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Matrix metalloproteinase (MMP)-2 and -9 are pivotal in remodeling many tissues. However, their functions and candidate substrates for brain development are poorly characterized. Intercellular adhesion molecule-5 (ICAM-5; Telencephalin) is a neuronal adhesion molecule that regulates dendritic elongation and spine maturation. We find that ICAM-5 is cleaved from hippocampal neurons when the cells are treated with N-methyl-d-aspartic acid (NMDA) or α-amino-3-hydroxy-5-methylisoxazole-propionic acid (AMPA). The cleavage is blocked by MMP-2 and -9 inhibitors and small interfering RNAs. Newborn MMP-2– and MMP-9–deficient mice brains contain more full-length ICAM-5 than wild-type mice. NMDA receptor activation disrupts the actin cytoskeletal association of ICAM-5, which promotes its cleavage. ICAM-5 is mainly located in dendritic filopodia and immature thin spines. MMP inhibitors block the NMDA-induced cleavage of ICAM-5 more efficiently in dendritic shafts than in thin spines. ICAM-5 deficiency causes retraction of thin spine heads in response to NMDA stimulation. Soluble ICAM-5 promotes elongation of dendritic filopodia from wild-type neurons, but not from ICAM-5–deficient neurons. Thus, MMPs are important for ICAM-5–mediated dendritic spine development.

Leukocyte motility is known to be dependent on both ␤2-integrins and matrix metalloproteinases MMP-2/-9 or gelatinases, which mediate leukocyte adhesion and the proteolysis needed for invasion, respectively. Gelatinases not only play an important role in cell migration, tissue remodeling, and angiogenesis during development, but are also involved in the progression and invasiveness of many cancers, including leukemias. The concept that MMPs associate with integrins, as well as their importance in some physiologic and pathologic conditions, has been advanced previously but has not been examined on leukocytes. This review will examine mainly the function of the MMP-integrin complexes in normal leukocyte migration and the effect of integrin and broad-spectrum MMP inhibitors in tumor

The promiscuous CD11b/CD18 (Mac-1) integrin has important roles in regulating many immunologic functions such as leukocyte adhesion and emigration from the bloodstream via interactions with the endothelial ligands ICAM-1 and ICAM-2, iC3b-mediated phagocytosis, and apoptosis. However, the mechanisms for Mac-1 inside-out activation have remained poorly understood. Phosphorylation of integrin cytoplasmic domains is emerging as an important mechanism of regulating integrin functions. Here, we have stud-

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...

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We have recently demonstrated that promatrix metalloproteinases (proMMPs), particularly proMMP-9, are potent ligands of the leukocyte β2 integrins. We studied here the complex formation between proMMP-9 and αMβ2, the major MMP and integrin of neutrophils. On resting neutrophils, the proMMP-9/αMβ2 complex was primarily detected in intracellular granules, but after cellular activation it became localized to the cell surface, as demonstrated by immunoprecipitation and double immunofluorescence. Further indication of the complex formation was that neutrophils and αMβ2-transfected L cells, but not the wild-type L cells or leukocyte adhesion deficiency cells, bound to immobilized proMMP-9 or its recombinant catalytic domain in a β2 integrin-dependent manner. Peptides that bound to the αM integrin-I domain and inhibited its complex formation with proMMP-9 prevented neutrophil migration in a transendothelial assay in vitro and in a thioglycolate-elicited peritonitis in vivo. These results suggest that the translocating proMMP-9/αMβ2 complex may be part of the cell surface machinery guiding neutrophil migration.

Acute myelogenous leukemias (AMLs) are characterized by medullary and extramedullary invasion. We hypothesized that a supramolecular complex, the leukemiacell invadosome, which contains certain integrins, matrix metalloproteinases (MMPs), and other as-yet unidentified proteins, is essential for tissue invasion and may be central to the phenotypic diversity observed in the clinic. Here we show that the specific binding of MMP-9 to leukocyte surface ␤ 2 integrin is required for pericellular proteolysis and migration of AMLderived cells. An efficient antileukemia effect was obtained by the hexapeptide HFDDDE, a motif of the MMP-9 catalytic domain that mediates integrin binding: HFDDDE prevented proMMP-9 binding, transmigration through a human endothelial cell layer, and extracellular matrix degradation. Notably, the functional protein anchorage between ␤ 2 integrin and proMMP-9 described in this study does not involve the enzymatic active sites targeted by known MMP inhibitors. Taken