Brain-death in rats increases neutrophil extracellular trap formation in donor organs

Abstract

During brain-death, increased numbers of neutrophils are recruited to organs as part of the inflammatory response. In the organ microenvironment, the recruited neutrophils may release neutrophil extracellular traps (NETs) through interaction with various pro-inflammatory stimuli, contributing to brain-death-induced endothelial activation, microthrombus formation and ultimately a decline in organ quality. To investigate whether NETs form in organs from brain-dead donors; kidneys, hearts, livers, and plasma samples were collected from brain-dead or sham-operated rats. The presence of NET-specific components, neutrophils and macrophages were analyzed through immunofluorescent microscopy. Endothelial activation and platelet infiltration were analyzed through immunohistochemistry and qRT-PCR analysis. Plasma free thiol levels were used to evaluate systemic oxidative stress. Increased neutrophils, NETs and NET/neutrophil ratios were observed in kidneys, hearts and livers of brain-dead rats compared to sham-operated rats. Numbers of NETs positively correlated with the extent of endothelial cell activation. Brain-dead animals also had increased kidney and liver macrophages, increased infiltrated platelets in the liver, and elevated systemic oxidative stress, compared to sham-operated animals. Our findings established the presence of NETs in organs from a brain-dead donor model and suggest that NETs, alongside increased inflammation and a redox imbalance, might prime organs for microvascular endothelial dysfunction and increased injury during brain-death.