Urokinase receptor mediates lung fibroblast attachment and migration toward provisional matrix proteins through interaction with multiple integrins

Abstract

MA. Urokinase receptor mediates lung fibroblast attachment and migration toward provisional matrix proteins through interaction with multiple integrins. Am J Physiol Lung Cell Mol Physiol 297: L97-L108, 2009. First published May 1, 2009 doi:10.1152/ajplung.90283.2008.-Fibroblasts from patients with pulmonary fibrosis express higher levels of the receptor for urokinase, and the extent of fibrosis in some animal models exhibits a dependence on the urokinase receptor. Recent observations have identified the urokinase receptor as a trans-interacting receptor with consequences on signaling and cell responses that vary depending on its interacting partner, the relative levels of expression, and the state of cellular transformation. We undertook this study to define the urokinase-type plasminogen activator cellular receptor (u-PAR)-integrin interactions and to determine the functional consequences of such interactions on normal human lung fibroblast attachment and migration. u-PAR colocalizes in lammelipodia/filopodia with relevant integrins that mediate fibroblast attachment and spreading on the provisional matrix proteins vitronectin, fibronectin, and collagens. Inhibitory antibody studies have revealed that human lung fibroblasts utilize ␣ v␤5 to attach to vitronectin, predominantly ␣5␤1 (and ␣v␤3) to attach to fibronectin, and ␣ 1␤1, ␣2␤1, and ␣3␤1 to attach to collagen. Blocking studies with ␣-integrin subunit decoy peptides and u-PAR neutralizing antibodies indicate that u-PAR modulates the integrinmediated attachment to purified provisional matrix proteins, to antiintegrin antibodies, or to fibroproliferative lesions from fibrotic lungs. Furthermore, these decoy peptides blunt fibroblast spreading and migration. We show that u-PAR can interact with multiple ␣-integrins but with a preference for ␣ 3. Taken together, these data demonstrate that u-PAR may interact with multiple integrins in normal human lung fibroblasts thereby promoting attachment, spreading, and migration. Modulation of fibroblast invasion would be expected to lead to amelioration of fibroproliferative diseases of the lung. pulmonary; fibrosis; plasmin STUDIES IN VITRO AND IN TRANSGENIC animals have demonstrated that the fibrinolytic system has protean effects on numerous disease processes including acute and chronic inflammatory disorders, cancer invasion and metastasis, angiogenesis, and fibroproliferative disorders of the vasculature, lung, kidney, and eye (3,4,11,16,22,23,27,29,37,55). These disorders have the common feature of a complex, yet coordinated movement of cells and the biochemical and structural remodeling of the extracellular matrix. The molecular interactions and cell biological processes that mediate the effects of the fibrinolytic system on cell-matrix interactions are active areas of investigation.Protease-protease inhibitor mechanisms of the urokinase/ plasmin system that relate to intravascular or alveolar fibrin turnover are deranged in pulmonary fibrosis but are clearly only a part of the relevant biology (38). The recep...

Kinase Activity is Required for Growth Regulation but not Invasion Suppression by Syk Kinase in Pancreatic Adenocarcinoma Cells

“…Integrin v3 suppresses uPA/uPAR expression in ovarian cancer cells (Hapke et al, 2001) and, as noted previously, the MMP2 axis has been associated with expression of the v3 integrin and v3 is required for MMP2 activation in some cell types (Deryugina et al, 2001); Nisato et al, 2005), including PDAC (Hosotani et al, 2002). Reciprocally, the uPA/uPAR axis is linked to v5 (Yebra et al, 1995; and other integrins including 31 (Zhu et al, 2009), which is also expressed by both MIAPaCa2 and BxPC3 cells (S. Silletti, unpublished data). Importantly, the differential integrin utilization by these cells extended to the regulation of invasion as well.…”

Plasminogen Activator Inhibitor-1 Suppresses Profibrotic Responses in Fibroblasts from Fibrotic Lungs

“…We (22) and others (7,8,37) have previously reported that the basal level of proliferation was significantly higher in hFL-fibroblasts than in hNL-fibroblasts, and IPF tissues showed increased Ki-67 antigen and reduced PAI-1 expression in fibrotic foci (Fig. 1).…”

TRPV4 mediates myofibroblast differentiation and pulmonary fibrosis in mice

“…To understand the pathophysiological relevance of our observations to fibrotic lung disease, we developed a model system to test the interaction of fibroblasts with actual normal and fibrotic lung tissue (38). The contribution of TRPV4 to myofibroblast differentiation was studied using fibroblasts plated on unfixed normal and fibrotic lung tissue sections from bleomycin-treated WT mice.…”

“…Furthermore, TRPV4 mediates the directional migration of endothelial cells noted upon local matrix compression (46). Here, we investigated whether TRPV4 will integrate mechanical (matrix stiffness) and soluble (TGF-β1) signals to mediate lung myofibroblast differentiation, using collagen-coated polyacrylamide gels of varying degrees of stiffness (47), in a model system we developed using actual lung tissue sections (38). We found that the TRPV4 channel activity (agonist-induced Ca 2+ influx) is potentiated when cells are plated on stiffer matrices within the pathophysiological range seen in diseased/fibrotic lung tissue.…”

“…Female C57BL/6 mice were injected with 4 U/kg bleomycin or saline via IT instillation, and, after 2 weeks, lungs were inflated with OCT to prepare 10-μm sections containing both fibrotic and normal areas as published previously (38,59). HLFs were allowed to attach on the 5% BSA-blocked lung sections, and unattached cells were washed off with PBS as published previously (38,59). After washing, lung section-containing cells were treated with vehicle or AB1 (20 μM) in 1% BSA/serum-free medium (24 hours), and staining for F-actin was performed using Alexa Fluor 594-labeled phalloidin (5 U/ml, 60-minute incubation at room temperature; Molecular Probes).…”