Tomohiro Ide

The fat-derived hormone adiponectin reverses insulin resistance associated with both lipoatrophy and obesity.

Adiponectin is an adipocyte-derived hormone. Recent genome-wide scans have mapped a susceptibility locus for type 2 diabetes and metabolic syndrome to chromosome 3q27, where the gene encoding adiponectin is located. Here we show that decreased expression of adiponectin correlates with insulin resistance in mouse models of altered insulin sensitivity. Adiponectin decreases insulin resistance by decreasing triglyceride content in muscle and liver in obese mice. This effect results from increased expression of molecules involved in both fatty-acid combustion and energy dissipation in muscle. Moreover, insulin resistance in lipoatrophic mice was completely reversed by the combination of physiological doses of adiponectin and leptin, but only partially by either adiponectin or leptin alone. We conclude that decreased adiponectin is implicated in the development of insulin resistance in mouse models of both obesity and lipoatrophy. These data also indicate that the replenishment of adiponectin might provide a novel treatment modality for insulin resistance and type 2 diabetes.

Targeted disruption of AdipoR1 and AdipoR2 causes abrogation of adiponectin binding and metabolic actions

Adiponectin plays a central role as an antidiabetic and antiatherogenic adipokine. AdipoR1 and AdipoR2 serve as receptors for adiponectin in vitro, and their reduction in obesity seems to be correlated with reduced adiponectin sensitivity. Here we show that adenovirus-mediated expression of AdipoR1 and R2 in the liver of Lepr−/− mice increased AMP-activated protein kinase (AMPK) activation and peroxisome proliferator-activated receptor (PPAR)-α signaling pathways, respectively. Activation of AMPK reduced gluconeogenesis, whereas expression of the receptors in both cases increased fatty acid oxidation and lead to an amelioration of diabetes. Alternatively, targeted disruption of AdipoR1 resulted in the abrogation of adiponectin-induced AMPK activation, whereas that of AdipoR2 resulted in decreased activity of PPAR-α signaling pathways. Simultaneous disruption of both AdipoR1 and R2 abolished adiponectin binding and actions, resulting in increased tissue triglyceride content, inflammation and oxidative stress, and thus leading to insulin resistance and marked glucose intolerance. Therefore, AdipoR1 and R2 serve as the predominant receptors for adiponectin in vivo and play important roles in the regulation of glucose and lipid metabolism, inflammation and oxidative stress in vivo.

Overexpression of Monocyte Chemoattractant Protein-1 in Adipose Tissues Causes Macrophage Recruitment and Insulin Resistance *

Adipose tissue expression and circulating concentrations of monocyte chemoattractant protein-1 (MCP-1) correlate positively with adiposity. To ascertain the roles of MCP-1 overexpression in adipose, we generated transgenic mice by utilizing the adipocyte P2 (aP2) promoter (aP2-MCP-1 mice). These mice had higher plasma MCP-1 concentrations and increased macrophage accumulation in adipose tissues, as confirmed by immunochemical, flow cytometric, and gene expression analyses. Tumor necrosis factor-α and interleukin-6 mRNA levels in white adipose tissue and plasma non-esterified fatty acid levels were increased in transgenic mice. aP2-MCP-1 mice showed insulin resistance, suggesting that inflammatory changes in adipose tissues may be involved in the development of insulin resistance. Insulin resistance in aP2-MCP-1 mice was confirmed by hyperinsulinemic euglycemic clamp studies showing that transgenic mice had lower rates of glucose disappearance and higher endogenous glucose production than wild-type mice. Consistent with this, insulin-induced phosphorylations of Akt were significantly decreased in both skeletal muscles and livers of aP2-MCP-1 mice. MCP-1 pretreatment of isolated skeletal muscle blunted insulin-stimulated glucose uptake, which was partially restored by treatment with the MEK inhibitor U0126, suggesting that circulating MCP-1 may contribute to insulin resistance in aP2-MCP-1 mice. We concluded that both paracrine and endocrine effects of MCP-1 may contribute to the development of insulin resistance in aP2-MCP-1 mice.

SREBPs suppress IRS-2-mediated insulin signalling in the liver

Insulin receptor substrate 2 (IRS-2) is the main mediator of insulin signalling in the liver, controlling insulin sensitivity. Sterol regulatory element binding proteins (SREBPs) have been established as transcriptional regulators of lipid synthesis. Here, we show that SREBPs directly repress transcription of IRS-2 and inhibit hepatic insulin signalling. The IRS-2 promoter is activated by forkhead proteins through an insulin response element (IRE). Nuclear SREBPs effectively replace and interfere in the binding of these transactivators, resulting in inhibition of the downstream PI(3)K/Akt pathway, followed by decreased glycogen synthesis. These data suggest a molecular mechanism for the physiological switching from glycogen synthesis to lipogenesis and hepatic insulin resistance that is associated with hepatosteatosis.

Impaired Insulin Signaling in Endothelial Cells Reduces Insulin-Induced Glucose Uptake by Skeletal Muscle

In obese patients with type 2 diabetes, insulin delivery to and insulin-dependent glucose uptake by skeletal muscle are delayed and impaired. The mechanisms underlying the delay and impairment are unclear. We demonstrate that impaired insulin signaling in endothelial cells, due to reduced Irs2 expression and insulin-induced eNOS phosphorylation, causes attenuation of insulin-induced capillary recruitment and insulin delivery, which in turn reduces glucose uptake by skeletal muscle. Moreover, restoration of insulin-induced eNOS phosphorylation in endothelial cells completely reverses the reduction in capillary recruitment and insulin delivery in tissue-specific knockout mice lacking Irs2 in endothelial cells and fed a high-fat diet. As a result, glucose uptake by skeletal muscle is restored in these mice. Taken together, our results show that insulin signaling in endothelial cells plays a pivotal role in the regulation of glucose uptake by skeletal muscle. Furthermore, improving endothelial insulin signaling may serve as a therapeutic strategy for ameliorating skeletal muscle insulin resistance.

Inhibition of RXR and PPARγ ameliorates diet-induced obesity and type 2 diabetes

PPARgamma is a ligand-activated transcription factor and functions as a heterodimer with a retinoid X receptor (RXR). Supraphysiological activation of PPARgamma by thiazolidinediones can reduce insulin resistance and hyperglycemia in type 2 diabetes, but these drugs can also cause weight gain. Quite unexpectedly, a moderate reduction of PPARgamma activity observed in heterozygous PPARgamma-deficient mice or the Pro12Ala polymorphism in human PPARgamma, has been shown to prevent insulin resistance and obesity induced by a high-fat diet. In this study, we investigated whether functional antagonism toward PPARgamma/RXR could be used to treat obesity and type 2 diabetes. We show herein that an RXR antagonist and a PPARgamma antagonist decrease triglyceride (TG) content in white adipose tissue, skeletal muscle, and liver. These inhibitors potentiated leptins effects and increased fatty acid combustion and energy dissipation, thereby ameliorating HF diet-induced obesity and insulin resistance. Paradoxically, treatment of heterozygous PPARgamma-deficient mice with an RXR antagonist or a PPARgamma antagonist depletes white adipose tissue and markedly decreases leptin levels and energy dissipation, which increases TG content in skeletal muscle and the liver, thereby leading to the re-emergence of insulin resistance. Our data suggested that appropriate functional antagonism of PPARgamma/RXR may be a logical approach to protection against obesity and related diseases such as type 2 diabetes.

TFE3 transcriptionally activates hepatic IRS-2, participates in insulin signaling and ameliorates diabetes.

Using an expression cloning strategy, we have identified TFE3, a basic helix-loop-helix protein, as a transactivator of metabolic genes that are regulated through an E-box in their promoters. Adenovirus-mediated expression of TFE3 in hepatocytes in culture and in vivo strongly activated expression of IRS-2 and Akt and enhanced phosphorylation of insulin-signaling kinases such as Akt, glycogen synthase kinase 3β and p70S6 kinase. TFE3 also induced hexokinase II (HK2) and insulin-induced gene 1 (INSIG1). These changes led to metabolic consequences, such as activation of glycogen and protein synthesis, but not lipogenesis, in liver. Collectively, plasma glucose levels were markedly reduced both in normal mice and in different mouse models of diabetes, including streptozotocin-treated, db/db and KK mice. Promoter analyses showed that IRS2, HK2 and INSIG1 are direct targets of TFE3. Activation of insulin signals in both insulin depletion and resistance suggests that TFE3 could be a therapeutic target for diabetes.

SREBP-1 Interacts with Hepatocyte Nuclear Factor-4α and Interferes with PGC-1 Recruitment to Suppress Hepatic Gluconeogenic Genes

The hepatocyte nuclear factor-4α (HNF-4α)/PGC-1 pathway plays a crucial role in the transcriptional regulation of hepatic gluconeogenic enzymes such as phosphoenolpyruvate carboxykinase (PEPCK) and Glc-6-Pase, genes that are activated at fasting and suppressed in a fed state. SREBP-1c dominates the nutritional regulation of lipogenic genes inverse to gluconeogenesis. Here we show the mechanism by which SREBP-1 suppresses expression of gluconeogenic genes. A series of luciferase reporter assays demonstrated that SREBP-1a and -1c effectively inhibited the PEPCK promoter activity that was induced by HNF-4α. The HNF-4α-binding site in the glucocorticoid-response unit was responsible for the SREBP-1 inhibition, although SREBP-1 did not bind to the PEPCK promoter as demonstrated by electrophoretic mobility shift assays. The inhibitory effect was more potent in the isoform of SREBP-1a than SREBP-1c and was eliminated by deletion of the amino-terminal transactivation domain of SREBP-1. Coimmunoprecipitation experiments demonstrated that these two transcription factors directly interact through the transactivation domain of SREBP-1 and the ligand binding/AF2 domains of HNF-4α. Estimation of coactivator recruitment using HNF-4α-Gal4DBD fusion assay showed that SREBP-1 competitively inhibited PGC-1 recruitment, a requirement for HNF-4α activation. Consistent with these results, hepatic PEPCK and Glc-6-Pase mRNA levels are suppressed by overexpression of SREBP-1a and -1c in the transgenic mice. Our data indicate that SREBP-1 has a novel role as negative regulator of gluconeogenic genes through a cross-talk with HNF-4α interference with PGC-1 recruitment.

Tissue-specific actions of antidiabetic thiazolidinediones on the reduced fatty acid oxidation in skeletal muscle and liver of zucker diabetic fatty rats

Fatty acid overload has been proposed as a cause of decreased responsiveness in the major insulin target tissues of the body such as muscle and liver tissue. We therefore investigated fatty acid oxidation in soleus muscle and liver isolated from Zucker diabetic fatty (ZDF) rats treated with thiazolidinediones, a new class of antidiabetic agents. 14CO2 production from [14C]palmitic (C16:0) acid was lower in the soleus muscle and liver of ZDF rats versus lean rats (P < .05). When administered orally to ZDF rats for 2 weeks, the thiazolidinediones troglitazone (300 mg/kg) and KRP-297 (10 mg/kg) increased palmitic acid oxidation in the soleus muscle of ZDF rats (P < .05). KRP-297, but not troglitazone, increased palmitic acid oxidation in the liver of ZDF rats (P < .05), and both troglitazone and KRP-297 inhibited triglyceride accumulation in the skeletal muscle of ZDF rats. Hepatic triglyceride accumulation in ZDF rats was inhibited by KRP-297, but not by troglitazone. A reduction of fatty acid oxidation in the liver of ZDF rats and an increase in response to KRP-297 were observed only when C16:0 and C18:0 fatty acids, not C8:0, were used as substrates. Thus, there were defects in fatty acid catabolic activity and triglyceride accumulation in the soleus muscle and liver of ZDF rats. These results indicate that KRP-297 has advantages over troglitazone in the amelioration of these lipid metabolic abnormalities in insulin resistance associated with obesity.

Design, synthesis and evaluation of substituted phenylpropanoic acid derivatives as peroxisome proliferator-activated receptor (PPAR) activators: novel human PPARα-selective activators

Abstract A series of substituted phenylpropanoic acid derivatives was prepared as part of a search for subtype-selective human peroxisome proliferator-activated receptor (PPAR) activators. Structure–activity relationship studies indicated that the substituent at the α-position of the carboxyl group plays a key role in determining the potency and the selectivity for PPAR transactivation.