Angiogenesis is essential for tumor growth and metastasis. In the process of angiogenesis, the interaction between adhesive proteins of endothelial cells and extracellular matrix components plays an important role by mediating cell attachment, which is indispensable for their motility, and by transmitting the regulatory signals for cell locomotion and proliferation. In this study, we examined the hypothesis that CD44 expressed on the endothelial cell surface is involved in the angiogenesis process. The experiments using calf pulmonary artery endothelial cells (CPAE) and a human microvascular endothelial cell line (HMEC-I) show that a monoclonal antibody against CD44 (clone J 173) inhibits endothelial cell proliferation by about 30% and migration by 25-50%, and abolishes the stimulating effect of hyaluronan polysaccharides on endothelial cell migration and proliferation. This antibody also suppresses the capillary formation of CPAE in an in vitro model of angiogenesis using fibrin matrix. These results provide evidence of the involvement of endothelial-cell-associated CD44 in angiogenesis.o 1996 Wiley-Liss, Inc.Angiogenesis plays a fundamental role in many physiological and pathological processes including wound healing, tissue repair and tumor growth (Folkman, 1995). For the formation of a new vessel, migration of stimulated endothelial cells and subsequent tube formation virtually depend on the occurrence of timely and locally coordinated extracellular proteolysis, cell adhesion and activation during the cell migration. Therefore, the interaction of adhesive proteins expressed on the endothelial cell surface with the components of extracellular matrix is one of the factors determining the occurrence of angiogenesis. Perturbation of these interactions may abolish an ongoing angiogenesis process. For example, blocking the binding of the integrin molecule to matrix-associated vitronectin led to endothelial cell apoptosis and abolished tumor-induced angiogenesis in vivo (Brooks et al., 1994). Furthermore, the interaction of adhesive proteins with their endothelial cell-surface receptors may also activate the process of angiogenesis. For instance, endothelial cells could be stimulated by the surrounding fibrin I1 matrix to form capillary tubes in vitro (Chalupowicz et al., 1995). However, due to the complexity of the mechanisms involved in cell adhesion, the functions of many adhesive molecules on the endothelial cell surface still remain to be determined.One of the examples is CD44, discovered through its role in mediating leukocyte adhesion and lymphocyte homing (Haynes et al., 1989). Although several studies reported the presence of CD44 on endothelial cells such as human microvascular endothelial cells (HMEC-1) (Xu et al., 1994), bovine microvascular endothelial cells (BME) (Bourguignon et al., 1992) and human umbilical endothelial cells (HUVEC) (Liesveld et al., 1994), the role of CD44 expressed on the endothelial cell surface still remains to be clarified. CD44 is a transmembrane molecule with multiple isofo...

The binding of urokinase (u-PA) to its cell surface receptor (u-PAR) is critical for tumor cell invasion. Here, we report that the disruption of this binding by an u-PAR antagonist ATF-HSA inhibits in vitro the motility of endothelial cells in a dose-dependent manner. This inhibition was also observed when the cells were first stimulated with potent angiogenic factors, including bFGF or VEGF.[3H]thymidine incorporation assay demonstrated that ATF-HSA did not affect the cell proliferation. ATF-HSA was more potent than plasmin inhibitors, suggesting that it exerts its effects not solely by inhibiting the remodeling of the extracellular matrix. In fact, analysis of the cell shape change during migration revealed for the first time that its effect is related to a decrease in cell deformability. These results suggest that u-PAR antagonist may be a new approach to control angiogenesis.

Receptor-bound urokinase is likely to be a crucial determinant in both tumor invasion and angiogenesis. We report here that a yeast-derived genetic conjugate between human serum albumin and the 1-135 N-terminal residues of urokinase (u-PA) competitively inhibits the binding of exogenous and endogenous u-PA to its cell-anchored receptor (u-PAR). This hybrid molecule (ATF-HSA) also inhibits in vitro pro-urokinasedependent plasminogen activation in the presence of u-PAR bearing cells. These effects are probably responsible for the observed in vitro inhibition of tumor cell invasion in a reconstituted basement membrane extract (Matrigel).