IKK and JNK pathway activation can blunt Akt activation and cause insulin resistance,36, 37 and ceramide accumulation also has been implicated in the development of hepatic insulin resistance,11 with activation of IKK and NF-κB triggering ceramide synthesis

and blunting Akt signaling.11 Here we report that reductions in mitochondrial fatty acid oxidation were associated with reduced Akt phosphorylation, although hepatic ceramide content was actually lower for HET-MTP than for WT mice. In addition, there was no apparent enhanced activation in selleck the JNK and NF-κB pathways, as indicated by the lack of differences between genotypes in JNK, phospho-JNK, or IKK-β. Hepatic DAGs are thought to activate classic and atypical PKCs and blunt insulin signaling at the insulin receptor and insulin receptor substrate.13 There are numerous studies implicating hepatic DAGs in potentially causing hepatic insulin resistance,38-42 although several recent studies refute DAGs role (see recent perspectives13, 14). While we observed dysregulated insulin signaling at IRS-2 (phosphorylation CHIR-99021 manufacturer at Ser731 is counterregulatory) and Akt, DAGs were not elevated in HET mice. In addition, PKC-ϵ (the predominant

isoform activated in the liver13) activation was not increased. Similar to our findings, deficiency in long-chain acyl-CoA dehydrogenase (LCAD−/−) results in hepatic insulin resistance,4 which the authors attributed to PKC-ϵ activation due to elevated hepatic DAG synthesis during insulin stimulation. However, in humans LCAD is a redundant enzyme and apparently has a limited role in mitochondrial long chain fatty acid oxidation, and to date there are no reports of its deficiency. Unfortunately, we did not assess hepatic DAG content after the hyperinsulinemic-euglycemic clamp due to the radiolabeled

tracer used during the procedures, but we did not Digestive enzyme see increases in activated PKC-ϵ (PKC-ϵ found in the membrane) following the insulin clamp, suggesting the lack of elevation in hepatic DAGs after insulin infusion and reducing the likelihood of DAGs as a cause for hepatic insulin resistance in this animal model. Due to the lack of differences in hepatic DAGs, ceramides, and the activation status of PKC-ϵ or JNK/IKKβ, we performed extensive examination of proteins involved in the mTOR pathway (RAPTOR, RICTOR, S6, S6 kinase) and found no differences between WT and HET mice. However, examination of phosphatases known to play a role in the regulation of insulin signaling (PTEN, PHLPP1, 2, and PP2A) revealed an increase in the methylation status of the catalytic subunit of PP2A in the HET mice.