Changes Of Decidual NK Cells Infected By T.GONDII:Implications For Abnormal Pregnancy

AIMs:To investigate the changes of subsets, receptors and associated factors in human decidual natural killer (dNK) cells with Toxoplasma gondii (T. gondii) infection in vitro and in vivo:implications for abnormal pregnancy outcomes.METHODS:Human dNK cells co-cultured with Trophoblast cells and then infected with T. gondii. The CD5brightCD6-dNK subset, CD56dimCD16+dNK subset and CD56brightCD16-/CD56dimCD16+dNK ratio were detected by flow cytometry. Infected pregnant mice were established. CD122+CD49b+NK1.1+NK subset and CD122+CD49b-NK1.1-NK subset were measured in placenta by flow cytometry; Human dNK cells were isolated and infected with T. gondii. Flow cytometry were used to measure levels of activating receptor NKG2D, inhibitory receptors KIR2DL4、ILT-2and the cytotoxicity to trophoblast cells; Human dNK cells co-cultured with Trophoblast cells and then infected with T. gondii, expression of inhibitory receptors KIR2DL4, ILT-2and its ligand HLA-G, activating receptor NKG2D were detected by flow cytometry and Real-time PCR. NKG2A, Qa-1and NKG2D in placenta of infected pregnant mice were also analyzed by flow cytometry and Real-time PCR; Human dNK cells co-cultured with Trophoblast cells and then infected with T. gondii. Real-tme PCR, flow cytometry, confocal microscopy, Western blot and ELISA were used to detect apoptosis of cells, caspase3, caspase8, and levels of IFN-y and sHAL-G respectively.RESULTS:The CD56brightCD16-/CD56dimCD16+dNK ratio was significantly decreased at12h,24h, and48h following YFP-T. gondii infection. CD56brightCD16-/CD56dimCD16+ratio was negatively correlated with dNK cytotoxic activity. In pregnanct mice, CD122+CD49b+NK1.1+dNK subset was increased, CD122+CD49b-NK1.1-dNK subset was decreased while CD122+CD49b+NK1.1+/CD122+CD49b-NK1.1-dNK ratio was significantly increased. CD56brightCD16-/CD56dimCD16+dNK ratio was correlated with dNK cytotoxic activity (r2=0.753, P<0.05); Expression of KIR2DL4, ILT-2, and NKG2D were increased after infection, but NKG2D were significantly higher than those of KIR2DL4and ILT-2. Further, HLA-G, which was the ligand of KIR2DL4and ILT-2, was up-regulated; the cytotoxic activity of dNK cells infected with YFP-T. gondii increased with the time after infection and was significantly higher than that of the uninfected cells. In pregnant mice infected with T. gondi, levels of NKG2A、Qa-1and NKG2D were up-regulated, meanwhile level of NKG2D was significantly higher than that of NKG2A. NKG2D expression was correlated with dNK cytotoxic activity (r2=0.701, P<0.05); T. gondii infection induced up-regulation of IFN-y secreated by dNK cells and sHAL-G secreated by trophoblast cells may promote trophoblast cells and dNK cells apoptosis respectively.CONCLUSIONS:Enhanced dNK cytotoxicity due to increased human CD56dimCD16+dNK subset and CD122+CD49b+NK1.1+dNK subset in mice may contribute to abnormal pregnancy outcomes observed upon maternal infection with T. gondii; Imbalances in the inhibitory signals and activating signals that regulate cytotoxic and cytokine production activities of dNK subsets likely lead to the failure of pregnancy or abnormal fetal development in mothers infected with T. gondii during the early pregnancy. Our results thus shed light on the molecular immune mechanism underlying abnormal pregnancy caused by T. gondii infection; The increased apoptosis of trophoblast and dNK cells induced by the changes of IFN-y and sHLA-G depended on caspase pathway may contribute to the abnormal pregnancy outcome with T. gondii infection.