Molecular events that result in loss of pain perception are poorly understood in diabetic neuropathy. Our results show that the receptor for advanced glycation end products (RAGE), a receptor associated with sustained NF-κB activation in the diabetic microenvironment, has a central role in sensory neuronal dysfunction. In sural nerve biopsies, ligands of RAGE, the receptor itself, activated NF-κBp65, and IL-6 colocalized in the microvasculature of patients with diabetic neuropathy. Activation of NF-κB and NF-κB–dependent gene expression was upregulated in peripheral nerves of diabetic mice, induced by advanced glycation end products, and prevented by RAGE blockade. NF-κB activation was blunted in RAGE-null (RAGE–/–) mice compared with robust enhancement in strain-matched controls, even 6 months after diabetes induction. Loss of pain perception, indicative of long-standing diabetic neuropathy, was reversed in WT mice treated with soluble RAGE. Most importantly, loss of pain perception was largely prevented in RAGE–/– mice, although they were not protected from diabetes-induced loss of PGP9.5-positive plantar nerve fibers. These data demonstrate, for the first time to our knowledge, that the RAGE–NF-κB axis operates in diabetic neuropathy, by mediating functional sensory deficits, and that its inhibition may provide new therapeutic approaches.
Angelika Bierhaus, Karl-Matthias Haslbeck, Per M. Humpert, Birgit Liliensiek, Thomas Dehmer, Michael Morcos, Ahmed A.R. Sayed, Martin Andrassy, Stephan Schiekofer, Jochen G. Schneider, Jörg B. Schulz, Dieter Heuss, Bernhard Neundörfer, Stefan Dierl, Jochen Huber, Hans Tritschler, Ann-Marie Schmidt, Markus Schwaninger, Hans-Ulrich Haering, Erwin Schleicher, Michael Kasper, David M. Stern, Bernd Arnold, Peter P. Nawroth
A critical defect in type 2 diabetes is impaired insulin-stimulated glucose transport and metabolism in muscle and adipocytes. To understand the metabolic adaptations this elicits, we generated mice with targeted disruption of the GLUT4 glucose transporter in both adipocytes and muscle (AMG4KO). In contrast to total body GLUT4-null mice, AMG4KO mice exhibit normal growth, development, adipose mass, and longevity. They develop fasting hyperglycemia and glucose intolerance and are at risk for greater insulin resistance than mice lacking GLUT4 in only one tissue. Hyperinsulinemic-euglycemic clamp studies showed a 75% decrease in glucose infusion rate and markedly reduced 2-deoxyglucose uptake into skeletal muscle (85–90%) and white adipose tissue (65%). However, AMG4KO mice adapt by preferentially utilizing lipid fuels, as evidenced by a lower respiratory quotient and increased clearance of lipids from serum after oral lipid gavage. While insulin action on hepatic glucose production and gluconeogenic enzymes is impaired, hepatic glucokinase expression, incorporation of 14C-glucose into lipids, and hepatic VLDL-triglyceride release are increased. The lipogenic activity may be mediated by increased hepatic expression of SREBP-1c and acetyl-CoA carboxylase. Thus, inter-tissue communication results in adaptations to impaired glucose transport in muscle and adipocytes that involve increased hepatic glucose uptake and lipid synthesis, while muscle adapts by preferentially utilizing lipid fuels. Genetic determinants limiting this “metabolic flexibility” may contribute to insulin resistance and type 2 diabetes in humans.
Ko Kotani, Odile D. Peroni, Yasuhiko Minokoshi, Olivier Boss, Barbara B. Kahn
The apolipoprotein apoC-III plays an important role in plasma triglyceride metabolism. It is predominantly produced in liver, and its hepatic expression is inhibited by insulin. To elucidate the inhibitory mechanism of insulin in apoC-III expression, we delivered forkhead box O1 (Foxo1) cDNA to hepatocytes by adenovirus-mediated gene transfer. Foxo1 stimulated hepatic apoC-III expression and correlated with the ability of Foxo1 to bind to its consensus site in the apoC-III promoter. Deletion or mutation of the Foxo1 binding site abolished insulin response and Foxo1-mediated stimulation. Likewise, Foxo1 also mediated insulin action on intestinal apoC-III expression in enterocytes. Furthermore, elevated Foxo1 production in liver augmented hepatic apoC-III expression, resulting in increased plasma triglyceride levels and impaired fat tolerance in mice. Transgenic mice expressing a constitutively active Foxo1 allele exhibited hypertriglyceridemia. Moreover, we show that hepatic Foxo1 expression becomes deregulated as a result of insulin deficiency or insulin resistance, culminating in significantly elevated Foxo1 production, along with its skewed nuclear distribution, in livers of diabetic NOD or db/db mice. While loss of insulin response is associated with unrestrained apoC-III production and impaired triglyceride metabolism, these data suggest that Foxo1 provides a molecular link between insulin deficiency or resistance and aberrant apoC-III production in the pathogenesis of diabetic hypertriglyceridemia.
Jennifer Altomonte, Lin Cong, Sonal Harbaran, Anja Richter, Jing Xu, Marcia Meseck, Hengjiang Henry Dong
Genetic and environmental factors contribute to age-dependent susceptibility to type 2 diabetes. Recent studies have reported reduced expression of PPARγ coactivator 1α (PGC-1α) and PGC-1β genes in skeletal muscle from type 2 diabetic patients, but it is not known whether this is an inherited or acquired defect. To address this question we studied expression of these genes in muscle biopsies obtained from young and elderly dizygotic and monozygotic twins without known diabetes before and after insulin stimulation and related the expression to a Gly482Ser variant in the PGC-1α gene. Insulin increased and aging reduced skeletal muscle PGC-1α and PGC-1β mRNA levels. This age-dependent decrease in muscle gene expression was partially heritable and influenced by the PGC-1α Gly482Ser polymorphism. In addition, sex, birth weight, and aerobic capacity influenced expression of PGC-1α in a complex fashion. Whereas expression of PGC-1α in muscle was positively related to insulin-stimulated glucose uptake and oxidation, PGC-1β expression was positively related to fat oxidation and nonoxidative glucose metabolism. We conclude that skeletal muscle PGC-1α and PGC-1β expression are stimulated by insulin and reduced by aging. The data also suggest different regulatory functions for PGC-1α and PGC-1β on glucose and fat oxidation in muscle cells. The finding that the age-dependent decrease in the expression of these key genes regulating oxidative phosphorylation is under genetic control could provide an explanation by which an environmental trigger (age) modifies genetic susceptibility to type 2 diabetes.
Charlotte Ling, Pernille Poulsen, Emma Carlsson, Martin Ridderstråle, Peter Almgren, Jørgen Wojtaszewski, Henning Beck-Nielsen, Leif Groop, Allan Vaag
Adipose tissue plays a central role in the control of energy homeostasis through the storage and turnover of triglycerides and through the secretion of factors that affect satiety and fuel utilization. Agents that enhance insulin sensitivity, such as rosiglitazone, appear to exert their therapeutic effect through adipose tissue, but the precise mechanisms of their actions are unclear. Rosiglitazone changes the morphological features and protein profiles of mitochondria in 3T3-L1 adipocytes. To examine the relevance of these effects in vivo, we studied white adipocytes from ob/ob mice during the development of obesity and after treatment with rosiglitazone. The levels of approximately 50% of gene transcripts encoding mitochondrial proteins were decreased with the onset of obesity. About half of those genes were upregulated after treatment with rosiglitazone, and this was accompanied by an increase in mitochondrial mass and changes in mitochondrial structure. Functionally, adipocytes from rosiglitazone-treated mice displayed markedly enhanced oxygen consumption and significantly increased palmitate oxidation. These data reveal mitochondrial remodeling and increased energy expenditure in white fat in response to rosiglitazone treatment in vivo and suggest that enhanced lipid utilization in this tissue may affect whole-body energy homeostasis and insulin sensitivity.
Leanne Wilson-Fritch, Sarah Nicoloro, My Chouinard, Mitchell A. Lazar, Patricia C. Chui, John Leszyk, Juerg Straubhaar, Michael P. Czech, Silvia Corvera
Casitas b-lineage lymphoma (c-Cbl) is an E3 ubiquitin ligase that has an important role in regulating the degradation of cell surface receptors. In the present study we have examined the role of c-Cbl in whole-body energy homeostasis. c-Cbl–/– mice exhibited a profound increase in whole-body energy expenditure as determined by increased core temperature and whole-body oxygen consumption. As a consequence, these mice displayed a decrease in adiposity, primarily due to a reduction in cell size despite an increase in food intake. These changes were accompanied by a significant increase in activity (2- to 3-fold). In addition, c-Cbl–/– mice displayed a marked improvement in whole-body insulin action, primarily due to changes in muscle metabolism. We observed increased protein levels of the insulin receptor (4-fold) and uncoupling protein-3 (2-fold) in skeletal muscle and a significant increase in the phosphorylation of AMP-activated protein kinase and acetyl-CoA carboxylase. These findings suggest that c-Cbl plays an integral role in whole-body fuel homeostasis by regulating whole-body energy expenditure and insulin action.
Juan C. Molero, Thomas E. Jensen, Phil C. Withers, Michelle Couzens, Herbert Herzog, Christine B.F. Thien, Wallace Y. Langdon, Ken Walder, Maria A. Murphy, David D.L. Bowtell, David E. James, Gregory J. Cooney
The accurate matching of caloric intake to caloric expenditure involves a complex system of peripheral signals and numerous CNS neurotransmitter systems. Syndecans are a family of membrane-bound heparan sulfate proteoglycans that modulate ligand-receptor interactions. Syndecan-3 is heavily expressed in several areas of the brain, including hypothalamic nuclei, which are known to regulate energy balance. In particular, syndecans have been implicated in modulation of the activity of the melanocortin system, which potently regulates energy intake, energy expenditure, and peripheral glucose metabolism. Our data demonstrate that syndecan-3–null mice have reduced adipose content compared with wild-type mice. On a high-fat diet, syndecan-3–null male and female mice exhibited a partial resistance to obesity due to reduced food intake in males and increased energy expenditure in females relative to that of wild-type mice. As a result, syndecan-3–null mice on a high-fat diet accumulated less adipose mass and showed improved glucose tolerance compared with wild-type controls. The data implicate syndecan-3 in the regulation of body weight and suggest that inhibition of syndecan-3 may provide a therapeutic approach for the treatment of obesity resulting from exposure to high-fat diets.
April D. Strader, Ofer Reizes, Stephen C. Woods, Stephen C. Benoit, Randy J. Seeley
We found that when a site-specific binding protein interacts with the “handle” region of the prorenin prosegment, the prorenin molecule undergoes a conformational change to its enzymatically active state. This nonproteolytic activation is completely blocked by a decoy peptide with the handle region structure, which competitively binds to such a binding protein. Given increased plasma prorenin in diabetes, we examined the hypothesis that the nonproteolytic activation of prorenin plays a significant role in diabetic organ damage. Streptozotocin-induced diabetic rats were treated with subcutaneous administration of handle region peptide. Metabolic and renal histological changes and the renin-Ang system components in the plasma and kidneys were determined at 8, 16, and 24 weeks following streptozotocin treatment. Kidneys of diabetic rats contained increased Ang I and II without any changes in renin, Ang-converting enzyme, or angiotensinogen synthesis. Treatment with the handle region peptide decreased the renal content of Ang I and II, however, and completely inhibited the development of diabetic nephropathy without affecting hyperglycemia. We propose that the nonproteolytic activation of prorenin may be a significant mechanism of diabetic nephropathy. The mechanism and substances causing nonproteolytic activation of prorenin may serve as important therapeutic targets for the prevention of diabetic organ damage.
Atsuhiro Ichihara, Matsuhiko Hayashi, Yuki Kaneshiro, Fumiaki Suzuki, Tsutomu Nakagawa, Yuko Tada, Yukako Koura, Akira Nishiyama, Hirokazu Okada, M. Nasir Uddin, A.H.M. Nurun Nabi, Yuichi Ishida, Tadashi Inagami, Takao Saruta
The melanocortin-4 receptor (MC4R), a centrally expressed G protein–coupled receptor (GPCR), is essential for the maintenance of long-term energy balance in humans. Mutations in MC4R are the most common genetic cause of obesity. Since activation of this receptor leads to a decrease in food intake, MC4R is also a major therapeutic target for the treatment of obesity. Control of MC4R activity in vivo is modulated by the opposing effects of the anorexigenic agonist α–melanocyte-stimulating hormone (α-MSH) and the orexigenic antagonist agouti-related protein (AGRP). In addition, experiments in vitro have demonstrated that the human MC4R has an intrinsic constitutive activity on which AGRP also acts as an inverse agonist. The physiological role of this constitutive activity in the control of energy balance as well as the domain of the protein implicated in its maintenance are unknown. By systematically studying functional defects in naturally occurring MC4R mutations from obese patients, we defined a cluster of N-terminal mutations that selectively impair the constitutive activity of the receptor. Further characterization of this domain demonstrated that it functions as a tethered intramolecular ligand that maintains the constitutive activity of MC4R and may provide novel avenues for the design of drugs targeting this receptor. Our results also suggest that the tonic satiety signal provided by the constitutive activity of MC4R may be required for maintaining long-term energy homeostasis in humans.
Supriya Srinivasan, Cecile Lubrano-Berthelier, Cedric Govaerts, Franck Picard, Pamela Santiago, Bruce R. Conklin, Christian Vaisse
The molecular link between obesity and β cell failure that causes diabetes is difficult to establish. Here we show that a conditional knockout of insulin receptor substrate 2 (Irs2) in mouse pancreas β cells and parts of the brain — including the hypothalamus —increased appetite, lean and fat body mass, linear growth, and insulin resistance that progressed to diabetes. Diabetes resolved when the mice were between 6 and 10 months of age: functional β cells expressing Irs2 repopulated the pancreas, restoring sufficient β cell function to compensate for insulin resistance in the obese mice. Thus, Irs2 signaling promotes regeneration of adult β cells and central control of nutrient homeostasis, which can prevent obesity and diabetes in mice.
Xueying Lin, Akiko Taguchi, Sunmin Park, Jake A. Kushner, Fan Li, Yedan Li, Morris F. White