Molecular and cellular evidence for T-cell stimulation by allogeneic retinal pigment epithelium cells in vitro

Purpose: The rejection of retinal pigment epithelium (RPE) allografts is a major barrier to longterm success after retinal transplantation. The aim of this study was to investigate the action of RPE cells on allogeneic T cell s in coculture with or without macrophages. For the detection of T-cell act ivation the interleukins IL-1 beta and IL-2, typical for this process, were investigated. Methods: Human RPE cells (6x10(5) cells/flask) were used as stimulator cells. To investigate the influence of MHC class II molecules th e RPE cells were pre-incubated with different concentrations of interferon- gamma (IFN-gamma; 0, 50, 100, or 250 U/ml) for 4 days. This was followed by coculture with either 6 x 10(6) T cells or, in a second trial, the T cells plus 6x10(5) macrophages. The mRNAs of the cytokines under study were dete cted using a reverse-transcriptase polymerase chain reaction and were quant ified by colorimetry after 6 h. The cytokine protein content in the superna tants was measured after 20 h using specific enzyme-linked immunoabsorbent assays. Results: Cytokine-specific mRNAs and proteins were found in all sam ples. After coculture the level of IL-1 beta mRNA was higher and that of cy tokine-specific protein was significantly increased. Furthermore, the addit ion of macrophages led to increased cytokine secretion but a general influe nce of the pre-activation with interferon could not be found. Similar resul ts were detected for IL-2; at the highest dose, IFN-gamma preactivation and , in combination with macrophages, a significant increase in the protein le vel could be found. Conclusion: These results show that RPE cells are able to activate allogeneic T cells in vitro. Professional antigen-presenting ce lls may promote this process, as may pre-treatment with IFN-gamma. The circ umstances modelled here are involved in the rejection process after RPE tra nsplantation in humans and help to explain this immune response.