Granulocyte transmigration through the endothelium is regulated by the oxidase activity of vascular adhesion protein-1 (VAP-1)

Abstract

IntroductionWhite blood cells of the polymorphonuclear series (polymorphonuclear leukocytes [PMNs] or granulocytes) are practically absent from healthy tissues. Upon inflammation, however, rapid influx of PMNs from the blood into the affected tissue takes place. PMNs leave the blood mainly via postcapillary venules and they do not recirculate to any significant extent. Lymphocytes, on the other hand, not only exit at the sites of inflammation but also continuously recirculate between the blood and lymphoid tissues. The physiologic recirculation of lymphocytes is supported by specialized high endothelial venules in lymphoid tissues, and this leukocyte type returns back to the blood via the lymphatic vasculature. Monocytes, the third major leukocyte type, normally leave the blood at low levels in various tissues under normal conditions to replenish the tissue macrophage pools, and at high levels during the later phase of inflammation. The extravasation process of all leukocyte classes consists of a series of carefully controlled steps. [1][2][3] First, the leukocyte makes initial tethers with the luminal surface of the blood vessel and starts to roll along the endothelial lining. If it receives appropriate activation signals, it can adhere in a shear-resistant manner to the endothelium. Finally, the leukocyte transmigrates through the vascular wall and continues its odyssey toward the chemotactic inflammation-induced signals within the tissue. Multiple adhesion and signaling molecules act in concert to execute the emigration cascade. One of the endothelial molecules involved in lymphocyte trafficking is vascular adhesion protein-1 (VAP-1). 4 VAP-1 is a cell-surface enzyme belonging to a specific group of amine oxidases (semicarbazide-sensitive amine oxidases [SSAOs] Enzyme Commission 1.4.3.6) that catalyze oxidative deamination of primary amines. 5,6 Here we questioned what the possible mechanistic connection is between the adhesive and enzymatic activity of VAP-1. By using adenoviral constructs encoding native or mutated VAP-1, chemical inhibitors of SSAO activity, and function-blocking anti-VAP-1 monoclonal antibodies (mAbs) in in vitro flow chamber assays we were able to study the role of VAP-1 during PMN rolling, firm adhesion, and transmigration. Interestingly, VAP-1 primarily supported the transmigration step during the PMN extravasation process. Experiments with the SSAO inhibitors and VAP-1 mutants showed that the enzymatic activity of VAP-1 is a prerequisite for the adhesive function of this molecule. Finally, we showed in a rat model of acute inflammation that the new, specific, and potent SSAO inhibitor also blocks the recruitment of PMNs to affected tissue in vivo. These data show that VAP-1 functions in leukocyte adhesion in a step-wise fashion via separate antibody epitope-dependent and oxidation-dependent steps. These experiments are the first to show that an oxidative enzymatic reaction on the luminal surface of endothelial cells regulates PMN emigration in vitro and in vivo. Patients, materials, ...

Vimentin regulates EMT induction by Slug and oncogenic H-Ras and migration by governing Axl expression in breast cancer

“…shAxl and shLuc MDA-MB-231 cells were stained with live cell dyes (shAxl red, CMTPX; shLuc green, CMFDA: Invitrogen) according to the manufacturer's instructions. Cells were harvested, suspended in 50 ml PBS (5 Â 10 5 each) and injected into lateral tail vein of mice (n ¼ 10); 48 h postinjection, the mice were injected with mouse anti-human HLA antibody (Hb116; Koskinen et al, 2004) 34 mg/ml for 5 min (to distinguish between human cells adhering to the vessel wall from those that have extravasated inside the tissue). Next, the pulmonary vasculature was perfused with PBS through the right ventricle (2 min) and blood was allowed to escape by a small incision in the left atrium.…”

Ischemia‐reperfusion injury is attenuated in VAP‐1‐deficient mice and by VAP‐1 inhibitors

“…Thus, leukocyte-endothelial interactions and development of inflammatory infiltrate in vivo have been inhibited to a similar extent either by anti-VAP-1 antibodies or small molecule SSAO inhibitors [20,32,33]. Moreover, combined use of both reagents has not led to additive or synergistic effects [20,32]. This has led to a hypothesis, in which leukocytes first use a counter-receptor to interact with surface-exposed antibodydefined epitopes of VAP-1.…”

Function-blocking antibodies to human vascular adhesion protein-1: A potential anti-inflammatory therapy

“…Recent work has also demonstrated that VAP-1 functions as a regulator of neutrophil transmigration [12]. These studies have shown that neutrophils bind to VAP-1 at two sites and that blockade of either site reduces neutrophil transmigration into the tissues [12]. The antibodies described in this study block one of these sites, the other is the SSAO enzyme active site of VAP-1.…”

“…Since VAP-1 functions also as a regulator of transmigration [12], we investigated the effect of Ch2D10 in an in vitro assay of transmigration. We examined the ability of lymphocytes to transmigrate through a monolayer of mock-transfected or VAP-1-expressing Ax cells grown on a co-culture insert, towards fMLP (Fig.…”

“…In addition to this, VAP-1 knockout mouse models have shown that lack of the protein leads to increased rolling velocity and a decreased rate of leukocyte transmigration [11]. Recent work has also demonstrated that VAP-1 functions as a regulator of neutrophil transmigration [12]. These studies have shown that neutrophils bind to VAP-1 at two sites and that blockade of either site reduces neutrophil transmigration into the tissues [12].…”