Platelet extracellular vesicles mediate transfusion-related acute lung injury by imbalancing the sphingolipid rheostat.

Transfusion-related acute lung injury (TRALI) is a hazardous transfusion complication with an associated mortality of 5% to 15%. We previously showed that stored (5 days) but not fresh platelets (1 day) cause TRALI via ceramide-mediated endothelial barrier dysfunction. As biological ceramides are hydrophobic, extracellular vesicles (EVs) may be required to shuttle these sphingolipids from platelets to endothelial cells. Adding to complexity, EV formation in turn requires ceramide. We hypothesized that ceramide-dependent EV formation from stored platelets and EV-dependent sphingolipid shuttling induces TRALI. EVs formed during storage of murine platelets were enumerated, characterized for sphingolipids, and applied in a murine TRALI model in vivo and for endothelial barrier assessment in vitro. Five-day EVs were more abundant, had higher long-chain ceramide (C16:0, C18:0, C20:0), and lower sphingosine-1-phosphate (S1P) content than 1-day EVs. Transfusion of 5-day, but not 1-day, EVs induced characteristic signs of lung injury in vivo and endothelial barrier disruption in vitro. Inhibition or supplementation of ceramide-forming sphingomyelinase reduced or enhanced the formation of EVs, respectively, but did not alter the injuriousness per individual EV. Barrier failure was attenuated when EVs were abundant in or supplemented with S1P. Stored human platelet 4-day EVs were more numerous compared with 2-day EVs, contained more long-chain ceramide and less S1P, and caused more endothelial cell barrier leak. Hence, platelet-derived EVs become more numerous and more injurious (more long-chain ceramide, less S1P) during storage. Blockade of sphingomyelinase, EV elimination, or supplementation of S1P during platelet storage may present promising strategies for TRALI prevention.

Gastrointestinal microbiota contributes to the development of murine transfusion-related acute lung injury.

Transfusion-related acute lung injury (TRALI) is a syndrome of respiratory distress upon blood transfusion and is the leading cause of transfusion-related fatalities. Whether the gut microbiota plays any role in the development of TRALI is currently unknown. We observed that untreated barrier-free (BF) mice suffered from severe antibody-mediated acute lung injury, whereas the more sterile housed specific pathogen-free (SPF) mice and gut flora-depleted BF mice were both protected from lung injury. The prevention of TRALI in the SPF mice and gut flora-depleted BF mice was associated with decreased plasma macrophage inflammatory protein-2 levels as well as decreased pulmonary neutrophil accumulation. DNA sequencing of amplicons of the 16S ribosomal RNA gene revealed a varying gastrointestinal bacterial composition between BF and SPF mice. BF fecal matter transferred into SPF mice significantly restored TRALI susceptibility in SPF mice. These data reveal a link between the gut flora composition and the development of antibody-mediated TRALI in mice. Assessment of gut microbial composition may help in TRALI risk assessment before transfusion.