title: Ligand occupancy of the alpha-V-beta3 integrin is necessary for smooth muscle cells to migrate in response to insulin-like growth factor.
abstract: Smooth muscle cells (SMCs) have been shown to migrate in response to insulin-like growth factor I (IGF-I). However, the mechanism mediating this response has not been determined. The migration rates of porcine and human vascular SMCs were assessed in a monolayer wounding assay. IGF-I and IGF-II induced increases of 141% and 97%, respectively, in the number of cells that migrated in 4 days. The presence of 0.2% fetal bovine serum in the culture medium was necessary for the IGFs to stimulate migration over uncoated plastic surfaces. However, if vitronectin was used as the substratum, IGF-I stimulated migration by 162% even in the absence of serum. To determine the role of integrins in mediating this migration, SMC surface proteins were labeled with 125I and immunoprecipitated with specific anti-integrin antibodies. Integrins containing alpha-V (vitronectin receptor), alpha5 (fibronectin receptor), and alpha3 (collagen/laminin receptor) subunits were the most abundant. IGF-I treatment caused a 73% reduction in alpha5-integrin subunit protein and a 25% increase in alpha-V subunit. More importantly, ligand binding of alpha-V-beta3 was increased by 2.4-fold. We therefore examined whether the function of the alpha-V-beta3 integrin was important for IGF-I-mediated migration. The disintegrin kistrin was shown by affinity crosslinking to specifically bind with high affinity to alpha-V-beta3 and not to alpha5-beta1 or other abundant integrins. The related disintegrin echistatin specifically inhibited 125I-labeled kistrin binding to alpha-V-beta3, while a structurally distinct disintegrin, decorsin, had 1000-fold lower affinity. The addition of increasing concentrations of either kistrin or echistatin inhibited IGF-I-induced migration, whereas decorsin had a minimal effect. The potency of these disintegrins in inhibiting IGF-I-induced migration paralleled their apparent affinity for the alpha-V integrin. Furthermore, an alpha-V-beta3 blocking antibody inhibited SMC migration by 80%. In summary, vitronectin receptor activation is a necessary component of IGF-I-mediated stimulation of smooth muscle migration, and alpha-V-beta3 integrin antagonists appear to be important reagents for modulating this process.
authors.affiliation: Department of Medicine, University of North Carolina School of Medicine, Chapel Hill 27599-7170, USA.
authors.forename: J I
authors.initials: JI
authors.lastname: Jones
authors.forename: T
authors.initials: T
authors.lastname: Prevette
authors.forename: A
authors.initials: A
authors.lastname: Gockerman
authors.forename: D R
authors.initials: DR
authors.lastname: Clemmons
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D015703:Antigens, CD / Q000378:metabolism*; D002465:Cell Movement; D002633:Chemotaxis; D007334:Insulin-Like Growth Factor I / Q000502:physiology*; D039564:Integrin alphaV; D039661:Integrin beta3; D036341:Intercellular Signaling Peptides and Proteins; D008024:Ligands; D009130:Muscle, Smooth / Q000166:cytology*; D010455:Peptides / Q000494:pharmacology; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism*; D017526:Receptor, IGF Type 1 / Q000502:physiology; D019030:Receptors, Vitronectin / Q000032:analysis / Q000378:metabolism; D013552:Swine; D019096:Vitronectin / Q000502:physiology
chemical_list: D015703:Antigens, CD; D039564:Integrin alphaV; D039661:Integrin beta3; D036341:Intercellular Signaling Peptides and Proteins; D008024:Ligands; D010455:Peptides; D010980:Platelet Membrane Glycoproteins; D019030:Receptors, Vitronectin; D019096:Vitronectin; C057986:echistatin; D007334:Insulin-Like Growth Factor I; C067623:kistrin; D017526:Receptor, IGF Type 1