Primary cultures of rat retina as a cell model for studying Toxoplasma gondii interconversion

Abstract

Toxoplasma gondii infection is an important cause of chorioretinitis, that may lead to blindness and glaucoma. The particular tropism of the parasite for retinal tissue suggests that retinal cell cultures, reproducing retinal microenvironment, may represent a suitable cell model for obtaining intracellular replication of the parasite and its stage conversion from tachyzoites to bradyzoites. To verify this hypothesis, we set up primary cultures of embryonic retina, obtained from Wistar rats at the 18th gestational day. Eyes were removed under sterile conditions and the retina was isolated under a stereomicroscope. After dissociation in trypsin, cells were seeded at high density (1−2 × 106 cells) onto poly‐l‐lysine‐coated 24‐well cell culture plates and grown in MEM with 10% foetal calf serum. Cultures were maintained in a humidified, 37°C incubator with 5% CO2. This culturing procedure yields a mixed cell population, where all the principal cell types of the retina are present, including neuronal cells (amacrine cells, photoreceptors, ganglionar cells), macroglial cells (Muller cells, astrocytes) and microglia. One week after T. gondii ME49 strain inoculation, cyst‐like structures were observed, firstly, enclosed in grape‐like clusters of round cells, then free in the medium. In these cysts, bradizoites were still infective up to 2 months, and they developed to the tachyzoite stage when re‐inoculated in a fibroblast monolayer. These results suggest that the microenvironment of retinal cell cultures may favour parasite interconversion from tachyzoite to bradyzoite. Studies are in course to monitor cell‐signallings (INF‐γ, TNF‐α and nitric oxide) that accompanies this phenomenon. This in vitro model may be a valuable tool to study the role of inflammatory process in the pathogenesis of ocular toxoplasmic infection and to test new drugs to prevent ocular toxoplasmosis.