Analysis of the Barrier Function of Cultured Corneal Endothelial Cells Isolated from Patients with Fuchs Endothelial Corneal Dystrophy.

Abstract

Purpose: The corneal endothelium forms a leaky barrier allowing the influx and outflux of aqueous humor between the anterior chamber and the stroma. Fuchs endothelial corneal dystrophy (FECD) is characterized by a progressive corneal endothelial decompensation and stromal edema. The goal of this study was to compare the capacity of FECD and healthy corneal endothelial cells to form a functional barrier in vitro, in order to gain insight into the mechanisms underlying the pathology.

Methods: Human corneal endothelial cells (HCEC), isolated from surgical specimens (FECD; n=6) and age-matched healthy Eye bank corneas (healthy; n=6) were cultured 24-38 days to obtain a post-confluent monolayer. Gene profiling analyses were performed using Agilent SurePrint G3 Human Gene expression microarrays. Transendothelial resistance (TER) and permeability to 10 kDa FITC-Dextran were measured in FECD (n=2) and healthy (n=4) HCEC populations cultured on 12 mm 0.4 µm polycarbonate Isopore membranes. Measures of membranes without cells were used as controls. Presence of cadherins (adherens junction marker) and zonula-occluden-1 (ZO-1; tight junction marker) was also determined by immunofluorescence.

Results: No significant differences were observed in the most abundant transcripts of the ZO-1 and cadherin family members (TJP1: +1.14X, CDH2: -1.05X, CDH11: -1.13X). Immunofluorescence analysis did not show any difference of expression between FECD and normal populations for ZO-1 and pan-cadherin. A lower TER was observed in FECD (22.3 ± 3.5 Ωcm²) compared to healthy (28.7 ± 8.0 Ωcm²) HCEC populations (p = 0.50). Higher permeability measurements were obtained for FECD (30.2 ± 5.2% relative to controls) compared to healthy (25.5 ± 3.4%) populations (p = 0.38).

Conclusions: Results show that within the frame of this study, cultured FECD and healthy cells had similar junction-related gene and protein expressions markers. However, the endothelium formed by FECD cells showed a higher permeability and a reduced TER compared to healthy cells. Although these early results did not reach statistical significance because of the small sample size, they suggest that the ability of FECD cells to form a functional barrier is hindered in this disease.<br />