The retinal pigment epithelium (RPE plays a critical role in the regulation of retinal and choroidal function in normal and disease states. Due to limited availability of human tissues, an in vitro cell culture system is desired. Therefore we have developed and characterized the primary cell lines of human RPE from donor eyes obtained from eye banks. Using human RPE cell cultures as a model, we conducted investigations to examine the various roles of R?E in the pathophysiology of retinal disorders. Human RPE cultures exposed to bacterial lipopolysaccharide (LPS), tumor necrosis factor alpha (TNF-alpha), and interleukins 1 alpha and 1 beta (IL-1alpha, IL-1beta) secreted large amounts of interleukin 6 (IL-6). Immunoblot and Northern blot analysis confirmed the presence of posttranslationally modified IL-6 protein and mRNA levels, respectively. Interferon-gamma (IFN-gamma) acted synergistically with other mediators to stimulate 3- to 5-fold increases in IL-6 secretion. We extended our studies to examine the expression and secretion of intercellular adhesion molecule-1 (ICAM-1), a cell surface ligand for lymphocyte function- associated antigen-1 (LFA-1) expressed during inflammatory reactions, by human RPE. IFN-gamma, TNF-alpha, and IL-1 increased significantly both cell surface expression (detected by immunofluorescence staining) and secretion into the medium (detected by ELISA). Secretion (shedding) of ICAM-1 by RPE cells in the presence of a mixture of F?-gamma (100 mu/ml), TNF-alpha (1 ng/ml), and IL-1 (1 ng/ml) was cumulative, suggesting that combining these cytokines results in potent inflammatory reactions. The response of RPE cells to inflammatory mediators was rapid and sustained in the presence of stimulants but reversed to control levels quickly upon withdrawal, suggesting the reversibility of the responses of RPE to inflammatory signals. The effects of transforming growth factor beta (TGF-beta) on RPE functions at cellular and molecular levels are also being studied. TGF-beta increased the expression of heme oxygenase-1, an enzyme reaction that generates the antioxidant bilirubin, which helps in cellular defense mechanisms against oxidative stress. The results clearly shows that human RPE cells respond to specific inflammatory signals or infections by increased cellular expression and secretion of IL-6 and ICAM-1, which may in turn perpetuate immune reactions in the pathogenesis and/or prevention of retinal and choroidal diseases.
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