HMGB1 promotes neutrophil extracellular trap formation through interactions with Toll-like receptor 4

Abstract

Although neutrophil extracellular traps (NETs) form to prevent dissemination of pathogenic microorganisms, excessive release of DNA and DNA-associated proteins can also perpetuate sterile inflammation. In this study, we found that the danger-associated molecular pattern protein high-mobility group box 1 (HMGB1) can induce NET formation. NET formation was found after exposure of wild-type and receptor for advanced glycation end products-deficient neutrophil to HMGB1, whereas deficiency of Toll-like receptor (TLR)4 diminished the ability of neutrophils to produce NETs. Incubation of neutrophils with HMGB1 significantly increased the amount of DNA and histone 3 released as well as intracellular histone 3 citrullination, a signaling event that precedes chromatin decondensation. In vivo, neutrophils isolated from bronchoalveolar lavages of mice exposed to LPS and HMGB1 showed consistently greater ability to produce NETs compared with pulmonary neutrophils from mice that received LPS alone. In contrast, mice treated with LPS and neutralizing antibody to HMGB1 had decreased amounts of the inflammatory cytokines TNF-α and macrophage inflammatory protein 2, as well as of free DNA and histone 3 in bronchoalveolar lavage fluids. Airway neutrophils from LPS-exposed mice that had been treated with anti-HMGB1 antibodies showed decreased citrullination of histone 3. These results demonstrate that interactions between HMGB1 and TLR4 enhance the formation of NETs and provide a novel mechanism through which HMGB1 may contribute to the severity of neutrophil-associated inflammatory conditions.

Antimicrobial Histones and DNA Traps in Invertebrate Immunity

“…HMGB1 is a DAMP of intracellular origin well known in vertebrates (34) but also in C. gigas, in which it enhances the Rel-dependent NF-B activation (35). Importantly, HMGB1 was recently shown to promote neutrophil ET formation through TLR-4 activation (36). Therefore, the ROS-dependent production of ETs by oyster hemocytes can be triggered by microbial agents and potentially by DAMPs released in the extracellular milieu in response to injury or cell lysis.…”

Neutrophil extracellular traps — the dark side of neutrophils

“…With this in mind, in vitro studies have identified several distinct activation pathways leading to NETosis that are believed, and in some cases have been shown, to be operative in vivo as well. These pathways include activation by integrins (complement receptor) and Toll-like receptors (14)(15)(16). Chemokine receptors are necessary for integrin activation and signaling (10,17).…”

Angiotensin converting enzyme inhibitors and angiotensin II receptor antagonist attenuate tumor growth via polarization of neutrophils toward an antitumor phenotype

“…[51][52][53] After stimulation with PMA (1 mg/mL) for 4 h, neutrophils were stained with Sytox Green (3 mM) for 30 min. Sytox Green fluorescence was measured using fluorescence microplate reader (Spectramax M2/e, Molecular Devices) at an excitation wavelength of 492 nm, and an emission wavelength of 530 nm.…”