Publications

Natural killer (NK) cells are a vital part of the innate immune system capable of rapidly clearing mutated or infected cells from the body and promoting an immune response. Here, we find that NK cells activated by viral infection or tumor challenge increase uptake of fatty acids and their expression of carnitine palmitoyltransferase I (CPT1A), a critical enzyme for long-chain fatty acid oxidation. Using a mouse model with an NK cell–specific deletion of CPT1A, combined with stable 13 C isotope tracing, we observe reduced mitochondrial function and fatty acid–derived aspartate production in CPT1A-deficient NK cells. Furthermore, CPT1A-deficient NK cells show reduced proliferation after viral infection and diminished protection against cancer due to impaired actin cytoskeleton rearrangement. Together, our findings highlight that fatty acid oxidation promotes NK cell metabolic resilience, processes that can be optimized in NK cell–based immunotherapies.

Abstract The growth arrest‐specific gene 6 ( Gas6 ) plays a role in pro‐atherogenic processes such as endothelial and leukocyte activation, smooth muscle cell migration and thrombosis, but its role in atherosclerosis remains uninvestigated. Here, we report that Gas6 is expressed in all stages of human and mouse atherosclerosis, in plaque endothelial cells, smooth muscle cells and macrophages. Gas6 expression is most abundant in lesions containing high amounts of macrophages, ie thin fibrous cap atheroma and ruptured plaque. Genetic loss of Gas6 does not affect the number and size of initial and advanced plaques in ApoE −/− mice, but alters its plaque composition. Compared to Gas6 +/+ : ApoE −/− mice, initial and advanced plaques of Gas6 −/− : ApoE −/− mice contained more smooth muscle cells and more collagen and developed smaller lipid cores, while the expression of TGFβ was increased. In addition, fewer macrophages were found in advanced plaques of Gas6 −/− : ApoE −/− mice. Hence, loss of Gas6 promotes the formation of more stable atherosclerotic lesions by increasing plaque fibrosis and by attenuating plaque inflammation. These findings identify a role for Gas6 in plaque composition and stability. Copyright © 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Abstract Gas6 downregulates the activation state of macrophages and thereby their production of proinflammatory cytokines induced by various stimuli. We aimed to determine whether Gas6 is involved in sepsis. We measured Gas6 plasma levels in 13 healthy subjects, 29 patients with severe sepsis, and 18 patients with non-infectious inflammatory diseases. Gas6 level was higher in septic patients than in control groups (P≤0.0001). The sensitivity and specificity of Gas6 levels to predict fatal outcome were 83% and 88%. We next investigated whether Gas6 affects cytokine production and outcome in experimental models of endotoxemia and peritonitis in wild-type (WT) and Gas6−/− mice. Circulating levels of Gas6 after LPS 25mg/kg i.p. peaked at 1 hour (P<0.001). Similarly, TNF-α was higher in Gas6−/− than in WT mice 1 hour after LPS (P<0.05). Furthermore, 62 anti- and pro-inflammatory cytokines were quantified in plasma after LPS injection. Their levels were globally higher in Gas6−/− plasma after LPS, 47/62 cytokines being at least 50% higher in Gas6−/− than in WT plasma after 1 hour. Mortality induced by 25mg/kg LPS was 25% in WT versus 87% in Gas6−/− mice (P<0.05). LPS-induced mortality in Gas6 receptors Axl−/−, Tyro3−/− and Merkd was also enhanced when compared to WT mice (P<0.001). In peritonitis models (cecal ligation and puncture, CLP, and i.p. injection of E. coli), Gas6 plasma levels increased and remained elevated at least 24 hours. CLP increased mortality in Gas6−/− mice. Finally, we explored the role of Gas6 in LPS-treated macrophages. We found that Gas6 was released by LPS-stimulated WT macrophages and that Gas6−/− macrophages produced more TNF-α and IL-6 than WT macrophages. Cytokine release by Gas6−/− macrophages was higher than by WT macrophages (cytokine array). Adjunction of recombinant Gas6 to the culture medium of Gas6−/− macrophages diminished the cytokine production to WT levels. In LPS-treated Gas6−/− macrophages, Akt and Erk1/2 phosphorylation was reduced whereas p38 and NFκB activation was enhanced. Thus, in septic patients, elevated Gas6 levels were associated with fatal outcome. In mice, they raised in experimental endotoxemia and peritonitis models, and correlated also with sepsis severity. However, Gas6−/− mice survival in these models was reduced compared to WT. Gas6 secreted by macrophages in response to LPS activated Akt and restrained p38 and NFκB activation, thereby dampening macrophage activation. Altogether these data suggest that, during endotoxemia, Gas6−/− mice phenotype resembles that of mice which have undergone PI3K inhibition, indicating that Gas6 is a major modulator of innate immunity.

These are the models and molecular dynamics trajectories obtained for the paper Role of glutamine synthetase in angiogenesis beyond glutamine synthesis [G. Eelen et al. Nature 2018 vol. 561 (7721) pp. 63-69].<br>The PDB files (human.pdb and bacterial.pdb) contain representative structures extracted from the molecular dynamics simulations of the S. Typhimurium (bacterial.pdb) and Human Glutamine Synthetase (human.pdb). For the latter, models of the two most relevant binding modes of Palmitoyl-CoA are included in the file.<br><br>The trajectories included are the FINAL 450 ns of 2 fully atomistic molecular dynamics simulations of Glutamine Synthetase (GS) performed with GROMACS and the Amber ff14SB force field. The Human_GS.x is the trajectory of human GS (reference PDB 2OJW) with Palmitoyl-CoA. In this trajecory the palmitate tail points upwards (which was the most interesting orientation found). "Bacterial_GS" is the bacterial GS trajectory (reference starting PDB 1FPY) with Palmitoyl-CoA - palmitate tail points downwards. The trajectories are in GROMACS trr format and have been saved every 200ps.<b> </b>Water molecules and the initial 50 ns (relaxation) are not included in the files.

Hyperthermia (HT) associated with radiotherapy or chemotherapy is a promising method for cancer treatment, although the molecular mechanisms of this process are not well understood. HT exhibits various antitumor effects, including damage of tumor vasculature. Here, we investigate the effect of HT on in vitro and in vivo angiogenesis. We show that heat treatment of endothelial cells (ECs) affect their differentiation into capillary-like structures in two models of in vitro angiogenesis. Furthermore, the formation of new vessels promoted by angiogenic inducers in the chick embryo chorioallantoic membrane assay is impaired after heat treatment. These effects cannot be explained by direct cytotoxicity but are dependent on modulation of angiogenesis-involved genes. Gene expression profile of ECs subjected to heat shock demonstrates that plasminogen activator inhibitor 1 (PAI-1), a protein involved in the control of extracellular matrix degradation, is specifically up-regulated. The use of anti-PAI-1-neutralizing antibodies reverts the effect of HT on the in vitro EC morphogenesis and in vivo vessel formation. Moreover, microvessel outgrowth from PAI-1(-/-) aortic rings was not affected by HT compared with aortic rings from PAI-1(+/+) mice. Heat treatment of murine mammary adenocarcinomas results in inhibition of tumor growth, associated with a reduction of microvessel number and an increase of PAI-1 expression. These results indicate that heat-mediated PAI-1 induction is an important pathway by which HT exerts its antitumor activity and may represent a rationale for a combined cancer therapy based on HT associated with antiangiogenic molecules.