The Role of Insulin-Like Growth Factor Binding Protein 2 (IGFBP2) in the Regulation of Corneal Fibroblast Differentiation

Abstract

Previously, we reported that keratocyte-conditioned medium (KCM) facilitates the differentiation of human mesenchymal stem cells (hMSCs) into corneal keratocyte-like cells. This study is designed to investigate the roles of insulin-like growth factor binding protein 2 (IGFBP2) for the regulation of corneal fibroblast differentiation as a newly unveiled component of KCM. Immunodot blot analysis was performed to identify the factors that are highly secreted, especially in KCM. Then, we investigated whether IGFBP2 differentiates hMSCs into keratocyte-like cells and whether maintains the phenotypes of keratocyte in human corneal fibroblasts (HCFs) by analyzing expression patterns of alpha-smooth muscle actin (α-SMA) and keratocyte markers including keratocan, lumican and aldehyde dehydrogenase 1 family member A1 (ALDH1A1). Furthermore, to specify the role of IGFBP2, the expression of α-SMA and keratocyte markers was determined in transforming growth factor β 1 (TGFβ1)-induced corneal myofibroblast and in HCFs after knockdown of IGFBP2. The most prominent factor in both KCM and amniotic membrane extract was IGFBP2. Insulin-like growth factor binding protein 2 increased the expression of IGFBP2, keratocan, and ALDH1A1, and decreased α-SMA expression in hMSCs and HCFs. Insulin-like growth factor binding protein 2 inhibited TGFβ1-induced upregulation of α-SMA and increased expressions of keratocan and ALDH1A1 in HCFs. Furthermore, the knockdown of IGFBP2 increased α-SMA expression and decreased ALDH1A1 level in HCFs. Insulin-like growth factor binding protein 2 is strongly associated with restoration of keratocyte phenotype in HCFs. Our results show an important novel role of IGFBP2 in regulation of corneal fibroblast differentiation and suggest that IGFBP2 can be a therapeutic candidate for corneal antifibrotic strategy.