Glutamine metabolism provides synergistic support for macrophage activation and elicitation of desirable immune responses; however, the underlying mechanisms regulated by glutamine metabolism to orchestrate macrophage activation remain unclear. Here we show that the production of α-ketoglutarate (αKG) via glutaminolysis is important for alternative (M2) activation of macrophages, including engagement of fatty acid oxidation (FAO) and Jmjd3-dependent epigenetic reprogramming of M2 genes. This M2-promoting mechanism is further modulated by a high αKG/succinate ratio, whereas a low ratio strengthens the proinflammatory phenotype in classically activated (M1) macrophages. As such, αKG contributes to endotoxin tolerance after M1 activation. This study reveals new mechanistic regulations by which glutamine metabolism tailors the immune responses of macrophages through metabolic and epigenetic reprogramming.

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Background: Storage of platelet concentrates (PCs) has an impact on platelet quality and possibly affects their functions after transfusion. The influence of processing and storage conditions of PCs on their in vivo function upon transfusion is unknown. One option for investigating this question is to implement an ex vivo labeling of human platelets, to analyze them after transfusion into heathy volunteers and/or patients. In this study, we developed two labeling methods employing biotin. Methods: Two methods of biotinylation were compared to a control (standard PC). The "Bio-Wash" process used washing steps to label all platelets within the PC; for the other method, "Bio-Direct," one fifth of the PC were directly labeled without washing steps. The control and the two biotinylated PCs were analyzed over 7 days of storage. Labeling efficiency, platelet counts, phenotypes, and functions, along with time and costs, were evaluated to select the best process.Results: Both methods achieved a stable labeling through the storage, with similar platelet counts and metabolism in comparison to control PCs. Bio-Wash showed higher activation phenotype and lower aggregation response in comparison to the Bio-Direct method. The Bio-Direct was performed within 1.5 h versus 3 h for the Bio-Wash. However, the Bio-Direct required 12 mg of biotin instead of 8 mg for the other process. Conclusion: We set up two methods of biotinylation that can be easily implemented in a blood bank environment. The Bio-Direct process was preferred to the Bio-Wash because of its similarity, from a functional and phenotypic point of view, with standard PCs.