Neutrophil extracellular traps, damage-associated molecular patterns, and cell death during sepsis

Abstract

In addition to pathogen-associated molecular patterns from invasive microorganisms, alarmins, which are major components of host defense mechanisms, are involved in the pathophysiology of sepsis. In fact, the magnitude of the insult is defined according to the damage-associated molecular pattern (DAMP), which is composed of alarmins as well as pathogen-associated molecular patterns, such as those involving nucleosomes, histones, and DNA. Regarding the antimicrobial mechanism of neutrophils, an alternative non-phagocytic mechanism was first recognized as "NETosis" in 2004. In this mechanism, microorganisms are trapped and eliminated by neutrophil extracellular traps (NETs). These NETs are composed of histones and DNA that have been expelled from the nucleus as well as antimicrobial proteases, including elastase and myeloperoxidase. NETosis, a cell death pathway reported to be distinct from apoptosis, is an active area of research. As NETs are composed of deleterious substances, they are extremely harmful to the host cells once they are released into the circulating blood. Therefore, the meanings and putative roles of these components in sepsis have attracted much attention.

Keywords: Alarmin; histone; innate immune system; necrosis; neutrophil.

Figure 1

Apoptosis, necrosis, and formation of neutrophil extracellular traps (NETs). Apoptotic neutrophils finally form apoptotic bodies and are phagocytosed by macrophages without eliciting inflammation. In contrast, necrotic neutrophils cause inflammation by releasing proteolytic enzymes. The other form of cell death is NETosis. Neutrophils expose NETs, formed by DNA, histone, and granules, and cause inflammation.

Figure 2

Representative images showing fluorescent immunostaining of neutrophil extracellular traps (NETs). Neutrophils from a healthy volunteer were treated with phorbol myristate acetate for 4 h, then fixed with 4% formalin on the cover glass, and stained with three different colors: green, anti‐histone H3 monoclonal antibody; blue, DNA labeled with 4′,6‐diamidino‐2‐phenylindole (DAPI); red, anti‐elastase monoclonal antibody (objective: ×40). Scale bar = 40 μm. Original data.

Figure 3

Mortality and histological changes after histone injection. Mice were treated with intravascular injection of different doses of histone H3. As a result, a dose‐dependent increase in mortality was recognized (left panel). Mice were killed 3 h after treatment, and macroscopic findings of the lung showed massive bleeding and edema in a dose‐dependent manner (top right). A representative microscopic image of the lung from animal number 12 shows significant bleeding surrounding the trachea and vessels at 3 h after histone H3 injection (bottom right). Original data.

Figure 4

Changes in circulating alanine aminotransferase ALT, nucleosomes, and high mobility group box 1(HMGB1), after lipopolysaccharide (LPS) injection. Plasma levels of ALT, nucleosomes, and HMGB1 were measured 3 h after LPS (8 mg/kg) injection. As cell death usually becomes evident several hours after the insult, significant increases were recognized in nucleosomes and HMGB1 but not in ALT. Therefore, the increases are thought to be mainly from the result of cell death by neutrophil extracellular traps (NETosis). Original data.