Inflammation, lipotoxicity and mitochondrial dysfunction have been implicated in the pathogenesis of obesity-induced insulin resistance and type 2 diabetes. However, how these factors are intertwined in the development of obesity/insulin resistance remains unclear. Here, we examine the role of mitochondrial fat oxidation on lipid-induced inflammation in skeletal muscle. We used skeletal muscle-specific Cpt1b knockout mouse model where the inhibition of mitochondrial fatty acid oxidation results in accumulation of lipid metabolites in muscle and elevated circulating free fatty acids. Gene expression of pro-inflammatory cytokines, chemokines, and cytokine- and members of TLR-signalling pathways were decreased in Cpt1bm−/− muscle. Inflammatory signalling pathways were not activated when evaluated by multiplex and immunoblot analysis. In addition, the inflammatory response to fatty acids was reduced in primary muscle cells derived from Cpt1bm−/− mice. Gene expression of Cd11c, the M1 macrophage marker, was decreased; while Cd206, the M2 macrophage marker, was increased in skeletal muscle of Cpt1bm−/− mice. Finally, expression of pro-inflammatory markers was decreased in white adipose tissue of Cpt1bm−/− mice. We show that the inflammatory response elicited by elevated intracellular lipids in skeletal muscle is repressed in Cpt1bm−/− mice, strongly supporting the hypothesis that mitochondrial processing of fatty acids is essential for the lipid-induction of inflammation in muscle.

Aims Bioactives of Artemisia dracunculus L. (termed PMI 5011) have been shown to improve insulin action by increasing insulin signalling in skeletal muscle. However, it has not known if PMI 5011’s effects are retained during an inflammatory condition. We examined the attenuation of insulin action and whether PMI 5011 enhances insulin signalling in the inflammatory environment with elevated cytokines. Methods Muscle cell cultures derived from lean, overweight and diabetic obese subjects were used. Expression of pro-inflammatory genes and inflammatory response of human myotubes were evaluated by RT-PCR. Insulin signalling and activation of inflammatory pathways in human myotubes were evaluated by Multiplex protein assays. Results We found increased gene expression of MCP1 and TNFα, and basal activity of the NFkB pathway in myotubes derived from diabetic-obese subjects as compared to myotubes derived from normal-lean subjects. In line with this, basal Akt phosphorylation (Ser473) was significantly higher, while insulin-stimulated phosphorylation of Akt (Ser473) was lower in myotubes from normal-overweight and diabetic-obese subjects compared to normal-lean subjects. PMI 5011 treatment reduced basal phosphorylation of Akt and enhanced insulin-stimulated phosphorylation of Akt in the presence of cytokines in human myotubes. PMI 5011 treatment led to an inhibition of cytokine-induced activation of inflammatory signalling pathways such as Erk1/2 and IkBα-NFkB and moreover, NFkB target gene expression, possibly by preventing further propagation of the inflammatory response within muscle tissue. Conclusions PMI 5011 improved insulin sensitivity in diabetic-obese myotubes to the level of normal-lean myotubes despite the presence of pro-inflammatory cytokines.

Obesity‐associated, low‐grade, systemic inflammation is major contributor to obesity‐induced insulin resistance. Impaired insulin signalling in skeletal muscle is a key feature of type 2 diabetes. Bioactives of Artemisia dracunculus L. (termed PMI 5011) have been shown to improve insulin action by increasing insulin signalling in skeletal muscle. However, it has not known if PMI 5011’s effects are retained during an inflammatory condition. We examined the attenuation of insulin action and whether PMI 5011 enhances insulin signalling in the inflammatory environment with elevated cytokines. We found increased inflammation in skeletal muscle tissue of diet‐induced obese mice with upregulated Mcp1, Tnfα, Cxcl5, Cxcl10 and Tnfr1, Tnfr2 gene expressions. In line with this we detected increases in gene expression of MCP1 and TNFα, and basal activity of the NFkB pathway in myotubes derived from diabetic‐obese subjects as compared to myotubes derived from normal‐lean subjects. Multiplex protein analysis revealed that basal Akt phosphorylation (Ser473) was significantly higher, while insulin‐stimulated phosphorylation of Akt (Ser473) was lower in myotubes from normal‐overweight and diabetic‐obese subjects compared to normal‐lean subjects. PMI 5011 treatment reduced basal phosphorylation of Akt and enhanced insulin‐stimulated phosphorylation of Akt in the presence of cytokines in human myotubes. PMI 5011 treatment led to an inhibition of cytokine‐induced activation of inflammatory signalling pathways such as Erk1/2 and IkBα‐NFkB and moreover, NFkB target gene expression, possibly by preventing further propagation of the inflammatory response within muscle tissue. Thus PMI 5011 improved insulin sensitivity in diabetic‐obese myotubes to the level of normal‐lean myotubes despite the presence of pro‐inflammatory cytokines. Grant Funding Source: Supported by grants NCAMM (P50AT002776 NIH), ADA (1‐10‐BS‐129), NIH (RO1DK089641), COBRE (NIH 8P20GM103528), NORC (NIH 2P30‐DK072476)

Obesity is characterized by adipose tissue expansion as well as macrophage infiltration of adipose tissue, resulting in chronic inflammation in adipose tissue, which in turn has a causative role in obesity‐induced insulin resistance. During state of chronic inflammation, levels of inflammatory markers, such as IL‐1b, TNF‐a, IL‐6 increase as a result of macrophage accumulation and activation, specifically a subset of inflammatory F4/80+CD11c+ macrophage/dendritic cells in adipose tissue. It is well recognized that increased production of free fatty acids and cytokine secretion from inflamed fat tissue induce insulin resistance in other insulin‐sensitive organs, such as skeletal muscle and liver. However, whether obesity also results in skeletal muscle inflammation and whether macrophage infiltration in muscle contributes to the development of insulin resistance has not been fully elucidated. We investigated inflammatory cytokine expression and mechanisms involved in the induction of insulin resistance and infiltrated local macrophages and other immune cells in skeletal muscle tissue in diet‐induced obese mice using qRT‐PCR. We found that F4/80+, Cd11b+ macrophages and Cd11c dendritic cells were most abundant immune cells infiltrated into skeletal muscle tissue and increased in response to high fat diet feeding, while mRNA expression of cell markers for CD4+, CD8+ T cells and CD19+ B cells were lower.