Role of triggering receptor expressed on myeloid cells 1/2 in secondary injury after cerebral hemorrhage

Abstract

Intracerebral hemorrhage (ICH) is a common severe emergency in neurosurgery, causing tremendous economic pressure on families and society and devastating effects on patients both physically and psychologically, especially among patients with poor functional outcomes. ICH is often accompanied by decreased consciousness and limb dysfunction. This seriously affects patients’ ability to live independently. Although rapid advances in neurosurgery have greatly improved patient survival, there remains insufficient evidence that surgical treatment significantly improves long-term outcomes. With in-depth pathophysiological studies after ICH, increasing evidence has shown that secondary injury after ICH is related to long-term prognosis and that the key to secondary injury is various immune-mediated neuroinflammatory reactions after ICH. In basic and clinical studies of various systemic inflammatory diseases, triggering receptor expressed on myeloid cells 1/2 (TREM-1/2), and the TREM receptor family is closely related to the inflammatory response. Various inflammatory diseases can be upregulated and downregulated through receptor intervention. How the TREM receptor functions after ICH, the types of results from intervention, and whether the outcomes can improve secondary brain injury and the long-term prognosis of patients are unknown. An analysis of relevant research results from basic and clinical trials revealed that the inhibition of TREM-1 and the activation of TREM-2 can alleviate the neuroinflammatory immune response, significantly improve the long-term prognosis of neurological function in patients with cerebral hemorrhage, and thus improve the ability of patients to live independently.

Key Words: Cerebral hemorrhage; Secondary injury; Triggering receptor expressed on myeloid cells 1/2; Neurosurgery; Inflammatory response

Core Tip: Based on the analysis of relevant research results in basic and clinical trials, this paper shows that inhibition of triggering receptor expressed on myeloid cells 1 and activation of triggering receptor expressed on myeloid cells 2 can alleviate neuroinflammatory immune response, and can significantly improve the long-term prognosis of neurological function in patients with cerebral hemorrhage, and significantly improve the ability of patients to live independently in daily life.

INTRODUCTION

Intracerebral hemorrhage (ICH) affects approximately 2 million people annually, with a mortality rate of over 40%, and only 20% of survivors live independently 6 months after the event[1]. The main cause of ICH is small cerebral vascular disease (SVD), which includes deep penetrating branch arteriosclerosis and cerebral amyloid vascular disease caused by hypertension. SVD accounts for approximately 77% of spontaneous cerebral hemorrhages[2]. However, people’s awareness of blood pressure management has significantly increased, and control measures are gradually improving. With the aging population and the widespread use of various anticoagulants, the incidence of ICH is expected to continue to increase[3]. The pathogenesis of SVD-induced ICH is slightly different depending on the location, but the main injuries to the body include primary and secondary brain injuries. Primary brain injury is tissue destruction caused by parenchymal hemorrhage, and mechanical injury is related to the space-occupying effect caused by hematoma, which is related to mortality after onset. Surgical removal of the hematoma is an important treatment option at this stage. Secondary brain injury involves a series of complex inflammatory and immune reactions triggered by damaged brain cells and related cytokines, blood and other components, which have been shown by many studies to be closely related to patients’ functional prognosis[4]. A comparison of the long-term prognosis of more than 200000 stroke patients indicated that the prognosis of hemorrhagic stroke patients was significantly worse than that of ischemic stroke patients[5]. Importantly, the secondary brain injury caused by ICH is more complex and severe than that caused by cerebral infarction, including not only neuronal necrosis and breakdown of the blood-brain barrier (BBB) but also the release of hemolysis and clotting products (e.g., hemoglobin, heme, iron, and thrombin), oxidative stress, and the toxicity of the inflammatory immune response. It is important to study the inflammatory and immune response processes associated with ICH to develop more targeted treatments for patients. During the early inflammatory response and innate immunity after ICH, microglia, centrocytes and monocytes/macrophages, which are activated to migrate from the peripheral blood to the site of injury, play a central role, and they all co-express the triggering receptor expressed on myeloid cell (TREM) receptor family[6]. Among these family members, TREM-1 represents the most studied inflammatory activating receptor, while TREM-2 is generally considered to be a suppressor of the inflammatory response. TREM-1 exists in two main forms: Membrane-bound (mTREM-1) and soluble (sTREM-1)[7]. At present, the roles of TREM-1 and TREM-2 in cerebral hemorrhage and other central nervous system (CNS)-related diseases are well studied and understood (Figure 1).

Figure 1 Triggering receptor expressed on myeloid cells signaling pathway. TLR: Toll-like receptor; TREM: Triggering receptor expressed on myeloid cells; sTREM-1: Soluble triggering receptor expressed on myeloid cells; Ig: Immunoglobulin; TM: Transmembrane; DAP12: DNAx-activating protein 12 kDa; MYD88: Myeloid differentiation factor 88; SYK: Spleen tyrosine kinase; ZAP70: Zeta chain-associated protein kinase 70; IRAK1: Interleukin 1 receptor associated kinase 1; TRAF: Tumor necrosis factor receptor-associated factor; TAK1: Transforming growth factor-beta-activated kinase 1; TAB: Transforming growth factor-beta-activated kinase 1-associated binding protein; ERK: Extracellular signal-regulated kinase; RIP2: Receptor-interacting protein 2; CARD9: Caspase recruitment domain family member 9; GRB2: Growth factor receptor bound protein 2; SOS: Son of sevenless; PI3K: Phosphoinositide 3-kinases; AKT: Protein kinase B; PLC: Phosphatidylinositol-specific phospholipase C; IKK: I-κB kinase; NF-κB: Nuclear factor κB; CREB: Cyclic adenosine monophosphate reactive element binding protein; HMGB1: High mobility group protein 1; PGLYRP1: Peptidoglycan recognition protein 1; HSP70: Heat shock 70 kDa protein; eCIRP: Extracellular cold-inducible RNA-binding protein; TLR4: Toll-like receptor 4; ROS: Reactive oxygen species; PKC: Protein kinase C; NLRP3: NLR family pyrin domain containing 3; MAPK: Mitogen-activated protein kinase; MMP9: Matrix metallopeptidase 9; ZO-1: Zonula occludens-1; BBB: Blood-brain barrier.

STRUCTURE AND FUNCTION OF TREM-1

TREM-1 is a transmembrane glycoprotein encoded by the TREM gene located on human chromosome 6. It has 3 distinct domains: An IG-like structure (most likely responsible for ligand binding), a transmembrane portion, and a cytoplasmic tail linked to DNAx-activating protein 12 kDa (DAP12). This complex is stabilized by a unique electrostatic interaction between the negatively charged (-) aspartate in DAP12 and the positively charged (+) lysine in the cytoplasm[8,9]. Following TREM-1 activation, the cytoplasmic portion of DAP12 containing the immunoreceptor tyrosine-activating motif is phosphorylated at its tyrosine residue, providing a docking site for protein tyrosine kinases: Zeta chain-associated protein kinase 70 (ZAP70) and spleen tyrosine kinase (SYK) subsequently transcribe genes encoding various inflammatory factors, chemokines, and cell surface molecules through a complex series of intracellular signaling processes. In addition, TREM-1-induced phosphoinositide 3-kinases (PI3K)/extracellular signal-regulated kinase (ERK) pathway activation can ensure mitochondrial integrity, thereby maintaining mitochondrial structure and function[10]. Stimulation of dendritic cells with lipopolysaccharide (LPS), bacteria, or fungi did not induce TREM-1 expression, whereas TREM-1 was strongly upregulated in neutrophils and monocytes. The selective expression of TREM-1 on neutrophils and monocytes suggests that it plays a role primarily in acute inflammatory responses. Studies have confirmed that sepsis, lung infection, urinary tract infection, and other organ system infectious diseases are strongly correlated with this disease and have become therapeutic targets. Moreover, TREM-1 plays an important role in many aseptic inflammatory processes, tumor processes, and immune system disease processes, including some inflammation-related degenerative diseases. For example, TREM-1 plays an important role in the occurrence and metastasis of lung cancer, liver cancer, prostate cancer, gastric cancer, and other diseases and provides a new target for future tumor intervention and treatment[11-14]. Recently, the expression of TREM-1 in glioma was shown to be related not only to the malignancy of the tumor but also a potential independent prognostic factor and immune target, providing a new approach to improve the efficacy of immunotherapy in glioma patients[15]. During ICH, TREM-1 amplifies inflammation and innate immune responses triggered by damage-associated molecular patterns (DAMPs) generated during secondary injury. After ICH, timely TREM-1 intervention therapy, such as the TREM-1 inhibitory proteins LP17 and LR12, can significantly improve the functional prognosis of patients. Although the role of TREM-1 in inflammation and the immune response has been confirmed, the ligand of TREM-1 remains unclear[16]. At present, many studies have focused on LPS, high mobility group protein 1 (HMGB-1), peptidoglycan recognition protein 1 (PGLYRP1) and N3. Initially, TREM-1 expression was found to be correlated with bacterial LPS and other microbial products. Recent reports implicate, a neurotoxic cycle between microglia and neurons in the subarachnoid hemorrhage (SAH) model. HMGB-1, a nonhistone chromosome-binding protein, is mainly located in the eukaryotic nucleus and widely expressed in the CNS, and exists in neurons, astrocytes, microglia, oligodendrocytes and microvascular endothelial cells. Known for its strong inflammatory effects, HMGB-1 belongs to the alarm protein family and is a protein DAMP[17]. Upon activation, HMGB-1 binds to receptors such as advanced glycosylation end product receptor and toll-like receptor (TLR) and subsequently activates signaling pathways, including mitogen-activated protein kinase (MAPK) and nuclear factor κB (NF-κB). This activation results in the release of many proinflammatory molecules, such as tumor necrosis factor α (TNF-α), interleukin-1β (IL-1β), and IL-6, which ultimately lead to pathogenic effects. The injured neurons then release large amounts of HMGB1 and heat shock protein 70, which further activate microglial pyrodeath via TREM-1. However, HMGB-1 alone does not seem to trigger TREM-1 activation, and coactivation of the molecule may be required[12]. PGLYRP1 has also been identified as a possible ligand, and the PGLYRP1 protein binds to peptidoglycan and other microbial cell wall components, such as LPS, to activate the TREM-1-SYK-ERK1/2-signal transducer and activator of transcription 3 pathway in microglia, thereby causing a neuroinflammatory response. Stimulation with soluble PGLYRP1 alone does not induce TREM-1 activation, suggesting that the PGLYRP1 protein amplifies the TREM-1-mediated neuroinflammatory response and is a potential therapeutic target[18]. Recently, a new TREM-1 receptor ligand was obtained. This peptide, called N3, is part of the native immune protein PGLYRP1/Tag7 and an N-terminal peptide of the Tag7 protein, which is responsible for activating the TREM-1 signaling pathway. The resulting peptide (N9) consists of nine amino acids and can be considered a potential peptide that blocks TREM-1 signaling[19].

STRUCTURE AND FUNCTION OF TREM-2

TREM-2 is a type I immunoglobulin superfamily of cell surface receptors that consists primarily of a deformable immunoglobulin domain, a transmembrane region, and a short cytoplasmic tail outside the cell. It is an activating receptor associated with DAP12 and DAP10. Many studies have confirmed that the opposing role of TREM-1 in the inflammatory process can inhibit the immune inflammatory response mediated by TLRs, thus playing an anti-inflammatory role in the disease process[20]. In mouse experiments, TREM-2 overexpression protected mice with sepsis from fatal outcomes and organ damage, and its beneficial effect was attributed to enhanced bacterial clearance in the body, suggesting that TREM-2 is a new and attractive therapeutic target for sepsis[21]. The soluble TREM-2 (sTREM-2) protein is elevated in the cerebrospinal fluid of subjects with multiple cerebrospinal sclerosis and inflammatory nervous system disease, and we suspect that sTREM-2 is similar to sTREM-1 as a result of protease digestion. Therefore, sTREM-2 can competitively inhibit the inflammatory inhibitory effect of TREM-2. Under homeostatic conditions, TREM-2 participates in an inflammatory suppressive state and promotes microglia and oligodendrocyte differentiation. Therefore, the absence of TREM-2 signaling in immature bone marrow cell precursors leads to partial defects in these cells and/or their function. For example, defects in TREM-2 can lead to neurodegeneration and dementia. Under inflammatory conditions, TREM-2 is downregulated, whereas other DAP12-related receptors, such as TREM-1, upregulate and redirect myeloid cell differentiation and function, promoting inflammatory responses in granulocytes, monocytes, and macrophages. TREM-2 is preferentially expressed in a subpopulation of bone marrow cells, including microglia, dendritic cells, and tissue-specific macrophages (osteoclasts, Kupffer cells, adipose tissue, and alveolar, intestinal, peritoneal, adrenal, and placental macrophages). TREM-2 is activated by a wide range of endogenous and exogenous ligands and regulates the proliferation, survival, and apoptosis of these bone marrow cells. With respect to the ligand of TREM-2, the specific ligand of TREM-2 remains unknown, although a variety of endogenous and exogenous factors, including anionic molecules, phospholipids, proteoglycans, apolipoproteins, and heat shock proteins, have been hypothesized to activate TREM-2-mediated signaling[22].

STRUCTURE AND FUNCTION OF STREM-1

sTREM-1 lacks transmembrane and intracellular regions and exists in soluble form in body fluids, with the same extracellular region as TREM-1. Therefore, the sTREM-1 can bind to the same ligand as TREM-1. The origin of sTREM-1 can be explained by two hypotheses: One hypotheses is that sTREM-1 is encoded by a splicing variant of TREM-1 mRNA, in which the third exon is deleted. In the other hypothesis, sTREM-1 is produced by proteolytic cleavage of anchored TREM-1 on the surface of mature cells, and metalloproteinases are formed by proteolytic cleavage of their long near-membrane linkers to shed part of the extracellular domain of TREM-1, which has been confirmed, and its generation depends on the activation and dimerization of TREM-1[23]. A synthetic peptide that mimics the highly conserved short domain of sTREM-1 appears to weaken cytokines produced by human monocytes and protect infected animals from hyperreactivity and death. This peptide appears to be not only effective in preventing but also in downregulating the deleterious effects of proinflammatory cytokines. sTREM-1 has been used to predict the prognosis of inflammatory diseases and has certain diagnostic and predictive roles in infectious diseases[24]. After SAH, the level of sTREM-1 in the patient’s cerebrospinal fluid is dynamically increased, and early sTREM-1 levels are significantly correlated with clinical severity and prognosis[25]. A comparison of the results of serum sTREM-1 in 104 patients with cerebral hemorrhage and healthy people revealed that serum sTREM-1 may be an inflammatory biomarker for evaluating the severity of cerebral hemorrhage, predicting early neurological deterioration and poor prognosis[26]. Cell experiments demonstrated that STREM-1 could be tested directly in human body fluids. sTREM-1 can promote the growth of activated microglia in vivo and in vitro. sTREM-1 induces the hippocampus through the activation of the PI3K-protein kinase B (AKT) pathway in microglia, which promotes NF-κB entry into the nucleus, thus inducing the expression of inflammatory factors and aggravating the deterioration of the brain environment. In addition, these events disrupt synaptic plasticity in the brain[27]. When the PI3K-AKT signaling pathway is inhibited, inflammation can be relieved. Studies using Mendelian randomization are the first to suggest a link between plasma sTREM-1 genetic susceptibility and Alzheimer disease (AD) risk. In addition, a potential causal relationship between higher plasma sTREM-1 levels and an increased risk of epilepsy has been identified[28]. Therefore, sTREM-1 has high value in disease risk prediction and prognosis. The TREM-1/TLR/sTREM-1 pathway interacts, and after TREM-1 activation binds to DAP12, the protein tyrosine kinases SYK and ZAP70 are recruited, leading to the activation of the phosphatidylinositol-specific phospholipase C, PI3K, and ERK pathways, which are upstream regulators of inflammatory gene transcription. The tyrosine kinase SYK/ZAP70 can also activate the NOD-like receptor (NLR) pathway, which merges with the TLR pathway activated by myeloid differentiation factor 88 (MYD88). The TREM-1 pathway relies on the synergistic action of the TLR pathway and MYD88. Binding stimulation of MYD88 involves a downstream signaling pathway for IL-1 receptor-associated kinase and another family of junction molecules, namely, TNF receptor-associated factor. This leads to the activation of MAPK, c-Jun N-terminal kinase, and p38, as well as the activation of transcription factors. The main family of transcription factors that are activated downstream of TLR signaling are NF-κB, cyclic adenosine monophosphate reactive element-binding protein, activator protein, and ETS domain-containing protein, which are responsible for the transcription of proinflammatory cytokines and chemokines. sTREM-1 inhibits the continuation of this inflammatory pathway by clearing the TREM-1 ligand. The secondary brain injury reaction after cerebral hemorrhage is predominantly due to the destruction of the BBB (mainly from endothelial cell injury) and the aseptic inflammation and immune response triggered by the components activated and released by the dissolution of blood and red blood cells (such as thrombin, heme and iron). Microglia, centrocytes, and monocytes play the main role in this reaction. These cells include endothelial cells in the BBB. They are the main functional performers of inflammation and the immune response, and they are also the main cells expressing TREM-1/2 in the nervous system. By studying the expression and function of the TREM receptor family in these cells, many new directions are provided for the treatment of secondary brain injury in patients with cerebral hemorrhage in the future.

THE EXPRESSION AND RESEARCH PROGRESS OF TREM-1 IN MICROGLIA

Microglia, as the resident immune cells of the brain, account for approximately 10% of the cells and are the main immune-active cells in the CNS, as well as the most abundant mononuclear phagocytes in the CNS[29,30]. Microglia are highly responsive to almost any form of CNS injury or disease, manifested by increased cell volume, increased branching, upregulated or de novo synthesis of molecules on the cell surface or within the cell, and increased cell proliferation[31]. Microglia express a variety of pattern recognition receptors (PRRSs), such as TLRs and NLRs, which play a central role in innate immune and inflammatory responses and are activated primarily by pathogen-associated molecular patterns (PAMPs) from external pathogens or DAMPs released from damaged tissue cells. After ICH injury, a series of complex reactions release DAMPs on TLRs to initiate the innate immune response, thus triggering a sterile neuroinflammatory response[32]. TLRs play a key role in the innate immune response. In the SAH model, the TLR4/MYD88/NF-κB signaling pathway has been identified as a key inflammatory pathway in the pathogenesis of neuronal injury after SAH[33]. TREM-1 amplifies the innate immune response to DAMPs. TREM-1 works synergistically with PRRSs such as TLRs and NLRs to increase the production of proinflammatory cytokines, proteases, and reactive oxygen species (ROS). This receptor pathway is extremely complex. The simultaneous activation of TREM-1 and TLR4 leads to the synergistic production of proinflammatory mediators via common signaling pathways, including the PI3K, ERK1/2, IL-1 receptor-associated kinase and NF-κB pathways, and the activation of ERK can promote the proliferation and survival of microglia. In addition to TLRs, the NLR is also a member of the PRR family and can detect microbial infections but also detect sterile tissue damage. NLR works synergistically with TLR to regulate inflammatory and apoptotic responses. NOD1 and NOD2 are two well-defined cytoplasmic PRRs associated with the NLR, and TREM-1 activation/crosslinking leads to increased NOD2 expression, caspase recruitment domain family member 9 (CARD9)/NF-κB pathway activation and cytokine production. TREM-1 can activate the CARD9/NF-κB signaling pathway through interaction with SYK. Microglia TREM-1 colocalizes with intracytoplasmic SYK. Mechanistically, as a tyrosine-based immunoreceptor activation motif-associated receptor, TREM-1 can be conjugated to DAP12 via its charged lysine residue and subsequently recruit and mobilize SYK. In a Parkinson’s disease model, the TREM-1 inhibitory peptide LP17 or SYK inhibitor r406 reduced neuroinflammation via CARD9/NF-κB signaling. In 2019, TREM-1 activation was found to promote M1 (proinflammatory) polarization of microglia in a mouse model of cerebral infarction, and another major observation of this study was that TREM-1 can activate the CARD9/NF-κB and NLR family pyrin domain containing 3 (NLRP3)/caspase-1 pathways by interacting with SYK. SYK initiation activates NF-κB signaling by controlling the CARD9/B cell lymphoma 10 complex and to activate the NLRP3 inflammasome through SYK-dependent ROS production[34]. CARD9/NF-κB is essential for the transcription of inflammatory genes and chemokines, whereas NLRP3 is responsible for neutrophil infiltration. In addition, NLRP3 provides the molecular platform for procaspase-1 cleavage, which subsequently cleaves pro-IL-1β and pro-IL-18 into biologically mature IL-1 and mature IL-18 for release into the extracellular environment. Enzyme-linked immunosorbent assay results confirmed that the IL-18 levels increased significantly 3 days after reperfusion. Recent studies have shown that cell pyrodeath caused by gasdermin D (GSDMD) is essential for IL-1β secretion. After GSDMD is cleaved, activated caspase-1 can release an N-terminal fragment, which then binds to the phosphoinositol cell membrane to form a 12-14 nm membrane pore. The formed pore acts as a gate for the extracellular release of mature IL-1β. In this study, the expression of GSDMD and GSDMD-N was sharply upregulated in microglia, indicating that pyrodeath occurred in microglia after cerebral hemorrhage[35]. In animal models, TREM-1 promotes the ability of microglia to phagocytose amyloid beta (Aβ), and the selective overexpression of TREM-1 on microglia or the activation of TREM-1 signaling by agitated antibodies can improve AD-related spatial cognitive impairment, establishing TREM-1 as a potential therapeutic target for AD. In vivo and in vitro experiments have demonstrated that sTREM-1 enhances the excessive phagocytosis of microglia at neuronal synapses by activating the PI3K-AKT pathway, thereby inducing hippocampal damage, indicating that sTREM-1 may be a potential therapeutic target for brain injury[36]. In addition, in diabetes-related cognitive impairment, the inhibition of TREM-1 was found to restore fatty autophagy damage and lipid droplet accumulation in high-sugar-stimulated microglia. Lipid metabolism disorders, impaired autophagy, and the resulting chronic neuroinflammation play key roles in the occurrence and development of cognitive impairment. Fatty autophagy injury and lipid droplets accumulation in microglia stimulated by high glucose and high TREM-1 generated by injury aggravate high glucose-induced neuroinflammation in microglia in vitro and intensify neuronal functional activation in vivo, forming a vicious cycle. The inhibition of TREM-1 can reverse this damage. Blocking the TREM-1 pathway provides a new strategy for diabetes-related cognitive impairment treatment[37]. Recently, in a mouse experiment at Xuzhou Medical University, the inhibition of TREM-1 after the administration of the TREM-1 inhibitor LP17 reduced brain edema, lowered the modified neurological severity scores, and improved neurobehavioral outcomes after traumatic brain injury (early brain injury). It has also been established that the inhibition of TREM-1 reduces neuroinflammation and is related to the SYK/p38 MAPK signaling pathway[38]. P38 is a subtype of MAPK that is widely involved in the regulation of cell growth, differentiation, proliferation, and apoptosis. P38 MAPK is associated with inflammation by promoting the production of inflammatory cytokines. For example, p38 not only regulates cytokine expression by regulating the transcription factor NF-κB but also induces and activates downstream matrix metallopeptidase 9 (MMP-9) to degrade the tightly linked protein latexin 1, leading to breakdown of the BBB and consequent cerebral edema. These findings suggest that TREM-1 may be an effective therapeutic target for reducing neuroinflammation after traumatic brain injury. The inhibition of TREM-1 induction appears to reverse these trends, with a significant reduction in p38 MAPK/MMP-9, thereby improving brain edema[39]. There are also data suggesting that various nerve injuries, such as trauma, infection, or autoimmune reactions, can induce microglial PGLYRP1 expression in the CNS. In multiple animal models of neuroinflammation, we observed that PGLYRP1 promotes neuroinflammation through the TREM-1-SYK-ERK1/2-signal transducer and activator of transcription 3 axis in cultured glial cells, revealing the role of microglial PGLYRP1 as a neuroinflammatory mediator. TREM-1-induced SYK activation is involved in the transformation of proinflammatory subtypes of microglia (type M1) and the formation of neutrophil extracellular traps (NETs)[40]. In general, microglia remain static and, once activated by relevant stimuli, can be transformed by TREM-1 regulation into an increased M1 phenotype with different physiological functions or a decreased anti-inflammatory phenotype (M2 type). Proinflammatory microglia can eventually amplify the immune response and lead to increased brain damage by secreting proinflammatory cytokines and chemokines (including IL-6, IL-1β, and TNF-α)[41]. In conclusion, TREM-1 is activated by microglia through the production of DAMPs after ICH. The expression of TREM-1 is subsequently increased and activated in microglia through a series of complex co-expression modes, thereby activating microglia to release some chemokines and inflammatory factors, thereby causing and amplifying the immune inflammatory response. It can even enhance the ability of microglia to phagocytose. In addition, both AD and cerebrovascular amyloidosis are caused by abnormal deposition of amyloid beta. In AD patients, TREM-1 can increase the ability of microglia to phagocytose amyloid beta. Accordingly, can TREM-1 enhance the deposition of amyloid beta by microglia on the vascular wall of the cerebrovascular amyloidosis? Further research is needed to confirm this theory.

THE EXPRESSION AND RESEARCH PROGRESS OF TREM-1 IN NEUTROPHILS

After ICH, microglia are activated within minutes to release cytokines, chemokines, and proteases, which then coordinate the infiltration of central granulocytes from the periphery to the brain tissue at the injured site. Once in the brain, neutrophils undergo inflammation and a congenital immune response, which can exacerbate brain damage. Neutrophils also release beneficial molecules, including iron-clearing lactoferrin, which may limit iron-mediated brain damage after brain hemorrhage. In the study of the pathogenesis of Parkinson’s disease, it has been confirmed in mouse models that activating microglial TREM-1 can over-activate TREM-1 expression in neutrophils, thereby amplifying and aggravating the inflammatory response. These results support the role of TREM-1 in the pathogenesis of PD through interactions between the central and peripheral immune systems. Neutrophils are rarely found in the CNS under normal conditions. However, neutrophils can be activated in response to CNS diseases or exogenous stimuli, thereby damaging the BBB. There is increasing evidence that pathological tissue and microenvironment changes after ICH promote the release of NETs, which play important roles in the neuroinflammatory response as a mechanism of neutrophil death, mainly destroying blood vessels and endothelial cells and promoting thrombosis[42]. TREM-1 activation enhances the activation of key pathways associated with NETosis after the involvement of TLR4[43]. Neutrophils also produce myeloperoxidase and MMP, which together damage the BBB. NETs released in the first week after ICH may enhance the inflammatory immune response and promote thrombosis[44]. Excessive sympathetic nerve activity following cerebral ischemia leads to intestinal barrier breakdown and microbial component translocation, and TREM-1 activation significantly increases intestinal permeability and bacterial translocation after bleeding. Thus, TREM-1 exacerbates the innate immune response after stroke in two spatially and temporally distinct ways. In the early stages after ICH, TREM-1 expands the local intestinal medullary response to PAMPs, and TREM-1 expands the inflammatory response to post-traumatic DAMP caused by brain tissue as medullary cells infiltrate the injured brain tissue. The bacterial antigens cross the intestinal barrier, and how much of the innate immune response that arises in brain tissue injury is driven by bacterial antigen translocation immune responses within the intestinal barrier remains to be determined. These findings support a dual and synergistic innate immune response after intracranial hemorrhage that begins early in the gut microbiome and later in the brain, resulting in an enhanced and toxic innate immune response that worsens brain injury[45]. In conclusion, although TREM-1 activation in the centrocyte after ICH is beneficial, it is more likely to aggravate the neuroinflammatory response after injury, and this amplification of inflammation involves not only DAMP but also PAMP.

THE EXPRESSION AND RESEARCH PROGRESS OF TREM-1 IN MONOCYTES/MACROPHAGES

The expression and activation of TREM-1 in human neutrophils and monocytes are significantly different, with slightly lower expression of TREM-1 in monocytes at rest than in neutrophils, which are rapidly recruited by the first wave to the site of inflammation after injury, whereas monocytes are recruited by the second wave to the site of injury within 1-7 days. DAP12 not only participates in the downstream signal transduction of TREM-1 but also stabilizes the expression of TREM-1 on the cell surface, which is conducive to its polymerization. LPS in monocytes triggers two-step clustering and polymerization of TREM-1, including upregulation of the membrane expression of TREM-1 and subsequent activation after polymerization. The levels of intracellular Ca²⁺ release, ROS and cytokine production are correlated with the degree of TREM-1 aggregation[46]. Monocytes/macrophages are the main phagocytes involved in the process of atherosclerosis, and abnormally elevated blood lipids activate TREM-1, resulting in an increase in monocytes/macrophages during bone marrow differentiation and the phagocytosis of lipids to form foam cells. Therefore, TREM-1 plays an important role in the process of atherosclerosis[47]. Therefore, in patients with SVD, TREM-1 is known to promote the chronic inflammatory process of the blood vessel wall, and it is speculated that it may also play a role in the hardening of the small arteries of the nervous system. Since TREM-1 has been shown to be associated with Aβ deposition in AD patients, there should also be a correlation between small vessel amyloidosis in SVD patients and TREM-1. TREM-1 and cerebrovascular diseases deserve more in-depth research, and there will be more breakthroughs and discoveries related to the occurrence and prevention of ICH. A study of the time course and phenotypic characteristics of monocyte recruitment in the brain after ischemia found that “inflammatory” monocytes/macrophages (CCR2⁺Ly6C^hi Mo/MΦ) are recruited into the brain during the acute phase of injury, resulting in the early accumulation of CCR2⁺Ly6C^hi Mo/MΦ. The secondary accumulation of “patrolling” monocytes/macrophages (CX3CR1⁺Ly6C^lo Mo/MΦ) occurred several weeks later and was likely the result of the phenotypic conversion of CCR2⁺Ly6C^hi Mo/MΦ rather than the recruitment of blood-derived CX3CR1⁺Ly6C^lo Mo/MΦ. The morphological diversity of CCR2⁺Ly6C^hi Mo/MΦ and CX3CR1⁺Ly6C^lo Mo/MΦ cells indicates that they play different roles in the development of injury[48]. Although the molecular cues that drive phenotypic transitions and their impact on tissue outcomes remain to be determined, these data provide new insights into the complexity of large monocyte transport to the brain and have important implications for immune-based approaches to improving brain injury by modulating the cellular basis of inflammation, particularly for the prognosis of cerebral hemorrhage.

RESEARCH PROGRESS OF TREM-1 EXPRESSION ON BBB ENDOTHELIAL CELLS

TREM-1 is expressed on the endothelial cells of collagenase-induced ICH mice and regulates the permeability of the BBB by controlling the expression of interendothelial adhesion molecules. Many studies have confirmed that destruction of the integrity of the BBB after cerebral hemorrhage and the subsequent increase in permeability are the key factors that induce secondary brain injury. Damage to the BBB can lead to brain edema, neuroinflammation caused by the susceptibility of white blood cells, and the entry of harmful molecules. Therefore, maintaining the integrity and permeability of the BBB is a potential target of ICH. Endothelial β-catenin is an essential intracellular mediator for maintaining the integrity of the BBB, and the phosphorylation of β-catenin, as a degenerate form, can lead to the downregulation of the interendothelial junction, leading to breakdown of the BBB. Studies have shown that the TREM-1 receptor increases ICH by regulating the permeability of the BBB after brain edema in mice, and this effect is mediated by the SYK/β-catenin signaling pathway to influence the expression of interendothelial junction molecules. Therefore, inhibition of the TREM-1 receptor by LP17 can reduce BBB permeability[15]. TREM-1 can not only increase the adhesion of bone marrow cells to endothelial cells and extracellular interstitial molecules but also stimulate the recruitment of other cells to the site of inflammatory injury and damage endothelial cells through microglia by producing MMPs or ROS, thus indirectly destroying the integrity and permeability of the BBB[37]. In addition, the inhibition of TREM-1 reduces the production of thrombin. The secondary injury caused by thrombin after cerebral hemorrhage can destroy the permeability of the BBB and aggravate cerebral edema. Therefore, TREM-1 inhibitors can reduce the toxic effects of thrombin on brain cells[49].

THE ROLE OF TREM-2 IN MICROGLIA AND MACROPHAGES

The myelin sheath surrounding the axons of neurons will undergo myelin remodeling during growth and injury, that is, the original myelin sheath will be shed to form a new myelin sheath. These detached myelin fragments are formed by microglia through the combination of TREM-2 and myelin lipid components and play a phagocytic role. Therefore, in the absence of TREM-2, neuronal and myelin regeneration are impeded during ICH recovery. Thus, TREM-2 can be used as a therapeutic target after ICH[50,51]. In addition, TREM-2 can also mediate the phagocytosis of apoptotic neurons and accumulated amyloid protein in microglia, regulate the relationship between microglia and lipid metabolism, and inhibit the production of inducible nitric oxide synthase, TNF-α, IL-1β, and IL-6 by negatively regulating TLR receptors. TREM-2 was positively correlated with C-X-C motif chemokine ligand 1 in relevant databases. Therefore, it also plays a certain role in mediating the chemotactic aggregation of the inflammatory site of neutrophil damage[52]. TREM-2 is not expressed in monocytes, but TREM-2 can be highly expressed in macrophage-derived foam-like cells that promote atherosclerosis. In addition, similar to sTREM-1, TREM-2 can also be used as a predictor of central system diseases after exocytosis and as a therapeutic target[52]. It has been confirmed in mouse models that TREM-2 alleviates post-ischemic injury by upregulating the transforming growth factor-β/Smad2/3 signaling pathway, promoting M-type polarization of microglia, enhancing phagocytosis of microglia, and reducing the cholesterol load[53]. In addition, M1 microglia/macrophages typically release destructive proinflammatory mediators, and M2 microglia/macrophages clear cell debris and release many protective/nutritional factors through phagocytosis. Although microglia/macrophage phenotyping offers a promising pathway for promoting CNS regeneration, this is a daunting task in the clinic. Our description of the microglia/macrophage phenotype remains unclear[54]. MicroRNAs are small, highly conserved endogenous noncoding RNAs (approximately 22 nt long). They accomplish cleavage or translational inhibition by binding to the 3’-untranslated region of the target gene and are abnormally expressed in neurological diseases. Thus, they can be used as diagnostic and therapeutic targets. A rat middle cerebral artery occlusion model was established to confirm that microRNA-26a targets TREM-1 and reduces neuronal apoptosis induced by inflammation by inhibiting the TREM-1/TLR4/MyD88/NF-κB pathway[55]. There have also been some findings about TREM-1/2 in some neurodegenerative diseases, such as AD pathology characterized by extracellular Aβ deposition and the hyperphosphorylation of tau (microtubule-associated protein) within neurons, causing the loss of neurons and synapses and ultimately cognitive impairment. Antibody-dependent TREM-2 activation in microglia, which are specialized phagocytes of the CNS, increases the density of oligodendrocyte precursors in the demyelinating region, as well as the formation of mature oligodendrocytes, thereby enhancing myelin regeneration and axon integrity and contributing to the development, homeostasis, and defense of the CNS. Studies have shown that the activation and overexpression of TREM-1 in AD mouse models can increase the ability of microglia to clear Aβ, thereby improving pathological changes and related spatial cognitive impairment in AD patients[56]. In a mouse model of AD in 2022, TREM-2 was found to enhance the ability of microglia to clear Aβ[57,58]. TREM-2 can maintain the clearance of the Aβ protein by microglia through the DAP10-PI3K-AKT-glycogen synthase kinase 3β-mammalian target of rapamycin pathway and the DAP12-SYK-PI3K-AKT-phosphatidylinositol-specific phospholipase C γ pathway[59-62]. Therefore, these findings seem to confirm that TREM-1 and TREM-2 are not completely opposed in some diseases.

TREM-RELATED THERAPIES

TREM-1 is a key regulator of inflammation. Lu et al[63] proved that TREM-1 raised neuroinflammation by modulating microglia polarization after ICH, and the regulation was partly mediated via protein kinase C δ/CARD9 signaling pathway and increased HMGB1 activation of TREM-1. The results shown intranasal administration of LP17 significantly decreased brain edema and improved neurobehavioral outcomes at 24 and 72 hours after ICH. TREM-1 monoclonal antibody increased neurobehavior deficits, proinflammatory cytokines, and reduced M2 microglia after ICH, which was reversed with rottlerin. HMBG1 interaction with TREM-1 enhanced after ICH, and glycyrrhizin relieved neuroinflammation and promoted M2 microglia which was reversed with TREM-1 monoclonal antibody[63]. In a study, two hundred and forty-two CD1 mice were used. The ICH model was established by collagenase injection. LP17 was administered intranasally at 2 or 8 hours after ICH to inhibit TREM-1. The results also shown the expression level of the TREM-1 receptor increased rapidly as early as 6 hours after ICH, and it was mainly expressed on the endotheliocytes in the neurovascular unit. Early and delayed administration of LP17 significantly decreased brain edema and improved neurobehavioral outcomes at 24 hours after ICH[64]. We firmly believe that the research on TREM-1/2 will have a good future in the treatment of cerebral hemorrhage.

ROLE OF TREM-1/2 IN IMMUNE RESPONSES

Studies have shown that TREM-2 gene silencing can intensify the immune response, increase neuronal apoptosis and infarct volume, and further exacerbate neurological dysfunction, but at present, the peripheral immune response of TREM-1/2 seems to be little in the study of cerebral hemorrhage, but it is also an interesting direction for future research.

CONCLUSION

Whether it is in vitro cell experiments, mouse experiments, or human experiments, the signaling pathway involved in TREM-1/2 signaling after ICH is very complex and co-expressed with a variety of PRRSs, such as TLRs and NLRs. The modes of cell death include not only apoptosis and necrosis but also pyrodeath. However, the specific mechanism involved needs to be further verified and explored in the future. In the secondary neuroinflammatory immune response after cerebral hemorrhage, the activation of various immune cells and the release of inflammatory factors, including the destruction of the BBB and the production of thrombin, participate in and regulate the TREM receptor family. Therefore, timely intervention and regulation of TREM-1/2 after ICH can prevent secondary brain injury and improve the long-term functional prognosis of patients. At present, four TREM-1 drugs are currently under preclinical study in China, including the TREM-1 inhibitors nangibotide and GF9 and the anti-TREM-1 monoclonal antibodies CEL383 and PY159, which are mainly used in observational studies of autoimmune diseases and tumors. Certain therapeutic effects have been achieved[65-69], and we look forward to subsequent research on the treatment of secondary injuries after cerebral hemorrhage. With the continuous progress of nanotechnology and the improvement of nano drug delivery system, nanotechnology is expected to be a new therapeutic strategy for the in-depth treatment of cerebral hemorrhage.

ACKNOWLEDGEMENTS

The authors give their thanks to all those who have helped with this issue.