- Email: [email protected]
PPARγ fundamental role in adipogenesis
International Congress Series 1262 (2004) 47 – 50
www.ics-elsevier.com
PPARg fundamental role in adipogenesis Terrie-Anne Cock a, Johan Auwerx a,b,* a
Institut de Ge´ne´tique et de Biologie Mole´culaire et Cellulaire, CNRS/INSERM/Universite´ Louis Pasteur, 67404 Illkirch, France b Institut Clinique de la Souris, Ge´nopole Strasbourg, 67404 Illkirch, France
Abstract. The peroxisomeproliferator-activated receptor gamma (PPARg) is a nuclear receptor that integrates the control of lipid and glucose homeostasis. The actions of PPARg are mediated by two protein isoforms, the widely expressed PPARg1 and adipose tissue-restricted PPARg2 with an additional 28 amino acids in the NH2-terminus. The effects of the absence of PPARg has not been extensively characterized in vivo since the PPARg mice are embryonic lethal, therefore we generated mice that carry a hypomorphic mutation at the PPARg2 locus, yielding a WAT-specific PPARg knockdown. The PPARg hyp/hyp mouse model demonstrates that PPARg is the master regulator of adipogenesis. Furthermore this model highlights PPARg fundamental roles in whole body physiology to maintain the metabolic and endocrine functions of white adipose tissue. D 2003 Elsevier B.V. All rights reserved. Keywords: Peroxisome proliferator-activated receptor gamma; PPARg; Nuclear receptor; Lipid and glucose metabolism; Mouse model; Adipogenesis; Metabolism
1. Introduction The peroxisome proliferator-activated receptor gamma (PPARg) is a prototypical metabolic nuclear receptor involved in the transcriptional control of energy, lipid, and glucose homeostasis. The activity of PPARg is governed by binding of small lipophilic ligands, mainly fatty acids, derived from nutrition or metabolic pathways, or synthetic agonists, like thiazolidinediones. Docking of these ligands in the ligand-binding pocket alters the conformation of PPARg resulting in transcriptional activation subsequent to the release of corepressors and the recruitment of coactivators. The actions of PPARg are mediated by two protein isoforms, the widely expressed PPARg1 and adipose tissuerestricted PPARg2, both produced from a single gene by alternative splicing and dif-
* Corresponding author. Institut de Ge´ne´tique et de Biologie Mole´culaire et Cellulaire, Orphan Nuclear Receptors, B.P. 10142, 1 rue Laurent Fries, F-67404 Illkirch, France. Tel.: +33-3-88-65-3425; fax: +33-3-88-653200. E-mail address: [email protected] (J. Auwerx). 0531-5131/ D 2003 Elsevier B.V. All rights reserved. doi:10.1016/S0531-5131(03)01767-9
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T.-A. Cock, J. Auwerx / International Congress Series 1262 (2004) 47–50
fering only by an additional 28 amino acids in the NH2-terminus of PPARg2 (reviewed Ref. [1]). 2. PPARg the master regulator of adipogenesis The initial suggestion that PPARg stimulated adipogenesis was based on the observation that overexpression of PPARg in cells was by itself sufficient to induce adipocyte differentiation [2]. Targeted mutagenesis of the PPARg gene in ES cells and knocking down the endogenous PPARg2 in cell lines confirmed the commanding role of PPARg in adipocyte differentiation [3,4]. Consistent with this, PPARg increases the expression of genes that promote fatty acid storage, whereas it represses genes that induce lipolysis in adipocytes. The lipodystrophy in the sole PPARg / mice, rescued by tetraploid aggregation [5], and the characterization of mice chimeric for PPARg / ES cells [3], further underscored the importance of PPARg in adipose tissue development in vivo. Human genetic studies further support the important role of PPARg in adipogenesis. One of the loci with suggestive linkage to obesity in the Pima Indians maps close to the location of PPARG on 3p25-p24[6]. Furthermore, subjects with the partial ‘‘loss-offunction’’ Pro12Ala mutation in the PPARg2-specific B exon have a lower body mass index (BMI), whereas carriers of the dominant negative Pro467Leu and Val290Met mutations or the Phe388Leu mutation that has a reduced affinity to ligand have partial lipodystrophy. Conversely, carriers of the Pro115Gln substitution, that renders PPARg constitutively active, are extremely obese [7,8]. A final line in favor of a role of PPARg in adipogenesis comes from pharmacological studies, which demonstrate that PPARg agonists, such as the thiazolidinediones, invariably increase WAT mass, whereas suboptimal PPARg activation, or PPARg antagonism, is neutral or even reverses weight gain [7]. The effects of the absence of PPARg has not been extensively characterized in vivo since the PPARg / mice die during intra-uterine development due to defects in cardiac and placental development in vivo [9 –11]. Therefore, we generated by homologous recombination, mice that carry a hypomorphic mutation at the PPARg2 locus, yielding a WAT-specific PPARg knockdown (PPARghyp/hyp) [12]. These mice have virtually no expression of PPARg in the sparse white adipose tissue (WAT), but not in other tissues. One-week-old mice with this WAT-specific PPARg knockdown display a lipodystrophic phenotype similar to that of humans with congenital generalized lipodystrophy (CGL). In CGL, the adipose deficiency normally is accompanied by severe insulin resistance, hyperglycemia, and fatty liver throughout life [13]. The neonatal PPARghyp/hyp phenotype mirrors most of these CGL characteristics, whereas the adult PPARghyp/hyp mice overcome some of the complications associated with the absence of WAT. In fact, adult PPARghyp/hyp mice do not have a fatty liver or show signs of hyperlipidemia [12]. The PPARghyp/hyp mice are furthermore only mildly glucose intolerant, which is distinct from most other lipodystrophy models [14 – 17]. This suggests that other physiological compensatory mechanisms ameliorate metabolic homeostasis in the PPARghyp/hyp mice. Overall the PPARghyp/hyp mouse model reveals that the PPARg locus is the critical regulator of adipogenesis in vivo [12].
T.-A. Cock, J. Auwerx / International Congress Series 1262 (2004) 47–50
49
3. WAT’s place in the scheme of energy metabolism The comparison of the physiological abnormalities in PPARghyp/hyp and PPARg heterozygote mice provides further insights into how the actions of PPARg in adipose tissue, liver, muscle and heart coordinate glucose and lipid homeostasis. Ubiquitous reduction of PPARg in all tissues, such as the PPARg heterozygote mice, results in normal body weight; these mice are insulin sensitive and resistant to diet-induced obesity [9,11]. In these PPARg heterozygote mice, circulating lipids can be stored in WAT, which avoids lipid overload in other tissues hence maintaining normal glucose and lipid metabolism [12]. In comparison, the PPARghyp/hyp mice with reduced PPARg levels in WAT have almost no WAT and are mildly glucose intolerant. Our study therefore implicates WAT as an important target tissue for PPARg activation to maintain glucose tolerance. Treatment with thiazolidinedione PPARg agonists can overcome the glucose intolerance in PPARghyp/hyp mice, although insulin resistance is not corrected. The PPARg agonist results from our WAT PPARghyp/hyp mice together with recent data in liver and muscle specific PPARg knockout mice indicate that primarily the WAT and not the liver and muscle are crucial for the protective effects of PPARg agonist on insulin resistance [12,18 –20]. 4. Selective modulation is the key The near absence of PPARg in WAT in adult PPARghyp/hyp mice is only causing a slightly abnormal metabolic profile and is rather well tolerated. Taken all together this lends further support to the hypothesis that selective PPARg modulators (or SPRMs) with a partial agonist or antagonist profile, rather than full agonists (reviewed in Ref. [7]), could be the preferred therapeutic strategy to treat the metabolic disorders associated with obesity, such as the metabolic syndrome, insulin resistance, or type 2 diabetes mellitus. The genetic evidence obtained in the PPARghyp/hyp mice arguments that selective PPARg modulators which have a reduced adipogenic drive, but which maintain agonist activity in other tissues, might maintain glucose homeostasis in such metabolic disorders. Acknowledgements This work was supported by grants from CNRS, INSERM, Hoˆpitaux Universitaires de Strasbourg, European Union, and NIH. References [1] J. Auwerx, PPARg, the ultimate thrifty gene, Diabetologia 42 (42) (1999) 1033 – 1049. [2] P. Tontonoz, E. Hu, B.M. Spiegelman, Stimulation of adipogenesis in fibroblasts by PPARg2, a lipidactivated transcription factor, Cell 79 (1994) 1147 – 1156. [3] E.D. Rosen, P. Sarraf, A.E. Troy, et al., PPARgamma is required for the differentiation of adipose tissue in vivo and in vitro, Mol. Cell 4 (1999) 611 – 617. [4] D. Ren, T.N. Collingwood, E.J. Rebar, et al., PPARgamma knockdown by engineered transcription factors: exogenous PPARgamma2 but not PPARgamma1 reactivates adipogenesis, Genes Dev. 16 (1) (2002) 27 – 32. [5] Y. Barak, D. Liao, W. Hem, et al., Effects of peroxisome proliferator-activated receptor delta on placentation, adiposity, and colorectal cancer, Proc. Natl. Acad. Sci. U. S. A. 99 (1) (2002) 303 – 308.
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T.-A. Cock, J. Auwerx / International Congress Series 1262 (2004) 47–50
[6] R.A. Norman, D.B. Thompson, T. Foroud, et al., Genomewide search for genes influencing percent body fat in Pima indians: suggestive linkage at chromosome 11Q21 Q22, Am. J. Hum. Genet. 60 (1997) 166 – 173. [7] F. Picard, J. Auwerx, PPAR(gamma) and glucose homeostasis, Annu. Rev. Nutr. 22 (2002) 167 – 197. [8] M. Gurnell, D.B. Savage, V.K. Chatterjee, S. O’Rahilly, The metabolic syndrome: peroxisome proliferatoractivated receptor gamma and its therapeutic modulation, J. Clin. Endocrinol. Metab. 88 (6) (2003) 2412 – 2421. [9] N. Kubota, Y. Terauchi, H. Miki, et al., PPARg mediates high-fat diet-induced adipocyte hypertrophy and insulin resistance, Mol. Cell 4 (1999) 597 – 609. [10] Y. Barak, M.C. Nelson, E.S. Ong, et al., PPARgamma is required for placental, cardiac, and adipose tissue development, Mol. Cell 4 (1999) 585 – 595. [11] P.D. Miles, Y. Barak, W. He, et al., Improved insulin-sensitivity in mice heterozygous for PPAR-gamma deficiency, J. Clin. Invest. 105 (3) (2000) 287 – 292. [12] H. Koutnikova, T.-A. Cock, M. Wanatabe, et al., Compensation by the muscle limits the metabolic consequences of lipodystrophy in PPARg hypomorphic mice, Proc. Natl. Acad. Sci. U. S. A. (2003). [13] A. Garg, Lipodystrophies, Am. J. Med. 108 (2) (2000) 143 – 152. [14] J. Moitra, M.M. Mason, M. Olive, et al., Life without fat: a transgenic mouse, Genes Dev. 12 (1998) 3168 – 3181. [15] I. Shimomura, R.E. Hammer, J.A. Richardson, et al., Insulin resistance and diabetes mellitus in transgenic mice expressing nuclear SREBP-1c in adipose tissue: model for congenital generalized lipodystrophy, Genes Dev. 12 (20) (1998) 3182 – 3194. [16] I. Shimomura, R.E. Hammer, S. Ikemoto, et al., Leptin reverses insulin resistance and diabetes mellitus in mice with congenital lipodystrophy, Nature 401 (6748) (1999) 73 – 76. [17] J.K. Kim, O. Gavrilova, Y. Chen, et al., Mechanism of insulin resistance in A-ZIP/F-1 fatless mice, J. Biol. Chem. 275 (12) (2000) 8456 – 8460. [18] K. Matsusue, M. Haluzik, G. Lambert, et al., Liver-specific disruption of PPARgamma in leptin-deficient mice improves fatty liver but aggravates diabetic phenotypes, J. Clin. Invest. 111 (5) (2003) 737 – 747. [19] O. Gavrilova, M. Haluzik, K. Matsusue, et al., Liver peroxisome proliferator activated receptor gamma contributes to hepatic steatosis, triglyceride clearance, and regulation of body fat mass, J. Biol. Chem. 278 (36) (2003) 34268 – 34276. [20] A.W. Norris, L. Chen, S.J. Fisher, et al., Muscle-specific PPARgamma-deficient mice develop increased adiposity and insulin resistance but respond to thiazolidinediones, J. Clin. Invest. 112 (4) (2003) 608 – 618.
www.ics-elsevier.com
PPARg fundamental role in adipogenesis Terrie-Anne Cock a, Johan Auwerx a,b,* a
Institut de Ge´ne´tique et de Biologie Mole´culaire et Cellulaire, CNRS/INSERM/Universite´ Louis Pasteur, 67404 Illkirch, France b Institut Clinique de la Souris, Ge´nopole Strasbourg, 67404 Illkirch, France
Abstract. The peroxisomeproliferator-activated receptor gamma (PPARg) is a nuclear receptor that integrates the control of lipid and glucose homeostasis. The actions of PPARg are mediated by two protein isoforms, the widely expressed PPARg1 and adipose tissue-restricted PPARg2 with an additional 28 amino acids in the NH2-terminus. The effects of the absence of PPARg has not been extensively characterized in vivo since the PPARg mice are embryonic lethal, therefore we generated mice that carry a hypomorphic mutation at the PPARg2 locus, yielding a WAT-specific PPARg knockdown. The PPARg hyp/hyp mouse model demonstrates that PPARg is the master regulator of adipogenesis. Furthermore this model highlights PPARg fundamental roles in whole body physiology to maintain the metabolic and endocrine functions of white adipose tissue. D 2003 Elsevier B.V. All rights reserved. Keywords: Peroxisome proliferator-activated receptor gamma; PPARg; Nuclear receptor; Lipid and glucose metabolism; Mouse model; Adipogenesis; Metabolism
1. Introduction The peroxisome proliferator-activated receptor gamma (PPARg) is a prototypical metabolic nuclear receptor involved in the transcriptional control of energy, lipid, and glucose homeostasis. The activity of PPARg is governed by binding of small lipophilic ligands, mainly fatty acids, derived from nutrition or metabolic pathways, or synthetic agonists, like thiazolidinediones. Docking of these ligands in the ligand-binding pocket alters the conformation of PPARg resulting in transcriptional activation subsequent to the release of corepressors and the recruitment of coactivators. The actions of PPARg are mediated by two protein isoforms, the widely expressed PPARg1 and adipose tissuerestricted PPARg2, both produced from a single gene by alternative splicing and dif-
* Corresponding author. Institut de Ge´ne´tique et de Biologie Mole´culaire et Cellulaire, Orphan Nuclear Receptors, B.P. 10142, 1 rue Laurent Fries, F-67404 Illkirch, France. Tel.: +33-3-88-65-3425; fax: +33-3-88-653200. E-mail address: [email protected] (J. Auwerx). 0531-5131/ D 2003 Elsevier B.V. All rights reserved. doi:10.1016/S0531-5131(03)01767-9
48
T.-A. Cock, J. Auwerx / International Congress Series 1262 (2004) 47–50
fering only by an additional 28 amino acids in the NH2-terminus of PPARg2 (reviewed Ref. [1]). 2. PPARg the master regulator of adipogenesis The initial suggestion that PPARg stimulated adipogenesis was based on the observation that overexpression of PPARg in cells was by itself sufficient to induce adipocyte differentiation [2]. Targeted mutagenesis of the PPARg gene in ES cells and knocking down the endogenous PPARg2 in cell lines confirmed the commanding role of PPARg in adipocyte differentiation [3,4]. Consistent with this, PPARg increases the expression of genes that promote fatty acid storage, whereas it represses genes that induce lipolysis in adipocytes. The lipodystrophy in the sole PPARg / mice, rescued by tetraploid aggregation [5], and the characterization of mice chimeric for PPARg / ES cells [3], further underscored the importance of PPARg in adipose tissue development in vivo. Human genetic studies further support the important role of PPARg in adipogenesis. One of the loci with suggestive linkage to obesity in the Pima Indians maps close to the location of PPARG on 3p25-p24[6]. Furthermore, subjects with the partial ‘‘loss-offunction’’ Pro12Ala mutation in the PPARg2-specific B exon have a lower body mass index (BMI), whereas carriers of the dominant negative Pro467Leu and Val290Met mutations or the Phe388Leu mutation that has a reduced affinity to ligand have partial lipodystrophy. Conversely, carriers of the Pro115Gln substitution, that renders PPARg constitutively active, are extremely obese [7,8]. A final line in favor of a role of PPARg in adipogenesis comes from pharmacological studies, which demonstrate that PPARg agonists, such as the thiazolidinediones, invariably increase WAT mass, whereas suboptimal PPARg activation, or PPARg antagonism, is neutral or even reverses weight gain [7]. The effects of the absence of PPARg has not been extensively characterized in vivo since the PPARg / mice die during intra-uterine development due to defects in cardiac and placental development in vivo [9 –11]. Therefore, we generated by homologous recombination, mice that carry a hypomorphic mutation at the PPARg2 locus, yielding a WAT-specific PPARg knockdown (PPARghyp/hyp) [12]. These mice have virtually no expression of PPARg in the sparse white adipose tissue (WAT), but not in other tissues. One-week-old mice with this WAT-specific PPARg knockdown display a lipodystrophic phenotype similar to that of humans with congenital generalized lipodystrophy (CGL). In CGL, the adipose deficiency normally is accompanied by severe insulin resistance, hyperglycemia, and fatty liver throughout life [13]. The neonatal PPARghyp/hyp phenotype mirrors most of these CGL characteristics, whereas the adult PPARghyp/hyp mice overcome some of the complications associated with the absence of WAT. In fact, adult PPARghyp/hyp mice do not have a fatty liver or show signs of hyperlipidemia [12]. The PPARghyp/hyp mice are furthermore only mildly glucose intolerant, which is distinct from most other lipodystrophy models [14 – 17]. This suggests that other physiological compensatory mechanisms ameliorate metabolic homeostasis in the PPARghyp/hyp mice. Overall the PPARghyp/hyp mouse model reveals that the PPARg locus is the critical regulator of adipogenesis in vivo [12].
T.-A. Cock, J. Auwerx / International Congress Series 1262 (2004) 47–50
49
3. WAT’s place in the scheme of energy metabolism The comparison of the physiological abnormalities in PPARghyp/hyp and PPARg heterozygote mice provides further insights into how the actions of PPARg in adipose tissue, liver, muscle and heart coordinate glucose and lipid homeostasis. Ubiquitous reduction of PPARg in all tissues, such as the PPARg heterozygote mice, results in normal body weight; these mice are insulin sensitive and resistant to diet-induced obesity [9,11]. In these PPARg heterozygote mice, circulating lipids can be stored in WAT, which avoids lipid overload in other tissues hence maintaining normal glucose and lipid metabolism [12]. In comparison, the PPARghyp/hyp mice with reduced PPARg levels in WAT have almost no WAT and are mildly glucose intolerant. Our study therefore implicates WAT as an important target tissue for PPARg activation to maintain glucose tolerance. Treatment with thiazolidinedione PPARg agonists can overcome the glucose intolerance in PPARghyp/hyp mice, although insulin resistance is not corrected. The PPARg agonist results from our WAT PPARghyp/hyp mice together with recent data in liver and muscle specific PPARg knockout mice indicate that primarily the WAT and not the liver and muscle are crucial for the protective effects of PPARg agonist on insulin resistance [12,18 –20]. 4. Selective modulation is the key The near absence of PPARg in WAT in adult PPARghyp/hyp mice is only causing a slightly abnormal metabolic profile and is rather well tolerated. Taken all together this lends further support to the hypothesis that selective PPARg modulators (or SPRMs) with a partial agonist or antagonist profile, rather than full agonists (reviewed in Ref. [7]), could be the preferred therapeutic strategy to treat the metabolic disorders associated with obesity, such as the metabolic syndrome, insulin resistance, or type 2 diabetes mellitus. The genetic evidence obtained in the PPARghyp/hyp mice arguments that selective PPARg modulators which have a reduced adipogenic drive, but which maintain agonist activity in other tissues, might maintain glucose homeostasis in such metabolic disorders. Acknowledgements This work was supported by grants from CNRS, INSERM, Hoˆpitaux Universitaires de Strasbourg, European Union, and NIH. References [1] J. Auwerx, PPARg, the ultimate thrifty gene, Diabetologia 42 (42) (1999) 1033 – 1049. [2] P. Tontonoz, E. Hu, B.M. Spiegelman, Stimulation of adipogenesis in fibroblasts by PPARg2, a lipidactivated transcription factor, Cell 79 (1994) 1147 – 1156. [3] E.D. Rosen, P. Sarraf, A.E. Troy, et al., PPARgamma is required for the differentiation of adipose tissue in vivo and in vitro, Mol. Cell 4 (1999) 611 – 617. [4] D. Ren, T.N. Collingwood, E.J. Rebar, et al., PPARgamma knockdown by engineered transcription factors: exogenous PPARgamma2 but not PPARgamma1 reactivates adipogenesis, Genes Dev. 16 (1) (2002) 27 – 32. [5] Y. Barak, D. Liao, W. Hem, et al., Effects of peroxisome proliferator-activated receptor delta on placentation, adiposity, and colorectal cancer, Proc. Natl. Acad. Sci. U. S. A. 99 (1) (2002) 303 – 308.
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T.-A. Cock, J. Auwerx / International Congress Series 1262 (2004) 47–50
[6] R.A. Norman, D.B. Thompson, T. Foroud, et al., Genomewide search for genes influencing percent body fat in Pima indians: suggestive linkage at chromosome 11Q21 Q22, Am. J. Hum. Genet. 60 (1997) 166 – 173. [7] F. Picard, J. Auwerx, PPAR(gamma) and glucose homeostasis, Annu. Rev. Nutr. 22 (2002) 167 – 197. [8] M. Gurnell, D.B. Savage, V.K. Chatterjee, S. O’Rahilly, The metabolic syndrome: peroxisome proliferatoractivated receptor gamma and its therapeutic modulation, J. Clin. Endocrinol. Metab. 88 (6) (2003) 2412 – 2421. [9] N. Kubota, Y. Terauchi, H. Miki, et al., PPARg mediates high-fat diet-induced adipocyte hypertrophy and insulin resistance, Mol. Cell 4 (1999) 597 – 609. [10] Y. Barak, M.C. Nelson, E.S. Ong, et al., PPARgamma is required for placental, cardiac, and adipose tissue development, Mol. Cell 4 (1999) 585 – 595. [11] P.D. Miles, Y. Barak, W. He, et al., Improved insulin-sensitivity in mice heterozygous for PPAR-gamma deficiency, J. Clin. Invest. 105 (3) (2000) 287 – 292. [12] H. Koutnikova, T.-A. Cock, M. Wanatabe, et al., Compensation by the muscle limits the metabolic consequences of lipodystrophy in PPARg hypomorphic mice, Proc. Natl. Acad. Sci. U. S. A. (2003). [13] A. Garg, Lipodystrophies, Am. J. Med. 108 (2) (2000) 143 – 152. [14] J. Moitra, M.M. Mason, M. Olive, et al., Life without fat: a transgenic mouse, Genes Dev. 12 (1998) 3168 – 3181. [15] I. Shimomura, R.E. Hammer, J.A. Richardson, et al., Insulin resistance and diabetes mellitus in transgenic mice expressing nuclear SREBP-1c in adipose tissue: model for congenital generalized lipodystrophy, Genes Dev. 12 (20) (1998) 3182 – 3194. [16] I. Shimomura, R.E. Hammer, S. Ikemoto, et al., Leptin reverses insulin resistance and diabetes mellitus in mice with congenital lipodystrophy, Nature 401 (6748) (1999) 73 – 76. [17] J.K. Kim, O. Gavrilova, Y. Chen, et al., Mechanism of insulin resistance in A-ZIP/F-1 fatless mice, J. Biol. Chem. 275 (12) (2000) 8456 – 8460. [18] K. Matsusue, M. Haluzik, G. Lambert, et al., Liver-specific disruption of PPARgamma in leptin-deficient mice improves fatty liver but aggravates diabetic phenotypes, J. Clin. Invest. 111 (5) (2003) 737 – 747. [19] O. Gavrilova, M. Haluzik, K. Matsusue, et al., Liver peroxisome proliferator activated receptor gamma contributes to hepatic steatosis, triglyceride clearance, and regulation of body fat mass, J. Biol. Chem. 278 (36) (2003) 34268 – 34276. [20] A.W. Norris, L. Chen, S.J. Fisher, et al., Muscle-specific PPARgamma-deficient mice develop increased adiposity and insulin resistance but respond to thiazolidinediones, J. Clin. Invest. 112 (4) (2003) 608 – 618.