Type 2 diabetes is associated with altered NF-κB DNA binding activity, JNK phosphorylation, and AMPK phosphorylation in skeletal muscle after LPS

Abstract

Systemic inflammation is often associated with impaired glucose metabolism. We therefore studied the activation of inflammatory pathway intermediates that interfere with glucose uptake during systemic inflammation by applying a standardised inflammatory stimulus in vivo. After ethical approval, informed consent and a thorough physical examination, 10 patients with type 2 diabetes and 10 participants with normal glucose tolerance (NGT) were given an intravenous bolus of E. coli lipopolysaccharide (LPS) of 0.3 ng/kg. Skeletal muscle biopsies and plasma were obtained at baseline and two, four and six hours after LPS. Nuclear factor (NF)-κB p65 DNA binding activity measured by ELISA, tumor necrosis factor-α and interleukin-6 mRNA expression analysed by real time reverse transcription polymerase chain reaction, and abundance of inhibitor of NF-κB (IκB)α, phosphorylated c-Jun-N-terminal kinase (JNK), AMP-activated protein kinase (AMPK), and acetyl-CoA carboxylase measured by Western blotting were detected in muscle biopsy samples. Relative to subjects with NGT, patients with type 2 diabetes exhibited a more pronounced increase in NF-κB binding activity and JNK phosphorylation after LPS, whereas skeletal muscle cytokine mRNA expression did not differ significantly between groups. AMPK phosphorylation increased in volunteers with NGT, but not in those with diabetes. The present findings indicate that pathways regulating glucose uptake in skeletal muscle may be involved in the development of inflammation-associated hyperglycemia. Patients with type 2 diabetes exhibit changes in these pathways, which may ultimately render such patients more prone to develop dysregulated glucose disposal in the context of systemic inflammation.

Trial registration: ClinicalTrials.gov NCT00412906.

Figure 1. Schematic overview of the study design.

Fig. 1 10 patients with diabetes and 10 volunteers with NGT were included in the study. After obtaining skeletal muscle biopsies and plasma samples at baseline (0 h), subjects received an intravenous bolus injection of LPS, 0.3 ng/kg. Muscle tissues biopsies and plasma samples were also obtained at 2, 4 and 6 hours after LPS. NGT: Normal glucose tolerance, LPS: lipopolysaccharide.

Figure 2. Fold increases in NF-κB, IκBα, p-JNK, TNFα mRNA and IL-6 mRNA after LPS.

Fig. 2 Fold increases (mean and 95% confidence interval) from baseline following an intravenous injection of LPS in 10 patients with diabetes (▪) and 10 volunteers with normal glucose tolerance (□). * P<0.05 compared to baseline. LPS: lipopolysaccharide, NF-κB: nuclear factor κB, IκBα: inhibitor of NF-κB α, p-JNK: phosphorylated c-Jun-N-terminal kinase, TNFα: tumour necrosis factor-alpha, IL-6: interleukin-6.

Figure 3. Fold increases in p-AMPK and p-ACC after LPS.

Fig. 3 Fold increases (mean and 95% confidence interval) from baseline following an intravenous injection of LPS in 10 patients with diabetes (▪) and 10 volunteers with NGT (□). LPS: lipopolysaccharide, NGT: normal glucose tolerance, p-AMPK: phosphorylated AMP-activated protein kinase, p-ACC: phosphorylated acetyl CoA carboxylase.