Role of Fibronectin-EDA in Retinal Wound Healing and Fibrosis

Abstract

Purpose:: The alternatively spliced embryonic isoform of fibronectin (FN-EDA) is one of the earliest extracellular matrix components expressed in response to injury and found in abundance in fibrotic disorders. We previously demonstrated that FN-EDA expression in RPE cells is upregulated by TGFß2 in a time- and dose-dependent manner. Here we propose a role for FN-EDA in retinal fibrosis.

Methods:: Immunohistochemistry and imunofluorescence microscopy were performed on frozen sections of surgically excised epiretinal membranes from six patients under treatment for PVR using a monoclonal antibody against FN-EDA and a polyclonal antibody against TGFß2. Immunohistochemistry was also performed on frozen sections of normal human retinas from six donors. Human fetal RPE cell cultures (passages 2-4) were treated with TGFß2 or cellular FN for 48 hours. Collagen content was determined using Human Type I Collagen Detection Kit. Culture samples were harvested and processed according to the ELISA kit protocol.

Results:: In normal human retinas, strong immunostaining for FN-EDA was detected along the walls of choroidal and retinal vessels. Staining was similar in all sections analyzed and absent in all retinal tissue except for the positive blood vessels. FN-EDA was abundantly expressed in all PVR membranes predominately within the extracellular matrix and cellular regions. Double staining of FN-EDA with TGFß2 showed strong co-localization primarily in cellular regions within the cytoplasm. In RPE cultures, cellular FN induced type I collagen expression, as did TGFß2, in a dose-dependent manner.

Conclusions:: These findings suggest that FN-EDA is an important factor in retinal fibrosis, making it a potential target for therapy because of its relative specific expression in tissue response to injury.

Keywords: extracellular matrix • proliferative vitreoretinopathy • retinal pigment epithelium