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 (HRPE) from donor eyes obtained from eye banks. Using human RPE cell cultures as a model, we conducted investigations to examine the various roles of RPE in the pathophysiology of retinal disorders. Human RPE cells secreted significant quantities of interleukin-6 (IL-6), intercellular adhesion molecule -1 (ICAM-1), granulocyte macrophage colony stimulating factor (GMCSF), and nitric oxide (NO) in response to the stimulation by inflammatory mediators such as interferon-gamma (IFN-gamma), interleukin-1 (IL-1), and tumor necrosis factor-alpha (TNF-alpha). Northern blot analyses for the mRNA expression of IL-6, ICAM-1, GMCSF, and nitric oxide synthase (iNOS) confirmed the increased secretion of these substances. Inflammation associated with retinochoroiditis due to ocular toxoplasmosis (OT) is one of the common causes of uveitis. Toxoplasma gondii ( T. gondii), an obligate intracellular protozoan parasite, is the causative agent of OT. In OT, neuro-retina and RPE are the primary targets where cysts and tachyzoites of T. gondii were detected. We have studied the replication of T. gondii in human RPE cells and the regulation of this replication by interferons and cytokines. Interferons alpha, beta, gamma, and TNF-alpha inhibited T. gondii replication in a dose-dependent manner. IFN-gamma is the most potent and can completely block T. gondii replication. Anti-toxoplasmotic activity of interferons and TNF-alpha is independent of NO pathway because they do not induce NO production in these cells. However, the addition of tryptophan to IFN-gamma treated cells significantly reversed the inhibitory effects of IFN-gamma, suggesting that IFN-gamma acts by depleting cellular tryptophan. This observation was confirmed by RT-PCR and Northern blot analysis, which indicated the induction of indoleamine 2,3-dioxygenase, an enzyme that converts tryptophan to kynurenine. This in vitro model of HRPE may be useful in evaluating the effects of cytokines and drugs on T. gondii replication within the retina.
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