Human Decidual Natural Killer Cells Research Articles

Pyruvate dehydrogenase kinase 1 regulates the function of human decidual natural killer cells.

Pyruvate dehydrogenase kinase 1 (PDK1) is an important enzyme for immune cell development. However, PDK1's role in human decidual natural killer (dNK) cells remains largely unknown. PDK1 expression in dNK cells from patients with recurrent spontaneous abortions (RSA) and age-matched healthy controls was analyzed by qRT-PCR, western bolt and flow cytometry. Moreover, dNK cells were treated with PDK1 inhibitoror the PDK1 siRNA followed by functional assays. The dNK cells from patients who underwent RSAs had higher mRNA expression and increased proteinof PDK1, perforin (PRF1), Granzyme B (GZMB), IFN-γ (IFNG), and CD107a expression compared to dNK cells from age-matched healthy controls. Perforin, Granzyme B, IFN-γ and CD107a expression levels in dNK cells were down-regulated when dNK cells were treated with a PDK1 inhibitor. As measured by the 51 Cr release assay, the killing activity of dNK cells was found to be decreased. We also demonstrated that PDK1 blockade could up-regulate the migration and adhesion of dNK cells. Furthermore, PDK1 inhibition reduced the glycolysis of dNK cells. This study suggested that PDK1 plays an important role in regulating dNK cell functions and human RSA.

Palmitic Acid Upregulates CD96 Expression to Mediate Maternal-Foetal Interface Immune Tolerance by Inhibiting Cytotoxic Activity and Promoting Adhesion Function in Human Decidual Natural Killer Cells.

Decidual natural killer cells (dNK cells) are an essential component of the immune cells present at the maternal-foetal interface during early pregnancy, and they play a vital role in various physiological processes. Abnormalities in the ratio or function of dNK cells have been linked to recurrent miscarriages. CD96 has been previously shown to regulate NK cell function in the tumour microenvironment; however, its role and mechanism at the maternal-foetal interface remains unclear. The present study aimed to investigate the immunomodulatory role of CD96 in dNK cells and its function at the maternal-foetal interface. Immunofluorescence staining and flow cytometry were used to detect the expression of cellular markers such as CD96. Furthermore, the secretory function, adhesion-function-related molecules, and cell proliferation markers of CD96+ and CD96- dNK cells were detected using flow cytometry. In addition, we performed cell culture experiments via the magnetic bead sorting of NK cells to detect changes in the expression of the aforementioned functional molecules in dNK cells after the CD96 blockade. Furthermore, we examined the functional characteristics of dNK cells after palmitic acid treatment at a concentration of 10 μM. We also examined the changes in dNK cell function when subjected to the combined effect of palmitic acid and CD96 antagonists. The results indicated that CD96, TIGIT, CD155, and CD112 were highly expressed at the maternal-foetal interface, with dNK cells predominantly expressing CD96, whereas TIGIT was mainly expressed on T cells, and CD155 and CD112 were mainly present in metaphase stromal and trophoblast cells. CD96+ dNK cells displayed low cytotoxic activity and a high adhesion phenotype, which mediated the immunosuppressive effect on dNK cells at the maternal-foetal interface. Palmitic acid upregulated CD96 expression on the surface of dNK cells in the coculture system, inhibiting dNK cell activity and increasing their adhesion molecule expression. CD96 antagonist treatment blocked the inhibitory effect of trophoblasts on dNK cells, resulting in enhanced cytokine secretion and reduced adhesion. The results of this study provide valuable insight into the immunomodulatory role of CD96 in dNK cells and its mechanism at the maternal-foetal interface, particularly in metaphase NK cells. This study sheds light on the mechanisms of immune regulation at the maternal-foetal interface and their implications for the study of recurrent miscarriages of unknown origin.

CD49a regulates the function of human decidual natural killer cells

The function of CD49a on human decidual natural killer (dNK) cells is unknown. The expression of CD49a on dNK cells from human patients with recurrent spontaneous abortions or age-matched healthy controls was analyzed by flow cytometry. DNK cells were treated with CD49a neutralizing antibody and analyzed for function (cytokines production and cytotoxic activity). Long non-coding RNA (lncRNA) microarray analysis was used to identify a potential regulator of CD49a. DNK cells from human patients who underwent recurrent spontaneous abortions had lower levels of CD49a and increased perforin, granzyme B, and IFN-r expression, when compared to dNK cells from age-matched healthy controls. Perforin, granzyme B, and IFN-r expression levels in dNK cells were upregulated, while the migration and adhesion of dNK cells were downregulated by CD49a-neutralizing antibody. By the 51 Cr release assay, the killing activity of dNK cells also increased with CD49a neutralizing antibody. Further, lnc-49a, a newly identified lncRNA, was shown to be a positive regulator of CD49a in primary human NK cells. CD49a is involved in the regulation of dNK cells functions, including cytotoxic activity, migration, and adhesion. Further, lnc-49a is a positive regulator of CD49a in human primary dNK cells.

In vivo generation of decidual natural killer cells from resident hematopoietic progenitors

Decidual natural killer cells accumulate at the fetal-maternal interface and play a key role in a successful pregnancy. However, their origin is still unknown. Do they derive from peripheral natural killer cells recruited in decidua or do they represent a distinct population that originates in situ? Here, we identified natural killer precursors in decidua and uterus of pregnant mice. These precursors underwent rapid in situ differentiation and large proportions of proliferating immature natural killer cells were present in decidua and uterus as early as gestation day 4.5. Here, we investigated the origin of decidua- and uterus-natural killer cells by performing transfer experiments of peripheral mature natural killer cells or precursors from EGFP(+) mice. Results showed that mature natural killer cells did not migrate into decidua and uterus, while precursors were recruited in these organs and differentiated towards natural killer cells. Moreover, decidua- and uterus-natural killer cells displayed unique phenotypic and functional features. They expressed high levels of the activating Ly49D receptor in spite of their immature phenotype. In addition, decidua- and uterus-natural killer cells were poorly cytolytic and produced low amounts of IFN-γ, while they released factors (GM-CSF, VEGF, IP-10) involved in neo-angiogenesis and tissue remodeling. Our data reveal in situ generation of decidual natural killer cells and provide an important correlation between mouse and human decidual natural killer cells, allowing further studies to be carried out on their role in pregnancy-related diseases.

Changes of human decidual natural killer cells cocultured with YFP-Toxoplasma gondii: implications for abnormal pregnancy

To investigate the changes of human decidual natural killer (dNK) cells cocultured with Toxoplasma gondii in vitro and to infer implications on pregnancy. Case-control study. College and hospital. Decidual tissue was obtained from 85 patients undergoing voluntary abortion during the first trimester of gestation (6-12 weeks). None. The dNK cells were isolated and infected with YFP-Toxoplasma gondii. Cells were observed by fluorescence and confocal microscopy. The CD56(bright)CD16(-)/CD56(dim)CD16(+) dNK ratio, expression of KIR2DL4, ILT-2, and NKG2D on dNK cells were analyzed by flow cytometry and the cytotoxic activity of infected dNK cells were evaluated. The CD56(dim)CD16(+)/CD56(bright)CD16(-) dNK ratio was significantly elevated at 12, 24, and 48 hours after YFP-T. gondii infection. Expression of KIR2DL4, ILT-2, and NKG2D were increased after infection, but NKG2D were significantly higher than those of KIR2DL4 and ILT-2. Both the CD56(dim)CD16(+)/CD56(bright)CD16(-) dNK ratio and NKG2D expression were correlated with dNK cytotoxic activity. Enhanced dNK cytotoxicity due to increased CD16 and NKG2D expression may contribute to abnormal pregnancy outcomes observed with maternal infection with T. gondii.

Effector functions of human decidual NK cells in healthy early pregnancy are dependent on the specific engagement of natural cytotoxicity receptors

Natural killer (NK) cells represent the major lymphocyte population in the decidua basalis of the human uterus during healthy early pregnancy. The activity of decidual NK (dNK) cells and their activation status are different from those of peripheral blood (PB)-NK cells; i.e. dNK cells exhibit a unique phenotype. Decidual NK cells have been defined as CD56(bright), CD16(neg), and more recently CD160(neg). They express a unique repertoire of NK cell receptors, identical among all donors tested. Decidual NK cells express in particular NKp46-, NKp30- and NKp44-activating receptors, contrasting with PB-NK cells which are devoid of NKp44-activating receptors. Specific engagement of each of these three so-called natural cytotoxicity receptors in dNK cells has important functional consequences in terms of cytokine, chemokine and angiogenic factor secretion as well as cytotoxic potential. Strikingly, and in contrast with PB-NK cells, engagement of NKp46- but not NKp30-activating receptor on freshly isolated dNK cells triggers cytotoxicity. Such cytotoxic potential of dNK cells is negatively controlled by NKG2A inhibitory receptor co-engagement. This suggests that in situ, dNK cells cannot kill trophoblast cells during normal pregnancy. Whether such NKG2A-mediated inhibition is abolished during pregnancies complicated by pathologies including viral infection is an interesting hypothesis that remains to be tested.

Su.103. Phenotipic and Functional Characterization of Human Decidual Natural Killer Cells

Su.103. Phenotipic and Functional Characterization of Human Decidual Natural Killer Cells

Human decidual natural killer cells as a source and target of macrophage migration inhibitory factor

The human uterine mucosa of early pregnancy is largely populated by CD56(bright) natural killer (NK) cells (uterine (u) NK cells). The specific functions of these cells are still unknown, but their interaction and response to foetal trophoblasts are thought to be important for the establishment of a successful pregnancy. The study reported herein shows that uNK cells respond to, and produce, macrophage migration inhibitory factor (MIF), a cytokine highly expressed in the human placenta and in the cyclic and pregnant endometrium. Recombinant human MIF reduced in a dose-dependent manner the cytolytic activity of purified uNK cells against K562 cells. RT-PCR, Western blot analysis and ELISA demonstrated the synthesis and secretion of the cytokine by uNK cells. Double immunofluorescence staining showed the presence of MIF in uterine CD56 + cells. Finally, neutralization of the endogenous cytokine by a polyclonal antibody resulted in a sharp increase in the cytolytic activity of uNK cells. These findings indicate the existence of a previously unrevealed paracrine and autocrine action of MIF on uNK cells and support its contribution to the immune privilege at the maternal-foetal interface.

Human Decidual Natural Killer Cells Are a Unique NK Cell Subset with Immunomodulatory Potential

Natural killer cells constitute 50–90% of lymphocytes in human uterine decidua in early pregnancy. Here, CD56bright uterine decidual NK (dNK) cells were compared with the CD56bright and CD56dim peripheral NK cell subsets by microarray analysis, with verification of results by flow cytometry and RT-PCR. Among the ∼10,000 genes studied, 278 genes showed at least a threefold change with P ≤ 0.001 when comparing the dNK and peripheral NK cell subsets, most displaying increased expression in dNK cells. The largest number of these encoded surface proteins, including the unusual lectinlike receptors NKG2E and Ly-49L, several killer cell Ig-like receptors, the integrin subunits αD, αX, β1, and β5, and multiple tetraspanins (CD9, CD151, CD53, CD63, and TSPAN-5). Additionally, two secreted proteins, galectin-1 and progestagen-associated protein 14, known to have immunomodulatory functions, were selectively expressed in dNK cells.

Human decidual natural killer cells express the receptor for and respond to the cytokine interleukin 15.

The natural killer (NK) cells that are present in the uterine mucosa (decidua) during early pregnancy have a distinctive phenotype, CD56(bright) CD16(-). These cells have previously been shown to proliferate and be activated by interleukin (IL)-2. However, IL-2 is absent from the decidua and placenta, and we have therefore investigated whether IL-15 is present in the uterus and can act on decidual NK cells. Both IL-15 mRNA and protein were found in a variety of cells but particularly in decidual macrophages. IL-15 induced a proliferative response in decidual NK cells that was blocked by anti-IL-15 and was augmented by stem cell factor. The cytolytic activity of decidual NK cells against K562 was augmented. Interestingly, in contrast to IL-2, although activation with IL-15 resulted in some killing of JEG-3 choriocarcinoma cells, normal trophoblast cells remained resistant to lysis. These findings suggest that IL-15 is a candidate cytokine responsible for NK cell proliferation in vivo in the progesterone-dominated secretory endometrium and early decidua.

Co-stimulation of human decidual natural killer cells by interleukin-2 and stromal cells.

At the late secretory phase of the menstrual cycle and in early pregnancy, the uterine mucosa is infiltrated by large numbers of natural killer (NK) cells with a distinctive phenotype (CD56bright CD16- CD3-) and large granular lymphocyte (LGL) morphology. Circulating CD56bright NK cells generally proliferate in the presence of interleukin-2 (IL-2), but it is clear that cofactors besides IL-2 are required for optimal response. In the bone marrow, this co-stimulating signal is provided by stromal cells. In the present study we observe that uterine CD56+ cells from early pregnancy decidua similarly proliferate vigorously when cultured with decidual stromal cells and a suboptimal dose of IL-2. This response is dependent on cell-cell contact, as no proliferation of decidual NK cells was observed when they were separated from stromal cells by a permeable cyclopore membrane. In addition, we have studied the expression of Bcl-2 by decidual CD56+ cells. Our results show that the microenvironment of the uterus is likely to have a significant influence on the proliferation and survival of uterine CD56+ cells.