Presence of Neutrophil Extracellular Traps and Citrullinated Histone H 3 in the Bloodstream of Critically Ill Patients

Neutrophil extracellular traps (NETs), a newly identified immune mechanism, are induced by inflammatory stimuli. Modification by citrullination of histone H3 is thought to be involved in the in vitro formation of NETs. The purposes of this study were to evaluate whether NETs and citrullinated histone H3 (Cit-H3) are present in the bloodstream of critically ill patients and to identify correlations with clinical and biological parameters. Blood samples were collected from intubated patients at the time of ICU admission from April to June 2011. To identify NETs, DNA and histone H3 were visualized simultaneously by immunofluorescence in blood smears. Cit-H3 was detected using a specific antibody. We assessed relationships of the presence of NETs and Cit-H3 with the existence of bacteria in tracheal aspirate, SIRS, diagnosis, WBC count, and concentrations of IL-8, TNF-a, cf-DNA, lactate, and HMGB1. Forty-nine patients were included. The median of age was 66.0 (IQR: 52.5–76.0) years. The diagnoses included trauma (7, 14.3%), infection (14, 28.6%), resuscitation from cardiopulmonary arrest (8, 16.3%), acute poisoning (4, 8.1%), heart disease (4, 8.1%), brain stroke (8, 16.3%), heat stroke (2, 4.1%), and others (2, 4.1%). We identified NETs in 5 patients and Cit-H3 in 11 patients. NETs and/or Cit-H3 were observed more frequently in ‘‘the presence of bacteria in tracheal aspirate’’ group (11/22, 50.0%) than in ‘‘the absence of bacteria in tracheal aspirate’’ group (4/27, 14.8%) (p,.01). Multiple logistic regression analysis showed that only the presence of bacteria in tracheal aspirate was significantly associated with the presence of NETs and/or Cit-H3. The presence of bacteria in tracheal aspirate may be one important factor associated with NET formation. NETs may play a pivotal role in the biological defense against the dissemination of pathogens from the respiratory tract to the bloodstream in potentially infected patients. Citation: Hirose T, Hamaguchi S, Matsumoto N, Irisawa T, Seki M, et al. (2014) Presence of Neutrophil Extracellular Traps and Citrullinated Histone H3 in the Bloodstream of Critically Ill Patients. PLoS ONE 9(11): e111755. doi:10.1371/journal.pone.0111755 Editor: Nades Palaniyar, The Hospital for Sick Children and The University of Toronto, Canada Received March 9, 2014; Accepted September 30, 2014; Published November 13, 2014 Copyright: 2014 Hirose et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Data Availability: The authors confirm that all data underlying the findings are fully available without restriction. All data are included within the manuscript. Funding: This work was supported by a Grant-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology in Japan (no. 21390163, no. 25293366 and no. 25861718) and by ZENKYOREN (National Mutual Insurance Federation of Agricultural Cooperatives). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * Email: htomoya1979@hp-emerg.med.osaka-u.ac.jp . These authors contributed equally to this work. " These authors are joint first authors on this work.