Osteopontin overexpression inhibits in vitro re-endothelialization via integrin engagement

Daria Leali, Emanuela Moroni, Federico Bussolino, Marco Presta

Research output: Contribution to journalArticle

Abstract

The extracellular matrix protein osteopontin (OPN) plays a nonredundant role in atherosclerosis and restenosis. Here we investigated the impact of OPN up-regulation in an in vitro model of re-endothelialization after mechanical injury of the endothelial cell monolayer. Murine aortic endothelial (MAE) cells interact via αv integrins with the integrin-binding Arg-Gly-Asp OPN sequence and adhere to immobilized OPN. On this basis, MAE cells were stably transfected with a wild-type OPN cDNA (OPN-MAE cells), with an OPN mutant lacking the Arg-Gly-Asp sequence (ΔRGD-OPN-MAE cells), or with vector alone (mock-MAE cells). When compared with mock-MAE and ΔRGD-OPN-MAE cells, OPN-MAE cells showed a reduced sprouting activity in fibrin gel, a reduced motility in a Boyden chamber assay, and a reduced capacity to repair the wounded monolayer. Accordingly, OPN-MAE cells at the edge of the wound were unable to form membrane ruffles, to reorganize their cytoskeleton, and to activate the focal adhesion kinase and the small GTPase Rac1, key regulators of the cell entry into the first phase of the cell migration cycle. Accordingly, wounded OPN-MAE cells failed to activate the intracellular signals RhoA and ERK1/2, involved in the later phases of the cell migration cycle. Also, parental MAE cells showed reduced re-endothelialization after wounding when seeded on immobilized OPN and exhibited increased adhesiveness to OPN-enriched extracellular matrix. In conclusion, OPN up-regulation impairs re-endothelialization by inhibiting the first phase of the cell migration cycle via αv integrin engagement by the extracellular matrix-immobilized protein. This may contribute to the adverse effects exerted by OPN in restenosis and atherosclerosis.

Original languageEnglish
Pages (from-to)19676-19684
Number of pages9
JournalJournal of Biological Chemistry
Volume282
Issue number27
DOIs
Publication statusPublished - Jul 6 2007

ASJC Scopus subject areas

  • Biochemistry

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