Abstract

P 151

Tight junction protein expression is affected by Trefoil Factor Family Peptide (TFF3) in a corneal in vitro scratch model

Ute Schulze1, Laura Contreras-Ruiz2, Yolanda Diebold2, Friedrich Paulsen1
1Institut für Anatomie und Zellbiologie, Martin-Luther-Universität Halle-Wittenberg, Halle/Saale, 2Ocular Surface Group, IOBA University of Valladolid, Valladolid, Spain

Objective
Adjacent epithelial cells are connected via tight junctions. This tight junction complex consists of transmembrane proteins, like occludin and claudins 1 and 2, as well as the cytoplasmatic plaque proteins zonula occludens ZO-1 and ZO-2. Trefoil Factor Family Peptide 3 (TFF3) is a mucin-associated molecule induced in cornea after lesions and corneal diseases.  It influences the rheology of the tear film and offers protective and antiapoptotic properties and promotes the wound healing. In this present study we tried to evaluate a potential TFF3-induced effect on the expression of these certain tight junction proteins in corneal epithelial cells to understand the TFF3-mediated wound healing promoting role at the ocular surface better.
Methods
The expression of several proteins of the tight junction complex in human SV40- immortalized corneal epithelial cells (Araki-Sasaki HCE cell line) was analyzed by RT-PCR. After stimulation with recombinant human TFF3 (rhTFF3, 0,3mg/ml) for 5min, 1h, 6h and 24h under certain additional conditions (with and without epithelial scratches - as defined simulated wound model) expression levels of the tight junction proteins were studied by real time PCR. Immunofluorescence studies are currently ongoing.
Results
All tight junction proteins (ZO-1, ZO-2, claudin 1, claudin 2 and occludin) could be detected on basal levels in HCE cells. After stimulation with rhTFF3 the transmembrane proteins occludin, claudin 1 and claudin 2 revealed significantly increased mRNA expression levels. Claudin 1 expression is upregulated after 6h rhTTF3 treatment whereas occludin expression level remained increased after 6h and 24h stimulation. Claudin 2 mRNA level is upregulated after 1h up to 24h rhTFF3 treatment. On the contrary, the cytoplasmatic plaque protein ZO-1 showed an upregulated expression after 24h, whereas ZO-2 expression was decreased only after 24h rhTFF3 stimulation. In contrast, after scratching the cells the mRNA expression level of all examined proteins of the tight junction complex is ancillary upregulated after almost all rhTFF3 treatment periods compared to scratched untreated controls.
Conclusions
TFF3 might play a role in regulating corneal barrier function which may also be due to its function in the restitution process.

Copyright: 2009 Porstmann Kongresse GmbH. All rights reserved. Impressum, Disclaimer.