; 296: R57-R66Crossref PubMed Scopus (35) Google Scholar). Therefore, when solicited, the PPARγ pathway brings about an integrated set of metabolic adaptations that leads to fat deposition in metabolically safe adipose compartments, a concomitant reduction in circulating lipids, and less exposure of nonadipose tissues to lipotoxicity. This process is thought to contribute to the powerful insulin-sensitizing action of PPARγ agonists, such as thiazolidinediones (8.Larsen T.M. Toubro S. Astrup A. PPARγ agonists in the treatment of type II diabetes: is increased fatness commensurate with long-term efficacy?.Int. J. Obes. Relat. Metab. Disord. 2003; 27: 147-161Crossref PubMed Scopus (246) Google Scholar). Although of definite importance, PPARγ is only one of many key modulators of adiposity. More specifically, robust evidence points to an important role of the mammalian target of rapamycin (mTOR) signaling pathway as a possible regulator of adipose tissue mass. mTOR is a conserved serine-threonine kinase that controls protein synthesis; cell size and proliferation according to the availability of amino acids; growth factors; nutrients; and cell energy status (9.Laplante M. Sabatini D.M. mTOR signaling at a glance.J. Cell Sci. 2009; 122: 3589-3594Crossref PubMed Scopus (1642) Google Scholar). mTOR is the catalytic core of two distinct multiprotein complexes, mTOR complex 1 (mTORC1) and 2 (mTORC2), that have different downstream targets, biological functions, and sensitivity to inhibition by the bacterial macrolide rapamycin. Whereas mTORC1 activity is broadly inhibited by rapamycin, mTORC2 is negatively affected by this molecule after prolonged treatment and in certain cell types only (10.Sarbassov D.D. Ali S.M. Sengupta S. Sheen J.H. Hsu P.P. Bagley A.F. Markhard A.L. Sabatini D.M. Prolonged rapamycin treatment inhibits mTORC2 assembly and Akt/PKB.Mol. Cell. 2006; 22: 159-168Abstract Full Text Full Text PDF PubMed Scopus (2178) Google Scholar). With regard to adiposity, expansion of fat mass in obesity, for example, is associated with marked activation of mTOR in adipose tissue (11.Um S.H. Frigerio F. Watanabe M. Picard F. Joaquin M. Sticker M. Fumagalli S. Allegrini P.R. Kozma S.C. Auwerx J. et al.Absence of S6K1 protects against age- and diet-induced obesity while enhancing insulin sensitivity.Nature. 2004; 431: 200-205Crossref PubMed Scopus (1358) Google Scholar), whereas fat mass retraction due to caloric restriction and fasting is associated with adipose tissue mTOR inhibition. Accordingly, chronic pharmacological or genetic inhibition of the mTORC1 signaling pathway is associated with a reduction in adipose tissue mass due to both reduced adipocyte size and number (11.Um S.H. Frigerio F. Watanabe M. Picard F. Joaquin M. Sticker M. Fumagalli S. Allegrini P.R. Kozma S.C. Auwerx J. et al.Absence of S6K1 protects against age- and diet-induced obesity while enhancing insulin sensitivity.Nature. 2004; 431: 200-205Crossref PubMed Scopus (1358) Google Scholar–13.Polak P. Cybulski N. Feige J.N. Auwerx J. Ruegg M.A. Hall M.N. Adipose-specific knockout of raptor results in lean mice with enhanced mitochondrial respiration.Cell Metab. 2008; 8: 399-410Abstract Full Text Full Text PDF PubMed Scopus (386) Google Scholar). Despite the direct association between mTOR activity and adiposity, little is known of the mechanisms by which mTOR modulates fat mass. Importantly, evidence suggests that mTOR may affect adiposity by modulating the activity of PPARγ. Pharmacological mTOR inhibition, for example, impairs in vitro preadipocyte differentiation into mature adipocytes through PPARγ inhibition (14.Kim J.E. Chen J. Regulation of peroxisome proliferator-activated receptor-γ activity by mammalian target of rapamycin and amino acids in adipogenesis.Diabetes. 2004; 53: 2748-2756Crossref PubMed Scopus (363) Google Scholar), an effect completely reversed by the presence of a synthetic PPARγ ligand. Likewise, in vivo rapamycin treatment reduces adipose tissue expression of several PPARγ target genes (12.Houde V.P. Brule S. Festuccia W.T. Blanchard P.G. Bellmann K. Deshaies Y. Marette A. Chronic rapamycin treatment causes glucose intolerance and hyperlipidemia by upregulating hepatic gluconeogenesis and impairing lipid deposition in adipose tissue.Diabetes. 2010; 59: 1338-1348Crossref PubMed Scopus (334) Google Scholar). The above evidence points to a possible cross-talk between PPARγ and mTOR; however, to our knowledge such interaction has not yet been explored directly in the in vivo setting. Considering that ligand-mediated PPARγ activation induces subcutaneous and brown fat accretion in vivo and that mTOR appears as an important regulator of adiposity, we tested in the present study the hypothesis that mTOR is a major mediator of the increased lipid uptake, fat accretion, and resulting reduction in lipemia induced by ligand-mediated PPARγ activation in vivo. To this end, rats treated with the PPARγ agonist rosiglitazone in combination or not with the mTORC1 inhibitor rapamycin were evaluated for plasma lipids; triacylglycerol (TAG) secretion; hydrolysis and uptake by adipose tissues; activity and expression of lipoprotein lipase (LPL), the major determinant of lipoprotein-derived fatty acid uptake; and expression of several other key genes involved in fatty acid uptake and deposition. Animal handling and treatment were approved by the Animal Care and Handling Committee of Laval University. Male Sprague-Dawley rats (200 g) purchased from Charles River Laboratories (St-Constant, QC, Canada) were housed individually in a room kept at 23 ± 1°C with a 12:12 h light-dark cycle. After a 4-day adaptation period, rats were matched by weight and divided into four groups: control, vehicle; rosiglitazone, vehicle; control, rapamycin; and rosiglitazone, rapamycin. Vehicle (0.1% Me2SO, 0.2% carboxymethylcellulose) or rapamycin (LC laboratories, Woburn, MA) (2 mg/kg/day) were injected intraperitoneally once daily. The dose of rapamycin was chosen based on previous studies showing its efficiency to completely block the mTOR pathway in rats and mice, a dose within the range of those used in human studies (15.Maeda K. Shioi T. Kosugi R. Yoshida Y. Takahashi K. Machida Y. Izumi T. Rapamycin ameliorates experimental autoimmune myocarditis.Int. Heart J. 2005; 46: 513-530Crossref PubMed Scopus (25) Google Scholar, 16.Shioi T. McMullen J.R. Tarnavski O. Converso K. Sherwood M.C. Manning W.J. Izumo S. Rapamycin attenuates load-induced cardiac hypertrophy in mice.Circulation. 2003; 107: 1664-1670Crossref PubMed Scopus (393) Google Scholar). Rats were fed a nonpurified powdered rodent diet (Charles River Rodent Diet #5075, Woodstock, ON, Canada) alone (control) or supplemented with the PPARγ agonist rosiglitazone (AVANDIA) at a dose of 15 mg/kg/day for 15 days. This dose of rosiglitazone was found in previous studies to be associated with subcutaneous fat accretion and improvement in the plasma lipid profile (3.Laplante M. Sell H. MacNaul K.L. Richard D. Berger J.P. Deshaies Y. PPAR-γ activation mediates adipose depot-specific effects on gene expression and lipoprotein lipase activity: mechanisms for modulation of postprandial lipemia and differential adipose accretion.Diabetes. 2003; 52: 291-299Crossref PubMed Scopus (134) Google Scholar). After 15 days of treatment, rats were euthanized by decapitation for tissue and blood harvesting after a 12 h fasting period followed or not by 3 h of ad libitum refeeding. Plasma adiponectin was measured by ELISA following supplier's recommendations (ALPCO Diagnostics, Salem, NH). Plasma TAG (Roche Diagnostics, Montreal, QC, Canada) and nonesterified fatty acids (NEFA) (Wako Chemicals, Richmond, VA) levels were measured by enzymatic methods according to the manufacturer's instructions. Tissue samples were homogenized in buffer, subjected to SDS-PAGE, and transferred to nitrocellulose membranes as previously described (12.Houde V.P. Brule S. Festuccia W.T. Blanchard P.G. Bellmann K. Deshaies Y. Marette A. Chronic rapamycin treatment causes glucose intolerance and hyperlipidemia by upregulating hepatic gluconeogenesis and impairing lipid deposition in adipose tissue.Diabetes. 2010; 59: 1338-1348Crossref PubMed Scopus (334) Google Scholar). Antibodies used for immunoblotting are listed in supplementary Table I. Densitometric analysis was performed with ImageQuant TL software (GE Healthcare, Little Chalfont, United Kingdom). The following procedure was carried out to assess the contribution to triglyceridemia of TAG rate of entry into the circulation from the intestine and liver (chylomicron- + VLDL-bound TAG). After a 12 h fast and 3 h of refeeding the habitual diet, an initial blood sample (0.15 ml) was withdrawn from the tail vein in an EDTA-containing syringe, and rats were injected through the tail vein with 1 ml/kg Triton WR-1339 (300 mg/ml saline; Sigma-Aldrich, St. Louis, MO), a detergent that prevents intravascular TAG hydrolysis (17.Otway S. Robinson D.S. The use of a non-ionic detergent (Triton WR 1339) to determine rates of triglyceride entry into the circulation of the rat under different physiological conditions.J. Physiol. 1967; 190: 321-332Crossref PubMed Scopus (217) Google Scholar). Blood samples (0.15 ml) were then taken 20, 40, and 60 min after the injection. Rats were then injected with a lethal dose of ketamine-xylazine. Blood samples were centrifuged and plasma was collected and stored at 20°C for later TAG quantification. The rate of appearance of TAG in the circulation was determined from regression analysis of TAG accumulation in plasma versus time corrected for plasma volume estimated from body weight and was expressed as micromoles per minute (7.Laplante M. Festuccia W.T. Soucy G. Blanchard P.G. Renaud A. Berger J.P. Olivecrona G. Deshaies Y. Tissue-specific postprandial clearance is the major determinant of PPARγ-induced triglyceride lowering in the rat.Am. J. Physiol. Regul. Integr. Comp. Physiol. 2009; 296: R57-R66Crossref PubMed Scopus (35) Google Scholar). Ex vivo measurement of [3H]TAG hydrolysis and incorporation of generated [3H]fatty acids into adipose tissue lipids was performed as previously described (18.Faraj M. Sniderman A.D. Cianflone K. ASP enhances in situ lipoprotein lipase activity by increasing fatty acid trapping in adipocytes.J. Lipid Res. 2004; 45: 657-666Abstract Full Text Full Text PDF PubMed Scopus (63) Google Scholar, 19.Faraj M. Cianflone K. Differential regulation of fatty acid trapping in mouse adipose tissue and muscle by ASP.Am. J. Physiol. Endocrinol. Metab. 2004; 287: E150-E159Crossref PubMed Scopus (35) Google Scholar). Adipose tissue explants were cut into small pieces (25–35 mg), weighed, and preincubated for 10 min with 1 ml of Krebs Ringer buffer [pH 7.2, composed of (in mM) 5 glucose, 0.51 MgCl2, 4.56 KCl, 119.8 NaCl, 0.7 Na2HPO4, 1.3 NaH2PO4, and 15.0 NaHCO3, and 1% fatty acid-free BSA]. Fat pieces were then incubated for 4 h in a well containing 1 ml of [3H]TAG-rich lipoprotein substrate prepared as previously described (20.Lago R.M. Singh P.P. Nesto R.W. Congestive heart failure and cardiovascular death in patients with prediabetes and type 2 diabetes given thiazolidinediones: a meta-analysis of randomised clinical trials.Lancet. 2007; 370: 1129-1136Abstract Full Text Full Text PDF PubMed Scopus (586) Google Scholar). Briefly, triolein (1.41 mM), phosphatidylcholine (0.08 mM), and [3H]triolein (2.5 μCi, specific activity 1.77 μCi/mM TAG) were emulsified by sonication in an aqueous buffer (0.54 M Tris-HCl, pH 7.2, 5.1% BSA, and 7.5% fasting porcine serum) into large phospholipid micelles with a TAG-to-phospholipid molar ratio similar to that of chylomicron particles. Incubations were conducted in a shaking water bath at 37°C under 95% O2–5% CO2 atmosphere. The adipose tissue pieces were washed twice with 2 ml of ice-cold Krebs Ringer buffer and extracted overnight at room temperature with 1 ml of heptane-isopropanol (2:3). The organic solvent was evaporated, and lipids were resuspended in chloroform-methanol (2:1, v/v). Total tissue 3H-lipids were quantified by direct counting of the tissue lipid extract. The activity of LPL, the enzyme responsible for the intracapillary hydrolysis of circulating lipoprotein-bound TAG, was measured by incubating 100 μl of adipose depot homogenates for 1 h at 28°C with 100 μl of a substrate mixture consisting of 0.2 mol/l Tris-HCl buffer, pH 8.6, which contained 10 MBq/l [carboxyl-14C]triolein (Amersham, Oakville, ON, Canada) and 2.52 mmol/l cold triolein emulsified in 50 g/l gum arabic, as well as 20 g/l fatty acid-free BSA, 10% porcine serum as a source of apolipoprotein C-II, and either 0.2 or 2 M NaCl. Free oleate released by LPL was then separated from intact triolein, and sample 14C radioactivity was determined in a scintillation counter. LPL activity was calculated by subtracting lipolytic activity determined in a final NaCl concentration of 1 M (non-LPL activity) from total lipolytic activity measured in a final NaCl concentration of 0.1 M. LPL activity was expressed as microunits (1 µU = 1 μmol NEFA released per hour of incubation at 28°C). RNA extraction and quantitative PCR analysis were performed as described previously (21.Festuccia W.T. Laplante M. Berthiaume M. Gelinas Y. Deshaies Y. PPARγ agonism increases rat adipose tissue lipolysis, expression of glyceride lipases, and the response of lipolysis to hormonal control.Diabetologia. 2006; 49: 2427-2436Crossref PubMed Scopus (111) Google Scholar). In addition to LPL, we quantified expression levels of key genes involved in NEFA uptake (including those generated by LPL-mediated TAG hydrolysis), intracellular trafficking and esterification [FAT/CD36, fatty acid-transport protein 1 (FATP1), fatty acid binding protein 4 (FABP4, also known as aP2), glycerol kinase (GyK), and phosphoenolpyruvate carboxykinase (PEPCK)], PPARγ1 itself, and two additional PPARγ target genes [fatty acid synthase (FAS) and the glucose transporter GLUT4]. The primers used are listed in supplementary Table II. Data are expressed as the ratio between the expression of the target gene and the housekeeping gene 36B4 (also known as ARBP), the expression of which was not significantly affected by either rosiglitazone or rapamycin treatments. Results are expressed as means ± SE. Multifactorial ANOVA followed by Newman-Keuls multiple-range test was used for multiple comparisons. P < 0.05 was taken as the threshold of significance. First, we evaluated treatment effects on the activation state of relevant signaling pathways, including mTORC1 and 2 and AMPK (activated by PPARγ and a known inhibitor of mTOR activity). As depicted in Fig. 1A, rosiglitazone significantly activated inguinal adipose tissue mTORC1 as evidenced by the increased ratio of p-S6(Ser240/4)/S6, a downstream target protein in the mTORC1 signaling pathway. Rosiglitazone also markedly reduced the ratio of p-Akt(Thr308)/Akt (Fig. 1C) without affecting that of the mTORC2 substrate p-Akt(Ser473)/Akt (Fig. 1B). Concomitant with activation of inguinal adipose tissue mTORC1 by rosiglitazone, there was a marked stimulation of AMPK, as evidenced by the increased content of total and p-AMPK(Thr172) (Fig. 1D). As previously reported (12.Houde V.P. Brule S. Festuccia W.T. Blanchard P.G. Bellmann K. Deshaies Y. Marette A. Chronic rapamycin treatment causes glucose intolerance and hyperlipidemia by upregulating hepatic gluconeogenesis and impairing lipid deposition in adipose tissue.Diabetes. 2010; 59: 1338-1348Crossref PubMed Scopus (334) Google Scholar), rapamycin inhibited adipose tissue mTORC1 and 2 as seen through reduced p-S6/S6 and p-Akt(Ser473)/Akt ratios without affecting p-Akt(Thr308). No effect of rapamycin, however, was seen on inguinal adipose tissue total AMPK content. Simultaneous administration of rosiglitazone and rapamycin completely blocked p-S6 upregulation and attenuated the increased p-AMPK induced by rosiglitazone in inguinal adipose tissue. As in inguinal WAT, rapamycin attenuated mTORC1 and 2 signaling and abolished p-S6 upregulation associated with rosiglitazone treatment in retroperitoneal WAT and BAT (supplementary Fig. I). Interestingly, rosiglitazone alone inhibited mTORC2 in BAT as evidenced by p-Akt(Ser473)/Akt, but it had no effect on the activity of this protein complex in subcutaneous and visceral WAT. As previously reported (22.Choo A.Y. Yoon S.O. Kim S.G. Roux P.P. Blenis J. Rapamycin differentially inhibits S6Ks and 4E–BP1 to mediate cell-type-specific repression of mRNA translation.Proc. Natl. Acad. Sci. USA. 2008; 105: 17414-17419Crossref PubMed Scopus (651) Google Scholar), supplementary Fig. I confirms the differential action of rapamycin on S6 versus 4EBP. To further assess tissue specificity of treatment actions, the above pathways were also evaluated in the liver (supplementary Fig. II). As previously described (12.Houde V.P. Brule S. Festuccia W.T. Blanchard P.G. Bellmann K. Deshaies Y. Marette A. Chronic rapamycin treatment causes glucose intolerance and hyperlipidemia by upregulating hepatic gluconeogenesis and impairing lipid deposition in adipose tissue.Diabetes. 2010; 59: 1338-1348Crossref PubMed Scopus (334) Google Scholar), rapamycin treatment inhibited mTOR signaling in the liver without affecting Akt phosphorylation (Thr308 and Ser473) (supplementary Fig. II, A–C). This was associated with activation of AMPK as revealed by an increase in p-AMPK/AMPK (supplementary Fig. II, D). In contrast to adipose tissue, rosiglitazone treatment did not increase mTOR signaling in the liver. Surprisingly, the combination of both drugs abolished the activation of AMPK induced by rapamycin alone. The above findings indicate that some of the effects of PPARγ activation and mTOR inhibition appear to be specific to selected adipose tissue depots, rather than generalized actions. Rats treated with rosiglitazone had higher body weight gain (13%) and showed a weak tendency to have higher food intake and efficiency than control vehicle-treated rats (Table 1), confirming previous studies (23.Festuccia W.T. Oztezcan S. Laplante M. Berthiaume M. Michel C. Dohgu S. Denis R.G. Brito M.N. Brito N.A. Miller D.S. et al.Peroxisome proliferator-activated receptor-γ-mediated positive energy balance in the at is associated with reduced sympathetic drive to adipose tissues and thyroid status.Endocrinology. 2008; 149: 2121-2130Crossref PubMed Scopus (100) Google Scholar). Rapamycin treatment, on the other hand, markedly reduced body weight gain (–87%) in control and rosiglitazone-treated rats, an effect due to a reduction in both food intake (–16%) and food efficiency (–82%).TABLE 1Body weight gain, food intake, and relative adipose depot masses corrected for body weight of rats treated with rapamycin (Rapa) and/or rosiglitazone (RSG) for 15 daysControlRapaRSGRapa + RSGBody weight gain (g)98.3 ± 2.7aMeans not sharing a common superscript are significantly different from each other, P < 0.05.12.6 ± 2.3bMeans not sharing a common superscript are significantly different from each other, P < 0.05.111.2 ± 2.7cMeans not sharing a common superscript are significantly different from each other, P < 0.05.9.61 ± 3.0bMeans not sharing a common superscript are significantly different from each other, P < 0.05.Food intake (g)214 ± 9.6aMeans not sharing a common superscript are significantly different from each other, P < 0.05.179 ± 10.1bMeans not sharing a common superscript are significantly different from each other, P < 0.05.242 ± 12.0aMeans not sharing a common superscript are significantly different from each other, P < 0.05.167 ± 8.6bMeans not sharing a common superscript are significantly different from each other, P < 0.05.Food efficiencydCalculated as grams of body weight gain per 100 g of food ingested. (%)65.3 ± 1.2aMeans not sharing a common superscript are significantly different from each other, P < 0.05.11.6 ± 2.3bMeans not sharing a common superscript are significantly different from each other, P < 0.05.71.6 ± 2.1aMeans not sharing a common superscript are significantly different from each other, P < 0.05.13.6 ± 2.6bMeans not sharing a common superscript are significantly different from each other, P < 0.05.Retroperitoneal fat (%)0.87 ± 0.05aMeans not sharing a common superscript are significantly different from each other, P < 0.05.0.53 ± 0.04bMeans not sharing a common superscript are significantly different from each other, P < 0.05.0.84 ± 0.05aMeans not sharing a common superscript are significantly different from each other, P < 0.05.0.79 ± 0.04aMeans not sharing a common superscript are significantly different from each other, P < 0.05.Inguinal fat (%)1.06 ± 0.08aMeans not sharing a common superscript are significantly different from each other, P < 0.05.0.90 ± 0.04aMeans not sharing a common superscript are significantly different from each other, P < 0.05.1.58 ± 0.06bMeans not sharing a common superscript are significantly different from each other, P < 0.05.1.26 ± 0.04cMeans not sharing a common superscript are significantly different from each other, P < 0.05.Brown fat (%)0.09 ± 0.01aMeans not sharing a common superscript are significantly different from each other, P < 0.05.0.08 ± 0.01aMeans not sharing a common superscript are significantly different from each other, P < 0.05.0.33 ± 0.02bMeans not sharing a common superscript are significantly different from each other, P < 0.05.0.18 ± 0.02cMeans not sharing a common superscript are significantly different from each other, P < 0.05.Adiponectin (μg/ml)2.44 ± 0.25aMeans not sharing a common superscript are significantly different from each other, P < 0.05.3.17 ± 0.28aMeans not sharing a common superscript are significantly different from each other, P < 0.05.9.38 ± 1.01bMeans not sharing a common superscript are significantly different from each other, P < 0.05.6.66 ± 1.45cMeans not sharing a common superscript are significantly different from each other, P < 0.05.Data are average ± SE, n = 12–20 rats. All groups had the same average body weight (230 g) at the onset of treatments.a,b,c Means not sharing a common superscript are significantly different from each other, P < 0.05.d Calculated as grams of body weight gain per 100 g of food ingested. Open table in a new tab Data are average ± SE, n = 12–20 rats. All groups had the same average body weight (230 g) at the onset of treatments. Because rapamycin dramatically affects growth and energy balance in rats, fat depot masses were expressed relative to body weight to minimize the impact of these rapamycin effects on the interpretation of the changes in adiposity. The higher body weight gain of rosiglitazone-treated rats was associated with an increase in relative adiposity (Fig. 2A), which could be mainly attributed to an enhanced fat accretion in the subcutaneous inguinal WAT depot (50%) and interscapular BAT (266%) (Table 1). As expected (3.Laplante M. Sell H. MacNaul K.L. Richard D. Berger J.P. Deshaies Y. PPAR-γ activation mediates adipose depot-specific effects on gene expression and lipoprotein lipase activity: mechanisms for modulation of postprandial lipemia and differential adipose accretion.Diabetes. 2003; 52: 291-299Crossref PubMed Scopus (134) Google Scholar, 4.Laplante M. Festuccia W.T. Soucy G. Gelinas Y. Lalonde J. Berger J.P. Deshaies Y. Mechanisms of the depot specificity of peroxisome proliferator-activated receptor γ action on adipose tissue metabolism.Diabetes. 2006; 55: 2771-2778Crossref PubMed Scopus (111) Google Scholar, 5.Festuccia W.T. Blanchard P.G. Turcotte V. Laplante M. Sariahmetoglu M. Brindley D.N. Richard D. Deshaies Y. The PPARgamma agonist rosiglitazone enhances rat brown adipose tissue lipogenesis from glucose without altering glucose uptake.Am. J. Physiol. Regul. Integr. Comp. Physiol. 2009; 296: R1327-R1335Crossref PubMed Scopus (52) Google Scholar, 6.Festuccia W.T. Blanchard P.G. Turcotte V. Laplante M. Sariahmetoglu M. Brindley D.N. Deshaies Y. Depot-specific effects of the PPARγ agonist rosiglitazone on adipose tissue glucose uptake and metabolism.J. Lipid Res. 2009; 50: 1185-1194Abstract Full Text Full Text PDF PubMed Scopus (65) Google Scholar), no effect of rosiglitazone was seen on visceral retroperitoneal WAT mass (Table 1). Rapamycin, on the other hand, significantly reduced relative adiposity (–20%, Fig. 2A) and retroperitoneal mass (–40%), and it attenuated the upregulation induced by rosiglitazone in relative adiposity and weights of inguinal WAT and BAT. Intriguingly, retroperitoneal WAT mass was preserved to control values in rats treated with both drugs. The upregulation of subcutaneous fat accretion by rosiglitazone treatment was associated with an increase in plasma adiponectin levels (2.8-fold), which was partially attenuated by rapamycin (Table 1). As depicted in Fig. 2, the actions of rosiglitazone and rapamycin on plasma lipids were strongly dependent upon the nutritional status. In fasting conditions (12 h ove