Divide-and-conquer analysis reveals hidden immune cell influencers across the placenta in preeclampsia.

BACKGROUND: Placenta is a focal point of cellular interactions of maternal and fetal immune systems, fostering immune tolerance essential for a successful pregnancy. However, the mechanisms underlying immune dysregulation in disorders such as preeclampsia remain poorly understood. METHODS: We constructed a multilayered atlas of the human placenta by performing single-cell RNA sequencing across distinct placental layers from both normal and severe preeclamptic pregnancies. Rather than focusing on specific regions of placenta, such as the villi and decidua, we explore placental architecture by collecting pair-matched tissues from individual pregnancies to suggest appropriate placental-wide immune atlas. To interpret intercellular communication in those unexplored placental layers, we designed two conceptual models: one based on immune interaction frequency (IIF) and another on immune tolerance influencers (IT). We applied machine learning classifiers to identify gene signatures associated with preeclampsia. RESULTS: We observed extensive admixture of semi-allogeneic fetal and maternal cells across all placental layers, regardless of disease status. This contradicts the IIF-based model, which is premised on that such intermixing frequency is a pathologic feature specific to preeclampsia. Instead, analysis under the IT framework revealed key molecular determinants of preeclampsia. Notably, classifier-prioritized genes associated with preeclampsia were enriched for ligands and receptors supporting a role for intercellular immune interactions. Among them, the ligand-receptor pair SPP1-CD44 between fetus and maternal immune cells emerged as peculiarly associated influencers of preeclampsia. Spatial image analysis confirmed co-localization of SPP1-CD44 expression within immune cell populations in preeclamptic placental tissue. Our study provides a comprehensive map of the human placenta and identifies disease-specific immune signaling pathways in preeclampsia using the divide and rule approach. The findings highlight SPP1-CD44 as a putative target of immune dysregulation, offering new insight into the cellular basis of maternal-fetal tolerance and its breakdown in pregnancy-related disorders. CLINICAL TRIAL NUMBER: Not applicable.