Liver-specific Inducible Nitric-oxide Synthase Expression Is Sufficient to Cause Hepatic Insulin Resistance and Mild Hyperglycemia in Mice

Abstract

Inducible nitric-oxide synthase (iNOS), a major mediator of inflammation, plays an important role in obesity-induced insulin resistance. Inhibition of iNOS by gene disruption or pharmacological inhibitors reverses or ameliorates obesity-induced insulin resistance in skeletal muscle and liver in mice. It is unknown, however, whether increased expression of iNOS is sufficient to cause insulin resistance in vivo. To address this issue, we generated liver-specific iNOS transgenic (L-iNOS-Tg) mice, where expression of the transgene, iNOS, is regulated under mouse albumin promoter. L-iNOS-Tg mice exhibited mild hyperglycemia, hyperinsulinemia, insulin resistance, and impaired insulin-induced suppression of hepatic glucose output, as compared with wild type (WT) littermates. Insulin-stimulated phosphorylation of insulin receptor substrate-1 (IRS-1) and -2, and Akt was significantly attenuated in liver, but not in skeletal muscle, of L-iNOS-Tg mice relative to WT mice without changes in insulin receptor phosphorylation. Moreover, liverspecific iNOS expression abrogated insulin-stimulated phosphorylation of glycogen synthase kinase-3␤, forkhead box O1, and mTOR (mammalian target of rapamycin), endogenous substrates of Akt, along with increased S-nitrosylation of Akt relative to WT mice. However, the expression of insulin receptor, IRS-1, IRS-2, Akt, glycogen synthase kinase-3␤, forkhead box O1, protein-tyrosine phosphatase-1B, PTEN (phosphatase and tensin homolog), and p85 phosphatidylinositol 3-kinase was not altered by iNOS transgene. Hyperglycemia was associated with elevated glycogen phosphorylase activity and decreased glycogen synthase activity in the liver of L-iNOS-Tg mice, whereas phosphoenolpyruvate carboxykinase, glucose-6-phosphatase, and proliferator-activated receptor ␥ coactivator-1␣ expression were not altered. These results clearly indicate that selective expression of iNOS in liver causes hepatic insulin resistance along with deranged insulin signaling, leading to hyperglycemia and hyperinsulinemia. Our data highlight a critical role for iNOS in the development of hepatic insulin resistance and hyperglycemia.The incidence of obesity and type 2 diabetes has been increasing in the United States and worldwide. Hepatic insulin resistance has a critical role in the progression of hyperglycemia in type 2 diabetes. Activation of inflammatory/stress signaling pathways have been recognized as a major culprit of hepatic insulin resistance (1, 2). These inflammatory/stress signaling pathways include the inhibitor of nuclear factor-B (IB) nuclear factor-B (NF-B), 2 c-Jun N-terminal kinase (JNK), and endoplasmic reticulum stress signaling cascades (3-7). Liver-specific activation of nuclear factor-B kinase causes hepatic insulin resistance in mice (8). Nonetheless, it remains to be determined how activation of inflammatory/stress signaling pathways induces hepatic insulin resistance.We and others have shown that the inhibition of inducible nitric-oxide synthase (iNOS, also known as NOS2) by gene disruption or pharma...

iNOS inhibitor, L-NIL, reverses burn-induced glycogen synthase kinase-3β activation in skeletal muscle of rats

“…Our results are in line with a recent study that lipid infusion decreases basal Akt/PKB phosphorylation in skeletal muscle of wild-type, but not iNOS knockout, mice [22]. Based on previous studies [17, 20], one can speculate that nitrosative protein modifications, such as S-nitrosylation and tyrosine nitration, may be possibly involved in iNOS-mediated decreases in phosphorylation of Akt/PKB. Further studies are required to clarify the precise mechanisms underlying iNOS-mediated alterations in Akt/PKB and GSK-3β activities after burn injury.…”

“…After 15-min incubation at 37°C, incorporation of 14 C radioactivity in glycogen was measured with the liquid scintillation counter. Glycogen content in muscle was measured as previously described [20] using purified glycogen (Sigma, St. Louis, MO) as a standard. Nitrotyrosine content was measured using ELISA kit (Cell Biolabs, San Diego, CA) according to the manufacturer’s instruction.…”

Regulation of obesity and insulin resistance by nitric oxide

“…Selective overexpression of NOS2 in liver is sufficient to cause hepatic insulin resistance, hyperglycemia and hyperinsulinemia [355], and the use of an NOS2-specific inhibitor (L-NIL) reversed hyperglycemia, hyperinsulinemia, and insulin resistance in ob/ob mice [150]. In obesity, proinflammatory macrophages accumulating in adipose tissue are responsible for the majority of NOS2 expression [151-153] and may propagate the inflammatory signaling implicated in insulin resistance [130].…”

Farnesyltransferase Inhibitor, Tipifarnib, Prevents Galactosamine/Lipopolysaccharide-Induced Acute Liver Failure

“…Liver homogenates were prepared as previously described (17). Protein levels in liver homogenates were determined using standard immunoblot techniques using primary antibodies (1:10,000, Cell Signaling Technology Inc., Danvers, MA, unless otherwise noted) against phosphorylated JNK at threonine183 (Thr 183 ) and tyrosine185 (Tyr 185 ) (1:1,000, Cell Signaling Technology Inc), JNK (1:5,000, Cell Signaling Technology Inc), Bcl-2 (1:1,000, Cell Signaling Technology Inc), BclxL (1:1,000, Cell Signaling Technology Inc), caspase 3, cleaved PARP, GAPDH and Vinculin.…”