Induction of neutrophil extracellular traps during tissue injury: Involvement of STING and Toll‐like receptor 9 pathways

Abstract

Objectives: Neutrophils are thought to release neutrophil extracellular traps (NETs) to form in response to exogenous bacteria, viruses and other pathogens. However, the mechanisms underlying NET formation during sterile inflammation are still unclear. In this study, we would like to identify neutrophil extracellular traps formation during sterile inflammation and tissue injury and associated pathways and its mechanism. Materials and methods:We identified different injuries such as chemical-induced and trauma-induced formation of NETs and investigated mechanism of the formation of NETs in vitro and in vivo during the treatment of mtDNA. Results:Here, we find the release of mitochondrial DNA (mtDNA) and oxidized mtDNA in acute peripheral tissue trauma models or other chemically induced lung injury, and moreover, endogenous mtDNA and oxidized mtDNA induce the formation of NETs and sterile inflammation. Oxidized mtDNA is a more potent inducer of NETs.Mitochondrial DNA activates neutrophils via cyclic GMP-AMP synthase (cGAS)-STING and the Toll-like receptor 9 (TLR9) pathways and increases the production of neutrophil elastase and extracellular neutrophil-derived DNA in NETs. Mitochondrial DNA also increases the production of reactive oxygen species (ROS) and expression of the NET-associated proteins Rac 2 and peptidylarginine deiminase 4 (PAD4). Conclusions:Altogether, these findings highlight that endogenous mitochondrial DNA inducted NETs formation and subsequent sterile inflammation and the mechanism associated with NET formation.

DAMPs and NETs in Sepsis

“…Neutrophils treated with mtDNA demonstrated increased NETosis in a manner which displayed significant increases of AKT and ERK1/2 phosphorylation and increased expression of Rac2 and PAD4. They further confirmed that both TLR9 and STING pathways are important in mtDA-induced NETosis via examination of the lungs of mice intravenously injected with mtDNA (191). Lungs displayed decreased NET formation in TLR9 KO and STING KO mice compared to wild type mice.…”

“…NETosis was completely inhibited by treatment with a TLR9 inhibitor (130). Liu et al further identified that mtDNA also activates NETosis via the STING pathway (191). Neutrophils treated with mtDNA demonstrated increased NETosis in a manner which displayed significant increases of AKT and ERK1/2 phosphorylation and increased expression of Rac2 and PAD4.…”

The role of neutrophil extracellular traps and TLR signaling in skeletal muscle ischemia reperfusion injury

“…However, growing evidence suggests that aberrant activation of these TLRs with self DNA and/or RNA occur when free nucleic material is present in the tissue, such as in autoimmune diseases, viral infections, and following traumatic injuries. 17,[25][26][27][28][29][30][31] Small molecule inhibitors of TLR7/8/9, such as hydroxychloroquine (HCQ, Plaquenil®), were developed initially as anti-malarial agents, but have shown to be efficacious for treating lupus, an autoimmune disease characterized by excess and impaired degradation of NETs. 13,24,[32][33][34] HCQ has also been shown to be an effective antiviral agent against RNA viruses including dengue virus and retroviruses 35 by de-acidifying endosomes required for endosome-mediated viral entry during TLR7/8/9 activation.…”

“…TLR 7/8 and 9 are pattern recognition receptors expressed on cells of the innate immune system, which are classically believed to recognize pathogenic RNA and DNA, respectively. However, growing evidence suggests that aberrant activation of these TLRs with self DNA and/or RNA occur when free nucleic material is present in the tissue, such as in autoimmune diseases, viral infections, and following traumatic injuries 17,25‐31 …”

Neutrophil Extracellular Traps: Signaling Properties and Disease Relevance

“…In addition, different PKC subtypes have different effects on inducing histone citrullination: PKA α inhibits histone citrullination and the formation of NETs, while PKC ζ promotes citrullination of histones and NET formation [ 55 ]. Studies have demonstrated that PI3K [ 56 ], Rac [ 57 ], TLR [ 58 ], and the Fc γ RIIIB receptor [ 59 ] participate in the formation process of NETs. Kenny et al [ 49 ] further demonstrated that the signaling mechanisms regulating NET formation were different depending on the different stimulants, indicating that the regulatory mechanism of NET formation is not a uniform process, which requires further investigation ( Figure 1 ).…”