Mechanisms of interferon-induced inhibition of Toxoplasma gondii replication in human retinal pigment epithelial cells

Inflammation associated with retinochoroiditis is a major complication of ocular toxoplasmosis in infants and immunocompetent individuals. Moreover, Toxoplasma gondii-induced retinal disease causes serious complications in patients with AIDS and transplant patients. The retinal pigment epithelial (RPE) cell is an important regulatory cell within the retina and is one of the cells infected with T. gondii in in vivo. We have developed a human RPE (HRPE) cell in vitro model system to evaluate T. gondii replication and the regulation of this replication by cytokines. T. gondii replication was quantitated by counting the foci of infection (plaque formation) and the numbers of tachyzoites released into the supernatant fluids. Pretreatment of cultures with recombinant human tumor necrosis factor alpha, alpha interferon (IFN-alpha), IFN-beta, or IFN-gamma for 24 h prior to inoculation inhibited T. gondii replication in a dose-dependent manner. Of these cytokines, IFN-gamma was the most potent, and T. gondii replication was completely inhibited at a concentration of 100 U/ml. The anti-toxoplasmotic activity of IFN-gamma was significantly blocked by monoclonal antibody to IFN-gamma. Treatment of the cultures with IFN-gamma from day 1 or 2 postinoculation with T. gondii also offered protection against the parasite. The anti toxoplasmotic activity of tumor necrosis factor alpha or IFN-alpha, -beta, or -gamma in these cultures was found to be independent of the nitric oxide (NO) pathway, since NO production was not found in HRPE cells treated with these cytokines. However, 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 effect was further confirmed by reverse transcription-PCR and Northern (RNA) blot analysis, which indicated induction of indoleamine 2,3-dioxygenase (IDO), an enzyme that converts tryptophan to kynurenine. These results indicated that interferons inhibited T. gondii replication in HRPE by NO-independent but IDO-dependent mechanisms. This in vitro model of T. gondii replication in HRPE may be useful in evaluating the effects of cytokines and drugs on T. gondii replication within the retina.