TREM-1 (triggering receptor expressed on myeloid cells-1) is an orphan immunoreceptor expressed on monocytes, macrophages, and neutrophils. TREM-1 associates with and signals via the adapter protein DAP12/TYROBP, which contains an ITAM. TREM-1 activation by receptor cross-linking has been shown to be proinflammatory and to amplify some cellular responses to TLR ligands such as bacterial LPS. To investigate the cellular consequences of TREM-1 activation, we have characterized global gene expression changes in human monocytes in response to TREM-1 cross-linking in comparison to and combined with LPS. Both TREM-1 activation and LPS up-regulate chemokines, cytokines, matrix metalloproteases, and PTGS/COX2, consistent with a core inflammatory response. However, other immunomodulatory factors are selectively induced, including SPP1 and CSF1 (i.e., M-CSF) by TREM-1 activation and IL-23 and CSF3 (i.e., G-CSF) by LPS. Additionally, cross-talk between TREM-1 activation and LPS occurs on multiple levels. Although synergy in GM-CSF protein production is reflected in commensurate mRNA abundance, comparable synergy in IL-1β protein production is not. TREM-1 activation also attenuates the induction of some LPS target genes, including those that encode IL-12 cytokine family subunits. Where tested, positive TREM-1 outputs are greatly reduced by the PI3K inhibitor wortmannin, whereas this attenuation is largely PI3K independent. These experiments provide a detailed analysis of the cellular consequences of TREM-1 activation and highlight the complexity in signal integration between ITAM- and TLR-mediated signaling.

␣ 4 ␤ 1 integrin (VLA-4) appears to be unique among the leukocyte integrins in that it can initiate the adhesion of circulating lymphocytes without cellular activation. It is not known how lymphocytes or other cell types maintain constitutive levels of ␣ 4 ␤ 1 integrin activity. The current report describes a monoclonal antibody, 15/7, that recognizes a high affinity or ligand-occupied conformation of ␤ 1 integrin. Studies with 15/7 revealed that ␣ 4 ␤ 1 integrin-dependent adhesion of leukocytic cell lines is mediated by a population of low affinity receptors that is conformationally responsive to ligand; the 15/7 epitope could be induced by nanomolar concentrations of soluble VCAM-1 or by micromolar concentrations of a peptide derived from the type III connecting segment domain of fibronectin (as ligands for ␣ 4 ␤ 1 integrin). The same receptors were also responsive to adhesion activating reagents, such as Mn 2؉ , activating anti-␤ 1 integrin antibodies, and phorbol myristate acetate, which induced the 15/7 epitope directly and/or decreased the concentration of ligand required for epitope induction. In addition to the responsive receptor pool, cells expressed a second population of ␣ 4 ␤ 1 integrin that was conformationally restrained, failing to respond to ligand or to any of the activating reagents. The relative size of the responsive and inactive receptor pools, as well as the affinity of the responsive receptors, represented a stable phenotype of different cell types and played important roles in defining the cells' adhesive capacity and ligand specificity. Similar receptor populations were measured on lymphocyte subsets in whole blood. These studies provide insight into how cells maintain different constitutive levels of ␣ 4 ␤ 1 integrin activity, and how the activity of ␤ 1 integrin can be modulated by activators of cell adhesion.Integrins are heterodimeric adhesion molecules that contribute to the specificity of cellular interactions through the recognition of numerous matrix and cell-associated ligands (1). Importantly, the ligand binding activity of integrins can be modulated rapidly, allowing cells to specify the timing and location of integrin-mediated adhesive interactions. On circulating leukocytes, for example, the ␤ 2 integrins LFA-1 and Mac-1 are thought to be activated by site-specific factors (cytokines or other adhesive interactions) during transient cell interactions with the vascular wall; the receptors then establish firm adhesive contacts and mediate leukocyte extravasation (2, 3). Likewise, platelets are stimulated at sites of vascular injury allowing ␣II b ␤ 3 integrin to bind to fibrinogen and initiate thrombosis (4).The regulation of ␣ 4 integrin activity on circulating immune cells appears to be different from that of the other leukocyte integrins described above. ␣ 4 ␤ 1 and ␣ 4 ␤ 7 integrin can mediate leukocyte adhesion to their endothelial ligands VCAM-1 (5) and MAdCAM-1 (6), respectively, without cellular activation, and can do so even in the presence of the shear forces encount...

Key Points• High anti-ADAMTS13 antibody and low ADAMTS13 antigen levels adversely affect outcome in immunemediated TTP with greater mortality seen.• A raised troponin at presentation confers a sixfold increase and reduced GCS a nine-fold increase in mortality in acute TTP.Immune-mediated thrombotic thrombocytopenic purpura (TTP) is a life-threatening disorder caused by antibodies against ADAMTS13. From the United Kingdom TTP registry, we undertook a prospective study investigating the impact of the presenting anti-ADAMTS13 IgG antibody and ADAMTS13 antigen on mortality. A total of 312 episodes involving 292 patients over 87 months were included; 68% were female, median age 46 (range, 11-88 years), and median presenting ADAMTS13 of <5% (range, <5%-18%). The mortality rate was 10.3% (n 5 32); 68% of patients had a raised troponin at presentation conferring a sixfold increase in mortality compared with those with normal troponin levels (12.1% vs 2.0%, P 5 .04). Twenty-four percent had a reduced Glasgow Coma Score (GCS) at presentation with a ninefold increase in mortality (20% vs 2.2% for normal GCS at presentation, P < .0001). Mortality increased with higher anti-ADAMTS13 antibody levels and lower ADAMTS13 antigen levels. Those with antibody levels in the upper quartile (antibody >77%) had a mortality of 16.9% compared with 5.0% for the lowest quartile (antibody <20%) (P 5 .004). Those with an antigen level in the lowest quartile (antigen <1.5%) had a mortality of 18% compared with 3.8% for the highest quartile (antigen >11%) (P 5 .005). The synergistic effect of anti-ADAMTS13 IgG antibody in the upper quartile and ADAMTS13 antigen in the lowest quartile had the highest mortality of 27.3%. We conclude that both anti-ADAMTS13 IgG antibody and ADAMTS13 antigen levels correlate with outcome in TTP with increased cardiac and neurological involvement and increased mortality. (Blood. 2017;130(4):466-471)

BackgroundAsthma exacerbations remain a major unmet clinical need. The difficulty in obtaining airway tissue and bronchoalveolar lavage samples during exacerbations has greatly hampered study of naturally occurring exacerbations. This study was conducted to determine if mRNA profiling of peripheral blood mononuclear cells (PBMCs) could provide information on the systemic molecular pathways involved during asthma exacerbations.Methodology/Principal FindingsOver the course of one year, gene expression levels during stable asthma, exacerbation, and two weeks after an exacerbation were compared using oligonucleotide arrays. For each of 118 subjects who experienced at least one asthma exacerbation, the gene expression patterns in a sample of peripheral blood mononuclear cells collected during an exacerbation episode were compared to patterns observed in multiple samples from the same subject collected during quiescent asthma. Analysis of covariance identified genes whose levels of expression changed during exacerbations and returned to quiescent levels by two weeks. Heterogeneity among visits in expression profiles was examined using K-means clustering. Three distinct exacerbation-associated gene expression signatures were identified. One signature indicated that, even among patients without symptoms of respiratory infection, genes of innate immunity were activated. Antigen-independent T cell activation mediated by IL15 was also indicated by this signature. A second signature revealed strong evidence of lymphocyte activation through antigen receptors and subsequent downstream events of adaptive immunity. The number of genes identified in the third signature was too few to draw conclusions on the mechanisms driving those exacerbations.Conclusions/SignificanceThis study has shown that analysis of PBMCs reveals systemic changes accompanying asthma exacerbation and has laid the foundation for future comparative studies using PBMCs.

SummaryThrombotic microangiopathies (TMAs) are frequently difficult to differentiate clinically, and measurement of ADAMTS13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13) remains vital in thrombotic thrombocytopenic purpura (TTP) diagnosis. We retrospectively reviewed cases referred for ADAMTS13 testing, using UK TTP Registry screening data. Of a total 810 cases, 350 were confirmed as TTP. The 460 non-TTP cases comprised secondary TMAs (24Á57%) and haemolytic uraemic syndrome (HUS) (27Á17% aHUS, 2Á83% Shiga-like toxin-producing E. coli [STEC]-HUS); the remainder were TMAs with no clear association, not TMAs, or had no confirmed diagnosis. ADAMTS13 levels were significantly lower in TTP than STEC-HUS, aHUS and other TMAs. TTP patients had significantly lower platelet count (15 9 10 9 /l; range 0-96) than aHUS (57 9 10 9 /l; range 13-145, P < 0Á0001) or STEC-HUS (35 9 10 9 /l; range 14-106, P < 0Á0001); they also had lower creatinine levels (92 lmol/l; range 43-374) than aHUS (255 lmol/l; range 23-941, P < 0Á0001) and STEC-HUS (324 lmol/l; range 117-639, P < 0Á0001). However, 12/34 (35Á3%) aHUS patients had a platelet count <30 9 10 9 /l and 26/150 (17Á3%) of TTP patients had a platelet count >30 9 10 9 /l; 23/ 150 (15Á3%) of TTP patients had a creatinine level >150 lmol/l. This study highlights the wide variety of TMA presentations, and confirms the utility of ADAMTS13 testing in TTP diagnosis.