TREM-like Transcript Research Articles

Relationship of proteins and subclinical cardiovascular traits in the population-based LIFE-Adult study

Background and aimsUnderstanding molecular processes of the early phase of atherosclerotic cardiovascular disease conditions is of utmost importance for early prediction and intervention measures. MethodsWe measured 92 cardiovascular-disease-related proteins (Olink, Cardiovascular III) in 2024 elderly participants of the population-based LIFE-Adult study. We analysed the impact of 27 covariables on these proteins including blood counts, cardiovascular risk factors and life-style-related parameters. We also analysed protein associations with 13 subclinical cardiovascular traits comprising carotid intima media thickness, plaque burden, three modes of Vicorder-based pulse-wave velocities, ankle-brachial index and ECLIA-based N-terminal prohormone of brain natriuretic peptide (NT-proBNP). ResultsEstimated glomerular filtration rate, triglycerides and sex where the most relevant covariables explaining more than 1 % variance of 49, 22 and 20 proteins, respectively. A total of 43 proteins were significantly associated with at least one of the analysed subclinical cardiovascular traits. NT-pro-BNP, brachial-ankle pulse-wave velocity (baPWV) and parameters of carotid plaque burden accounted for the largest number of associations. Association overlaps were relatively sparse. Only growth/differentiation factor 15, low density lipoprotein receptor and interleukin-1 receptor type 2 are associated with these three different cardiovascular traits. We confirmed several literature findings and found yet unreported associations for carotid plaque presence (von-Willebrand factor, galectin 4), carotid intima-media thickness (carboxypeptidase A1 andB1), baPWV (cathepsin D) and NT-proBNP (cathepsin Z, low density lipoprotein receptor, neurogenic locus homolog protein 3, trem-like transcript 2). Sex-interaction effects were observed, e.g. for spondin-1 and growth/differentiation factor 15 likely regulated by androgen response elements. ConclusionsWe extend the catalogue of proteome biomarkers possibly involved in early stages of cardiovascular disease pathologies providing targets for early risk prediction or intervention strategies.

A Phase 1/2, First-in-Human, Study to Investigate the Safety, Tolerability, Pharmacokinetics, Pharmacodynamics, and Efficacy of HMB-001 in Participants with Glanzmann Thrombasthenia

Background & Significance: Glanzmann thrombasthenia (GT) is a rare bleeding disorder resulting from a deficiency of integrin αIIbβ3 (also known as glycoprotein [GP] IIb/IIIa), a receptor for fibrinogen on platelets. Fibrinogen binding to αIIbβ3 bridges platelets and is a required step for normal platelet aggregation and hemostasis. GT is considered a severe bleeding disorder with approximately 50% of the patients reporting 1 bleed every day, and 13% reporting over 500 bleeds per year. The current standard of care for bleeding in patients with GT is reactive and on-demand, with no approved therapies for primary prophylaxis. HMB-001 is a bispecific antibody being developed by Hemab for prophylactic treatment to prevent and reduce bleeding events in patients with GT that can be administered subcutaneously. One arm of HMB-001 binds to and accumulates endogenous activated coagulation factor VII (FVIIa) while the second arm binds to the TREM-like transcript 1 receptor (TLT-1) on activated platelets. The combined effect of FVIIa accumulation and targeting to the surface of activated platelets via HMB-001 brings the activity of FVIIa to levels that are considered therapeutically effective based on clinical experience with recombinant FVIIa (rFVIIa). This is a first-in-human, Phase 1/2, dose escalation, safety, pharmacokinetic (PK), pharmacodynamic (PD), and preliminary efficacy study of HMB-001 in participants with Glanzmann thrombasthenia. Study Design and Methods: The study consists of 3 parts. Part A is a Phase 1, open-label, single ascending dose study, which will evaluate the safety, tolerability, PK, and PD of HMB-001 in participants with GT.Part B is a Phase 2, open-label, multiple ascending dose study evaluating the safety, tolerability, PK, PD, and preliminary effects on bleeding of repeat doses of HMB-001 monotherapy.Part C is a Phase 2, open-label, treatment extension portion study. It will be open to participants who have fulfilled the requirements of Part B and are considered eligible to continue by the Investigator. Part C will evaluate longer term safety, and preliminary efficacy of repeat doses of HMB-001 monotherapy for 9 months. Study Population: Male and female participants 18 to 65 years of age with GT. Part A is single-center, while Part B/C will be a multicenter study in various countries. Major inclusion and exclusion criteria include: Criteria for Inclusion: Part A and B Age 18 to 65Diagnosis of GTVital signs within normal rangeNegative pregnancy testMust meet the following baseline organ function eGFR >45 mL/min/1.73m 2LFTs within normal rangeHgb >85 g/L and platelet count >150 x 10 9/L Part B only 2 bleeding events per week on average of any severity and type and at least 1 spontaneous or traumatic bleed within the last 12 months requiring treatment with rFVIIa, platelets, medical or surgical procedure Criteria for Exclusion: Part A Active severe infection or inflammationHistory of clinically significant hypersensitivity associated with monoclonal antibody therapies.Personal or family history of venous or arterial thrombosis or thromboembolic disease.Other risk factors that substantially increase risk of venous or arterial thrombosisCongenital or acquired bleeding disorders other than GTConcurrent disease, treatment, medications, or abnormality in clinical laboratory tests that may pose additional riskAddiction or other diseases that prevent the participant from appropriately assessing the nature and scope of the clinical study or participating in study procedures Participants included in Part B are eligible for Part C following completion of their dosing in Part B.

Comprehensive analyses of circulating cardiometabolic proteins and objective measures of fat mass

BackgroundThe underlying molecular pathways for the effect of excess fat mass on cardiometabolic diseases is not well understood. Since body mass index is a suboptimal measure of body fat content, we investigated the relationship of fat mass measured by dual-energy X-ray absorptiometry with circulating cardiometabolic proteins.MethodsWe used data from a population-based cohort of 4950 Swedish women (55–85 years), divided into discovery and replication samples; 276 proteins were assessed with three Olink Proseek Multiplex panels. We used random forest to identify the most relevant biomarker candidates related to fat mass index (FMI), multivariable linear regression to further investigate the associations between FMI characteristics and circulating proteins adjusted for potential confounders, and principal component analysis (PCA) for the detection of common covariance patterns among the proteins.ResultsTotal FMI was associated with 66 proteins following adjustment for multiple testing in discovery and replication multivariable analyses. Five proteins not previously associated with body size were associated with either lower FMI (calsyntenin-2 (CLSTN2), kallikrein-10 (KLK10)), or higher FMI (scavenger receptor cysteine-rich domain-containing group B protein (SSC4D), trem-like transcript 2 protein (TLT-2), and interleukin-6 receptor subunit alpha (IL-6RA)). PCA provided an efficient summary of the main variation in FMI-related circulating proteins involved in glucose and lipid metabolism, appetite regulation, adipocyte differentiation, immune response and inflammation. Similar patterns were observed for regional fat mass measures.ConclusionsThis is the first large study showing associations between fat mass and circulating cardiometabolic proteins. Proteins not previously linked to body size are implicated in modulation of postsynaptic signals, inflammation, and carcinogenesis.

Novel biomarkers associated with thoracic aortic disease

BackgroundBiomarkers might help to improve diagnosis, surveillance and risk stratification of thoracic aortic disease (TAD). We explored the association between a broad spectrum of cardiovascular biomarkers with clinical characteristics and thoracic aortic diameter in TAD patients. MethodsVenous blood-samples were obtained in 158 clinically stable TAD patients visiting our outpatient clinic (2017–2020). TAD was defined as a thoracic aortic diameter ≥ 40 mm, or genetic confirmation (hereditary TAD). The cardiovascular panel III of the Olink multiplex platform was used for batch analysis of 92 proteins. A comparison was made between biomarker levels in patients with and without previous aortic dissection and/or surgery, and with and without hereditary TAD. Linear regression analyses were applied to identify (relative, normalized) biomarker concentrations associated with the absolute thoracic aortic diameter (ADmax), and thoracic aortic diameter indexed for body surface area (IDmax). ResultsMedian age of study patients was 61.0 (IQR 50.3–68.8) years, 37.3% females. Mean ADmax and IDmax were 43.3 ± 5.4 mm and 21.3 ± 3.3 mm/m2. After multivariable adjustment, Matrix Metalloproteinase-3 (MMP-3) and Insulin-like growth factor binding protein 2 (IGFBP-2) showed a significant positive association with ADmax and IDmax, respectively. Patients with previous aortic surgery/dissection had higher N-terminal-pro hormone BNP (NTproBNP) (median 3.67 [IQR 3.01–3.99] vs 2.84 [2.32–3.26], p ≤0.001). Patients with hereditary TAD had higher Trem-like transcript protein 2 (TLT-2) (median 4.64 [IQR 4.45–4.84]) than those with non-heriditary TAD (4.40 [4.17–4.64]; p = 0.00042). ConclusionsAmong a broad range of biomarkers, MMP-3 and IGFBP-2 were associated with disease severity in TAD patients. The pathophysiological pathways uncovered by these biomarkers, and their potential clinical use warrants further research.

Platelet-derived TLT-1 promotes tumor progression by suppressing CD8+ T cells.

Current understanding of tumor immunosuppressive mechanisms forms the basis for modern day immunotherapies. Immunoregulatory role of platelets in cancer remains largely elusive. Platelets from non-small cell lung cancer (NSCLC) patients revealed a distinct activation phenotype. TREM-like transcript 1 (TLT-1), a platelet protein, was increased along with enhanced extracellular release from NSCLC platelets. The increased platelet TLT-1 was also evident in humanized mice with patient-derived tumors. In immunocompetent mice with syngeneic tumors, TLT-1 binding to T cells, in vivo, led to suppression of CD8 T cells, promoting tumor growth. We identified direct interaction between TLT-1 and CD3ε on T cells, implicating the NF-κB pathway in CD8 T cell suppression. Anti-TLT-1 antibody rescued patients' T cells from platelet-induced suppression ex vivo and reduced tumors in mice in vivo. Clinically, higher TLT-1 correlated with reduced survival of NSCLC patients. Our findings thus identify TLT-1 as a platelet-derived immunosuppressor that suppresses CD8 T cells and demonstrate its therapeutic and prognostic significance in cancer.

Cytokine storm induced coagulopathy in septic shock and critical Covid-19: head-to-head comparison

BackgroundSeptic shock generates an important inflammatory reaction, endothelial activation and a procoagulant state leading to microvascular thrombosis and subsequent organ impairment [1].Similarly, a severe inflammatory reaction and a coagulopathy with pulmonary micro-thrombosis eventually leading to acute lung injury, is a typical feature of critical form of Coronavirus disease 2019 (Covid-19) [2].Our aim was to compare coagulation, platelet activation and platelets-neutrophils interplay between control, septic shock and critical Covid-19 patients.Methods/MaterialsA total of 118 patients were included in our prospective, monocentric, observational study between February 2019 and June 2020. Septic shock (n=48) and Covid-19 (n=22) patients were consecutively included at admission in our ICU department. Control patients (n=48) with matched gender and co-morbidities were recruited at central lab consultation.ResultsSeptic shock patients had worse severity scores due to multiple organ failure (assessed by APACHE II and SOFA score) whereas Covid-19 patients had more severe respiratory failure and a longer ICU length-of-stay (Table 1).At the time of inclusion, CRP and lymphocyte count were comparable between septic shock and Covid-19 patients. White cell count ad neutrophil count was higher for septic shock patients.Analysis of coagulation showed a prolonged INR, TT and aPTT in septic shock although only INR was prolonged in Covid-19. Thrombin antithrombin complex (TATc) formation was similar in both pathologies, whereas consumption of antithrombin III (ATIII) and D-dimers formation was more pronounced in septic shock.Platelet count was lower in septic shock and platelet activation, assessed via plasmatic levels of soluble P-selectin (sCD62P) and Trem-like transcript 1 (sTLT-1), was more important in septic shock.Neutrophil activation and NETosis, evaluated by levels of circulating myeloperoxidase (MPO) and citrullinated histone 3 (H3-Cit), was similarly increased in both groups (Figure 1).ConclusionsThis study confirmed an activation of coagulation cascade, platelet activation and NETosis in both septic shock and critical Covid-19, compared with control patients. Importantly, the extent of these changes was similar or less pronounced in critical COVID-19 compared with septic shock.Funding AcknowledgementType of funding sources: Foundation. Main funding source(s): Fondation Saint Luc Figure 1

DIA proteomics analysis through serum profiles reveals the significant proteins as candidate biomarkers in women with PCOS

BackgroundThe aim of this study was to apply proteomic methodology for the analysis of proteome changes in women with polycystic ovary syndrome (PCOS).Material and methodsAll the participators including 31 PCOS patients and 31 healthy female as controls were recruited, the clinical characteristics data was recorded at the time of recruitment, the laboratory biochemical data was detected. Then, a data-independent acquisition (DIA)-based proteomics method was performed to compare the serum protein changes between PCOS patients and controls. In addition, Western blotting was used to validate the expression of identified proteomic biomarkers.ResultsThere were 80 proteins differentially expressed between PCOS patients and controls significantly, including 54 downregulated and 26 upregulated proteins. Gene ontology and Kyoto Encyclopedia of Genes and Genomes analysis showed that downregulated proteins were enriched in platelet degranulation, cell adhesion, cell activation, blood coagulation, hemostasis, defense response and inflammatory response terms; upregulated proteins were enriched in cofactor catabolic process, hydrogen peroxide catabolic process, antioxidant activity, cellular oxidant detoxification, cellular detoxification, antibiotic catabolic process and hydrogen peroxide metabolic process. Receiver operating characteristic curves analysis showed that the area under curve of Histone H4 (H4), Histone H2A (H2A), Trem-like transcript 1 protein (TLT-1) were all over than 0.9, indicated promising diagnosis values of these proteins. Western blotting results proved that the detected significant proteins, including H4, H2A, TLT-1, Peroxiredoxin-1, Band 3 anion transport protein were all differently expressed in PCOS and control groups significantly.ConclusionThese proteomic biomarkers provided the potentiality to help us understand PCOS better, but future studies comparing systemic expression and exact role of these candidate biomarkers in PCOS are essential for confirmation of this hypothesis.

PROTEOMIC ASSESSMENT OF CIRCULATING MICROPARTICLES IN HYPERTENSION AND DIABETES

Objective: It is estimated that 415 million individuals are living with diabetes and an estimated 1.13 billion individuals worldwide have hypertension. Hypertension and diabetes are both associated with increased cardiovascular risk however the molecular changes to the vasculature associated these conditions are not fully understood. In the present study we examined the protein composition of circulating microparticles (MPs) in hypertensive, diabetic and healthy mice to identify common and disease-specific molecular changes. Design and method: Protein content was assessed in microparticles isolated from platelet-poor plasma of transgenic mice expressing human renin in the liver (TtRhRen, hypertension model), OVE26 type 1 diabetic mice and wild-type (WT) FVBN mice. Microparticles were isolated by differential centrifugation and protein content was analyzed using liquid chromatography mass spectrometry (LC-MS/MS). Results: We identified a total of 297 independent proteins with at least 2 peptides per protein. Of these, 163 proteins were found in all groups studied, 48 were exclusive to WT mice, 23 were exclusive to OVE26 mice and 4 were exclusive to TtRhRen mice. In addition, 22 proteins were observed with > 1.5 fold change (FC) compared to wild-type mice, and 68 proteins were reduced by > 50%. Amongst the top ten differentially expressed proteins, fibrinogen was upregulated in both OVE26 and TtRhRen mice compared with wild-type controls. Similarly Trem-like transcript 1, sarcoplasmic/ endoplasmic reticulum calcium ATPase 3 and junction plakoglobin were all downregulated in both OVE26 mice and TtRhRen mice. Conversely, arginase was up-regulated in diabetic, but not hypertensive mice while carboxypeptidase was up-regulated in hypertensive but not diabetic mice. Gene ontology (GO) analysis of 96 proteins unique or enriched in MPs from OVE26 mice indicated the presence of proteins involved in arginine biosynthesis and blood coagulation. GO analysis of TtRhRen-enriched proteins indicated presence of proteins associated with integrin signaling and inflammation. Conclusions: In summary, assessment of MP protein composition revealed common and disease-specific changes in diabetes and hypertension. Further analysis of these changes may lead to the identification of novel pathways associated with the pathogenesis of vascular injury in hypertension and diabetes.

Defining the TLT-1 interactome from resting and activated human platelets

The triggering receptor expressed on myeloid cells (TREM) protein family forms a class of type I transmembrane proteins expressed in immune cells that play important roles in innate and adaptive immune responses. The TREM family member TREM-like transcript 1 (TLT-1, also TREML1) is expressed in megakaryocytes and packaged into platelet granules. TLT-1 binds fibrinogen and plays a role in bleeding initiated by inflammatory insults. Here, we describe a proteomics screen that maps the TLT-1 interactome in resting and activated human platelets. Several identified TLT-1 interactors are involved in cell adhesion and migration, as well as platelet activation. Select interactors, including β3-integrin, RACK1, GRB2, and Rabs 5A, 7, and 11A, were additionally characterized in co-immunoprecipitation/immunoblotting experiments. Finally, several phosphorylation sites were found on immunoprecipitated TLT-1, including Thr280, a novel, regulated site on a conserved residue near the TLT-1 ITIM regulatory sequence. SignificancePlatelet function relies on the secretion of active molecules from intracellular vesicles, or granules, which contain soluble and membrane-bound proteins that are essential for platelet aggregation, coagulation reactions, and pathogen defense mechanisms. TLT-1 is sequestered in α-granules and transported to the plasma membrane, where it plays a unique role in hemostasis after inflammatory insults. Despite the known importance of TLT-1 in platelet biology, our knowledge of TLT-1 mechanistic signaling is limited. This study defines the TLT-1 interactome in resting and active human platelets, identifying several novel TLT-1 interactors, as well as TLT-1 phosphorylation sites, all with likely signaling implications in platelet aggregation dynamics.

Enhanced potency of recombinant factor VIIa with increased affinity to activated platelets

BackgroundRecombinant factor VIIa (rFVIIa) enhances thrombin generation in a platelet‐dependent manner; however, rFVIIa binds activated platelets with relatively low affinity. Triggering receptor expressed on myeloid cells (TREM)‐like transcript (TLT)‐1 is expressed exclusively on activated platelets. ObjectiveTo enhance the potency of rFVIIa via binding TLT‐1. MethodsRecombinant FVIIa was conjugated to a TLT‐1 binding Fab. In vitro potency of this platelet‐targeted rFVIIa (PT‐rFVIIa) was evaluated using factor X activation assays and by measuring viscoelastic changes in whole blood. In vivo potency was evaluated using a tail vein transection model in F8−/− mice expressing human TLT‐1. ResultsPT‐rFVIIa and rFVIIa had similar dissociation constant values for tissue factor binding and similar tissue factor‐dependent factor X activation. However, PT‐rFVIIa had increased catalytic efficiency on TLT‐1‐loaded vesicles and activated platelets. The in vitro potency in normal human blood with antibody‐induced hemophilia A was dependent on assay conditions used; with maximally activated platelets, the half maximal effective concentration for clot time for PT‐rFVIIa was 49‐fold lower compared with rFVIIa. In the murine bleeding model, a 53‐fold lower half maximal effective concentration was observed for blood loss for PT‐rFVIIa, supporting the relevance of the assay conditions with maximally activated platelets. In vitro analysis of blood from subjects with hemophilia A confirmed the data obtained with normal blood. ConclusionsIncreasing the affinity of rFVIIa to activated platelets resulted in approximately 50‐fold increased potency both in vitro and in the mouse model. The correlation of in vivo with in vitro data using maximally activated platelets supports that these assay conditions are relevant when evaluating platelet‐targeted hemostatic concepts.

Elevated level of circulatory sTLT1 induces inflammation through SYK/MEK/ERK signalling in coronary artery disease.

The role of inflammation in all phases of atherosclerotic process is well established and soluble TREM-like transcript 1 (sTLT1) is reported to be associated with chronic inflammation. Yet, no information is available about the involvement of sTLT1 in atherosclerotic cardiovascular disease. Present study was undertaken to determine the pathophysiological significance of sTLT1 in atherosclerosis by employing an observational study on human subjects (n=117) followed by experiments in human macrophages and atherosclerotic apolipoprotein E (apoE)-/- mice. Plasma level of sTLT1 was found to be significantly (P<0.05) higher in clinical (2342 ± 184 pg/ml) and subclinical cases (1773 ± 118 pg/ml) than healthy controls (461 ± 57 pg/ml). Moreover, statistical analyses further indicated that sTLT1 was not only associated with common risk factors for Coronary Artery Disease (CAD) in both clinical and subclinical groups but also strongly correlated with disease severity. Ex vivo studies on macrophages showed that sTLT1 interacts with Fcɣ receptor I (FcɣRI) to activate spleen tyrosine kinase (SYK)-mediated downstream MAP kinase signalling cascade to activate nuclear factor-κ B (NF-kB). Activation of NF-kB induces secretion of tumour necrosis factor-α (TNF-α) from macrophage cells that plays pivotal role in governing the persistence of chronic inflammation. Atherosclerotic apoE-/- mice also showed high levels of sTLT1 and TNF-α in nearly occluded aortic stage indicating the contribution of sTLT1 in inflammation. Our results clearly demonstrate that sTLT1 is clinically related to the risk factors of CAD. We also showed that binding of sTLT1 with macrophage membrane receptor, FcɣR1 initiates inflammatory signals in macrophages suggesting its critical role in thrombus development and atherosclerosis.

A Candidate Regulatory Variant at the TREM Gene Cluster Confer Alzheimer's Disease Risk by Modulating Both Amyloid-β Pathology and Neuronal Degeneration.

Background: rs9357347 located at the triggering receptor expressed on myeloid cells (TREM) gene cluster could increase TREM2 and TREM-like transcript 1 (TREML1) brain gene expression, which is considered to play a protective role against Alzheimer’s disease (AD).Objectives: To investigate the role of rs9357347 in AD pathogenesis by exploring the effects of rs9357347 on AD specific biomarkers.Methods: This study analyzed the association of rs9357347 with AD-related cerebrospinal fluid (CSF) and neuroimaging markers from 201 cognitively normal (CN) older adults, 349 elders with mild cognitive impairment (MCI), and 172 elders with AD dementia from the Alzheimer’s Disease Neuroimaging Initiative (ADNI). We next analyzed the association in 259 amyloid-β positive (Aβ+) elders and 117 amyloid-β negative (Aβ-) elders (Aβ+: CSF Aβ1-42 ≤ 192 pg/ml; Aβ-: CSF Aβ1-42 > 192 pg/ml). Associations were tested using multiple linear regression models at baseline. Furthermore, multiple mixed-effects models were used in a longitudinal study which lasted 4 years.Results: At baseline, we found that rs9357347 had association with CSF Aβ1-42 in CN group (β = 0.357, P = 0.009). In AD group, rs9357347 was associated with total tau (T-tau) level (β = -0.436, P = 0.007). Moreover, the strong influence exerted by rs9357347 on T-tau was also seen in Aβ+ group (β = -0.202, P = 0.036). In the longitudinal study, rs9357347 was also found to be associated with Aβ1-42 in CN group (β = 0.329, P = 0.023). In AD group, the mutation of rs9357347 was associated with slower accumulation of T-tau (β = -0.472, P = 0.002) and tau phosphorylated at threonine 181 [P-tau 181 (β = -0.330, P = 0.019)]. Furthermore, the obvious influence exerted by rs9357347 on T-tau was also seen in Aβ+ group (β = -0.241, P = 0.013).Conclusion: This study suggested that rs9357347 reduced the risk of AD by modulating both amyloid-β pathology and neuronal degeneration.

Downregulation of TREM-like transcript-1 and collagen receptor α2 subunit, two novel RUNX1-targets, contributes to platelet dysfunction in familial platelet disorder with predisposition to acute myelogenous leukemia.

Germline RUNX1 mutations lead to thrombocytopenia and platelet dysfunction in familial platelet disorder with predisposition to acute myelogenous leukemia (AML). Multiple aspects of platelet function are impaired in these patients, associated with altered expression of genes regulated by RUNX1. We aimed to identify RUNX1-targets involved in platelet function by combining transcriptome analysis of patient and shRUNX1-transduced megakaryocytes (MK). Down-regulated genes included TREM-like transcript (TLT)-1 (TREML1) and the integrin subunit alpha (α)-2 (ITGA2) of collagen receptor α2-beta (β)-1, which are involved in platelet aggregation and adhesion, respectively. RUNX1 binding to regions enriched for H3K27Ac marks was demonstrated for both genes using chromatin immunoprecipitation. Cloning of these regions upstream of the respective promoters in lentivirus allowing mCherry reporter expression showed that RUNX1 positively regulates TREML1 and ITGA2, and this regulation was abrogated after deletion of RUNX1 sites. TLT-1 content was reduced in patient MK and platelets. A blocking anti-TLT-1 antibody was able to block aggregation of normal but not patient platelets, whereas recombinant soluble TLT-1 potentiated fibrinogen binding to patient platelets, pointing to a role for TLT-1 deficiency in the platelet function defect. Low levels of α2 integrin subunit were demonstrated in patient platelets and MK, coupled with reduced platelet and MK adhesion to collagen, both under static and flow conditions. In conclusion, we show that gene expression profiling of RUNX1 knock-down or mutated MK provides a suitable approach to identify novel RUNX1 targets, among which downregulation of TREML1 and ITGA2 clearly contribute to the platelet phenotype of familial platelet disorder with predisposition to AML.

TREM-like transcript 1: a more sensitive marker of platelet activation than P-selectin in humans and mice

Key Points Platelet activation in vitro results in a more rapid and greater upregulation of TLT-1 surface expression compared with P-selectin. TLT-1 is more rapidly translocated to the surface of activated platelets than P-selectin during thrombus formation in vivo.

B7-H3 in tumors: friend or foe for tumor immunity?

B7-H3 is a type I transmembrane co-stimulatory molecule of the B7 family. B7-H3 mRNA is widely distributed in most tissues; however, B7-H3 protein is not constitutively expressed. Few molecules have been shown to mediate the regulation of B7-H3 expression, and their regulatory mechanisms remain unexplored. Recently, TREM-like transcript 2 (TLT-2) has been identified as a potential receptor of B7-H3. However, TLT-2 may not be the only receptor of B7-H3, as B7-H3 has many contradictory roles. As a co-stimulatory molecule, B7-H3 increases the proliferation of both CD4+ and CD8+ T-cells and enhances cytotoxic T-cell activity. However, greatly increased T-cell proliferation and IL-2 levels have been observed in the absence of B7-H3. Thus far, it has been shown that various tumors test positive for B7-H3 expression and that B7-H3 levels correlate with tumor growth, invasion, metastasis, malignant stage, and recurrence rate. Furthermore, transfection of cells with a B7-H3 plasmid and treatment with monoclonal antibodies to block B7-H3 are the main immunotherapeutic strategies for cancer treatment. Several groups have generated anti-B7-H3 antibodies and observed tumor growth suppression in vitro and in vivo. Therefore, it is likely that B7-H3 plays an important role in cancer diagnosis and treatment, aside from its role as a co-stimulatory molecule.

Triggering receptor expressed on myeloid cells 1 (TREM-1) and cytokine gene variants in complicated and uncomplicated malaria.

Malaria elicits inflammatory responses, which, if not well regulated, may exert detrimental effects. When activated, triggering receptor expressed on myeloid cells 1 (TREM-1) enhances inflammatory responses by increasing secretion of IL-8 and other Th1 cytokines. In contrast, TREM-like transcript 1 (TREML-1) promotes anti-inflammatory responses by binding to TREM-1 ligands and competing with TREM-1, thus antagonizing TREM-1 activation to reduce inflammation. Endothelial protein C receptor (EPCR) also mediates anti-inflammatory responses by activating endothelial protein C (PC). Upon microbial stimulation, soluble forms of TREM-1 (sTREM-1) and soluble EPCR (sEPCR) are released. Their plasma levels reflect the degree of inflammation and the severity of infection. In a cross-sectional study comparing patients with severe with uncomplicated malaria, sTREM-1, soluble TREML-1 (sTREML-1) and sEPCR plasma levels as well as plasma levels of sEPCR derived from convalescent patients were quantified. Samples were collected on admittance of paediatric patients infected with Plasmodium falciparum to hospitals in Accra, Ghana. Distinct genetic regions of the genes encoding TREM-1, EPCR, interleukin (IL)-8 and IL-18 encompassing known genetic polymorphisms that influence plasma levels underwent DNA sequencing. Higher sTREM-1 levels were observed among children suffering from severe malaria compared to those with uncomplicated malaria (P = 0.049). Low TREM-1 to TREML-1 ratios were associated with uncomplicated malaria (P = 0.033). The TREM1 rs2234237T variant causing the amino acid exchange Thr25Ser, which has been associated with higher TREM-1 plasma levels, was significantly more frequent among patients with severe malaria than in those with uncomplicated malaria (P = 0.036). Low levels of sEPCR were observed in severe and uncomplicated malaria, while variant genotypes of IL8, IL18 and EPCR did not show any association. Higher plasma levels of sTREM-1 alone or relative to sTREML-1 during malaria predispose to the phenotype of severe malaria. Carriage of the TREM1 rs2234237T allele appears to be a risk factor for the development of severe malaria.

Opposing roles of the triggering receptor expressed on myeloid cells 2 and triggering receptor expressed on myeloid cells-like transcript 2 in microglia activation

Mutations in triggering receptor expressed on myeloid cells 2 (TREM2), which has been proposed to regulate the inflammatory responses and the clearance of apoptotic neurons and/or amyloid-β, are genetically linked to increased risk for late-onset Alzheimer's disease (AD). Interestingly, a missense variant in TREM-like transcript 2 (TREML2), a structurally similar protein encoded by the same gene cluster with TREM2 on chromosome 6, has been shown to protect against AD. However, the molecular mechanisms by which TREM2 and TREML2 regulate the pathogenesis of AD, and their functional relationship, if any, remain unclear. Here, we show that lipopolysaccharide (LPS) stimulation significantly suppressed TREM2 but increased TREML2 expression in mouse brain. Consistent with this in vivo result, LPS or oligomeric amyloid-β treatment down regulated TREM2 but up-regulated TREML2 expression in primary microglia. Most important, modulation of TREM2 or TREML2 levels had opposing effects on inflammatory responses with enhancement or suppression of LPS-induced proinflammatory cytokine gene expression observed on TREM2 or TREML2 down regulation, respectively. In addition, the proliferation of primary microglia was significantly decreased when TREM2 was down regulated, whereas it was increased on TREML2 knockdown. Together, our results suggest that several microglial functions are strictly regulated by TREM2 and TREML2, whose dysfunctions likely contribute to AD pathogenesis by impairing brain innate immunity. Our findings provide novel mechanistic insights into the functions of TREM2 and TREML2 in microglia and have implications on designing new therapeutic strategies to treat AD.

TREM-like transcript 2 is stored in human neutrophil primary granules and is up-regulated in response to inflammatory mediators.

The triggering receptor expressed on myeloid cell locus encodes a family of receptors that is emerging as an important class of molecules involved in modulating the innate immune response and inflammation. Of the 4 conserved members, including triggering receptor expressed on myeloid cells 1 and 2 and triggering receptor expressed on myeloid cell-like transcripts 1 and 2, relatively little is known about triggering receptor expressed on myeloid cell-like transcript 2 expression and function, particularly in humans. In this study, experiments were performed to determine if triggering receptor expressed on myeloid cell-like transcript 2 expression is conserved between mouse and human, demonstrating that human triggering receptor expressed on myeloid cell-like transcript 2 is expressed on cells of the lymphoid, as well as myeloid/granuloid lineages, similar to murine triggering receptor expressed on myeloid cell-like transcript 2. Consistent with studies in the mouse, triggering receptor expressed on myeloid cell-like transcript 2 expression is up-regulated in response to inflammatory mediators on human neutrophils. Importantly, it was shown that triggering receptor expressed on myeloid cell-like transcript 2, in resting human neutrophils, is predominantly localized to intracellular vesicles, including secretory vesicles and primary granules; with the majority of triggering receptor expressed on myeloid cell-like transcript 2 stored in primary granules. In contrast to other primary granule proteins, triggering receptor expressed on myeloid cell-like transcript 2 is not expelled on neutrophil extracellular traps but is retained in the plasma membrane following primary granule exocytosis. In summary, these findings establish that triggering receptor expressed on myeloid cell-like transcript 2 expression is conserved between species and is likely to be important in regulating neutrophil antimicrobial function following primary granule exocytosis.

A New Triggering Receptor Expressed on Myeloid Cells (TREM) Family Member, TLT-6, is Involved in Activation and Proliferation of Macrophages.

The triggering receptor expressed on myeloid cells (TREM) family, which is abundantly expressed in myeloid lineage cells, plays a pivotal role in innate and adaptive immune response. In this study, we aimed to identify a novel receptor expressed on hematopoietic stem cells (HSCs) by using in silico bioinformatics and to characterize the identified receptor. We thus found the TREM-like transcript (TLT)-6, a new member of TREM family. TLT-6 has a single immunoglobulin domain in the extracellular region and a long cytoplasmic region containing 2 immunoreceptor tyrosine-based inhibitory motif-like domains. TLT-6 transcript was expressed in HSCs, monocytes and macrophages. TLT-6 protein was up-regulated on the surface of bone marrow-derived and peritoneal macrophages by lipopolysaccharide stimulation. TLT-6 exerted anti-proliferative effects in macrophages. Our results demonstrate that TLT-6 may regulate the activation and proliferation of macrophages.

Levels of soluble TREM-like transcript 1 in patients presenting to the emergency department with chest pain.

Recent studies suggest that the soluble triggering receptor expressed on myeloid cells-like transcript 1 (sTLT-1) facilitate atherothrombosis. Therefore, we evaluated sTLT-1 as a functional measure of atherothrombosis in acute coronary syndrome (ACS). Levels of sTLT-1 were determined by enzyme-linked immunosorbent assay on plasma from patients with potential ACS and compared with an age-matched control group with similar risk factors for cardiovascular disease. Of 53 patients enrolled, 19 patients were undergoing ACS (15 unstable angina, 2 non-ST-segment elevated myocardial infarction, and 2 ST-segment elevated myocardial infarction), 5 patients were found with noncardiac chest pain, and 29 were in the control group. The mean plasma sTLT-1 values in the ACS group were 4.644 ng/mL ± 1.277 standard error of the mean (SEM), in the noncardiac chest pain group were 0.708 ng/mL ± 0.427 SEM, and in the control group were 1.007 ng/mL ± 0.098 SEM. A statistically significant difference exists between patients experiencing cardiogenic chest pain versus controls (P < .05), suggesting sTLT-1 as a potential tool for understanding atherothrombosis in ACS.