Evidence for P2Y2 Receptor Facilitation of Hyperglycemia‐induced Insulin Resistance in Human Hepatocytes

Abstract

It is known that deletion of the ectonucleotidase CD39 leads to liver insulin resistance in mice; however, it remains unknown whether this phenotype is due to a lack of extracellular adenosine generation or an excessive nucleotides accumulation and signaling. We hypothesized that the nucleotide P2Y2 receptor mediates insulin resistance in human hepatocytes. RT‐PCR showed that among the eight known P2Y receptors, only the P2Y2, P2Y6 and P2Y11 mRNAs were detected in cultured HepG2 cells, a human hepatocytes model; however, stimulation of the cells with P2Y6‐ and P2Y11‐selective agonists all failed to induce intracellular Ca2+ mobilization, whereas both ATP and UTP induced significant Ca2+ signaling in a dose‐dependent manner, which was abolished by pretreatment of the cells with ARC‐118925XX, a selective and competitive P2Y2 receptor antagonist, suggesting that the P2Y2 receptor is the only functional P2Y receptor expressed in HepG2 cells. In addition, we found that stimulation of the P2Y2 receptor induced MAPK signaling including phosphorylated ERK1/2, JNK and p38, but not the AKT pathway. Interestingly, stimulation of the cells with ATP or UTP dose‐dependently blocked insulin‐induced AKT phosphorylation, but potentiated ERK1/2 signaling, indicating a selective disruption of the insulin‐AKT signaling axis by P2Y2 receptor. Furthermore, we found that a notable release of ATP into the extracellular space was observed when exposing HepG2 cells to high glucose concentrations (25mM) as compared with normal glucose (6.1 mM). Consistent with this, we also found that extracellular nucleotides signaling through P2Y2 receptor induced a dose‐dependent reduction in hepatocyte glucose uptake. We conclude that the P2Y2 receptor mediates liver insulin resistance in response to hyperglycemia, possibly through activation of the JNK pathway and inhibition of insulin signaling to the AKT pathway, highlighting that P2Y2 receptor may be a new drug target for type‐2 diabetes.Support or Funding InformationSupported in part by NIH funding 1R01HL125279‐01A1 (JS)