POSTER 3Controlled recruitment and proliferation of placental specific CD8+ T cells in the pregnant uterus underlie fetal immune tolerance

In physiological pregnancies, the semi-allogenic fetus and placenta do not induce adverse immune responses by the maternal CD8+T cells in the pregnant uterus. Yet, we previously showed that uterine CD8+T cell counts escalate across pregnancy and their shift towards inflammation has detrimental consequences for fetal growth/survival. Here, we aimed to bridge central gaps in knowledge about the homeostasis of the CD8+T cell compartment in the uterus in the context of healthy pregnancies, as a fundamental step to understand their involvement in pregnancy pathology. Blood, spleen, uterine-draining lymph nodes and uterus were harvested on gestation days 6.5, 10.5 and 14.5 from Balb/c mated C57Bl/6 females, and virgin controls. Leucocytes were isolated, quantified and characterized by flow cytometry using supervised and dimension reduction analyses. Furthermore, the Act-mOVA/OT-I system served to investigate the migration, proliferation, and phenotype of adoptively transferred placental-specific OT-I T cells in Rag2-/-gc-/- or C57Bl/6 pregnant females containing OVAexpressing placentas. In all experiments, mice were injected with anti-CD45 antibody prior to tissue collection, for exclusion of intravascular leucocytes. Characterization of CD8+T cells across pregnancy demonstrated particular CD8+T cell dynamics in the uterus, which were largely not replicated in peripheral blood and lymphoid organs. This included signs of immune activation in the uterus following critical pregnancy milestones such as implantation and placental formation with enrichment of effector and memory subsets. Moreover, the massive expansion of uterine CD8+T cell counts across pregnancy involved the recruitment of circulating cells and their local proliferation. In fact, adoptively transferred CD8+T cells homed in the pregnant uterus of congenic immune-deficient and immune-competent females, through a migration pathway that may involve adhesion through CD49d, which was abundantly expressed in uterine CD8+T cells. The observed migration and proliferation were exclusively detectable in an antigen specific manner in mice carrying OVA+ placentas and injected with OT-I cells, but not in pregnancies lacking placental specific CD8+T cells. Among CD8+T cells, those expressing CD122+and presenting a regulatory phenotype were enriched in the uterus in physiological pregnancies. The frequency of CD8+CD122+T cells appear to be under a tight homeostatic control, as in Rag2-/-gc-/-pregnancies, adoptively transferred OT-I CD8+T cells homing the uterus reached a similar CD122+:CD122-ratio as observed in healthy wild-type pregnancies, whereas this ratio was highly variable in blood and spleen. Of note, uterine CD8+CD122+ T cells responded more promptly to pregnancy events compared to their CD122- counterparts. Adoptive transfer of CD8+CD122+T cells significantly enhanced pregnancy performance. The upregulation of inhibitory receptors such as PD1 or members of the Ly49 family of receptors in CD8+CD122+ T cells in the uterus but not in other analysed organs suggests tissue-specific immune regulatory traits. Placental antigen-specific CD8+T cells are prone to seed the uterus during pregnancy, where they can further proliferate without posing a threat for pregnancy success. This is at least partly due to the homeostatic crosstalk with CD8+CD122+T cells exhibiting immune regulatory/suppressive tissue-specific features. Further research is required to mechanistically understand this crosstalk in health but also in the context of intrauterine inflammation and pregnancy pathology