Blood Natural Killer Cell Subsets Research Articles

Characterization of NK Cells Using Single-Cell RNA Sequencing in Patients With Acute-On-Chronic Liver Failure.

Acute-on-chronic liver failure (ACLF) is characterized by fast progression and high mortality, with systemic inflammation and immune paralysis as its key events. While natural killer (NK) cells are key innate immune cells, their unique function and subpopulation heterogeneity in ACLF have not been fully elucidated. This study aimed to investigate the characteristics of NK cell subsets in the peripheral blood of patients with ACLF and determine their roles in the inflammatory responses. Circulating NK cells (14 751 cells) from patients with ACLF and healthy controls (HCs) were subjected to single-cell RNA sequencing (scRNA-seq). Clustering and annotation were used to identify the features of NK cell subsets and the characteristics of disease progression in ACLF. Four NK cell subsets were obtained, including adaptive NK cells, mature NK cells, inflamed NK cells, and CD56bright NK cells. Compared with the HCs, the patients with ACLF had a significantly lower proportion of Mature NK cells and a higher proportion of Inflamed NK cells. Quasi-temporal analysis showed that Inflamed NK cells were highly enriched in the late quasi-temporal sequence, and genes related to pro-inflammatory were significantly up-regulated in Inflamed NK cells. In addition, scRNA-seq and flow cytometry confirmed that the expression level of cell migration inducing hyaluronidase 2 (CEMIP2) in NK cells progressively increased from the HC group to the ACLF survival group and then to the ACLF death group. scRNA-seq reveals that Inflamed NK cell subsets are associated with ACLF progression and poor prognosis. CEMIP2 may be a molecular marker for ACLF progression.

The Expression pattern of NK cells in systemic lupus erythematosus patients with different disease activities

Data demonstrated the role of natural killer (NK) cells in systemic lupus erythematosus (SLE). We aimed to determine the immunophenotype and frequency of NK cells and their subsets, CXCR3, CD161 expression in blood and renal tissue of SLE patients with and without lupus nephritis and their relationship with disease activity. The study included 31 SLE patients and 11 controls. Study participants underwent full history and thorough clinical examination. SLE patients underwent routine laboratory investigations. Renal tissue biopsies were taken from patients with lupus nephritis. The frequency of NK cell subsets in blood of patients and controls and renal tissue from patients was performed by flow cytometry. An increase in circulatory CD56bright NK cells and its CD56bright CD16dim subtype was associated with the severity of systemic manifestations in SLE patients. Total CD56bright NK cells and its CD56bright CD16bright subtypes in renal tissues were related to renal damage. We detected decreased CD161 expression on NK cells related to renal damage and severity of systemic manifestations in SLE patients. Decreased expression of CXCR3 on NK cell surface in renal tissues causes misdirected trafficking of NK cells that seems to reduce the severity of lupus nephritis. In conclusion, our results paved the way to understanding the role of NK cells in the pathogenesis of SLE which may represent a future target for immune therapy of SLE. NK and CD56dim subset were decreased in the blood and increased in renal tissue of SLE patients, while the CD56bright subset was increased in blood and decreased in renal tissue reflecting their effects on renal damage and severity of manifestations in SLE.

High-dimensional single-cell analysis of human natural killer cell heterogeneity

Natural killer (NK) cells are innate lymphoid cells (ILCs) contributing to immune responses to microbes and tumors. Historically, their classification hinged on a limited array of surface protein markers. Here, we used single-cell RNA sequencing (scRNA-seq) and cellular indexing of transcriptomes and epitopes by sequencing (CITE-seq) to dissect the heterogeneity of NK cells. We identified three prominent NK cell subsets in healthy human blood: NK1, NK2 and NK3, further differentiated into six distinct subgroups. Our findings delineate the molecular characteristics, key transcription factors, biological functions, metabolic traits and cytokine responses of each subgroup. These data also suggest two separate ontogenetic origins for NK cells, leading to divergent transcriptional trajectories. Furthermore, we analyzed the distribution of NK cell subsets in the lung, tonsils and intraepithelial lymphocytes isolated from healthy individuals and in 22 tumor types. This standardized terminology aims at fostering clarity and consistency in future research, thereby improving cross-study comparisons.

Aberrant NK cell profile in gestational diabetes mellitus with fetal growth restriction.

Gestational diabetes mellitus (GDM) is a gestational disorder characterized by hyperglycemia, that can lead to dysfunction of diverse cells in the body, especially the immune cells. It has been reported that immune cells, specifically natural killer (NK) cells, play a crucial role in normal pregnancy. However, it remains unknown how hyperglycemia affects NK cell dysfunction thus participates in the development of GDM. In this experiment, GDM mice were induced by an intraperitoneal injection of streptozotocin (STZ) after pregnancy and it has been found that the intrauterine growth restriction occurred in mice with STZ-induced GDM, accompanied by the changed proportion and function of NK cells. The percentage of cytotoxic CD27-CD11b+ NK cells was significantly increased, while the proportion of nourished CD27-CD11b- NK cells was significantly reduced in the decidua of GDM mice. Likewise, the same trend appeared in the peripheral blood NK cell subsets of GDM patients. What's more, after intrauterine reinfusion of NK cells to GDM mice, the fetal growth restriction was alleviated and the proportion of NK cells was restored. Our findings provide a theoretical and experimental basis for further exploring the pathogenesis of GDM.

Natural Killer Cells Regulate Acute SIV Replication, Dissemination, and Inflammation, but Do Not Impact Independent Transmission Events.

Natural killer (NK) cells are potent effector cells of the innate immune system possessing both cytotoxic and immunoregulatory capabilities, which contribute to their crucial role in controlling human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) infections. However, despite significant evidence for NK cell modulation of HIV disease, their specific contribution to transmission and control of acute infection remains less clear. To elucidate the contribution of NK cells during acute SIV infection, we performed an acute necropsy study, where rhesus macaques (RM) were subjected to preinfection depletion of systemic NK cells using established methods of IL-15 neutralization, followed by subsequent challenge with barcoded SIVmac239X. Our study showed that depletion was highly effective, resulting in near total ablation of all NK cell subsets in blood, liver, oral, and rectal mucosae, and lymph nodes (LN) that persisted through the duration of the study. Meanwhile, frequencies and phenotypes of T cells remained virtually unchanged, indicating that our method of NK cell depletion had minimal off-target effects. Importantly, NK cell-depleted RM demonstrated an early and sustained 1 to 2 log increase in viremia over controls, but sequence analysis suggested no difference in the number of independent transmission events. Acute bulk, central memory (CM), and CCR5+ CD4+ T cell depletion was similar between experimental and control groups, while CD8+ T cell activation was higher in NK cell-depleted RM as measured by Ki67 and PD-1 expression. Using 27-plex Luminex analyses, we also found modestly increased inflammatory cytokines in NK cell-depleted RM compared to control animals. In the effort to determine the impact of NK cells on HIV/SIV transmission and acute viremia, future studies will be necessary to better harness these cells for future viral therapies. Collectively, these data suggest NK cells are important modulators of lentivirus dissemination and disease but may not have the capacity to independently eliminate individual transmission events. IMPORTANCE Natural killer (NK) cells as major effector cells of the innate immune system can contribute significantly to human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) control. However, a specific role for NK cells in blocking lentivirus transmission remains incompletely clear. In this study, we depleted NK cells prior to challenge with a barcoded SIV. Importantly, our studied showed systemic NK cell depletion was associated with a significant increase in acute viremia, but did not impact the number of independent transmission events. Collectively, these data suggest NK cells are critical modulators of early lentivirus replication but may not regulate individual transmission events at mucosal portals of entry.

Identification of the CD16low CD56low CD38pos Natural Killer Cell as a Key Subset in Patients with Multiple Myeloma.

Natural killer (NK) cells are classified as innate immune cells which can directly recognize and kill tumor cells without antigen sensitization. NK cell-based adoptive immunotherapy for blood malignancies has attracted more attention in recent years. To analyze different NK cell subsets in the peripheral blood and bone marrow (BM) of patients with multiple myeloma (MM). Using flow cytometry we analyzed: (i) the distribution of distinct NK cell subpopulations (i.e. CD16low CD56low, CD16pos CD56high, CD16neg CD56high, CD16high CD56low, CD16neg CD56low, CD16low CD56low CD38pos) in the BM from MM patients at distinct disease stages. (ii) the expression of NKG2D, DNAM-1 and NKp30, and (iii) the expression of CD107a in CD16low CD56low CD38pos and CD16low CD56low CD38neg NK cells subsets. CD16low CD56low CD38pos was the dominant subset in BM from patients with MM at the CR stage with a decreased expression of NKp30. CD16low CD56low CD38pos subset showed a higher proportion of CD107a expression compared to CD16low CD56low CD38neg cells. In vitro experiments indicated that the CD16low CD56low CD38pos NK cell subset possesses more cytotoxicity than CD16low CD56low CD38neg NK cells. Our data suggest that CD16low CD56low CD38pos NK cells may reflect as an effector population with the potential therapeutic target in patients with MM. This group of cells may be useful for adoptive immunotherapy in MM in the future.

CXCR6+ and NKG2C+ Natural Killer Cells Are Distinct With Unique Phenotypic and Functional Attributes Following Bone Marrow Transplantation.

Reactivation of human cytomegalovirus (HCMV) is a life-threatening complication in transplant patients. Natural Killer (NK) cells are the first lymphocyte lineage to reconstitute following an allogeneic hematopoietic stem cell transplant (HSCT). Amongst them, NK cell Group 2 isoform C/Killer cell lectin-like receptor subfamily C, member 2 (NKG2C)-expressing NK cells contribute significantly to patient protection upon HCMV reactivation. NKG2C+ NK cells are capable of immunological memory, albeit NK cell memory is not restricted to them. Hepatic C-X-C Motif Chemokine Receptor 6 (CXCR6)-expressing NK cells also mediate memory responses in mice and humans. Small numbers of them circulate and can thus be studied in peripheral blood samples. We hypothesize that NKG2C+ and CXCR6+ NK cell subsets are distinct. To test our hypothesis, we used multi-parametric flow cytometry to determine the phenotypes and effector functions of CD56bright vs. CD56dim and NKG2C+ vs. CXCR6+ human NK cell subsets in the peripheral blood (PB) of pediatric transplant recipients monthly while monitoring patients for HCMV reactivation. Interestingly, we did not find any NKG2C+CXCR6+ NK cells in the transplant recipients’ peripheral blood, suggesting that NKG2C+ and CXCR6+ NK cells are distinct. Also, NKG2C–CXCR6– NK cells, rather than NKG2C+ NK cells, made up most NK cells post-transplant, even in transplant recipients with HCMV viremia. In contrast to NKG2C+ NK cells, CXCR6+ NK cells appeared phenotypically less differentiated but were highly proliferative and produced IFN-γ and TNF α . Our findings contribute to our understanding of post-transplant NK cell development and its implications for human health.

Natural killer cells promote the differentiation of follicular helper T cells instead of inducing apoptosis in myasthenia gravis

Recent evidence has shown that natural killer (NK) cells have an immunoregulatory function in the pathogenesis of myasthenia gravis (MG). In this study, the phenotype and function of NK cell subsets in peripheral blood of new-onset MG (N-MG) and stable MG (S-MG) patients were explored. Circulating CD56dim and CD56bright NK cells were increased and decreased, respectively, in patients with N-MG and S-MG compared with healthy control (HC). Moreover, all circulating NK cell subsets from N-MG patients showed significantly lower expression of activating receptor NKG2D and production of Interferon (IFN) -γ than that from HC. The killing effects of NK cells on CD4+ T cells and Tfh cells were impaired in MG patients, whereas, they promoted the differentiation and activation of Tfh cells. These data indicated that the immune-regulation of NK cells on CD4+ T cells and Tfh cells in MG patients was abnormal, which may contribute to the immune-pathological mechanism of MG.

Assessment of Natural Killer Cell Subsets in HCV Infection and Hepatocellular Carcinoma

Aim of work: In this study the NK cell CD markers testing were offer the possibility and potentiality of some types of NK cells to target HCV and HCC immunotherapy. Background: Natural Killer (NK) cells are important for responding to and destroying cells that have become infected with virus and cancer cells. We evaluated the effects of hepatitis C virus (HCV) only infected patients and HCV-related hepatocellular carcinoma (HCC) of Egyptian cases on peripheral blood NK cell subsets CD56bright and CD56Dim NK cells, CD56dim16+, CD56dim16 and with emphasis on the CD3 cells. Methods: There are NK subpopulations levels that can easily be distinguished by flow-cytometry as depicted. Results: A significant increase in CD3 HCV patients and HCC were observed compared with healthy control individuals.CD56 subsets were non-significantly elevated in HCV only infected patients, but highly significantly lowered in HCV infected cases that associated with HCC, in addition to HCC cases were recorded decreasing levels for CD56 subtypes. Finally the hallmark of this NK cells dysfunction is persistent activation promoting ongoing hepatitis and hepatocytes damage which contribute to the progression of HCC, while being unable to clear HCV due to impaired IFN- γ response. Conclusions: Testing of NK cell subtypes will offers the possibility and potential to target HCC by immunotherapy and personalizing immunotherapy theory in different tumors.

Altered Expression of Natural Cytotoxicity Receptors and NKG2D on Peripheral Blood NK Cell Subsets in Breast Cancer Patients

Human natural killer (NK) cells are considered professional cytotoxic cells that are integrated into the effector branch of innate immunity during antiviral and antitumoral responses. The purpose of this study was to examine the peripheral distribution and expression of NK cell activation receptors from the fresh peripheral blood mononuclear cells of 30 breast cancer patients prior to any form of treatment (including surgery, chemotherapy, and radiotherapy), 10 benign breast pathology patients, and 24 control individuals. CD3−CD56dimCD16bright NK cells (CD56dim NK) and CD3−CD56brightCD16dim/− NK cells (CD56bright NK) were identified using flow cytometry. The circulating counts of CD56dim and CD56bright NK cells were not significantly different between the groups evaluated, nor were the counts of other leukocyte subsets between the breast cancer patients and benign breast pathology patients. However, in CD56dim NK cells, NKp44 expression was higher in breast cancer patients (P = .0302), whereas NKp30 (P = .0005), NKp46 (P = .0298), and NKG2D (P = .0005) expression was lower with respect to healthy donors. In CD56bright NK cells, NKp30 (P = .0007), NKp46 (P = .0012), and NKG2D (P = .0069) expression was lower in breast cancer patients compared with control group. Only NKG2D in CD56bright NK cells (P = .0208) and CD56dim NK cells (P = .0439) showed difference between benign breast pathology and breast cancer patients. Collectively, the current study showed phenotypic alterations in activation receptors on CD56dim and CD56bright NK cells, suggesting that breast cancer patients have decreased NK cell cytotoxicity.

Increased CD56(bright) NK cells in HIV-HCV co-infection and HCV mono-infection are associated with distinctive alterations of their phenotype.

BackgroundHIV-HCV co-infection is associated with accelerated progression to hepatic fibrosis, cirrhosis and hepatocellular carcinoma than HCV mono-infection. The contribution of innate immunity during HIV-HCV co-infection has been a relatively under-investigated area. Natural killer (NK) cells are pivotal sentinels of innate immunity against viruses and tumour cells. In this study we evaluated the effect of HIV-HCV co-infection on peripheral blood NK cell subsets with emphasis on the phenotype of CD56bright NK cells.MethodsSixty patients were included in the study; HIV mono-infected (n = 12), HCV mono-infected (n = 15), HCV-HIV co-infected (n = 21) and healthy controls (n = 16). PBMCs were isolated and immunophenotyping of NK cells was performed by flowcytometry.ResultsWe observed an expansion of CD56bright NK cell subset in HIV-HCV co-infection as compared to healthy controls and HIV mono-infected group. All the infected groups had an upregulated expression of the activating receptor NKG2D on CD56bright NK cells in comparison to healthy controls while not differing amongst themselves.The expression of NKp46 in HIV-HCV co-infected group was significantly upregulated as compared to both HIV as well as HCV mono-infections while NKp30 expression in the HIV-HCV co-infected group significantly differed as compared to HIV mono-infection. The CD56bright NK cell subset was activated in HIV-HCV co-infection as assessed by the expression of CD69 as compared to healthy controls but was significantly downregulated in comparison to HIV mono-infection. CD95 expression on CD56bright NK cells followed the same pattern where there was an increased expression of CD95 in HIV mono-infection and HIV-HCV co-infection as compared to healthy controls. In contrast to CD69 expression, CD95 expression in HCV mono-infection was decreased when compared to HIV mono-infection and HIV-HCV co-infection. Finally, expression of CXCR3 on CD56bright NK cells was increased in HIV-HCV co-infection in comparison to HIV mono-infection while remaining similar to HCV mono-infection.ConclusionThus, HIV-HCV co-infection is able to modulate the phenotype of CD56bright NK cell subset in a unique way such that NKp46 and CXCR3 expressions are distinct for co-infection while both mono-infections have an additive effect on CD56bright, CD69 with CD95 expressions. HCV mono-infection has a dominant effect on NKp30 expression while NKG2D and CD127 expressions remained same in all the groups.

Tumor-associated and immunochemotherapy-dependent long-term alterations of the peripheral blood NK cell compartment in DLBCL patients

Natural Killer (NK) cells are a key component of tumor immunosurveillance and thus play an important role in rituximab-dependent killing of lymphoma cells via an antibody-dependent cellular cytotoxicity (ADCC) mechanism. We evaluated the phenotypic and functional assets of peripheral blood NK cell subsets in 32 newly-diagnosed diffuse large B-cell lymphoma (DLBCL) patients and in 27 healthy controls. We further monitored long-term modifications of patient NK cells for up to 12 months after rituximab-based immunochemotherapy. At diagnosis, patients showed a higher percentage of CD56dim and CD16+ NK cells, and a higher frequency of GrzB+ cells in CD56dim, CD56bright, and CD16+ NK cell subsets than healthy controls. Conversely, DLBCL NK cell killing and interferon γ (IFNγ) production capability were comparable to those derived from healthy subjects. Notably, NK cells from refractory/relapsed patients exhibited a lower “natural” cytotoxicity. A marked and prolonged therapy-induced reduction of both “natural” and CD16-dependent NK cytotoxic activities was accompanied by the down-modulation of CD16 and NKG2D activating receptors, particularly in the CD56dim subset. However, reduced NK cell killing was not associated with defective lytic granule content or IFNγ production capability. This study firstly describes tumor-associated and therapy-induced alterations of the systemic NK cell compartment in DLBCL patients. As these alterations may negatively impact rituximab-based therapy efficacy, our work may provide useful information for improving immunochemotherapeutic strategies.

NK Cell Subset Redistribution during the Course of Viral Infections.

Natural killer (NK) cells are important effectors of innate immunity that play a critical role in the control of human viral infections. Indeed, given their capability to directly recognize virally infected cells without the need of specific antigen presentation, NK cells are on the first line of defense against these invading pathogens. By establishing cellular networks with a variety of cell types such as dendritic cells, NK cells can also amplify anti-viral adaptive immune responses. In turn, viruses evolved and developed several mechanisms to evade NK cell-mediated immune activity. It has been reported that certain viral diseases, including human immunodeficiency virus-1 as well as human cytomegalovirus infections, are associated with a pathologic redistribution of NK cell subsets in the peripheral blood. In particular, it has been observed the expansion of unconventional CD56neg NK cells, whose effector functions are significantly impaired as compared to that of conventional CD56pos NK cells. In this review, we address the impact of these two chronic viral infections on the functional and phenotypic perturbations of human NK cell compartment.

Comparative analysis of NK cell subsets in menstrual and peripheral blood of patients with unexplained recurrent spontaneous abortion and fertile subjects

Natural killer (NK) cells play a fundamental role in maintaining pregnancy. Based on the availability and non-invasive method of collection of menstrual blood (MB), here we investigated for the first time a comparative analysis of NK cell subsets in MB and peripheral blood (PB) of women with recurrent spontaneous abortion (RSA) and fertile women. PB and MB of healthy fertile (n=15) and RSA women (n=15) were sampled simultaneously on the second day of the menstrual cycle. Proportions of CD56+CD3−CD16+/−, CD56+CD3−CCR7+/−, and CD56+CD3−CD45RO+/− cells were analyzed using flow cytometry. In the MB of both groups, proportions of CD16+ and CD45RO− NK cells were significantly lower than in the PB. In parallel, CD56+CD16+CCR7− and CCR7+ cells were present in significantly smaller amounts in MB than in PB. However, the amounts of CD56+CD16−CCR7− and CCR7+ cells were greater in MB. In comparison to the fertile group, the percentage of MB CD45RO+ NK cells was significantly lower and frequencies of PB CD16−, CD45RO− and CD56+CD16+CCR7+ subsets were significantly higher in RSA patients. Different subsets of NK cells are differentially distributed in MB in comparison with PB in women with RSA and fertile subjects. Population differences of NK cell subsets in RSA patients and normal controls were more clearly reflected at the systemic level.

The Natural Killer Cell Interferon-Gamma Response to Bacteria Is Diminished in Untreated HIV-1 Infection and Defects Persist Despite Viral Suppression

Natural killer (NK) cells are important in innate immune responses to bacterial and viral pathogens. HIV-1 infection is associated with opportunistic bacterial infections and with microbial translocation, but the nature of the NK cell response to bacteria during HIV-1 infection has not been studied extensively. The objective of this study was to compare NK cell responses to bacteria in HIV-1-infected versus that in uninfected individuals. Multicolor flow cytometry was used to evaluate the ability of blood NK cell subsets (CD56CD16, CD56CD16, and CD56CD16) from treated, virally suppressed, and untreated viremic subjects with chronic HIV-1 infection and uninfected controls, to secrete interferon gamma (IFN-γ) in response to the in vitro stimulation of peripheral blood mononuclear cells with heat-killed commensal Escherichia coli or pathogenic Salmonella typhimurium. All 3 NK cell subsets produced IFN-γ in response to bacteria, but CD56CD16 NK cells were least responsive. Untreated HIV-1-infected donors had increased frequencies of CD56CD16 NK cells and lower overall frequencies of IFN-γ-producing NK cells responding to E. coli and S. typhimurium than did NK cells from uninfected donors. These NK cell defects were not fully restored in antiretroviral therapy-treated donors. Monocytes were necessary for NK cells to respond to bacteria, but the HIV-associated defect was intrinsic to NK cells because the addition of normal monocytes did not restore IFN-γ production in response to bacteria. Functional defects and numeric alterations of NK cell subsets lead to decreased frequencies of bacteria-reactive, IFN-γ-producing NK cells in HIV-1-infected subjects, even those on antiretroviral therapy.

Frequency and Phenotype of Human Circulating and Intrahepatic Natural Killer Cell Subsets Is Differentially Regulated according to Stage of Chronic Liver Disease

Background/Aims: Experimental liver injury models have indicated that natural killer (NK) cells are critical regulators of inflammation and fibrosis. However, data on NK cells and subsets in patients with liver diseases are limited. We thus comprehensively characterized peripheral and hepatic NK cell subsets in patients with chronic liver diseases (CLDs) of different etiologies and fibrosis stages. Methods: NK cells and other lymphocyte populations were characterized by FACS in 189 CLD patients (71 non-cirrhosis, 118 cirrhosis) and 153 healthy controls in blood and liver biopsies (n = 40). Results: In contrast to other lymphocyte subsets, circulating NK cells were generally reduced in CLD patients. Patients with fibrosis displayed a distinct increase of CD16^- NK cells in blood and of the CD16⁺ NK cell subset in liver. Patients with cirrhosis had overall lymphopenia, including reduced peripheral NK cells. Most pronounced shifts in NK cell subsets in blood and liver were found in cholestatic and autoimmune CLDs. Blood NK cells and subsets correlated with liver function, and inversely with fibrosis markers and inflammatory cytokines. Conclusions: The close association of human NK cells with disease severity and the intrahepatic accumulation of CD16⁺ NK cells in early fibrosis favor the concept of beneficial NK cell functions in hepatofibrogenesis.

Intracellular cytokine expression of peripheral blood natural killer cell subsets in women with recurrent spontaneous abortions and implantation failures

To investigate the cytokine expression by peripheral blood natural killer (NK) cells of women with recurrent spontaneous abortion (SAB) or implantation failures. Prospective cohort study. University clinic. Twenty-five women with recurrent SAB, 20 women with implantation failures, and 15 healthy controls. None. Cytokine expression (interferon-gamma, tumor necrosis factor [TNF]-alpha, interleukin [IL]-4, IL-5, IL-10, IL-13, granulocyte-macrophage colony-stimulating factor [GM-CSF]) in NK cells and their subsets (CD56(dim) and CD56(bright)). Proportion (percentage) of CD56(bright)/interferon-gamma(+)/TNF-alpha(+) cells was significantly higher in women with recurrent SAB and implantation failures as compared with that of healthy controls. Proportion of CD56(bright)/IL-4(+)/IL-10(+) cells was very low (<2%) in all groups but was significantly lower in women with recurrent SAB than that of controls. The TNF-alpha/GM-CSF expressing CD56(bright) cell ratio was significantly higher in women with recurrent SAB and implantation failures than in controls. Natural killer-1 shift in peripheral blood NK cells was identified in nonpregnant women with recurrent SAB and implantation failures. Tumor necrosis factor-alpha/GM-CSF expressing CD56(bright) cell ratio can be applicable for the diagnosis of recurrent SAB or implantation failures. Further studies are needed as to whether cytokine expression of NK cells during pregnancy can affect pregnancy outcome.

Trafficking of peripheral blood CD56 bright cells to the decidualizing uterus—new tricks for old dogmas?

CD56 bright lymphocytes become abundant in the human uterus during every menstrual cycle, following the surge in pituitary-derived luteinizing hormone (LH), which initiates final oocyte maturation. While the uterus is host to some CD56 bright cells prior to ovulation, the rapid increase is thought to be due to proliferation of the resident population, accompanied by recruitment of CD56 bright lymphocytes from the circulation. The rapid increase in CD56 bright cells is concurrent with the onset of decidualization, the transformation of uterine stromal cells into secretory decidual cells. Uterine CD56 bright cells proliferate and differentiate to become the predominant lymphocytes of the post-ovulatory uterus. These distinct, tissue-specific natural killer (NK) cells either die prior to menses or increase in number during early pregnancy, and then decline toward the end of the first trimester. Since lymphocytes home to tissues from the circulation, we investigated mechanisms of NK cell traffic over the course of natural menstrual cycles by measuring functional interactions between CD56+ cells from blood and endothelial cells using the Stamper–Woodruff assay of lymphocyte adhesion to frozen tissue sections. While a baseline level of adhesion was maintained throughout the cycle, elevated l-selectin-dependent adhesion of peripheral blood CD56 bright cells occurred during a peri-ovulatory window. However, there were no significant menstrual cycle-induced changes in the transcription of l-selectin, alpha 4 integrin or LFA-1, or in expression of these proteins by NK cells, suggesting that the enhanced adhesion was due to post-translational modifications of these molecules. Quantitative RT-PCR failed to amplify the message for LH receptor or the alpha or beta forms of progesterone or estrogen receptors from blood NK cell subsets. Thus, we conclude that the actions of LH, E 2, and P 4 on NK cells that promote interactions with endothelium and potential uterine homing are indirectly mediated through the responsiveness of other cell types.

Expression of Killer Immunoglobulin‐like Receptors on Peripheral Blood NK Cell Subsets of Women with Recurrent Spontaneous Abortions or Implantation Failures

Decidual natural killer (NK) cells express inhibitory receptors (killer immunoglobulin-like receptors, KIRs), which bind to ligands on trophoblast cells (human leucocyte antigen, HLA-C). This interaction appears to block NK cytotoxicity against trophoblast cells. In this study, we investigated the expression of inhibitory and activating receptors in peripheral blood NK cells of women with recurrent spontaneous abortion (RSA) or implantation failures. CD56(dim)/CD16(+), CD56(bright)/CD16(-) NK cells and CD56(+)/CD3(+) NKT cells of women with RSA or in vitro fertilization (IVF) failures and normal controls were analyzed for the expression of CD158a, CD158b inhibitory KIRs or CD161-activating receptors, by flow cytometric analysis. CD158a and CD158b inhibitory receptor expression by CD56(dim)/CD16(+) and CD56(bright)/CD16(-) NK cells were significantly decreased, and CD161-activating receptor expression by CD56(+)/CD3(+) NKT cells was significantly increased in women with implantation failures when compared with normal controls. An imbalance between inhibitory and activating receptor expression was found in NK cells of women with implantation failures. This imbalance may explain the adverse reproductive outcome.

Differential expression of KIR/NKAT2 and CD94 molecules on decidual and peripheral blood CD56 bright and CD56 dim natural killer cell subsets

Objective To investigate killer inhibitory receptor (KIR) expression by natural killer (NK) cells in early pregnancy. Design Case-control study of immunologic markers. Setting University hospital. Patient(s) Thirty pregnant women and 22 nonpregnant women. Intervention(s) Peripheral venous blood sampling and decidual tissue collection after elective abortion. Main outcome measure(s) Flow cytometry was used to assess expression of KIR by NK cells in the cell samples. Result(s) In contrast to CD56 bright peripheral blood NK cells, CD56 dim cells express killer cell Ig-like receptor KIR/NKAT2. However, KIR/NKAT2 and lectin-like CD94 are present on both subsets of decidual NK cells. We found no differences between peripheral blood NK cell subsets from pregnant and nonpregnant women. Conclusion(s) Our findings demonstrate that NK cell subsets, distributed in accordance with CD56 molecule density on cell surface, express killer inhibitory receptors CD94 and KIR/NKAT2 in a different way. Our data support the view that CD56 brightKIR/NKAT2 +CD94 + decidual NK cells are specialized NK cells that have an important role to play in early pregnancy.