TREM-1 Expression Research Articles

Construction of Durvalumab/carbon nanotube/PEI/aptamer-siRNA chimera for the immunotherapy of hepatocellular carcinoma

Immunotherapy is the most promising treatment for hepatocellular carcinoma (HCC). However, the immunosuppressive microenvironment and necrosis limit its therapeutic effectiveness. Carbon nanotubes (CNTs) have good tissue permeability and can penetrate tumor necrosis area. Here we constructed a Durvalumab/CNT/PEI/aptamer-siRNA chimera (chimera/Durmab/CNT) nanoparticles for the immunotherapy of HCC. In vivo and in vitro experiments showed that aptamer-siRNA chimeras could specifically bind HCC cells and inhibit the triggering receptor expressed on myeloid cells-2 (Trem2) expression, but had no effect on Trem2 expression in normal liver and lung. Transmission electron microscope results showed that the CNT/PEI nanoparticles were 20–30 nm in diameter and 200–350 nm in length. Dense PEI attachment can be observed on CNTs. CNT/PEI nanoparticles could control the sustained release of Durvalumab for 48 h. In vitro experimental results showed that chimera/Durmab/CNT could increase the proportion of T cells and CD8 + T cells, and then promote the apoptosis of HepG2 cells, and the therapeutic effect was superior to aptamer/Durmab/CNT and Durmab/CNT. We constructed a tumor-bearing mouse model, and the results showed that chimera/Durmab/CNT significantly inhibited the growth of transplanted tumor, and the volume and proliferation was further reduced in the chimera/Durmab/CNT group compared with the aptamer/Durmab/CNT group. T cells and CD8 + T cells infiltration, and HCC cell apoptosis were significantly increased in the chimera/Durmab/CNT group. In conclusion, we constructed a Durvalumab/CNT/PEI/chimera, which can effectively treat HCC by activating anti-tumor immunity.

Circular RNA hsa_circ_0083756 promotes intervertebral disc degeneration by sponging miR-558 and regulating TREM1 expression.

ObjectivesIntervertebral disc degeneration (IVDD) is a leading cause of low back pain. Circular RNAs (circRNAs) have been demonstrated to exert vital functions in IVDD. However, the role and mechanism of hsa_circ_0083756 in the development of IVDD remain unclear.Materials and methodsRT‐qPCR was performed to detect expressions of hsa_circ_0083756, miR‐558 and TREM1 in nucleus pulposus (NP) tissues and cells. CCK8 assay, flow cytometry, TUNEL assay, RT‐qPCR and WB were used to clarify the roles of hsa_circ_0083756 in NP cells proliferation and extracellular matrix (ECM) formation. Bioinformatics analyses, dual‐luciferase reporter gene experiment, RNA immunoprecipitation (RIP) assay and FISH assay were performed to predict and verify the targeting relationship between hsa_circ_0083756 and miR‐558, as well as that between miR‐558 and TREM1. Ultimately, the effect of hsa_circ_0083756 on IVDD was tested through anterior disc‐puncture IVDD animal model in rats.Resultshsa_circ_0083756 was upregulated in degenerative NP tissues and cells. In vitro loss‐of‐function and gain‐of‐function studies suggested that hsa_circ_0083756 knockdown promoted, whereas hsa_circ_0083756 overexpression inhibited NP cells proliferation and ECM formation. Mechanistically, hsa_circ_0083756 acted as a sponge of miR‐558 and subsequently promoted the expression of TREM1. Furthermore, in vivo study indicated that silencing of hsa_circ_0083756 could alleviate IVDD in rats.Conclusionshsa_circ_0083756 promoted IVDD via targeting the miR‐558/TREM1 axis, and hsa_circ_0083756 may serve as a potential therapeutic target for the treatment of IVDD.

Tumor-Infiltrating Myeloid Cells Co-Express TREM1 and TREM2 and Elevated TREM-1 Associates With Disease Progression in Renal Cell Carcinoma

Myeloid-derived suppressor cells (MDSC) and tumor-associated macrophages (TAM) contribute to cancer-related inflammation and tumor progression. While several myeloid molecules have been ascribed a regulatory function in these processes, the triggering receptors expressed on myeloid cells (TREMs) have emerged as potent modulators of the innate immune response. While various TREMs amplify inflammation, others dampen it and are emerging as important players in modulating tumor progression—for instance, soluble TREM-1 (sTREM-1), which is detected during inflammation, associates with disease progression, while TREM-2 expression is associated with tumor-promoting macrophages. We hypothesized that TREM-1 and TREM-2 might be co-expressed on tumor-infiltrating myeloid cells and that elevated sTREM-1 associates with disease outcomes, thus representing a possibility for mutual modulation in cancer. Using the 4T1 breast cancer model, we found TREM-1 and TREM-2 expression on MDSC and TAM and that sTREM-1 was elevated in tumor-bearing mice in multiple models and correlated with tumor volume. While TREM-1 engagement enhanced TNF, a TREM-2 ligand was detected on MDSC and TAM, suggesting that both TREM could be functional in the tumor setting. Similarly, we detected TREM-1 and Trem2 expression in myeloid cells in the RENCA model of renal cell carcinoma (RCC). We confirmed these findings in human disease by demonstrating the expression of TREM-1 on tumor-infiltrating myeloid cells from patients with RCC and finding that sTREM-1 was increased in patients with RCC. Finally, The Cancer Genome Atlas analysis shows that TREM1 expression in tumors correlates with poor outcomes in RCC. Taken together, our data suggest that manipulation of the TREM-1/TREM-2 balance in tumors may be a novel means to modulate tumor-infiltrating myeloid cell phenotype and function.

Construction of Durvalumab/carbon nanotube/PEI/aptamer-siRNA chimera for the immunotherapy of hepatocellular carcinoma.

Immunotherapy is the most promising treatment for hepatocellular carcinoma (HCC). However, the immunosuppressive microenvironment and necrosis limit its therapeutic effectiveness. Carbon nanotubes (CNTs) have good tissue permeability and can penetrate tumor necrosis area. Here we constructed a Durvalumab/CNT/PEI/ aptamer-siRNA chimera (chimera/Durmab/CNT) nanoparticles for the immunotherapy of HCC. In vivo and in vitro experiments showed that aptamer-siRNA chimeras could specifically bind HCC cells and inhibit the triggering receptor expressed on myeloid cells-2 (Trem2) expression, but had no effect on Trem2 expression in normal liver and lung. Transmission electron microscope (TEM) results showed that the CNT/PEI nanoparticles were 20-30 nm in diameter and 200-350nm in length. Dense PEI attachment can be observed on CNTs. CNT/PEI nanoparticles could control the sustained release of Durvalumab for 48 hours. In vitro experimental results showed that chimera/Durmab/CNT could increase the proportion of T cells and CD8+T cells, and then promote the apoptosis of HepG2 cells, and the therapeutic effect was superior to aptamer/Durmab/CNT and Durmab/CNT. We constructed a tumor-bearing mouse model, and the results showed that chimera/Durmab/CNT significantly inhibited the growth of transplanted tumor, and the volume and proliferation was further reduced in the chimera/Durmab/CNT group compared with the aptamer/Durmab/CNT group. T cells and CD8+T cells infiltration, and HCC cell apoptosis were significantly increased in the chimera/Durmab/CNT group. In conclusion, we constructed a Durvalumab/CNT/PEI/chimera, which can effectively treat HCC by activating anti-tumor immunity.

TREM1 fosters an immunosuppressive tumor microenvironment in papillary thyroid cancer.

The immunosuppressive microenvironment is associated with poor prognosis in papillary thyroid cancer (PTC); however, the molecular mechanisms involved are unknown. Among the triggering receptors expressed on myeloid cell (TREM) family, we found that TREM1 expression in PTC was significantly higher than that in normal tissues. TREM1 overexpression was associated with BRAFV600E profiles and advanced tumor stages. Furthermore, TREM1 mRNA expression was negatively correlated with promoter methylation status. Specifically, hypomethylation of CpG site cg06196379 in the TREM1 promoter was related with poor patient disease-free survival (DFS) and a high PTC recurrence rate. Mechanistically, TREM1 was mainly expressed in malignant epithelial cells but not in macrophages in PTC by single-cell analysis. PTC tissues with high TREM1 levels had enhanced infiltration of regulatory T cells (Tregs) and decreased infiltration of CD8+ T cells. Our study confirms that hypomethylation-mediated overexpression of TREM1 in PTC cells promotes an immunosuppressive microenvironment by enhancing Treg infiltration. We recommend the future use of therapeutic strategy targeting TREM1 for the treatment of PTC.

POS-379 TO INVESTIGATE THE EFFECT OF HERBA LYSIMACHIAE IN THE TREATMENT OF OBSTRUCTIVE NEPHROPATHY BASED ON STAT-1/TREM-1 PATHWAY

Obstructive nephropathy is a major cause of chronic renal failure in children and adults.Clinical and animal experiments have found that renal damage still exists after obstruction is relieved, which is mainly related to irreversible fibrosis changes caused by persistent inflammatory infiltration, and there is no specific medicine for western medicine treatment. Herba lysimachiae can detoxify detumescence, diuresis tonglin, has a certain kidney protection. However, there is still a lack of systematic knowledge about the herba lysimachiae components and mechanism of s. chinensis in the treatment of obstructive nephropathy. Twenty SD rats were randomly divided into Sham operation group (Sham), unilateral ureteral obstruction operation group (UUO), Pennyswort intervention Sham operation group (Sham+Lys), herba lysimachiae intervention model group (UUO+ Lys).From the 1st day after surgery, the decoction of herba lysimachiae was given 24g/kg, once a day, for continuous 2w.Or intragastric administration of normal saline at the same time.Routine biochemical kit was used to detect renal function 2 weeks later.Renal tubule dilatation, mesangial hyperplasia and renal interstitial fibrosis were detected by HE, PAS and Masson staining.Immunohistochemistry and immunofluorescence were used to detect the activation of STAT-1/ TreM-1 pathway, macrophage aggregation (CD68+ iNOS+), renal tubular injury markers (KIM-1), and fibrosis related molecules (α-SMA, TGF-beta) in renal tissues. In the model group, the renal tubules were oppressed by urine retention on the obstructed side. Pathological staining showed that the epithelial cells of the renal tubules were necrotic to varying degrees, and the interstitial was infiltrated by acute and chronic inflammatory cells. Some of the necrotic renal tubules were gradually replaced by fibrous scar, and the formation of renal interstitial fibrosis was visible.Biochemical tests showed that the creatinine and renal injury scores were significantly higher than those of the normal group.Immunohistochemistry/immunofluorescence assay showed increased stat-1 /TREM-1 expression.Abnormal aggregation of M1-type macrophages in renal interstitium, accompanied by renal injury and fibrosis, showed increased expressions of KIM-1, α-SMA and TGF-beta (all P<0.05).In contrast, stat-1 /TREM-1 expression was decreased in renal tissue after herba lysimachiae intervention, and the above inflammatory and fibrosis related indicators were improved (P<0.05). Herba lysimachiae can protect renal function and inhibit renal inflammation and fibrosis in UUO rats, and this mechanism is related to the STAT-1/TREM-1 pathway.

DOP81 Baseline whole-blood gene expression of TREM1 does not predict clinical or endoscopic outcomes following adalimumab treatment in patients with Ulcerative Colitis or Crohn’s Disease in the SERENE studies

Abstract Background Conflicting evidence exists regarding whether low baseline Triggering Receptor Expressed on Myeloid cells (TREM)1 whole-blood gene expression predicts response or non-response to anti-tumour necrosis factor agents in patients with Ulcerative Colitis (UC) or Crohn’s Disease (CD). This study investigated whether baseline TREM1 expression predicted outcomes following adalimumab treatment in patients with UC or CD in the SERENE Phase 3 studies. Methods The SERENE studies (SERENE-UC, NCT02065622; and SERENE-CD, NCT02065570) compared a higher adalimumab induction dosing regimen versus the standard regimen in patients with UC or CD. Whole-blood TREM1 expression was analysed by RNA sequencing in patients who received standard-dose adalimumab and had clinical and endoscopic outcomes at Week 8 or 52 in SERENE-UC (n=95 for Week 8 and n=70 for Week 52 outcomes), or Week 12 or 56 in SERENE-CD (n=106 for all Week 12 outcomes; n=48 for clinical and n=50 for endoscopic outcomes at Week 56). Outcome definitions are summarised in Table 1. A 2-sample t-test was used to compare baseline TREM1 gene expression (log2 counts per million) in clinical or endoscopic remitters or responders versus non-remitters or non-responders at Weeks 8 and 52 (SERENE-UC) or Weeks 12 and 56 (SERENE-CD). Results In SERENE-UC, baseline TREM1 expression did not predict clinical remission at Week 8 (Figure 1a; p=0.7942) and was only weakly predictive of clinical remission at Week 52 (Figure 1b; p=0.04997). Further, it was not predictive of clinical response at Week 8 or Week 52, nor of endoscopic remission or endoscopic response (Figure 1c) at Week 8 (p&amp;gt;0.05 in all cases), although a weak correlation was observed with both endoscopic remission (p=0.0484) and endoscopic response (p=0.01162; Figure 1d) at Week 52. In SERENE-CD, baseline TREM1 expression was not linked to endoscopic or clinical outcomes at either Week 12 or Week 56 (p&amp;gt;0.05 in all cases; Figure 2a–d). Stratification by adalimumab quartiles did not increase the ability of baseline TREM1 expression to predict clinical outcomes in either study. Of note, a highly positive correlation was observed between baseline TREM1 expression and neutrophils in endoscopic responders and non-responders in both studies. Regardless of clinical or endoscopic response or non-response, TREM1 expression decreased over time following adalimumab treatment. All results obtained with standard-dose adalimumab were replicated with the higher dose (data not shown). Conclusion The findings of this study suggest that baseline whole-blood TREM1 gene expression is not a robust predictor of clinical or endoscopic outcomes following adalimumab treatment in patients with UC or CD.

Liraglutide Inhibits Osteoclastogenesis and Improves Bone Loss by Downregulating Trem2 in Female Type 1 Diabetic Mice: Findings From Transcriptomics.

BackgroundThe mechanisms of bone fragility in type 1 diabetes (T1D) are not fully understood. Whether glucagon-like peptide-1 receptor (GLP-1R) agonists could improve bone quality in T1D context also remains elusive.AimsWe aimed to explore the possible mechanisms of bone loss in T1D and clarify whether liraglutide has effects on bone quality of T1D mice using transcriptomics.MethodsFemale streptozotocin-induced diabetic C57BL/6J mice were randomly divided into four groups and received the following treatments daily for 8 weeks: saline as controls, insulin, liraglutide, and liraglutide combined with insulin. These groups were also compared with non-STZ-treated normal glucose tolerance (NGT) group. Trunk blood and bone tissues were collected for analysis. Three tibia from each of the NGT, saline-treated, and liraglutide-treated groups were randomly selected for transcriptomics.ResultsCompared with NGT mice, saline-treated T1D mice manifested markedly hyperglycemia and weight loss, and micro-CT revealed significantly lower bone mineral density (BMD) and deficient microarchitectures in tibias. Eight weeks of treatment with liraglutide alone or combined with insulin rescued the decreased BMD and partly corrected the compromised trabecular microarchitectures. Transcriptomics analysis showed there were 789 differentially expressed genes mainly mapped to osteoclastogenesis and inflammation pathways. The RT-qPCR verified that the gene expression of Trem2, Nfatc1, Trap, and Ctsk were significantly increased in the tibia of T1D compared with those in the NGT group. Liraglutide treatment alone or combined with insulin could effectively suppress osteoclastogenesis by downregulating the gene expression of Trem2, Nfatc1, Ctsk, and Trap.ConclusionsTaken together, increased osteoclastogenesis with upregulated expression of Trem2 played an important role in bone loss of T1D mice. Liraglutide provided protective effects on bone loss in T1D mice by suppressing osteoclastogenesis.

Bulk and Single-Cell Profiling of Breast Tumors Identifies TREM-1 as a Dominant Immune Suppressive Marker Associated With Poor Outcomes.

BackgroundTriggering receptor expressed on myeloid cells (TREM)-1 is a key mediator of innate immunity previously associated with the severity of inflammatory disorders, and more recently, the inferior survival of lung and liver cancer patients. Here, we investigated the prognostic impact and immunological correlates of TREM1 expression in breast tumors.MethodsBreast tumor microarray and RNAseq expression profiles (n=4,364 tumors) were analyzed for associations between gene expression, tumor immune subtypes, distant metastasis-free survival (DMFS) and clinical response to neoadjuvant chemotherapy (NAC). Single-cell (sc)RNAseq was performed using the 10X Genomics platform. Statistical associations were assessed by logistic regression, Cox regression, Kaplan-Meier analysis, Spearman correlation, Student’s t-test and Chi-square test.ResultsIn pre-treatment biopsies, TREM1 and known TREM-1 inducible cytokines (IL1B, IL8) were discovered by a statistical ranking procedure as top genes for which high expression was associated with reduced response to NAC, but only in the context of immunologically “hot” tumors otherwise associated with a high NAC response rate. In surgical specimens, TREM1 expression varied among tumor molecular subtypes, with highest expression in the more aggressive subtypes (Basal-like, HER2-E). High TREM1 significantly and reproducibly associated with inferior distant metastasis-free survival (DMFS), independent of conventional prognostic markers. Notably, the association between high TREM1 and inferior DMFS was most prominent in the subset of immunogenic tumors that exhibited the immunologically hot phenotype and otherwise associated with superior DMFS. Further observations from bulk and single-cell RNAseq analyses indicated that TREM1 expression was significantly enriched in polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) and M2-like macrophages, and correlated with downstream transcriptional targets of TREM-1 (IL8, IL-1B, IL6, MCP-1, SPP1, IL1RN, INHBA) which have been previously associated with pro-tumorigenic and immunosuppressive functions.ConclusionsTogether, these findings indicate that increased TREM1 expression is prognostic of inferior breast cancer outcomes and may contribute to myeloid-mediated breast cancer progression and immune suppression.

TREM2-induced activation of microglia contributes to synaptic resilience in non-demented individuals with Alzheimer's neuropathology.

The existence of individuals who remain cognitively intact despite presenting histopathological signs of Alzheimer's disease (AD), here referred to as "Non demented with AD neuropathology" (NDAN), suggests that some unknown mechanisms are triggered to resist cognitive impairment. Synaptic dysfunction has been identified as one of the major AD causes and it is established that microglia, attracted to plaques, engulf damaged synapses. In this context, exposed phosphatidylserine (ePS) represents a neuronal "eat-me" signal involved in microglial-mediated phagocytosis of damaged synapses. A possible candidate mediator of this process is represented by TREM2, a microglial surface receptor activated by ligands including PS. Based on TREM2 role in the scavenging function of microglia, we hypothesize that an efficient microglial phagocytosis of damaged synapses underlies synaptic resilience in NDAN, thus protecting from memory deficits. Using immunofluorescence microscopy, a comparative study of human post-mortem frontal cortices of aged-matched individuals, AD and NDAN individuals has been performed. The distribution of activated microglia (IBA1 and IBA1/CD68 positive cells) and the expression of phagocytic microglia-related proteins (TREM2 and DAP12) were evaluated. To test the efficacy of microglia in removing debris and damaged synapses, preservation of synapses around plaques was assessed using MAP2 and tubulin βIII as dendritic and axonal markers respectively, and PSD95 as a post synaptic marker. Furthermore, taking advantage of flow cytometry techniques, a study of the ePS using Annexin V assay has been performed. Immunofluorescence analyses indicate higher microglial activation and TREM2 expression in NDAN individuals, as well as preserved axonal and dendritic structure around plaques vs. AD. High levels of PSD95 around NDAN plaques and the colocalization of PSD95 and CD68 may suggest a prompt removal of damaged synapses by hyperactive phagocytic microglia. Annexin V assay on synaptosomes isolated from aged-matched, AD and NDAN individuals indicates changes in the ePS in NDAN individuals, confirming the engulfment of damaged synapses. Our results suggest a higher efficiency of TREM2-induced phagocytic microglia in removing damaged synapses, underlying synaptic resilience in NDAN individuals.

Pharmacological inhibition of carbonic anhydrases ameliorates cognitive dysfunction, rescuing Aβ‐induced neurovascular pathology in vivo

Cerebrovascular dysfunction (CVD) is an early feature of Alzheimer's disease (AD), contributing to the pathology progression, and suggesting a strict association between CVD and neurodegeneration. The majority of AD cases present cerebral amyloid angiopathy (CAA), neuropathological feature characterized by abnormal vasculotropic deposition of amyloid beta, mainly Aβ40. Severe CAA is also induced by familial Aβ variants, such as the Dutch-Q22. Our previous in vitro studies demonstrated that brain vascular amyloidosis elicits mitochondrial dysregulation and caspase-mediated apoptosis, in cells composing the neurovascular unit, including neurons, endothelial, glial and smooth muscle cells. Additionally, we showed that acetazolamide (ATZ) and methazolamide (MTZ), FDA-approved carbonic anhydrase inhibitors (CAIs) which cross the blood-brain barrier (BBB), hamper these detrimental processes. Carbonic anhydrases (CAs) represent a family of metalloenzymes catalyzing the reversible hydration of carbon dioxide, and their inhibition improves cerebral blood flow, vasoreactivity and neuronal excitability, pointing to CAs as CVD targets in AD.We employed Tg-SwDI mice (expressing human amyloid precursor protein, APP, carrying the Swedish, Dutch and Iowa mutations), which develop fibrillar amyloid burden, primarily in the cerebral microvasculature, starting at 6 months. We fed the animals (from 8 to 16 months of age) a CAI-diet, following which we performed behavioral analysis, and harvested the brains for both biochemical and immunohistochemical examination.Compared to untreated Tg mice, ATZ- and MTZ-fed animals showed reduced cognitive impairment, along with decreased vascular Aβ overload and astrogliosis, main pathological hallmarks of the disease. Tg animals exhibited Aβ deposition in endothelial and glial cells, leading to cell-specific caspase-3 activation. Interestingly, CAI-diet reduced endothelial and astrocytic Aβ deposits, and Aβ-induced caspase activation. Interestingly, the augmented expression of cerebral TREM2 and CD68 in CAI-treated mice suggests an attenuated inflammatory response, concomitantly with a boost of the cerebral clearance and active Aβ removal from brain microglia/macrophages, respectively, which may underlie the observed Aβ reduction. Furthermore, preliminary data suggest that both ATZ and MTZ promote microglial endocytosis, possibly facilitating Aβ degradation.CAIs provide neuroprotection, fostering neurovascular and glial physiological functions and clearance mechanisms, and attenuating CVD.

Impact of Galectin-Receptor Interactions on Liver Pathology During the Erythrocytic Stage of Plasmodium berghei Malaria.

Hepatopathy is frequently observed in patients with severe malaria but its pathogenesis remains unclear. Galectins are evolutionarily conserved glycan-binding proteins with pleiotropic roles in innate and adaptive immune responses, and exhibit pivotal roles during Plasmodium spp. infection. Here, we analyzed the impact of blockage of galectin-receptor interactions by treatment with alpha (α)-lactose on liver immunopathology during the erythrocytic stage of malaria in mice infected with Plasmodium berghei ANKA (PbANKA). Our results found that compared with PbANKA-infected mice (malarial mice), blockage of galectin-receptor interactions led to decreased host survival rate and increased peripheral blood parasitemia; exacerbated liver pathology, increased numbers of CD68+ macrophages and apoptotic cells, and increased parasite burden in the livers on days 5 and 7 post infection (p.i.) as well as increased mRNA expression levels of galectin-9 (Gal-9) and its receptor, the T cell immunoglobulin domain and mucin domain protein 3 (Tim-3), interferon (IFN)α, IFNγ, and the triggering receptor expressed on myeloid cells (TREM)-1 in the livers or spleens of PbANKA-infected mice on day 7 p.i. Observed by transmission electron microscopy, the peritoneal macrophages isolated from malarial mice with α-lactose treatment had more pseudopodia than those from malarial mice. Measured by using quantitative real-time reverse transcription-polymerase chain reaction assay, the mRNA expression levels of Gal-9, IFNα, IFNβ, IFNγ, and TREM-1 were increased in the peritoneal macrophages isolated from malarial mice with α-lactose treatment in comparison of those from malarial mice. Furthermore, significant positive correlations existed between the mRNA levels of Gal-9 and Tim-3/IFNγ/TREM-1 in both the livers and the peritoneal macrophages, and between Gal-9 and Tim-3/TREM-1 in the spleens of malarial mice; significant positive correlations existed between the mRNA levels of Gal-9 and IFNγ in the livers and between Gal-9 and IFNα in the peritoneal macrophages from malarial mice treated with α-lactose. Our data suggest a potential role of galectin-receptor interactions in limiting liver inflammatory response and parasite proliferation by down-regulating the expressions of IFNα, IFNγ, and TREM-1 during PbANKA infection.

Triggering Receptor Expressed on Myeloid Cells 2 Protects Dopaminergic Neurons by Promoting Autophagy in the Inflammatory Pathogenesis of Parkinson's Disease.

Parkinson’s disease is a neurodegenerative disorder with an inflammatory response as the core pathogenic mechanism. Previous human genetics findings support the view that the loss of TREM2 function will aggravate neurodegeneration, and TREM2 is one of the most highly expressed receptors in microglia. However, the role of TREM2 in the inflammatory mechanism of PD is not clear. In our study, it was found both in vivo and in vitro that the activation of microglia not only promoted the secretion of inflammatory factors but also decreased the level of TREM2 and inhibited the occurrence of autophagy. In contrast, an increase in the level of TREM2 decreased the expression of inflammatory factors and enhanced the level of autophagy through the p38 MAPK/mTOR pathway. Moreover, increased TREM2 expression significantly decreased the apoptosis of dopaminergic (DA) neurons and improved the motor ability of PD mice. In summary, TREM2 is an important link between the pathogenesis of PD and inflammation. Our study provides a new view for the mechanism of TREM2 in PD and reveals TREM2 as a potential therapeutic target for PD.

SLAMF7 and TREM1 Mediate Immunogenic Cell Death in Colorectal Cancer Cells: Focus on Microsatellite Stability.

The aim of this study was to identify the association between SLAMF7 and TREM1 and anti-PD-1 drugs, and to determine whether they are molecular targets or predictors of responses to immunotherapy through induction of immunogenic cell death. CRC cell lines over-expressing SLAMF7 and TREM1 were used to examine immunogenic and biological traits (e.g., proliferation and invasiveness) associated with factors related to anti-cancer immunity. In addition, multiplex immunofluorescence was used to examine immune cells in microsatellite instability-high (MSI-H) CRC and microsatellite stable (MSS) CRC. Proliferation rate and invasiveness of TREM1-over-expressing CRC cells were significantly greater than those of control cells (p<0.001 and 0.031, respectively), whereas SLAMF7-over-expressing CRC cells showed the opposite traits (p=0.005 and 0.002, respectively). SLAMF7-over-expressing DLD-1 cells harboring MSI-H showed increased apoptosis when treated with anti-PD-1 drugs, unlike SLAMF7-over-expressing SW480 cells harboring MSS. SLAMF7-over-expressing DLD1 and SW480 cells showed a marked increase in expression of the major cytokine mediator HMGB1 when exposed to anti-PD-1 drugs. Co-administration of anti-PD-1 drugs and TREM1 inhibitors induced apoptosis only in MSI-H HCT116 cells; HMGB1 was over-expressed regardless of microsatellite status. Expression of TREM1 and SLAMF7 is closely associated with immunogenic cell death, and TREM1 inhibitors may be an effective adjuvant that enhances anti-PD-1-mediated immunogenic cell death in MSS CRC.

Interdependence of neural network dysfunction and microglial alterations in Alzheimer’s disease-related models

SummaryNonconvulsive epileptiform activity and microglial alterations have been detected in people with Alzheimer’s disease (AD) and related mouse models. However, the relationship between these abnormalities remains to be elucidated. We suppressed epileptiform activity by treatment with the antiepileptic drug levetiracetam or by genetic ablation of tau and found that these interventions reversed or prevented aberrant microglial gene expression in brain tissues of aged human amyloid precursor protein transgenic mice, which simulate several key aspects of AD. The most robustly modulated genes included multiple factors previously implicated in AD pathogenesis, including TREM2, the hypofunction of which increases disease risk. Genetic reduction of TREM2 exacerbated epileptiform activity after mice were injected with kainate. We conclude that AD-related epileptiform activity markedly changes the molecular profile of microglia, inducing both maladaptive and adaptive alterations in their activities. Increased expression of TREM2 seems to support microglial activities that counteract this type of network dysfunction.

The mannose receptor (CD206) identifies a population of colonic macrophages in health and inflammatory bowel disease

To understand the contribution of mononuclear phagocytes (MNP), which include monocyte-derived intestinal macrophages, to the pathogenesis of inflammatory bowel disease (IBD), it is necessary to identify functionally-different MNP populations. We aimed to characterise intestinal macrophage populations in patients with IBD. We developed 12-parameter flow cytometry protocols to identify and human intestinal MNPs. We used these protocols to purify and characterize colonic macrophages from colonic tissue from patients with Crohn’s disease (CD), ulcerative colitis (UC), or non-inflamed controls, in a cross-sectional study. We identify macrophage populations (CD45+CD64+ HLA-DR+) and describe two distinct subsets, differentiated by their expression of the mannose receptor, CD206. CD206+ macrophages expressed markers consistent with a mature phenotype: high levels of CD68 and CD163, higher transcription of IL-10 and lower expression of TREM1. CD206− macrophages appear to be less mature, with features more similar to their monocytic precursors. We identified and purified macrophage populations from human colon. These appear to be derived from a monocytic precursor with high CCR2 and low CD206 expression. As these cells mature, they acquire expression of IL-10, CD206, CD63, and CD168. Targeting the newly recruited monocyte-derived cells may represent a fruitful avenue to ameliorate chronic inflammation in IBD.

Targeting TREM2 on tumor-associated macrophages enhances immunotherapy.

Converting checkpoint inhibitor (CPI)-resistant individuals to being responsive requires identifying suppressive mechanisms. We identify TREM2+ tumor-associated macrophages (TAMs) as being correlated with exhausted CD8+ tumor-infiltrating lymphocytes (TILs) in mouse syngeneic tumor models and human solid tumors of multiple histological types. Fc domain-enhanced anti-TREM2 monoclonal antibody (mAb) therapy promotes anti-tumor immunity by elimination and modulation of TAM populations, which leads to enhanced CD8+ TIL infiltration and effector function. TREM2+ TAMs are most enriched in individuals with ovarian cancer, where TREM2 expression corresponds to disease grade accompanied by worse recurrence-free survival. In an aggressive orthotopic ovarian cancer model, anti-TREM2 mAb therapy drives potent anti-tumor immunity. These results highlight TREM2 as a highly attractive target for immunotherapy modulation in individuals who are refractory to CPI therapy and likely have a TAM-rich tumor microenvironment.

Effects of microglial depletion and TREM2 deficiency on A\u03b2 plaque burden and neuritic plaque tau pathology in 5XFAD mice

Dystrophic neuronal processes harboring neuritic plaque (NP) tau pathology are found in association with Aβ plaques in Alzheimer’s disease (AD) brain. Microglia are also in proximity to these plaques and microglial gene variants are known risk factors in AD, including loss-of-function variants of TREM2. We have further investigated the role of Aβ plaque-associated microglia in 5XFAD mice in which NP tau pathology forms after intracerebral injection of AD brain-derived pathologic tau (AD-tau), focusing on the consequences of reduced TREM2 expression and microglial depletion after treatment with the colony-stimulating factor 1 (CSFR1) inhibitor, PLX3397. Young 5XFAD mice treated with PLX3397 had a large reduction of brain microglia, including cortical plaque-associated microglia, with a significant reduction of Aβ plaque burden in the cortex. A corresponding decrease in cortical APP-positive dystrophic processes and NP tau pathology were observed after intracerebral AD-tau injection in the PLX3397-treated 5XFAD mice. Consistent with prior reports, 5XFAD × TREM2−/− mice showed a significant reduction of plaque-associated microglial, whereas 5XFAD × TREM2+/− mice had significantly more plaque-associated microglia than 5XFAD × TREM2−/− mice. Nonetheless, AD-tau injected 5XFAD × TREM2+/− mice showed greatly increased AT8-positive NP tau relative to 5XFAD × TREM2+/+ mice. Expression profiling revealed that 5XFAD × TREM2+/− mice had a disease-associated microglial (DAM) gene expression profile in the brain that was generally intermediate between 5XFAD × TREM2+/+ and 5XFAD × TREM2−/− mice. Microarray analysis revealed significant differences in cortical and hippocampal gene expression between AD-tau injected 5XFAD × TREM2+/− and 5XFAD × TREM2−/− mice, including pathways linked to microglial function. These data suggest there is not a simple correlation between the extent of microglia plaque interaction and plaque-associated neuritic damage. Moreover, the differences in gene expression and microglial phenotype between TREM2+/− and TREM2−/− mice suggest that the former may better model the single copy TREM2 variants associated with AD risk.

TREM2 Regulates High Glucose-Induced Microglial Inflammation via the NLRP3 Signaling Pathway.

Background: TREM2 expressed on microglia plays an important role in modulating inflammation in neurodegenerative diseases. It remains unknown whether TREM2 modulates hyperglycemia-induced microglial inflammation. Methods: We investigated the molecular function of TREM2 in high glucose-induced microglial inflammation using western blotting, qPCR, ELISA, pulldown, and co-IP methods. Results: Our data showed that in high glucose-induced BV2 cells, TREM2 was increased, and the proinflammatory cytokine IL-1β was increased. TREM2 knockout (KO) attenuated the proinflammatory cytokine IL-1β; conversely, TREM2 overexpression (OE) exacerbated IL-1β expression. Furthermore, we found that high glucose promoted the interaction of TREM2 with NLRP3. TREM2 KO abolished the interaction of TREM2 with NLRP3, while TREM2 OE enhanced the interaction. Moreover, TREM2 KO reduced high glucose-induced NLRP3 inflammasome activation, and TREM2 OE augmented high glucose-induced NLRP3 inflammasome activation, indicating that high glucose enhances the expression of TREM2, which activates the NLRP3 inflammasome. To further clarify whether the NLRP3 signaling pathway mediates the TREM2-regulated inflammatory response, we blocked the NLRP3 inflammasome by knocking out NLRP3 and treating cells with a caspase1 inhibitor, which decreased the levels of the IL-1β proinflammatory cytokine but did not affect the high glucose-induced expression of TREM2. Conclusions: TREM2 modulates high glucose-induced microglial inflammation via the NLRP3 signaling pathway.

Abstract 64: Characterization of molecular and spatial diversity of macrophages in hepatocellular carcinoma

Abstract Hepatocellular carcinoma (HCC) has a dismal prognosis, and though checkpoint blocking antibodies have significantly improved patient outcome, many patients remain left out highlighting the need to identify additional immune target to enhance therapeutic immunity. Macrophages (MΦ) are an abundant and heterogeneous population in the tumor microenvironment (TME), and are associated with a poor prognosis in multiple tumor types, including HCC, however, their molecular and functional diversity is still poorly understood. We analyzed the molecular and spatial organization patterns of immune cells within the TME and adjacent tissue of 23 resected HCC lesions using single-cell RNA sequencing and multiplex immunohistochemistry (IHC). Strikingly, we found that Kupffer cells (KC), the self-renewing tissue-resident macrophages in liver tissue, are lacking from the TME, which is dominated by monocyte-derived macrophages. ScRNAseq followed by high-resolution clustering identified distinct MΦ molecular programs within the monocyte-derived macrophage compartment. One MΦ subset expressed a shared signature with monocytes including FCN1 and S100A8 in addition to T cell activation genes like CXCL9 and IL32. Conversely, one subset of MΦ expressed FOLR2, SEPP1 and genes of the complement (C1Qs), a program shared with KC in the adjacent uninvolved tissue, and include another intratumoral subset enriched for the expression of TREM2 and GPNMB. Guided by these results, we are developing an IHC antibody panel that allows to visualize distinct MΦ localization in the TME. Intratumoral MΦ interface with, and potentially regulate, the T cell compartment within the TME. We are analyzing HCC tumor lesions in our treatment-naïve cohort and in patients treated with neoadjuvant anti-PD-1 therapy (NCT03916627) to study direct interaction of MΦ and T cells using an innovative technology of physically-interacting cell sequencing (PICseq). This analysis enables us to identify MΦ with direct cell-cell contact with T cells, and our preliminary analysis demonstrates an enrichment in MΦ with a highly immunosuppressive phenotype. We are also using PICseq to map the molecular program of MΦ that are interacting with T cells and that we will corroborated with additional patients. Taken together, our data provide a new understanding of intratumoral MΦ diversity and highlight the presence of specific immunoregulatory MΦ programs unique to tumor lesions, with subsets of these MΦ found to be directly interacting with T cells, potentially modulating anti-tumor responsiveness. Our analysis of resected tumor from anti-PD-1 treated patients, will allow us to correlate MΦ programs, and direct T cell interaction, with clinical response, and will inform therapeutic trials targeting specific MΦ populations so as to improve clinical efficacy of cancer immunotherapy. Citation Format: Pauline Hamon, Assaf Magen, Merav Cohen, Alexandra Tabachnikova, Christie Chang, Mark Buckup, Leanna Troncoso, Zhen Zhao, Joel Kim, Amir Giladi, Nausicaa Malissen, Fiona Desland, Jessica Le Berichel, Ido Amit, Ephraim Kenigsberg, Myron Schwartz, Thomas Marron, Miriam Merad. Characterization of molecular and spatial diversity of macrophages in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 64.