Use of a Rabbit Cornea Model for the Development of a Cell Transfer System for Limbal Epithelial Cells

Abstract

 

Purpose:

 

The aim of this work is to develop a contact lens system for transferring laboratory expanded limbal epithelial cells for treatment of the cornea.

 

Methods:

 

The approach taken is to use a chemically defined engineered surface using plasma polymerisation technology to develop a coating which supports both the initial attachment of epithelial cells and their subsequent transfer onto the denuded cornea. To assist in this development we have established a rabbit organ culture model to examine transfer of cells on to the cornea. Initial studies identified the most appropriate plasma coating for support of epithelial cells. We are also examining the contribution of stromal cells to the survival of epithelial cells under serum-free conditions in this model. First we examined the culture of a human corneal epithelial cell line (HCEC) and primary rabbit limbal epithelial cells on a range of plasma coatings. Acrylic acid, allyl amine and allyl alcohol surfaces were synthesised at different power and flow rates and their surface chemistry examined by XPS analysis. From these, the surface which best supported epithelial cell culture (both human and rabbit cells) was identified. Cells were cultured on contact lenses coated with this surface. Rabbit corneal organ cultures with the intact epithelium were then denuded of epithelial cells (Fig. 1A & B). Lenses with cells were placed onto the cornea and kept in place for 4 days. Transferof cells from lenses was examined by pre-staining cells with CellTracker ^TMRed CMTPX and also by subsequent staining of cells on the cornea with DAPI and phalloidin FITC.

 

Results:

 

The surface that best supported the epithelial cell culture was acrylic acid. Preliminary results using this model show that the primary rabbit limbal epithelial cells and the human cell line will transfer from the lens onto the cornea (Fig. 1C & D).

 

Conclusions:

 

This model can be used to develop a culture and transfer protocol which we hope to use for future clinical studies.  

 

Keywords: cornea: epithelium