Toxoplasma gondii suppress human cord blood cell differentiation to the NK cell population

Abstract

Background: Toxoplasma gondii is an obligate intracellular protozoan parasite that can invade all mammalian cells. It is well established that natural killer (NK) cells have critical protective roles in innate immunity during infections by intracellular pathogens. In the current study, we conducted an in vitro experiment to evaluate NK cell differentiation and activation from human umbilical cord blood mononuclear cells (UCB-MNCs) after infection with T. gondii tachyzoites.

Methods: UCB-MNCs were infected by fresh tachyzoites of type I (RH) or type II (PTG) strains of T. gondii pre-expanded in mesenchymal stem cells for 2 weeks in a medium enriched with stem cell factor, Flt3, IL-2, and IL-15. Flow cytometry analysis and western blot analysis were performed to measure the CD57⁺, CD56⁺, and Granzyme A (GZMA).

Results: Data revealed that incubation of UCB-MNCs with NK cell differentiation medium increased the CD57⁺, CD56⁺, and GZMA. UCB-MNCs cocultured with PTG tachyzoites showed a significant reduction of CD56⁺ and GZMA, but nonsignificant changes, in the levels of CD56⁺ compared to the control UCB-MNCs (p > .05). Noteworthy, 2-week culture of UCB-MNCs with type I (RH) tachyzoites significantly suppressed CD57⁺, CD56⁺, and GZMA, showing reduction of NK cell differentiation from cord blood cells.

Conclusion: Our findings suggest that virulent T. gondii tachyzoites with cytopathic effects inhibit NK cell activation and eliminate innate immune responses during infection, and consequently enable the parasite to continue its survival in the host body.

Keywords: NK cells; Toxoplasma gondii; differentiation; functional activity; maturation; umbilical cord blood mononuclear cells.

Figure 1

Overview of the experimental approach and the main findings of the study. BM, bone marrow; MNC, mononuclear cells; UCB, umbilical cord blood.

Figure 2

Incubation of human BM‐MSCs with Toxoplasma gondii tachyzoites at different ratios (A) 1:10 and (B) 1:100 for 2 weeks (arrow points to BM‐MSCs). By increasing tachyzoites numbers, BM‐MSCs exhibited cytotoxic indicated with the reduction of cell number per cm² and loss of spindle shape morphology. Incubation of human BM‐MSCs with PTG (C) and RH (D) T. gondii tachyzoites for 2 weeks. (E) Control noninfected MSCs. BM‐MSCs, bone marrow mesenchymal stem cells.

Figure 3

CD57⁺ expression was evaluated on infected UCB‐MNCs using flow cytometry analysis after 14 days. In the MNC group, the highest expression of CD57⁺ was achieved (left). The expression of CD57⁺ on UCB‐MNCs exposed to PTG stain was high compared to the group that received the RH strain. The lowest CD57⁺ expression rate was obtained in human BM‐MSCs after exposure to tachyzoites (right). All experiments were performed in triplicate. Significant differences were observed between the MNC group and other groups (*p < .05; **p < .01, ****p < .0001). BM‐MSCs, bone marrow mesenchymal stem cells; UCB‐MNCs, umbilical cord blood mononuclear cells.

Figure 4

Measuring protein levels of CD56 (A) and GZMA (B) in UCB‐MNCs after exposure to RH and PTG Toxoplasma gondii tachyzoites. The results are shown as the mean ± SD of three independent experiments. There was a statistically significant difference in MNC and other groups (*p < .05; **p < .01, ***p < .001, ****p < .0001). GZMA, granzyme A; UCB‐MNCs, umbilical cord blood mononuclear cells.