Endogenous IGF-I and alphaVbeta3 integrin ligands regulate increased smooth muscle hyperplasia in stricturing Crohn's disease.

Abstract

Insulin-like growth factor-I (IGF-I) regulates human intestinal smooth muscle growth by stimulating proliferation and inhibiting apoptosis. IGF-I-stimulated growth is augmented when alphaVbeta3 integrin is occupied by its ligands, fibronectin and vitronectin. Increased IGF-I expression and muscle cell hyperplasia are features of stricturing Crohn's disease (CD); however, the role of IGF-I in stricture formation is unknown. The aim was to identify the functional role of endogenous IGF-I and alphaVbeta3 integrin ligands in regulating muscle cell hyperplasia in stricturing CD. Smooth muscle cells were isolated from muscularis propria of stricturing CD or normal margins. Quantitative polymerase chain reaction, immunoblot analysis, and enzyme-linked immunosorbent assay were used to measure fibronectin, vitronectin, alphaVbeta3 integrin, and IGF-I levels. Activation of the IGF-I receptor, Erk1/2, p70S6 kinase, and GSK-3beta was measured by immunoblot. Proliferation was quantified by Ki67 immunostaining and [(3)H]thymidine incorporation. Apoptosis was measured from caspase-3 cleavage and nucleosome accumulation. IGF-I, vitronectin, and fibronectin RNA and protein levels were increased 1.8- to 3.4-fold in muscle cells from strictures over normal margins. Basal IGF-I receptor phosphorylation was increased 320% in strictured over normal muscle, and basal Erk1/2, p70S6 kinase, and GSK-3beta phosphorylation were increased 205%-292% in strictures. In muscle cells from strictures, Ki67 immunoreactivity and [(3)H]thymidine incorporation were increased and apoptosis was decreased compared with normal margins. Antagonists of the IGF-I receptor or alphaVbeta3 integrin reversed these changes. Smooth muscle cell hyperplasia in stricturing CD is regulated by increased endogenous IGF-I and alphaVbeta3 integrin ligands that regulate augmented proliferation and diminished apoptosis.