Up-Regulation of Integrin α5 Expression by Combination of Substance P and Insulin-like Growth Factor-1 in Rabbit Corneal Epithelial Cells

Abstract

To clarify the mechanisms by which substance P (SP) and insulin-like growth factor-1 (IGF-1) synergistically facilitate corneal epithelial wound healing, we tested the hypothesis that the combination promotes cell attachment to a fibronectin matrix through up-regulation of expression of integrin α5β1, the major cell surface fibronectin receptor in rabbit corneal epithelial cells. Cultured rabbit corneal epithelial cells were treated with SP and/or IGF-1 and then plated on wells coated with fibronectin and bovine serum albumin. After incubation, the number of cells attached to the wells was counted. In a second experiment, reverse transcription-polymerase chain reaction was used to determine the expression of integrin α5 and β1 by cells pretreated with SP and/or IGF-1. The combination of SP and IGF-1 significantly increased the number of cells attached to the fibronectin matrix and the expression of integrin α5. However, attachment to the fibronectin matrix was inhibited by the addition of GRGDSP, a synthetic peptide that mimics fibronectin. Thus, the synergistic enhancing effect of SP and IGF-1 on the attachment of corneal epithelial cells to the fibronectin matrix and on corneal epithelial migration is partly due to the up-regulation of integrin α5 expression in corneal epithelial cells.