Insulin resistance is often associated with obesity and can precipitate type 2 diabetes. To date, most known approaches that improve insulin resistance must be preceded by the amelioration of obesity and hepatosteatosis. Here, we show that this provision is not mandatory; insulin resistance and hyperglycemia are improved by the modification of hepatic fatty acid composition, even in the presence of persistent obesity and hepatosteatosis. Mice deficient for Elovl6, the gene encoding the elongase that catalyzes the conversion of palmitate to stearate, were generated and shown to become obese and develop hepatosteatosis when fed a high-fat diet or mated to leptin-deficient ob/ob mice. However, they showed marked protection from hyperinsulinemia, hyperglycemia and hyperleptinemia. Amelioration of insulin resistance was associated with restoration of hepatic insulin receptor substrate-2 and suppression of hepatic protein kinase C epsilon activity resulting in restoration of Akt phosphorylation. Collectively, these data show that hepatic fatty acid composition is a new determinant for insulin sensitivity that acts independently of cellular energy balance and stress. Inhibition of this elongase could be a new therapeutic approach for ameliorating insulin resistance, diabetes and cardiovascular risks, even in the presence of a continuing state of obesity.

GH deficiency is known to be clinically associated with a high incidence of nonalcoholic fatty liver disease, and this can be reversed by GH administration. Here we investigated the mechanistic basis for this phenomenon using engineered male mice lacking different signaling elements of the GH receptor, hepatic stat5a/b(-/-) mice and a mouse hepatoma line. We found deficient GH-dependent signal transducer and activator of transcription (STAT)-5 signaling correlates with steatosis, and through microarray analysis, quantitative PCR, and chromatin immunoprecipitation, identified putative targets of STAT5 signaling responsible for the steatosis seen on a normal diet. These targets were verified with liver-specific stat5a/b deletion in vivo, and in vitro we show that dominant-negative (DN) STAT5 increases lipid uptake in a mouse hepatoma line. Because loss of STAT5 signaling results in elevated STAT1 and STAT3 activity and intracellular lipid accumulation, we have used DN-STAT5a/b, DN-STAT1, constitutively active (CA)-STAT3, or addition of oleate/palmitate in the hepatoma line to assign which of these apply to individual targets in STAT5 signaling deficiency. These findings and published mouse models of steatosis enable us to propose elevated cd36, pparγ, and pgc1α/β expression as primary instigators of the steatosis along with elevated fatty acid synthase, lipoprotein lipase, and very low-density lipoprotein receptor expression. Decreased fgf21 and insig2 expression may also contribute. In conclusion, despite normal plasma free fatty acids and minimal obesity, absent GH activation leads to steatosis because activated STAT5 prevents hepatic steatosis. These results raise the possibility of low-dose GH treatment for nonalcoholic fatty liver disease.

To determine the role of cholesterol synthesis in pancreatic b-cells, a transgenic model of in vivo activation of sterol-regulatory element binding protein 2 (SREBP-2) specifically in b-cells (TgRIP-SREBP-2) was developed and analyzed. Expression of nuclear human SREBP-2 in b-cells resulted in severe diabetes as evidenced by greater than 5-fold elevations in glycohemoglobin compared with C57BL/6 controls. Diabetes in TgRIP-SREBP-2 mice was primarily due to defects in glucose-and potassium-stimulated insulin secretion as determined by glucose tolerance test. Isolated islets of TgSREBP-2 mice were fewer in number, smaller, deformed, and had decreased insulin content. SREBP-2-expressing islets also contained increased esterified cholesterol and unchanged triglycerides with reduced ATP levels. Consistently, these islets exhibited elevated expression of HMG-CoA synthase and reductase and LDL receptor, with suppression of endogenous SREBPs. Genes involved in b-cell differentiation, such as PDX1 and BETA2, were suppressed, explaining loss of b-cell mass, whereas IRS2 expression was not affected. These phenotypes were dependent on the transgene expression. Taken Sterol-regulatory element binding protein 1c (SREBP-1c) is a membrane-bound transcription factor of the basic HLH (bHLH) leucine zipper family and has been established as a nutritional regulator of lipogenic enzymes in the liver (3, 4). Expression of SREBP-1c is highly upregulated by dietary intake of carbohydrates, sugars, and saturated FAs, whereas PUFAs, such as eicosapentaenoic acid, have been shown to inhibit hepatic SREBP-1c through multiple mechanisms (5, 6). These nutritional regulations of SREBP-1c are also observed in a cultured b-cell line and in isolated islets of mice (7,8). SREBP-1c also plays a role in insulin signaling by inhibiting insulin receptor substrate 2 (IRS-2), the major insulin-signaling mediator in the liver and in b-cells (9, 10).As a model for lipotoxicity by endogenous FAs in pancreatic b-cells, we previously developed transgenic mice overexpressing the active form of SREBP-1c under the insulin promoter expression (10). These mice exhibited impaired glucose tolerance in vivo due to both decreased b-cell mass and impaired insulin secretion estimated in isolated islets, which was enhanced by feeding the mice a high-fat, high-sucrose diet. The SREBP-1c-overexpressing islets had ATP depletion caused by enhanced lipogenesis and increased uncoupling protein 2 (UCP-2). Explaining the loss of b-cell mass, these islets had decreased expression of IRS-2 and PDX1. In addition to inhibition of GSIS,

GH is generally believed to signal exclusively through Janus tyrosine kinases (JAK), particularly JAK2, leading to activation of signal transducers and activators of transcription (STAT), ERK and phosphatidylinositol 3-kinase pathways, resulting in transcriptional regulation of target genes. Here we report the creation of targeted knock-in mice wherein the Box1 motif required for JAK2 activation by the GH receptor (GHR) has been disabled by four Pro/Ala mutations. These mice are unable to activate hepatic JAK2, STAT3, STAT5, or Akt in response to GH injection but can activate Src and ERK1/2. Their phenotype is identical to that of the GHR(-/-) mouse, emphasizing the key role of JAK2 in postnatal growth and the minimization of obesity in older males. In particular, they show dysregulation of the IGF-I/IGF-binding protein axis at transcript and protein levels and decreased bone length. Because no gross phenotypic differences were evident between GHR(-/-) and Box1 mutants, we undertook transcript profiling in liver from 4-month-old males. We compared their transcript profiles with our 391-GHR truncated mice, which activate JAK2, ERK1/2, and STAT3 in response to GH but not STAT5a/b. This has allowed us for the first time to identify in vivo Src/ERK-regulated transcripts, JAK2-regulated transcripts, and those regulated by the distal part of the GHR, particularly by STAT5.

Both adipocyte hyperplasia and hypertrophy are determinant factors for adipocyte differentiation during the development of obesity. p21 WAF1/CIP1 , a cyclin-dependent kinase inhibitor, is induced during adipocyte differentiation; however, its precise contribution to this process is unknown. Using both in vitro and in vivo systems, we show that p21 is crucial for maintaining adipocyte hypertrophy and obesity-induced insulin resistance. The absence of p21 in 3T3-L1 fibroblasts by RNA-mediated interference knockdown or in embryonic fibroblasts from p21 ؊/؊ mice impaired adipocyte differentiation, resulting in smaller adipocytes. Despite normal adipose tissue mass on a normal diet, p21 ؊/؊ mice fed high energy diets had reduced adipose tissue mass and adipocyte size accompanied by a marked improvement in insulin sensitivity. Knockdown of p21 in enlarged epididymal fat of diet-induced obese mice and also in fully differentiated 3T3-L1 adipocytes caused vigorous apoptosis by activating p53. Thus, p21 is involved in both adipocyte differentiation and in protecting hypertrophied adipocytes against apoptosis. Via both of these mechanisms, p21 promotes adipose tissue expansion during high fat diet feeding, leading to increased downstream pathophysiological consequences such as insulin resistance.The process of adipocyte differentiation has been extensively characterized in cultures of preadipocyte clonal cell lines such as mouse 3T3-L1 and 3T3-F442A (1, 2). Adipogenesis requires a sequence of events including growth arrest of proliferating preadipocytes, coordinated reentry into the cell cycle with limited clonal expansion, and growth arrest before terminal differentiation during which lipid accumulation occurs. Thus, it is reasonable to assume that factors involved in cell cycle regulation may have important roles in the adipocyte differentiation process. Cell cycle progression in mammals is governed by various complexes of cyclins and cyclin-dependent kinases (CDKs) 2 as well as their inhibitors, of which p21 and p27 are the most widely studied. It was previously suggested that expression of p21 and p27 is changed during the adipogenesis (3). p21 has been reported to be induced by Foxo1 and has been implicated in entry of adipocytes into the clonal expansion phase of adipogenesis; meanwhile, p27 is critical for adipocyte hyperplasia (3-5). The expression of both p21 and p27 is altered during adipocyte differentiation; however, their precise role in adipogenesis, especially at the time of terminal differentiation, remains unclear.Obesity is caused by over-nutrition and decreased physical activity and is characterized by excess storage of lipids in adipose tissue, which can be accounted for by both adipocyte hyperplasia and hypertrophy. Obesity often precedes insulin resistance and precipitates type 2 diabetes and cardiovascular diseases (6). Impaired adipocyte differentiation is also related to insulin resistance. Based upon the potential roles of CDK inhibitors in cell cycle progression and apoptosis, it is reasonable t...