BeWo trophoblast cell susceptibility to Toxoplasma gondii is increased by interferon-gamma, interleukin-10 and transforming growth factor-beta1

Abstract

The present study aimed to investigate BeWo trophoblast cell susceptibility to Toxoplasma gondii infection under stimulation with anti-inflammatory cytokines in comparison with HeLa cells. Both cell types were submitted to different treatments with recombinant cytokines [interleukin (IL)-10 and transforming growth factor (TGF)-beta1] or the respective antibodies (anti-IL-10 and anti-TGF-beta) before and after T. gondii infection. The effect of interferon (IFN)-gamma was also assessed alone or in combination with anti-inflammatory cytokines or the respective antibodies after the parasite infection. Cells were fixed, stained and parasites quantified under light microscopy to evaluate intracellular replication (mean number of parasites per cell in 100 infected cells) and infection index (percentage of infected cells per 100 examined cells). In contrast with HeLa cells, treatments with IL-10 or TGF-beta1 induced a considerable augmentation in both T. gondii intracellular replication and invasion into BeWo cells. In addition, treatment with IFN-gamma alone or associated with IL-10 or TGF-beta1 increased the same parameters in BeWo cells, whereas the opposite effect was observed in HeLa cells. When endogenous IL-10 or TGF-beta was blocked, both BeWo and HeLa cells were able to control the parasite infection only in the presence of IFN-gamma. Together, these results indicate that the higher susceptibility of BeWo cells to T. gondii may be due to immunomodulation mechanisms, suggesting that the role of trophoblast cells in maintaining a placental microenvironment favourable to pregnancy may facilitate the infection into the placental tissues.

Fig. 1

Photomicrograph of BeWo (a and b) and HeLa (c and d) cells after 24 h of Toxoplasma gondii infection. Arrows indicate parasites inside the parasitophorous vacuoles. Toluidine blue staining; 450×. Bars: 22 μm.

Fig. 2

Levels of interleukin (IL)-10 (a) and transforming growth factor (TGF)-β1 (b) in supernatants from BeWo and HeLa cells uninfected or infected by Toxoplasma gondii. The median, 25th and 75th percentiles and minimum and maximum values are shown. Levels of IL-10 were higher in supernatants from infected rather than uninfected BeWo cells. Significant differences were considered when P < 0·05.

Fig. 3

Effect of treatment with anti-inflammatory cytokines interleukin (IL)-10 or transforming growth factor (TGF)-β1 associated or not with interferon (IFN)-γ on Toxoplasma gondii intracellular replication (number of parasites per cell) and index of infection (%) in BeWo (a and b) and HeLa (c and d) cells after 24 h of infection. A significant increase in both infection parameters was observed in BeWo but not HeLa cells. (*) Significant differences in comparison with medium (P < 0·05).

Fig. 4

Effect of treatment with anti-cytokine antibodies [anti-interleukin (IL)-10 or anti-transforming growth factor (TGF)-β] associated or not with interferon (IFN)-γ on Toxoplasma gondii intracellular replication (number of parasites per cell) and index of infection (%) in BeWo (a and b) and HeLa (c and d) cells after 24 h of infection. A significant decrease in both infection parameters was observed in BeWo cells only when antibodies were associated with IFN-γ. However, IFN-γ alone is enough to HeLa cells decrease both infection parameters. Significant differences in comparison with medium (*) or IFN-γ (#) were considered when P < 0·05.

Fig. 5

Effect of treatment with isotype controls (M30 CytoDeath: mouse monoclonal IgG2b or H-240: rabbit polyclonal IgG) associated or not with interferon (IFN)-γ on Toxoplasma gondii intracellular replication (number of parasites per cell) and index of infection (%) in BeWo (a and b) and HeLa (c and d) cells after 24 h of infection. No significant results in both infection parameters were observed in BeWo cells treated with clone M30 or H-240 but, in the presence of IFN-γ, this cytokine increased significantly both infection parameters. However, IFN-γ alone is enough to HeLa cells decrease both infection parameters. Significant differences in comparison with medium (*) were considered when P < 0·05.