Endothelial CD47 interaction with SIRPγ is required for human T-cell transendothelial migration under shear flow conditions in vitro

Abstract

Leukocyte transendothelial migration (TEM) is a critical event during inflammation. CD47 has been implicated in myeloid cell migration across endothelium and epithelium. CD47 binds to signal regulatory protein (SIRP), SIRP␣ and SIRP␥. So far, little is known about the role of endothelial CD47 in T-cell TEM in vivo or under flow conditions in vitro. Fluorescence-activated cell sorting and biochemical analysis show that CD3 ؉ T cells express SIRP␥ but not SIRP␣, and fluorescence microscopy showed that CD47 was enriched at endothelial junctions. These expression patterns suggested that CD47 plays a role in T-cell TEM through binding interactions with SIRP␥. We tested, therefore, whether CD47-SIRP␥ interactions affect T-cell transmigration using blocking mAb against CD47 or SIRP␥ in an in vitro flow model. These antibodies inhibited T-cell TEM by 70% plus or minus 6% and 82% plus or minus 1%, respectively, but had no effect on adhesion. In agreement with human mAb studies, transmigration of murine wild-type T helper type 1 cells across TNF-␣-activated murine CD47 ؊/؊ endothelium was reduced by 75% plus or minus 2% even though murine T cells appear to lack SIRP␥. Nonetheless, these findings suggest endothelial cell CD47 interacting with T-cell ligands, such as SIRP␥, play an important role in T-cell transendothelial migration. (Blood. 2008; 112:1280-1289) IntroductionRecruitment of leukocytes from the bloodstream into tissues during an inflammatory response involves a multistep adhesive and signaling cascade composed of selectin-mediated rolling and initial attachment, subsequent integrin-mediated arrest and migration on the apical surface, followed by transendothelial migration (TEM; diapedesis). 1 TEM involves multiple adhesion molecule pathways such as intracellular adhesion molecule (ICAM)-1-CD18, vascular cell adhesion molecule (VCAM)-1-VLA-4, CD47, platelet endothelial cell adhesion molecule (PECAM)-1, CD99, endothelial cell adhesion molecule (ESAM), junctional adhesion molecules (JAMs), and DNAX accessory molecule 1 [CD226]-polio virus receptor [CD155] (DNAM-1-PVR), although the cellular mechanisms controlling TEM are incompletely understood. 2 CD47 or integrin-associated protein (IAP) belongs to the immunoglobulin superfamily (IgSF) and is highly expressed in most cell types. It is a 50-kDa transmembrane protein that consists of an extracellular amino-terminal Ig domain, 5 highly hydrophobic putative membrane-spanning segments, and a short cytoplasmic tail. 3 CD47 has been shown to associate with integrins at the cell surface and signal through G-coupled proteins. 4 Important cellular ligands for CD47 are the signal regulatory proteins (SIRPs) SIRP␣ and SIRP␥. The SIRPs comprise a family of several transmembrane glycoproteins that belong to the IgSF and are present in the immune and central nervous system. 5 Each molecule contains 3 homologous extracellular Ig-like domains with distinct transmembrane and cytoplasmic domains. SIRP␣ was the first member to be identified and was detected on hematopoietic progeni...

JAM-related proteins in mucosal homeostasis and inflammation

“…After initial adhesion, PMN negotiate the paracellular space along the basolateral epithelial membrane through interactions with CD47, a transmembrane glycoprotein that binds to its receptor on PMN termed SIRPα (Signal Regulatory Protein alpha) [116–118]. CD47 and SIRPα binding interactions have also been shown to regulate the rate of PMN migration across the endothelium [119,120]. As PMNs migrate across epithelial intercellular junctions, they disrupt E-cadherin-mediated cell contacts in an elastase-dependent manner [121].…”

Signal regulatory protein alpha (SIRPα)/CD47 interaction and function

“…The third member SIRPγ binds CD47 albeit weaker than SIRPα but it lacks an extensive cytoplasmic region and is unlikely to signal. Recent data suggest it has a role in T cell migration in endothelium [9]. In considering the CD47 / SIRPα interaction it is necessary to consider signals through SIRPα, why SIRPβ does not bind along with how CD47 signals itself and the role of additional ligands for the SIRPs and CD47.…”

Revisiting the role of B cells in skin immune surveillance

“…The exact signals that control the final stage of lymphocyte trafficking, which involves the transmigration of cells across vascular endothelia, are still incompletely understood. However multiple adhesion molecule pathways including ICAM-1, vascular cell adhesion molecule (VCAM)-1, and CD47 have been described and are still being investigated [108,109].…”