The integrin α3β1 mediates cellular adhesion to the matrix ligand laminin-5. A second integrin ligand, the urokinase receptor (uPAR), associates with α3β1 via a surface loop within the α3 β-propeller (residues 242–246) but outside the laminin binding region, suggesting that uPAR–integrin interactions could signal differently from matrix engagement. To explore this, α3−/− epithelial cells were reconstituted with wild-type (wt) α3 or α3 with Ala mutations within the uPAR-interacting loop (H245A or R244A). Wt or mutant-bearing cells showed comparable expression and adhesion to laminin-5. Cells expressing wt α3 and uPAR dissociated in culture, with increased Src activity, up-regulation of SLUG, and down-regulation of E-cadherin and γ-catenin. Src kinase inhibition or expression of Src 1–251 restored the epithelial phenotype. The H245A and R244A mutants were unaffected by coexpression of uPAR. We conclude that α3β1 regulates both cell–cell contact and matrix adhesion, but through distinct protein interaction sites within its β-propeller. These studies reveal an integrin- and Src-dependent pathway for SLUG expression and mesenchymal transition.

Background-Elevated plasma levels of plasminogen activator inhibitor type 1 (PAI-1) are associated with myocardial infarction, atherosclerosis, and restenosis. PAI-1 is increased in atherosclerotic arteries and failed vein grafts. No experimental data, however, support a causal relationship between elevated PAI-1 expression and vascular lesions. Paradoxically, data generated in PAI-1 knockout mice suggest that PAI-1 might decrease lesion formation after arterial injury and that PAI-1 gene transfer might prevent restenosis. Methods and Results-Using the rat carotid balloon injury model and a PAI-1-expressing adenoviral vector, we tested whether elevated arterial PAI-1 expression would alter neointima formation. Compared with control-transduced arteries, neointima formation in PAI-1-transduced arteries was initially retarded. By 14 days, however, the intimas of PAI-1-transduced arteries were significantly larger than intimas of control-transduced arteries (1.6Ϯ0.1ϫ10 5 versus 1.2Ϯ0.1ϫ10 5 m 2 , nϭ18 to 19, PϽ0.03). PAI-1 expression in individual arteries correlated with increased cell proliferation at 4 and 8 days after injury (Rϭ0.6, PϽ0.02 and PϽ0.006). PAI-1 expression also correlated with fibrin(ogen) accumulation (Rϭ0.77, PϽ0.001), and fibrin(ogen) accumulation correlated strongly with proliferation (Rϭ0.86, PϽ0.00001). Conclusions-Increased expression of PAI-1 in the artery wall promotes neointima growth after balloon injury. Therefore, despite encouraging data generated in other animal models, PAI-1 is not a promising agent for gene therapy to prevent restenosis. Moreover, our data associate elevated PAI-1 expression with fibrin(ogen) accumulation and increased cell proliferation. These data suggest a mechanism to explain the association between elevated PAI-1 expression and the progression of arterial disease.

Overexpression of urokinase plasminogen activator (uPA) in endothelial cells can decrease intravascular thrombosis. However, expression of uPA is increased in atherosclerotic human arteries, which suggests that uPA might accelerate atherogenesis. To investigate whether elevated uPA expression accelerates atherogenesis, we cloned a rabbit uPA cDNA and expressed it in carotid arteries of cholesterol-fed rabbits. uPA gene transfer increased artery-wall uPA activity for at least 1 week, with a return to baseline by 2 weeks. One week after gene transfer, uPA-transduced arteries were constricted, with significantly smaller lumens and thicker walls, but no difference in intimal or medial mass. Two weeks after gene transfer, uPA-and control-transduced arteries were morphologically indistinguishable. By 4 weeks, however, uPA-transduced arteries had 70% larger intimas than control-transduced arteries (P < 0.01) and smaller lumens (P < 0.05). Intimal lesions appeared to be of similar composition in uPA-and control-transduced arteries, except that degradation of elastic laminae was evident in uPA-transduced arteries. These data suggest that elevated uPA expression in atherosclerotic arteries contributes to intimal growth and constrictive remodeling leading to lumen loss. Antagonists of uPA activity might, therefore, be useful in limiting intimal growth and preventing constrictive remodeling. Overexpression of uPA in endothelial cells to prevent intravascular thrombosis must be reconsidered, because this intervention could worsen underlying vascular disease.