apyrase and Insulin-Resistance

Extracellular nucleotides are important mediators of inflammatory responses and could also impact metabolic homeostasis. Type 2 purinergic (P2) receptors bind extracellular nucleotides and are expressed by major peripheral tissues responsible for glucose homeostasis. CD39/ENTPD1 is the dominant vascular and immune cell ectoenzyme that hydrolyzes extracellular nucleotides to regulate purinergic signaling.. We have studied Cd39/Entpd1-null mice to determine whether any associated changes in extracellular nucleotide concentrations influence glucose homeostasis.. Cd39/Entpd1-null mice have impaired glucose tolerance and decreased insulin sensitivity with significantly higher plasma insulin levels. Hyperinsulinemic-euglycemic clamp studies indicate altered hepatic glucose metabolism. These effects are mimicked in vivo by injection into wild-type mice of either exogenous ATP or an ecto-ATPase inhibitor, ARL-67156, and by exposure of hepatocytes to extracellular nucleotides in vitro. Increased serum interleukin-1beta, interleukin-6, interferon-gamma, and tumor necrosis factor-alpha levels are observed in Cd39/Entpd1-null mice in keeping with a proinflammatory phenotype. Impaired insulin sensitivity is accompanied by increased activation of hepatic c-Jun NH(2)-terminal kinase/stress-activated protein kinase in Cd39/Entpd1 mice after injection of ATP in vivo. This results in decreased tyrosine phosphorylation of insulin receptor substrate-2 with impeded insulin signaling.. CD39/Entpd1 is a modulator of extracellular nucleotide signaling and also influences metabolism. Deletion of Cd39/Entpd1 both directly and indirectly impacts insulin regulation and hepatic glucose metabolism. Extracellular nucleotides serve as "metabolokines," indicating further links between inflammation and associated metabolic derangements.

Topics: Adenosine Triphosphate; Adipose Tissue, Brown; Animals; Antigens, CD; Apyrase; Blood Glucose; Cells, Cultured; Cytokines; Energy Metabolism; Glucose Clamp Technique; Hepatocytes; Hyperinsulinism; Insulin; Insulin Receptor Substrate Proteins; Insulin Resistance; Intracellular Signaling Peptides and Proteins; JNK Mitogen-Activated Protein Kinases; Liver; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Muscle, Skeletal; Oxygen Consumption; Pancreas; Phosphoproteins; Phosphorylation; Signal Transduction

Extracellular ATP is an ubiquitous mediator that regulates several cellular functions via specific P2 plasma membrane receptors (P2Rs), for which a role in modulating intracellular glucose metabolism has been recently suggested. We have investigated glucose uptake in response to P2Rs stimulation in fibroblasts from type 2 diabetic (T2D) patients and control subjects. P2Rs expression was evaluated by RT-PCR; intracellular calcium release by fluorometry; glucose transporter (GLUT1) translocation by immunoblotting and chemiluminescence; glucose uptake was measured with 2-deoxy-D-[1-(3)H]glucose (2-DOG) and ATP by luminometry. Cells from T2D patients, in contrast to those from healthy controls, showed no increase in glucose uptake after ATP stimulation; extracellular ATP caused, however, a similar GLUT1 recruitment to the plasma membrane in both groups. P2Rs expression did not differ between fibroblasts from diabetic and healthy subjects, but while plasma membrane depolarization, a P2X-mediated response was similar in both groups, no evident intracellular calcium increase was detectable in the cells from the former group. The calcium response in fibroblasts from diabetics was restored by co-incubation with apyrase or hexokinase, suggesting that P2YRs in those cells were normally expressed but chronically desensitised. In support to this finding, fibroblasts from T2D subjects secreted a two-fold larger amount of ATP compared to controls. Pre-treatment with apyrase or hexokinase also restored ATP stimulated glucose uptake in fibroblasts from diabetic subjects. These results suggest that extracellular ATP plays a role in the modulation of glucose transport via GLUT1, and that the P2Y-dependent GLUT1 activation is deficient in fibroblasts from T2D individuals. Our observations may point to additional therapeutic targets for improving glucose utilization in diabetes.

Topics: Adenosine Triphosphate; Apyrase; Biological Transport; Calcium; Calcium Signaling; Cell Membrane; Cells, Cultured; Diabetes Mellitus, Type 2; Energy Metabolism; Extracellular Fluid; Fibroblasts; Glucose; Glucose Transporter Type 1; Hexokinase; Humans; Insulin Resistance; Membrane Potentials; Monosaccharide Transport Proteins; Receptors, Purinergic P2; Receptors, Purinergic P2Y1; RNA, Messenger