Abstract
BackgroundMacrophage-derived factors contribute to whole-body insulin resistance, partly by impinging on metabolically active tissues. As proof of principle for this interaction, conditioned medium from macrophages treated with palmitate (CM-PA) reduces insulin action and glucose uptake in muscle cells. However, the mechanism whereby CM-PA confers this negative response onto muscle cells remains unknown.Methodology/Principal FindingsL6-GLUT4myc myoblasts were exposed for 24 h to palmitate-free conditioned medium from RAW 264.7 macrophages pre-treated with 0.5 mM palmitate for 6 h. This palmitate-free CM-PA, containing selective cytokines and chemokines, inhibited myoblast insulin-stimulated insulin receptor substrate 1 (IRS1) tyrosine phosphorylation, AS160 phosphorylation, GLUT4 translocation and glucose uptake. These effects were accompanied by a rise in c-Jun N-terminal kinase (JNK) activation, degradation of Inhibitor of κBα (IκBα), and elevated expression of proinflammatory cytokines in myoblasts. Notably, CM-PA caused IRS1 phosphorylation on Ser1101, and phosphorylation of novel PKCθ and ε. Co-incubation of myoblasts with CM-PA and the novel and conventional PKC inhibitor Gö6983 (but not with the conventional PKC inhibitor Gö6976) prevented PKCθ and ε activation, JNK phosphorylation, restored IκBα mass and reduced proinflammatory cytokine production. Gö6983 also restored insulin signalling and glucose uptake in myoblasts. Moreover, co-silencing both novel PKC θ and ε isoforms in myoblasts by RNA interference, but not their individual silencing, prevented the inflammatory response and restored insulin sensitivity to CM-PA-treated myoblasts.Conclusions/Clinical SignificanceThe results suggest that the block in muscle insulin action caused by CM-PA is mediated by novel PKCθ and PKCε. This study re-establishes the participation of macrophages as a relay in the action of fatty acids on muscle cells, and further identifies PKCθ and PKCε as key elements in the inflammatory and insulin resistance responses of muscle cells to macrophage products. Furthermore, it portrays these PKC isoforms as potential targets for the treatment of fatty acid-induced, inflammation-linked insulin resistance.
Highlights
Low grade inflammation provoked by immune cells contributes to insulin resistance, a major cause of Type 2 diabetes [1,2]
We recently reported that conditioned medium from palmitate-treated macrophages (CMPA) provokes insulin resistance in muscle cells at the levels of Akt signalling, glucose transporter 4 (GLUT4) translocation and glucose uptake [11], but how conditioned media (CM)-PA confers this negative response onto muscle cells is yet unknown
We have previously reported a drop in insulin-stimulated phosphorylation of Akt on Ser473 in L6 myoblasts treated with ceramides as a model of lipotoxicity [30] or with conditioned medium from palmitate-treated macrophages (CM-PA) as a model of inflammatory toxicity [11]
Summary
Low grade inflammation provoked by immune cells contributes to insulin resistance, a major cause of Type 2 diabetes [1,2]. Macrophage infiltration of the expanding adipose tissue renders adipocytes insulin-resistant, elevating circulating fatty acids and inflammatory cytokines that in turn contribute to insulin resistance in skeletal muscle [3,4,5,6]. There is growing evidence that fatty acids trigger macrophages to secrete factors that impair insulin action [5,10] In this context, we and others have shown that fatty acids activate macrophages to release cytokines that cause insulin resistance in adipocytes and muscle cells [3,5,10,11]. We recently reported that conditioned medium from palmitate-treated macrophages (CMPA) provokes insulin resistance in muscle cells at the levels of Akt signalling, glucose transporter 4 (GLUT4) translocation and glucose uptake [11], but how CM-PA confers this negative response onto muscle cells is yet unknown. The mechanism whereby CM-PA confers this negative response onto muscle cells remains unknown
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