Triggering Receptor Expressed On Myeloid Cells 2 Expression Research Articles

Triggering receptor expressed on myeloid Cells-2 (TREM2) inhibits steroidogenesis in adrenocortical cell by macrophage-derived exosomes in lipopolysaccharide-induced septic shock

PurposeEndogenously produced glucocorticoids exhibit immunomodulating properties and are of pivotal importance for sepsis outcome. Uncontrolled activation of the immune-adrenal crosstalk increases the risk of sepsis-related death. Triggering receptor expressed on myeloid cells-2 (TREM2) is richly expressed on macrophages and has been demonstrated to improve outcome of sepsis by enhancing elimination of pathogens. However, the role and mode of action of macrophage TREM2 on adrenocortical steroidogenesis remains unclear in septic shock. MethodsThe acute septic shock model was established by intraperitoneally challenging wild-type (WT) and TREM2 knock-out (Trem2−/−) mice with lipopolysaccharide (LPS, 30 mg/kg). The mice were assessed for TREM2 expression and local inflammation in adrenal gland and for synthesis of corticotropin releasing hormone (CRH) and adrenocorticotropic hormone (ACTH) in vivo. Bone marrow-derived macrophages or macrophage-derived exosomes were isolated from WT and Trem2−/− mice and were co-cultured with adrenocortical cells. The expression of steroidogenic enzymes and corticosterone production was assessed. ResultsGenetic deficiency of TREM2 caused significantly higher corticosterone levels at the early stage of LPS-induced septic shock; whereas TREM2 deficiency neither increased CRH and ACTH nor exacerbated the inflammation in adrenocortical tissue during septic shock. Ex vivo study revealed that Trem2−/− macrophages significantly promoted the expression of steroidogenic enzymes and increased production of corticosterone. Furthermore, Trem2−/− macrophage-derived exosomes were able to mimic Trem2−/− macrophages in enhancing adrenocortical steroidogenesis. ConclusionsAt the early stage of LPS-induced septic shock, corticosterone biosynthesis can be inhibited by macrophage TREM2 in adrenocortical cells, which might partially associate with macrophage-derived exosomes.

Microglial Hyperreactivity Evolved to Immunosuppression in the Hippocampus of a Mouse Model of Accelerated Aging and Alzheimer's Disease Traits.

Neuroinflammation is a risk factor for Alzheimer’s disease (AD). We sought to study the glial derangement in AD using diverse experimental models and human brain tissue. Besides classical pro-inflammatory cytokines, we analyzed chitinase 3 like 1 (CHI3L1 or YKL40) and triggering receptor expressed on myeloid cells 2 (TREM2) that are increasingly being associated with astrogliosis and microgliosis in AD, respectively. The SAMP8 mouse model of accelerated aging and AD traits showed elevated pro-inflammatory cytokines and activated microglia phenotype. Furthermore, 6-month-old SAMP8 showed an exacerbated inflammatory response to peripheral lipopolysaccharide in the hippocampus and null responsiveness at the advanced age (for this strain) of 12 months. Gene expression of TREM2 was increased in the hippocampus of transgenic 5XFAD mice and in the cingulate cortex of autosomal dominant AD patients, and to a lesser extent in aged SAMP8 mice and sporadic early-onset AD patients. However, gene expression of CHI3L1 was increased in mice but not in human AD brain samples. The results support the relevance of microglia activation in the pathways leading to neurodegeneration and suggest diverse neuroinflammatory responses according to the AD process. Therefore, the SAMP8 mouse model with marked alterations in the dynamics of microglia activation and senescence may provide a complementary approach to transgenic mouse models for the study of the neuroinflammatory mechanisms underlying AD risk and progression.

Effect of acupuncture on microglia activation in prefrontal cortex of chronic stress-induced depression rats

To observe the effect of acupuncture on the expression of proinflammatory cytokines interleukin-1β (IL-1β), interleukin-6 (IL-6), microglia marker ion calcium adaptor protein (Iba-1) and triggering receptor expressed on myeloid cells 2 (TREM2) in the prefrontal cortex of chronic stress-induced depression rats, so as to explore its antidepressant mechanism. Thirty-two SD rats were randomly divided into normal control, model, acupuncture and fluoxetine groups, with 8 rats in each group. The depression model was established by using chronic mild unpredictable stress methods for 6 weeks. Manual acupuncture stimulation was applied to "Baihui" (GV20) and "Yintang" (GV29) for 10 min before modeling for 6 weeks. Fluoxetine (10 mg/kg, 1 mg/mL) was given to rats of the fluoxetine group by gavage before stress stimulation, once every day for 6 weeks. The open field test was used to evaluate the behavioral changes of rats. The contents of IL-1β, IL-6 in the prefrontal cortex were determined by enzyme-linked immunosorbent assay (ELISA). Immunohistochemistry was used to determine the expression of Iba-1 in the prefrontal cortex. The TREM2 gene expression in the prefrontal cortex was determined by real time fluorescent quantitative polymerase chain reaction (RT-PCR). After modeling, the crossing numbers and rearing times were significantly decreased in the model group relevant to the control group (P<0.05). After the treatment, the crossing numbers were significantly increased in the acupuncture and fluoxetine groups (P<0.05), while the rearing times in the acupuncture group were significantly increased (P<0.05). Compared with the control group, the contents of IL-1β, IL-6 and the expression of Iba-1 positive cells in the prefrontal cortex were significantly increased in the model group (P<0.05), while the expression of TREM2 gene in the prefrontal cortex was significantly decreased (P<0.05). After the treatment, the increased levels of IL-1β, IL-6 and Iba-1 positive cells and the decreased TREM2 gene expression were considerably reversed in both acupuncture and fluoxetine groups compared with the model group (P<0.05). Acupuncture intervention plays a positive role in anti-depression in rats, which may be related to its effects in inhibiting the activation of microglia, reducing the expression of proinflammatory cytokines, and increasing TREM2 expression in the prefrontal cortex.

High expression of TREM2 promotes EMT via the PI3K/AKT pathway in gastric cancer: bioinformatics analysis and experimental verification.

Background: To date, the pathogenesis of gastric cancer (GC) remains unclear. We combined public database resources and bioinformatics analysis methods, explored some novel genes and verified the experiments to further understand the pathogenesis of GC and to provide a promising target for anti-tumor therapy.Methods: We downloaded the chip data related to GC from the Gene Expression Omnibus (GEO) database, extracted differentially expressed genes (DEGs), and then determined the key genes in the development of GC via PPI networks and model analysis. Functional annotation via GO and KEGG enrichment of DEGs was used to understand the latent roles of DEGs. The expression of the triggering receptor expressed on myeloid cells 2 (TREM2) gene in GC cell lines was verified via RT-PCR and western blotting. Moreover, the CCK-8, wound healing assay, and transwell migration and invasion assays were used to understand the changes in the proliferation, migration, and invasion abilities of GC cells after silencing TREM2. Western blotting verified the interaction between TREM2 and PI3K predict of the string website, as well as the effect of TREM2 on EMT. Finally, a lung metastasis model was used to explore the relationship between TREM2 and metastasis.Results: Our study identified 16 key genes, namely BGN, COL1A1, COL4A1, COL5A2, NOX4, SPARC, HEYL, SPP1, TIMP1, CTHRC1, TREM2, SFRP4, FBXO32, GPX3, KIF4A, and MMP9 genes associated with GC. The EMT-related pathway was the most significantly altered pathway. TREM2 expression was higher in GC cell lines and was remarkably associated with tumor invasion depth, TNM stage, histological grade, histological type, anatomic subdivision, and Helicobacter pylori state. Knockdown of TREM2 expression inhibited the proliferation, migration, and invasion of GC cells as well as the progression of EMT by PI3K/AKT signaling in vitro. In addition, lung metastasis were decreased in vivo.Conclusions: We identified some important genes associated with the progression of GC via public database analysis, explored and verified the effects of proto-oncogene TREM2 on EMT via the PI3K/AKT pathway. TREM2 may be a novel target in the GC therapy.

Resveratrol mediates mechanical allodynia through modulating inflammatory response via the TREM2-autophagy axis in SNI rat model

BackgroundNeuropathic pain (NeuP) is a chronic and challenging clinical problem, with little effective treatment. Resveratrol has shown neuroprotection by inhibiting inflammatory response in NeuP. Recently, the triggering receptor expressed on myeloid cells 2 (TREM2) expressed by microglia was identified as a critical factor of inflammation in nervous system diseases. In this study, we explored whether resveratrol could ameliorate neuroinflammation and produce anti-mechanical allodynia effects via regulating TREM2 in spared nerve injury rats, as well as investigated the underlying mechanisms.MethodsA spared nerve injury (SNI) rat model was performed to investigate whether resveratrol could exert anti-mechanical allodynia effects via inhibiting neuroinflammation. To evaluate the role of TREM2 in anti-neuroinflammatory function of resveratrol, lentivirus coding TREM2 was intrathecally injected into SNI rats to activate TREM2, and the pain behavior was detected by the von Frey test. Furthermore, 3-methyladenine (3-MA, an autophagy inhibitor) was applied to study the molecular mechanisms of resveratrol-mediated anti-neuroinflammation using Western blot, qPCR, and immunofluorescence.ResultsThe TREM2 expression and number of the microglial cells were significantly increased in the ipsilateral spinal dorsal horn after SNI. We found that intrathecal administration of resveratrol (300ug/day) alleviated mechanical allodynia; obviously enhanced autophagy; and markedly reduced the levels of interleukin-1β, interleukin-6, and tumor necrosis factor-α in the ipsilateral spinal dorsal horn after SNI. Moreover, the number of Iba-1+ microglial cells and TREM2 expression were downregulated after resveratrol treatment. Intrathecal administration of lentivirus coding TREM2 and/or 3-MA in those rats induced deficiencies in resveratrol-mediated anti-inflammation, leading to mechanical allodynia that could be rescued via administration of Res. Furthermore, 3-MA treatment contributed to TREM2-mediated mechanical allodynia.ConclusionsTaken together, these data reveal that resveratrol relieves neuropathic pain through suppressing microglia-mediated neuroinflammation via regulating the TREM2-autophagy axis in SNI rats.

TREM2 ameliorates neuroinflammatory response and cognitive impairment via PI3K/AKT/FoxO3a signaling pathway in Alzheimer's disease mice.

Triggering receptor expressed on myeloid cells 2 (TREM2) has been shown with a neuroprotective function against inflammation and neuronal injury in Alzheimer’s disease (AD). However, the TREM2 induced anti-inflammatory mechanism is still not well known. In this study it has been demonstrated that the expression of TREM2 was upregulated in hippocampus of 5xFAD mice, whereas TREM2 knock-out mediated by AAV significantly increased the levels of pro-inflammatory cytokines and aggravated cognitive defect. Additionally, FoxO3a, a downstream member of the PI3K/AKT pathway, could be activated by TREM2 defect via the PI3K/AKT signaling in 5xFAD mice. That suggests TREM2-induced protection is associated with the PI3K-FoxO3a axis. On the contrary, overexpression of TREM2 alleviated the LPS-induced inflammatory response and induced M2 phenotype microglia in vitro. This phenomenon can be abolished by applying the PI3K inhibitor LY294002, suggesting FoxO3a not only participates in TREM2-induced anti-inflammation response, but is also involved in regulating the phenotype of microglia. Taken together, our results show that the protective functions of TREM2, both in inflammatory response and cognitive impairment as well as in the decrease of M1 phenotype microglia, are related to PI3K/AKT/FoxO3a signaling pathway in AD mice.

Triggering Receptor Expressed on Myeloid Cells-2 Protects Aged Mice Against Sepsis by Mitigating the IL-23/IL-17A Response.

Advancing age is an independent predictor of mortality in septic patients. Recent animal studies were unable to reflect this clinical pathophysiological process, largely hampering the development of new efficacious therapies. Triggering receptor expressed on myeloid cells-2 (TREM-2) is a novel immune regulator with multiple activities. However, very little is known about the regulatory role of TREM-2 in sepsis upon aging. Blood samples were collected from septic patients within 24 h after intensive care unit admission. The patients were preselected into two groups based on the age (age with ≥60 years old and age with <60 years old). Sepsis in aged mice was induced by cecal ligation and puncture. The expression of TREM-2 was evaluated in septic patients and aged septic mice. Aged macrophages overexpressing TREM-2 and green fluorescent protein (GFP) were administered to aged septic mice after cecal ligation and puncture. Survival rate was monitored, and bacterial load and inflammatory mediators levels were evaluated. In vivo IL-23 function was blocked using appropriate monoclonal antibodies. The expression levels of TREM-2 were downregulated in both aged septic patients and aged septic mice. The administration of TREM-2-overexpressing macrophages significantly prolonged survival and alleviated organ injury in the aged septic mice. The protective effect did not affect host bacterial burden, but markedly inhibited the host IL-17A response, as determined by a multiplex cytokine assay. Screening the expression of IL-17A-related activating factors revealed that the IL-23 level in TREM-2-overexpressing macrophages was significantly lower than that in GFP-expressing macrophages. Blocking IL-23 after the administration of GFP-expressing macrophages protected aged mice against sepsis. TREM-2 prolonged survival of aged mice from sepsis by finely modulating the IL-23/IL-17A immune pathway. These results provide previously unidentified mechanistic insight into immune regulation by TREM-2 and new therapeutic targets in sepsis upon aging.

Microglia TREM2: A Potential Role in the Mechanism of Action of Electroacupuncture in an Alzheimer's Disease Animal Model.

Alzheimer's disease (AD) is one of the most serious public health concerns facing the world. Its characteristic feature is neuroinflammation due to microglial activation. Electroacupuncture is one of the therapies employed to improve the condition of patients with AD, although its mechanism of action is still to be determined. Triggering receptor expressed on myeloid cells 2 (TREM2) is a microglia-specific receptor that is involved in regulating neuroinflammation in AD. In this study, we applied senescence-accelerated mouse-prone 8 mice as the AD animal model, used the Morris water maze, and applied hematoxylin and eosin staining, immunofluorescence double staining, and Western blotting, to explore the effects and potential mechanisms of action of electroacupuncture. In summary, this study suggested that electroacupuncture treatment could improve the learning and memory abilities (p < 0.05) and protect neurons. These effects result from acupuncture could upregulate TREM2 expression in the hippocampus (p < 0.01), which was essential for the anti-inflammatory effects in the AD animal model. However, further studies are needed to conclusively demonstrate the mechanism of action of electroacupuncture in AD.

IL-4 and IL-10 promotes phagocytic activity of microglia by up-regulation of TREM2

Triggering receptor expressed on myeloid cells-2 (TREM2) is an innate immune receptor that promotes phagocytosis by microglia. However, whether TREM2 is related to the stimulus-dependent phagocytic activity of microglia is unclear. In this study, the primary cultured microglia were stimulated with interferon (IFN)-γ, interleukin (IL)-4, and interleukin (IL)-10, respectively, and their phagocytic activity against microbeads and apoptotic neural stem cells (NSCs) was measured. TREM2 of microglia was detected by qPCR and western blotting. The TREM2 signal was blocked in microglia using the siRNA technique. The results showed that IL-4 or IL-10 treatment significantly increased the number of microglia gathered around the apoptotic neurosphere. IL-4 and IL-10 treatment also promoted phagocytosis of microbeads and apoptotic NSCs by primary cultured microglia. The TREM2 expression was up-regulated in IL-4- or IL-10- treated microglia. TREM2 siRNA treatment blocked the phagocytic activity of IL-4- or IL-10-treated microglia. In conclusion, these results indicated that IL-4 and IL-10 promote the phagocytic activity of microglia by the up-regulation of TREM2, which suggested a new potential therapeutic target for neurodegenerative disease.

Low-Dose Ionizing Radiation Modulates Microglia Phenotypes in the Models of Alzheimer's Disease.

Alzheimer’s disease (AD) is the most common type of dementia. AD involves major pathologies such as amyloid-β (Aβ) plaques and neurofibrillary tangles in the brain. During the progression of AD, microglia can be polarized from anti-inflammatory M2 to pro-inflammatory M1 phenotype. The activation of triggering receptor expressed on myeloid cells 2 (TREM2) may result in microglia phenotype switching from M1 to M2, which finally attenuated Aβ deposition and memory loss in AD. Low-dose ionizing radiation (LDIR) is known to ameliorate Aβ pathology and cognitive deficits in AD; however, the therapeutic mechanisms of LDIR against AD-related pathology have been little studied. First, we reconfirm that LDIR (two Gy per fraction for five times)-treated six-month 5XFAD mice exhibited (1) the reduction of Aβ deposition, as reflected by thioflavins S staining, and (2) the improvement of cognitive deficits, as revealed by Morris water maze test, compared to sham-exposed 5XFAD mice. To elucidate the mechanisms of LDIR-induced inhibition of Aβ accumulation and memory loss in AD, we examined whether LDIR regulates the microglial phenotype through the examination of levels of M1 and M2 cytokines in 5XFAD mice. In addition, we investigated the direct effects of LDIR on lipopolysaccharide (LPS)-induced production and secretion of M1/M2 cytokines in the BV-2 microglial cells. In the LPS- and LDIR-treated BV-2 cells, the M2 phenotypic marker CD206 was significantly increased, compared with LPS- and sham-treated BV-2 cells. Finally, the effect of LDIR on M2 polarization was confirmed by detection of increased expression of TREM2 in LPS-induced BV2 cells. These results suggest that LDIR directly induced phenotype switching from M1 to M2 in the brain with AD. Taken together, our results indicated that LDIR modulates LPS- and Aβ-induced neuroinflammation by promoting M2 polarization via TREM2 expression, and has beneficial effects in the AD-related pathology such as Aβ deposition and memory loss.

Triggering receptor expressed on myeloid cells 2 activation downregulates toll-like receptor 4 expression and ameliorates cognitive impairment in the Aβ1-42 -induced Alzheimer's disease mouse model.

Increasing evidence suggests that changes in the triggering receptor expressed on myeloid cells 2 (TREM2) is closely correlated with the pathological development of Alzheimer's disease (AD). However, the biological function and related role of this change remain poorly understood. Higher TREM2 expression has been reported in the brain of AD patients than in normal controls. Here, levels of TREM2 gene and protein levels were observed to be higher in both cortex and hippocampus of the Aβ1-42 -induced AD mice than in those of the wild type mice. Together with in vitro experimental data, we found that the anti-inflammatory role of TREM2 was, to some extent, limited and potentially counteracted by the hyperactive toll-like receptor 4 (TLR4) in the AD mice. In this context, Interleukin 4 (IL-4), as an agonist of TREM2, was administered to the AD mice to persistently activate TREM2. Interestingly, TREM2 activation in IL-4-treated AD mice led to an elevation in lysosomes and microtubule-associated protein 1 light chain 3 (LC3) II/I expression, demonstrating that the level of microglia autophagy was increased. Increased autophagy significantly downregulated the expression levels of caspase recruitment domain-containing protein 9 (CARD9) and TLR4, potentially weakening the CARD9-TLR4 pathway and suppressing the TLR4-mediated pro-inflammatory effect in IL-4-treated AD mice. Furthermore, data acquired from Morris water maze testing indicated that IL-4 administration could ameliorate cognitive impairment in the AD mice. In conclusion, the findings from in vitro and in vivo experiments suggest that TREM2 might represent a potential drug target to treat neuroinflammation in AD.

Echinacoside promotes the proliferation of human renal tubular epithelial cells by blocking the HBX/TREM2‑mediated NF‑κB signalling pathway.

Hepatitis B virus X (HBX) protein is required for the replication of HBV and plays a role in the progression of hepatitis in humans. However, the underlying function of HBX during HBV-induced chronic glomerulonephritis (HBV-GN) is unknown. Echinacoside (ECH) is a phenylethanoid glycoside from the Cistanche genus, which possesses strong antiapoptosis and neuroprotective activities. In the present study, the function of HBX and the relationship between HBX and ECH in human renal tubular epithelial cells (RTECs; HK-2 cell line) were explored. Reverse transcription-quantitative PCR and western blot analyses were used to quantify the mRNA and protein expression levels of HBX in HK-2 cells, respectively. The Cell Counting Kit-8 assay was performed to analyse cell proliferation. Flow cytometry analysis was used to determine the rate of apoptosis. HBX showed antiproliferative and proapoptotic effects in HK-2 cells and was positively associated with triggering receptor expressed on myeloid cells 2 (TREM2) expression. Furthermore, ECH disrupted the function of HBX in HK-2 cells, functioning as an HBX suppressor. Moreover, a specific NF-κB inhibitor, PDTC, was used to further examine the relationship between HBX and NF-κB. The results suggested that NF-κB was involved in the HBX/TREM2 signaling pathway and negatively regulated TREM2 expression in RTECs. The present study provided novel insights into the function of HBX, and also indicated the potential value of ECH as a therapeutic agent for HBV-GN.

TREM2 activation attenuates neuroinflammation and neuronal apoptosis via PI3K/Akt pathway after intracerebral hemorrhage in mice

BackgroundNeuroinflammation is an important host defense response to secondary brain injury after intracerebral hemorrhage (ICH). Triggering receptor expressed on myeloid cells 2 (TREM2) confers strong neuroprotective effects by attenuating neuroinflammation in experimental ischemic stroke. Recent studies suggest that apolipoprotein E (apoE) is a novel, high-affinity ligand of TREM2. This study aimed to investigate the effects of TREM2 activation on neuroinflammation and neuronal apoptosis in a mouse model of ICH.MethodsAdult male CD1 mice (n = 216) were subjected to intrastriatal injection of bacterial collagenase. The TREM2 ligand, apoE-mimetic peptide COG1410 was administered intranasally at 1 h after ICH induction. To elucidate the underlying mechanism, TREM2 small interfering RNA (siRNA) and the phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 were administered intracerebroventricularly prior to COG1410 treatment. Neurobehavioral tests, brain water content, immunofluorescence, western blotting, and Fluoro-Jade C- and terminal deoxynucleotidyl transferase dUTP nick end labeling staining were performed.ResultsEndogenous TREM2 expression was increased and peaked at 24 h after ICH. TREM2 was expressed on microglia, astrocytes, and neurons. COG1410 improved both short-term and long-term neurological functions, reduced brain edema, inhibited microglia/macrophage activation and neutrophil infiltration, and suppressed neuronal apoptotic cell death in perihematomal areas after ICH. Knockdown of endogenous TREM2 by TREM2 siRNA aggravated neurological deficits and decreased the expression of TREM2 in naïve and ICH mice. COG1410 was associated with upregulation of TREM2, PI3K, phosphorylated-Akt, and Bcl-2 and downregulation of TNF-α, IL-1β, and Bax after ICH. The neuroprotective effects of COG1410 were abolished by both TREM2 siRNA and PI3K inhibitor LY294002.ConclusionsOur finding demonstrated that TREM2 activation improved neurological functions and attenuated neuroinflammation and neuronal apoptosis after ICH, which was, at least in part, mediated by activation of PI3K/Akt signaling pathway. Therefore, activation of TREM2 may be a potential therapeutic strategy for the management of ICH patients.

TREM2 suppresses the proinflammatory response to facilitate PRRSV infection via PI3K/NF-κB signaling.

Triggering receptor expressed on myeloid cells 2 (TREM2) serves as an anti-inflammatory receptor, negatively regulating the innate immune response. TREM2 is mainly expressed on dendritic cells and macrophages, the target cells of porcine reproductive and respiratory syndrome virus (PRRSV). Thus, we investigated the potential role of TREM2 in PRRSV infection in porcine alveolar macrophages (PAMs). We found that there was an increased expression of TREM2 upon PRRSV infection in vitro. TREM2 silencing restrained the replication of PRRSV, whereas TREM2 overexpression facilitated viral replication. The cytoplasmic tail domain of TREM2 interacted with PRRSV Nsp2 to promote infection. TREM2 downregulation led to early activation of PI3K/NF-κB signaling, thus reinforcing the expression of proinflammatory cytokines and type I interferons. Due to the enhanced cytokine expression, a disintegrin and metalloproteinase 17 was activated to promote the cleavage of membrane CD163, which resulted in suppression of infection. Furthermore, exogenous soluble TREM2 (sTREM2)-mediated inhibition of PRRSV attachment might be attributed to its competitive binding to viral envelope proteins. In pigs, following PRRSV challenge in vivo, the expression of TREM2 in lungs and lymph nodes as well as the production of sTREM2 were significantly increased. These novel findings indicate that TREM2 plays a role in regulating PRRSV replication via the inflammatory response. Therefore, our work describes a novel antiviral mechanism against PRRSV infection and suggests that targeting TREM2 could be a new approach in the control of the PRRSV infection.

Surgery-induced cognitive dysfunction is alleviated through triggering receptor expressed on myeloid cells 2

Neuroinflammation plays a key role in perioperative neurocognitive disorders (PND). Increased evidences indicate that triggering receptor expressed on myeloid cells 2 (TREM2) can mitigate inflammatory response in the brain, and the aim of this study is to investigate whether TREM2 is involved in surgery-induced cognitive dysfunction in adult mice. We used adult C57BL/6 mice subjected to intramedullary fixation surgery, and found that surgery did not impair the motor ability of mice, but worsened the learning and memory function, and reduced the expression of TREM2. Meanwhile, up-regulated TREM2 expression in the brain of mice, induced by selective TREM2 agonist HSP60, significantly improved the learning and memory, alleviated the neuroinflammation, and decreased the neuronal cell apoptosis in mice. Meanwhile, TREM2-siRNA abolished the increased expression of TREM2 induced by HSP60, and reversed all the HSP60-induced beneficial effects. Therefore, our study indicated that up-regulation of TREM2 alleviated neuroinflammation and improved learning and memory function after surgery in mice.

Enhancing protective microglial activities with a dual function TREM2 antibody to the stalk region.

Triggering receptor expressed on myeloid cells 2 (TREM2) is essential for the transition of homeostatic microglia to a disease‐associated microglial state. To enhance TREM2 activity, we sought to selectively increase the full‐length protein on the cell surface via reducing its proteolytic shedding by A Disintegrin And Metalloproteinase (i.e., α‐secretase) 10/17. We screened a panel of monoclonal antibodies against TREM2, with the aim to selectively compete for α‐secretase‐mediated shedding. Monoclonal antibody 4D9, which has a stalk region epitope close to the cleavage site, demonstrated dual mechanisms of action by stabilizing TREM2 on the cell surface and reducing its shedding, and concomitantly activating phospho‐SYK signaling. 4D9 stimulated survival of macrophages and increased microglial uptake of myelin debris and amyloid β‐peptide in vitro. In vivo target engagement was demonstrated in cerebrospinal fluid, where nearly all soluble TREM2 was 4D9‐bound. Moreover, in a mouse model for Alzheimer's disease‐related pathology, 4D9 reduced amyloidogenesis, enhanced microglial TREM2 expression, and reduced a homeostatic marker, suggesting a protective function by driving microglia toward a disease‐associated state.

AGE-RELATED CHANGES TO MACROPHAGES AFFECT FRACTURE HEALING

Fracture healing follows a strict temporal sequence characterized by an initial inflammatory phase. Perturbation of the inflammatory phase may be responsible for the poorer fracture healing outcomes in older adults. Herein, we examine age-related changes to the macrophage during fracture healing. Macrophages regulate inflammation through pro-inflammatory (M1) and anti-inflammatory (M2) phenotypes. Anti-inflammatory activity is promoted via activation of triggering receptor expressed on myeloid cells 2 (TREM2). Tibia fractures were made in old (24 months) and young (3 months) mice. Immune cells from the fracture callus were analyzed via RNAseq and FACS, and fracture healing was evaluated histologically. Old mice demonstrated significantly delayed fracture healing compared to young (p<0.05). The quantity of infiltrating macrophages into the fracture callus was similar in young and old mice. However, 1222 genes were significantly differentially regulated (FDR<0.1) in callus macrophages from old mice compared to young, and old macrophages demonstrated a more pro-inflammatory phenotype. TREM2 expression was increased in macrophages after fracture in both groups but was significantly less in old mice compared to young via RNAseq and FACS (FDR<0.1, p<0.05). TREM2-/- mice demonstrated increased pro-inflammatory cytokine expression within the callus with resulting significant delays in fracture healing compared to age-matched controls (p<0.05). Inhibition of macrophage infiltration into the fracture callus significantly improved fracture healing in old mice compared to age-matched controls. Age-related changes to macrophages, including increased pro-inflammatory cytokine expression and dysregulated TREM2 expression, may explain fracture healing deficits observed in older adults. Therapeutically targeting macrophages may improve management of fractures in older adults.

Distinct Signaling Pathways Regulate TREM2 Phagocytic and NFκB Antagonistic Activities.

Several genetic variants of the Triggering Receptor Expressed on Myeloid Cells-2 (TREM2) have been shown to increase the risk of developing Alzheimer’s disease (AD) supporting a role of microglia and immune cells in the pathobiology of AD. We have employed an ectopic model of TREM2 and DAP12 expression in HEK293 cells to study selectively TREM2 dependent signaling and phagocytic functions and evaluated the effects of some of the TREM2 mutations associated with AD. We show that shedding of the TREM2 N-terminal domain does not affect the inhibition of NFκB activation induced by TREM2 while it completely blocks phagocytosis suggesting that TREM2 anti-inflammatory properties can be mediated by the TREM2 C-terminal fragment while the phagocytic activity requires the full-length receptor. In addition, we confirm in that model that apolipoprotein E (APOE) is a ligand for TREM2 and triggers TREM2 signaling. In particular, we show that APOE4 stimulates spleen tyrosine kinase (SYK) activation more potently than APOE2 in a TREM2 dependent manner. Interestingly, TREM2 appears to antagonize NFκB activation induced by phorbol ester but is unable to prevent TNFα induction of NFκB activation suggesting that TREM2 antagonizes inflammatory events triggered downstream of PKC. TREM2 mutations drastically impact TREM2 phagocytosis as well as its ability to antagonize NFκB activation and notably prevent the activation of the PI3K/AKT pathway observed with wild-type TREM2. Overall our data suggest that TREM2 dependent phagocytosis requires an activation of the SYK/PI3K/AKT/PLCγ pathways while the suppression of NFκB activation by TREM2 is independent of SYK, PI3K, and PLCγ activities. This model of ectopic TREM2-DAP12 co-expression appears suitable to study TREM2 signaling as several biological functions of TREM2 and TREM2 mutations that have been previously described in myeloid and microglial cells were also replicated in this model.

Curcumin inhibits LPS-induced neuroinflammation by promoting microglial M2 polarization via TREM2/ TLR4/ NF-κB pathways in BV2 cells

Microglia mediate multiple facets of neuroinflammation, which plays a double-edged role in various brain diseases via distinct microglial phenotypes (deleterious M1 and neuroprotective M2). Therefore, the inhibition of overactivated inflammatory M1 microglia by switching to the protective M2 phenotype appears to be a potential therapeutic strategy in neuroinflammatory disorders. Curcumin has been shown to exhibit anti-inflammatory and neuroprotective activities. The present study investigated the potential effects of curcumin on microglial M1/M2 polarization and elucidated the possible molecular mechanisms of action in vitro. In this study, the BV2 microglial cell line was pretreated with different curcumin concentrations in the presence or absence of lipopolysaccharide (LPS) to assess the anti-inflammatory efficacy of curcumin based on the morphological and inflammatory changes. The cytotoxicity of curcumin for BV2 cells was evaluated using the CCK-8 assay. Further, the effect of curcumin concentrations on LPS-induced BV2 cells was studied. The morphological changes were observed using an optical microscope and immunofluorescent staining. Nitric oxide (NO) expression was determined using the Griess reagent. The expression of cytokines and inflammatory mediators was also measured by ELISA, qRT-PCR, flow cytometry, and immunofluorescence. Western blot analysis was used to determine the levels of triggering receptor expressed on myeloid cells 2 (TREM2), toll-like receptor 4 (TLR4), nuclear factor-kappa B (NF-κB) p65, p-NF-κB p65, IκB, and p-IκB expression. Results showed that curcumin concentrations less than 10 μM did not induce any detectable cytotoxicity but decreased BV2 cell viability up to 20 μM. Curcumin inhibited LPS-induced microglial activation. Curcumin treatment switched the M1 pro-inflammatory phenotype to the M2 anti-inflammatory phenotype by decreasing the expression of M1 markers (i.e., iNOS, IL-1β, IL-6, and CD16/32) and elevating the expression of M2 markers (i.e., arginase 1, IL-4, IL-10, and CD206). Interestingly, curcumin attenuated the activation of TLR4/NF-κB pathways and the downregulation of TREM2 expression in LPS-activated BV2 cells. Collectively, these results suggest that curcumin significantly alleviates LPS-induced inflammation by regulating microglial (M1/M2) polarization by reducing the imbalance of TREM2 and TLR4 and balancing the downstream NF-κB activation.

Triggering Receptor Expressed on Myeloid Cells 2 (TREM2) Mutations: a Potential Common Cause of Alzheimer's Disease and Musculoskeletal Disorders

Alzheimer's disease (AD), a worldwide growing health issue, is a purely neurodegenerative form of age‐induced dementia. Interestingly, weight loss is frequently associated with disease severity and clinical progression and has been shown to begin at least ten years before the diagnosis of the disease. Bone frailty and sarcopenia have each also been linked to the onset of this disease, although very little is known about the pathogenesis of these conditions in subjects with AD. Indeed, although a common link between dementia and osteoporosis has been suggested, it has never been conclusively demonstrated. The prevalence of both conditions increase with aging, and some genetic conditions can lead to both AD and osteoporosis. Examples for the potential common cause of the 2 conditions are mutations of the Triggering Receptor Expressed on Myeloid cells 2 (TREM2) gene, primarily expressed in myeloid cells or microglia in brain and in osteoclasts in bone, which result in AD and dementia, as well as in bone fragility. In a recent Genome Wide Association Study, a single nucleotide polymorphism (SNP) encoding the R47H variant in TREM2 was linked with strong single allele genetic risk factors for AD, as well as with increased risk for Parkinson's disease, amyotrophic lateral sclerosis and frontotemporal dementia. In addition, recent studies showed that mice expressing the R47H TREM2 variant exhibit loss of TREM2 function and neuritic dystrophy. In a similar manner, another TREM2 mutation, the Y38C variant, was previously described in the pathogenesis of Nasu‐Hakola disease (also known as PLOSL) which is characterized by extensive white matter loss and frontotemporal‐like dementia and polycystic lipomembranous osteodysplasia. To investigate whether mutations in the TREM2 gene, previously associated with AD and frontotemporal dementia, also affect the skeleton, we analyzed the musculoskeletal phenotype of mice in which the Y38C and R47H variants of the TREM2 gene are ubiquitously expressed, generated by Drs. Lamb and Landreth at the Stark Neuroscience Research Institute, Indiana University School of Medicine. Our studies showed decreased bone mass in mice expressing the TREM2 mutants at 6–8 months of age. Interestingly, even though TREM2 expression was absent in skeletal muscle of wild type mice, mice expressing the R47H TREM2 variant exhibit decrease muscle mass. In summary, our results suggest a common cause of cognitive disorders and bone loss, and support an indirect role of TREM2 mutations on the loss of muscle mass, secondary to the altered bone remodeling.Support or Funding InformationNIH/NIAMS R01AR067210ICMH Collaborative Pilot Project AwardThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.