NEUTROPHIL-DERIVED CIRCULATING FREE DNA (cf-DNA/NETs)

Abstract

The release of "neutrophil extracellular traps" (NETs) has been identified as a novel immune response in innate immunity. Neutrophil extracellular traps are composed of neutrophil-derived circulating free DNA (cf-DNA), histones, and neutrophil cytoplasm-derived proteins such as proteases. Here, we studied the putative predictive value of plasma cf-DNA/NETs for the development of sepsis and mortality after multiple trauma. In a prospective pilot study with 45 multiple trauma (Injury Severity Score>16) patients, cf-DNA was directly quantified in plasma. Blood samples were sequentially obtained daily from admission to our Trauma Center until day 10. Because of limited intensive care unit (ICU) stay of less than 3 days, 8 patients have been excluded, resulting in 37 patients that were evaluated. Time kinetics of cf-DNA/NETs was compared with C-reactive protein (CRP), interleukin (IL) 6, leukocyte counts, and myeloperoxidase. The severity of the injury was calculated on the basis of the Injury Severity Score, as well as Multiple Organ Dysfunction Score, Sequential Organ Failure Assessment, and Simplified Acute Physiology Score II on ICU. Initially high cf-DNA/NETs values (>800 ng/mL) with recurrent increased values between days 5 to 9 were associated with subsequent sepsis, multiple organ failure, and death. In conjunction with cf-DNA/NETs, IL-6 was significantly elevated after admission. However, the development of a second hit was not indicated by IL-6. In contrast to cf-DNA/NETs, no difference in CRP kinetics was observed between patients with and without development of sepsis. Circulating free DNA/NETs kinetics rather followed kinetics of Multiple Organ Dysfunction Score, Sepsis-related Organ Failure Assessment, leukocyte counts, and partially of myeloperoxidase. Circulating free DNA/NETs seems to be a valuable additional marker for the calculation of injury severity and/or prediction of inflammatory second hit on ICU. However, a large clinical trial with severely injured patients should confirm the prognostic value of neutrophil-derived cf-DNA/NETs.

Peptidylarginine deiminase 4 concentration, but not PADI4 polymorphisms, is associated with ICU mortality in septic shock patients

“…As we mentioned above, the citrullination of histones, in particular histone H3, is a post‐translational modification that regulates gene expression and plays a critical role in NET development and in inflammatory and host defense response 2, 3, 4, 5, 6. This reaction is catalyzed by PAD4.…”

“…Citrullination of histones, in particular histone H3, is one of the major points for inflammatory signals that trigger the neutrophil response to infections 2, 3, 4, 5. Nevertheless, citrullinated histone H3 is also important in the development of neutrophils extracellular traps (NETs) 5, 6. It has been reported that elevated levels of NETs in the plasma may predict multiple organ dysfunction and sepsis in trauma patients 6…”

Induction of Neutrophil Extracellular Traps in Shiga Toxin-Associated Hemolytic Uremic Syndrome

“…After the addition of EDTA (5 m M ) to prevent further MNase activity, the samples were centrifuged and the supernatants were collected. The DNA concentration in the PMN supernatants and the cf-DNA in the plasma samples were determined using SYBR® Gold Nucleic Acid Gel Stain (Invitrogen, Carlsbad, Calif., USA) staining as previously reported [15]. Fluorescence intensity reflects the amount of DNA and was measured using a microplate reader (VICTOR 3 ; USA).…”

“…NET induce platelet adhesion, activation, and aggregation that finally promote fibrin deposition, indicating that NET are a link between inflammation and thrombosis [14]. In patients with sepsis, there is an increase in the level of circulating free DNA (cf-DNA) in peripheral blood [15], which has been considered a marker for NETosis.…”

Cell death during sepsis: integration of disintegration in the inflammatory response to overwhelming infection

“…NET formation depends on generation of reactive oxygen species, and consequently patients with NADPH oxidase deficiency (chronic granulomatous disease) exhibit a deficiency in NET formation that may further contribute to their predisposition to severe and chronic infections. The presence and kinetics of free circulating neutrophilderived DNA/NETs appear to serve as a useful marker of sepsis disease severity and multiple organ failure in patients following multiple traumas [84]. Specific bacterial products, such as the surface expressed M protein of S. pyogenes, or host chemokines, such as IL-8, can induce the production of NETs [85,86].…”