The Mechanism Of LILRB4 In Abnormal Pregnancy Outcomes Induced By T.gondii Infection

Aims: In this study, we want to investigate the mechanism of LILRB4 expressed on uterine dendritic cells (uDCs) in normal pregnancy and in abnormal pregnancy outcomes following by Toxoplasma gondii infection.Materials: Normal pregnant mice and LILRB4-/- pregnant mice were established with a C57BL/6 genetic background respectively and observed the pregnancy outcomes.The expression of LILRB4, MHC class Ⅱ molecules, CD80 and CD86 expressed on uterine DCs were analyzed by flow cytometry. The levels of TGF-β, IL-10, IL-12,TNF-α as well as IFN-γ in plancenta supernatant were detected by ELISA. The mice model with T. gondii infection during early pregnancy in vivo were established and divided into control group, infection group and LILRB4-/- infection group. The infected group and LILRB4-/- infected group were infected with 400 T. gondii tachyzoite on gestational day (GD) 8. The mice in each group were sacrificed on gestational day 14. Single-cell suspensions were prepared from placenta and uterus.The surface expression of LILRB4; and presence of MHC Ⅱ, CD80/CD86, which are functional molecules of uDCs were analyzed by flow cytometry. Purified uDCs from human first-trimester decidual tissues were divided into control group, infection group and LILRB4-neutralized infection group. Both infection groups were co-cultured with T. gondii tachyzoites a ratio of 1:1. The expression of CD80/CD86 and HLA-DR were analyzed in three DC subsets respectively by flow cytometry.Results: The expression of LILRB4 was detected on CD11c+uterine DCs at GD 5,reached its peak on GD 14, and quickly declined by GD 16. Moreover, the expression level of LILRB4 on uDCs was very low in non-pregnant mice. Higher levels of MHC-II, CD80 and CD86 were found in LILRB4-/- pregnant mice compared to WT mice. In addition, the level of TGF-β was decreased, while IL-12, TNF-a as well as IFN-y were increased in LILRB4-/- pregnant mice compared to WT mice. In animal models, T. gondii infection could lead to the pregnancy failure in both WT and LILRB4-/- mice during the first trimester of pregnancy. More importantly, in contrast to fetuses from infected WT mice, which exhibited a reduction in body weight, the fetuses from infected LILRB4-/- mice display significant weight loss compared to WT-infected mice. The relative of CD11c+CD8α-uDCs was higher when compared with CD8a+ uDCs and the expression of LILRB4 was obviously higher on CD11c+CD8α- uDCs subset than that on CD11c+CD8α+ uDCs subset in WT pregnant group. The level of LILRB4 expressed on both two subsets of uDCs was down-regulated in the WT infected group when compared with WT normal group,whereas levels of effector molecules MHC Ⅱ, CD80 and CD86 were significantly increased. Additionally, LILRB41/- infected group have robust co-stimulatory molecules production compared to the WT infected group. Locally in human early pregnancy decidual dendritic cells, level of LILRB4 was expressed on the three DCs subsets. The expression of LILRB4 reduced significantly in infection group compared with the control group, whereas levels of effector molecules such as CD80/CD86 and HLA-DR were significantly increased in LILRB4-neutralized infection group compared with control and single infected group.Conclusion: 1. The inhibitory receptor of LILRB4 on decidual dendritic cells at the feto-maternal interface may play an important role in nomal pregnancy. 2. T.gondii infection induces a decline in the expression of LILRB4 on DCs that initiate of immune responses may contribute abnormal pregnancy outcomes during T. gondii infection.