CD32 captures committed haemogenic endothelial cells during human embryonic development.

During embryonic development, blood cells emerge from specialized endothelial cells, named haemogenic endothelial cells (HECs). As HECs are rare and only transiently found in early developing embryos, it remains difficult to distinguish them from endothelial cells. Here we performed transcriptomic analysis of 28- to 32-day human embryos and observed that the expression of Fc receptor CD32 (FCGR2B) is highly enriched in the endothelial cell population that contains HECs. Functional analyses using human embryonic and human pluripotent stem cell-derived endothelial cells revealed that robust multilineage haematopoietic potential is harboured within CD32+ endothelial cells and showed that 90% of CD32+ endothelial cells are bona fide HECs. Remarkably, these analyses indicated that HECs progress through different states, culminating in FCGR2B expression, at which point cells are irreversibly committed to a haematopoietic fate. These findings provide a precise method for isolating HECs from human embryos and human pluripotent stem cell cultures, thus allowing the efficient generation of haematopoietic cells in vitro.

The transcriptional repressor ID2 supports natural killer cell maturation by controlling TCF1 amplitude.

Gaining a mechanistic understanding of the expansion and maturation program of natural killer (NK) cells will provide opportunities for harnessing their inflammation-inducing and oncolytic capacity for therapeutic purposes. Here, we demonstrated that ID2, a transcriptional regulatory protein constitutively expressed in NK cells, supports NK cell effector maturation by controlling the amplitude and temporal dynamics of the transcription factor TCF1. TCF1 promotes immature NK cell expansion and restrains differentiation. The increased TCF1 expression in ID2-deficient NK cells arrests their maturation and alters cell surface receptor expression. Moreover, TCF1 limits NK cell functions, such as cytokine-induced IFN-γ production and the ability to clear metastatic melanoma in ID2-deficient NK cells. Our data demonstrate that ID2 sets a threshold for TCF1 during NK cell development, thus controlling the balance of immature and terminally differentiated cells that support future NK cell responses.

Transcription factor ID2 prevents E proteins from enforcing a naïve T lymphocyte gene program during NK cell development.

All innate lymphoid cells (ILCs) require the small helix-loop-helix transcription factor ID2, but the functions of ID2 are not well understood in these cells. We show that mature natural killer (NK) cells, the prototypic ILCs, developed in mice lacking ID2 but remained as precursor CD27+CD11b- cells that failed to differentiate into CD27-CD11b+ cytotoxic effectors. We show that ID2 limited chromatin accessibility at E protein binding sites near naïve T lymphocyte-associated genes including multiple chemokine receptors, cytokine receptors, and signaling molecules and altered the NK cell response to inflammatory cytokines. In the absence of ID2, CD27+CD11b- NK cells expressed ID3, a helix-loop-helix protein associated with naïve T cells, and they transitioned from a CD8 memory precursor-like to a naïve-like chromatin accessibility state. We demonstrate that ID3 was required for the development of ID2-deficient NK cells, indicating that completely unfettered E protein function is incompatible with NK cell development. These data solidify the roles of ID2 and ID3 as mediators of effector and naïve gene programs, respectively, and revealed a critical role for ID2 in promoting a chromatin state and transcriptional program in CD27+CD11b- NK cells that supports cytotoxic effector differentiation and cytokine responses.

Cytochrome P450 26A1 modulates natural killer cells in mouse early pregnancy.

Cytochrome P450 26A1 (CYP26A1) has a spatiotemporal expression pattern in the uterus, with a significant increase in mRNA and protein levels during peri-implantation. Inhibiting the function or expression of CYP26A1 can cause pregnancy failure, suggesting an important regulatory role of CYP26A1 in the maintenance of pregnancy. However, little is known about the exact mechanism involved. In this study, using a pCR3.1-cyp26a1 plasmid immunization mouse model and a Cyp26a1-MO (Cyp26a1-specific antisense oligos) knockdown mouse model, we report that the number of Dolichos biflorus agglutinin (DBA) lectin-positive uterine natural killer (uNK) cells was reduced in pCR3.1-cyp26a1 plasmid immunized and Cyp26a1-MO-treated mice. In contrast, the percentage of CD3- CD49b+ NK cells in the uteri from the treatment group was significantly higher than that of the control group in both models. Similarly, significantly up-regulated expression of CD49b (a pan-NK cell marker), interferon gamma, CCL2, CCR2 (CCL2 receptor) and CCL3 were detected in the uteri of pCR3.1-cyp26a1- and Cyp26a1-MO-treated mice. Transcriptome analysis suggested that CYP26A1 might regulate NK cells through chemokines. In conclusion, the present data suggest that silencing CYP26A1 expression/function can decrease the number of uNK cells and significantly increase the percentage of CD3- CD49b+ NK cells in the uteri of pregnant mice. These findings provide a new line of evidence correlating the deleterious effects of blocking CYP26A1 in pregnancy with the aberrant regulation of NK cells in the uterus.

Seminal plasma induces inflammation in the uterus through the γδ T/IL-17 pathway.

After insemination, a large number of leukocytes migrate into the uterus, which is accompanied by intense inflammation. However, the details of how seminal plasma interacts with the uterus are still not very clear. Here, we present that neutrophils migrate and accumulate around the uterine epithelium following insemination, which is accompanied by an increase in interleukin (IL) 17A levels. Additionally, we find that γδ T cells are the major source of IL-17A, and the seminal plasma could induce the γδ T cells to secret IL-17A. Blocking IL-17A could reduce the number of neutrophils in the uterus and prevent them from migrating to the epithelium by decreasing the chemokines CXCL1, CXCL2 and CXCL5. Blocking IL-17A did not affect the Th1/Th2 balance but actually diminished the inflammation in the uterus by reducing the expression of IL-1ß and TNF-α. In summary, we found a new mechanism by which seminal plasma could influence the inflammation in the uterus through the γδ T/IL-17 pathway to regulate the expression of various chemokines and cytokines.

IFN-γ modulates Ly-49 receptors on NK cells in IFN-γ-induced pregnancy failure.

We have previously shown that interferon gamma (IFN-γ) induces aberrant CD49b(+) natural killer (NK) cell recruitment by regulating CX3CL1 and eventually provokes foetal loss. In this study, we show that IFN-γ also modulates Ly-49 receptors on NK cells during pregnancy failure. The percentages of Ly-49A(+) and Ly-49G2(+) NK cells in the uteri of the IFN-γ-treated group were significantly lower than those observed in the control group. Moreover, the median fluorescence intensity (MFI) values of Ly-49A and Ly-49G2 expression on NK cells in the uteri of the IFN-γ-treated group were significantly lower than those of the control group. Using isolated spleen leucocytes, we further found that IFN-γ significantly reduced the percentage of Ly-49A(+) NK cells in vitro. However, CX3CL1 was not involved in the modulation of Ly-49 receptors, and the expression of CX3CR1 was not regulated by IFN-γ in spleen leucocytes. Collectively, our data indicate that IFN-γ can modulate Ly-49 receptors on NK cells and this process may play a role in IFN-γ-induced pregnancy failure. Thus, we provide a new line of evidence correlating the deleterious effects of IFN-γ with its role in regulating NK cell Ly-49 receptors during pregnancy failure.