Electroporating Human Corneal Epithelial Cells With Interleukin 10 and Fas Ligand pDNA
Nadia Zakaria1, Nathalie Cools, Zwi Berneman
1From the *Department of Ophthalmology and †Center for Cell Therapy and Regenerative Medicine, Antwerp University Hospital; and ‡Laboratory of Experimental Haematology, Vaccine and Infectious Disease Institute (Vaxinfectio), University of Antwerp, Edegem, Belgium.
Purpose:
To develop an optimal electroporation protocol for plasmid (pDNA) transfection of a human corneal epithelial cell (hCEC) line and investigate the immunomodulatory capacity of interleukin 10 and Fas ligand (FasL) transfection on hCECs.
Design:
A controlled experimental study.
Methods:
Human corneal epithelial cells were electroporated with pDNA encoding enhanced green fluorescent protein, interleukin 10, or FasL. Supernatants were analyzed for cytokine secretion using enzyme-linked immunosorbent assay. To test potential immunosuppression, electroporated hCECs were cocultured with allogeneic peripheral blood mononuclear cells, and the supernatants analyzed for interferon γ production.
Results:
Maximum transfection efficiencies were obtained using optimized settings, and transgene expression was detected up to 13 days following transfection. Interleukin 10 levels peaked at day 4 and FasL at day 2 following electroporation. Coculture supernatants showed significantly lower levels of interferon γ in the modulated groups compared with control.
Conclusions:
Our results demonstrate highly efficient transfection of hCECs using an optimized electroporation protocol. Interleukin 10 and FasL may provide a means of immune modulation of corneal epithelial cells.
