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Presence of neutrophil extracellular traps and citrullinated histone H3 in the bloodstream of critically ill patients.

Hirose T, Hamaguchi S, Matsumoto N, Irisawa T, Seki M, Tasaki O, Hosotsubo H, Yamamoto N, Yamamoto K, Akeda Y, Oishi K, Tomono K, Shimazu T - PLoS ONE (2014)

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: Department of Traumatology and Acute Critical Medicine, Osaka University Graduate School of Medicine, Osaka, Japan.

ABSTRACT
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-α, 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.

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Representative images of immunofluorescence staining to detect neutrophil extracellular traps (NETs).NETs were visualized in the blood smear samples by immunocytochemistry and identified as extracellular string-like structures composed of chromatin (DNA and histone H3). NETs were present in the bloodstream of critically ill patients. Citrullination of histone H3 was not recognized in these images. In the blood smears surveyed in this study, we identified NETs in five patients (5/49, 10.2%). Blue, 4′,6-diamidino-2-phenylindole (DAPI); red, histone H3; green, citrullinated histone H3. Arrowheads indicate the double-stained areas containing NETs (Magnification ×400). Scale bar; 50 µm.
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pone-0111755-g002: Representative images of immunofluorescence staining to detect neutrophil extracellular traps (NETs).NETs were visualized in the blood smear samples by immunocytochemistry and identified as extracellular string-like structures composed of chromatin (DNA and histone H3). NETs were present in the bloodstream of critically ill patients. Citrullination of histone H3 was not recognized in these images. In the blood smears surveyed in this study, we identified NETs in five patients (5/49, 10.2%). Blue, 4′,6-diamidino-2-phenylindole (DAPI); red, histone H3; green, citrullinated histone H3. Arrowheads indicate the double-stained areas containing NETs (Magnification ×400). Scale bar; 50 µm.

Mentions: NETs were identified as extracellular string-like structures that were simultaneously immunoreactive for DNA and histone H3 (Fig. 2). Cit-H3 was detected by a specific antibody, and its presence was confirmed to be located inside lobulated nuclei and histone H3 (Fig. 3). In the blood smears surveyed in this study, we identified NETs in 5 patients and Cit-H3 in 11 patients (Table 2). Both NETs and Cit-H3 were identified concurrently in one patient with infection. We detected the presence of circulating NETs and/or Cit-H3-positive cells in samples from patients with infection (4/14, 28.6%), resuscitation from CPA (5/8, 62.5%), acute poisoning (1/4, 25.0%), brain stroke (3/8, 37.5%), and heat stroke (1/2, 50.0%). We found no NETs or Cit-H3-positive cells in samples from patients with trauma (0/7) or heart disease (0/4).


Presence of neutrophil extracellular traps and citrullinated histone H3 in the bloodstream of critically ill patients.

Hirose T, Hamaguchi S, Matsumoto N, Irisawa T, Seki M, Tasaki O, Hosotsubo H, Yamamoto N, Yamamoto K, Akeda Y, Oishi K, Tomono K, Shimazu T - PLoS ONE (2014)

Representative images of immunofluorescence staining to detect neutrophil extracellular traps (NETs).NETs were visualized in the blood smear samples by immunocytochemistry and identified as extracellular string-like structures composed of chromatin (DNA and histone H3). NETs were present in the bloodstream of critically ill patients. Citrullination of histone H3 was not recognized in these images. In the blood smears surveyed in this study, we identified NETs in five patients (5/49, 10.2%). Blue, 4′,6-diamidino-2-phenylindole (DAPI); red, histone H3; green, citrullinated histone H3. Arrowheads indicate the double-stained areas containing NETs (Magnification ×400). Scale bar; 50 µm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4230949&req=5

pone-0111755-g002: Representative images of immunofluorescence staining to detect neutrophil extracellular traps (NETs).NETs were visualized in the blood smear samples by immunocytochemistry and identified as extracellular string-like structures composed of chromatin (DNA and histone H3). NETs were present in the bloodstream of critically ill patients. Citrullination of histone H3 was not recognized in these images. In the blood smears surveyed in this study, we identified NETs in five patients (5/49, 10.2%). Blue, 4′,6-diamidino-2-phenylindole (DAPI); red, histone H3; green, citrullinated histone H3. Arrowheads indicate the double-stained areas containing NETs (Magnification ×400). Scale bar; 50 µm.
Mentions: NETs were identified as extracellular string-like structures that were simultaneously immunoreactive for DNA and histone H3 (Fig. 2). Cit-H3 was detected by a specific antibody, and its presence was confirmed to be located inside lobulated nuclei and histone H3 (Fig. 3). In the blood smears surveyed in this study, we identified NETs in 5 patients and Cit-H3 in 11 patients (Table 2). Both NETs and Cit-H3 were identified concurrently in one patient with infection. We detected the presence of circulating NETs and/or Cit-H3-positive cells in samples from patients with infection (4/14, 28.6%), resuscitation from CPA (5/8, 62.5%), acute poisoning (1/4, 25.0%), brain stroke (3/8, 37.5%), and heat stroke (1/2, 50.0%). We found no NETs or Cit-H3-positive cells in samples from patients with trauma (0/7) or heart disease (0/4).

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: Department of Traumatology and Acute Critical Medicine, Osaka University Graduate School of Medicine, Osaka, Japan.

ABSTRACT
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-α, 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.

Show MeSH
Related in: MedlinePlus