In the epithelium of the lower airways, a cell type of unknown function has been termed "brush cell" because of a distinctive ultrastructural feature, an apical tuft of microvilli. Morphologically similar cells in the nose have been identified as solitary chemosensory cells responding to taste stimuli and triggering trigeminal reflexes. Here we show that brush cells of the mouse trachea express the receptors (Tas2R105, Tas2R108), the downstream signaling molecules (α-gustducin, phospholipase C β2 ) of bitter taste transduction, the synthesis and packaging machinery for acetylcholine, and are addressed by vagal sensory nerve fibers carrying nicotinic acetylcholine receptors. Tracheal application of an nAChR agonist caused a reduction in breathing frequency. Similarly, cycloheximide, a Tas2R108 agonist, evoked a drop in respiratory rate, being sensitive to nicotinic receptor blockade and epithelium removal. This identifies brush cells as cholinergic sensors of the chemical composition of the lower airway luminal microenvironment that are directly linked to the regulation of respiration.airway sensory innervation | respiratory epithelium | jugular-nodose ganglion | bitter-tasting substances

Leukocyte migration to sites of inflammation is regulated by several endothelial adhesion molecules. Vascular adhesion protein-1 (VAP-1) is unique among the homing-associated molecules as it is both an enzyme that oxidizes primary amines and an adhesin. Although granulocytes can bind to endothelium via a VAP-1-dependent manner, the counter-receptor(s) on this leukocyte population is(are) not known. Here we used a phage display approach and identified Siglec-9 as a candidate ligand on granulocytes. The binding between Siglec-9 and VAP-1 was confirmed by in vitro and ex vivo adhesion assays. The interaction sites between VAP-1 and Siglec-9 were identified by molecular modeling and confirmed by further binding assays with mutated proteins. Although the binding takes place in the enzymatic groove of VAP-1, it is only partially dependent on the enzymatic activity of VAP-1. In positron emission tomography, the 68 Gallium-labeled peptide of Siglec-9 specifically detected VAP-1 in vasculature at sites of inflammation and cancer. Thus, the peptide binding to the enzymatic groove of VAP-1 can be used for imaging conditions, such as inflammation and cancer. (Blood. 2011;118(13):3725-3733) IntroductionLeukocyte migration from the blood into the nonlymphoid tissues is a hallmark of inflammation. Several molecules on the endothelial cell surface and their counter-receptors on leukocytes mediate a multistep adhesion cascade featuring tethering, rolling, activation, adhesion, crawling, and transmigration phases. 1,2 Vascular adhesion protein-1 (VAP-1/AOC3) is an endothelial cell molecule that is rapidly translocated from the intracellular storage granules to the endothelial cell surface on inflammation. It contributes to several steps in the extravasation cascade and controls trafficking of lymphocytes, granulocytes, and monocytes to sites of inflammation. VAP-1 has unique features distinct from other conventional adhesion molecules because, besides being an adhesin, it is also an enzyme. It catalyzes oxidative deamination of primary amines and produces hydrogen peroxide, aldehyde, and ammonium. 3 The end products of the enzymatic activity are highly potent inflammatory mediators and can up-regulate other adhesion molecules, such as E-and P-selectin, ICAM-1, and VCAM-1. 4,5 We recently found the first lymphocyte ligand for VAP-1, Siglec-10. 6 It is expressed on B cells, monocytes, and eosinophils but is absent from granulocytes. 7 However, VAP-1 is also involved in granulocyte migration to sites of inflammation. This has been demonstrated in studies with acute inflammation models (peritonitis, lung, and air pouch inflammation) in mouse. In these studies, significant reduction in granulocyte migration to sites of inflammation was obtained with a function blocking anti-VAP-1 antibody and a small molecular inhibitor against VAP-1. [8][9][10] Contribution of VAP-1 both at the rolling and transmigration steps during leukocyte extravasation has been demonstrated, and the enzymatic activity of VAP-1 seems to be important in these proc...

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Neuroimmune interactions play a critical role in the pathogenesis of asthma. Symptoms like wheezing and cough have been attributed to neural dysregulation, whereas sensitization and the induction of allergic inflammation have been linked with the activity of dendritic cells. Neuropeptides were previously shown to control dendritic cell function in vitro, suggesting interactions between dendritic cells and sensory nerves. Here we characterized the anatomical basis of the interactions between dendritic cells and nerves in the airways of mice and monitored the changes during allergic inflammation. Airway microdissection, whole-mount immunohistology, and confocal microscopy were used for the three-dimensional quantitative mapping of airway nerves and dendritic cells along the main axial pathway of nonsensitized versus ovalbumin-sensitized and -challenged CD11c-enhanced yellow fluorescent protein (CD11c-EYFP) transgenic mice. CD11c-EYFP-positive airway mucosal dendritic cells were contacted by calcitonin gene-related peptide-immunoreactive sensory fibers and their co-localization increased in allergic inflammation. Moreover, protein gene product 9.5-positive neuroepithelial bodies and airway ganglia were associated with dendritic cells. In human airways, human leukocyte antigen DR-positive mucosal dendritic cells were found in the close proximity of sensory nerves and neuroepithelial cells. These results provide morphologic evidence of the interactions between dendritic cells and the neural network of the airways at multiple anatomical sites.

The molecular mechanisms leading to reactivation of latent cytomegalovirus are not well understood. To study reactivation, the few cells in an organ tissue that give rise to reactivated virus need to be identified, ideally at the earliest possible time point in the process. To this end, mouse cytomegalovirus (MCMV) reporter mutants were designed to simultaneously express the red fluorescent protein mCherry and the secreted Gaussia luciferase (Gluc). Whereas Gluc can serve to assess infection at the level of individual mice by measuring luminescence in blood samples or by in vivo imaging, mCherry fluorescence offers the advatage of detection of infection at the single cell level. To visualize cells in which MCMV was being reactivated, precision-cut lung slices (PCLS) that preserve tissue microanatomy were prepared from the lungs of latently infected mice. By day 3 of cultivation of the PCLS, reactivation was revealed by Gluc expression, preceding the detection of infectious virus by approximately 4 days. Reactivation events in PCLS could be identified when they were still confined to single cells. Notably, using fractalkine receptor-GFP reporter mice, we never observed reactivation originating from CX3CR1 + monocytes or pulmonary dendritic cells derived therefrom. Furthermore, latent viral genome in the lungs was not enriched in sorted bone-marrow-derived cells expressing CD11b. Taken together, these complementary approaches suggest that CD11b + and CX3CR1 + subsets of the myeloid differentiation lineage are not the main reservoirs and cellular sites of MCMV latency and reactivation in the lungs. INTRODUCTIONHuman cytomegalovirus (HCMV) infection is highly prevalent in the human population (Mocarski et al., 2007). Primary infection often occurs in early childhood, usually without symptoms or with mild symptoms only. The acute infection is cleared by the immune system, but viral genomes remain in specific cells of the organism in a latent state. Cells of the myeloid lineage such as monocytes, which are the progenitors of dendritic cells and macrophages, but also other cell types such as endothelial cells, are considered to form the reservoir for latent HCMV genomes (reviewed by Jarvis & Nelson, 2002;Sinclair, 2008). Latency is operationally defined by the absence of detectable infectious virus and the capacity of the latent viral genomes to give rise to recurrent infection (Roizman & Sears, 1987). Reactivation of viral gene expression may be a relatively frequent event if viewed over long periods of time, however, the immune system in healthy individuals can recognize and terminate reactivation events even before virion assembly (Simon et al., 2006; reviewed by Reddehase et al., 2008). In addition, the immune system limits the spread of already reactivated virus, thus preventing recurrent viraemia and cytomegalovirus (CMV) disease (Polić et al., 1998). Consequently, HCMV reactivation is primarily a health risk for immunocompromised individuals such as transplant patients.The molecular events that lead to activati...