The Effects of CTGF on HCE Cell Re–epithelialisation and Migration

Abstract

Abstract: : Purpose: Connective tissue growth factor (CTGF) is a cysteine–rich, heparin–binding protein thought to be a downstream mediator of the profibrotic action of transforming growth factor beta (TGFß) in fibroblasts. CTGF’s effect on human corneal epithelial cells (HCEC) has not been completely characterised. Previously, we have found that HCEC produce CTGF in a temporal manner during differentiation into multi–layered epithelium and that CTGF stimulates proliferation. This study investigated the effects of CTGF and CTGF antisense oligonucleotides on HCEC wound healing and migration in comparision with TGFß1 and TGFß2. Methods: Confluent monolayers of HCE cell line (HCE–T) were wounded using a pipette tip. Digital photographs were taken immediately after wounding and at 5, 8, and 11 hours post wounding. Image tool was used to determine wound width at each time point. HCE–T migration was assessed using a colony dispersion assay. Using these two assays, under serum free conditions, the effects of various concentrations of CTGF, CTGF antisense, TGFß1 and TGFß2on re–epithelialisation and migration was compared with controls. Results: Percentage wound closure of HCE–T was increased significantly with CTGF and TGFß2 in a time dependant manner, with no effect on migration. CTGF antisense inhibited wound closure at concentrations of 10 & 100µM when examined at 5 & 11 hours and it did not affect migration. Comparatively, TGFß1 had no significant effect on wound closure and but induced a marked decreased in migration. Conclusions: CTGF and TGFß2 promote re–epithelialisation in vitro but did not affect migration of HCE–T cells. On this basis, we suggest that these growth factors may have an important role in human corneal epithelial wound healing.

Keywords: cornea: epithelium • wound healing • growth factors/growth factor receptors