自然杀伤细胞的基础研究和进展

人们对免疫系统的认识早已从单一的线性结构到充满着反馈性因素的网络结构,代表着固有免疫细胞的NK细胞是淋巴细胞中具有重要意义的第三类亚群.NK细胞具有特征性的功能包括杀伤病毒感染细胞或变异的肿瘤细胞,因而被称之为溶细胞性效应细胞,并且还有重要的分泌细胞因子和趋化因子功能,是固有免疫中重要的组成部分.     

Natural killer cell immune responses

Natural killer (NK) cells play a vital role in innate immune responses to infection; they express activation receptors that recognize virus-infected cells. Highly related to receptors recognizing tumor cells, the activation receptors trigger cytotoxicity and cytokine production. NK cells also express inhibitory receptors for major histocompatibility complex (MHC) class I molecules that block the action of the activation receptors. Although many ligands for NK cell receptors have MHC class I folds, recent studies also indicate ligands resembling the NK cell receptors themselves. A combination of immunologic, genetic, biophysical, and in vivo approaches is being employed to understand fully how these receptors contribute to NK cell activities in innate immunity to pathogens and tumors.     

Characterization of age-related changes in natural killer cells during primary influenza infection in mice

The current investigation examined the importance of natural killer (NK) cells during the innate immune response to primary influenza infection in young and aged mice. Young (6-8 weeks) and aged (22 months) C57BL/6 mice were infected intranasally with influenza A virus, and NK cell-mediated cytotoxicity was determined in lung and spleen during the first 4 days of infection. Aged mice demonstrated both a decrease in influenza-inducible NK activity and a reduction in the percentage and number of NK1.1+ cells in response to primary influenza infection, relative to young mice. In order to further establish a role for NK cells in controlling influenza infection, young mice were depleted of NK cells in vivo by injecting rabbit anit-NK1.1 antibody 2 days and 1 day prior to influenza infection. Young mice depleted of NK cells exhibited increased weight loss and lung virus titers during the course of infection, compared to young mice infected with influenza virus. These data indicate that NK cell function is impaired in response to primary influenza infection in aged mice. More importantly, these results underscore the essential role of NK cells in controlling virus titers in lung during the early course of influenza infection, regardless of age.     

Differences in activation and tissue homing markers of natural killer cell subsets during acute dengue infection

Dengue virus (DENV) infection is considered one of the most important mosquito‐borne diseases. It causes a spectrum of illness that could be due to qualitative and/or quantitative difference(s) of the natural killer (NK) cell responses during acute DENV infection. This view prompted us to perform a detailed phenotypic comparative characterization of NK cell subsets from DENV‐infected patients with dengue fever (DF), patients with dengue haemorrhagic fever (DHF) and healthy controls. The activation/differentiation molecules, CD69 and CD57 and a variety of tissue homing molecules were analysed on the CD56^hi CD16^? and CD56^lo CD16⁺ NK cells. Although there was no increase in the frequency of the total NK cells during DENV infection compared with the healthy individuals, there was a significant increase in the frequency of the CD56^hi CD16^? subset and the frequency of CD69 expression by both NK cell subsets during the febrile phase of infection. We also found an increase in the frequencies of cells expressing CD69 and CD57 in the CD56^lo CD16⁺ subset compared with those in the CD56^hi CD16^? subset. Moreover, although the CD56^lo CD16⁺ subset contained a high frequency of cells expressing skin‐homing markers, the CD56^hi CD16^? subset contained a high frequency of cells expressing bone marrow and lymph node trafficking markers. Interestingly, no differences of these NK cell subsets were noted in samples from patients with DF versus those with DHF. These findings suggest that activation and differentiation and the patterns of tissue homing molecules of the two major NK cell subsets are different and that these might play a critical role in the immune response against acute DENV infection.     

Dendritic cells and natural killer cells in the pathogenesis of HIV infection

Dendritic cells (DC) and natural killer (NK) cells, the main cellular components of the innate immune system, participate in the most ancient first line of defense against infections. Both types of cells patrol peripheral tissues, whereas their rapid recruitment and activation at mucosal surfaces [the major entry point for the human immunodeficiency virus (HIV)] is a hallmark of acute inflammatory response. The ability of HIV to survive and replicate in the human host relies upon several molecular mechanisms eluding the immune surveillance of both adaptive immunity and of DC and NK cells beginning with the acute phase of primary HIV infection. DC and NK cells, unlike CD4⁺ T cells, are impaired more functionally rather than being depleted by HIV infection. In this article we will review some of the aspects of DC/NK cells interaction with HIV infection both in vitro and in vivo, and we will also speculate on the potential consequence for HIV pathogenesis and for the capacity of the virus to escape the surveillance of the innate immune system.     

TGF‐β affects development and differentiation of human natural killer cell subsets

Human peripheral blood NK cells may be divided into two main subsets: CD56^brightCD16^? and CD56^dimCD16⁺. Since TGF‐β is known to influence the development of many leukocyte lineages, its effects on NK cell differentiation either from human CD34⁺Lin^? hematopoietic progenitor/stem cells in vitro or from peripheral blood NK cells were investigated. TGF‐β represses development of NK cells from CD34⁺ progenitors and inhibits differentiation of CD16⁺ NK cells. Moreover, TGF‐β also results in conversion of a minor fraction of CD56^brightCD16⁺ cells found in peripheral blood into CD56^brightCD16^? cells, highlighting a possible role of the former as a developmental intermediate and of TGF‐β in influencing the genesis of NK subsets found in blood.     

Pro- and antiinflammatory cytokine signaling: reciprocal antagonism regulates interferon-gamma production by human natural killer cells

Activated monocytes produce proinflammatory cytokines (monokines) such as interleukin (IL)-12, IL-15, and IL-18 for induction of interferon-gamma (IFN-gamma) by natural killer (NK) cells. NK cells provide the antiinflammatory cytokine transforming growth factor (TGF)-beta, an autocrine/negative regulator of IFN-gamma. The ability of one signaling pathway to prevail over the other is likely important in controlling IFN-gamma for the purposes of infection and autoimmunity, but the molecular mechanism(s) of how this counterregulation occurs is unknown. Here we show that in isolated human NK cells, proinflammatory monokines antagonize antiinflammatory TGF-beta signaling by downregulating the expression of the TGF-beta type II receptor, and its signaling intermediates SMAD2 and SMAD3. In contrast, TGF-beta utilizes SMAD2, SMAD3, and SMAD4 to suppress IFN-gamma and T-BET, a positive regulator of IFN-gamma. Indeed, activated NK cells from Smad3(-/-) mice produce more IFN-gamma in vivo than NK cells from wild-type mice. Collectively, our data suggest that pro- and antiinflammatory cytokine signaling reciprocally antagonize each other in an effort to prevail in the regulation of NK cell IFN-gamma production.     

IL-21 enhances dendritic cell ability to induce interferon-gamma production by natural killer T cells

Interleukin (IL)-21 shows pleiotropic effects on the proliferation, differentiation, and effector functions of leukocytes. However, the influence of IL-21 on dendritic cell (DC) activation of natural killer T (NKT) cells has not yet been elucidated. In the present study, we examined the effect of IL-21 on murine myeloid DC ability to induce NKT cell production of interferon-γ (IFN-γ) and IL-4. Pretreatment of DCs with IL-21 and α-galactosylceramide (α-GalCer), an NKT cell-specific ligand, resulted in the enhanced ability of the DCs to induce NKT cell production of IFN-γ but not IL-4 in vitro compared to DCs pretreated with α-GalCer alone. A similar effect of IL-21 was observed when DCs pretreated with IL-21 and α-GalCer in vitro were transferred into naïve mice. Direct administration of IL-21 to the mice also enhanced IFN-γ production after injection of α-GalCer. Thus, IL-21 can modify DC ability to selectively enhance NKT cell production of IFN-γ upon stimulation with α-GalCer.     

Mature resting Ly6Chigh natural killer cells can be reactivated by IL‐15

Mature NK cells are heterogeneous as to their expression levels of cell surface molecules. However, the functional differences and physiological roles of each NK‐cell subset are not fully understood. In this study, we report that based on the Ly6C expression levels, mature C57BL/6 murine NK cells can be subdivided into Ly6C^low and Ly6C^high subsets. Ly6C^high NK cells are in an inert state as evidenced by the production of lower levels of IFN‐γ and granzyme B, and they exhibit poorer proliferative potential than Ly6C^low NK cells. In addition, adoptive transfer experiments revealed that Ly6C^high NK cells are derived from Ly6C^low NK cells in the steady state. These results strongly suggest that Ly6C^high NK cells are resting cells in the steady state. However, in vitro, Ly6C^high NK cells become Ly6C^low NK cells with strong effector functions upon stimulation with IL‐15. Moreover, Ly6C^high NK cells also revert to Ly6C^low NK cells in vivo upon injection of the IL‐15 inducers polyI:C and CpG. Taken together, these results demonstrate the plasticity of mature NK cells and suggest that Ly6C^high NK cells are a reservoir of potential NK cells that allow effective and strong response to infections.     

β-galactosylceramide alters invariant natural killer T cell function and is effective treatment for lupus

NZB/W female mice spontaneously develop systemic lupus, an autoantibody mediated disease associated with immune complex glomerulonephritis. Natural killer (NK) T cells augment anti-dsDNA antibody secretion by NZB/W B cells in vitro, and blocking NKT cell activation in vivo with anti-CD1 mAb ameliorates lupus disease activity. In the current study, we show that β-galactosylceramide reduces the in vivo induction of serum IFN-γ and/or IL-4 by the potent NKT cell agonist α-galactosylceramide and reduces NKT cell helper activity for IgG secretion. Treatment of NZB/W mice with the β-galactosylceramide ameliorated lupus disease activity as judged by improvement in proteinuria, renal histopathology, IgG anti-dsDNA antibody formation, and survival. In conclusion, β-galactosylceramide, a glycolipid that reduces the cytokine secretion induced by a potent NKT cell agonist ameliorates lupus in NZB/W mice.     

Innate sensing and cell-autonomous resistance pathways in Legionella pneumophila infection

Legionella pneumophila is a facultative intracellular bacterium which can cause a severe pneumonia called Legionnaires' disease after inhalation of contaminated water droplets and replication in alveolar macrophages. The innate immune system is generally able to sense and −in most cases- control L. pneumophila infection. Comorbidities and genetic risk factors, however, can compromise the immune system and high infection doses might overwhelm its capacity, thereby enabling L. pneumophila to grow and disseminate inside the lung. The innate immune system mediates sensing of L. pneumophila by employing e.g. NOD-like receptors (NLRs), Toll-like receptors (TLRs), as well as the cGAS/STING pathway to stimulate death of infected macrophages as well as production of proinflammatory cytokines and interferons (IFNs). Control of pulmonary L. pneumophila infection is largely mediated by inflammasome-, TNFα- and IFN-dependent macrophage-intrinsic resistance mechanisms. This article summarizes the current knowledge of innate immune responses to L. pneumophila infection in general, and of macrophage-intrinsic defense mechanisms in particular.     

Host defense peptides and their antimicrobial-immunomodulatory duality

Host defence peptides (HDPs) are short cationic molecules produced by the immune systems of most multicellular organisms and play a central role as effector molecules of innate immunity. Host defence peptides have a wide range of biological activities from direct killing of invading pathogens to modulation of immunity and other biological responses of the host. HDPs have important functions in multiple, clinically relevant disease processes and their imbalanced expression is associated with pathology in different organ systems and cell types. Furthermore, HDPs are now evaluated as model molecules for the development of novel natural antibiotics and immunoregulatory compounds. This review provides an overview of HDPs focused on their antimicrobial-immunomodulatory duality.     

Inhibition of human natural killer cell activity by Pseudomonas aeruginosa alkaline protease and elastase.

The present study was designed to examine the effect of Pseudomonas aeruginosa alkaline protease (AP) and elastase (Ela) on human natural killer (NK) cell activity in vitro. AP and Ela were found to inhibit NK cell function. Addition of alpha interferon and interleukin-2 did not abolish this inhibition of NK cell activity. Adhesion of effector to target cells was studied in a single-cell agarose assay of monocyte-depleted NK-cell-enriched cell populations. AP and Ela were shown to inhibit effector/target cell conjugate formation. Furthermore, AP and Ela inhibited the binding of the monoclonal antibody Leu-11, which reacts with the Fc receptor of NK cells. The inhibition of NK cell binding to the target cell by P. aeruginosa proteases is most likely due to proteolytic cleavage of the surface receptors involved in the binding of the effector cell to the target cell.     

Association between natural killer cell activity and infection in immunologically normal elderly people

Congenital patients who lack natural killer (NK) cell activity experience repeated polymicrobial infections. NK cell activity varies significantly among normal people, but it is unknown whether this variation influences their ability to fight infections. This study examined this concern. NK cell activity and other variables, i.e. age, sex, performance status (PS), serum albumin value, lymphocyte and neutrophil counts, various lymphocyte subsets, etc. were determined for 108 immunologically normal elderly subjects who were in nursing homes due to an impaired PS. We analysed for correlations between these variables and the follow-up results of the subjects. Forty-eight subjects developed infection(s) during the first year of follow-up. A low NK cell activity was associated with the development of infection (P = 0.0105, multivariate logistic regression analysis). The relative risk for the development of infection increased in accordance with the decrease in the NK cell activity. Eleven subjects died of infection during the study period. A low NK cell activity was associated with short survival due to infection (P = 0.0056, multivariate Cox's proportional-hazards regression analysis). Our data indicate that low NK cell activity is associated with development of infections and death due to infection in immunologically normal elderly subjects with an impaired PS.     

Controlled infection with a therapeutic virus defines the activation kinetics of human natural killer cells in vivo

Human natural killer (NK) cells play an important role in anti‐viral immunity. However, studying their activation kinetics during infection is highly problematic. A clinical trial of a therapeutic virus provided an opportunity to study human NK cell activation in vivo in a controlled manner. Ten colorectal cancer patients with liver metastases received between one and five doses of oncolytic reovirus prior to surgical resection of their tumour. NK cell surface expression of the interferon‐inducible molecules CD69 and tetherin peaked 24–48 h post‐infection, coincident with a peak of interferon‐induced gene expression. The interferon response and NK cell activation were transient, declining by 96 h post‐infection. Furthermore, neither NK cell activation nor the interferon response were sustained in patients undergoing multiple rounds of virus treatment. These results show that reovirus modulates human NK cell activity in vivo and suggest that this may contribute to any therapeutic effect of this oncolytic virus. Detection of a single, transient peak of activation, despite multiple treatment rounds, has implications for the design of reovirus‐based therapy. Furthermore, our results suggest the existence of a post‐infection refractory period when the interferon response and NK cell activation are blunted. This refractory period has been observed previously in animal models and may underlie the enhanced susceptibility to secondary infections that is seen following viral infection.     

Differential effects of ifosfamide on dendritic cell-mediated stimulation of T cell interleukin-2 production, natural killer cell cytotoxicity and interferon-gamma production

Ifosfamide is a DNA-alkylating agent used frequently in chemotherapy of human malignancies. Ifosfamide and its major decomposition products deplete intracellular glutathione (GSH). Glutathione is the major intracellular thiol reductant that protects cells against oxidative injury. Ifosfamide depletion of intracellular GSH in human dendritic cells (DC), T cells and natural killer (NK) cells impairs their functional activity which can be restored by reconstituting GSH. Here we assessed the effect of ifosfamide on DC-mediated stimulation of NK cell proliferation via T cells and on direct DC stimulation of NK cell cytotoxicity and interferon (IFN)-gamma production. Indirect DC stimulation of NK cell proliferation via T cells and T cell-derived interleukin (IL)-2 were reduced by ifosfamide treatment of DC and reconstitution of GSH in DC restored both responses. When DC and NK cells were treated with ifosfamide, DC could overcome the negative effect of ifosfamide on NK cytotoxic function whereas NK cell IFN-gamma production was less efficiently restored. The ability of IL-2 activated NK cells to kill autologous immature DC or to induce DC maturation was reduced moderately by treatment of both cell types with ifosfamide. Overall, our results suggest that DC may stimulate anti-tumour effector cells in patients even if they had received treatment with chemotherapeutic agents such as ifosfamide.     

Effects of Chlamydia trachomatis infection on the expression of natural killer (NK) cell ligands and susceptibility to NK cell lysis

Natural killer (NK) cells are an important component of the immediate immune response to infections, including infection by intracellular bacteria. We have investigated recognition of Chlamydia trachomatis (CT) by NK cells and show that these cells are activated to produce interferon (IFN)-gamma when peripheral blood mononuclear cells (PBMC) are stimulated with CT organisms. Furthermore, infection of epithelial cell lines with CT renders them susceptible to lysis by human NK cells. Susceptibility was observed 18-24 h following infection and required protein synthesis by the infecting chlamydiae, but not by the host cell; heat or UV inactivated chlamydiae did not induce susceptibility to NK cell lysis. CT infection was also shown to decrease the expression of classical and non-classical major histocompatibility complex (MHC) molecules on infected cells, thus allowing recognition by NK cells when combined with an activating signal. A candidate activating signal is MICA/B, which was shown to be expressed constitutively on epithelial cells.     

CsSAP,a teleost serum amyloid P component,interacts with bacteria,promotes phagocytosis,and enhances host resistance against bacterial and viral infection

Serum amyloid P component (SAP) is a member of the pentraxins family that plays important roles in innate immunity in vertebrates. In fish, the biological function of SAP is essentially unknown. In this study, we examined the expression and function of a SAP homologue (CsSAP) from tongue sole Cynoglossus semilaevis. CsSAP shares 46%–58% overall sequence identities with known fish SAP and was upregulated in expression by bacterial and viral infection. Recombinant CsSAP (rCsSAP) exhibited differential binding capacities to a wide range of Gram-positive and Gram-negative bacteria and promoted uptake of the bound bacteria by host phagocytes. When introduced in vivo, rCsSAP enhanced host resistance not only to bacterial infection but also to viral infection. Consistently, antibody blockage of CsSAP significantly weakened the ability of tongue sole to clear invading bacteria. These results provide the first evidence that fish SAP contributes significantly to both antibacterial and antiviral immunities.