Elevation of KIR+CD8+ T cells during human pregnancy

During pregnancy, immune responses must balance fetal protection from infections with tolerance of the semi-allogeneic fetus. However, the mechanisms regulating maternal-fetal tolerance remain poorly understood. Recently, we identified KIR+CD8+ T cells as a previously underappreciated regulatory subset important for suppressing self-reactivity in human autoimmune and infectious diseases. To better understand what other roles these cells might play, we asked whether they are active during pregnancy. We first observed an increased frequency of KIR+CD8+ T cells in the peripheral blood of pregnant women at the second trimester, especially in those carrying a male fetus. In vitro, KIR+CD8+ T cells inhibited the alloreactive responses of maternal T cells against irradiated cord blood cells and suppressed HY-specific CD8+ T cells from mothers with a male fetus. Therefore, the higher induction of KIR+CD8+ T cells in mothers carrying a male fetus may help suppress additional allogenic responses triggered by Y chromosome antigens. Longitudinal analysis showed that KIR+CD8+ T cells undergo expansion and differentiate into functional cytotoxic cells during pregnancy. Single cell RNA-seq analysis of decidual CD8+ T cells from early pregnancy revealed elevated number and activity of KIR+ CD8+ T cells at the maternal-fetal interface. In addition, increased levels of KIR+CD8+ T cells correlated with pregnancy disorders (e.g., spontaneous abortion and preeclampsia). Taken together, our findings suggest an important role of KIR+CD8+ T cells in the maintenance of maternal tolerance by suppressing fetal-specific alloreactive T cells. They may also be useful as predictive biomarkers or drug targets for human pregnancy disorders. Overall design: PBMCs from pregnant women collected at different time points throughout pregnancy or in the postpartum period were labeled with unique tags using a BD™ Hu Single Cell Sample Multiplexing Kit (no. 633781) for 20 min at room temperature. Cells were then washed three times with PBS, after which the barcoded PBMCs were pooled and stained with viability dye and flow antibodies. Live KIR+CD8+ T cells were sorted into sample buffer and immediately loaded onto the BD Rhapsody Cartridge using the BD Rhapsody Cartridge Reagent Kit (no. 633731) per manufacturer's instructions. Single-cell cDNA was prepared using the BD Rhapsody cDNA Kit (no. 633773). This was followed by the preparation of single-cell mRNA, TCR, and tag libraries using the BD Rhapsody™ TCR/BCR Multiomic Assay for Targeted Kit (catalog no. 666622), along with primers from the BD Rhapsody Immune Response Panel (399 genes; catalog no. 633750) and custom primers. Sequencing was carried out using NovaSeq (Illumina, San Diego, CA) in Stanford Genomics Sequencing Center and Novogene.