Innate lymphoid cells (ILCs), including NK cells, contribute to barrier immunity and tissue homeostasis. In addition to the role of uterine NK cells in placentation and fetal growth, other uterine ILCs (uILCs) are likely to play roles in uterine physiology and pathology. In this article, we report on the composition of uILCs in the endometrium during the luteal phase and in the decidua during early pregnancy. Whereas nonkiller uILC1s and uILC2s are barely detectable in mouse and not detected in humans, a sizeable population of uILC3s is found in human endometrium and decidua, which are mostly NCR+ and partially overlap with previously described IL-22–producing uterine NK cells. Development of mouse uILC3 is Nfil3 independent, suggesting unique features of uILCs. Indeed, although the cytokine production profile of mouse uILCs recapitulates that described in other tissues, IL-5, IL-17, and IL-22 are constitutively produced by uILC2s and uILC3s. This study lays the foundation to understand how ILCs function in the specialized uterine mucosa, both in tissue homeostasis and barrier immunity and during pregnancy.

Uterine NK cells are innate lymphoid cells (ILC) that populate the uterus and expand during pregnancy, regulating placental development and fetal growth in humans and mice. We have recently characterized the composition of uterine ILCs (uILCs), some of which require the transcription factor NFIL3, but the extent to which NFIL3-dependent cells support successful reproduction in mice is unknown. By mating Nfil3−/− females with wild-type males, here we show the effects of NFIL3 deficiency in maternal cells on both the changes in uILCs during pregnancy and the downstream consequences on reproduction. Despite the presence of CD49a+Eomes− uILC1s and the considerable expansion of residual CD49a+Eomes+ tissue-resident NK cells and uILC3s in pregnant Nfil3−/− mice, we found incomplete remodeling of uterine arteries and decidua, placental defects, and fetal growth restriction in litters of normal size. These results show that maternal NFIL3 mediates non-redundant functions in mouse reproduction.

Determining the function of uterine lymphocytes is challenging because of the dynamic changes in response to sex hormones and, during pregnancy, to the invading foetal trophoblast cells. Here we provide a genome-wide transcriptome atlas of mouse uterine group 1 innate lymphoid cells (ILCs) at mid-gestation. Tissue-resident Eomes+CD49a+ NK cells (trNK), which resemble human uterine NK cells, are most abundant during early pregnancy, and have gene signatures associated with TGF-β responses and interactions with trophoblast, epithelial, endothelial, smooth muscle cells, leucocytes and extracellular matrix. Conventional NK cells expand late in gestation and may engage in crosstalk with trNK cells involving IL-18 and IFN-γ. Eomes−CD49a+ ILC1s dominate before puberty, and specifically expand in second pregnancies when the expression of the memory cell marker CXCR6 is upregulated. These results identify trNK cells as the cellular hub of uterine group 1 ILCs, and mark CXCR6+ ILC1s as potential memory cells of pregnancy.

Highlights d CD94/NKG2A educates uterine NK cells d NKG2A-deficient dams display reduced utero-placental hemodynamic adaptations d Asymmetric growth restriction and abnormal brain development in NKG2A-deficient dams d Non-functional HLA-B/HLA-E/NKG2A pathway exposes women to greater pre-eclampsia risk

Determining the function of uterine lymphocytes is challenging because of the rapidly changing nature of the organ in response to sex hormones and, during pregnancy, to the invading fetal trophoblast cells. Here we provide the first genome-wide transcriptome atlas of mouse uterine group 1 innate lymphoid cells (g1 ILCs) at mid-gestation. The composition of g1 ILCs fluctuates throughout reproductive life, with Eomes-veCD49a+ ILC1s dominating before puberty and specifically expanding in second pregnancies, when the expression of CXCR6, a marker of memory cells, is upregulated. Tissue-resident Eomes+CD49a+ NK cells (trNK), which resemble human uterine NK cells, are most abundant during early pregnancy, and showcase gene signatures of responsiveness to TGF-β, connections with trophoblast, epithelial, endothelial and smooth muscle cells, leucocytes, as well as extracellular matrix. Unexpectedly, trNK cells express genes involved in anaerobic glycolysis, lipid metabolism, iron transport, protein ubiquitination, and recognition of microbial molecular patterns. Conventional NK cells expand late in gestation and may engage in crosstalk with trNK cells involving IL-18 and IFN-γ. These results identify trNK cells as the cellular hub of uterine g1 ILCs at mid-gestation and mark CXCR6+ ILC1s as potential memory cells of pregnancy.

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Fetal growth restriction (FGR) causes a wide variety of defects in the neonate which can lead to increased risk of heart disease, diabetes, anxiety and other disorders later in life. However, the effect of FGR on the immune system, is poorly understood. We used a well-characterized mouse model of FGR in which placental Igf-2 production is lost due to deletion of the placental specific Igf-2 P0 promotor. The thymi in such animals were reduced in mass with a ~70% reduction in cellularity. We used single cell RNA sequencing (Drop-Seq) to analyze 7,264 thymus cells collected at postnatal day 6. We identified considerable heterogeneity among the Cd8/Cd4 double positive cells with one subcluster showing marked upregulation of transcripts encoding a sub-set of proteins that contribute to the surface of the ribosome. The cells from the FGR animals were underrepresented in this cluster. Furthermore, the distribution of cells from the FGR animals was skewed with a higher proportion of immature double negative cells and fewer mature T-cells. Cell cycle regulator transcripts also varied across clusters. The T-cell deficit in FGR mice persisted into adulthood, even when body and organ weights approached normal levels due to catch-up growth. This finding complements the altered immunity found in growth restricted human infants. This reduction in T-cellularity may have implications for adult immunity, adding to the list of adult conditions in which the in utero environment is a contributory factor.

SummaryMaternal immune cells are an integral part of reproduction, but how they might cause pregnancy complications remains elusive. Macrophages and their dual function in inflammation and tissue repair are thought to play key yet undefined roles. Altered perinatal growth underpins adult morbidity, and natural killer (NK) cells may sustain fetal growth by establishing the placental blood supply. Using a mouse model of genetic inactivation of PI3K p110δ, a key intracellular signaling molecule in leukocytes, we show that p110δ regulates macrophage dynamics and NK-cell-mediated arterial remodeling. The uterus of dams with inactive p110δ had decreased IFN-γ and MHC class IIlow macrophages but enhanced IL-6. Poor vascular remodeling and a pro-inflammatory uterine milieu resulted in fetal death or growth retardation. Our results provide one mechanism that explains how imbalanced adaptations of maternal innate immune cells to gestation affect offspring well-being with consequence perinatally and possibly into adulthood.