RESUMO
The pancreas is critical for maintaining glucose homeostasis. Activating transcription factor 3 (ATF3) is an adaptive response transcription factor. There are major discrepancies in previous reports on pancreatic ATF3; therefore, its role in the pancreas is unclear. To better elucidate the role of ATF3 in the pancreas, we conducted in vitro studies using pancreatic α and ß cell lines, and also evaluated the use of ATF3 antibodies for immunohistochemistry. We determined ATF3 expression was increased by low glucose and decreased by high glucose in both αTC-1.6 and ßTC3 cells. We also showed that adenovirus-mediated ATF3 overexpression increased glucagon promoter activity and glucagon mRNA levels in αTC-1.6 cells; whereas, it had no effect on insulin promoter activity and insulin mRNA levels in ßTC3 cells. Although immunostaining with the C-19 ATF3 antibody demonstrated predominant expression in α cells rather than ß cells, ATF3 staining was still detected in ATF3 knockout mice as clearly as in control mice. On the other hand, another ATF3 antibody (H-90) detected ATF3 in both α cells and ß cells, and was clearly diminished in ATF3 knockout mice. These results indicate that previous discrepancies in ATF3 expression patterns in the pancreas were caused by the varying specificities of the ATF3 antibodies used, and that ATF3 is actually expressed in both α cells and ß cells.
Assuntos
Fator 3 Ativador da Transcrição/genética , Expressão Gênica/efeitos dos fármacos , Glucagon/genética , Glucose/administração & dosagem , Insulina/genética , Ilhotas Pancreáticas/metabolismo , Fator 3 Ativador da Transcrição/análise , Animais , Linhagem Celular , Células Secretoras de Glucagon/química , Células Secretoras de Glucagon/metabolismo , Células Secretoras de Insulina/química , Células Secretoras de Insulina/metabolismo , Camundongos , Camundongos Knockout , Regiões Promotoras Genéticas/genética , RNA Mensageiro/análiseRESUMO
OBJECTIVE: Nicotinamide rescues ß-cell damage and diabetes in rodents, but a large-scale clinical trial failed to show the benefit of nicotinamide in the prevention of type 1 diabetes. Recent studies have shown that Sirt1 deacetylase, a putative protector of ß-cells, is inhibited by nicotinamide. We investigated the effects of isonicotinamide, which is a derivative of nicotinamide and does not inhibit Sirt1, on streptozotocin (STZ)-induced diabetes in mice. RESEARCH DESIGN AND METHODS: Male C57BL/6 mice were administered with three different doses of STZ (65, 75, and 100 mg/kg BW) alone or in combination with subsequent high-fat feeding. The mice were treated with isonicotinamide (250 mg/kg BW/day) or phosphate-buffered saline for 10 days. The effects of isonicotinamide on STZ-induced diabetes were assessed by blood glucose levels, glucose tolerance test, and immunohistochemistry. RESULTS: Isonicotinamide effectively prevented hyperglycemia induced by higher doses of STZ (75 and 100mg/kg BW) alone and low-dose STZ (65 mg/kg BW) followed by 6-week high-fat diet in mice. The protective effects of isonicotinamide were associated with decreased apoptosis of ß-cells and reductions in both insulin content and insulin-positive area in the pancreas of STZ-administered mice. In addition, isonicotinamide inhibited STZ-induced apoptosis in cultured isolated islets. CONCLUSIONS: These data clearly demonstrate that isonicotinamide exerts anti-diabetogenic effects by preventing ß-cell damage after STZ administration. These findings warrant further investigations on the protective effects of isonicotinamide and related compounds against ß-cell damage in diabetes.
Assuntos
Citoproteção , Diabetes Mellitus Experimental/prevenção & controle , Hipoglicemiantes/administração & dosagem , Células Secretoras de Insulina/efeitos dos fármacos , Niacinamida/administração & dosagem , Animais , Apoptose/efeitos dos fármacos , Diabetes Mellitus Experimental/induzido quimicamente , Dieta Hiperlipídica/efeitos adversos , Células Secretoras de Insulina/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Sirtuína 1/antagonistas & inibidores , Estreptozocina/administração & dosagemRESUMO
In liver, glucose utilization and lipid synthesis are inextricably intertwined. When glucose availability exceeds its utilization, lipogenesis increases, leading to increased intrahepatic lipid content and lipoprotein secretion. Although the fate of three-carbon metabolites is largely determined by flux rate through the relevant enzymes, insulin plays a permissive role in this process. But the mechanism integrating insulin receptor signaling to glucose utilization with lipogenesis is unknown. Forkhead box O1 (FoxO1), a downstream effector of insulin signaling, plays a central role in hepatic glucose metabolism through the regulation of hepatic glucose production. In this study, we investigated the mechanism by which FoxO1 integrates hepatic glucose utilization with lipid synthesis. We show that FoxO1 overexpression in hepatocytes reduces activity of carbohydrate response element binding protein (Chrebp), a key regulator of lipogenesis, by suppressing O-linked glycosylation and reducing the protein stability. FoxO1 inhibits high glucose- or O-GlcNAc transferase (OGT)-induced liver-pyruvate kinase (L-PK) promoter activity by decreasing Chrebp recruitment to the L-PK promoter. Conversely, FoxO1 ablation in liver leads to the enhanced O-glycosylation and increased protein level of Chrebp owing to decreased its ubiquitination. We propose that FoxO1 regulation of Chrebp O-glycosylation is a mechanism linking hepatic glucose utilization with lipid synthesis.
Assuntos
Fatores de Transcrição Forkhead/metabolismo , Glucose/metabolismo , Fígado/metabolismo , Proteínas Nucleares/metabolismo , Fatores de Transcrição/metabolismo , Animais , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos , Western Blotting , Proteína Forkhead Box O1 , Fatores de Transcrição Forkhead/genética , Glicosilação , Imunoprecipitação , Camundongos , Camundongos Knockout , Proteínas Nucleares/genética , Regiões Promotoras Genéticas/genética , Estabilidade Proteica , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Transcrição/genéticaRESUMO
OBJECTIVE: Atherosclerotic cardiovascular disease is the leading cause of death among people with diabetes. Generation of oxidized LDLs and reduced nitric oxide (NO) availability because of endothelial NO synthase (eNOS) dysfunction are critical events in atherosclerotic plaque formation. Biochemical mechanism leading from hyperglycemia to oxLDL formation and eNOS dysfunction is unknown. RESEARCH DESIGN AND METHODS: We show that glucose, acting through oxidative stress, activates the transcription factor Foxo1 in vascular endothelial cells. RESULTS: Foxo1 promotes inducible NOS (iNOS)-dependent NO-peroxynitrite generation, which leads in turn to LDL oxidation and eNOS dysfunction. We demonstrate that Foxo1 gain-of-function mimics the effects of hyperglycemia on this process, whereas conditional Foxo1 knockout in vascular endothelial cells prevents it. CONCLUSIONS: The findings reveal a hitherto unsuspected role of the endothelial iNOS-NO-peroxynitrite pathway in lipid peroxidation and eNOS dysfunction and suggest that Foxo1 activation in response to hyperglycemia brings about proatherogenic changes in vascular endothelial cell function.
Assuntos
Endotélio Vascular/fisiologia , Fatores de Transcrição Forkhead/fisiologia , Hiperglicemia/fisiopatologia , Lipoproteínas LDL/metabolismo , Microcirculação/fisiologia , Óxido Nítrico Sintase Tipo III/metabolismo , Animais , Aorta/enzimologia , Glicemia/metabolismo , Doenças Cardiovasculares/epidemiologia , Doenças Cardiovasculares/genética , Doenças Cardiovasculares/mortalidade , Doenças Cardiovasculares/prevenção & controle , Proteínas de Ciclo Celular , Angiopatias Diabéticas/epidemiologia , Angiopatias Diabéticas/genética , Angiopatias Diabéticas/mortalidade , Angiopatias Diabéticas/prevenção & controle , Endotélio Vascular/enzimologia , Endotélio Vascular/fisiopatologia , Proteína Forkhead Box O1 , Proteína Forkhead Box O3 , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Humanos , Hiperglicemia/enzimologia , Hiperglicemia/genética , Resistência à Insulina/fisiologia , Camundongos , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo III/genética , Oxirredução , RNA/genética , RNA/isolamento & purificação , Fatores de Risco , Fatores de Transcrição/genéticaRESUMO
Forkhead transcription factors of the FoxO family have important roles in cellular proliferation, apoptosis, differentiation and stress resistance. FoxO proteins also play important roles in metabolism of complex organisms. FoxO1 regulates glucose and lipid metabolism in liver, as well as preadipocyte, myoblast and vascular endothelial cell differentiation. In the hypothalamus, FoxO controls food intake. In this chapter, we review the role of FoxO in pancreatic beta cells. Pancreatic beta cells secrete insulin to maintain the plasma glucose levels in a strict physiological range. Defects of beta cell function cause diabetes. The expression pattern of FoxO1 during pancreatic organogenesis is similar to that of Pdx1, Nkx2.2 and Pax4, transcription factors known to be critical for beta cell development. FoxO1 is expressed in a subset of pancreatic duct cells, in which insulin and/or Pdx1 are occasionally expressed. FoxO1 inhibits beta cell proliferation through suppression of Pdx1 by competing with FoxA2 and protects against beta cell failure induced by oxidative stress through NeuroD and MafA induction. Thus, a series of FoxO1 studies in pancreas suggested that FoxO1 plays important roles in pancreatic beta cell differentiation, neogenesis, proliferation and stress resistance. Genetic or pharmacological manipulation of FoxO can be used to prevent beta cell failure or aid in the differentiation of uncommitted endocrine progenitors into beta cells for transplantation.