Dataset Information

Molecular characterization of insulin-mediated suppression of hepatic glucose production in vivo.

ABSTRACT:

Objective

Insulin-mediated suppression of hepatic glucose production (HGP) is associated with sensitive intracellular signaling and molecular inhibition of gluconeogenic (GNG) enzyme mRNA expression. We determined, for the first time, the time course and relevance (to metabolic flux) of these molecular events during physiological hyperinsulinemia in vivo in a large animal model.

Research design and methods

24 h fasted dogs were infused with somatostatin, while insulin (basal or 8 x basal) and glucagon (basal) were replaced intraportally. Euglycemia was maintained and glucose metabolism was assessed using tracer, (2)H(2)O, and arterio-venous difference techniques. Studies were terminated at different time points to evaluate insulin signaling and enzyme regulation in the liver.

Results

Hyperinsulinemia reduced HGP due to a rapid transition from net glycogen breakdown to synthesis, which was associated with an increase in glycogen synthase and a decrease in glycogen phosphorylase activity. Thirty minutes of hyperinsulinemia resulted in an increase in phospho-FOXO1, a decrease in GNG enzyme mRNA expression, an increase in F2,6P(2), a decrease in fat oxidation, and a transient decrease in net GNG flux. Net GNG flux was restored to basal by 4 h, despite a substantial reduction in PEPCK protein, as gluconeogenically-derived carbon was redirected from lactate efflux to glycogen deposition.

Conclusions

In response to acute physiologic hyperinsulinemia, 1) HGP is suppressed primarily through modulation of glycogen metabolism; 2) a transient reduction in net GNG flux occurs and is explained by increased glycolysis resulting from increased F2,6P(2) and decreased fat oxidation; and 3) net GNG flux is not ultimately inhibited by the rise in insulin, despite eventual reduction in PEPCK protein, supporting the concept that PEPCK has poor control strength over the gluconeogenic pathway in vivo.

SUBMITTER: Ramnanan CJ

PROVIDER: S-EPMC2874690 | biostudies-literature | 2010 Jun

REPOSITORIES: biostudies-literature

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Publications

Molecular characterization of insulin-mediated suppression of hepatic glucose production in vivo.

Ramnanan Christopher J CJ Edgerton Dale S DS Rivera Noelia N Irimia-Dominguez Jose J Farmer Ben B Neal Doss W DW Lautz Margaret M Donahue E Patrick EP Meyer Catalina M CM Roach Peter J PJ Cherrington Alan D AD

Diabetes 20100225 6

<h4>Objective</h4>Insulin-mediated suppression of hepatic glucose production (HGP) is associated with sensitive intracellular signaling and molecular inhibition of gluconeogenic (GNG) enzyme mRNA expression. We determined, for the first time, the time course and relevance (to metabolic flux) of these molecular events during physiological hyperinsulinemia in vivo in a large animal model.<h4>Research design and methods</h4>24 h fasted dogs were infused with somatostatin, while insulin (basal or 8 ...[more]

PMID: 20185816

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Project description:MicroRNAs (miRNAs) are known to contribute to many metabolic diseases, including diabetes. In this study, we investigated the role of miR199a-5p in the regulation of hepatic insulin sensitivity. Ad-anti-miR199a-5p adenoviruses were injected into male C57BL/6J WT mice fed a high-fat diet to inhibit miR199a-5p expression before the glucose levels and insulin resistance were assessed. Similarly, Ad-miR199a-5p adenoviruses were injected into male C57BL/6J WT mice to cause the overexpression of miR199a-5p. To investigate the roles of autophagy-related protein 14 (ATG14) and miR199a-5p in the regulation of insulin sensitivity, we injected Ad-miR199a-5p with or without Ad-ATG14 viruses into WT C57BL/6J mice before performing functional assays. Moreover, we infected HepG2 cells or primary hepatocytes with Ad-anti-miR199a-5p or Ad-miR199a-5p viruses to determine the effect of miR199a-5p on insulin resistance in vitro. Finally, we explored the clinical relevance of miR199a-5p by examining the expression level of miR199a-5p in liver samples derived from diabetes patients. We first demonstrated that knocking down miR199a-5p led to decreased glucose tolerance and clearance in vivo, whereas the overexpression of miR199a-5p had the opposite effect. We further identified ATG14 as the target of miR199a-5p, and ATG14 partially rescued miR199a-5p-potentiated glucose and insulin tolerance. In addition, transmission electron microscopy data and western blot data regarding ATG14, LC3 and BECLIN1 illustrated that miR199a-5p regulates autophagy via ATG14. Knocking down miR199a-5p in primary hepatocytes and HepG2 cells suppressed the insulin-stimulated phosphorylation of insulin receptor β, glycogen synthase kinase 3β and protein kinase B, whereas the overexpression of miR199a-5p further potentiated their phosphorylation. Finally, we detected upregulated miR199a-5p levels, which were correlated with reduced ATG14 mRNA levels and downregulated autophagy in liver samples obtained from diabetes patients. Our study uncovered a novel biological role of miR199a-5p in the regulation of hepatic insulin sensitivity via ATG14-mediated autophagy.

Project description:BACKGROUND:Ghrelin modulates many physiological processes. However, the effects of intestinal ghrelin on hepatic glucose production (HGP) are still unclear. The current study was to explore the roles of intestinal ghrelin on glucose homeostasis and insulin signaling in the liver. METHODS:The system of intraduodenal infusion and intracerebral microinfusion into the nucleus of the solitary tract (NTS) in the normal chow-diet rats and pancreatic-euglycemic clamp procedure (PEC) combined with [3-3H] glucose as a tracer were used to analyze the effect of intestinal ghrelin. Intraduodenal co-infusion of ghrelin, tetracaine and Activated Protein Kinase (AMPK) activator (AICAR), or pharmacologic and molecular inhibitor of N-methyl-D-aspartate receptors within the dorsal vagal complex, or hepatic vagotomy in rats were used to explore the possible mechanism of the effect of intestinal ghrelin on HGP. RESULTS:Our results demonstrated that gut infusion of ghrelin inhibited duodenal AMP-dependent protein kinase (AMPK) signal pathways, increased HGP and expression of gluconeogenic enzymes, and decreased insulin signaling in the liver of the rat. Intraduodenal co-infusion of ghrelin receptor antagonist [D-Lys3]-GHRP-6 and AMPK agonist with ghrelin diminished gut ghrelin-induced increase in HGP and decrease in glucose infusion rate (GIR) and hepatic insulin signaling. The effects of gut ghrelin were also negated by co-infusion with tetracaine, or MK801, an N-methyl-D-aspartate (NMDA) receptor inhibitor, and adenovirus expressing the shRNA of NR1 subunit of NMDA receptors (Ad-shNR1) within the dorsal vagal complex, and hepatic vagotomy in rats. When ghrelin and lipids were co-infused into the duodenum, the roles of gut lipids in increasing the rate of glucose infusion (GIR) and lowering HGP were reversed. CONCLUSIONS:The current study provided evidence that intestinal ghrelin has an effect on HGP and identified a neural glucoregulatory function of gut ghrelin signaling.

Dataset Information

Molecular characterization of insulin-mediated suppression of hepatic glucose production in vivo.

Objective

Research design and methods

Results

Conclusions

Publications

Molecular characterization of insulin-mediated suppression of hepatic glucose production in vivo.

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