New Insight Into Neutrophils: A Potential Therapeutic Target for Cerebral Ischemia

Abstract

Ischemic stroke is one of the main issues threatening human health worldwide, and it is also the main cause of permanent disability in adults. Energy consumption and hypoxia after ischemic stroke leads to the death of nerve cells, activate resident glial cells, and promote the infiltration of peripheral immune cells into the brain, resulting in various immune-mediated effects and even contradictory effects. Immune cell infiltration can mediate neuronal apoptosis and aggravate ischemic injury, but it can also promote neuronal repair, differentiation and regeneration. The central nervous system (CNS), which is one of the most important immune privileged parts of the human body, is separated from the peripheral immune system by the blood-brain barrier (BBB). Under physiological conditions, the infiltration of peripheral immune cells into the CNS is controlled by the BBB and regulated by the interaction between immune cells and vascular endothelial cells. As the immune response plays a key role in regulating the development of ischemic injury, neutrophils have been proven to be involved in many inflammatory diseases, especially acute ischemic stroke (AIS). However, neutrophils may play a dual role in the CNS. Neutrophils are the first group of immune cells to enter the brain from the periphery after ischemic stroke, and their exact role in cerebral ischemia remains to be further explored. Elucidating the characteristics of immune cells and their role in the regulation of the inflammatory response may lead to the identification of new potential therapeutic strategies. Thus, this review will specifically discuss the role of neutrophils in ischemic stroke from production to functional differentiation, emphasizing promising targeted interventions, which may promote the development of ischemic stroke treatments in the future.

Keywords: NETs; ROS; blood-brain barrier; ischemia; neuroinflammation; neutrophils; stroke.

Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Copyright © 2021 Chen, Zhang, Gu, Zhu, Zhong, Ye, Xiong and Jian.

Figures

Figure 1

The inflammatory response after cerebral ischemia. Acute ischemic events lead to oxidative stress and excitotoxicity, which lead to the activation of microglia and astrocytes and then promote the secretion of cytokines, MMPs and GFAP(glial fibrillary acidic protein). These proinflammatory signaling molecules lead to upregulation of the expression of ICAM-1 and selectins on endothelial cells and promote the entry of neutrophils, macrophages, lymphocytes and other blood-derived inflammatory cells into the ischemic area. Neutrophils are the first group of leukocytes to enter the CNS. At inflammatory sites, high levels of proinflammatory cytokines such as GM-CSF and TNF-α can enhance the release of ROS. ROS can induce the production of MMP-9 through signaling pathways, which leads to tissue damage and excessive inflammatory responses, such as microvascular basement membrane damage and BBB damage, and then cause more peripheral neutrophils to enter the damaged area, aggravating the nerve injury response. The activation of neutrophils can also lead to the release of nuclear and particulate matter, which form a wide network of DNA complexes (NETs) that further aggravate neuronal damage. In addition, DAMPs, which in turn activate microglia and peripheral immune cells (neutrophils, macrophages and lymphocytes), are released by dying neurons, resulting in the production of proinflammatory factors and thus leading to further activation of neutrophils. These pathological events lead to neuronal death and further increase damage to the ischemic brain.

Figure 2

(A) Active NADPH oxidase complexes promote ROS production. The activated NADPH oxidase complex uses fad to transfer electrons (e-) to oxygen to form O2− from phagocytes. O2− is produced by the activation of NADPH oxidase. O2−. O2− from phagocytes. O2− is produced by the activation of NADPH oxidase. O2− produced by NADPH oxidase can react with protons to form H2O2, which in turn produces HOCl under the action of MPO. O2− from phagocytes. O2− is produced by the activation of NADPH oxidase. O2− can react with H2O2 in the presence of Fe2+ or Cu2+ to form OH- or ROS. (B) The role of neutrophil activation in host defense and the inflammatory response. As shown on the left side, the initiation of proinflammatory cytokines or microbial molecules under physiological conditions is an immune-monitoring mechanism that ultimately enhances the antibacterial activity of neutrophils. However, as shown on the right side, excessive activation of neutrophil NADPH oxidase leads to excessive production of ROS, causing tissue damage and an excessive inflammatory response. Mediated by various proinflammatory mediators, receptor signals on the vascular endothelial surface are involved in neutrophil rolling, adhesion and endothelial barrier crossing. Phagocytosis of neutrophils leads to the activation of a series of processes, leading to the release of antimicrobial peptides, proteases, MPO and O2− from phagocytes. O2− is produced by the activation of NADPH oxidase. O2−, which is produced by the activation of NADPH oxidase, from phagocytes.

Figure 3

In the process of atherosclerosis, platelet-derived chemokines, such as CC chemokine ligand 5, promote the activation and recruitment of neutrophils. On the lumen side, activated neutrophils secrete granule proteins, including cathepsin G, which directly or indirectly promote the recruitment of myeloid cells. ROS and proteases secreted by neutrophils in the lumen and intima of atherosclerotic plaques lead to activation and dysfunction of the extracellular matrix in the layer and bottom of endothelial cells (ECs), resulting in leukocyte infiltration and low-density lipoprotein (LDL) extravasation. In the process of atherosclerotic plaque formation, the neutrophil-derived granule proteins antimicrobial peptides and α-defensin activate macrophages and cause them adopt a proinflammatory state. Neutrophils secrete MPO, which mediates the oxidation of low-density lipoprotein (oxLDL) and promotes the formation of foam cells. NETs stimulate plasma-like dendritic cells (pDCs) to produce atherogenic interferon (IFN)-α through NLRP3, and macrophages produce IL-1β and IL-18 but not AIM2 In the late stage of atherosclerosis, neutrophils can destroy plaque stability by secreting a network of proteins including cytotoxic histone H4, which penetrates and eventually dissolves vascular smooth muscle cells (VSMCs). Neutrophil-derived metalloproteinases can also induce VSMC death by degrading ECM. The death of VSMCs and degradation of ECM lead to the thinning of fiber caps and the formation of vulnerable plaques. Neutrophils can trigger epithelial cell desquamation, which is specifically manifested by the stimulation of endothelial cell stress and apoptosis by neutrophils, leading to endothelial cell detachment. This process is regulated by Toll-like receptor 2 (TLR2) signaling and other neural network signals in endothelial cells.

Figure 4

(A) The classic pathway of NETosis. Upon stimulation, ROS produced by NADPH oxidase activate the binding of PAD4 to citrullinated histones, leading to the unfolding of chromatin, and induce the translocation of enzymes such as NE and MPO in azurophilic granules to the nucleus, where they exacerbate the dissociation of histones and DNA. Finally, after the nuclear membrane and cytomembrane are dissolved, cell components including DNA, histones and granzymes are released into the intercellular space. (B) The effect of NETosis in ischemic stroke. Upon stimulation, neutrophils in the thrombus release NETs, which act as scaffolds to consolidate the thrombus. Some circulating neutrophils migrate from the blood stream into brain tissue, and they also release NETs to exacerbate neural damage.

Figure 5

The mechanism by which neutrophils repair the vascular endothelium. First, activated neutrophils deposit the antimicrobial peptide cathelicidin, which promotes the adhesion of circulating endothelial progenitor cells through FPR2, at the site of arterial injury. Endothelial progenitor cells recruited in this way can directly cover the injury site but also release angiogenic growth factor in a paracrine manner, thus promoting reendothelialization. Cathelicidin stimulates endothelial progenitor cells to secrete VEGF and EGF, which can promote the repair of damaged endothelial cells. Neutrophils penetrate the vascular endothelium by reacting with endothelial adhesion molecules to reduce blood flow velocity in the vasculature, and then neutrophils cross the vascular wall with the help of a series of adhesion molecules, such as P-, E-, and L-selectin; ICAM-1; and integrins (CD11b, a, and c).