Antibody- and interferon-dependent killer cells are part of the NK cell receptor positive subpopulation of human peripheral blood cells.

Cytotoxicity by human non-adherent peripheral blood lymphocytes was analysed after effector cell activation with either interferon (IF) or by target cell specific IgG antibodies (T-IgG). Four different cell lines were used as target cells that differed in susceptibility to natural killer cell (NK) activity; a highly susceptible T cell line (Molt-4), a medium susceptible B lymphoma line (Daudi), a resistant B cell line established by Epstein-Barr virus transformation (LCL-LS) and a resistant mouse mastocytoma line (P815). Three different parameters influencing killing were investigated; lytic potential, target cell binding and efficiency of the lytic phase from which the absolute number of effector cells and their recycling capacity could be estimated. It was found that, when using human target cell lines, IF and T-IgG influenced the system in a similar way by activating the lytic phase and the effector cell recycling but not the early binding phase. With the NK resistant mouse mastocytoma cell line P815 a comparatively small target binding population was found which, however, increased markedly with T-IgG treatment. Taken together, the results indicate that the effector population responsible for antibody-induced killing belong to a subpopulation of cells that have the ability to spontaneously conjugate to the present target cells by virtue of naturally occuring undefined cell surface receptors designated NKR (NK receptor) and that the role of T-IgG in the present system is similar to that of IF. In contrast, if target cells are used that do not express binding structures for NKR receptors, T-IgG may also fulfill a receptor function through Fc receptors for IgG.     

肝癌Bel7402细胞HLA-E基因上调对NK细胞体外杀伤作用的影响

目的: 探讨肝癌Bel7402细胞人类白细胞抗原-E(HLA-E)基因上调对NK细胞体外杀伤作用的影响。方法: 体外构建Lentivirus/CMV/GFP-HLA-E慢病毒表达载体并转导肝癌Bel7402细胞,采用RT-PCR和Western blotting方法检测肝癌Bel7402细胞HLA-E基因mRNA水平和蛋白水平的表达,分析肝癌Bel7402细胞HLA-E基因上调及HLA-ABC抗体封闭靶细胞相应位点对NK细胞体外杀伤作用的影响。结果: Lentivirus/CMV/GFP-HLA-E重组慢病毒载体感染Bel7402细胞后不同时段(24 h、48 h、72 h、96 h)的HLA-E mRNA的表达明显上调,除在感染后24 h(P＜0.05)外,慢病毒过表达组在感染后48 h、72 h、96 h与Bel7402组比较,显著差异 (P＜0.01)。蛋白水平的表达亦明显上调:24 h、48 h、72 h、96 h外源性/内源性HLA-E吸光度比值与12 h比较显著差异 (P＜0.01)。未封闭的Bel7402组和Bel7402 Lenti HLA-E组比较,NK细胞的杀瘤活性差异显著(P＜0.05或P<0.01);封闭组和未封闭组比较,NK细胞的杀瘤活性差异显著(P＜0.05);Bel7402(封闭组) 和Bel7402 Lenti HLA-E(封闭组)比较,NK细胞的杀瘤活性有显著差异(P＜0.01)。结论: Lentivirus/CMV/GFP-HLA-E慢病毒表达载体能有效增加HLA-E的表达。NK细胞对HLA-E基因表达上调的Bel7402细胞的靶杀伤作用降低;抗HLA-ABC单抗封闭靶细胞相应位点后,NK细胞对靶细胞的杀伤能力普遍有所提高。     

C-reactive protein is involved in natural killer cell-mediated lysis but does not mediate effector-target cell recognition.

Anti-CRP and complement treatment of human peripheral blood lymphocytes significantly reduces natural killer (NK) cell-mediated cytotoxicity to K562 target cells as well as to MOLT-4 target cells. Although not all activity is eliminated by treatment of effector cells with antibody and complement, the reduction of NK function indicates that C-reactive protein (CRP) is present on a significant proportion of NK cells. Higher concentrations of anti-CRP or anti-CRP F(ab')2 fragments also reduce NK function; this suggests that CRP is not only present on these effector cells but may also play a role in NK-mediated killing. We initially suspected that CRP-ligand interactions might be involved in effector-target cell recognition. Several lines of evidence suggest that this is not the case. While F(ab')2 anti-CRP will block NK function, Fab anti-CRP will not, suggesting that the NK response is not impaired when surface CRP (S-CRP) is blocked but is only inhibited when the S-CRP is cross-linked and modulated. Neither CRP-C polysaccharide complexes (CRP-CPS) nor concentrations of CPS ranging from 0.1 microgram/ml to 200 micrograms/ml have any effect on NK cell-mediated killing. Treatment of target cells with a ligand for CRP or CRP prior to co-culture with NK effectors does not augment NK function. Single cell assays clearly demonstrate that high concentrations of anti-CRP have no effect on the formation of effector-target cell conjugates. Although these concentrations of anti-CRP do not block effector-target cell conjugation in the single cell assay, they do block the killing of conjugated target cells. In total, this evidence strongly suggests that although CRP appears to be involved in NK-mediated killing, it is not involved in effector-target cell-mediated recognition.     

Apoptosis induced by NK cells is modulated by the NK-active cytokines IL-2 and IL-12

NK cells kill sensitive targets via exocytosis of cytoplasmic granules containing membrane damaging perforins and DNA damaging granzymes. Therefore, the target cell can either die by necrosis or by apoptosis. A third, non-secretory, mechanism of killing mediated by Fas-FasL interaction can be used by activated NK cells to kill Fas+ targets. Here, we have studied the modulation exerted by two NK active cytokines, IL-2 and IL-12, on the apoptosis-inducing activity of NK cells in NK-sensitive Fas- K562 and NK-insensitive Fas+ Raji cells. Our results show that apoptosis is preferentially induced at low target:effector ratios. Resting NK cells virtually do not induce apoptosis in target cells while IL-2 stimulation endows NK cells with the capacity of inducing apoptosis and IL-12 further enhances this effect against both targets. Finally, we demonstrate that cytokine- stimulated NK cells use both granzyme and Fas-FasL pathways of apoptosis induction.      

The two‐pore domain K2P channel TASK2 drives human NK‐cell proliferation and cytolytic function

Natural killer (NK) cells are a subset of cytotoxic lymphocytes that recognize and kill tumor‐ and virus‐infected cells without prior stimulation. Killing of target cells is a multistep process including adhesion to target cells, formation of an immunological synapse, and polarization and release of cytolytic granules. The role of distinct potassium channels in this orchestrated process is still poorly understood. The current study reveals that in addition to the voltage‐gated K_V1.3 and the calcium‐activated K_Ca3.1 channels, human NK cells also express the two‐pore domain K₂P channel TASK2 (TWIK‐related acid‐sensitive potassium channel). Expression of Task2 varies among NK‐cell subsets and depends on their differentiation and activation state. Despite its different expression in TASK2^highCD56^brightCD16^? and TASK2^lowCD56^dimCD16⁺ NK cells, TASK2 is involved in cytokine‐induced proliferation and cytolytic function of both subsets. TASK2 is crucial for leukocyte functional antigen (LFA‐1) mediated adhesion of both resting and cytokine‐activated NK cells to target cells, an early step in killing of target cells. With regard to the following mechanism, TASK2 plays a role in release of cytotoxic granules by resting, but not IL‐15‐induced NK cells. Taken together, our data exhibit two‐pore potassium channels as important players in NK‐cell activation and effector function.     

Interferon-Induced NK Augmentation in Humans

By combining a single-cell cytotoxicity assay in agarose with estimations of the maximal natural killer (NK) cell potential (Vmax) by 51Cr release, the mechanism behind interferon augmentation of human NK cells were analysed. The number of target-binding cells (TBCs) the fraction of active TBCs and NK cell recycling were studied after short-term interferon treatment. The results demonstrate a dual effect of interferon on human NK cells: effector cell recruitment and increased effector cell recycling. Both of these variables were increased when NK cells were tested against the standard target K-562 and against Daudi and BJAB cells, derived from B-type lymphomas. However, when T cell lines derived from acute lymphocytic leukaemia (Molt-4 and 1310) were used as targets, a larger fraction of active NK cells were found among untreated TBCs, whereas interferon treatment only resulted in increased effector cell recycling and not in effector cell recruitment. No increase in TBCs after interferon treatment could be detected with any cell line tested. The difference seen between T and non-T cell lines with regard to interferon-induced effector cell recruitment is discussed in relation to known characteristics of the human NK system.     

Human natural killer-like activity against mouse spleen cells

Unstimulated human peripheral lymphocytes were cytotoxic for normal mouse spleen cells and suppressed the in vitro antibody plaque-forming cell (PFC) response of these cells to sheep red blood cells and dinitrophenylated Ficoll. The cells in the lymphocyte population that were responsible for the immunosuppression had properties of natural killer (NK) or NK-like cells in that they were: (a) non-E-rosetting, (b) nonadherent, (c) unaffected by treatment with anti-human immunoglobulin plus complement, (d) cytotoxic against an established human NK target, K562 leukemic cells, and (e) partially inactivated by mitomycin C. Addition of the human NK-like cells to mouse spleen cell cultures at the time of antigen addition and at an effector cell to target cell ratio as low as 0.67:1 resulted in 85 to 96% suppression of the PFC response. Addition of NK-like cells to cultures 18 h before harvesting in 5-day cultures required higher concentrations and ratios (2.7 : 1) of effector to target cells to significantly suppress the PFC response. The data suggest that human NK-like activity in suppression of the mouse PFC response is due to killing of the targets. The mouse spleen cell PFC system represents a potential model for assessment of human NK activity that is quite dramatic in its effect and can be used in addition to the well known ⁵¹Cr-release assay. Also, since the mouse spleen cell is a normal cell, it provides a model in the PFC system for studying the mechanism of NK regulation of normal cellular function. An additional finding of this study was the observation that E-rosetting T cells significanty enhanced the mouse PFC response. Thus, human peripheral lymphocytes contain discrete cellular populations that either enhance or suppress the mouse PFC response.     

Vγ2Vδ2 T-cell co-stimulation increases natural killer cell killing of monocyte-derived dendritic cells

Interactions between natural killer (NK) cells and dendritic cells (DC) affect maturation and function of both cell populations, including NK cell killing of DC (editing), which is important for controlling the quality of immune responses. We also know that antigen-stimulated Vγ2Vδ2 T cells co-stimulate NK cells via 4-1BB to enhance the killing of tumour cell lines but we do not know what regulates 4-1BB expression or whether other NK effector functions including DC killing, might also be influenced by NK–γδ T-cell cross-talk. Here we show that antigen-stimulated γδ T cells co-stimulate NK cells through inducible T-cell co-stimulator (ICOS)– ICOS ligand (ICOSL) and this signal increases NK cell killing of autologous DC. Effects of NK–γδ T-cell co-culture, which could be reproduced with soluble ICOS-Fc fusion protein, included increased CD69 and 4-1BB expression, interferon-γ, tumour necrosis factor-α, macrophage inflammatory protein-1β, I-309, RANTES and sFas ligand production, as well as elevated mRNA levels for co-stimulatory receptors OX40 (TNFRSF4) and GITR (TNFRSF18). Hence, ICOS–ICOSL co-stimulation of NK by Vγ2Vδ2 T cells had broad effects on NK phenotype and effector functions. The NK–γδ T-cell cross-talk links innate and antigen-specific lymphocyte responses in the control of cytotoxic effector function and DC killing.     

Multiple natural killer cell-activating signals are inhibited by major histocompatibility complex class I expression in target cells

Several lines of evidence indicate that major histocompatibility complex class I molecules expressed by target cells can prevent natural killer cell (NK) lysis, possibly by engaging inhibitory receptors expressed by NK cells. On the other hand it is likely that NK cells must be activated to lysis by the recognition of unidentified NK target structures on target cells. To investigate the relationship between positive activation of NK cells by NK target structures versus inhibition by target cell class I molecules, we have examined various NK/target cell interactions for which the expression of inhibitory class I molecules by the target cells is known. The results suggest that specific properties of the target cell other than the absence of class I expression are necessary to activate NK-mediated lysis. Furthermore, different effector cell populations, i.e. freshly isolated versus interleukin-2 activated NK cells, differ in their capacity to kill class I-deficient lymphoblast target cells. In general, class I-deficient target cells that are resistant to direct lysis by a given NK population can be lysed by the NK cells when the reaction is mediated by antibody-dependent cellular cytotoxicity (ADCC). Most significantly, all types of NK-mediated lysis of lymphoblasts, of tumor cells and of almost any target by ADCC can be inhibited by appropriate class I gene expression in the target cell. These results suggest a model in which lysis by NK cells must be triggered by any one of a set of distinct target cell ligands, but that all of these signals can be overruled by class I-mediated inhibition.     

Autoantibodies as Predictive and Diagnostic Markers of Idiopathic Inflammatory Myopathies

The purpose of the present study was to determine whether patients with euthyroid Graves’ exophthal-mopathy have an impaired NK cell function compared to patients with Graves’ hyperthyroidism and healthy controls.

The NK cell activity measured against K562 target cells was significantly suppressed (p < 0.01) in patients with euthyroid Graves’ ophthalmopathy, whereas the NK cell activity of patients with Graves’ hyperthyroidism was not. Although interferon-α, interleukin-2 and indomethacin significantly enhanced (p < 0.01) the NK cell activity in all three groups, none of these agents fully restored the defective NK cell activity in euthyroid Graves’ ophthalmopathy. The concentrations in the blood of large granular lymphocytes and CD 16 positive cells did not differ between the three groups, furthermore an immunosuppressive serum factor was not detected. The number of effector/target cell conjugates did not differ between patients and controls, whereas the interferon-α -induced production of a soluble natural killer cytotoxic factor (NKCF) with specificity for NK sensitive target cells was suppressed in patients with Graves’ euthyroid ophthalmopathy. We conclude that one of the mechanisms underlying the defective NK cell activity in patients with euthyroid ophthalmopathy may be an impairment of the release of NKCF from the NK cells.     

Natural killer cell function in patients with acquired immunodeficiency syndrome and related diseases

This review describes current knowledge of changes in natural killer (NK) cell function in acquired immunodeficiency syndrome (AIDS)-related disorders, vis-à-vis associated abnormalities in NK cytolytic function, NK cell subset distribution, NK cytopathology, and lymphokine regulation. NK cells, which are closely associated with large granular lymphocytes, are spontaneously cytotoxic to tumor and virally infected targets. As such, they may play a role in natural resistance to human immunodeficiency virus type 1 (HIV-1)-associated disorders and other opportunistic infections. Yet, peripheral blood NK activity is frequently reduced in patients with HIV-1-induced disease. NK cells are heterogeneous both with respect to their expression of serologically defined membrane antigens and functional activity. In AIDS-related syndromes, there appears to be a diminution of the NK pool (CD16+ cells) involved in cytolytic function, while there is an elevation of the NK pool that coexpresses NK (Leu 7+) and T (CD8+) cell markers, which show little or no involvement in cytolytic function. The impairment of in vitro NK function is not associated with a reduced frequency of lytic conjugates of effectors and target cells nor with the recycling capacity of these effector cells but rather is associated with defects in the NK cell lytic machinery following formation of such conjugates. NK cells in AIDS patients show an impairment in effector cell microtubule rearrangement following target cell interaction. The causes of NK cell dysfunction in AIDS-related disorders remain unknown. NK cells do not appear to express the CD4 epitope of the HIV receptor, nor have they been demonstrated to be susceptible to infection by HIV-1. There appears to be a preponderance of immature NK cells and a lymphokine imbalance in patients with HIV-1 associated disease. Interleukin-2 can partially restore diminished in vitro NK function. Elucidation of the involvement of the NK compartment in natural resistance to HIV-1 merits further investigation.     

Human T Lymphocyte Clones: Influence of Culture Conditions and Optimization of Proliferative Assays

Phosphatidylinositol-specific phospholipase C (PI-PLC) treatment of the human natural killer (NK) target cells Molt-4, Jurkat, and U937 reduced their susceptibility to killing by human NK cells in a dose-dependent fashion. This indicates that a cell surface structure, anchored by a glycosyl-phosphatidylinositol (G-PI) moiety, is important in NK cytotoxicity. In contrast, another common NK target cell line, K562, remained susceptible to NK killing after enzyme treatment, suggesting that distinct target structures are expressed by this cell line. PI-PLC treatment of Molt-4 cells also reduced their sensitivity to human lymphokine activated killer (LAK) cells, suggesting that NK and LAK cells share common specificity in the killing of Molt-4. In contrast, PI-PLC had no effect on the killing of the LAK target cell line, Daudi, which is only weakly sensitive to unactivated NK cells. Killing of a variety of murine target cells by murine NK cells was not affected by PI-PLC treatment, but cross-species killing of Molt-4 by murine NK cells was inhibited by PI-PLC, suggesting a common mechanism in the killing of this human target cell line. The PI-PLC treatment of effector cells from either species did not reduce their NK activity. The reduction in sensitivity of the Molt-4, Jurkat, and U937 target cells probably results from a loss of a target specific G-PI linked membrane molecule, but other possible explanations for these results are also discussed.     

Cytotoxic effector cells of the immune system

Summary The organism contains several types of cytotoxic cells which are able to lyse host and foreign cells. Cytotoxic T-lymphocytes (CTL) appear to play the most important role among the killer cells but other lymphatic cells, such as natural killer (NK) cells and lymphokine-activated killer (LAK) cells as well as macrophages are also highly effective in the lysis of appropriate targets. The various cytotoxic effector cells differ distinctly concerning origin, phenotype, morphology and target cell specificity, but they bear the common feature that they destroy the target cells in a contact-dependent non-phagocytotic process.CTL are characterized by typical lysosomal granules and by the expression of a characteristic pattern of surface molecules. They recognize specific antigens which are presented in context with molecules of class I major histocompatibility complex (MHC). NK cells, on the other hand, kill the appropriate targets without prior immunisation and without requiring recognition of MHC molecules at the target cells. They also bear a typical pattern of surface markers which differ in several aspects from that of CTL. Human NK cells are further characterized by peculiar cytoplasmic granules with parallel tubular arrays which are not present in other cytotoxic cells. LAK cells constitute an additional, only recently described, killer cell population which arise from lymphatic cells in the presence of interleukin-2. They appear to represent a functional unique cytotoxic effector cell system with an exceptionally wide target cell spectrum including normal and malignant cells of different origin. LAK cells, however, show a profound heterogeneity concerning the expression of phenotype surface markers and it is not yet clear whether they are a unique cell line. By electron microscopy they display peculiar intranuclear inclusion bodies which may be associated with prolonged stimulation by interleukin-2. CTL, NK and LAK cells appear to possess similar mechanisms for cytolysis including secretion of pore-forming proteins, serine proteases and other proteins. Furthermore, they are able to trigger the cleavage of DNA in the target cell nucleus by a hitherto unknown pathway.Macrophages differ substantially from other cytotoxic effector cells concerning morphology, phenotype, kinetic of activation and target cell spectrum. They perform a variety of functions whereby contact-dependent target cell lysis represents only one of their properties. After target cell binding they release over 20 different molecules such as interleukin-1 and tumor necrosis-factor-alpha as mediators for cytolysis. Thus, macrophages appear like other cytotoxic effector cells to destroy their appropriate targets by different factors.Taken together the data obtained hitherto suggest that cellular cytotoxicity is mediated by various effector cells which may make a major contribution in the defence against infections and malignancies.Abbreviations ADCC antibody dependent cellular cytotoxicity - CD clone of differentiation - CTL cytotoxic T-lymphocyte - HTL helper T-lymphocyte - IL-2 interleukin-2 - LAK lymphokine-activated killer cell - LGL large granular lymphocytes - MHC major histocompatibility complex - NK natural killer cell - P1 perforin 1 - PTA parallel tubular arrays - TCR T-cell receptor complex     

Influence of autologous dendritic cells on the in-vitro expansion and functions of peripheral blood NK cells

Abstract

Context: Allogeneic reactive NK cells were previously shown to exert a graft-versus-leukemia (GVL) effect during allogeneic hematopoietic stem cell transplantation, as well as reduce the incidence of graft-versus-host disease (GVHD).

Objective: We used autologous immature DCs as feeder cells for the in-vitro expansion of NK cells and studied the function of the NK cell cultures.

Materials and methods: NK cells were cultured for 15 days in the presence of autologous, immature DCs. Fold expansion, killing activity and expression of IFN-γ, perforin and granzyme B were evaluated.

Results: The highest NK cell expansion efficiency was observed when the ratio of NK cells:DCs was 2:1 and when cells were cultured in a contact-dependent manner. The killing activity of NK cells was highest when the NK:DC ratio was 10:1. NK cell cultures exhibited a significant upregulation in the mRNA expression of IFN-γ, perforin and granzyme B when the ratio of NK cells to DCs was 10:1.

Discussion: We successfully amplified NK cells using autologous immature DCs derived from human peripheral monocytes after induction as feeder cells. The use of autologous immature DCs for ex-vivo expansion of NK cells can be clinically applied to overcome limitations, such as the small number of NK cells in peripheral blood, and the high cost of NK cell sorting. Transfusion of allogeneic reactive NK cells has been suggested as a potential adjunctive therapeutic strategy after transplantation.

Conclusion: Autologous immature DCs can be used as feeder cells for ex-vivo expansion of functional NK cells.     

Virus-Dependent Cellular Cytotoxicity in Vitro

When human peripheral blood lymphocytes were incubated with 51Cr-labelled tissue culture cells (T24 bladder carcinoma cells or Chang liver cells), their natural cytotoxicity (NK) usually stopped after 8 h of incubation. The 51Cr release induced by lymphocytes treated with small amounts of live or ultraviolet-inactivated mumps virus was strongly enhanced and lasted longer. When the lymphocytes were fractionated by Percoll gradient centrifugation, the highest NK activity was found in the low-density fraction enriched in large granular lymphocytes, whereas that of the T-cell-enriched high-density fractions was low. In contrast, the virus-dependent cellular cytotoxic (VDCC) activity was more evenly distributed between these fractions. However, there was a difference between the target cells in that the T24 cells were more susceptible to the cytotoxicity of lymphocytes in the high-density fractions than the Chang cells. Studies of Percoll fractions in the single-cell agarose assay showed that virus treatment increased the proportion of both target binding cells and killer cells in all fractions. Moreover, in the high-density fractions the increase in the number of killer cells was greater than that in binding cells, suggesting that the enhanced target cell killing induced by the virions reflected both increased binding and effector cell activation. Surface marker analysis of unfractionated lymphocytes indicated that the number of T3+ effector cells was greater than that of the HNK-1+ effector cells, regardless of whether the lymphocytes were treated with virus or not. However, for both NK and VDCC, the T3 to HNK-1 distribution ratio on the effector cells was 5-8:1 for T24 and 2:1 for Chang. Taken together, the results indicate that both NK and VDCC effector cells are phenotypically heterogeneous and that the target cells may play an active role in the recruitment of those effector cells that are most efficient in that system. The enhancement of lymphocyte cytotoxicity primarily reflects effector cell recruitment.     

Impaired release of natural killer cytotoxic factor in patients with primary Sjögren''s syndrome.

The impaired natural killer (NK) cell activity against K562 target cells of patients with primary Sjögren''s syndrome (primary SS) was re-examined in a 2-year follow-up study of 10 patients and 10 normal controls. The ability of blood mononuclear cells (BMNC) to form effector/target cell conjugates and to release NK cytotoxic factor (NKCF) were studied. NK cell activity of the patients was unchanged low (P less than 0.01) compared with the controls. The number of effector/target cell conjugates did not differ between patients and controls, whereas NKCF-release from interferon-stimulated BMNC was significantly (P less than 0.01) reduced in the patients with primary SS and positively correlated to the reduced NK cell activity (r = 0.85, P = 0.0002). The permanently low NK cell activity of patients with primary SS appears therefore, at least in part, to be due to an impaired release of NKCF and not to a defective ability of effector cells to recognize and/or adhere to target cells.     

Regulation of human natural killing by lysosomotropic and thiol-reactive agents.

Treatment of human peripheral blood lymphocytes (PBL) with ammonium or other lysosomotropic amines eliminates natural killing (NK) activity of effector cells without affecting their viability. Inhibition of NK by these reagents is probably due to blocking of events subsequent to effector cell target cell binding but before programming for cytolysis. Treatment of K562 target cells with the same reagents has no effect on target cell sensitivity to NK. Pretreatment of effector cells with the NK inhibitory lysosomotropic agents decreases lysosomal thiol cathepsin activity without affecting the lysosomal acid phosphatase activity. Thiol-reactive agents, tosyl-L-lysine chloromethyl ketone (TLCK) and iodoacetamide also inhibit NK and thiol cathepsin activity in PBL. These data suggest a role for thiol-containing enzymes in the early stages of NK cytolysis.     

IL‐18, but not IL‐15, contributes to the IL‐12‐dependent induction of NK‐cell effector functions by Leishmania infantum in vivo

Activation of NK cells is a hallmark of infections with intracellular pathogens. We previously showed that the protozoan parasite Leishmania infantum triggered a rapid NK‐cell response in mice that required TLR9‐positive myeloid DC and IL‐12, but no IFN‐α/β. Here, we investigated whether IL‐15 or IL‐18 mediate the activity of IL‐12 or function as independent activators of NK cells. In contrast to earlier studies that described IL‐15 as crucial for NK‐cell priming in response to TLR ligands, the expression of IFN‐γ, FasL, perforin and granzyme B by NK cells in L. infantum‐infected mice was completely preserved in the absence of IL‐15, whereas the proliferative capacity of NK cells was lower than in WT mice. IFN‐γ secretion, cytotoxicity and FasL expression of NK cells from infected IL‐18^?/? mice were significantly reduced compared with controls, but, unlike IL‐12, IL‐18 was not essential for NK‐cell effector functions. Part of the NK‐cell‐stimulatory effect of IL‐12 was dependent on IL‐18. We conclude that IL‐15 is not functioning as a universal NK‐cell priming signal and that IL‐18 contributes to the NK‐cell response in visceral leishmaniasis. The cytokine requirements for NK‐cell activation appear to differ contingent upon the infectious pathogen.     

Impaired neonatal natural killer-cell activity to herpes simplex virus: Decreased inhibition of viral replication and altered response to lymphokines

Human adult natural killer (NK) cells were recently demonstrated to inhibit herpes simplex virus (HSV) replicationin vitro. In this study we compared the ability of newborn and adult NK cells to inhibit HSV replication. Cord blood mononuclear cells (MNCs) from healthy, term newborns and MNCs from adults were analyzed for their percentage of Leu-11⁺ cells and comparedin vitro for their NK-cell activity against HSV-infected fibroblasts and the tumor cell line K562. Cord blood MNCs, compared with adult MNCs, had significantly lower percentages of Leu-11⁺ cells (5 vs 11%;P<0.01), less anti-K562 NK activity (6 vs 54 lytic units/10⁷ cells;P<0.001), and less anti-HSV NK activity (5 vs 52% HSV plaque inhibition;P<0.02). Comparing individual neonates and adults with equal percentages of Leu-11⁺ cells, neonatal MNCs still had less NK activity against either target. When Leu-11⁺ MNCs were isolated using the fluorescence-activated cell sorter, neonatal Leu-11⁺ MNCs still inhibited HSV replication less than adult Leu-11⁺ MNCs (P<0.01). MNCs from some neonates had significant anti-K562 NK activity but poor anti-HSV NK activity, suggesting either nonidentical NK-cell subpopulations or specific suppression. Whereas neonatal NK activity against K562 was always augmented by prior exposure to either interferon (IFN) or interleukin-2 (IL-2), the neonatal NK activity against HSV-infected cells was only augmented for half of the neonates tested. Endogenous production of alpha-IFN and gamma-IFN by MNCs exposed to HSV-infected fibroblasts was the same for cells from neonates or from HSV-seronegative adults. More gamma-IFN was produced by MNCs from HSV-seropositive adults than from neonates or HSV-seronegative adults. These results suggest that although newborns have phenotypically identifiable NK cells and the capacity for IFN production, the ability of the NK cells to inhibit HSV replication is impaired, and their level of response and augmentation by specific lymphokines is target specific.     

Effect of Human Seminal Plasma on the Lytic Activity of Natural Killer Cells and Presumptive Identification of Participant Macromolecules

ABSTRACT: Using the lytic activity of natural killer (NK) cells as an in vitro parameter, the immunoregulatory properties of human seminal plasma (SeP1) and participant macromolecules have been investigated. Significant (P < 0.05) suppression of NK cell activity by SeP1 and chromatographically separable fractions was demonstrated in association with high and low molecular weight (M_r) macromolecules. SePl suppression was retained after heating to 56°C for 30 min, and appeared to function at the level of the effector, rather than target cell. Physicochemical characterization of high and low M_r fractions provided presumptive identification of the participation of transglutaminase and prostaglandins as the principal molecules contributory to SeP1 immunosuppression.