Human Siglec-10 can bind to vascular adhesion protein-1 and serves as its substrate

Abstract

Leukocytes migrate from the blood into areas of inflammation by interacting with various adhesion molecules on endothelial cells. Vascular adhesion protein-1 (VAP-1) is a glycoprotein expressed on inflamed endothelium where it plays a dual role: it is both an enzyme that oxidizes primary amines and an adhesin that is involved in leukocyte trafficking to sites of inflammation. Although VAP-1 was identified more than 15 years ago, the counterreceptor(s) for VAP-1 on leukocytes has remained unknown. Here we have identified Siglec-10 as a leukocyte ligand for VAP-1 using phage display screenings. The binding between Siglec-10 and VAP-1 was verified by different adhesion assays, and this interaction was also consistent with molecular modeling. Moreover, the interaction between Siglec-10 and VAP-1 led to increased hydrogen peroxide production, indicating that Siglec-10 serves as a substrate for VAP-1. Thus, the Siglec-10-VAP-1 interaction seems to mediate lymphocyte adhesion to endothelium and has the potential to modify the inflammatory microenvironment via the enzymatic end products. IntroductionMaintenance of an adequate immune defense is largely dependent on migration of circulating leukocytes from the vasculature into the surrounding tissue. Extravasation of leukocytes from the blood into the tissues is controlled by a multistep cascade, which involves numerous adhesion and signaling molecules. First, leukocytes tether and roll on the vascular endothelium, after which they adhere more strongly, arrest, and finally diapedese through the vessel wall. 1,2 One of the endothelial molecules involved in this cascade is a 180-kDa homodimeric glycoprotein, vascular adhesion protein-1 (VAP-1). 3 VAP-1 is mainly expressed on vascular endothelium, on smooth muscle cells, and on adipocytes. 4 It is rapidly translocated to the endothelial cell surface on inflammation, where it supports recruitment of leukocytes. 5,6 VAP-1 is involved in the rolling, firm adhesion, and transmigration phases of the extravasation cascade. Besides being an adhesin, VAP-1 is also an enzyme (semicarbazide sensitive amine oxidase, SSAO) and catalyzes oxidative deamination of a primary amine to an aldehyde with concomitant release of hydrogen peroxide and ammonium. 4 We and others have shown that VAP-1 supports leukocyte recruitment to sites of inflammation via both enzyme-activity-dependent and enzyme-activity-independent ways. Monoclonal anti-VAP-1 antibodies, which do not block the enzymatic activity of VAP-1, still block the binding of leukocytes to the endothelium in vitro and in vivo, and inhibitors of the enzymatic function of VAP-1 are able to effectively prevent the interaction between leukocytes and endothelium in vitro and in vivo. 7,8 Moreover, it has been recently shown that the end products generated by the catalytic activity of VAP-1, especially hydrogen peroxide, play a role in the regulation of other homing-associated molecules and thus enhance the inflammatory response. 9,10 Although VAP-1 has been known already for a long time and ...

Soluble Vascular Adhesion Protein-1 Correlates With Cardiovascular Risk Factors and Early Atherosclerotic Manifestations

“…16 Recombinant VAP-1 was purified from stable CHO-VAP-1 transfectants using immunoaffinity purification as described. 26 We found that the optimal final serum dilutions are 1:100 and 1:34 for the 96-and 384-well formats, respectively, and the linear range of the assay is from 2.0 to 86.0 nmol/mL/h. The intraassay and interassay variations were 4% and 10%, respectively, for the 96-well format (tested by measuring triplicates of 9 serum samples in 2 to 4 independent experiments), and 8% and 15%, respectively, for the 384-well format (tested by measuring duplicates of 32 serum samples in 7 independent experiments).…”

Reaction of Vascular Adhesion Protein-1 (VAP-1) with Primary Amines

“…Bovine VAP has been shown to bind, but not oxidize, lysine and soluble elastin in vitro in the presence of H 2 O 2 and thereby inhibit the catalytic turnover of benzylamine (13). Recently, a binding partner for VAP-1 on the surface of lymphocytes has been identified from phage libraries displaying decapeptides (14). Siglec-10 (sialic acid-binding Ig-like lectin-10), shown to bind specifically to recombinant VAP-1, is the first leukocyte ligand identified for the enzyme (14).…”

“…Recently, a binding partner for VAP-1 on the surface of lymphocytes has been identified from phage libraries displaying decapeptides (14). Siglec-10 (sialic acid-binding Ig-like lectin-10), shown to bind specifically to recombinant VAP-1, is the first leukocyte ligand identified for the enzyme (14). Binding results in H 2 O 2 production, suggesting that Siglec-10 acts as a substrate.…”

Immune regulation by CD52-expressing CD4 T cells

“…Given its broad immune distribution, suppression by CD52 released from CD52-expressing regulatory CD4 T cells may not be restricted to T cells only, but may also extend to a broader population of peripheral blood mononuclear cells that include dendritic cells, monocytes and B lymphocytes. As human Siglec-10 also binds to vascular adhesion protein-1 to act as its substrate and to mediate lymphocyte binding to the endothelium, 9 binding of CD52 to siglec-10 could also potentially modify the inflammatory microenvironment.…”