Killer Cell Inhibitory Receptors Research Articles

Inhibition of Ly-49/LILRB-MHC-I interactions by anti-MHC-I augments innate and adaptive immunity.

Abstract Mouse NK cells recognize MHC-I antigens via stochastically expressed inhibitory Ly49 receptors. The loss of MHC-I expression on tumor cells (“missing self”) abrogates inhibitory signals, resulting in NK activation. We have identified an anti-mouse pan MHC-I mAb (M1/42), which blocks Ly49-MHC-I interactions but not TCR-MHC-I interactions. Administration of M1/42 in vivo markedly activated IFNg-producing NK cells that further drove the proliferation of NK cells and memory phenotype (MP) T cells and profoundly augmented adaptive immunity against viral infection and cancer metastasis. In contrast to mouse Ly-49 antigens, human killer cell inhibitory receptors bind to HLA at a site overlapping the TCR binding site and blocking of KIR binding to MHC-I would inhibit TCR recognition. However, two anti-pan-human MHC-I abs (DX17 and W6/32) markedly induced NK cell proliferation and IFNg production in cultures of human PBMC. DX17 potently blocked the interaction of leukocyte Ig-like inhibitory receptors LILRB1, B2, B3 and B5 with human MHC-I but had no effect on TCR-MHC-I interactions. The crystal structure of a DX17 Fab/MHC-I complex reveals that the footprint of DX17 overlaps the LILRB1 binding site on MHC-I. Administration of W6/32 Fab to humanized mice (PBMC reconstituted NSG or NK cell reconstituted NOG-IL-15) induced human NK and MP T cell proliferation. Marked reduction of tumor size was seen when NOG-hIL-15Tg mice were reconstituted with CD3 −cells and then treated with W6/32 Fab. DX17 Fc silenced mAb (DX17 LALAPG) activated human tumor infiltrating lymphocytes and monocytes. These results strongly suggest that inhibition of LILRB-MHC-I interactions by anti-MHC-I in humans may result in marked augmentation of anti-tumor immunity. This work was supported by the Intramural Research Program of NIAID, NIH.

Structure of the human NK cell NKR-P1:LLT1 receptor:ligand complex reveals clustering in the immune synapse

Signaling by the human C-type lectin-like receptor, natural killer (NK) cell inhibitory receptor NKR-P1, has a critical role in many immune-related diseases and cancer. C-type lectin-like receptors have weak affinities to their ligands; therefore, setting up a comprehensive model of NKR-P1-LLT1 interactions that considers the natural state of the receptor on the cell surface is necessary to understand its functions. Here we report the crystal structures of the NKR-P1 and NKR-P1:LLT1 complexes, which provides evidence that NKR-P1 forms homodimers in an unexpected arrangement to enable LLT1 binding in two modes, bridging two LLT1 molecules. These interaction clusters are suggestive of an inhibitory immune synapse. By observing the formation of these clusters in solution using SEC-SAXS analysis, by dSTORM super-resolution microscopy on the cell surface, and by following their role in receptor signaling with freshly isolated NK cells, we show that only the ligation of both LLT1 binding interfaces leads to effective NKR-P1 inhibitory signaling. In summary, our findings collectively support a model of NKR-P1:LLT1 clustering, which allows the interacting proteins to overcome weak ligand-receptor affinity and to trigger signal transduction upon cellular contact in the immune synapse.

Advances in generating HLA-Universal platelets for transfusion medicine

Objectives‘Universal’ blood typically refers to blood components that can be donated to most individuals. Allogenic platelet refractoriness is often caused by incompatibility of human leukocyte antigens (HLA) and antibodies in the recipient binding donor epitopes. HLA-matched platelets are not always readily available for transfusions, hindered by a general rise in platelet demand, logistical challenges of HLA-selected platelet provisions and the limited HLA repertoire of current stocks. Several groups have therefore progressed towards making in vitro platelets lacking HLA class I molecules, to avoid immunogenic reactions. This review will outline the progress to date in establishing these ‘HLA-universal platelets,’ taking a comparative view of the gene-editing technologies used for their creation while also evaluating the resulting products and the immunogenic properties of HLA-deficient cells. Key findingsUniversal platelets have been differentiated from HLA class I knockdown or knockout progenitors, generated mainly through gene modification techniques targeting B2M, which encodes the β2m-light chain component of HLA class I antigens. B2M-knockdown through RNA interference technology reduces but does not completely deplete HLA class I expression, whereas B2M-knockout via endonucleases (TALENs or CRISPR/Cas9 systems) creates HLA-negative cells. Cultured HLA-silenced or ablated megakaryocytes and platelets are functional and can circulate in mouse models, while evading immune detection. HLA class I proteins serve as ligands for Natural Killer (NK) cell inhibitory receptors. A key concern for patient welfare, using HLA-universal cells, is the triggering of immune reactions including NK cytotoxicity; therefore, a possible approach to circumvent this has been to either retain or co-express alternative inhibitory or immunoregulatory molecules. ConclusionsThe collection of studies brought together and reviewed here show proof-of-concept that functional HLA-universal platelets, inert to immune responses, can be produced using powerful gene-editing techniques. CRISPR/Cas9, in particular, offers versatility to co-engineer multiple specific gene knockouts in combination with options to knock in immunomodulating proteins or platelet factors, to correspondingly further enhance immune tolerance or improve platelet functions. Together these data demonstrate the rapid development and feasibility of moving HLA-universal platelets towards the clinic.

Natural Killer Cell Subpopulations and Inhibitory Receptor Dynamics in Myelodysplastic Syndromes and Acute Myeloid Leukemia.

Natural killer (NK) cells are key innate immunity effectors that play a major role in malignant cell destruction. Based on expression patterns of CD16, CD56, CD57, and CD94, three distinct NK cell maturation stages have been described, which differ in terms of cytokine secretion, tissue migration, and the ability to kill target cells. Our study addressed NK cell maturation in bone marrow under three conditions: a normal developmental environment, during pre-leukemic state (myelodysplastic syndrome, MDS), and during leukemic transformation (acute myeloblastic leukemia, AML). In this study, we used a new tool to perform multicolor flow cytometry data analysis, based on principal component analysis, which allowed the unsupervised, accurate discrimination of immature, mature, and hypermature NK subpopulations. An impaired NK/T cell distribution was observed in the MDS bone marrow microenvironment compared with the normal and AML settings, and a phenotypic shift from the mature to the immature state was observed in NK cells under both the MDS and AML conditions. Furthermore, an impaired NK cell antitumor response, resulting in changes in NK cell receptor expression (CD159a, CD158a, CD158b, and CD158e1), was observed under MDS and AML conditions compared with the normal condition. The results of this study provide evidence for the failure of this arm of the immune response during the pathogenesis of myeloid malignancies. NK cell subpopulations display a heterogeneous and discordant dynamic on the spectrum between normal and pathological conditions. MDS does not appear to be a simple, intermediate stage but rather serves as a decisive step for the mounting of an efficient or ineffective immune response, leading to either the removal of the tumor cells or to malignancy.

Quantification of low affinity binding interactions between natural killer cell inhibitory receptors and targeting ligands with a self-induced back-action actuated nanopore electrophoresis (SANE) sensor

A plasmonic nanopore sensor enabling detection of bimodal optical and electrical molecular signatures was fabricated and tested for its ability to characterize low affinity ligand-receptor interactions. This plasmonic nanosensor uses self-induced back-action (SIBA) for optical trapping to enable SIBA-actuated nanopore electrophoresis (SANE) through a nanopore located immediately below the optical trap volume. A natural killer (NK) cell inhibitory receptor heterodimer molecule CD94/NKG2A was synthesized to target a specific peptide-presenting Qa-1b Qdm ligand as a simplified model of low-affinity interactions between immune cells and peptide-presenting cancer cells that occurs during cancer immunotherapy. A cancer-irrelevant Qa-1b GroEL ligand was also targeted by the same receptor as a control experiment to test for non-specific binding. The analysis of different pairs of bimodal SANE sensor signatures enabled discrimination of ligand, receptor and their complexes and enabled differentiating between specific and non-specific ligand interactions. We were able to detect ligand-receptor complex binding at concentrations over 500 times lower than the free solution equilibrium binding constant (K D ). Additionally, SANE sensor measurements enabled estimation of the fast dissociation rate (k off) for this low-affinity specific ligand-receptor system, previously shown to be challenging to quantify with commercial technologies. The k off value of targeted peptide-presenting ligands is known to correlate with the subsequent activation of immune cells in vivo, suggesting the potential utility of the SANE senor as a screening tool in cancer immunotherapy.

Control of Viral Infection by Natural Killer Cell Inhibitory Receptors

SUMMARYMajor histocompatibility complex class I (MHC-I)-restricted immune responses are largely attributed to cytotoxic T lymphocytes (CTLs). However, natural killer (NK) cells, as predicted by the missing-self hypothesis, have opposing requirements for MHC-I, suggesting that they may also demonstrate MHC-I-restricted effects. In mice, the Ly49 inhibitory receptors prevent NK cell killing of missing-self targets in effector responses, and they have a proposed second function in licensing or educating NK cells via self-MHC-I in vivo. Here we show MHC-I-restricted control of murine cytomegalovirus (MCMV) infection in vivo that is NK cell dependent. Using mice lacking specific Ly49 receptors, we show that control of MCMV requires inhibitory Ly49 receptors and an inhibitory signaling motif and the capacity for MCMV to downregulate MHC-I. Taken together, these data provide definitive evidence that the inhibitory receptors are required for missing-self rejection and are relevant to MHC-I-restricted NK cell control of a viral infection in vivo.

Targeting NK Cell Checkpoint Receptors or Molecules for Cancer Immunotherapy.

Checkpoint blockade therapy, for example using antibodies against CTLA-4 and PD-1/PD-L1, relieves T cells from the suppression by inhibitory checkpoints in the tumor microenvironment; thereby achieving good outcomes in the treatment of different cancer types. Like T cells, natural killer (NK) cell inhibitory receptors function as checkpoints for NK cell activation. Upon interaction with their cognate ligands on infected cells, tumor cells, dendritic cells and regulatory T cells, signals from these receptors severely affect NK cells' activation and effector functions, resulting in NK cell exhaustion. Checkpoint inhibition with antagonistic antibodies (Abs) can rescue NK cell exhaustion and arouse their robust anti-tumor capacity. Most notably, the response to anti-PD-1 therapy can be enhanced by the increased frequency and activation of NK cells, thereby increasing the overall survival of patients with multiple types of cancer. In addition, rescue of NK cell activity could enhance adaptive T cells' anti-tumor activity. Some antagonistic Abs (e.g., anti-TIGIT and anti-NKG2A monoclonal Abs) have extraordinary potential in cancer therapy, as evidenced by their induction of potent anti-tumor immunity through recovering both NK and T cell function. In this review, we summarize the dysfunction of NK cells in the tumor microenvironment and the key NK cell checkpoint receptors or molecules that control NK cell function. We particularly focus on recent advances in the most promising strategies through blockade of NK cell checkpoints or their combination with other approaches to more effectively reject tumors.

Natural Killer Cells Offer Differential Protection From Leukemia in Chinese Southern Han.

Interactions of human natural killer (NK) cell inhibitory receptors with polymorphic HLA-A, -B and -C molecules educate NK cells for immune surveillance against tumor cells. The KIR A haplotype encodes a distinctive set of HLA-specific NK cell inhibiting receptors having strong influence on immunity. We observed higher frequency of KIR A homozygosity among 745 healthy Chinese Southern Han than 836 adult patients representing three types of leukemia: ALL (OR = 0.68, 95% CI = 0.52–0.89, p = 0.004), AML (OR = 0.76, 95% CI = 0.59–0.98, p = 0.034), and CML (OR = 0.72 95% CI = 0.51–1.0, ns). We observed the same trend for NHL (OR = 0.47 95% CI = 0.26–0.88 p = 0.017). For ALL, the protective effect of the KIR AA genotype was greater in the presence of KIR ligands C1 (Pc = 0.01) and Bw4 (Pc = 0.001), which are tightly linked in East Asians. By contrast, the C2 ligand strengthened protection from CML (Pc = 0.004). NK cells isolated from KIR AA individuals were significantly more cytotoxic toward leukemic cells than those from other KIR genotypes (p < 0.0001). These data suggest KIR allotypes encoded by East Asian KIR A haplotypes are strongly inhibitory, arming NK cells to respond to leukemogenic cells having altered HLA expression. Thus, the study of populations with distinct KIR and HLA distributions enlightens understanding of immune mechanisms that significantly impact leukemia pathogenesis.

The Role of Soluble HLA-G in the Vertical Transmission of Toxoplasma gondii

Soluble human leukocyte antigen G (sHLA-G) plays a key role in pregnancy through interaction with decidual natural killer (dNK) cell inhibitory receptors at the maternal-fetal interface. To demonstrate the possible role of sHLA-G during the pregnancy with Toxoplasma gondii infection, we compared the concentration of a murine functional homolog of sHLA-G, Qa-2, in T. gondii infected and non-infected pregnant C57BL/6 mice, and that of sHLA-G in BeWo culture supernatant. In addition, the levels of KIR2DL4 expressed on human dNK cells and NKG2A in pregnant mice were evaluated. We showed that T. gondii infection result in significant increase in the level of Qa-2 and NKG2A in pregnant mice. sHLA-G and KIR2DL4 in human samples were also significantly upregulated under the condition of T. gondii infection. The further treatment with sHLA-G antibody could reduce the expression level of KIR2DL4 which was upregulated by T. gondii infection. In summary, sHLA-G could upregulate the expression level of KIR2DL4 which lead to excessive immunological tolerance, and further contributed to T. gondii immunity escaping and affecting fetus via vertical transmission which may lead to adverse outcomes.

Recognition of host Clr-b by the inhibitory NKR-P1B receptor provides a basis for missing-self recognition

The interaction between natural killer (NK) cell inhibitory receptors and their cognate ligands constitutes a key mechanism by which healthy tissues are protected from NK cell-mediated lysis. However, self-ligand recognition remains poorly understood within the prototypical NKR-P1 receptor family. Here we report the structure of the inhibitory NKR-P1B receptor bound to its cognate host ligand, Clr-b. NKR-P1B and Clr-b interact via a head-to-head docking mode through an interface that includes a large array of polar interactions. NKR-P1B:Clr-b recognition is extremely sensitive to mutations at the heterodimeric interface, with most mutations severely impacting both Clr-b binding and NKR-P1B receptor function to implicate a low affinity interaction. Within the structure, two NKR-P1B:Clr-b complexes are cross-linked by a non-classic NKR-P1B homodimer, and the disruption of homodimer formation abrogates Clr-b recognition. These data provide an insight into a fundamental missing-self recognition system and suggest an avidity-based mechanism underpins NKR-P1B receptor function.

Abstract 4624: Natural killer cell licensing contributes to clearance of lung cancer

Abstract a) Natural Killer (NK) cell Inhibitory receptors function in a dichotomous role. Their engagement by MHC Class I upon encountering a target results in the inhibition of cytotoxicity. During NK development, however, their interaction with MHC Class I mediates the process of licensing or education, leading to an increase in responsiveness and cytotoxicity. For this reason the role of inhibitory receptors in controlling the clearance of MHC Class I+ solid tumors such as lung cancer is poorly defined. Since NK cells comprise the primary cytotoxic lymphocyte responsible for clearance of lung cancer, we set out to determine whether the licensing status of NK cells contributes to lung cancer susceptibility. b) Growth of MHC Class I+ murine lung cancer was evaluated in MHC Class I- mice, with unlicensed NK cells in direct comparison to MHC Class I+ mice with licensed NK cells. IFN-γ production was evaluated after plate bound antibody stimulation through the NKp46 and NKG2D pathways both flow cytometrically and by quantitative RT-PCR. Bone marrow transplantation was performed by injection of 10x106 unfractionated bone marrow cells after 500cGy irradiation and NK and CD4+ T cell depletion of recipient host. c) Injection of Lewis Lung Carcinoma (LLC) revealed a significant decrease in pulmonary tumor burden in mice with licensed NK cells (230.4±21.6 vs. 442.7±53.8 mg) that disappeared after NK depletion (421.4±36.8 vs. 421.9±66.3 mg). Freshly isolated licensed NK cells lysed LLC more robustly than unlicensed NK cells (28.7±.16% vs. 4.9±.2% specific lysis at 100:1 ratio respectively). Lung cancer expresses stress ligands recognized by NKG2D and NKp46 activating receptors. Licensed NK cells produced significantly more IFN-γ than their unlicensed counterparts after plate-bound stimulation with anti-NKp46 and anti-NKG2D (17.9±2.8% vs. 5.4±.8% respectively for dual NKp46 and NKG2D stimulation). No differences were evident between licensed and unlicensed NK cells in ribosomal mass, making it unlikely that translational defects were responsible for altered cytokine production. Licensing status also did not affect stability of IFN-γ but did affect mRNA levels. This suggests transcriptional control of licensing-mediated NK activity. IFN-γ production could be significantly increased in unlicensed NK cells by reintroduction of MHC Class I through non-myeloablative bone marrow transplantation from MHC Class I-sufficient congenic donors. Blockade of H2Kb-Ly49c (MHC Class I-inhibitory receptor) interaction during NK development in C57BL/6 mice using a non-depleting anti-Ly49c antibody (clone 4LO) decreased IFN-γ production in NK cells. d) The role of NK cell licensing and tumor-specific cytotoxicity is highly controversial. We now demonstrate that licensed NK cells mediate a superior lung cancer-specific immune response in a murine model. Our data suggests that altering the licensing status of NK cells may be an avenue of lung cancer-specific immunotherapy. Citation Format: Shi Lei, Kang Li, Saeed Arefanian, Alexander S. Krupnick. Natural killer cell licensing contributes to clearance of lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4624. doi:10.1158/1538-7445.AM2017-4624

Expression of Natural Killer Cell Inhibitory Receptors is Associated with Significant Liver Injury in Chronic Hepatitis C in Children

Introduction and aimNatural Killer (NK) cells play an important role in innate immune response to viral infections and their high proportion is situated in the liver. The aim of this study was to analyze possible relation between the expression of NK cell receptors and varied intensity of liver lesions in chronic hepatitis C (CHC) in children. Material and methodsStudy included 105 children with CHC - 54 boys and 51 girls, age 13.62 +3.48 years. Blood specimens were taken at the day of the liver biopsy. Histological evaluation was performed according to METAVIR scoring system. Circulating NK cells were evaluated by flow cytometry. The results were shown as a proportion of cells expressing evaluated receptor and its’ mean fluorescent intensity (MFI). ResultsIn 58 children with CHC (55.2%) significant liver fibrosis was observed (>F2). Higher proportion of cells expressing CD158e inhibitory receptors was observed in the group of children with ALT > 2UNL (21.11 + 14.60 vs. 12.22 +8.99%; p = 0.037). While higher proportion of cells expressing inhibitory CD158b receptor was observed in children with significant fibrosis (F >2) compared to minimal fibrosis (F < 2) - (34.14 +12.44 vs. 27.48+8.71%; p = 0.049). Children with advanced fibrosis (F >3) had higher MFI of NK cell CD 158b receptor than children with fibrosis scored F < 3 - (5344.20 +3407.49 vs. 2979.67 +1190.64; p = 0.049). Proportion of NK cells expressing CD158b was found a predictor of significant fibrosis in univariate analysis - [OR 1.065; 95%CI (1.07-1.15); p = 0.046]. ConclusionsHigher proportion of NK cells expressing inhibitory CD158b and CD158e receptors is associated with significant liver injury.

Lack of Siglec-7 expression identifies a dysfunctional natural killer cell subset associated with liver inflammation and fibrosis in chronic HCV infection

ObjectiveSialic-acid-binding immunoglobulin-like lectin-7 (Siglec-7) is a natural killer (NK) cell inhibitory receptor associated with NK phenotypic and functional abnormalities in HIV-1 infection. We investigated the significance of NK-expressed and serum...

Daratumumab-mediated lysis of primary multiple myeloma cells is enhanced in combination with the human anti-KIR antibody IPH2102 and lenalidomide.

Despite recent treatment improvements, multiple myeloma remains an incurable disease. Since antibody-dependent cell-mediated cytotoxicity is an important effector mechanism of daratumumab, we explored the possibility of improving daratumumab-mediated cell-mediated cytotoxicity by blocking natural killer cell inhibitory receptors with the human monoclonal anti-KIR antibody IPH2102, next to activation of natural killer cells with the immune modulatory drug lenalidomide. In 4-hour antibody-dependent cell-mediated cytotoxicity assays, IPH2102 did not induce lysis of multiple myeloma cell lines, but it did significantly augment daratumumab-induced myeloma cell lysis. Also in an ex vivo setting, IPH2102 synergistically improved daratumumab-dependent lysis of primary myeloma cells in bone marrow mononuclear cells (n=21), especially in patients carrying the FcγRIIIa-158F allele or the FcγRIIa-131R allele, who bind IgG1 with lower affinity than patients carrying the FcγRIIIa-158V allele or the FcγRIIa-131H allele. Finally, a further synergistically improved myeloma cell lysis with the daratumumab-IPH2102 combination was observed by adding lenalidomide, which suggests that more effective treatment strategies can be designed for multiple myeloma by combining daratumumab with agents that independently modulate natural killer cell function.

Differential Expression of Immune-Modulatory Molecule HLA-E in Non-Neoplastic and Neoplastic Lesions of the Thyroid

Human leukocyte antigen (HLA)–E is a non-classical molecule of the histocompatibility complex that functions as one of the main ligands of the Natural Killer (NK) cell inhibitory receptor CD94/NKG2A and inhibits its potent cytotoxic activity. Due to the important role of NK cells in combating neoplasm, we hypothesized that the differential expression of HLA-E could favor the progression of heterogeneous thyroid tumors.Using an immunohistochemistry technique in 143 biopsies of thyroid tumors, including benign and malignant neoplasms and goiters, we evaluated the expression of HLA-E among various tumor types and its association with the clinicopathological factors of diseases. We verified high HLA-E expression in all types of neoplastic tumors, although no significant differences between the groups were found. Low expression was observed in 95 percent of the goiter samples, showing significant differences between neoplastic and non-neoplastic lesions. Furthermore, a significant result was found with regard to the tumor size, with high HLA-E expression being related to smaller tumors. Therefore, our data suggest that an increase in HLA-E may be associated with the establishment of thyroid neoplasms, with either benign or malignant features.

Natural Killer Cell Inhibitory Receptor Expression in Humans and Mice: A Closer Look

The Natural Killer (NK) cell population is composed of subsets of varying sizes expressing different combinations of inhibitory receptors for MHC class I molecules. Genes within the NK gene complex, including the inhibitory receptors themselves, seem to be the primary intrinsic regulators of inhibitory receptor expression, but the MHC class I background is an additional Modulating factor. In this paper, we have performed a parallel study of the inhibitory receptor repertoire in inbred mice of the C57Bl/6 background and in a cohort of 44 humans. Deviations of subset frequencies from the “product rule (PR),” i.e., differences between observed and expected frequencies of NK cells, were used to identify MHC-independent and MHC-dependent control of receptor expression frequencies. Some deviations from the PR were similar in mice and humans, such as the decreased presence of NK cell subset lacking inhibitory receptors. Others were different, including a role for NKG2A in determining over- or under-representation of specific subsets in humans but not in mice. Thus, while human and murine inhibitory receptor repertoires differed in details, there may also be shared principles governing NK cell repertoire formation in these two species.

Analysis of immunoglobulin-like transcripts (ILTs) in lymphocytes with sHLA-G and IL10 from SLE patients

The aim of this work was to study the expression of human leukocyte antigen G (HLA-G) and interleukin 10 (IL-10) in conjunction with expression of HLA-G killer-cell inhibitory receptor ligand immunoglobulin-like transcript 2 (ILT2) in CD3+, CD19+, CD56+ lymphomas, and ILT4 in CD14+ cells from patients with systemic lupus erythematosus (SLE). Thirty-one SLE patients and 36 healthy controls were studied. ILTs expression was analyzed by flow cytometry in peripheral blood mononuclear cells (PBMCs). The plasma sHLA-G and IL10 were evaluated by enzyme-linked immunosorbent assay (ELISA). We found a significant increased expression of ILT2 by lymphocytes in SLE patients. When the expression of this receptor was assessed in cell subsets, significantly higher ILT2 MRFI levels were detected in CD3+ cells, CD19+ cells, CD56+ cells (P<0.05), but no change with ILT4 MRFI in CD14+ cells, neither did the percentages of ILT2/4+ lymphocytes change in SLE patients compared with healthy controls (P>0.05). The upregulation of ILT2 expression was related to IL10 and anti-ds-DNA antibodies (P<0.05), but not sHLA-G and steroid therapy (P>0.05). IL-10 and sHLA-G were increased, but did not change remarkably (P>0.05); however, they were quite related (P<0.05). ILT2 might be one of the factors accounting for the evasion of immunosurveillance, thus participate in the pathogenesis of SLE, and the upregulation of ILT2 may be associated with its disease activity.

The Adaptor Protein Crk Controls Activation and Inhibition of Natural Killer Cells

Natural killer (NK) cell inhibitory receptors recruit tyrosine phosphatases to prevent activation, induce phosphorylation and dissociation of the small adaptor Crk from cytoskeleton scaffold complexes, and maintain NK cells in a state of responsiveness to subsequent activation events. How Crk contributes to inhibition is unknown. We imaged primary NK cells over lipid bilayers carrying IgG1 Fc to stimulate CD16 and human leukocyte antigen (HLA)-E to inhibit through receptor CD94-NKG2A. HLA-E alone induced Crk phosphorylation in NKG2A(+) NK cells. At activating synapses with Fc alone, Crk was required for the movement of Fc microclusters and their ability to trigger activation signals. At inhibitory synapses, HLA-E promoted central accumulation of both Fc and phosphorylated Crk and blocked the Fc-induced buildup of F-actin. We propose a unified model for inhibitory receptor function: Crk phosphorylation prevents essential Crk-dependent activation signals and blocks F-actin network formation, thereby reducing constraints on subsequent engagement of activation receptors.

Expression patterns of NKG2A, KIR, and CD57 define a process of CD56dim NK-cell differentiation uncoupled from NK-cell education

AbstractNatural killer (NK) cells are lymphocytes of the innate immune system that, following differentiation from CD56bright to CD56dim cells, have been thought to retain fixed functional and phenotypic properties throughout their lifespan. In contrast to this notion, we here show that CD56dim NK cells continue to differentiate. During this process, they lose expression of NKG2A, sequentially acquire inhibitory killer cell inhibitory immunoglobulin-like receptors and CD57, change their expression patterns of homing molecules, and display a gradual decline in proliferative capacity. All cellular intermediates of this process are represented in varying proportions at steady state and appear, over time, during the reconstitution of the immune system, as demonstrated in humanized mice and in patients undergoing hematopoietic stem cell transplantation. CD56dim NK-cell differentiation, and the associated functional imprint, occurs independently of NK-cell education by interactions with self–human leukocyte antigen class I ligands and is an essential part of the formation of human NK-cell repertoires.

Multiple, Non-conserved, Internal Viral Ligands Naturally Presented by HLA-B27 in Human Respiratory Syncytial Virus-infected Cells

Cytotoxic T lymphocyte (CTL)-mediated death of virus-infected cells requires prior recognition of short viral peptide antigens that are presented by human leukocyte antigen (HLA) class I molecules on the surface of infected cells. The CTL response is critical for the clearance of human respiratory syncytial virus (HRSV) infection. Using mass spectrometry analysis of complex HLA-bound peptide pools isolated from large amounts of HRSV-infected cells, we identified nine naturally processed HLA-B27 ligands. The isolated peptides are derived from six internal, not envelope, proteins of the infective virus. The sequences of most of these ligands are not conserved between different HRSV strains, suggesting a mechanism to explain recurrent infection with virus of different HRSV antigenic subgroups. In addition, these nine ligands represent a significant fraction of the proteome of this virus, which is monitored by the same HLA class I allele. These data have implications for vaccine development as well as for analysis of the CTL response.