Toxoplasma gondii infection of neurons induces neuronal cytokine and chemokine production, but gamma interferon- and tumor necrosis factor-stimulated neurons fail to inhibit the invasion and growth of T. gondii

Abstract

The intracellular parasite Toxoplasma gondii has the capacity to persist in the brain within neurons. In this study we demonstrated that T. gondii infected murine cerebellar neurons in vitro and replicated within these cells. Stimulation with gamma interferon (IFN-gamma) and/or tumor necrosis factor (TNF) did not enable neurons to inhibit parasite invasion and replication. Cultured neurons constitutively produced interleukin 1 (IL-1), IL-6, macrophage inflammatory protein 1alpha (MIP-1alpha), and MIP-1beta but not transforming growth factor beta1 (TGF-beta1), IL-10, and granulocyte-macrophage colony-stimulating factor. Neuronal expression of some cytokines (IL-6, TGF-beta1) and chemokines (MIP-1beta) was regulated by infection and/or by IFN-gamma and TNF.

FIG. 1

Cultured cerebellar neurons after infection with T. gondii RH (MOI, 1) for 48 h: PV containing numerous tachyzoites in the cytoplasm of a neuron with a prominent nucleolus. Note the intimate contact of the PV with the nuclear membrane. Anti-T. gondii immunostaining and slight counterstaining with hemalum were used. Magnification, ×625.

FIG. 2

Proliferation of T. gondii in neurons 24 h (A) and 48 h (B) after infection. After 7 days of cultivation neurons were treated with the cytokines indicated for 12 h. After this, RH toxoplasms were added at an MOI of 1. At 24 and 48 h after infection neurons were fixed with 4% paraformaldehyde, and T. gondii was stained immunohistochemically. The number of toxoplasms per PV was determined microscopically for 100 infected neurons per group; the data are means ± standard deviations based on three wells per group. Similar data were obtained in two repeat experiments.

FIG. 3

Cytokine and chemokine production by neurons. Neurons were treated as indicated with IFN-γ and/or TNF for 12 h before RH toxoplasms were added at an MOI of 1. At 48 h after infection the supernatant was harvested, and IL-1β, IL-6, IL-10, GM-CSF, TGF-β1, TGF-β2, MIP-1α, and MIP-1β contents were determined by enzyme-linked immunosorbent assays. IL-10 and GM-CSF were not detected (data not shown). The data are means ± standard deviations based on three wells per group. A second experiment yielded comparable results.