α v β 3 Integrin Induces Tyrosine Phosphorylation–Dependent Ca 2+ Influx in Pulmonary Endothelial Cells

Abstract—The endothelial αvβ3 integrin occurs luminally, where its ligation by soluble agents may induce inflammatory signaling. We tested this hypothesis in bovine pulmonary artery endothelial cell monolayers with the use of vitronectin and cross-linking antibodies to ligate and aggregate the integrin. We quantified the endothelial cytosolic Ca2+ concentration ([Ca2+]i) according to the Fura 2 ratio imaging method in single cells of confluent monolayers. At baseline, endothelial [Ca2+]i levels remained steady at 86 nmol/L for >20 minutes. Cross-linking of the αvβ3 integrin through the sequential exposure of monolayers to anti-αvβ3 monoclonal antibody LM609 and secondary IgG resulted in a [Ca2+]i increase of 100% above baseline. This increase commenced in <0.5 minute, peaked in <2 minutes, and decayed to baseline in ≈5 minutes. Similar responses occurred after the addition of vitronectin (400 μg/mL). In contrast, external Ca2+ depletion blunted the cross-linking–induced [Ca2+]i increase by 60%, a response...