We have recently reported that the inhibition of endothelial cell COX-2 by non-steroidal anti-inflammatory drugs suppresses ␣ V ␤ 3 -(but not ␣ 5 ␤ 1 -) dependent Rac activation, endothelial cell spreading, migration, and angiogenesis (Dormond, O., Foletti, A., Paroz, C., and Ruegg, C. (2001) Nat. Med. 7, 1041-1047). Here we investigated the role of the COX-2 metabolites PGE 2 and TXA2 in regulating human umbilical vein endothelial cell (HUVEC) adhesion and spreading. We report that PGE 2 accelerated ␣ V ␤ 3 -mediated HUVEC adhesion and promoted Rac activation and cell spreading, whereas the TXA2 agonist U46619 retarded adhesion and inhibited spreading. We show that the cAMP level and the cAMP-regulated protein kinase A (PKA) activity are critical mediators of these PGE 2 effects. ␣ V ␤ 3 -mediated adhesion induced a transient COX-2-dependent rise in cAMP levels, whereas the cellpermeable cAMP analogue 8-brcAMP accelerated adhesion, promoted Rac activation, and cell spreading in the presence of the COX-2 inhibitor NS-398. Pharmacological inhibition of PKA completely blocked ␣ V ␤ 3 -mediated adhesion. A constitutively active Rac mutant (L61Rac) rescued ␣ V ␤ 3 -dependent spreading in the presence of NS398 or U46691, but did not accelerate adhesion, whereas a dominant negative Rac mutant (N17Rac) suppressed spreading without affecting adhesion. ␣ 5 ␤ 1 -mediated HU-VEC adhesion, Rac activation, and spreading were not affected by PGE 2 , U46691, 8-brcAMP, or the inhibition of PKA. In conclusion, these results demonstrate that PGE 2 accelerates ␣ V ␤ 3 -mediated endothelial cell adhesion through cAMP-dependent PKA activation and induces ␣ V ␤ 3 -dependent spreading via cAMP-and PKA-dependent Rac activation and may contribute to the further understanding of the regulation of vascular integrins ␣ V ␤ 3 by COX-2/PGE 2 during tumor angiogenesis and inflammation.Tumor angiogenesis, i.e. the formation of new blood vessels in response to angiogenic stimuli, promotes tumor progression by stimulating tumor cell survival, tumor invasion, and metastasis formation (1). Many molecules involved in mediating or regulating angiogenesis have been identified (2). They include growth factors (i.e. vascular endothelial growth factors, VEGF) 1 and their cell surface receptors, matrix-degrading enzymes (e.g. matrix metalloproteinases), vascular remodeling ligands, and receptors (i.e. angiopoietins and Tie receptors) and adhesion receptors of the integrin and cadherin families. Integrins are the main receptors for extracellular matrix proteins and consist of two non-covalently associated ␣ and ␤ subunits (3). Integrin ligand binding affinity and adhesion-promoting activity are regulated by intracellular events ("inside out" signaling) (4). Upon ligand binding, integrins rapidly cluster and recruit structural (e.g. ␣-actinin, talin, vinculin) and signaling (e.g. focal adhesion kinase, paxillin, c-Src) proteins to form characteristic structures called focal contacts or focal adhesions (5). Integrins and focal adhesions propagate tensional ...

Bisphosphonates are potent inhibitors of osteoclast function widely used to treat conditions of excessive bone resorption, including tumor bone metastases. Recent evidence indicates that bisphosphonates have direct cytotoxic activity on tumor cells and suppress angiogenesis, but the associated molecular events have not been fully characterized. In this study we investigated the effects of zoledronate, a nitrogen-containing bisphosphonate, and clodronate, a non-nitrogen-containing bisphosphonate, on human umbilical vein endothelial cell (