Single-Cell Analysis To Define Protective and Tolerizing Immune Cell Populations in the Human Placenta

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Maternal immune adaptation to pregnancy requires maintenance of immune competence with concurrent specific tolerance of the semi-allogeneic fetus. Immune maladaptation has been associated with several adverse pregnancy outcomes, with both short- and long-term impact on maternal and offspring health. Maternal exposure to fetal-placental antigens during pregnancy is well-established; however, many questions remain about the nature of both antigen presentation and the resultant adaptive immune response. Eliciting a de novo adaptive immune response requires the activation of professional antigen presenting cells (APCs), especially dendritic cells (DCs). Importantly, DCs can also induce T cell tolerance and thus play a critical role in avoiding unwanted immune responses. This has been typically studied in the context of preventing responses against self-antigens and is also relevant for protection of the fetus from maternal rejection. It remains unknown how APCs in the human placenta and decidua provide protection against infection and at the same time prevent immune responses against the fetus. A major roadblock to understanding protection and tolerance in human reproduction is the limited information about the functional profile of these APCs, and their inter-individual variation. We propose to define human APC subsets in maternal blood, placenta, decidua, and umbilical cord blood to test the hypothesis that APCs involved in human reproduction consist of several subsets that are functionally specialized to contribute to either protection or tolerance. To accomplish this, we will take advantage of recent technological advances in the field of single-cell analysis, namely high- dimensional cytometry and single-cell gene expression analysis to dissect the functional heterogeneity within this immune compartment. Our lab has developed and validated multiple 28-color fluorescence cytometry assays, which interrogate cell phenotype, activation, and functional capacity (i.e. expression of exhaustion markers, inhibitory receptors, co-stimulatory molecules). We will employ these assays to determine the composition and capabilities of cells directly ex vivo. These experiments will provide critical insight to define functional specialization of DCs in healthy human reproductive tissues during pregnancy and also provide the basis to study pathologies such as preeclampsia in the future.