Fisp12 was first identified as a secreted protein encoded by a growth factor-inducible immediate-early gene in mouse fibroblasts, whereas its human ortholog, CTGF (connective tissue growth factor), was identified as a mitogenic activity in conditioned media of human umbilical vein endothelial cells. Fisp12/CTGF is a member of a family of secreted proteins that includes CYR61, Nov, Elm-1, Cop-1/WISP-2, and WISP-3. Fisp12/CTGF has been shown to promote cell adhesion and mitogenesis in both fibroblasts and endothelial cells and to stimulate cell migration in fibroblasts. These findings, together with the localization of Fisp12/CTGF in angiogenic tissues, as well as in atherosclerotic plaques, suggest a possible role for Fisp12/CTGF in the regulation of vessel growth during development, wound healing, and vascular disease. In this study, we show that purified Fisp12 (mCTGF) protein promotes the adhesion of microvascular endothelial cells through the integrin receptor ␣ v ␤ 3 . Furthermore, Fisp12 stimulates the migration of microvascular endothelial cells in culture, also through an integrin-␣ v ␤ 3 -dependent mechanism. In addition, the presence of Fisp12 promotes endothelial cell survival when cells are plated on laminin and deprived of growth factors, a condition that otherwise induces apoptosis. In vivo, Fisp12 induces neovascularization in rat corneal micropocket implants. These results demonstrate that Fisp12 is a novel angiogenic inducer and suggest a direct role for Fisp12 in the adhesion, migration, and survival of endothelial cells during blood vessel growth. Taken together with the recent finding that the related protein CYR61 also induces angiogenesis, we suggest that Fisp12/mCTGF and CYR61 comprise prototypes of a new family of angiogenic regulators that function, at least in part, through integrin-␣ v ␤ 3 -dependent pathways.

The angiogenic inducers cysteine-rich angiogenic protein 61 (Cyr61) and connective tissue growth factor (CTGF) are structurally related, extracellular matrixassociated heparin-binding proteins. Both can stimulate chemotaxis and promote proliferation in endothelial cells and fibroblasts in culture and induce neovascularization in vivo. Encoded by inducible immediate early genes, Cyr61 and CTGF are synthesized upon growth factor stimulation in cultured fibroblasts and during cutaneous wound healing in dermal fibroblasts. Recently, we have shown that adhesion of primary human fibroblasts to immobilized Cyr61 is mediated through integrin ␣ 6 ␤ 1 and cell surface heparan sulfate proteoglycans (HSPGs) (Chen, N., Chen, C.-C., and Lau, L.F. (2000) J. Biol. Chem. 275, 24953-24961), providing the first demonstration of an absolute requirement for HSPGs in integrin-mediated cell attachment. We show in this study that CTGF also mediates fibroblast adhesion through the same mechanism and demonstrate that fibroblasts adhesion to immobilized Cyr61 or CTGF induces distinct adhesive signaling responses consistent with their biological activities. Compared with fibroblast adhesion to fibronectin, laminin, or type I collagen, cell adhesion to Cyr61 or CTGF induces 1) more extensive and prolonged formation of filopodia and lamellipodia, concomitant with formation of integrin ␣ 6 ␤ 1 -containing focal complexes localized at leading edges of pseudopods; 2) activation of intracellular signaling molecules including focal adhesion kinase, paxillin, and Rac with similar rapid kinetics; 3) sustained activation of p42/p44 MAPKs lasting for at least 9 h; and 4) prolonged gene expression changes including up-regulation of MMP-1 (collagenase-1) and MMP-3 (stromelysin-1) mRNAs and proteins sustained for at least 24 h. Together, these results establish Cyr61 and CTGF as bona fide adhesive substrates with specific signaling capabilities, provide a molecular basis for their activities in fibroblasts through integrin ␣ 6 ␤ 1 and HSPG-mediated signaling during attachment and indicate that these proteins may function in matrix remodeling through the activation of metalloproteinases during angiogenesis and wound healing.

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