Origin of human uterine natural killer cells

Abstract Uterine NK cells (uNK) are a distinct immune population which influence pregnancy outcome. While murine studies have suggested that tissue-resident uNKs predominate in virgin endometrium, the origin of uNKs in humans is unknown. To define subsets of uterine NKs in humans and determine their developmental origin, we performed single-cell RNA-sequencing on CD56+ uNKs sorted from endometrial biopsies of 4 healthy controls (HCs) (17,146 cells) and 5 uterus transplant (UTx) recipients (30,351 cells). SNP polymorphisms and HLA alleles were used to identify cells of donor (tissue-resident) or recipient (peripheral) origin in UTx recipients. Developmental trajectories were built with Monocle v.3.0. Ten major clusters were identified in HCs, with 48% of endometrial NK cells (eNKs) occupying 3 clusters (eNK1–3) that shared gene signatures with known human decidual NK cell subsets (dNK1–3). Pseudotime analysis revealed that the eNK1–3 clusters arose from proliferating subsets and were less mature than cytotoxic uNKs expressing ZEB2, TBX21, and FCGR3A (CD16). uNKs from UTx recipients were similarly distributed across the clusters, with a trend towards fewer cells in the FCGR3A-expressing mature subset. Notably, recipient-derived uNKs dominated all uNK subsets within four months post-transplant but became less frequent in UTx recipients >1 year after transplant (95% vs. 72%; early vs. late). The proportion of donor-derived to recipient-derived cells remained constant across subsets in all UTx recipients. In contrast to mouse models, our study suggests that human uNKs derive primarily from blood-borne peripheral immigrants and that environmental factors influence uNK differentiation more than genotype or developmental origin.