Design and development of a photoswitchable DFG-out kinase inhibitor

We report the synthesis and characterisation of a photoswitchable DFG-out kinase inhibitor. Photocontrol of the target kinase in both enzymatic and living cell assays is demonstrated

Palmitoleic acid prevents palmitic acid-induced macrophage activation and consequent p38 MAPK-mediated-skeletal muscle insulin resistance

Obesity and saturated fatty acid (SFA) treatment are both associated with skeletal muscle insulin resistance (IR) and increased macrophage infiltration. However, the relative effects of SFA and unsaturated fatty acid (UFA)-activated macrophages on muscle are unknown. Here, macrophages were treated with palmitic acid, palmitoleic acid or both and the effects of the conditioned medium (CM) on C2C12 myotubes investigated. CM from palmitic acid-treated J774s (palm-mac-CM) impaired insulin signalling and insulin-stimulated glycogen synthesis, reduced Inhibitor κBα and increased phosphorylation of p38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase in myotubes. p38 MAPK inhibition or siRNA partially ameliorated these defects, as did addition of tumour necrosis factor-α blocking antibody to the CM. Macrophages incubated with both FAs generated CM that did not induce IR, while palmitoleic acid-mac-CM alone was insulin sensitising. Thus UFAs may improve muscle insulin sensitivity and counteract SFA-mediated IR through an effect on macrophage activation

Lactoferrin prevents LPS-induced decrease of the iron exporter ferroportin in human monocytes/macrophages.

Iron balance is tightly linked to inflammation and it has been demonstrated that many proteins involved in cellular iron management are up- or down-regulated by inflammatory stimuli, ultimately leading to iron retention in the reticuloendothelial system. Ferroportin is a key player in maintenance of correct iron homeostasis, because it is the only known mammalian cellular iron exporter. In this work we show that incubation of THP-1 monocytes/macrophages with lactoferrin prevents the LPS-induced decrease of ferroportin by reducing secretion of IL-6. © 2014 Springer Science+Business Media New York.Iron balance is tightly linked to inflammation and it has been demonstrated that many proteins involved in cellular iron management are up- or downregulated by inflammatory stimuli, ultimately leading to iron retention in the reticuloendothelial system. Ferroportin is a key player in maintenance of correct iron homeostasis, because it is the only known mammalian cellular iron exporter. In this work we show that incubation of THP-1 monocytes/macrophages with lactoferrin prevents the LPS-induced decrease of ferroportin by reducing secretion of IL-6

ARAP2 promotes GLUT1-mediated basal glucose uptake through regulation of sphingolipid metabolism

Lipid droplet formation, which is driven by triglyceride synthesis, requires several droplet-associated proteins. We identified ARAP2 (an ADP-ribosylation factor 6 GTPase-activating protein) in the lipid droplet proteome of NIH-3T3 cells and showed that knockdown of ARAP2 resulted in decreased lipid droplet formation and triglyceride synthesis. We also showed that ARAP2 knockdown did not affect fatty acid uptake but reduced basal glucose uptake, total levels of the glucose transporter GLUT1, and GLUT1 levels in the plasma membrane and the lipid micro-domain fraction (a specialized plasma membrane domain enriched in sphingolipids). Microarray analysis showed that ARAP2 knockdown altered expression of genes involved in sphingolipid metabolism. Because sphingolipids are known to play a key role in cell signaling, we performed lipidomics to further investigate the relationship between ARAP2 and sphingolipids and potentially identify a link with glucose uptake. We found that ARAP2 knockdown increased glucosylceramide and lactosylceramide levels without affecting ceramide levels, and thus speculated that the rate-limiting enzyme in glycosphingolipid synthesis, namely glucosylceramide synthase (GCS), could be modified by ARAP2. In agreement with our hypothesis, we showed that the activity of GCS was increased by ARAP2 knockdown and reduced by ARAP2 overexpression. Furthermore, pharmacological inhibition of GCS resulted in increases in basal glucose uptake, total GLUT1 levels, triglyceride biosynthesis from glucose, and lipid droplet formation, indicating that the effects of GCS inhibition are the opposite to those resulting from ARAP2 knockdown. Taken together, our data suggest that ARAP2 promotes lipid droplet formation by modifying sphingolipid metabolism through GCS