Circulating H3Cit is elevated in a human model of endotoxemia and can be detected bound to microvesicles

Abstract

Early diagnosis of sepsis is crucial since prompt interventions decrease mortality. Citrullinated histone H3 (H3Cit), released from neutrophil extracellular traps (NETs) upon binding of platelets to neutrophils following endotoxin stimulation, has recently been proposed a promising blood biomarker in sepsis. Moreover, microvesicles (MVs), which are released during cell activation and apoptosis and carry a variety of proteins from their parental cells, have also been shown to be elevated in sepsis. In a randomized and placebo-controlled human model of endotoxemia (lipopolysaccharide injection; LPS), we now report significant LPS-induced elevations of circulating H3Cit in 22 healthy individuals. We detected elevations of circulating H3Cit by enzyme-linked immunosorbent assay (ELISA), as well as bound to MVs quantified by flow cytometry. H3Cit-bearing MVs expressed neutrophil and/or platelet surface markers, indicating platelet-neutrophil interactions. In addition, in vitro experiments revealed that H3Cit can bind to phosphatidylserine exposed on platelet derived MVs. Taken together; our results demonstrate that NETs can be detected in peripheral blood during endotoxemia by two distinct H3Cit-specific methods. Furthermore, we propose a previously unrecognized mechanism by which H3Cit may be disseminated throughout the vasculature by the binding to MVs.

Citrullinated histone H3, a biomarker for neutrophil extracellular trap formation, predicts the risk of mortality in patients with cancer

“…Previous studies in the non-cancer setting have indicated an association of NETs with arterial thrombosis (de Boer et al, 2013;Borissoff et al, 2013;Mangold et al, 2015;Perez-de-Puig et al, 2015;Riegger et al, 2016). However, most studies investigated NETs at the site of thrombus formation (de Boer et al, 2013;Mangold et al, 2015;Perez-de-Puig et al, 2015;Riegger et al, 2016).…”

“…Previous studies in the non-cancer setting have indicated an association of NETs with arterial thrombosis (de Boer et al, 2013;Borissoff et al, 2013;Mangold et al, 2015;Perez-de-Puig et al, 2015;Riegger et al, 2016). However, most studies investigated NETs at the site of thrombus formation (de Boer et al, 2013;Mangold et al, 2015;Perez-de-Puig et al, 2015;Riegger et al, 2016). One study that evaluated circulating doublestranded DNA (dsDNA), nucleosomes, citrullinated Histone 4 (H4Cit) and myeloperoxidase-DNA complexes (MPO-DNA complexes) as NET formation markers found that baseline levels of dsDNA, nucleosomes and MPO-DNA are associated with the occurrence of major arterial cardiovascular events (Borissoff et al, 2013).…”

“…Therefore, we have indirectly quantified NETs by measuring biomarkers that have been proposed to reflect NET formation (Brinkmann et al , ; Martinod & Wagner, ; Vorobjeva & Pinegin, ). We used an H3Cit ELISA that has been employed in previous clinical studies (Thålin et al , , ; Paues Göranson et al , ). As no biological cut‐offs for elevated levels of biomarkers for NET formation have been established, we chose an empirical cut‐off at the 75th percentile of the distribution of H3Cit, cfDNA and nucleosome levels in the study population to define elevated levels of these biomarkers.…”

Serum citrullinated histone H3 concentrations differentiate patients with septic verses non-septic shock and correlate with disease severity

“…Studies from other investigators have unveiled CitH3 as potential diagnostic and prognostic blood marker associated with an exacerbated inflammatory response in patients with advanced cancer [ 20 ] and elevated in serval diseases like pneumonia, sepsis, and experimental endotoxemia [ 21 – 23 ]. Recently, Zuo et.al found that serum levels of CitH3 were increased in coronavirus disease 2019 (COVID-19) patients [ 24 ].…”

Reduced Neutrophil Extracellular Trap (NET) Formation During Systemic Inflammation in Mice With Menkes Disease and Wilson Disease: Copper Requirement for NET Release

“…Herein we report that in mice with either phenotype, of high (Wilson disease) and low (Menkes disease) copper levels in the liver, NET formation during endotoxemia was impaired whereas NET release during this condition is one of the hallmarks of the human reaction to LPS and has been proposed to serve as its promising blood biomarker (6,52). The fact that less NETs were formed in inflamed livers of Menkes mice correlates with hardly occurring neutrophil infiltration and impaired neutropoiesis is most probably directly responsible for this phenomenon.…”