RESUMO
ABSTRACT: Severe burns are associated with massive tissue destruction and cell death where nucleus histones and other damage-associated molecular patterns are released into the circulation and contribute to the pathogenesis of multiple-organ dysfunction. Currently, there is limited information regarding the pathophysiology of extracellular histones after burns, and the mechanisms underlying histone-induced vascular injury are not fully understood. In this study, by comparing the blood samples from healthy donors and burn patients, we confirmed that burn injury promoted the release of extracellular histones into the circulation, evidenced by increased plasma levels of histones correlating with injury severity. The direct effects of extracellular histones on human endothelial monolayers were examined, and the results showed that histones caused cell-cell adherens junction discontinuity and barrier dysfunction in a dose-related manner. Like burn patients, mice subjected to a scald burn covering 25% total body surface area also displayed significantly increased plasma histones. Intravital microscopic analysis of mouse mesenteric microcirculation indicated that treatment with a histone antibody greatly attenuated burn-induced plasma leakage in postcapillary venules, supporting the pathogenic role of extracellular histones in the development of microvascular barrier dysfunction during burns. At the molecular level, intrigued by the recent discovery of C-type lectin domain family 2 member D (Clec2d) as a novel receptor of histones, we tested its potential involvement in the histone interaction with endothelial cells. Indeed, we identified abundant expression of Clec2d in vascular endothelial cells. Further proximity ligation assay demonstrated a close association between extracellular histones and endothelial expressing Clec2d. Functionally, in vivo administration of an anti-Clec2d antibody attenuated burn-induced plasma leakage across mesenteric microvessels. Consistently, Clec2d knockdown in endothelial cells partially inhibited histone-induced endothelial barrier dysfunction. Together, our data suggest that burn injury-induced increases in circulating histones contribute to microvascular leakage and endothelial barrier dysfunction via a mechanism involving the endothelial Clec2d receptor.
