Electroporating human corneal epithelial cells with interleukin 10 and Fas ligand pDNA
Faculty of Medicine and Health Sciences
Asia-Pacific journal of ophthalmology / National University Hospital of Singapore. Department of Ophthalmology. - Singapore
, p. 56-63
University of Antwerp
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.