ABSTRACT
All pancreatic endocrine cells, producing glucagon, insulin, somatostatin, or PP, differentiate from Pdx1+ progenitors that transiently express Neurogenin3. To understand whether the competence of pancreatic progenitors changes over time, we generated transgenic mice expressing a tamoxifen-inducible Ngn3 fusion protein under the control of the pdx1 promoter and backcrossed the transgene into the ngn3(-/-) background, devoid of endogenous endocrine cells. Early activation of Ngn3-ER(TM) almost exclusively induced glucagon+ cells, while depleting the pool of pancreas progenitors. As from E11.5, Pdx1+ progenitors became competent to differentiate into insulin+ and PP+ cells. Somatostatin+ cells were generated from E14.5, while the competence to make glucagon+ cells was dramatically decreased. Hence, pancreas progenitors, similar to retinal or cortical progenitors, go through competence states that each allow the generation of a subset of cell types. We further show that the progenitors acquire competence to generate late-born cells in a mechanism that is intrinsic to the epithelium.
Subject(s)
Basic Helix-Loop-Helix Transcription Factors/genetics , Cell Differentiation/genetics , Endocrine System/embryology , Epithelial Cells/metabolism , Nerve Tissue Proteins/genetics , Pancreas/embryology , Stem Cells/metabolism , Animals , Endocrine System/cytology , Endocrine System/metabolism , Epithelial Cells/cytology , Female , Gene Expression Regulation, Developmental/genetics , Glucagon/metabolism , Homeodomain Proteins/genetics , Insulin/metabolism , Insulin Secretion , Islets of Langerhans/cytology , Islets of Langerhans/embryology , Islets of Langerhans/metabolism , Male , Mice , Mice, Knockout , Pancreas/cytology , Pancreas/metabolism , Pancreatic Hormones/metabolism , Promoter Regions, Genetic/genetics , Somatostatin/metabolism , Stem Cells/cytology , Time Factors , Trans-Activators/geneticsABSTRACT
Accumulation of lipids in non-adipose tissues is often associated with Type 2 diabetes and its complications. Elevated expression of the lipogenic transcription factor, sterol regulatory element binding protein-1c (SREBP-1c), has been demonstrated in islets and liver of diabetic animals. To elucidate the molecular mechanisms underlying SREBP-1c-induced beta-cell dysfunction, we employed the Tet-On inducible system to achieve tightly controlled and conditional expression of the nuclear active form of SREBP-1c (naSREBP-1c) in INS-1 cells. Controlled expression of naSREBP-1c induced massive accumulation of lipid droplets and blunted nutrient-stimulated insulin secretion in INS-1 cells. K(+)-evoked insulin exocytosis was unaltered. Quantification of the gene expression profile in this INS-1 stable clone revealed that naSREBP-1c induced beta-cell dysfunction by targeting multiple genes dedicated to carbohydrate metabolism, lipid biosynthesis, cell growth, and apoptosis. naSREBP-1c elicits cell growth-arrest and eventually apoptosis. We also found that the SREBP-1c processing in beta-cells was irresponsive to acute stimulation of glucose and insulin, which was distinct from that in lipogenic tissues. However, 2-day exposure to these agents promoted SREBP-1c processing. Therefore, the SREBP-1c maturation could be implicated in the pathogenesis of beta-cell glucolipotoxicity.
Subject(s)
CCAAT-Enhancer-Binding Proteins/biosynthesis , DNA-Binding Proteins/biosynthesis , Islets of Langerhans/physiopathology , Protein Processing, Post-Translational/physiology , Animals , CCAAT-Enhancer-Binding Proteins/physiology , Clone Cells , DNA-Binding Proteins/physiology , Diabetes Mellitus, Type 2/etiology , Diabetes Mellitus, Type 2/metabolism , Gene Expression Regulation/drug effects , Gene Expression Regulation/physiology , Glucose/metabolism , Glucose/pharmacology , Hypoglycemic Agents/pharmacology , Insulin/metabolism , Insulin/pharmacology , Islets of Langerhans/metabolism , Protein Processing, Post-Translational/drug effects , Rats , Sterol Regulatory Element Binding Protein 1 , Transcription Factors/physiologyABSTRACT
The transcription factor Foxa2 is implicated in blood glucose homeostasis. Conditional expression of Foxa2 or its dominant-negative mutant DN-Foxa2 in INS-1 cells reveals that Foxa2 regulates the expression of genes important for glucose sensing in pancreatic beta-cells. Overexpression of Foxa2 results in blunted glucose-stimulated insulin secretion, whereas induction of DN-Foxa2 causes a left shift of glucose-induced insulin release. The mRNA levels of GLUT2 and glucokinase are drastically decreased after induction of Foxa2. In contrast, loss of Foxa2 function leads to up-regulation of hexokinase (HK) I and II and glucokinase (HK-IV) mRNA expression. The glucokinase and the low K(m) hexokinase activities as well as glycolysis are increased proportionally. In addition, induction of DN-Foxa2 also reduces the expression of beta-cell K(ATP) channel subunits Sur1 and Kir6.2 by 70%. Furthermore, in contrast to previous reports, induction of Foxa2 causes pronounced decreases in the HNF4alpha and HNF1alpha mRNA levels. Foxa2 fails to regulate the expression of Pdx1 transcripts. The expression of insulin and islet amyloid polypeptide is markedly suppressed after induction of Foxa2, while the glucagon mRNA levels are significantly increased. Conversely, Foxa2 is required for glucagon expression in these INS-1-derived cells. These results suggest that Foxa2 is a vital transcription factor evolved to control the expression of genes essential for maintaining beta-cell glucose sensing and glucose homeostasis.
