Toxoplasma gondii Infection Induces Gene Expression and Secretion of Interleukin 1 (IL-1), IL-6, Granulocyte-Macrophage Colony-Stimulating Factor, and Intercellular Adhesion Molecule 1 by Human Retinal Pigment Epithelial Cells

Abstract

We have used human retinal pigment epithelial (HRPE) cultures to investigate the primary cellular responses of retinal resident cells to intracellular Toxoplasma gondii replication. At 4 days postinoculation, when all of the cells were infected, the secretion of interleukin 1␤ (IL-1␤), IL-6, granulocyte-macrophage colonystimulating factor (GM-CSF), and intercellular adhesion molecule 1 (ICAM-1) was augmented by 23-, 10-, 8-, and 5-fold, respectively, over the control. Northern and reverse transcriptase PCR analyses showed significant upregulation of steady-state levels of mRNA for IL-1␤, IL-6, GM-CSF, and ICAM-1. The secretion of these molecules by HRPE cells may play a critical immunoregulatory role in the pathophysiological processes associated with T. gondii-induced retinochoroiditis.Retinochoroiditis caused by Toxoplasma gondii during ocular toxoplasmosis results in inflammation and disorganization of the retina, occasionally leading to severe loss of vision (11,20,23,30). The intensity of damage to the retina and choroid depends on the severity of the infection and the associated inflammatory reaction (11,13,20). Inflammatory cells, predominantly macrophages and lymphocytes, infiltrate the retina, the subretinal space, and the vitreous (6,11,20). In the severe form of T. gondii-induced uveitis, destruction of large segments of the outer retina and pigment epithelium is observed (6, 9, 13, 24). Within the retina, lysosomal and other autolytic enzymes released by inflammatory cells are thought to contribute to the pathogenic mechanisms of retinal tissue damage (6, 11).Retinal pigment epithelium (RPE), an integral part of the neuroretina in the posterior pole of the eye, acts as a barrier between the highly vascularized choroid and the retina with a complex architecture of neuronal cells (2, 31). In addition to its role in the transport of metabolites between retina and choroid, the RPE phagocytose the shed outer segments of retinal rods and cones (2, 31). Many of these activities of the RPE are essential for the structural and functional integrity of the retina and choroid. The RPE, because of its critical location and physiological activities, is constantly subjected to contact with various infectious agents and inflammatory mediators (12,14).Tachyzoites of T. gondii injected into the peritonial cavities of mice were thought to reach the retina via both choroidal and retinal circulation (8,21,26). In a rabbit model, injection of tachyzoites into the suprachoroidal space resulted in outer retinal lesions and localized foci of retinal pigment epitheliosis within 48 h. This suggests the crossing of the parasite through the RPE-Bruchs membrane barrier from the choroid to the retina (27). Histopathological examination of the eyes of patients with toxoplasma-induced retinochoroiditis revealed the presence of free tachyzoites and cysts in the RPE and the retina (9,19,24). Hence, studies of the mechanisms of T. gondii replication within retinal cells and the responses of the host cells to parasite invasion would be ...

Transforming growth factor-β expression in human retinal pigment epithelial cells is enhanced by Toxoplasma gondii: a possible role in the immunopathogenesis of retinochoroiditis

“…HRPE cultures at passages 6-12 were used for the experiments reported in this study. HRPE cultures were grown to confluence in 24-well tissue culture plates in MEM containing 10% FBS and other components as described [16,24]. Tachyzoites of T. gondii (RH strain) were grown in HRPE cultures and prepared for inoculation as reported previously [23,24].…”

“…HRPE cultures were grown to confluence in 24-well tissue culture plates in MEM containing 10% FBS and other components as described [16,24]. Tachyzoites of T. gondii (RH strain) were grown in HRPE cultures and prepared for inoculation as reported previously [23,24]. The cultures were washed with serum free medium (SFM) and incubated in SFM overnight before inoculating with tachyzoites of T. gondii, suspended in SFM (0·5 ml/well), at a multiplicity of infection (moi) of 5.…”

“…Tachyzoites of T. gondii (RH strain) were harvested from infected HRPE cells as described earlier [23,24]. Parasites were suspended in PBS at 2 ¥ 10 8 /ml, and lysed by freeze-thaw three times on dry-ice followed by sonication for three cycles of 30 s duration.…”

“…Total cellular RNA from the control and T. gondii-infected cultures was prepared by using RNASTAT-60 solution [24]. A RNA PCR kit (Perkin Elmer, Applied Biosystems Division, Foster City, CA, USA) was used according to the manufacturer's instructions.…”

“…Histopathological examination of the eyes of patients with toxoplasma-induced retinochoroiditis revealed the presence of free tachyzoites and tissue cysts in the RPE and the neuroretina [20][21][22]. In our previous studies, we have characterized the replication of T. gondii in human RPE cultures [23] and the secretion of inflammatory molecules upon T. gondii infection [24]. In this report, we have identified retinal host cell-parasite interactions in which T. gondii infection stimulates and alters the active form of TGF-b, while administration of TGF-b augments parasite replication.…”