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A novel mechanism of ultraviolet A and riboflavin mediated corneal cross-linking via induction of tissue transglutaminases

Poster Details

First Author: N.Kopsachilis UK

Co Author(s):    M. Tsatsos   D. Parmar   U. Welge Luessen        

Abstract Details



Purpose:

Collagen crosslinking using ultraviolet- A (UVA) -irradiation combined with the photosensitizer riboflavin is a new technique for treating progressive keratoconus. The purpose of this study is to examine whether primary human corneal keratocytes (HCK) are capable of expressing and secreting fibronectin and tissue transglutaminase (tTgase), an enzyme cross-linking extracellular matrix (ECM) proteins, and whether fibronectin and tTgase are increased after treatment of HCK cells with ultraviolet- A (UVA) -irradiation combined with riboflavin (RFUVA), thus providing another possible physiological mechanism of the cross-linking pathway.

Setting:

University of Erlangen Nuremberg and Moorfields Eye Hospital

Methods:

Cell cultures established from human keratocytes were treated with 0.025% riboflavin solution and UVA (370 nm)-irradiance 3 mW/cm2 for 30 min. Induction of fibronectin and tTgase was investigated by immunohistochemistry, Real-Time PCR and Western blot analysis. Cell viability was quantified by a microscopic live - dead assay. External tTgase activity was measured by the ability to form polymerized fibronectin and the incorporation of biotinylated cadaverine into fibronectin.

Results:

Treatment of cultured HCK cells with RFUVA increased the fibronectin and tTgase mRNA and protein levels. This effect was not observed in cells treated with riboflavin or UVA-radiation alone. Incorporation of biotinylated cadaverine was significantly increased when HCK cells were treated with RFUVA.

Conclusions:

The enzymes tTgase and fibronectin are expressed by RFUVA treatment in cultured HCK cells. This mechanism provides more information about the physiology of corneal crosslinking. FINANCIAL INTEREST: NONE

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