JNK1 in hematopoietically derived cells contributes to diet-induced inflammation and insulin resistance without affecting obesity

Obesity-induced insulin resistance is a major factor in the etiology of type 2 diabetes, and Jun kinases (JNKs) are key negative regulators of insulin sensitivity in the obese state. Activation of JNKs (mainly JNK1) in insulin target cells results in phosphorylation of insulin receptor substrates (IRSs) at serine and threonine residues that inhibit insulin signaling. JNK1 activation is also required for accumulation of visceral fat. Here we used reciprocal adoptive transfer experiments to determine whether JNK1 in myeloid cells, such as macrophages, also contributes to insulin resistance and central adiposity. Our results show that deletion of Jnk1 in the nonhematopoietic compartment protects mice from high-fat diet (HFD)-induced insulin resistance, in part through decreased adiposity. By contrast, Jnk1 removal from hematopoietic cells has no effect on adiposity but confers protection against HFD-induced insulin resistance by decreasing obesity-induced inflammation.     

IRS1-independent defects define major nodes of insulin resistance

Insulin resistance is a common disorder caused by a wide variety of physiological insults, some of which include poor diet, inflammation, anti-inflammatory steroids, hyperinsulinemia, and dyslipidemia. The common link between these diverse insults and insulin resistance is widely considered to involve impaired insulin signaling, particularly at the level of the insulin receptor substrate (IRS). To test this model, we utilized a heterologous system involving the platelet-derived growth factor (PDGF) pathway that recapitulates many aspects of insulin action independently of IRS. We comprehensively analyzed six models of insulin resistance in three experimental systems and consistently observed defects in both insulin and PDGF action despite a range of insult-specific defects within the IRS-Akt nexus. These findings indicate that while insulin resistance is associated with multiple deficiencies, the most deleterious defects and the origin of insulin resistance occur independently of IRS.     

Hepatic insulin resistance is sufficient to produce dyslipidemia and susceptibility to atherosclerosis

Insulin resistance plays a central role in the development of the metabolic syndrome, but how it relates to cardiovascular disease remains controversial. Liver insulin receptor knockout (LIRKO) mice have pure hepatic insulin resistance. On a standard chow diet, LIRKO mice have a proatherogenic lipoprotein profile with reduced high-density lipoprotein (HDL) cholesterol and very low-density lipoprotein (VLDL) particles that are markedly enriched in cholesterol. This is due to increased secretion and decreased clearance of apolipoprotein B-containing lipoproteins, coupled with decreased triglyceride secretion secondary to increased expression of Pgc-1 beta (Ppargc-1b), which promotes VLDL secretion, but decreased expression of Srebp-1c (Srebf1), Srebp-2 (Srebf2), and their targets, the lipogenic enzymes and the LDL receptor. Within 12 weeks on an atherogenic diet, LIRKO mice show marked hypercholesterolemia, and 100% of LIRKO mice, but 0% of controls, develop severe atherosclerosis. Thus, insulin resistance at the level of the liver is sufficient to produce the dyslipidemia and increased risk of atherosclerosis associated with the metabolic syndrome.     

The insulin-regulated CREB coactivator TORC promotes stress resistance in Drosophila

In fasted mammals, glucose homeostasis is maintained through induction of the cAMP response element-binding protein (CREB) coactivator transducer of regulated CREB activity 2 (TORC2), which stimulates the gluconeogenic program in concert with the forkhead factor FOXO1. Here we show that starvation also triggers TORC activation in Drosophila, where it maintains energy balance through induction of CREB target genes in the brain. TORC mutant flies have reduced glycogen and lipid stores and are sensitive to starvation and oxidative stress. Neuronal TORC expression rescued stress sensitivity as well as CREB target gene expression in TORC mutants. During refeeding, increases in insulin signaling inhibited TORC activity through the salt-inducible kinase 2 (SIK2)-mediated phosphorylation and subsequent degradation of TORC. Depletion of neuronal SIK2 increased TORC activity and enhanced stress resistance. As disruption of insulin signaling also augmented TORC activity in adult flies, our results illustrate the importance of an insulin-regulated pathway that functions in the brain to maintain energy balance.     

Glucose-induced cyclic AMP oscillations regulate pulsatile insulin secretion

Cyclic AMP (cAMP) and Ca²⁺ are key regulators of exocytosis in many cells, including insulin-secreting β cells. Glucose-stimulated insulin secretion from β cells is pulsatile and involves oscillations of the cytoplasmic Ca²⁺ concentration ([Ca²⁺]_i), but little is known about the detailed kinetics of cAMP signaling. Using evanescent-wave fluorescence imaging we found that glucose induces pronounced oscillations of cAMP in the submembrane space of single MIN6 cells and primary mouse β cells. These oscillations were preceded and enhanced by elevations of [Ca²⁺]_i. However, conditions raising cytoplasmic ATP could trigger cAMP elevations without accompanying [Ca²⁺]_i rise, indicating that adenylyl cyclase activity may be controlled also by the substrate concentration. The cAMP oscillations correlated with pulsatile insulin release. Whereas elevation of cAMP enhanced secretion, inhibition of adenylyl cyclases suppressed both cAMP oscillations and pulsatile insulin release. We conclude that cell metabolism directly controls cAMP and that glucose-induced cAMP oscillations regulate the magnitude and kinetics of insulin exocytosis.     

Nampt/PBEF/Visfatin regulates insulin secretion in beta cells as a systemic NAD biosynthetic enzyme

Intracellular nicotinamide phosphoribosyltransferase (iNampt) is an essential enzyme in the NAD biosynthetic pathway. An extracellular form of this protein (eNampt) has been reported to act as a cytokine named PBEF or an insulin-mimetic hormone named visfatin, but its physiological relevance remains controversial. Here we show that eNampt does not exert insulin-mimetic effects in vitro or in vivo but rather exhibits robust NAD biosynthetic activity. Haplodeficiency and chemical inhibition of Nampt cause defects in NAD biosynthesis and glucose-stimulated insulin secretion in pancreatic islets in vivo and in vitro. These defects are corrected by administration of nicotinamide mononucleotide (NMN), a product of the Nampt reaction. A high concentration of NMN is present in mouse plasma, and plasma eNampt and NMN levels are reduced in Nampt heterozygous females. Our results demonstrate that Nampt-mediated systemic NAD biosynthesis is critical for beta cell function, suggesting a vital framework for the regulation of glucose homeostasis.     

SirT1 inhibition reduces IGF-I/IRS-2/Ras/ERK1/2 signaling and protects neurons

Sirtuins are known to protect cells and extend life span, but our previous studies indicated that S. cerevisiae Sir2 can also increase stress sensitivity and limit life-span extension. Here we provide evidence for a role of the mammalian Sir2 ortholog SirT1 in the sensitization of neurons to oxidative damage. SirT1 inhibition increased acetylation and decreased phosphorylation of IRS-2; it also reduced activation of the Ras/ERK1/2 pathway, suggesting that SirT1 may enhance IGF-I signaling in part by deacetylating IRS-2. Either the inhibition of SirT1 or of Ras/ERK1/2 was associated with resistance to oxidative damage. Markers of oxidized proteins and lipids were reduced in the brain of old SirT1-deficient mice, but the life span of the homozygote knockout mice was reduced under both normal and calorie-restricted conditions. These results are consistent with findings in S. cerevisiae and other model systems, suggesting that mammalian sirtuins can play both protective and proaging roles.     

Dissociation of hepatic steatosis and insulin resistance in mice overexpressing DGAT in the liver

Hepatic steatosis, the accumulation of lipids in the liver, is widely believed to result in insulin resistance. To test the causal relationship between hepatic steatosis and insulin resistance, we generated mice that overexpress acyl-CoA:diacylglycerol acyltransferase 2 (DGAT2), which catalyzes the final step of triacylglycerol (TG) biosynthesis, in the liver (Liv-DGAT2 mice). Liv-DGAT2 mice developed hepatic steatosis, with increased amounts of TG, diacylglycerol, ceramides, and unsaturated long-chain fatty acyl-CoAs in the liver. However, they had no abnormalities in plasma glucose and insulin levels, glucose and insulin tolerance, rates of glucose infusion and hepatic glucose production during hyperinsulinemic-euglycemic clamp studies, or activities of insulin-stimulated signaling proteins in the liver. DGAT1 overexpression in the liver also failed to induce glucose or insulin intolerance. Our results indicate that DGAT-mediated lipid accumulation in the liver is insufficient to cause insulin resistance and show that hepatic steatosis can occur independently of insulin resistance.     

A conserved role for phosphatidylinositol 3-kinase but not Akt signaling in mitochondrial adaptations that accompany physiological cardiac hypertrophy

Physiological cardiac hypertrophy is associated with mitochondrial adaptations that are characterized by activation of PGC-1alpha and increased fatty acid oxidative (FAO) capacity. It is widely accepted that phosphatidylinositol 3-kinase (PI3K) signaling to Akt1 is required for physiological cardiac growth. However, the signaling pathways that coordinate physiological hypertrophy and metabolic remodeling are incompletely understood. We show here that activation of PI3K is sufficient to increase myocardial FAO capacity and that inhibition of PI3K signaling prevents mitochondrial adaptations in response to physiological hypertrophic stimuli despite increased expression of PGC-1alpha. We also show that activation of the downstream kinase Akt is not required for the mitochondrial adaptations that are secondary to PI3K activation. Thus, in physiological cardiac growth, PI3K is an integrator of cellular growth and metabolic remodeling. Although PI3K signaling to Akt1 is required for cellular growth, Akt-independent pathways mediate the accompanying mitochondrial adaptations.     

Regulation of insulin biosynthesis in pancreatic beta cells by an endoplasmic reticulum-resident protein kinase IRE1

In pancreatic beta cells, the endoplasmic reticulum (ER) is an important site for insulin biosynthesis and the folding of newly synthesized proinsulin. Here, we show that IRE1alpha, an ER-resident protein kinase, has a crucial function in insulin biosynthesis. IRE1alpha phosphorylation is coupled to insulin biosynthesis in response to transient exposure to high glucose; inactivation of IRE1alpha signaling by siRNA or inhibition of IRE1alpha phosphorylation hinders insulin biosynthesis. IRE1 activation by high glucose does not accompany XBP-1 splicing and BiP dissociation but upregulates its target genes such as WFS1. Thus, IRE1 signaling activated by transient exposure to high glucose uses a unique subset of downstream components and has a beneficial effect on pancreatic beta cells. In contrast, chronic exposure of beta cells to high glucose causes ER stress and hyperactivation of IRE1, leading to the suppression of insulin gene expression. IRE1 signaling is therefore a potential target for therapeutic regulation of insulin biosynthesis.     

Enhanced leptin sensitivity and improved glucose homeostasis in mice lacking suppressor of cytokine signaling-3 in POMC-expressing cells

Suppressor of cytokine signaling-3 (Socs-3) negatively regulates the action of various cytokines, as well as the metabolic hormones leptin and insulin. Mice with haploinsufficiency of Socs-3, or those with neuronal deletion of Socs-3, are lean and more leptin and insulin sensitive. To examine the role of Socs-3 within specific neurons critical to energy balance, we created mice with selective deletion of Socs-3 within pro-opiomelanocortin (POMC)-expressing cells. These mice had enhanced leptin sensitivity, measured by weight loss and food intake after leptin infusion. On chow diet, glucose homeostasis was improved despite normal weight gain. On a high-fat diet, the rate of weight gain was reduced, due to increased energy expenditure rather than decreased food intake; glucose homeostasis and insulin sensitivity were substantially improved. These studies demonstrate that Socs-3 within POMC neurons regulates leptin sensitivity and glucose homeostasis, and plays a key role in linking high-fat diet to disordered metabolism.     

Increased energy expenditure, dietary fat wasting, and resistance to diet-induced obesity in mice lacking renin

An overactive renin-angiotensin system is associated with obesity and the metabolic syndrome. However, the mechanisms behind it are unclear. Cleaving angiotensinogen to angiotensin I by renin is a rate-limiting step of angiotensin II production, but renin is suggested to have angiotensin-independent effects. We generated mice lacking renin (Ren1c) using embryonic stem cells from C57BL/6 mice, a strain prone to diet-induced obesity. Ren1c^−/− mice are lean, insulin sensitive, and resistant to diet-induced obesity without changes in food intake and physical activity. The lean phenotype is likely due to a higher metabolic rate and gastrointestinal loss of dietary fat. Most of the metabolic changes in Ren1c^−/− mice were reversed by angiotensin II administration. These results support a role for angiotensin II in the pathogenesis of diet-induced obesity and insulin resistance.     

Deorphanization of a G protein-coupled receptor for oleoylethanolamide and its use in the discovery of small-molecule hypophagic agents

The endogenous lipid signaling agent oleoylethanolamide (OEA) has recently been described as a peripherally acting agent that reduces food intake and body weight gain in rat feeding models. This paper presents evidence that OEA is an endogenous ligand of the orphan receptor GPR119, a G protein-coupled receptor (GPCR) expressed predominantly in the human and rodent pancreas and gastrointestinal tract and also in rodent brain, suggesting that the reported effects of OEA on food intake may be mediated, at least in part, via the GPR119 receptor. Furthermore, we have used the recombinant receptor to discover novel selective small-molecule GPR119 agonists, typified by PSN632408, which suppress food intake in rats and reduce body weight gain and white adipose tissue deposition upon subchronic oral administration to high-fat-fed rats. GPR119 therefore represents a novel and attractive potential target for the therapy of obesity and related metabolic disorders.     

Cannabinoid Receptor Type 1 Protects against Age- Related Osteoporosis by Regulating Osteoblast and Adipocyte Differentiation in Marrow Stromal Cells

Age-related osteoporosis is characterized by reduced bone formation and accumulation of fat in the bone marrow compartment. Here, we report that the type 1 cannabinoid receptor (CB1) regulates this process. Mice with CB1 deficiency (CB1^−/−) had increased peak bone mass due to reduced bone resorption, but developed age-related osteoporosis with reduced bone formation and accumulation of adipocytes in the bone marrow space. Marrow stromal cells from CB1^−/− mice had an enhanced capacity for adipocyte differentiation, a reduced capacity for osteoblast differentiation, and increased expression of phosphorylated CREB (pCREB) and PPARγ. Pharmacological blockade of CB1 receptors stimulated adipocyte differentiation, inhibited osteoblast differentiation, and increased cAMP and pCREB in osteoblast and adipocyte precursors. The CB1 receptor is therefore unique in that it regulates peak bone mass through an effect on osteoclast activity, but protects against age-related bone loss by regulating adipocyte and osteoblast differentiation of bone marrow stromal cells.