The plasminogen activation system plays an integral role in the migration of macrophages in response to an inflammatory stimulus, and the binding of plasminogen to its cell-surface receptor initiates this process. Although previous studies from our laboratory have shown the importance of the plasminogen receptor S100A10 in cancer cell plasmin production, the potential role of this protein in macrophage migration has not been investigated. Using thioglycollate to induce a peritoneal inflammatory response, we demonstrate, for the first time, that compared with wild-type (WT) mice, macrophage migration across the peritoneal membrane into the peritoneal cavity in S100A10-deficient (S100A10 ؊/؊ ) mice was decreased by up to 53% at 24, 48, and 72 hours. Furthermore, the number of S100A10-deficient macrophages that infiltrated Matrigel plugs was reduced by 8-fold compared with their WT counterpart in vivo. Compared with WT macrophages, macrophages from S100A10 ؊/؊ mice demonstrated a 50% reduction in plasmin-dependent invasion across a Matrigel barrier and a 45% reduction in plasmin generation in vitro. This loss in plasmin-dependent invasion was in part the result of a decreased generation of plasmin and a decreased activation of pro-MMP-9 by S100A10-deficient macrophages. This study establishes a direct involvement of S100A10 in macrophage recruitment in response to inflammatory stimuli. (Blood. 2010;116(7): 1136-1146) IntroductionMonocytes/macrophages play a central role in pathogenic inflammatory responses associated with atherosclerosis, restenosis, tumor surveillance, and arthritis. [1][2][3] In response to changes in the cellular environment, monocytes and monocytoid cells undergo extensive phenotypic alterations, including marked changes in their fibrinolytic properties. Synthesis and activation of matrix-degrading proteinases by monocytes and macrophages play an essential role in their migration through tissue. A key proteinase that participates in pericellular proteolysis is the serine proteinase plasmin. Plasmin is a broad substrate proteinase that is formed from the inactive zymogen plasminogen (Plg) by the Plg activators, tissue Plg activator (tPA) and urokinase-type Plg activator (uPA). 4,5 The participation of plasmin in cell invasion and migration is dependent on the ability of plasmin not only to degrade extracellular matrix (ECM) proteins but to also activate other proteinases that have matrix-degrading activity. Plasmin can degrade a variety of matrix proteins, such as laminin and fibronectin, and appears to activate matrix metalloproteinase-1 (MMP-1), MMP-3, and MMP-13 directly, and to activate MMP-2 and MMP-9 indirectly, thereby facilitating cell migration through ECMs. 6 The assembly of Plg and its activators on the cell surface is facilitated by the protein S100A10 (also referred to as p11). S100A10 is a member of the S100 family of calcium-binding proteins and is typically found in most cells bound to its annexin A2 (p36) ligand as the heterotetrameric (S100A10) 2 -(annexin A2) 2 complex, annexi...

The plasminogen receptors mediate the production and localization to the cell surface of the broad spectrum proteinase, plasmin. S100A10 is a key regulator of cellular plasmin production and may account for as much as 50% of cellular plasmin generation. In parallel to plasminogen, the plasminogen-binding site on S100A10 is highly conserved from mammals to fish. S100A10 is constitutively expressed in many cells and is also induced by many diverse factors and physiological stimuli including dexamethasone, epidermal growth factor, transforming growth factor-α, interferon-γ, nerve growth factor, keratinocyte growth factor, retinoic acid, and thrombin. Therefore, S100A10 is utilized by cells to regulate plasmin proteolytic activity in response to a wide diversity of physiological stimuli. The expression of the oncogenes, PML-RARα and KRas, also stimulates the levels of S100A10, suggesting a role for S100A10 in pathophysiological processes such as in the oncogenic-mediated increases in plasmin production. The S100A10-null mouse model system has established the critical role that S100A10 plays as a regulator of fibrinolysis and oncogenesis. S100A10 plays two major roles in oncogenesis, first as a regulator of cancer cell invasion and metastasis and secondly as a regulator of the recruitment of tumor-associated cells, such as macrophages, to the tumor site.

The vascular endothelial cells line the inner surface of blood vessels and function to maintain blood fluidity by producing the protease plasmin that removes blood clots from the vasculature, a process called fibrinolysis. Plasminogen receptors play a central role in the regulation of plasmin activity. The protein complex annexin A2 heterotetramer (AIIt) is an important plasminogen receptor at the surface of the endothelial cell. AIIt is composed of 2 molecules of annexin A2 (ANXA2) bound together by a dimer of the protein S100A10. Recent work performed by our laboratory allowed us to clarify the specific roles played by ANXA2 and S100A10 subunits within the AIIt complex, which has been the subject of debate for many years. The ANXA2 subunit of AIIt functions to stabilize and anchor S100A10 to the plasma membrane, whereas the S100A10 subunit initiates the fibrinolytic cascade by colocalizing with the urokinase type plasminogen activator and receptor complex and also providing a common binding site for both tissue-type plasminogen activator and plasminogen via its C-terminal lysine residue. The AIIt mediated colocalization of the plasminogen activators with plasminogen results in the rapid and localized generation of plasmin to the endothelial cell surface, thereby regulating fibrinolysis. (Blood. 2011;118(18):4789-4797) IntroductionThe vascular endothelium consists of a single cell layer lining all vessels that separates the blood from the tissues. It is estimated to be composed of ϳ 10 13 cells, representing a weight of 1.5 kg and an area of 4000 to 7000 m 2 . 1 Endothelial cells play a role in primary hemostasis, coagulation, fibrinolysis, and regulation of vasomotor tone. In addition to regulating the flow of nutrients, the vascular endothelium regulates many diverse biologically active molecules. These functions of the endothelium are achieved through the presence of membrane-bound receptors for various proteins, lipidtransporting complexes, hormones, and metabolites, as well as through specific extracellular proteins and receptors that regulate cell-cell and cell-matrix interactions. 2 Whereas exposure to inflammatory and/or septic stimuli rapidly leads to procoagulant behavior, unperturbed endothelial cells provide an anticoagulant environment. After vascular insult, endothelial cells express tissue factor and initiate the coagulation cascade that results in thrombin activation and fibrin clot deposition. At the same time, anticoagulant pathways and fibrinolysis are activated to avoid disseminated coagulation and to also limit fibrin accumulation. [3][4][5] Fibrinogen is a large glycoprotein that constitutes the main component of a fibrin clot. Each fibrinogen molecule is composed of 2 sets of A␣-, B␤-, and ␥-polypeptide chains that form a protein containing 2 distal D regions connected to a central E region by a coiled-coil segment. 6 Fibrin is produced on cleavage of the fibrinopeptides by thrombin, which results in the formation of double-stranded half-staggered oligomers that lengthen into protofibrils...

Endothelial cells form the inner lining of vascular networks and maintain blood fluidity by inhibiting blood coagulation and promoting blood clot dissolution (fibrinolysis). Plasmin, the primary fibrinolytic enzyme, is generated by the cleavage of the plasma protein, plasminogen, by its activator, tissue plasminogen activator. This reaction is regulated by plasminogen receptors at the surface of the vascular endothelial cells. Previous studies have identified the plasminogen receptor protein S100A10 as a key regulator of plasmin generation by cancer cells and macrophages. Here we examine the role of S100A10 and its annexin A2 binding partner in endothelial cell function using a homozygous S100A10-null mouse. Compared with wild-type mice, S100A10-null mice displayed increased deposition of fibrin in the vasculature and reduced clearance of batroxobin-induced vascular thrombi, suggesting a role for S100A10 in fibrinolysis in vivo. Compared with wild-type cells, endothelial cells from S100A10-null mice demonstrated a 40% reduction in plasminogen binding and plasmin generation in vitro. Furthermore, S100A10-deficient endothelial cells demonstrated impaired neovascularization of Matrigel plugs in vivo, suggesting a role for S100A10 in angiogenesis. These results establish an important role for S100A10 in the regulation of fibrinolysis and angiogenesis in vivo, suggesting S100A10 plays a critical role in endothelial cell function. (Blood. 2011;118(11): 3172-3181)

Macrophages are critical drivers of tumor growth, invasion, and metastasis. Movement of macrophages into tumors requires the activity of cell surface proteases such as plasmin. In this study, we offer genetic evidence that plasminogen receptor S100A10 is essential for recruitment of macrophages to the tumor site. Growth of murine Lewis lung carcinomas or T241 fibrosarcomas was dramatically reduced in S100A10-deficient mice compared with wild-type mice. The tumor growth deficit corresponded with a decrease in macrophage density that could be rescued by intraperitoneal injection of wild-type but not S100A10-deficient macrophages. Notably, macrophages of either genotype could rescue tumor growth if they were injected into the tumor itself, establishing that S100A10 was required specifically for the migratory capability needed for tumor homing. Conversely, selective depletion of macrophages from wild-type mice phenocopied the tumor growth deficit seen in S100A10-deficient mice. Together, our findings show that S100A10 is essential and sufficient for macrophage migration to tumor sites, and they define a novel rate-limiting step in tumor progression. Cancer Res; 71(21); 6676-83. Ó2011 AACR.