Neutrophil extracellular trap induced by HMGB1 exacerbates damages in the ischemic brain

Abstract

It has been reported that neutrophil extracellular traps (NETs) play important roles in non-infectious diseases. In ischemic stroke, neutrophils infiltrate damaged brain tissue soon after injury and aggravate inflammation. Using a rat permanent MCAO model, we showed citrullinated histone H3 + (CitH3, a marker of NETosis) induction in neutrophils in leptomeninges and in peripheral blood soon after MCAO. Entry of CitH3 + cells occurred through leptomeninges after 6 h of MCAO and these cells were observed in cerebral cortex from 12 h and subsequently in striatum. It is interesting to note that CitH3 + induction began in circulating neutrophils before they migrated to brain parenchyma and they were detected as intact or lysed form. High mobility group box 1 (HMGB1), a danger associated molecular pattern (DAMP) molecule, was accumulated massively in serum after permanent MCAO and plays a critical role in CitH3 inductions in neutrophils in brain parenchyma and in peripheral blood. Both the all-thiol and disulfide types of HMGB1 induced CitH3 via their specific receptors, CXCR4 and TLR4, respectively. Importantly, HMGB1 not only induced NETosis but was included as a part of the extruded NETs, and contribute to NETosis-mediated neuronal death. Therefore, it would appear a vicious cycle exists between neuronal cell death and NETosis and HMGB1 mediates detrimental effects exerted by this cycle. When NETosis was suppressed by a PAD inhibitor in MCAO animals, delayed immune cell infiltrations were markedly suppressed and damages in blood vessels were significantly mitigated. The study shows NETosis with the involvement of HMGB1 as a mediator in a vicious cycle aggravates inflammation and subsequent damage in the ischemic brain. Electronic supplementary material The online version of this article (10.1186/s40478-019-0747-x) contains supplementary material, which is available to authorized users.

Adenosine Triphosphate Accumulated Following Cerebral Ischemia Induces Neutrophil Extracellular Trap Formation

“…Expression of PAD4, which induces citrullination of histones H3 (CitH3) and initiates NETosis, was found to be increased in PMNs isolated from peripheral blood (blood PMNs) following MCAO, peaking at 12 h ( Figure 1 B). Additionally, level of CitH3 (a NETosis marker) was also significantly increased at 12 h after MCAO in blood PMNs ( Figure 1 B), which is consistent with our previous report [ 7 ]. In brain parenchyma, robust inductions of PAD4 and CitH3 were also detected in cortical penumbra, where NETosis was shown to occur following MCAO [ 7 ], at 24 h after MCAO ( Figure 1 C), which is later than those in blood PMNs.…”

“…Additionally, level of CitH3 (a NETosis marker) was also significantly increased at 12 h after MCAO in blood PMNs ( Figure 1 B), which is consistent with our previous report [ 7 ]. In brain parenchyma, robust inductions of PAD4 and CitH3 were also detected in cortical penumbra, where NETosis was shown to occur following MCAO [ 7 ], at 24 h after MCAO ( Figure 1 C), which is later than those in blood PMNs. It is notable that PAD4 level was slightly decreased until 12 h post-MCAO and then markedly increased at 24 h ( Figure 1 C).…”

“…Interleukin-33 released from liver sinusoidal endothelial cells aggravates liver damage during hepatic ischemia/reperfusion by inducing excessive inflammation and NETosis [ 6 ]. Recently, we reported that high mobility group box 1 (HMGB1), a danger associated molecular patterns (DAMPs), triggers NET formation and exacerbates neuronal damage in the post-ischemic brain [ 7 ].…”

“…Similarly, CitH3-positive neutrophils were found in perivascular spaces, in brain parenchyma near the blood vessels, and in lumen of the capillary in an animal model of permanent middle cerebral artery occlusion (MCAO) [ 10 ]. In our previous study, we have demonstrated an induction of NETosis by HMGB1 (a prototypical DAMP) in the post-ischemic brain and shown that suppression of NET formation mitigates delayed inflammation and vessel damage [ 7 ]. Recently, Kang et al [ 11 ] also reported that NETs impair revascularization and vascular remodeling after stroke.…”

Protein Deimination Signatures in Plasma and Plasma-EVs and Protein Deimination in the Brain Vasculature in a Rat Model of Pre-Motor Parkinson’s Disease

“…Deiminated proteins also expose neo-epitopes which, in addition to leakage of deiminated proteins from dying cells, can further contribute to neuro-inflammatory responses. This also includes histone deimination which, in addition to gene regulatory effects, may cause extracellular trap formation which can contribute to local tissue damage [118]. Increased levels of deiminated histone H3 were here observed in the dentate gyrus as well as in cortex of the pre-motor PD brains.…”