Diabetes is associated with defective β cell function and altered β cell mass. The mechanisms regulating β cell mass and its adaptation to insulin resistance are unknown. It is unclear whether compensatory β cell hyperplasia is achieved via proliferation of existing β cells or neogenesis from progenitor cells embedded in duct epithelia. We have used transgenic mice expressing a mutant form of the forkhead-O1 transcription factor (FoxO1) in both pancreatic ductal and endocrine β cells to assess the contribution of these 2 compartments to islet expansion. We show that the mutant FoxO1 transgene prevents β cell replication in 2 models of β cell hyperplasia, 1 due to peripheral insulin resistance (Insulin receptor transgenic knockouts) and 1 due to ectopic local expression of IGF2 (Elastase-IGF2 transgenics), without affecting insulin secretion. In contrast, we failed to detect a specific effect of the FoxO1 transgene on the number of periductal β cells. We propose that β cell compensation to insulin resistance is a proliferative response of existing β cells to growth factor signaling and requires FoxO1 nuclear exclusion.
Haruka Okamoto, Marta Letizia Hribal, Hua V. Lin, William R. Bennett, ndrew Ward,, Domenico Accili
James L. Smart, Virginie Tolle, Malcolm J. Low
The growth hormone (GH) secretagogue receptor (GHSR) was cloned as the target of a family of synthetic molecules endowed with GH release properties. As shown recently through in vitro means, this receptor displays a constitutive activity whose clinical relevance is unknown. Although pharmacological studies have demonstrated that its endogenous ligand — ghrelin — stimulates, through the GHSR, GH secretion and appetite, the physiological importance of the GHSR-dependent pathways remains an open question that gives rise to much controversy. We report the identification of a GHSR missense mutation that segregates with short stature within 2 unrelated families. This mutation, which results in decreased cell-surface expression of the receptor, selectively impairs the constitutive activity of the GHSR, while preserving its ability to respond to ghrelin. This first description, to our knowledge, of a functionally significant GHSR mutation, which unveils the critical importance of the GHSR-associated constitutive activity, discloses an unusual pathogenic mechanism of growth failure in humans.
Jacques Pantel, Marie Legendre, Sylvie Cabrol, Latifa Hilal, Yassir Hajaji, Séverine Morisset, Sylvie Nivot, Marie-Pierre Vie-Luton, Dominique Grouselle, Marc de Kerdanet, Abdelkrim Kadiri, Jacques Epelbaum, Yves Le Bouc, Serge Amselem
Hepatic steatosis is a core feature of the metabolic syndrome and type 2 diabetes and leads to hepatic insulin resistance. Malonyl-CoA, generated by acetyl-CoA carboxylases 1 and 2 (Acc1 and Acc2), is a key regulator of both mitochondrial fatty acid oxidation and fat synthesis. We used a diet-induced rat model of nonalcoholic fatty liver disease (NAFLD) and hepatic insulin resistance to explore the impact of suppressing Acc1, Acc2, or both Acc1 and Acc2 on hepatic lipid levels and insulin sensitivity. While suppression of Acc1 or Acc2 expression with antisense oligonucleotides (ASOs) increased fat oxidation in rat hepatocytes, suppression of both enzymes with a single ASO was significantly more effective in promoting fat oxidation. Suppression of Acc1 also inhibited lipogenesis whereas Acc2 reduction had no effect on lipogenesis. In rats with NAFLD, suppression of both enzymes with a single ASO was required to significantly reduce hepatic malonyl-CoA levels in vivo, lower hepatic lipids (long-chain acyl-CoAs, diacylglycerol, and triglycerides), and improve hepatic insulin sensitivity. Plasma ketones were significantly elevated compared with controls in the fed state but not in the fasting state, indicating that lowering Acc1 and -2 expression increases hepatic fat oxidation specifically in the fed state. These studies suggest that pharmacological inhibition of Acc1 and -2 may be a novel approach in the treatment of NAFLD and hepatic insulin resistance.
David B. Savage, Cheol Soo Choi, Varman T. Samuel, Zhen-Xiang Liu, Dongyan Zhang, Amy Wang, Xian-Man Zhang, Gary W. Cline, Xing Xian Yu, John G. Geisler, Sanjay Bhanot, Brett P. Monia, Gerald I. Shulman
Developmental exposure to appropriate levels of thyroid hormones (THs) in a timely manner is critical to normal development in vertebrates. Among the factors potentially affecting perinatal exposure of tissues to THs is type 3 deiodinase (D3). This enzyme degrades THs and is highly expressed in the pregnant uterus, placenta, and fetal and neonatal tissues. To determine the physiological role of D3, we have generated a mouse D3 knockout model (D3KO) by a targeted inactivating mutation of the Dio3 gene in mouse ES cells. Early in life, D3KO mice exhibit delayed 3,5,3′-triiodothyronine (T3) clearance, a markedly elevated serum T3 level, and overexpression of T3-inducible genes in the brain. From postnatal day 15 to adulthood, D3KO mice demonstrate central hypothyroidism, with low serum levels of 3,5,3′,5′-tetraiodothyronine (T4) and T3, and modest or no increase in thyroid-stimulating hormone (TSH) concentration. Peripheral tissues are also hypothyroid. Hypothalamic T3 content is decreased while thyrotropin-releasing hormone (TRH) expression is elevated. Our results demonstrate that the lack of D3 function results in neonatal thyrotoxicosis followed later by central hypothyroidism that persists throughout life. These mice provide a new model of central hypothyroidism and reveal a critical role for D3 in the maturation and function of the thyroid axis.
Arturo Hernandez, M. Elena Martinez, Steven Fiering, Valerie Anne Galton, Donald St. Germain
Null mutations of the proopiomelanocortin gene (Pomc–/–) cause obesity in humans and rodents, but the contributions of central versus pituitary POMC deficiency are not fully established. To elucidate these roles, we introduced a POMC transgene (Tg) that selectively restored peripheral melanocortin and corticosterone secretion in Pomc–/– mice. Rather than improving energy balance, the genetic replacement of pituitary POMC in Pomc–/–Tg+ mice aggravated their metabolic syndrome with increased caloric intake and feed efficiency, reduced oxygen consumption, increased subcutaneous, visceral, and hepatic fat, and severe insulin resistance. Pair-feeding of Pomc–/–Tg+ mice to the daily intake of lean controls normalized their rate of weight gain but did not abolish obesity, indicating that hyperphagia is a major but not sole determinant of the phenotype. Replacement of corticosterone in the drinking water of Pomc–/– mice recapitulated the hyperphagia, excess weight gain and fat accumulation, and hyperleptinemia characteristic of genetically rescued Pomc–/–Tg+ mice. These data demonstrate that CNS POMC peptides play a critical role in energy homeostasis that is not substituted by peripheral POMC. Restoration of pituitary POMC expression to create a de facto neuronal POMC deficiency exacerbated the development of obesity, largely via glucocorticoid modulation of appetite, metabolism, and energy partitioning.
James L. Smart, Virginie Tolle, Malcolm J. Low
Diabetes can result in loss of enteric neurons and subsequent gastrointestinal complications. The mechanism of enteric neuronal loss in diabetes is not known. We examined the effects of hyperglycemia on enteric neuronal survival and the effects of glial cell line–derived neurotrophic factor (GDNF) on modulating this survival. Exposure of primary enteric neurons to 20 mM glucose (hyperglycemia) for 24 hours resulted in a significant increase in apoptosis compared with 5 mM glucose (normoglycemia). Exposure to 20 mM glucose resulted in decreased Akt phosphorylation and enhanced nuclear translocation of forkhead box O3a (FOXO3a). Treatment of enteric neurons with GDNF ameliorated these changes. In streptozotocin-induced diabetic mice, there was evidence of myenteric neuronal apoptosis and reduced Akt phosphorylation. Diabetic mice had loss of NADPH diaphorase–stained myenteric neurons, delayed gastric emptying, and increased intestinal transit time. The pathophysiological effects of hyperglycemia (apoptosis, reduced Akt phosphorylation, loss of inhibitory neurons, motility changes) were reversed in diabetic glial fibrillary acidic protein–GDNF (GFAP-GDNF) Tg mice. In conclusion, we demonstrate that hyperglycemia induces neuronal loss through a reduction in Akt-mediated survival signaling and that these effects are reversed by GDNF. GDNF may be a potential therapeutic target for the gastrointestinal motility disorders related to diabetes.
Mallappa Anitha, Chetan Gondha, Roy Sutliff, Alexander Parsadanian, Simon Mwangi, Shanthi V. Sitaraman, Shanthi Srinivasan
Insulin inhibits glucose production through both direct and indirect effects on the liver; however, considerable controversy exists regarding the relative importance of these effects. The first aim of this study was to determine which of these processes dominates the acute control of hepatic glucose production (HGP). Somatostatin and portal vein infusions of insulin and glucagon were used to clamp the pancreatic hormones at basal levels in the nondiabetic dog. After a basal sampling period, insulin infusion was switched from the portal vein to a peripheral vein. As a result, the arterial insulin level doubled and the hepatic sinusoidal insulin level was reduced by half. While the arterial plasma FFA level and net hepatic FFA uptake fell by 40–50%, net hepatic glucose output increased more than 2-fold and remained elevated compared with that in the control group. The second aim of this study was to determine the effect of a 4-fold rise in head insulin on HGP during peripheral hyperinsulinemia and hepatic insulin deficiency. Sensitivity of the liver was not enhanced by increased insulin delivery to the head. Thus, this study demonstrates that the direct effects of insulin dominate the acute regulation of HGP in the normal dog.
Dale S. Edgerton, Margaret Lautz, Melanie Scott, Carrie A. Everett, Kathryn M. Stettler, Doss W. Neal, Chang A. Chu, Alan D. Cherrington
Thyroid hormone (TH) action is mediated by TH receptors (TRs), which are members of the nuclear hormone receptor superfamily. In vitro studies have demonstrated that TR activity is regulated by interactions with corepressor and coactivator proteins (CoRs and CoAs, respectively). TH stimulation is thought to involve dissociation of CoRs and recruitment of CoAs to the liganded TR. In contrast, negative regulation by TH is thought to occur via recruitment of CoRs to the liganded TR. The physiological role of CoAs bound to TRs, however, has yet to be defined. In this study, we used gene-targeting techniques to mutate the TR-β locus within its activation function–2 (AF-2) domain (E457A). This mutation was chosen because it completely abolished CoA recruitment in vitro, while preserving normal triiodothyronine (T3) binding and CoR interactions. As expected, TH-stimulated gene expression was reduced in homozygous E457A mice. However, these animals also displayed abnormal regulation of the hypothalamic-pituitary-thyroid axis. Serum thyroxine, T3, and thyroid-stimulating hormone (TSH) levels and pituitary Tshb mRNA levels were inappropriately elevated compared with those of WT animals, and L-T3 treatment failed to suppress serum TSH and pituitary Tshb mRNA levels. Therefore, the AF-2 domain of TR-β is required for positive and, paradoxically, for negative regulation by TH in vivo.
Tania M. Ortiga-Carvalho, Nobuyuki Shibusawa, Amisra Nikrodhanond, Karen J. Oliveira, Danielle S. Machado, Xiao-Hui Liao, Ronald N. Cohen, Samuel Refetoff, Fredric E. Wondisford
The relative roles of the types 1 and 2 iodothyronine deiodinases (D1 and D2) in extrathyroidal 3,5,3′-triiodothyronine (T3) production in humans are unknown. We calculated the rate of thyroxine (T4) to T3 conversion by intact cells transiently expressing D1 or D2 at low (2 pM), normal (20 pM), and high (200 pM) free T4 concentrations. Deiodinase activities were then assayed in cell sonicates. The ratio of T3 production in cell sonicates (catalytic efficiency) was multiplied by the tissue activities reported in human liver (D1) and skeletal muscle (D2). From these calculations, we predict that in euthyroid humans, D2-generated T3 is 29 nmol/d, while that of D1-generated T3 is 15 nmol/d, from these major deiodinase-expressing tissues. The total estimated extrathyroidal T3 production, 44 nmol/d, is in close agreement with the 40 nmol T3/d based on previous kinetic studies. D2-generated T3 production accounts for approximately 71% of the peripheral T3 production in hypothyroidism, but D1 for approximately 67% in thyrotoxic patients. We also show that the intracellular D2-generated T3 has a greater effect on T3-dependent gene transcription than that from D1, which indicates that generation of nuclear T3 is an intrinsic property of the D2 protein. We suggest that impairment of D2-generated T3 is the major cause of the reduced T3 production in the euthyroid sick syndrome.
Ana Luiza Maia, Brian W. Kim, Stephen A. Huang, John W. Harney, P. Reed Larsen