Science.gov

Sample records for nk cell responses

  1. The Human NK Cell Response to Yellow Fever Virus 17D Is Primarily Governed by NK Cell Differentiation Independently of NK Cell Education.

    PubMed

    Marquardt, Nicole; Ivarsson, Martin A; Blom, Kim; Gonzalez, Veronica D; Braun, Monika; Falconer, Karolin; Gustafsson, Rasmus; Fogdell-Hahn, Anna; Sandberg, Johan K; Michaëlsson, Jakob

    2015-10-01

    NK cells play an important role in the defense against viral infections. However, little is known about the regulation of NK cell responses during the first days of acute viral infections in humans. In this study, we used the live attenuated yellow fever virus (YFV) vaccine 17D as a human in vivo model to study the temporal dynamics and regulation of NK cell responses in an acute viral infection. YFV induced a robust NK cell response in vivo, with an early activation and peak in NK cell function at day 6, followed by a delayed peak in Ki67 expression, which was indicative of proliferation, at day 10. The in vivo NK cell response correlated positively with plasma type I/III IFN levels at day 6, as well as with the viral load. YFV induced an increased functional responsiveness to IL-12 and IL-18, as well as to K562 cells, indicating that the NK cells were primed in vivo. The NK cell responses were associated primarily with the stage of differentiation, because the magnitude of induced Ki67 and CD69 expression was distinctly higher in CD57(-) NK cells. In contrast, NK cells expressing self- and nonself-HLA class I-binding inhibitory killer cell Ig-like receptors contributed, to a similar degree, to the response. Taken together, our results indicate that NK cells are primed by type I/III IFN in vivo early after YFV infection and that their response is governed primarily by the differentiation stage, independently of killer cell Ig-like receptor/HLA class I-mediated inhibition or education. PMID:26283480

  2. Local Microenvironment Controls the Compartmentalization of NK Cell Responses during Systemic Inflammation in Mice.

    PubMed

    Rasid, Orhan; Ciulean, Ioana Sonya; Fitting, Catherine; Doyen, Noelle; Cavaillon, Jean-Marc

    2016-09-15

    Systemic inflammatory response syndrome is a whole-body reaction to a triggering insult that often results in life-threatening illness. Contributing to the development of this inflammatory cascade are numerous cellular partners, among which NK cells were shown to play a key role. Accumulating evidence points to organ-specific properties of systemic inflammation and NK cells. However, little is known about compartment-specific activation of NK cells during systemic inflammatory response syndrome or the relative contribution of NK cell-intrinsic properties and microenvironmental cues. In this study, we undertook a sequential characterization of NK responses in the spleen, lungs, bone marrow, peritoneum, and blood using a mouse model of endotoxemia. We report that, despite similar systemic dynamics of NK cell responses, expression of activation markers (CD69 and CD25) and effector molecules (IFN-γ, granzyme B, and IL-10) display organ-specific thresholds of maximum activation. Using adoptive transfers of spleen and lung NK cells, we found that these cells have the capacity to quickly adapt to a new environment and adjust their response levels to that of resident NK cells. This functional adaptation occurs without significant alterations in phenotype and independently of subpopulation-specific trafficking. Thus, using a dynamic in vivo-transfer system, to our knowledge our study is the first to report the compartmentalization of NK cells responses during systemic inflammation and to show that NK cell-intrinsic properties and microenvironmental cues are involved in this process, in a sequential manner. PMID:27521338

  3. Evaluation of Functional NK Cell Responses in Vaccinated and SIV-Infected Rhesus Macaques

    PubMed Central

    Vargas-Inchaustegui, Diego A.; Ying, Olivia; Demberg, Thorsten; Robert-Guroff, Marjorie

    2016-01-01

    NK cells are crucial components of the innate immune system due to their capacity to exert rapid cytotoxic and immunomodulatory function in the absence of prior sensitization. NK cells can become activated by exposure to target cells and/or by cytokines produced by antigen-presenting cells. In this study, we examined the effects of a simian immunodeficiency virus (SIV) vaccine regimen and subsequent SIV infection on the cytotoxic and immunomodulatory functions of circulatory NK cells. While vaccination did not significantly impact the capacity of NK cells to kill MHC-devoid 721.221 target cells, SIV-infection led to a significant decrease in target cell killing. NK cells from uninfected macaques were responsive to a low dose (5 ng/ml) of IL-15 pre-activation, leading to significant increases in their cytotoxic potential, however, NK cells from SIV-infected macaques required a higher dose (50 ng/ml) of IL-15 pre-activation in order to significantly increase their cytotoxic potential. By contrast, no differences were observed in the capacity of NK cells from vaccinated and SIV-infected macaques to respond to IL-12 and IL-18. Similarly, NK cells both before and after infection exhibited equivalent responses to Fc-mediated activation. Collectively, our results show that early SIV-infection impairs the natural cytotoxic capacity of circulatory NK cells without affecting Fc-mediated or cytokine-producing function.

  4. Two photon microscopy intravital study of DC-mediated anti-tumor response of NK cells

    NASA Astrophysics Data System (ADS)

    Caccia, Michele; Gorletta, Tatiana; Sironi, Laura; Zanoni, Ivan; Salvetti, Cristina; Collini, Maddalena; Granucci, Francesca; Chirico, Giuseppe

    2010-02-01

    Recent studies have demonstrated that dendritic cells (DCs) play a crucial role in the activation of Natural Killer cells (NKs) that are responsible for anti-tumor innate immune responses. The focus of this report is on the role of pathogen associated molecular pattern (PAMP) activated-DCs in inducing NK cell-mediated anti-tumor responses. Mice transplanted sub-cute (s.c.) with AK7 cells, a mesothelioma cell line sensitive to NK cell responses, are injected with fluorescent NK cells and DC activation is then induced by s.c. injection of Lipopolysaccharide (LPS). Using 4 dimensional tracking we follow the kinetic behavior of NK cells at the Draining Lymph-Node (DLN). As control, noninflammatory conditions are also evaluated. Our data suggest that NK cells are recruited to the DLN where they can interact with activated-DCs with a peculiar kinetic behavior: short lived interactions interleaved by rarer longer ones. We also found that the changes in the NK dynamic behavior in inflammatory conditions clearly affect relevant motility parameters such as the instantaneous and average velocity and the effective diffusion coefficient. This observation suggests that NK cells and activated-DCs might efficiently interact in the DLN, where cells could be activated. Therefore the interaction between activated-DCs and NK cells in DLN is not only a reality but it may be also crucial for the start of the immune response of the NKs.

  5. Antibody-mediated response of NKG2Cbright NK cells against human cytomegalovirus.

    PubMed

    Costa-Garcia, Marcel; Vera, Andrea; Moraru, Manuela; Vilches, Carlos; López-Botet, Miguel; Muntasell, Aura

    2015-03-15

    Human CMV (HCMV) infection promotes a variable and persistent expansion of functionally mature NKG2C(bright) NK cells. We analyzed NKG2C(bright) NK cell responses triggered by Abs from HCMV(+) sera against HCMV-infected MRC5 fibroblasts. Specific Abs promoted the degranulation (i.e., CD107a expression) and the production of cytokines (TNF-α and IFN-γ) by a significant fraction of NK cells, exceeding the low natural cytotoxicity against HCMV-infected targets. NK cell-mediated Ab-dependent cell-mediated cytotoxicity was limited by viral Ag availability and HLA class I expression on infected cells early postinfection and increased at late stages, overcoming viral immunoevasion strategies. Moreover, the presence of specific IgG triggered the activation of NK cells against Ab-opsonized cell-free HCMV virions. As compared with NKG2A(+) NK cells, a significant proportion of NKG2C(bright) NK cells was FcεR γ-chain defective and highly responsive to Ab-driven activation, being particularly efficient in the production of antiviral cytokines, mainly TNF-α. Remarkably, the expansion of NKG2C(bright) NK cells in HCMV(+) subjects was related to the overall magnitude of TNF-α and IFN-γ cytokine secretion upon Ab-dependent and -independent activation. We show the power and sensitivity of the anti-HCMV response resulting from the cooperation between specific Abs and the NKG2C(bright) NK-cell subset. Furthermore, we disclose the proinflammatory potential of NKG2C(bright) NK cells, a variable that could influence the individual responses to other pathogens and tumors. PMID:25667418

  6. Interleukin-21 enhances NK cell activation in response to antibody-coated targets.

    PubMed

    Roda, Julie M; Parihar, Robin; Lehman, Amy; Mani, Aruna; Tridandapani, Susheela; Carson, William E

    2006-07-01

    NK cells express an activating FcR (FcgammaRIIIa) that mediates Ab-dependent cellular cytotoxicity and the production of immune modulatory cytokines in response to Ab-coated targets. IL-21 has antitumor activity in murine models that depends in part on its ability to promote NK cell cytotoxicity and IFN-gamma secretion. We hypothesized that the NK cell response to FcR stimulation would be enhanced by the administration of IL-21. Human NK cells cultured with IL-21 and immobilized IgG or human breast cancer cells coated with a therapeutic mAb (trastuzumab) secreted large amounts of IFN-gamma. Increased secretion of TNF-alpha and the chemokines IL-8, MIP-1alpha, and RANTES was also observed under these conditions. NK cell IFN-gamma production was dependent on distinct signals mediated by the IL-21R and the FcR and was abrogated in STAT1-deficient NK cells. Supernatants derived from NK cells that had been stimulated with IL-21 and mAb-coated breast cancer cells were able to drive the migration of naive and activated T cells in an in vitro chemotaxis assay. IL-21 also enhanced NK cell lytic activity against Ab-coated tumor cells. Coadministration of IL-21 and Ab-coated tumor cells to immunocompetent mice led to synergistic production of IFN-gamma by NK cells. Furthermore, the administration of IL-21 augmented the effects of an anti-HER2/neu mAb in a murine tumor model, an effect that required IFN-gamma. These findings demonstrate that IL-21 significantly enhances the NK cell response to Ab-coated targets and suggest that IL-21 would be an effective adjuvant to administer in combination with therapeutic mAbs. PMID:16785506

  7. NK cells enhance dendritic cell response against parasite antigens via NKG2D pathway.

    PubMed

    Guan, Hongbing; Moretto, Magali; Bzik, David J; Gigley, Jason; Khan, Imtiaz A

    2007-07-01

    Recent studies have shown that NK-dendritic cell (DC) interaction plays an important role in the induction of immune response against tumors and certain viruses. Although the effect of this interaction is bidirectional, the mechanism or molecules involved in this cross-talk have not been identified. In this study, we report that coculture with NK cells causes several fold increase in IL-12 production by Toxoplasma gondii lysate Ag-pulsed DC. This interaction also leads to stronger priming of Ag-specific CD8+ T cell response by these cells. In vitro blockade of NKG2D, a molecule present on human and murine NK cells, neutralizes the NK cell-induced up-regulation of DC response. Moreover, treatment of infected animals with Ab to NKG2D receptor compromises the development of Ag-specific CD8+ T cell immunity and reduces their ability to clear parasites. These studies emphasize the critical role played by NKG2D in the NK-DC interaction, which apparently is important for the generation of robust CD8+ T cell immunity against intracellular pathogens. To the best of our knowledge, this is the first work that describes in vivo importance of NKG2D during natural infection. PMID:17579080

  8. Toll-like receptor 3 regulates NK cell responses to cytokines and controls experimental metastasis

    PubMed Central

    Guillerey, Camille; Chow, Melvyn T; Miles, Kim; Olver, Stuart; Sceneay, Jaclyn; Takeda, Kazuyoshi; Möller, Andreas; Smyth, Mark J

    2015-01-01

    The Toll-like receptor 3 (TLR3) agonist poly(I:C) is a promising adjuvant for cancer vaccines due to its induction of potent antitumor responses occurring primarily through the activation of dendritic cells (DCs) and natural killer (NK) cells. However, little is known about the role of TLR3 sensing of endogenous ligands in innate tumor immunosurveillance. Here, we investigated whether TLR3 could modulate immune responses and facilitate tumor control without administration of an agonist. We observed only limited impact of TLR3 deficiency on spontaneous carcinogenesis and primary growth of B16F10, E0771 or MC38 tumors when injected subcutaneously to mice. Nevertheless, TLR3 was observed to limit experimental B16F10 lung metastasis, an immunologic constraint dependent on both IFNγ secretion and NK cells. Interestingly, we observed that NK cells derived from Tlr3 null (Tlr3−/−) mice were hyporesponsive to cytokine stimulation. Indeed, compared with NK cells with intact TLR3, Tlr3−/− NK cells produced significantly reduced pro-inflammatory cytokines, including IFNγ, when incubated in the presence of different combinations of IL-12, IL-18 and IL-15. Bone-marrow chimera experiments established that competent NK cell responses required TLR3 sensing on radio-sensitive immune cells. Intriguingly, although CD8α DCs robustly express high levels of TLR3, we found that those cells were not necessary for efficient IFNγ production by NK cells. Moreover, the defective NK cell phenotype of Tlr3−/− mice appeared to be independent of the gut microbiota. Altogether, our data demonstrate a pivotal role of endogenous TLR3 stimulation for the acquisition of full NK cell functions and immune protection against experimental metastasis. PMID:26405596

  9. Multidirectional interactions are bridging human NK cells with plasmacytoid and monocyte-derived dendritic cells during innate immune responses.

    PubMed

    Della Chiesa, Mariella; Romagnani, Chiara; Thiel, Andreas; Moretta, Lorenzo; Moretta, Alessandro

    2006-12-01

    During innate immune responses, natural killer (NK) cells may interact with both plasmacytoid dendritic cells (pDCs) and monocyte-derived dendritic cells (MDDCs). We show that freshly isolated NK cells promote the release by pDCs of IFN-alpha, in a CpG-dependent manner, whereas they induce IL-6 production in a CpG-independent manner. In turn pDC-derived IFN-alpha up-regulates NK-mediated killing, whereas IL-6 could promote B-cell differentiation. We also show that exposure to exogenous IL-12 or coculture with maturing MDDCs up-regulates the NK-cell-dependent IFN-alpha production by pDCs. On the other hand, NK cells cocultured with pDCs acquire the ability to kill immature MDDCs, thus favoring their editing process. Finally, we show that activated NK cells are unable to lyse pDCs because these cells display an intrinsic resistance to lysis. The exposure of pDCs to IL-3 increased their susceptibility to NK-cell cytotoxicity resulting from a de novo expression of ligands for activating NK-cell receptors, such as the DNAM-1 ligand nectin-2. Thus, different cell-to-cell interactions and various cytokines appear to control a multidirectional network between NK cells, MDDCs, and pDCs that is likely to play an important role during the early phase of innate immune responses to viral infections and to tumors. PMID:16873676

  10. NK Cell-Mediated Regulation of Protective Memory Responses against Intracellular Ehrlichial Pathogens

    PubMed Central

    Habib, Samar; El Andaloussi, Abdeljabar; Hisham, Ahmed; Ismail, Nahed

    2016-01-01

    Ehrlichiae are gram-negative obligate intracellular bacteria that cause potentially fatal human monocytic ehrlichiosis. We previously showed that natural killer (NK) cells play a critical role in host defense against Ehrlichia during primary infection. However, the contribution of NK cells to the memory response against Ehrlichia remains elusive. Primary infection of C57BL/6 mice with Ehrlichia muris provides long-term protection against a second challenge with the highly virulent Ixodes ovatus Ehrlichia (IOE), which ordinarily causes fatal disease in naïve mice. Here, we show that the depletion of NK cells in E. muris-primed mice abrogates the protective memory response against IOE. Approximately, 80% of NK cell-depleted E. muris-primed mice succumbed to lethal IOE infection on days 8–10 after IOE infection, similar to naïve mice infected with the same dose of IOE. The lack of a recall response in NK cell-depleted mice correlated with an increased bacterial burden, extensive liver injury, decreased frequency of Ehrlichia-specific IFN-γ-producing memory CD4+ and CD8+ T-cells, and a low titer of Ehrlichia-specific antibodies. Intraperitoneal infection of mice with E. muris resulted in the production of IL-15, IL-12, and IFN-γ as well as an expansion of activated NKG2D+ NK cells. The adoptive transfer of purified E. muris-primed hepatic and splenic NK cells into Rag2-/-Il2rg-/- recipient mice provided protective immunity against challenge with E. muris. Together, these data suggest that E. muris-induced memory-like NK cells, which contribute to the protective, recall response against Ehrlichia. PMID:27092553

  11. Anti-ovarian tumor response of donor peripheral blood mononuclear cells is due to infiltrating cytotoxic NK cells

    PubMed Central

    Pandey, Veethika; Oyer, Jeremiah L.; Igarashi, Robert Y.; Gitto, Sarah B.; Copik, Alicja J.; Altomare, Deborah A.

    2016-01-01

    Treatment of ovarian cancer, a leading cause of gynecological malignancy, has good initial efficacy with surgery and platinum/taxane-based chemotherapy, but poor long-term survival in patients. Inferior long-term prognosis is attributed to intraperitoneal spreading, relapse and ineffective alternate therapies. Adoptive cell therapy is promising for tumor remission, although logistical concerns impede widespread implementation. In this study, healthy PBMCs were used to examine the immune response in a mouse model with human ovarian cancer, where natural killer (NK) cells were found to be the effector cells that elicited an anti-tumor response. Presence of tumor was found to stimulate NK cell expansion in mice treated intraperitoneally with PBMC+Interleukin-2 (IL-2), as compared to no expansion in non-tumor-bearing mice given the same treatment. PBMC+IL-2 treated mice exhibiting NK cell expansion had complete tumor remission. To validate NK cell mediated anti-tumor response, the intratumoral presence of NK cells and their cytotoxicity was confirmed by immunohistochemistry and granzyme activity of NK cells recovered from the tumor. Collectively, this study highlights the significance of NK cell-cytotoxic response to tumor, which may be attributed to interacting immune cell types in the PBMC population, as opposed to clinically used isolated NK cells showing lack of anti-tumor efficacy in ovarian cancer patients. PMID:26802025

  12. Viral MHC class I-like molecule allows evasion of NK cell effector responses in vivo.

    PubMed

    Pyzik, Michal; Dumaine, Anne; Dumaine, Anne A; Charbonneau, Benoît; Fodil-Cornu, Nassima; Jonjic, Stipan; Vidal, Silvia M

    2014-12-15

    The outcome of mouse CMV (MCMV) infection varies among different inbred mouse strains depending on NK cell effector functions governed through recognition receptor triggering. NK cells from different mouse strains possess diverse repertoires of activating or inhibitory Ly49 receptors, which share some of their polymorphic MHC class I (MHC-I) ligands. By examining the NK cell response to MCMV infection in novel BALB substrains congenic for different MHC (or H-2 in mice) haplotypes, we show that recognition of viral MHC-I-like protein m157 by inhibitory Ly49C receptor allows escape from NK cell control of viral replication. Dominant inhibition by Ly49C bound to self-H-2(b) encoded MHC-I molecules masks this effect, which only becomes apparent in distinct H-2 haplotypes, such as H-2(f). The recognition of m157-expressing cells by Ly49C resulted in both decreased NK cell killing in vitro and reduced rejection in vivo. Further, control of infection with m157-deletant (Δm157) MCMV was improved in mice carrying H-2 molecules unrecognized by Ly49C but allowing expansion of NK cell effectors expressing activating Ly49L receptors. Hence, our study is the first, to our knowledge, to demonstrate that MHC-I mimicry strategies used by MCMV to avoid NK cell control are biologically relevant during in vivo viral infection. Of value for human studies is that only a few genetic assortments conditional on the repertoires of viral MHC-I-like proteins/host NK receptors/MHC haplotypes should allow efficient protection against CMV infection. PMID:25392524

  13. The Exonuclease Domain of Lassa Virus Nucleoprotein Is Involved in Antigen-Presenting-Cell-Mediated NK Cell Responses

    PubMed Central

    Russier, Marion; Reynard, Stéphanie; Carnec, Xavier

    2014-01-01

    ABSTRACT Lassa virus is an Old World Arenavirus which causes Lassa hemorrhagic fever in humans, mostly in West Africa. Lassa fever is an important public health problem, and a safe and effective vaccine is urgently needed. The infection causes immunosuppression, probably due to the absence of activation of antigen-presenting cells (dendritic cells and macrophages), low type I interferon (IFN) production, and deficient NK cell function. However, a recombinant Lassa virus carrying D389A and G392A substitutions in the nucleoprotein that abolish the exonuclease activity and IFN activation loses its inhibitory activity and induces strong type I IFN production by dendritic cells and macrophages. We show here that during infection by this mutant Lassa virus, antigen-presenting cells trigger efficient human NK cell responses in vitro, including production of IFN-γ and cytotoxicity. NK cell activation involves close contact with both antigen-presenting cells and soluble factors. We report that infected dendritic cells and macrophages express the NKG2D ligands major histocompatibility complex (MHC) class I-related chains A and B and that they may produce interleukin-12 (IL-12), IL-15, and IL-18, all involved in NK cell functions. NK cell degranulation is significantly increased in cocultures, suggesting that NK cells seem to kill infected dendritic cells and macrophages. This work confirms the inhibitory function of Lassa virus nucleoprotein. Importantly, we demonstrate for the first time that Lassa virus nucleoprotein is involved in the inhibition of antigen-presenting cell-mediated NK cell responses. IMPORTANCE The pathogenesis and immune responses induced by Lassa virus are poorly known. Recently, an exonuclease domain contained in the viral nucleoprotein has been shown to be able to inhibit the type I IFN response by avoiding the recognition of viral RNA by cell sensors. Here, we studied the responses of NK cells to dendritic cells and macrophages infected with a

  14. The Effects of Age and Latent Cytomegalovirus Infection on NK-Cell Phenotype and Exercise Responsiveness in Man

    PubMed Central

    Bigley, Austin B.; Spielmann, Guillaume; Agha, Nadia; Simpson, Richard J.

    2015-01-01

    The redeployment of NK-cells in response to an acute bout of exercise is thought to be an integral component of the “fight-or-flight” response, preparing the body for potential injury or infection. We showed previously that CMV seropositivity impairs the redeployment of NK-cells with exercise in the young. In the current study, we examined the effect of aging on the redeployment of NK-cells with exercise in the context of CMV. We show here that CMV blunts the exercise-induced redeployment of NK-cells in both younger (23–39 yrs) and older (50–64 yrs) subjects with older CMVneg subjects showing the largest postexercise mobilization and 1 h postexercise egress of NK-cells. The blunted exercise response in CMVpos individuals was associated with a decreased relative redeployment of the CD158a+ and CD57+ NK-cell subsets in younger and older individuals. In addition, we show that aging is associated with a CMV-independent increase in the proportion of NK-cells expressing the terminal differentiation marker CD57, while CMV is associated with an age-dependent decrease in the proportion of NK-cells expressing the inhibitory receptors KLRG1 (in the younger group) and CD158a (in the older group). Collectively, these data suggest that CMV may decrease NK-cell mediated immunosurveillance after exercise in both younger and older individuals. PMID:26583066

  15. The Effects of Age and Latent Cytomegalovirus Infection on NK-Cell Phenotype and Exercise Responsiveness in Man.

    PubMed

    Bigley, Austin B; Spielmann, Guillaume; Agha, Nadia; Simpson, Richard J

    2015-01-01

    The redeployment of NK-cells in response to an acute bout of exercise is thought to be an integral component of the "fight-or-flight" response, preparing the body for potential injury or infection. We showed previously that CMV seropositivity impairs the redeployment of NK-cells with exercise in the young. In the current study, we examined the effect of aging on the redeployment of NK-cells with exercise in the context of CMV. We show here that CMV blunts the exercise-induced redeployment of NK-cells in both younger (23-39 yrs) and older (50-64 yrs) subjects with older CMV(neg) subjects showing the largest postexercise mobilization and 1 h postexercise egress of NK-cells. The blunted exercise response in CMV(pos) individuals was associated with a decreased relative redeployment of the CD158a+ and CD57+ NK-cell subsets in younger and older individuals. In addition, we show that aging is associated with a CMV-independent increase in the proportion of NK-cells expressing the terminal differentiation marker CD57, while CMV is associated with an age-dependent decrease in the proportion of NK-cells expressing the inhibitory receptors KLRG1 (in the younger group) and CD158a (in the older group). Collectively, these data suggest that CMV may decrease NK-cell mediated immunosurveillance after exercise in both younger and older individuals. PMID:26583066

  16. Development of the adaptive NK cell response to human cytomegalovirus in the context of aging.

    PubMed

    López-Botet, Miguel; Muntasell, Aura; Martínez-Rodríguez, José E; López-Montañés, María; Costa-García, Marcel; Pupuleku, Aldi

    2016-09-01

    Human cytomegalovirus (HCMV) establishes a highly prevalent life-long latent infection. Though generally subclinical, HCMV infection may have severe consequences during fetal development and in immunocompromised individuals. Based on epidemiological studies HCMV(+) serology has been associated with the development of atherosclerosis, immune senescence and an increase mortality rate in elderly people. Such long-term detrimental effects of the viral infection presumably result from an inefficient immune control of the pathogen, depending on the quality and evolution of the individual host-pathogen relationship. Together with antigen-specific T lymphocytes, NK cells play an important role in anti-viral immune defense. HCMV promotes in some individuals the differentiation and persistent steady state expansion of an NK cell subset bearing the CD94/NKG2C activating receptor. The relationship between this adaptive NK cell response to HCMV and aging is overviewed. PMID:27349430

  17. Chemokine Receptor Expression on Normal Blood CD56+ NK-Cells Elucidates Cell Partners That Comigrate during the Innate and Adaptive Immune Responses and Identifies a Transitional NK-Cell Population

    PubMed Central

    Lima, Margarida; Leander, Magdalena; Santos, Marlene; Santos, Ana Helena; Lau, Catarina; Queirós, Maria Luís; Gonçalves, Marta; Fonseca, Sónia; Moura, João; Teixeira, Maria dos Anjos; Orfao, Alberto

    2015-01-01

    Studies of chemokine receptors (CKR) in natural killer- (NK-) cells have already been published, but only a few gave detailed information on its differential expression on blood NK-cell subsets. We report on the expression of the inflammatory and homeostatic CKR on normal blood CD56+low CD16+ and CD56+high  CD16−/+low NK-cells. Conventional CD56+low and CD56+high NK-cells present in the normal PB do express CKR for inflammatory cytokines, although with different patterns CD56+low NK-cells are mainly CXCR1/CXCR2+ and CXCR3/CCR5−/+, whereas mostly CD56+high NK-cells are CXCR1/CXCR2− and CXCR3/CCR5+. Both NK-cell subsets have variable CXCR4 expression and are CCR4− and CCR6−. The CKR repertoire of the CD56+low NK-cells approaches to that of neutrophils, whereas the CKR repertoire of the CD56+high NK-cells mimics that of Th1+ T cells, suggesting that these cells are prepared to migrate into inflamed tissues at different phases of the immune response. In addition, we describe a subpopulation of NK-cells with intermediate levels of CD56 expression, which we named CD56+int NK-cells. These NK-cells are CXCR3/CCR5+, they have intermediate levels of expression of CD16, CD62L, CD94, and CD122, and they are CD57− and CD158a−. In view of their phenotypic features, we hypothesize that they correspond to a transitional stage, between the well-known CD56+high and CD56+low NK-cells populations. PMID:26543875

  18. CAM and NK Cells

    PubMed Central

    2004-01-01

    It is believed that tumor development, outgrowth and metastasis are under the surveillance of the immune system. Although both innate and acquired immune systems play roles, innate immunity is the spearhead against tumors. Recent studies have revealed the critical role of natural killer (NK) cells in immune surveillance and that NK cell activity is considerably influenced by various agents, such as environmental factors, stress, foods and drugs. Some of these NK cell stimulants have been used in complementary and alternative medicine (CAM) since ancient times. Therefore, the value of CAM should be re-evaluated from this point of view. In this review, we overview the intimate correlation between NK cell functions and CAM agents, and discuss possible underlying mechanisms mediating this. In particular, neuro-immune crosstalk and receptors for CAM agents are the most important and interesting candidates for such mechanisms. PMID:15257322

  19. Microwell-Based Live Cell Imaging of NK Cell Dynamics to Assess Heterogeneity in Motility and Cytotoxic Response.

    PubMed

    Vanherberghen, Bruno; Frisk, Thomas; Forslund, Elin; Olofsson, Per E; Guldevall, Karolin; Önfelt, Björn

    2016-01-01

    NK cell heterogeneity has primarily been studied either on the population level, measuring average responses, or on the single cell level by flow cytometry, providing static snapshots. These approaches have certain drawbacks, not enabling dynamic observations of single cells over extended periods of time. One of the primary limitations of single cell imaging has been throughput; it has been challenging to collect data for many cells due to their dynamic nature and migrating out of the field of view. Spatially confining cells combined with automated fluorescence microscopy enables the simultaneous monitoring of many NK cells in parallel for extended periods of time (>12 h). Such an approach allows us to dissect how the sum of individual NK cell responses translates to the global average response typically observed. PMID:27177659

  20. Coordinated expansion of both memory T cells and NK cells in response to CMV infection in humans.

    PubMed

    Bayard, Charles; Lepetitcorps, Hélène; Roux, Antoine; Larsen, Martin; Fastenackels, Solène; Salle, Virginie; Vieillard, Vincent; Marchant, Arnaud; Stern, Marc; Boddaert, Jacques; Bajolle, Fanny; Appay, Victor; Sauce, Delphine

    2016-05-01

    NK cells are key players in the fight against persistent viruses. Human cytomegalovirus (HCMV) infection is associated with the presence of a population of CD16(+) CD56(dim) NKG2C(+) NK cells in both acutely and latently infected individuals. Here, we studied the nature of these terminally differentiated NK cells in different human populations infected with HCMV: healthy donors stratified by age, thymectomized individuals, pregnant women suffering from primary CMV infection, and lung transplant patients. Both CD16(+) CD56(dim) NK- and CD8 T-cell phenotypes as well as functional capacities were determined and stratified according to age and/or CMV event. Similarly to T-cell responsiveness, we observe an accumulation over time of NKG2C(+) NK cells, which preferentially expressed CD57. This accumulation is particularly prominent in elderly and amplified further by CMV infection. Latent HCMV infection (without replication) is sufficient for NKG2C(+) CD57(+) NK cells to persist in healthy individuals but is not necessarily required in old age. Collectively, the present work supports the emerging concept that CMV shapes both innate and adaptive immunity in humans. PMID:26910859

  1. Candida albicans Induces Metabolic Reprogramming in Human NK Cells and Responds to Perforin with a Zinc Depletion Response

    PubMed Central

    Hellwig, Daniela; Voigt, Jessica; Bouzani, Maria; Löffler, Jürgen; Albrecht-Eckardt, Daniela; Weber, Michael; Brunke, Sascha; Martin, Ronny; Kurzai, Oliver; Hünniger, Kerstin

    2016-01-01

    As part of the innate immune system, natural killer (NK) cells are directly involved in the response to fungal infections. Perforin has been identified as the major effector molecule acting against many fungal pathogens. While several studies have shown that perforin mediated fungicidal effects can contribute to fungal clearance, neither the activation of NK cells by fungal pathogens nor the effects of perforin on fungal cells are well-understood. In a dual approach, we have studied the global gene expression pattern of primary and cytokine activated NK cells after co-incubation with Candida albicans and the transcriptomic adaptation of C. albicans to perforin exposure. NK cells responded to the fungal pathogen with an up-regulation of genes involved in immune signaling and release of cytokines. Furthermore, we observed a pronounced increase of genes involved in glycolysis and glycolysis inhibitor 2-deoxy-D-glucose impaired C. albicans induced NK cell activation. This strongly indicates that metabolic adaptation is a major part of the NK cell response to C. albicans infections. In the fungal pathogen, perforin induced a strong up-regulation of several fungal genes involved in the zinc depletion response, such as PRA1 and ZRT1. These data suggest that fungal zinc homeostasis is linked to the reaction to perforin secreted by NK cells. However, deletion mutants in PRA1 and ZRT1 did not show altered susceptibility to perforin. PMID:27242763

  2. Candida albicans Induces Metabolic Reprogramming in Human NK Cells and Responds to Perforin with a Zinc Depletion Response.

    PubMed

    Hellwig, Daniela; Voigt, Jessica; Bouzani, Maria; Löffler, Jürgen; Albrecht-Eckardt, Daniela; Weber, Michael; Brunke, Sascha; Martin, Ronny; Kurzai, Oliver; Hünniger, Kerstin

    2016-01-01

    As part of the innate immune system, natural killer (NK) cells are directly involved in the response to fungal infections. Perforin has been identified as the major effector molecule acting against many fungal pathogens. While several studies have shown that perforin mediated fungicidal effects can contribute to fungal clearance, neither the activation of NK cells by fungal pathogens nor the effects of perforin on fungal cells are well-understood. In a dual approach, we have studied the global gene expression pattern of primary and cytokine activated NK cells after co-incubation with Candida albicans and the transcriptomic adaptation of C. albicans to perforin exposure. NK cells responded to the fungal pathogen with an up-regulation of genes involved in immune signaling and release of cytokines. Furthermore, we observed a pronounced increase of genes involved in glycolysis and glycolysis inhibitor 2-deoxy-D-glucose impaired C. albicans induced NK cell activation. This strongly indicates that metabolic adaptation is a major part of the NK cell response to C. albicans infections. In the fungal pathogen, perforin induced a strong up-regulation of several fungal genes involved in the zinc depletion response, such as PRA1 and ZRT1. These data suggest that fungal zinc homeostasis is linked to the reaction to perforin secreted by NK cells. However, deletion mutants in PRA1 and ZRT1 did not show altered susceptibility to perforin. PMID:27242763

  3. A poxvirus protein that binds to and inactivates IL-18, and inhibits NK cell response.

    PubMed

    Born, T L; Morrison, L A; Esteban, D J; VandenBos, T; Thebeau, L G; Chen, N; Spriggs, M K; Sims, J E; Buller, R M

    2000-03-15

    IL-18 induces IFN-gamma and NK cell cytotoxicity, making it a logical target for viral antagonism of host defense. We demonstrate that the ectromelia poxvirus p13 protein, bearing homology to the mammalian IL-18 binding protein, binds IL-18, and inhibits its activity in vitro. Binding of IL-18 to the viral p13 protein was compared with binding to the cellular IL-18R. The dissociation constant of p13 for murine IL-18 is 5 nM, compared with 0.2 nM for the cellular receptor heterodimer. Mice infected with a p13 deletion mutant of ectromelia virus had elevated cytotoxicity for YAC-1 tumor cell targets compared with control animals. Additionally, the p13 deletion mutant virus exhibited decreased levels of infectivity. Our data suggest that inactivation of IL-18, and subsequent impairment of NK cell cytotoxicity, may be one mechanism by which ectromelia evades the host immune response. PMID:10706717

  4. NK Cell Responsiveness is Tuned Commensurate with the Number of Inhibitory Receptors for Self MHC Class I: the Rheostat Model

    PubMed Central

    Joncker, Nathalie T.; Fernandez, Nadine C.; Treiner, Emmanuel; Vivier, Eric; Raulet, David H.

    2010-01-01

    Inhibitory receptors that engage self-MHC class I molecules enable NK cells to detect disease-associated loss of MHC class I on surrounding cells. Previous studies showed that some NK cells lack all receptors for self-MHC class I, yet fail to exhibit autoimmunity because they are generally hyporesponsive to stimulation. We asked whether NK cells exist in only two states, responsive and hyporesponsive, corresponding to cells that express or fail to express inhibitory receptors for self-MHC class I. The alternative model is that NK cells vary continuously in their responsiveness, based on variations in the number of different inhibitory and stimulatory receptors they express, which is known to vary. Here we show in the murine system that NK cell responsiveness increases quantitatively with each added self MHC-specific inhibitory receptor. Genetic analysis demonstrated that interactions of each of the receptors with self-MHC class I were necessary to observe augmented responsiveness. These findings suggest that NK cell responsiveness is comparable to a rheostat: it is tuned to an optimal set point depending on the inhibitory and stimulatory interactions encountered in the normal environment, so as to ensure self-tolerance and yet optimize sensitivity to changes in normal cells. PMID:19342631

  5. Interferon Alpha Induces Sustained Changes in NK Cell Responsiveness to Hepatitis B Viral Load Suppression In Vivo

    PubMed Central

    Gill, Upkar S.; Peppa, Dimitra; Micco, Lorenzo; Singh, Harsimran D.; Carey, Ivana; Foster, Graham R.; Maini, Mala K.; Kennedy, Patrick T. F.

    2016-01-01

    NK cells are important antiviral effectors, highly enriched in the liver, with the potential to regulate immunopathogenesis in persistent viral infections. Here we examined whether changes in the NK pool are induced when patients with eAg-positive CHB are ‘primed’ with PegIFNα and importantly, whether these changes are sustained or further modulated long-term after switching to nucleos(t)ides (sequential NUC therapy), an approach currently tested in the clinic. Longitudinal sampling of a prospectively recruited cohort of patients with eAg+CHB showed that the cumulative expansion of CD56bright NK cells driven by 48-weeks of PegIFNα was maintained at higher than baseline levels throughout the subsequent 9 months of sequential NUCs. Unexpectedly, PegIFNα-expanded NK cells showed further augmentation in their expression of the activating NK cell receptors NKp30 and NKp46 during sequential NUCs. The expansion in proliferating, functional NK cells was more pronounced following sequential NUCs than in comparison cohorts of patients treated with de novo NUCs or PegIFNα only. Reduction in circulating HBsAg concentrations, a key goal in the path towards functional cure of CHB, was only achieved in those patients with enhancement of NK cell IFNγ and cytotoxicity but decrease in their expression of the death ligand TRAIL. In summary, we conclude that PegIFNα priming can expand a population of functional NK cells with an altered responsiveness to subsequent antiviral suppression by NUCs. Patients on sequential NUCs with a distinct NK cell profile show a decline in HBsAg, providing mechanistic insights for the further optimisation of treatment strategies to achieve sustained responses in CHB. PMID:27487232

  6. Transcriptional Control of NK Cells.

    PubMed

    Sun, Joseph C

    2016-01-01

    Natural killer (NK) cells are innate lymphocytes that survey the environment and protect the host from infected and cancerous cells. As their name implies, NK cells represent an early line of defense during pathogen invasion by directly killing infected cells and secreting inflammatory cytokines. Although the function of NK cells was first described more than four decades ago, the development of this cytotoxic lineage is not well understood. In recent years, we have begun to identify specific transcription factors that control each stage of development and maturation, from ontogeny of the NK cell progenitor to the effector functions of activated NK cells in peripheral organs. This chapter highlights the transcription factors that are unique to NK cells, or shared between NK cells and other hematopoietic cell lineages, but govern the biology of this cytolytic lymphocyte. PMID:26177585

  7. An NK Cell Perforin Response Elicited via IL-18 Controls Mucosal Inflammation Kinetics during Salmonella Gut Infection.

    PubMed

    Müller, Anna A; Dolowschiak, Tamas; Sellin, Mikael E; Felmy, Boas; Verbree, Carolin; Gadient, Sandra; Westermann, Alexander J; Vogel, Jörg; LeibundGut-Landmann, Salome; Hardt, Wolf-Dietrich

    2016-06-01

    Salmonella Typhimurium (S.Tm) is a common cause of self-limiting diarrhea. The mucosal inflammation is thought to arise from a standoff between the pathogen's virulence factors and the host's mucosal innate immune defenses, particularly the mucosal NAIP/NLRC4 inflammasome. However, it had remained unclear how this switches the gut from homeostasis to inflammation. This was studied using the streptomycin mouse model. S.Tm infections in knockout mice, cytokine inhibition and -injection experiments revealed that caspase-1 (not -11) dependent IL-18 is pivotal for inducing acute inflammation. IL-18 boosted NK cell chemoattractants and enhanced the NK cells' migratory capacity, thus promoting mucosal accumulation of mature, activated NK cells. NK cell depletion and Prf-/- ablation (but not granulocyte-depletion or T-cell deficiency) delayed tissue inflammation. Our data suggest an NK cell perforin response as one limiting factor in mounting gut mucosal inflammation. Thus, IL-18-elicited NK cell perforin responses seem to be critical for coordinating mucosal inflammation during early infection, when S.Tm strongly relies on virulence factors detectable by the inflammasome. This may have broad relevance for mucosal defense against microbial pathogens. PMID:27341123

  8. An NK Cell Perforin Response Elicited via IL-18 Controls Mucosal Inflammation Kinetics during Salmonella Gut Infection

    PubMed Central

    Müller, Anna A.; Dolowschiak, Tamas; Sellin, Mikael E.; Felmy, Boas; Verbree, Carolin; Gadient, Sandra; Westermann, Alexander J.; Vogel, Jörg; LeibundGut-Landmann, Salome; Hardt, Wolf-Dietrich

    2016-01-01

    Salmonella Typhimurium (S.Tm) is a common cause of self-limiting diarrhea. The mucosal inflammation is thought to arise from a standoff between the pathogen's virulence factors and the host's mucosal innate immune defenses, particularly the mucosal NAIP/NLRC4 inflammasome. However, it had remained unclear how this switches the gut from homeostasis to inflammation. This was studied using the streptomycin mouse model. S.Tm infections in knockout mice, cytokine inhibition and –injection experiments revealed that caspase-1 (not -11) dependent IL-18 is pivotal for inducing acute inflammation. IL-18 boosted NK cell chemoattractants and enhanced the NK cells' migratory capacity, thus promoting mucosal accumulation of mature, activated NK cells. NK cell depletion and Prf-/- ablation (but not granulocyte-depletion or T-cell deficiency) delayed tissue inflammation. Our data suggest an NK cell perforin response as one limiting factor in mounting gut mucosal inflammation. Thus, IL-18-elicited NK cell perforin responses seem to be critical for coordinating mucosal inflammation during early infection, when S.Tm strongly relies on virulence factors detectable by the inflammasome. This may have broad relevance for mucosal defense against microbial pathogens. PMID:27341123

  9. Higher Frequency of NK and CD4+ T-Cells in Mucosa and Potent Cytotoxic Response in HIV Controllers.

    PubMed

    Taborda, Natalia Andrea; González, Sandra Milena; Alvarez, Cristiam Mauricio; Correa, Luis Alfonso; Montoya, Carlos Julio; Rugeles, María Teresa

    2015-01-01

    HIV infection induces immune alterations, mainly in gut mucosa, where the main target cells reside. However, the evolution of the infection is variable among infected individuals, as evidenced by HIV controllers who exhibit low or undetectable viral load in the absence of treatment. The aim of this study was to evaluate the frequency, phenotype and activity of T and NK cells in peripheral blood and gut mucosa in a cohort of Colombian HIV controllers. Blood and gut biopsies were included. The frequency and the activation status of T and NK cells were performed by flow cytometry. In addition, Gag-stimulated CD8+ T-cells and cytokine-stimulated NK cells were tested for cytotoxic activity. Finally, microbial translocation was measured by plasma lipopolysaccharide quantification. Compared with HIV-progressors, HIV controllers exhibited higher frequency of CD4+ T and NK cells, and lower expression of activation molecules in blood and mucosal immune cells, as well as lower microbial translocation. An increased production of molecules associated with cytotoxic activity of CD8+ T-cells in blood and mucosa and a higher percentage of polyfunctional CD8+ T cells in blood were also observed in HIV controllers. In addition, an increased activity of NK cells was observed in blood. These findings suggest that HIV controllers have a potent immune response, mainly mediated by cytotoxic cells that control HIV replication, which contribute to reducing alterations at the gut mucosa. PMID:26291824

  10. Higher Frequency of NK and CD4+ T-Cells in Mucosa and Potent Cytotoxic Response in HIV Controllers

    PubMed Central

    Taborda, Natalia Andrea; González, Sandra Milena; Alvarez, Cristiam Mauricio; Correa, Luis Alfonso; Montoya, Carlos Julio; Rugeles, María Teresa

    2015-01-01

    HIV infection induces immune alterations, mainly in gut mucosa, where the main target cells reside. However, the evolution of the infection is variable among infected individuals, as evidenced by HIV controllers who exhibit low or undetectable viral load in the absence of treatment. The aim of this study was to evaluate the frequency, phenotype and activity of T and NK cells in peripheral blood and gut mucosa in a cohort of Colombian HIV controllers. Blood and gut biopsies were included. The frequency and the activation status of T and NK cells were performed by flow cytometry. In addition, Gag-stimulated CD8+ T-cells and cytokine-stimulated NK cells were tested for cytotoxic activity. Finally, microbial translocation was measured by plasma lipopolysaccharide quantification. Compared with HIV-progressors, HIV controllers exhibited higher frequency of CD4+ T and NK cells, and lower expression of activation molecules in blood and mucosal immune cells, as well as lower microbial translocation. An increased production of molecules associated with cytotoxic activity of CD8+ T-cells in blood and mucosa and a higher percentage of polyfunctional CD8+ T cells in blood were also observed in HIV controllers. In addition, an increased activity of NK cells was observed in blood. These findings suggest that HIV controllers have a potent immune response, mainly mediated by cytotoxic cells that control HIV replication, which contribute to reducing alterations at the gut mucosa. PMID:26291824

  11. CpG-containing oligodeoxynucleotides act through TLR9 to enhance the NK cell cytokine response to antibody-coated tumor cells.

    PubMed

    Roda, Julie M; Parihar, Robin; Carson, William E

    2005-08-01

    Bacterial DNA contains a high frequency of unmethylated CpG motifs that stimulate immune cells via TLR9. NK cells express a low-affinity activating receptor for the Fc portion of IgG (FcgammaRIIIa), but were not thought to express TLR9 protein. The direct response of NK cells to CpG oligodeoxynucleotides (ODN) in the presence of FcR stimulation was investigated. Human NK cells cultured in the presence of CpG ODN plus immobilized IgG or Ab-coated tumor cells secreted large amounts of IFN-gamma (>2000 pg/ml), whereas cells stimulated with Ab alone, CpG ODN alone, or Ab and control ODN produced negligible amounts. Enhanced secretion of IL-8, macrophage-derived chemokine, and MIP-1alpha was also observed after costimulation. NK cell cytokine production was not the result of interactions with APCs or their cytokine products. Flow cytometric analysis revealed that 36 +/- 3.5% of human NK cells expressed basal levels of TLR9. TLR9 expression in human NK cells was confirmed by immunoblot analysis. Only TLR9-expressing NK cells responded to CpG ODN and Ab, because cytokine production was not observed in NK cells from TLR9-deficient mice. Mice receiving CpG ODN and HER2/neu-positive tumor cells treated with an anti-HER2 Ab exhibited enhanced systemic levels of IFN-gamma compared with mice receiving either agent alone. TLR9-/- animals reconstituted with TLR9+/+ NK cells secreted IFN-gamma in response to CpG ODN and Ab-coated tumor cells. These findings indicate that CpG ODN can directly enhance the NK cell cytokine response to Ab-coated targets via activation of TLR9. PMID:16034101

  12. Induction of potent NK cell-dependent anti-myeloma cytotoxic T cells in response to combined mapatumumab and bortezomib

    PubMed Central

    Neeson, Paul J; Hsu, Andy K; Chen, Yin R; Halse, Heloise M; Loh, Joanna; Cordy, Reece; Fielding, Kate; Davis, Joanne; Noske, Josh; Davenport, Alex J; Lindqvist-Gigg, Camilla A; Humphreys, Robin; Tai, Tsin; Prince, H Miles; Trapani, Joseph A; Smyth, Mark J; Ritchie, David S

    2015-01-01

    There is increasing evidence that some cancer therapies can promote tumor immunogenicity to boost the endogenous antitumor immune response. In this study, we used the novel combination of agonistic anti-TRAIL-R1 antibody (mapatumumab, Mapa) with low dose bortezomib (LDB) for this purpose. The combination induced profound myeloma cell apoptosis, greatly enhanced the uptake of myeloma cell apoptotic bodies by dendritic cell (DC) and induced anti-myeloma cytotoxicity by both CD8+ T cells and NK cells. Cytotoxic lymphocyte expansion was detected within 24 h of commencing therapy and was maximized when myeloma-pulsed DC were co-treated with low dose bortezomib and mapatumumab (LDB+Mapa) in the presence of NK cells. This study shows that Mapa has two distinct but connected modes of action against multiple myeloma (MM). First, when combined with LDB, Mapa produced powerful myeloma cell apoptosis; secondly, it promoted DC priming and an NK cell-mediated expansion of anti-myeloma cytotoxic lymphocyte (CTL). Overall, this study indicates that Mapa can be used to drive potent anti-MM immune responses. PMID:26405606

  13. NK cell-based immunotherapy for malignant diseases

    PubMed Central

    Cheng, Min; Chen, Yongyan; Xiao, Weihua; Sun, Rui; Tian, Zhigang

    2013-01-01

    Natural killer (NK) cells play critical roles in host immunity against cancer. In response, cancers develop mechanisms to escape NK cell attack or induce defective NK cells. Current NK cell-based cancer immunotherapy aims to overcome NK cell paralysis using several approaches. One approach uses expanded allogeneic NK cells, which are not inhibited by self histocompatibility antigens like autologous NK cells, for adoptive cellular immunotherapy. Another adoptive transfer approach uses stable allogeneic NK cell lines, which is more practical for quality control and large-scale production. A third approach is genetic modification of fresh NK cells or NK cell lines to highly express cytokines, Fc receptors and/or chimeric tumor-antigen receptors. Therapeutic NK cells can be derived from various sources, including peripheral or cord blood cells, stem cells or even induced pluripotent stem cells (iPSCs), and a variety of stimulators can be used for large-scale production in laboratories or good manufacturing practice (GMP) facilities, including soluble growth factors, immobilized molecules or antibodies, and other cellular activators. A list of NK cell therapies to treat several types of cancer in clinical trials is reviewed here. Several different approaches to NK-based immunotherapy, such as tissue-specific NK cells, killer receptor-oriented NK cells and chemically treated NK cells, are discussed. A few new techniques or strategies to monitor NK cell therapy by non-invasive imaging, predetermine the efficiency of NK cell therapy by in vivo experiments and evaluate NK cell therapy approaches in clinical trials are also introduced. PMID:23604045

  14. NK cell receptor imbalance and NK cell dysfunction in HBV infection and hepatocellular carcinoma

    PubMed Central

    Sun, Cheng; Sun, Haoyu; Zhang, Cai; Tian, Zhigang

    2015-01-01

    Hepatocellular carcinoma (HCC) is currently the third leading cause of cancer mortality and a common poor-prognosis malignancy due to postoperative recurrence and metastasis. There is a significant correlation between chronic hepatitis B virus (HBV) infection and hepatocarcinogenesis. As the first line of host defense against viral infections and tumors, natural killer (NK) cells express a large number of immune recognition receptors (NK receptors (NKRs)) to recognize ligands on hepatocytes, liver sinusoidal endothelial cells, stellate cells and Kupffer cells, which maintain the balance between immune response and immune tolerance of NK cells. Unfortunately, the percentage and absolute number of liver NK cells decrease significantly during the development and progression of HCC. The abnormal expression of NK cell receptors and dysfunction of liver NK cells contribute to the progression of chronic HBV infection and HCC and are significantly associated with poor prognosis for liver cancer. In this review, we focus on the role of NK cell receptors in anti-tumor immune responses in HCC, particularly HBV-related HCC. We discuss specifically how tumor cells evade attack from NK cells and how emerging understanding of NKRs may aid the development of novel treatments for HCC. Novel mono- and combination therapeutic strategies that target the NK cell receptor–ligand system may potentially lead to successful and effective immunotherapy in HCC. PMID:25308752

  15. Dendritic cells and NK cells stimulate bystander T cell activation in response to TLR agonists through secretion of IFN-alpha beta and IFN-gamma.

    PubMed

    Kamath, Arun T; Sheasby, Christopher E; Tough, David F

    2005-01-15

    Recognition of conserved features of infectious agents by innate pathogen receptors plays an important role in initiating the adaptive immune response. We have investigated early changes occurring among T cells after injection of TLR agonists into mice. Widespread, transient phenotypic activation of both naive and memory T cells was observed rapidly after injection of molecules acting through TLR3, -4, -7, and -9, but not TLR2. T cell activation was shown to be mediated by a combination of IFN-alphabeta, secreted by dendritic cells (DCs), and IFN-gamma, secreted by NK cells; notably, IFN-gamma-secreting NK cells expressed CD11c and copurified with DCs. Production of IFN-gamma by NK cells could be stimulated by DCs from TLR agonist-injected mice, and although soluble factors secreted by LPS-stimulated DCs were sufficient to induce IFN-gamma, maximal IFN-gamma production required both direct contact of NK cells with DCs and DC-secreted cytokines. In vitro, IFN-alphabeta, IL-18, and IL-12 all contributed to DC stimulation of NK cell IFN-gamma, whereas IFN-alphabeta was shown to be important for induction of T cell bystander activation and NK cell IFN-gamma production in vivo. The results delineate a pathway involving innate immune mediators through which TLR agonists trigger bystander activation of T cells. PMID:15634897

  16. DNA damage response and evasion from immunosurveillance in CLL: new options for NK cell-based immunotherapies

    PubMed Central

    Shatnyeva, Olga M.; Hansen, Hinrich P.; Reiners, Katrin S.; Sauer, Maike; Vyas, Maulik; von Strandmann, Elke Pogge

    2015-01-01

    Chronic lymphocytic leukemia (CLL) is the most prominent B cell malignancy among adults in the Western world and characterized by a clonal expansion of B cells. The patients suffer from severe immune defects resulting in increased susceptibility to infections and failure to generate an antitumor immune response. Defects in both, DNA damage response (DDR) pathway and crosstalk with the tissue microenvironment have been reported to play a crucial role for the survival of CLL cells, therapy resistance and impaired immune response. To this end, major advances over the past years have highlighted several T cell immune evasion mechanisms in CLL. Here, we discuss the consequences of an impaired DDR pathway for detection and elimination of CLL cells by natural killer (NK) cells. NK cells are considered to be a major component of the immunosurveillance in leukemia but NK cell activity is impaired in CLL. Restoration of NK cell activity using immunoligands and immunoconstructs in combination with the conventional chemotherapy may provide a future perspective for CLL treatment. PMID:25699074

  17. Memory-Like Antigen-Specific Human NK Cells from TB Pleural Fluids Produced IL-22 in Response to IL-15 or Mycobacterium tuberculosis Antigens

    PubMed Central

    Fu, Xiaoying; Yu, Sifei; Yang, Binyan; Lao, Suihua; Li, Baiqing; Wu, Changyou

    2016-01-01

    Our previous result indicated that memory-like human natural killer (NK) cells from TB pleural fluid cells (PFCs) produced large amounts of IFN-γ in response to Bacille Calmette Guerin (BCG). Furthermore, recent studies have shown that human lymphoid tissues harbored a unique NK cell subset that specialized in production of interleukin (IL)-22, a proinflammatory cytokine that mediates host defense against pathogens. Yet little information was available with regard to the properties of IL-22 production by memory-like human NK cells. In the present study, we found that cytokines IL-15 induced and IL-12 enhanced the levels of IL-22 by NK cells from TB PFCs. In addition, IL-22 but not IL-17 was produced by NK cells from PFCs in response to BCG and M.tb-related Ags. More importantly, the subset of specific IL-22-producing NK cells were distinct from IFN-γ-producing NK cells in PFCs. CD45RO+ or CD45RO- NK cells were sorted, co-cultured with autologous monocytes and stimulated with BCG for the production of IL-22. The result demonstrated that CD45RO+ but not CD45RO- NK cells produced significantly higher level of IL-22. Anti-IL-12Rβ1 mAbs (2B10) partially inhibit the expression of IL-22 by NK cells under the culture with BCG. Consistently, BCG specific IL-22-producing NK cells from PFCs expressed CD45ROhighNKG2Dhighgranzyme Bhigh. In conclusion, our data demonstrated that memory-like antigen-specific CD45RO+ NK cells might participate in the recall immune response for M. tb infection via producing IL-22, which display a critical role to fight against M. tb. PMID:27031950

  18. Stepwise phosphorylation of p65 promotes NF-κB activation and NK cell responses during target cell recognition

    PubMed Central

    Kwon, Hyung-Joon; Choi, Go-Eun; Ryu, Sangryeol; Kwon, Soon Jae; Kim, Sun Chang; Booth, Claire; Nichols, Kim E.; Kim, Hun Sik

    2016-01-01

    NF-κB is a key transcription factor that dictates the outcome of diverse immune responses. How NF-κB is regulated by multiple activating receptors that are engaged during natural killer (NK)-target cell contact remains undefined. Here we show that sole engagement of NKG2D, 2B4 or DNAM-1 is insufficient for NF-κB activation. Rather, cooperation between these receptors is required at the level of Vav1 for synergistic NF-κB activation. Vav1-dependent synergistic signalling requires a separate PI3K-Akt signal, primarily mediated by NKG2D or DNAM-1, for optimal p65 phosphorylation and NF-κB activation. Vav1 controls downstream p65 phosphorylation and NF-κB activation. Synergistic signalling is defective in X-linked lymphoproliferative disease (XLP1) NK cells entailing 2B4 dysfunction and required for p65 phosphorylation by PI3K-Akt signal, suggesting stepwise signalling checkpoint for NF-κB activation. Thus, our study provides a framework explaining how signals from different activating receptors are coordinated to determine specificity and magnitude of NF-κB activation and NK cell responses. PMID:27221592

  19. Stepwise phosphorylation of p65 promotes NF-κB activation and NK cell responses during target cell recognition.

    PubMed

    Kwon, Hyung-Joon; Choi, Go-Eun; Ryu, Sangryeol; Kwon, Soon Jae; Kim, Sun Chang; Booth, Claire; Nichols, Kim E; Kim, Hun Sik

    2016-01-01

    NF-κB is a key transcription factor that dictates the outcome of diverse immune responses. How NF-κB is regulated by multiple activating receptors that are engaged during natural killer (NK)-target cell contact remains undefined. Here we show that sole engagement of NKG2D, 2B4 or DNAM-1 is insufficient for NF-κB activation. Rather, cooperation between these receptors is required at the level of Vav1 for synergistic NF-κB activation. Vav1-dependent synergistic signalling requires a separate PI3K-Akt signal, primarily mediated by NKG2D or DNAM-1, for optimal p65 phosphorylation and NF-κB activation. Vav1 controls downstream p65 phosphorylation and NF-κB activation. Synergistic signalling is defective in X-linked lymphoproliferative disease (XLP1) NK cells entailing 2B4 dysfunction and required for p65 phosphorylation by PI3K-Akt signal, suggesting stepwise signalling checkpoint for NF-κB activation. Thus, our study provides a framework explaining how signals from different activating receptors are coordinated to determine specificity and magnitude of NF-κB activation and NK cell responses. PMID:27221592

  20. Ionomycin Treatment Renders NK Cells Hyporesponsive.

    PubMed

    Romera-Cárdenas, Gema; Thomas, L Michael; Lopez-Cobo, Sheila; García-Cuesta, Eva M; Long, Eric O; Reyburn, Hugh T

    2016-01-01

    Natural killer cells are cytotoxic lymphocytes important in immune responses to cancer and multiple pathogens. However, chronic activation of NK cells can induce a hyporesponsive state. The molecular basis of the mechanisms underlying the generation and maintenance of this hyporesponsive condition are unknown, thus an easy and reproducible mechanism able to induce hyporesponsiveness on human NK cells would be very useful to gain understanding of this process. Human NK cells treated with ionomycin lose their ability to degranulate and secrete IFN-γ in response to a variety of stimuli, but IL-2 stimulation can compensate these defects. Apart from reductions in the expression of CD11a/CD18, no great changes were observed in the activating and inhibitory receptors expressed by these NK cells, however their transcriptional signature is different to that described for other hyporesponsive lymphocytes. PMID:27007115

  1. Ionomycin Treatment Renders NK Cells Hyporesponsive

    PubMed Central

    Romera-Cárdenas, Gema; Thomas, L. Michael; Lopez-Cobo, Sheila; García-Cuesta, Eva M.; Long, Eric O.; Reyburn, Hugh T.

    2016-01-01

    Natural killer cells are cytotoxic lymphocytes important in immune responses to cancer and multiple pathogens. However, chronic activation of NK cells can induce a hyporesponsive state. The molecular basis of the mechanisms underlying the generation and maintenance of this hyporesponsive condition are unknown, thus an easy and reproducible mechanism able to induce hyporesponsiveness on human NK cells would be very useful to gain understanding of this process. Human NK cells treated with ionomycin lose their ability to degranulate and secrete IFN-γ in response to a variety of stimuli, but IL-2 stimulation can compensate these defects. Apart from reductions in the expression of CD11a/CD18, no great changes were observed in the activating and inhibitory receptors expressed by these NK cells, however their transcriptional signature is different to that described for other hyporesponsive lymphocytes. PMID:27007115

  2. Infection with foot-and-mouth disease virus (FMDV) induces a natural killer (NK) cell response in cattle that is lacking following vaccination

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Natural killer (NK) cells play a role in innate antiviral immunity by directly lysing virus-infected cells and producing antiviral cytokines such as interferon gamma (IFNgamma). We developed a system for characterizing the bovine NK response to foot-and-mouth disease virus (FMDV), which causes a dis...

  3. Increased B Cell and Cytotoxic NK Cell Proportions and Increased T Cell Responsiveness in Blood of Natalizumab-Treated Multiple Sclerosis Patients

    PubMed Central

    Jenmalm, Maria C.; Dahle, Charlotte; Vrethem, Magnus; Ernerudh, Jan

    2013-01-01

    Background Changes in the blood lymphocyte composition probably both mediate and reflect the effects of natalizumab treatment in multiple sclerosis, with implications for treatment benefits and risks. Methods A broad panel of markers for lymphocyte populations, including states of activation and co-stimulation, as well as functional T cell responses to recall antigens and mitogens, were assessed by flow cytometry in 40 patients with relapsing multiple sclerosis before and after one-year natalizumab treatment. Results Absolute numbers of all major lymphocyte populations increased after treatment, most markedly for NK and B cells. The fraction of both memory and presumed regulatory B cell subsets increased, as did CD3-CD56dim cytotoxic NK cells, whereas CD3-CD56bright regulatory NK cells decreased. The increase in cell numbers was further associated with a restored T cell responsiveness to recall antigens and mitogens in functional assays. Conclusions Our data confirms that natalizumab treatment increases the number of lymphocytes in blood, likely mirroring the expression of VLA-4 being highest on NK and B cells. This finding supports reduction of lymphocyte extravasation as a main mode of action, although the differential effects on subpopulation composition suggests that cell-signalling may also be affected. The systemic increase in T cell responsiveness reflects the increase in numbers, and while augmenting anti-infectious responses systemically, localized responses may become correspondingly decreased. PMID:24312575

  4. NK and CD4+ T cell co-operative immune responses correlate with control of disease in a macaque SIV infection model1

    PubMed Central

    Vargas-Inchaustegui, Diego A.; Xiao, Peng; Tuero, Iskra; Patterson, L. Jean; Robert-Guroff, Marjorie

    2012-01-01

    Control of infectious disease may be accomplished by successful vaccination, or by complex immunologic and genetic factors favoring antigen-specific multicellular immune responses. Using a rhesus macaque model, we evaluated antigen-specific T cell-dependent NK cell immune responses in SIV-infected macaques, designated controlling or non-controlling based on long-term chronic viremia levels, to determine if NK cell effector functions contribute to control of SIV infection. We observed that Gag stimulation of macaque PBMCs induced subset-specific NK cell responses in SIV-controlling, but not non-controlling animals, and that circulatory NK cell responses were dependent on antigen-specific IL-2 production by CD4+ central memory T cells. NK cell activation was blocked by anti-IL-2 neutralizing antibody and by CD4+ T cell depletion which abrogated the Gag-specific responses. Among tissue-resident cells, splenic and circulatory NK cells displayed similar activation profiles, whereas liver and mucosal NK cells displayed a decreased activation profile, similar in SIV controlling and non-controlling macaques. Lack of T cell-dependent NK cell function was rescued in SIV non-controlling macaques through drug-mediated control of viremia. Our results indicate that control of disease progression in SIV controlling macaques is associated with co-operation between antigen-specific CD4+ T cells and NK cell effector function, highlight the importance of such cell-to-cell co-operativity in adaptive immunity and suggest this interaction should be further investigated in HIV vaccine development and other prophylactic vaccine approaches. PMID:22798665

  5. Regulation of experimental autoimmune encephalomyelitis by natural killer (NK) cells.

    PubMed

    Zhang, B; Yamamura, T; Kondo, T; Fujiwara, M; Tabira, T

    1997-11-17

    In this report, we establish a regulatory role of natural killer (NK) cells in experimental autoimmune encephalomyelitis (EAE), a prototype T helper cell type 1 (Th1)-mediated disease. Active sensitization of C57BL/6 (B6) mice with the myelin oligodendrocyte glycoprotein (MOG)35-55 peptide induces a mild form of monophasic EAE. When mice were deprived of NK cells by antibody treatment before immunization, they developed a more serious form of EAE associated with relapse. Aggravation of EAE by NK cell deletion was also seen in beta 2-microglobulin-/- (beta 2m-/-) mice, indicating that NK cells can play a regulatory role in a manner independent of CD8+ T cells or NK1.1+ T cells (NK-T cells). The disease enhancement was associated with augmentation of T cell proliferation and production of Th1 cytokines in response to MOG35-55. EAE passively induced by the MOG35-55-specific T cell line was also enhanced by NK cell deletion in B6, beta 2m-/-, and recombination activation gene 2 (RAG-2)-/- mice, indicating that the regulation by NK cells can be independent of T, B, or NK-T cells. We further showed that NK cells inhibit T cell proliferation triggered by antigen or cytokine stimulation. Taken together, we conclude that NK cells are an important regulator for EAE in both induction and effector phases. PMID:9362528

  6. NK Cell-extrinsic IL-18 Signaling Is Required for Efficient NK Cell Activation to Vaccinia Virus

    PubMed Central

    Brandstadter, Joshua D.; Huang, Xiaopei; Yang, Yiping

    2014-01-01

    Summary NK cells are important for the control of vaccinia virus (VV) in vivo. Recent studies have shown that multiple pathways are required for effective activation of NK cells. These include both TLR-dependent and -independent pathways, as well as the NKG2D activating receptor that recognizes host stress-induced NKG2D ligands. However, it remains largely unknown what controls the upregulation of NKG2D ligands in response to VV infection. In this study, we first showed that IL-18 is critical for NK cell activation and viral clearance. We then demonstrated that IL-18 signaling on both NK cells and DCs is required for efficient NK cell activation upon VV infection in vitro. We further showed in vivo that efficient NK cell activation to VV is dependent on DCs and IL-18 signaling in non-NK cells, suggesting an essential role for NK cell-extrinsic IL-18 signaling in NK cell activation. Mechanistically, IL-18 signaling in DCs promotes expression of Rae-1, an NKG2D ligand. Collectively, our data reveal a previously unrecognized role for NK cell-extrinsic IL-18 signaling in NK cell activation through upregulation of NKG2D ligands. These observations may provide insights into the design of effective NK cell-based therapies for viral infections and cancer. PMID:24846540

  7. NK Cell Activation in the Antitumor Response Induced by IFN-α Dendritic Cells Loaded with Apoptotic Cells from Follicular Lymphoma Patients.

    PubMed

    Lapenta, Caterina; Donati, Simona; Spadaro, Francesca; Castaldo, Paolo; Belardelli, Filippo; Cox, Maria C; Santini, Stefano M

    2016-08-01

    Follicular lymphoma (FL) is the most common form of indolent non-Hodgkin lymphoma. This malignancy is considered virtually incurable, with high response rates to therapy but frequent relapses. We investigated the ability of monocyte-derived dendritic cells generated in the presence of IFN-α and GM-CSF (IFN-DC) and loaded with apoptotic lymphoma cells to activate immune responses against FL cells, with the ultimate goal of designing novel patient-specific vaccination strategies for the treatment of FL. In this article, we show that apoptotic tumor cell-loaded IFN-DC from FL patients, which were cultured for 2 wk with autologous lymphocytes, led to Th1 response skewing, based on significantly higher levels of IFN-γ production and a remarkable increase in CD8(+) and NK cell frequency, consistent with the detection of enhanced cytotoxic effector function toward autologous FL cells. IFN-DC were found to promote efficient NK cell activation, increased expression of cytotoxicity receptors, and extensive IFN-γ production in the virtual absence of IL-10. Moreover, direct recognition and killing of primary autologous lymphoma cells by activated NK cells from FL patients was also demonstrated. A critical role was demonstrated for MHC class I-related chain A and B and membrane-bound IL-15 in IFN-DC-mediated NK cell activation and early IFN-γ production. The overall results indicate that IFN-DC loaded with autologous apoptotic FL cells represent a valuable tool for improving the potency of therapeutic cancer vaccines through the efficient induction of NK cell activation and promotion of CD8(+) T cell antitumor immunity. PMID:27357153

  8. PTEN Is a Negative Regulator of NK Cell Cytolytic Function

    PubMed Central

    Briercheck, Edward L.; Trotta, Rossana; Chen, Li; Hartlage, Alex S.; Cole, Jordan P.; Cole, Tyler D.; Mao, Charlene; Banerjee, Pinaki P.; Hsu, Hsiang-Ting; Mace, Emily M.; Ciarlariello, David; Mundy-Bosse, Bethany L.; Garcia-Cao, Isabel; Scoville, Steven D.; Yu, Lianbo; Pilarski, Robert; Carson, William E.; Leone, Gustavo; Pandolfi, Pier Paolo; Yu, Jianhua; Orange, Jordan S.; Caligiuri, Michael A.

    2015-01-01

    Human NK cells are characterized by their ability to initiate an immediate and direct cytolytic response to virally infected or malignantly transformed cells. Within human peripheral blood, the more mature CD56dim NK cell efficiently kills malignant targets at rest, whereas the less mature CD56bright NK cells cannot. In this study, we show that resting CD56bright NK cells express significantly more phosphatase and tensin homolog deleted on chromosome 10 (PTEN) protein when compared with CD56dim NK cells. Consistent with this, forced overexpression of PTEN in NK cells resulted in decreased cytolytic activity, and loss of PTEN in CD56bright NK cells resulted in elevated cytolytic activity. Comparable studies in mice showed PTEN overexpression did not alter NK cell development or NK cell–activating and inhibitory receptor expression yet, as in humans, did decrease expression of downstream NK activation targets MAPK and AKT during early cytolysis of tumor target cells. Confocal microscopy revealed that PTEN overexpression disrupts the NK cell’s ability to organize immunological synapse components including decreases in actin accumulation, polarization of the microtubule organizing center, and the convergence of cytolytic granules. In summary, our data suggest that PTEN normally works to limit the NK cell’s PI3K/AKT and MAPK pathway activation and the consequent mobilization of cytolytic mediators toward the target cell and suggest that PTEN is among the active regulatory components prior to human NK cells transitioning from the noncytolytic CD56bright NK cell to the cytolytic CD56dim NK cells. PMID:25595786

  9. Killer Ig-like receptor 2DL4 does not mediate NK cell IFN-γ responses to soluble HLA-G preparations.

    PubMed

    Le Page, Michael E L; Goodridge, Jodie P; John, Elisabeth; Christiansen, Frank T; Witt, Campbell S

    2014-01-15

    The MHC class Ib molecule HLA-G has previously been reported to be the ligand for the NK cell receptor killer Ig-like receptor (KIR)2DL4, but this remains controversial. In this study, we investigated IFN-γ production by freshly isolated NK cells in response to both soluble and solid-phase ligands, including anti-KIR2DL4 mAbs and rHLA-G. Although freshly isolated CD56(bright) NK cells produced IFN-γ in response to soluble HLA-G preparations, the response was found to be absolutely dependent on the presence of small numbers of contaminating CD56(-), CD14(-), CD11c(+) myeloid dendritic cells (mDCs). HLA-G tetramers bound only to the contaminating mDCs in the NK preparations, and Abs to KIR2DL4 and HLA-G did not block NK cell IFN-γ production. NK cells did not respond to plate-bound HLA-G. Freshly isolated NK cells also produced IFN-γ in response to unpurified soluble anti-KIR2DL4 mAb but not to low endotoxin affinity-purified Ab. The data suggest that previous reports of functional interactions between KIR2DL4 and HLA-G may have resulted from the use of purified NK cells that were contaminated with mDCs and HLA-G preparations that were contaminated with material capable of stimulating mDCs to produce cytokines that stimulate NK cells to produce IFN-γ. PMID:24337374

  10. NK cells and exercise: implications for cancer immunotherapy and survivorship.

    PubMed

    Bigley, Austin B; Simpson, Richard J

    2015-06-01

    Natural Killer (NK) cells are cytotoxic effectors of the innate immune system that are able to recognize and eradicate tumor cells without prior antigenic exposure. Tumor infiltration by NK-cells is associated with prolonged survival in cancer patients and high NK-cell cytotoxicity has been linked to decreased cancer risk. Allogeneic adoptive transfer of NK-cells from healthy donors to cancer patients has shown promise as a means of controlling or reversing the spread of multiple human malignancies including multiple myeloma and acute myeloid leukemia. However, multiple issues remain that undermine the efficacy of long-term cancer treatment using adoptive transfer of NK-cells including loss of activating receptors and cytotoxic potential in transferred NK-cells. Moreover, chronic exercise has been linked to improved NK-cell cytotoxicity, prognosis, and survival in cancer patients, and cytomegalovirus (CMV) reactivation is associated with enhanced NK-cell function after hematopoietic stem cell transplantation and decreased relapse risk in AML patients. In this work, we explore the potential of exercise- and CMV-driven alterations in NK-cell phenotype and function to increase the efficacy of NK-cells for cancer immunotherapy and prolong survival in cancer patients. We conclude that acute exercise and CMV are both capable of enhancing NK-cell cytotoxicity through distinct mechanisms; however, these effects are not additive as CMV infection is associated with an impaired acute exercise response. Thus, we suggest that either acute exercise or in vitro expansion of NKG2C+/NKG2A- NK-cells (as seen in those with CMV) could serve as a simple strategy for enhancing the anti-tumor cytotoxicity of NK-cells for immunotherapy, and that exercise training could be used to improve survivorship in cancer patients being treated with either HSCT or NK-cell infusions. PMID:26175401

  11. Models to study NK cell biology and possible clinical application

    PubMed Central

    Zamora, Anthony E.; Grossenbacher, Steven K.; Aguilar, Ethan G.; Murphy, William J.

    2016-01-01

    Natural killer (NK) cells are large granular lymphocytes of the innate immune system, responsible for direct targeting and killing of both virally infected and transformed cells. NK cells rapidly recognize and respond to abnormal cells in the absence of prior sensitization due to their wide array of germline-encoded inhibitory and activating receptors, which differs from the receptor diversity found in B and T lymphocytes resulting from the use of recombination-activation gene (RAG) enzymes. Although NK cells have traditionally been described as natural killers that provide a first line of defense prior to the induction of adaptive immunity, a more complex view of NK cells is beginning to emerge indicating they may also function in various immunoregulatory roles and have the capacity to shape adaptive immune responses. With the growing appreciation for the diverse functions of NK cells and recent technological advancements that allow for a more in-depth understanding of NK cell biology, we can now begin to explore new ways to manipulate NK cells to increase their clinical utility. In this overview unit, we introduce the reader to various aspects of NK cell biology by reviewing topics ranging from NK cell diversity and function, mouse models and the roles of NK cells in health and disease, to potential clinical applications. PMID:26237009

  12. Models to Study NK Cell Biology and Possible Clinical Application.

    PubMed

    Zamora, Anthony E; Grossenbacher, Steven K; Aguilar, Ethan G; Murphy, William J

    2015-01-01

    Natural killer (NK) cells are large granular lymphocytes of the innate immune system, responsible for direct targeting and killing of both virally infected and transformed cells. NK cells rapidly recognize and respond to abnormal cells in the absence of prior sensitization due to their wide array of germline-encoded inhibitory and activating receptors, which differs from the receptor diversity found in B and T lymphocytes that is due to the use of recombination-activation gene (RAG) enzymes. Although NK cells have traditionally been described as natural killers that provide a first line of defense prior to the induction of adaptive immunity, a more complex view of NK cells is beginning to emerge, indicating they may also function in various immunoregulatory roles and have the capacity to shape adaptive immune responses. With the growing appreciation for the diverse functions of NK cells, and recent technological advancements that allow for a more in-depth understanding of NK cell biology, we can now begin to explore new ways to manipulate NK cells to increase their clinical utility. In this overview unit, we introduce the reader to various aspects of NK cell biology by reviewing topics ranging from NK cell diversity and function, mouse models, and the roles of NK cells in health and disease, to potential clinical applications. © 2015 by John Wiley & Sons, Inc. PMID:26237009

  13. PDK1 orchestrates early NK cell development through induction of E4BP4 expression and maintenance of IL-15 responsiveness

    PubMed Central

    Yang, Meixiang; Li, Dan; Chang, Zai; Yang, Zhongzhou; Tian, Zhigang

    2015-01-01

    E4BP4, a circadian protein, is indispensable for NK cell development. It remains largely unknown which signal is required to induce E4BP4 expression and what effects it has during NK cell differentiation. Here, we reveal that PDK1, a kinase upstream of mTOR, connects IL-15 signaling to E4BP4. Early deletion of PDK1 caused a severe loss of NK cells and compromised antitumor activity in vivo. PDK1-deficient NK cells displayed much weaker IL-15–induced mTOR activation and E4BP4 induction, as well as remarkable reduction in CD122, a receptor subunit specifying NK cell responsiveness to IL-15. The phenotypes were partially reversible by ectopic expression of E4BP4 or bypassed activation of mTOR. We also determined that PDK1-mediated metabolic signaling was dispensable for NK cell terminal maturation and survival. Thus, we identify a role for PDK1 signaling as a key mediator in regulating E4BP4 expression during early NK cell development. Our findings underscore the importance of IL-15 self-responsiveness through a positive feedback loop that involves PDK1–mTOR–E4BP4–CD122 signaling. PMID:25624444

  14. PDK1 orchestrates early NK cell development through induction of E4BP4 expression and maintenance of IL-15 responsiveness.

    PubMed

    Yang, Meixiang; Li, Dan; Chang, Zai; Yang, Zhongzhou; Tian, Zhigang; Dong, Zhongjun

    2015-02-01

    E4BP4, a circadian protein, is indispensable for NK cell development. It remains largely unknown which signal is required to induce E4BP4 expression and what effects it has during NK cell differentiation. Here, we reveal that PDK1, a kinase upstream of mTOR, connects IL-15 signaling to E4BP4. Early deletion of PDK1 caused a severe loss of NK cells and compromised antitumor activity in vivo. PDK1-deficient NK cells displayed much weaker IL-15-induced mTOR activation and E4BP4 induction, as well as remarkable reduction in CD122, a receptor subunit specifying NK cell responsiveness to IL-15. The phenotypes were partially reversible by ectopic expression of E4BP4 or bypassed activation of mTOR. We also determined that PDK1-mediated metabolic signaling was dispensable for NK cell terminal maturation and survival. Thus, we identify a role for PDK1 signaling as a key mediator in regulating E4BP4 expression during early NK cell development. Our findings underscore the importance of IL-15 self-responsiveness through a positive feedback loop that involves PDK1-mTOR-E4BP4-CD122 signaling. PMID:25624444

  15. IL-15 complexes induce NK- and T-cell responses independent of type I IFN signaling during rhinovirus infection.

    PubMed

    Jayaraman, A; Jackson, D J; Message, S D; Pearson, R M; Aniscenko, J; Caramori, G; Mallia, P; Papi, A; Shamji, B; Edwards, M; Westwick, J; Hansel, T; Stanciu, L A; Johnston, S L; Bartlett, N W

    2014-09-01

    Rhinoviruses are among the most common viruses to infect man, causing a range of serious respiratory diseases including exacerbations of asthma and COPD. Type I IFN and IL-15 are thought to be required for antiviral immunity; however, their function during rhinovirus infection in vivo is undefined. In RV-infected human volunteers, IL-15 protein expression in fluid from the nasal mucosa and in bronchial biopsies was increased. In mice, RV induced type I IFN-dependent expressions of IL-15 and IL-15Rα, which in turn were required for NK- and CD8(+) T-cell responses. Treatment with IL-15-IL-15Rα complexes (IL-15c) boosted RV-induced expression of IL-15, IL-15Rα, IFN-γ, CXCL9, and CXCL10 followed by recruitment of activated, IFN-γ-expressing NK, CD8(+), and CD4(+) T cells. Treating infected IFNAR1(-/-) mice with IL-15c similarly increased IL-15, IL-15Rα, IFN-γ, and CXCL9 (but not CXCL10) expression also followed by NK-, CD8(+)-, and CD4(+)-T-cell recruitment and activation. We have demonstrated that type I IFN-induced IFN-γ and cellular immunity to RV was mediated by IL-15 and IL-15Rα. Importantly, we also show that IL-15 could be induced via a type I IFN-independent mechanism by IL-15 complex treatment, which in turn was sufficient to drive IFN-γ expression and lymphocyte responses. PMID:24472849

  16. IL-2 induces STAT4 activation in primary NK cells and NK cell lines, but not in T cells.

    PubMed

    Wang, K S; Ritz, J; Frank, D A

    1999-01-01

    IL-2 exerts potent but distinct functional effects on two critical cell populations of the immune system, T cells and NK cells. Whereas IL-2 leads to proliferation in both cell types, it enhances cytotoxicity primarily in NK cells. In both T cells and NK cells, IL-2 induces the activation of STAT1, STAT3, and STAT5. Given this similarity in intracellular signaling, the mechanism underlying the distinct response to IL-2 in T cells and NK cells is not clear. In this study, we show that in primary NK cells and NK cell lines, in addition to the activation of STAT1 and STAT5, IL-2 induces tyrosine phosphorylation of STAT4, a STAT previously reported to be activated only in response to IL-12 and IFN-alpha. This activation of STAT4 in response to IL-2 is not due to the autocrine production of IL-12 or IFN-alpha. STAT4 activated in response to IL-2 is able to bind to a STAT-binding DNA sequence, suggesting that in NK cells IL-2 is capable of activating target genes through phosphorylation of STAT4. IL-2 induces the activation of Jak2 uniquely in NK cells, which may underlie the ability of IL-2 to activate STAT4 only in these cells. Although the activation of STAT4 in response to IL-2 occurs in primary resting and activated NK cells, it does not occur in primary resting T cells or mitogen-activated T cells. The unique activation of the STAT4-signaling pathway in NK cells may underlie the distinct functional effect of IL-2 on this cell population. PMID:9886399

  17. Natural Killer (NK)/melanoma cell interaction induces NK-mediated release of chemotactic High Mobility Group Box-1 (HMGB1) capable of amplifying NK cell recruitment

    PubMed Central

    Parodi, Monica; Pedrazzi, Marco; Cantoni, Claudia; Averna, Monica; Patrone, Mauro; Cavaletto, Maria; Spertino, Stefano; Pende, Daniela; Balsamo, Mirna; Pietra, Gabriella; Sivori, Simona; Carlomagno, Simona; Mingari, Maria Cristina; Moretta, Lorenzo; Sparatore, Bianca; Vitale, Massimo

    2015-01-01

    In this study we characterize a new mechanism by which Natural Killer (NK) cells may amplify their recruitment to tumors. We show that NK cells, upon interaction with melanoma cells, can release a chemotactic form of High Mobility Group Box-1 (HMGB1) protein capable of attracting additional activated NK cells. We first demonstrate that the engagement of different activating NK cell receptors, including those mainly involved in tumor cell recognition can induce the active release of HMGB1. Then we show that during NK-mediated tumor cell killing two HMGB1 forms are released, each displaying a specific electrophoretic mobility possibly corresponding to a different redox status. By the comparison of normal and perforin-defective NK cells (which are unable to kill target cells) we demonstrate that, in NK/melanoma cell co-cultures, NK cells specifically release an HMGB1 form that acts as chemoattractant, while dying tumor cells passively release a non-chemotactic HMGB1. Finally, we show that Receptor for Advanced Glycation End products is expressed by NK cells and mediates HMGB1-induced NK cell chemotaxis. Proteomic analysis of NK cells exposed to recombinant HMGB1 revealed that this molecule, besides inducing immediate chemotaxis, also promotes changes in the expression of proteins involved in the regulation of the cytoskeletal network. Importantly, these modifications could be associated with an increased motility of NK cells. Thus, our findings allow the definition of a previously unidentified mechanism used by NK cells to amplify their response to tumors, and provide additional clues for the emerging role of HMGB1 in immunomodulation and tumor immunity. PMID:26587323

  18. Bridging innate NK cell functions with adaptive immunity.

    PubMed

    Marcenaro, Emanuela; Carlomagno, Simona; Pesce, Silvia; Moretta, Alessandro; Sivori, Simona

    2011-01-01

    Killer Ig-like receptors (KIRs) are major human NK receptors displaying either inhibitory or activating functions which recognize allotypic determinants of HLA-class I molecules. Surprisingly, NK cell treatment with CpG-ODN (TLR9 ligands) results in selective down-modulation of KIR3DL2, its co-internalization with CpG-ODN and its translocation to TLR9-rich early endosomes. This novel KIR-associated function may offer clues to better understand the possible role of certain KIRs and also emphasizes the involvement of NK cells in the course of microbial infections. NK cells are involved not only in innate immune responses against viruses and tumors but also participate in the complex network of cell-to cell interaction that leads to the development of adaptive immune responses. In this context the interaction of NK cells with DC appears to play a crucial role in the acquisition of CCR7, a chemokine receptor that enables NK cells to migrate towards lymph nodes in response to CCL19 and/or CCL21. Analysis of NK cell clones revealed that KIR-mismatched but not KIR-matched NK cells acquire CCR7. These data have important implications in haploidentical haematopoietic stem cell transplantation (HSCT), in which KIR-mismatched NK cells may acquire the ability to migrate to secondary lymphoid compartments (SLCs), where they can kill recipient's antigen presenting cells (APCs) and T cells thus preventing graft versus host (and host vs. graft) reactions. PMID:21842364

  19. Altered NK Cell Development and Enhanced NK Cell-Mediated Resistance to MCMV in NKG2D-Deficient Mice

    PubMed Central

    Zafirova, Biljana; Mandarić, Sanja; Antulov, Ronald; Krmpotić, Astrid; Jonsson, Helena; Yokoyama, Wayne M.; Jonjić, Stipan; Polić, Bojan

    2009-01-01

    Summary NKG2D is a potent activating receptor on NK cells which acts as a molecular sensor for stressed cells expressing NKG2D ligands such as infected or tumor transformed cells. Although NKG2D is expressed on NK cell precursors, its role in NK cell development is still not known. We have generated NKG2D-deficient mice by targeting the Klrk1 locus. Here we provide evidence for an important regulatory role of NKG2D in the development of NK cells. The absence of NKG2D causes faster division of NK cells, perturbation in size of some NK cell subpopulations and their augmented sensitivity to apoptosis. As expected, NKG2D−/− NK cells are less responsive to tumor targets expressing NKG2D ligands. NKG2D−/− mice, however, show an enhanced NK cell-mediated resistance to MCMV infection as a consequence of NK cell dysregulation. Altogether, these findings provide evidence for yet unknown regulatory function of NKG2D in NK cell physiology. PMID:19631564

  20. Exercise-Dependent Regulation of NK Cells in Cancer Protection.

    PubMed

    Idorn, Manja; Hojman, Pernille

    2016-07-01

    Natural killer (NK) cells are the most responsive immune cells to exercise, displaying an acute mobilization to the circulation during physical exertion. Recently, exercise-dependent mobilization of NK cells was found to play a central role in exercise-mediated protection against cancer. Here, we review the link between exercise and NK cell function, focusing on circulating exercise factors and additional effects, including vascularization, hypoxia, and body temperature in mediating the effects on NK cell functionality. Exercise-dependent mobilization and activation of NK cells provides a mechanistic explanation for the protective effect of exercise on cancer, and we propose that exercise represents a potential strategy as adjuvant therapy in cancer, by improving NK cell recruitment and infiltration in solid tumors. PMID:27262760

  1. Recognition and Regulation of T Cells by NK Cells.

    PubMed

    Pallmer, Katharina; Oxenius, Annette

    2016-01-01

    Regulation of T cell responses by innate lymphoid cells (ILCs) is increasingly documented and studied. Direct or indirect crosstalk between ILCs and T cells early during and after T cell activation can affect their differentiation, polarization, and survival. Natural killer (NK) cells that belong to the ILC1 group were initially described for their function in recognizing and eliminating "altered self" and as source of early inflammatory cytokines, most notably type II interferon. Using signals conveyed by various germ-line encoded activating and inhibitory receptors, NK cells are geared to sense sudden cellular changes that can be caused by infection events, malignant transformation, or cellular stress responses. T cells, when activated by TCR engagement (signal 1), costimulation (signal 2), and cytokines (signal 3), commit to a number of cellular alterations, including entry into rapid cell cycling, metabolic changes, and acquisition of effector functions. These abrupt changes may alert NK cells, and T cells might thereby expose themselves as NK cell targets. Here, we review how activated T cells can be recognized and regulated by NK cells and what consequences such regulation bears for T cell immunity in the context of vaccination, infection, or autoimmunity. Conversely, we will discuss mechanisms by which activated T cells protect themselves against NK cell attack and outline the significance of this safeguard mechanism. PMID:27446081

  2. Recognition and Regulation of T Cells by NK Cells

    PubMed Central

    Pallmer, Katharina; Oxenius, Annette

    2016-01-01

    Regulation of T cell responses by innate lymphoid cells (ILCs) is increasingly documented and studied. Direct or indirect crosstalk between ILCs and T cells early during and after T cell activation can affect their differentiation, polarization, and survival. Natural killer (NK) cells that belong to the ILC1 group were initially described for their function in recognizing and eliminating “altered self” and as source of early inflammatory cytokines, most notably type II interferon. Using signals conveyed by various germ-line encoded activating and inhibitory receptors, NK cells are geared to sense sudden cellular changes that can be caused by infection events, malignant transformation, or cellular stress responses. T cells, when activated by TCR engagement (signal 1), costimulation (signal 2), and cytokines (signal 3), commit to a number of cellular alterations, including entry into rapid cell cycling, metabolic changes, and acquisition of effector functions. These abrupt changes may alert NK cells, and T cells might thereby expose themselves as NK cell targets. Here, we review how activated T cells can be recognized and regulated by NK cells and what consequences such regulation bears for T cell immunity in the context of vaccination, infection, or autoimmunity. Conversely, we will discuss mechanisms by which activated T cells protect themselves against NK cell attack and outline the significance of this safeguard mechanism. PMID:27446081

  3. Natural killer cell (NK) subsets and NK-like T-cell populations in acute myeloid leukemias and myelodysplastic syndromes.

    PubMed

    Aggarwal, N; Swerdlow, S H; TenEyck, S P; Boyiadzis, M; Felgar, R E

    2016-07-01

    The impact of the immune microenvironment on the behavior and therapeutic strategies for hematopoietic and lymphoid neoplasms is being increasingly recognized. Many functional studies of natural killer (NK) cell cytotoxic responses in myelodysplasia (MDS) and acute myeloid leukemia (AML) exist, but with limited data on these lymphocyte proportions and related T-cell subsets. The proportions of these cells and their prognostic implications were therefore investigated in 89 AML, 51 MDS, and 20 control marrows by flow cytometry. The median proportion of NK cells (relative to the total lymphocytes) was lower in AML versus controls (P = 0.01). Among AML, a lower proportion of NK cells predicted better survival, whereas a higher NK cell proportion was associated with the poor prognostic AML category (P = 0.002). NK cell proportions were similar in MDS, MDS subgroups, and control marrows. The relative proportion of the mature NK cell subset (CD56(dim) CD16/57(bright) ) was lower in AML and MDS versus controls (P = 0.006, P = 0.0002, respectively). The proportion of mature NK cells was not a prognostic indicator although fewer were seen in poor prognosis AML. In contrast, a lower proportion of mature NK cells correlated with worse survival in MDS (P = 0.027). A higher proportion of NK-like T-cells (of total lymphoid cells) was found in MDS compared to controls (P = 0.01). A lower proportion of NK-like T-cells predicted better survival in AML but not in MDS. Thus, the proportions of NK, NK-cell subsets, and NK-like T-cells vary in myeloid neoplasms, may potentially impact immunomodulatory therapies, and may impact outcome. © 2016 International Clinical Cytometry Society. PMID:26648320

  4. Axitinib induces DNA damage response leading to senescence, mitotic catastrophe, and increased NK cell recognition in human renal carcinoma cells.

    PubMed

    Morelli, Maria Beatrice; Amantini, Consuelo; Santoni, Matteo; Soriani, Alessandra; Nabissi, Massimo; Cardinali, Claudio; Santoni, Angela; Santoni, Giorgio

    2015-11-01

    Tyrosine kinase inhibitors (TKIs) including axitinib have been introduced in the treatment of renal cell carcinoma (RCC) because of their anti-angiogenic properties. However, no evidence are presently available on a direct cytotoxic anti-tumor activity of axitinib in RCC.Herein we reported by western blot analysis that axitinib treatment induces a DNA damage response (DDR) initially characterized by γ-H2AX phosphorylation and Chk1 kinase activation and at later time points by p21 overexpression in A-498 and Caki-2 RCC cells although with a different potency. Analysis by immunocytochemistry for the presence of 8-oxo-7,8-dihydro-2'-deoxyguanosine in cellular DNA and flow cytometry using the redox-sensitive fluorescent dye DCFDA, demonstrated that DDR response is accompanied by the presence of oxidative DNA damage and reactive oxygen species (ROS) generation. This response leads to G2/M cell cycle arrest and induces a senescent-like phenotype accompanied by enlargement of cells and increased senescence-associated β-galactosidase activity, which are abrogated by N-acetyl cysteine (NAC) pre-treatment. In addition, axitinib-treated cells undergo to cell death through mitotic catastrophe characterized by micronucleation and abnormal microtubule assembly as assessed by fluorescence microscopy.On the other hand, axitinib, through the DDR induction, is also able to increase the surface NKG2D ligand expression. Accordingly, drug treatment promotes NK cell recognition and degranulation in A-498 RCC cells in a ROS-dependent manner.Collectively, our results indicate that both cytotoxic and immunomodulatory effects on RCC cells can contribute to axitinib anti-tumor activity. PMID:26474283

  5. NK Cells, Tumor Cell Transition, and Tumor Progression in Solid Malignancies: New Hints for NK-Based Immunotherapy?

    PubMed Central

    Huergo-Zapico, Leticia; Parodi, Monica; Pedrazzi, Marco; Mingari, Maria Cristina; Sparatore, Bianca; Gonzalez, Segundo; Olive, Daniel; Bottino, Cristina

    2016-01-01

    Several evidences suggest that NK cells can patrol the body and eliminate tumors in their initial phases but may hardly control established solid tumors. Multiple factors, including the transition of tumor cells towards a proinvasive/prometastatic phenotype, the immunosuppressive effect of the tumor microenvironment, and the tumor structure complexity, may account for limited NK cell efficacy. Several putative mechanisms of NK cell suppression have been defined in these last years; conversely, the cross talk between NK cells and tumor cells undergoing different transitional phases remains poorly explored. Nevertheless, recent in vitro studies and immunohistochemical analyses on tumor biopsies suggest that NK cells could not only kill tumor cells but also influence their evolution. Indeed, NK cells may induce tumor cells to change the expression of HLA-I, PD-L1, or NKG2D-L and modulate their susceptibility to the immune response. Moreover, NK cells may be preferentially located in the borders of tumor masses, where, indeed, tumor cells can undergo Epithelial-to-Mesenchymal Transition (EMT) acquiring prometastatic phenotype. Finally, the recently highlighted role of HMGB1 both in EMT and in amplifying the recruitment of NK cells provides further hints on a possible effect of NK cells on tumor progression and fosters new studies on this issue. PMID:27294158

  6. Cytomegalovirus Infection Drives Adaptive Epigenetic Diversification of NK Cells with Altered Signaling and Effector Function

    PubMed Central

    Schlums, Heinrich; Cichocki, Frank; Tesi, Bianca; Theorell, Jakob; Beziat, Vivien; Holmes, Tim D.; Han, Hongya; Chiang, Samuel C.C.; Foley, Bree; Mattsson, Kristin; Larsson, Stella; Schaffer, Marie; Malmberg, Karl-Johan; Ljunggren, Hans-Gustaf; Miller, Jeffrey S.; Bryceson, Yenan T.

    2015-01-01

    SUMMARY The mechanisms underlying human natural killer (NK) cell phenotypic and functional heterogeneity are unknown. Here, we describe the emergence of diverse subsets of human NK cells selectively lacking expression of signaling proteins after human cytomegalovirus (HCMV) infection. The absence of B and myeloid cell-related signaling protein expression in these NK cell subsets correlated with promoter DNA hyperme-thylation. Genome-wide DNA methylation patterns were strikingly similar between HCMV-associated adaptive NK cells and cytotoxic effector T cells but differed from those of canonical NK cells. Functional interrogation demonstrated altered cytokine responsiveness in adaptive NK cells that was linked to reduced expression of the transcription factor PLZF. Furthermore, subsets of adaptive NK cells demonstrated significantly reduced functional responses to activated autologous T cells. The present results uncover a spectrum of epigenetically unique adaptive NK cell subsets that diversify in response to viral infection and have distinct functional capabilities compared to canonical NK cell subsets. PMID:25786176

  7. Human breast cancer cells enhance self tolerance by promoting evasion from NK cell antitumor immunity.

    PubMed

    Mamessier, Emilie; Sylvain, Aude; Thibult, Marie-Laure; Houvenaeghel, Gilles; Jacquemier, Jocelyne; Castellano, Rémy; Gonçalves, Anthony; André, Pascale; Romagné, François; Thibault, Gilles; Viens, Patrice; Birnbaum, Daniel; Bertucci, François; Moretta, Alessandro; Olive, Daniel

    2011-09-01

    NK cells are a major component of the antitumor immune response and are involved in controlling tumor progression and metastases in animal models. Here, we show that dysfunction of these cells accompanies human breast tumor progression. We characterized human peripheral blood NK (p-NK) cells and malignant mammary tumor-infiltrating NK (Ti-NK) cells from patients with noninvasive and invasive breast cancers. NK cells isolated from the peripheral blood of healthy donors and normal breast tissue were used as controls. With disease progression, we found that expression of activating NK cell receptors (such as NKp30, NKG2D, DNAM-1, and CD16) decreased while expression of inhibitory receptors (such as NKG2A) increased and that this correlated with decreased NK cell function, most notably cytotoxicity. Importantly, Ti-NK cells had more pronounced impairment of their cytotoxic potential than p-NK cells. We also identified several stroma-derived factors, including TGF-β1, involved in tumor-induced reduction of normal NK cell function. Our data therefore show that breast tumor progression involves NK cell dysfunction and that breast tumors model their environment to evade NK cell antitumor immunity. This highlights the importance of developing future therapies able to restore NK cell cytotoxicity to limit/prevent tumor escape from antitumor immunity. PMID:21841316

  8. Functional Analysis of Human NK cells by Flow Cytometry

    PubMed Central

    Bryceson, Yenan T.; Fauriat, Cyril; Nunes, João M.; Wood, Stephanie M.; Björkström, Niklas K.; Long, Eric O.; Ljunggren, Hans-Gustaf

    2016-01-01

    Natural killer (NK) cells are a subset of lymphocytes that contribute to innate immunity through cytokine secretion and target cell lysis. NK cell function is regulated by a multiplicity of activating and inhibitory receptors. The advance in instrumentation for multi-color flow cytometry and the generation of specific mAbs for different epitopes related to phenotypic and functional parameters have facilitated our understanding of NK cell responses. Here, we provide protocols for flow cytometric evaluation of degranulation and cytokine production by human NK cells from peripheral blood at the single cell level. In addition to offering insight into the regulation of human NK cell responses, these techniques are applicable to the assessment of various clinical conditions, including the diagnosis of immunodeficiency syndromes. PMID:20033652

  9. Signalling through NK1.1 triggers NK cells to die but induces NK T cells to produce interleukin-4.

    PubMed Central

    Asea, A; Stein-Streilein, J

    1998-01-01

    In vivo inoculation of specific antibody is an accepted protocol for elimination of specific cell populations. Except for anti-CD3 and anti-CD4, it is not known if the depleted cells are eliminated by signalling through the target molecule or through a more non-specific mechanism. C57BL/6 mice were inoculated with anti-natural killer (NK1.1) monoclonal antibody (mAb). Thereafter spleen cells were harvested, stained for both surface and intracellular markers, and analysed by flow cytometry. As early as 2 hr post inoculation, NK cells were signalled to become apoptotic while signalling through the NK1.1 molecule activated NK1.1+ T-cell receptor (TCR)+ (NK T) cells to increase in number, and produce interleukin-4 (IL-4). Anti NK1.1 mAb was less efficient at signalling apoptosis in NK cells when NK T-cell deficient [beta 2-microglobulin beta 2m-deficient] mice were used compared with wild type mice. Efficient apoptotic signalling was restored when beta 2m-deficient mice were reconstituted with NK T cells. NK-specific antibody best signals the apoptotic process in susceptible NK cells when resistant NK T cells are present, activated, and secrete IL-4. Images Figure 4 PMID:9616382

  10. Development of Murine Hepatic NK Cells during Ontogeny: Comparison with Spleen NK Cells

    PubMed Central

    Wu, Xian; Chen, Yongyan; Wei, Haiming; Sun, Rui; Tian, Zhigang

    2012-01-01

    The phenotype of developing liver NK cells (CD3−NK1.1+) was investigated during mouse ontogeny comparing with spleen NK cells. The highest percentage of hepatic CD27−CD11b− NK cells occurred at the fetal stage. After birth, the percentage of CD27−CD11b−NK cells in both the liver and spleen gradually decreased to their lowest level at 6 weeks. More CD27+CD11b−NK cells were detected in the liver than that in spleen from week 1 to 6. Expression of NKG2A on liver NK cells was decreased but still much higher than that of spleen NK cells after 1 week. The NKG2D expression on liver NK cells increased to its highest level and was significantly higher than on spleen NK cells till 4 weeks. During mouse ontogeny, weaker expression of NKp46 and CD2 and stronger expression of CD69, CD11c, 2B4, and CD73 were observed on liver NK cells. Furthermore, neonatal liver NK cells express higher IFN-γ and perforin than adult .These results suggest that the maturation process of NK cells is unique in the livers, and liver microenvironments might play critical roles to keep NK cells in an immature status. PMID:22203859

  11. NK Cell Subset Redistribution during the Course of Viral Infections

    PubMed Central

    Lugli, Enrico; Marcenaro, Emanuela; Mavilio, Domenico

    2014-01-01

    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. PMID:25177322

  12. NK cell biology: An update and future directions

    PubMed Central

    Campbell, Kerry S.; Hasegawa, Jun

    2013-01-01

    Natural killer (NK) cells constitute a minor subset of normal lymphocytes that initiate innate immune responses toward tumor and virus-infected cells. They can mediate spontaneous cytotoxicity toward these abnormal cells and rapidly secrete numerous cytokines and chemokines to promote subsequent adaptive immune responses. Significant progress has been made in the past two decades to improve our understanding of NK cell biology. Here we review recent discoveries, including a better comprehension of the “education” of NK cells to achieve functional competence during their maturation and the discovery of “memory” responses by NK cells suggesting that they may also contribute to adaptive immunity. The improved understanding of NK cell biology has forged greater awareness that these cells play integral early roles in immune responses. In addition, several promising clinical therapies have been used to exploit NK cell functions in treating cancer patients. As our molecular understanding improves, these and future immunotherapies should continue to provide promising strategies to exploit the unique functions of NK cells to treat cancer, infections, and other pathological conditions. PMID:23906377

  13. Combination of NK Cells and Cetuximab to Enhance Anti-Tumor Responses in RAS Mutant Metastatic Colorectal Cancer

    PubMed Central

    Spanholtz, Jan; Tordoir, Marleen; Thijssen, Victor L.; Heideman, Daniëlle A. M.; Verheul, Henk M. W.; de Gruijl, Tanja D.; van der Vliet, Hans J.

    2016-01-01

    The ability of Natural Killer (NK) cells to kill tumor targets has been extensively studied in various hematological malignancies. However, NK cell therapy directed against solid tumors is still in early development. Epidermal Growth Factor Receptor (EGFR) targeted therapies using monoclonal antibodies (mAbs) such as cetuximab and panitumumab are widely used for the treatment of metastatic colorectal cancer (mCRC). Still, the clinical efficacy of this treatment is hampered by mutations in RAS gene, allowing tumors to escape from anti-EGFR mAb therapy. It is well established that NK cells kill tumor cells by natural cytotoxicity and can in addition be activated upon binding of IgG1 mAbs through Fc receptors (CD16/FcγRIIIa) on their surface, thereby mediating antibody dependent cellular cytotoxicity (ADCC). In the current study, activated Peripheral Blood NK cells (PBNK) were combined with anti-EGFR mAbs to study their effect on the killing of EGFR+/- cancer cell lines, including those with RAS mutations. In vitro cytotoxicity experiments using colon cancer primary tumors and cell lines COLO320, Caco-2, SW620, SW480 and HT-29, demonstrated that PBNK cells are cytotoxic for a range of tumor cells, regardless of EGFR, RAS or BRAF status and at low E:T ratios. Cetuximab enhanced the cytotoxic activity of NK cells on EGFR+ tumor cells (either RASwt, RASmut or BRAFmut) in a CD16 dependent manner, whereas it could not increase the killing of EGFR- COLO320. Our study provides a rationale to strengthen NK cell immunotherapy through a combination with cetuximab for RAS and BRAF mutant mCRC patients. PMID:27314237

  14. Hepatitis B virus antigens impair NK cell function.

    PubMed

    Yang, Yinli; Han, Qiuju; Zhang, Cai; Xiao, Min; Zhang, Jian

    2016-09-01

    An inadequate immune response of the host is thought to be a critical factor causing chronic hepatitis B virus (CHB) infection. Natural killer (NK) cells, as one of the key players in the eradication and control of viral infections, were functionally impaired in CHB patients, which might contribute to viral persistence. Here, we reported that HBV antigens HBsAg and HBeAg directly inhibited NK cell function. HBsAg and/or HBeAg blocked NK cell activation, cytokine production and cytotoxic granule release in human NK cell-line NK-92 cells, which might be related to the downregulation of activating receptors and upregulation of inhibitory receptor. Furthermore, the underlying mechanisms likely involved the suppression of STAT1, NF-κB and p38 MAPK pathways. These findings implicated that HBV antigen-mediated inhibition of NK cells might be an efficient strategy for HBV evasion, targeting the early antiviral responses mediated by NK cells and resulting in the establishment of chronic virus infection. Therefore, this study revealed the relationship between viral antigens and human immune function, especially a potential important interaction between HBV and innate immune responses. PMID:27341035

  15. Pegylated Interferon α-2a Triggers NK-Cell Functionality and Specific T-Cell Responses in Patients with Chronic HBV Infection without HBsAg Seroconversion

    PubMed Central

    Bruder Costa, Juliana; Dufeu-Duchesne, Tania; Leroy, Vincent; Bertucci, Inga; Bouvier-Alias, Magali; Pouget, Noelle; Brevot-Lutton, Ophelie; Bourliere, Marc; Zoulim, Fabien

    2016-01-01

    Pegylated interferon α-2a (Peg-IFN-α) represents a therapeutic alternative to the prolonged use of nucleos(t)ide analog (NA) in chronic hepatitis B (CHB) infection. The mechanisms leading to a positive clinical outcome remain unclear. As immune responses are critical for virus control, we investigated the effects of Peg-IFN-α on both innate and adaptive immunity, and related it to the clinical evolution. The phenotypic and functional features of the dendritic cells (DCs), natural killer (NK) cells and HBV-specific CD4/CD8 T cells were analyzed in HBeAg-negative CHB patients treated for 48-weeks with NA alone or together with Peg-IFN-α, before, during and up to 2-years after therapy. Peg-IFN-α induced an early activation of DCs, a potent expansion of the CD56bright NK subset, and enhanced the activation and functionality of the CD56dim NK subset. Peg-IFN-α triggered an increase in the frequencies of Th1- and Th17-oriented HBV-specific CD4/CD8 T cells. Peg-IFN-α reversed the unresponsiveness of patients to a specific stimulation. Most of the parameters returned to baseline after the stop of Peg-IFN-α therapy. Peg-IFN-α impacts both innate and adaptive immunity, overcoming dysfunctional immune responses in CHB patients. These modulations were not associated with seroconversion, which questioned the benefit of the add-on Peg-IFN-α treatment. PMID:27348813

  16. Pegylated Interferon α-2a Triggers NK-Cell Functionality and Specific T-Cell Responses in Patients with Chronic HBV Infection without HBsAg Seroconversion.

    PubMed

    Bruder Costa, Juliana; Dufeu-Duchesne, Tania; Leroy, Vincent; Bertucci, Inga; Bouvier-Alias, Magali; Pouget, Noelle; Brevot-Lutton, Ophelie; Bourliere, Marc; Zoulim, Fabien; Plumas, Joel; Aspord, Caroline

    2016-01-01

    Pegylated interferon α-2a (Peg-IFN-α) represents a therapeutic alternative to the prolonged use of nucleos(t)ide analog (NA) in chronic hepatitis B (CHB) infection. The mechanisms leading to a positive clinical outcome remain unclear. As immune responses are critical for virus control, we investigated the effects of Peg-IFN-α on both innate and adaptive immunity, and related it to the clinical evolution. The phenotypic and functional features of the dendritic cells (DCs), natural killer (NK) cells and HBV-specific CD4/CD8 T cells were analyzed in HBeAg-negative CHB patients treated for 48-weeks with NA alone or together with Peg-IFN-α, before, during and up to 2-years after therapy. Peg-IFN-α induced an early activation of DCs, a potent expansion of the CD56bright NK subset, and enhanced the activation and functionality of the CD56dim NK subset. Peg-IFN-α triggered an increase in the frequencies of Th1- and Th17-oriented HBV-specific CD4/CD8 T cells. Peg-IFN-α reversed the unresponsiveness of patients to a specific stimulation. Most of the parameters returned to baseline after the stop of Peg-IFN-α therapy. Peg-IFN-α impacts both innate and adaptive immunity, overcoming dysfunctional immune responses in CHB patients. These modulations were not associated with seroconversion, which questioned the benefit of the add-on Peg-IFN-α treatment. PMID:27348813

  17. Human NK Cell Diversity in Viral Infection: Ramifications of Ramification

    PubMed Central

    Strauss-Albee, Dara M.; Blish, Catherine A.

    2016-01-01

    Natural killer (NK) cells are a unique lymphocyte lineage with remarkable agility in the rapid destruction of virus-infected cells. They are also the most poorly understood class of lymphocyte. A spectrum of activating and inhibitory receptors at the NK cell surface leads to an unusual and difficult-to-study mechanism of cellular recognition, as well as a very high capacity for diversity at the single-cell level. Here, we review the evidence for the role of NK cells in the earliest stage of human viral infection, and in its prevention. We argue that single-cell diversity is a logical evolutionary adaptation for their position in the immune response and contributes to their ability to kill virus-infected cells. Finally, we look to the future, where emerging single-cell technologies will enable a new generation of rigorous and clinically relevant studies on NK cells accounting for all of their unique and diverse characteristics. PMID:26973646

  18. “Natural Regulators”: NK Cells as Modulators of T Cell Immunity

    PubMed Central

    Schuster, Iona S.; Coudert, Jerome D.; Andoniou, Christopher E.; Degli-Esposti, Mariapia A.

    2016-01-01

    Natural killer (NK) cells are known as frontline responders capable of rapidly mediating a response upon encountering transformed or infected cells. Recent findings indicate that NK cells, in addition to acting as innate effectors, can also regulate adaptive immune responses. Here, we review recent studies on the immunoregulatory function of NK cells with a specific focus on their ability to affect the generation of early, as well as long-term antiviral T cell responses, and their role in modulating immune pathology and disease. In addition, we summarize the current knowledge of the factors governing regulatory NK cell responses and discuss origin, tissue specificity, and open questions about the classification of regulatory NK cells as classical NK cells versus group 1 innate lymphoid cells. PMID:27379097

  19. Host protective ASP-based vaccine against the parasitic nematode Ostertagia ostertagi triggers NK cell activation and mixed IgG1-IgG2 response.

    PubMed

    González-Hernández, Ana; Van Coppernolle, Stefanie; Borloo, Jimmy; Van Meulder, Frederik; Paerewijck, Oonagh; Peelaers, Iris; Leclercq, Georges; Claerebout, Edwin; Geldhof, Peter

    2016-01-01

    The mucus-dwelling parasite Ostertagia ostertagi is one of the most important gastrointestinal nematodes in cattle. Our group has previously demonstrated the protective capacity of a vaccine against this parasite based on a native activation-associated secreted protein ASP1 (nASP) in combination with the saponin adjuvant QuilA. The aim of the current study was to analyse the effect of both antigen and adjuvant on the cellular and humoral vaccine-induced immune responses by comparing the native ASP to a recombinant version expressed in Pichia pastoris (pASP) and replacing QuilA by Al(OH)3. Immunization of cattle with the protective nASP+QuilA vaccine was associated with antigen-induced proliferation of natural killer (NK) cells combined with IFN-γ secretion and the induction of a mixed IgG1/IgG2 antibody response. ASP-specific activation and proliferation of NK cells was also observed in mice following the same vaccination regime. Replacing QuilA by Al(OH)3 or nASP by pASP significantly decreased the capacity of the vaccines to trigger both NK cell activation and antibody responses and failed to induce protection against a challenge infection. Reduction of the structurally anchoring disulphide bonds of the nASP completely abolished its ability to induce NK cell activation and antibody responses, highlighting the importance of protein conformation for the immunostimulatory activity. PMID:27403891

  20. Host protective ASP-based vaccine against the parasitic nematode Ostertagia ostertagi triggers NK cell activation and mixed IgG1-IgG2 response

    PubMed Central

    González-Hernández, Ana; Van Coppernolle, Stefanie; Borloo, Jimmy; Van Meulder, Frederik; Paerewijck, Oonagh; Peelaers, Iris; Leclercq, Georges; Claerebout, Edwin; Geldhof, Peter

    2016-01-01

    The mucus-dwelling parasite Ostertagia ostertagi is one of the most important gastrointestinal nematodes in cattle. Our group has previously demonstrated the protective capacity of a vaccine against this parasite based on a native activation-associated secreted protein ASP1 (nASP) in combination with the saponin adjuvant QuilA. The aim of the current study was to analyse the effect of both antigen and adjuvant on the cellular and humoral vaccine-induced immune responses by comparing the native ASP to a recombinant version expressed in Pichia pastoris (pASP) and replacing QuilA by Al(OH)3. Immunization of cattle with the protective nASP+QuilA vaccine was associated with antigen-induced proliferation of natural killer (NK) cells combined with IFN-γ secretion and the induction of a mixed IgG1/IgG2 antibody response. ASP-specific activation and proliferation of NK cells was also observed in mice following the same vaccination regime. Replacing QuilA by Al(OH)3 or nASP by pASP significantly decreased the capacity of the vaccines to trigger both NK cell activation and antibody responses and failed to induce protection against a challenge infection. Reduction of the structurally anchoring disulphide bonds of the nASP completely abolished its ability to induce NK cell activation and antibody responses, highlighting the importance of protein conformation for the immunostimulatory activity. PMID:27403891

  1. Bacillus Calmette-Guérin (BCG) Revaccination of Adults with Latent Mycobacterium tuberculosis Infection Induces Long-Lived BCG-Reactive NK Cell Responses.

    PubMed

    Suliman, Sara; Geldenhuys, Hennie; Johnson, John L; Hughes, Jane E; Smit, Erica; Murphy, Melissa; Toefy, Asma; Lerumo, Lesedi; Hopley, Christiaan; Pienaar, Bernadette; Chheng, Phalkun; Nemes, Elisa; Hoft, Daniel F; Hanekom, Willem A; Boom, W Henry; Hatherill, Mark; Scriba, Thomas J

    2016-08-15

    One third of the global population is estimated to be latently infected with Mycobacterium tuberculosis We performed a phase I randomized controlled trial of isoniazid preventive therapy (IPT) before revaccination with bacillus Calmette-Guérin (BCG) in healthy, tuberculin skin test-positive (≥15-mm induration), HIV-negative South African adults. We hypothesized that preclearance of latent bacilli with IPT modulates BCG immunogenicity following revaccination. Frequencies and coexpression of IFN-γ, TNF-α, IL-2, IL-17, and/or IL-22 in CD4 T cells and IFN-γ-expressing CD8 T, γδ T, CD3(+)CD56(+) NKT-like, and NK cells in response to BCG were measured using whole blood intracellular cytokine staining and flow cytometry. We analyzed 72 participants who were revaccinated with BCG after IPT (n = 33) or without prior IPT (n = 39). IPT had little effect on frequencies or cytokine coexpression patterns of M. tuberculosis- or BCG-specific responses. Revaccination transiently boosted BCG-specific Th1 cytokine-expressing CD4, CD8, and γδ T cells. Despite high frequencies of IFN-γ-expressing BCG-reactive CD3(+)CD56(+) NKT-like cells and CD3(-)CD56(dim) and CD3(-)CD56(hi) NK cells at baseline, BCG revaccination boosted these responses, which remained elevated up to 1 y after revaccination. Such BCG-reactive memory NK cells were induced by BCG vaccination in infants, whereas in vitro IFN-γ expression by NK cells upon BCG stimulation was dependent on IL-12 and IL-18. Our data suggest that isoniazid preclearance of M. tuberculosis bacilli has little effect on the magnitude, persistence, or functional attributes of lymphocyte responses boosted by BCG revaccination. Our study highlights the surprising durability of BCG-boosted memory NKT-like and NK cells expressing antimycobacterial effector molecules, which may be novel targets for tuberculosis vaccines. PMID:27412415

  2. Cancer-induced immunosuppression: IL-18-elicited immunoablative NK cells.

    PubMed

    Terme, Magali; Ullrich, Evelyn; Aymeric, Laetitia; Meinhardt, Kathrin; Coudert, Jérôme D; Desbois, Mélanie; Ghiringhelli, François; Viaud, Sophie; Ryffel, Bernard; Yagita, Hideo; Chen, Lieping; Mécheri, Salaheddine; Kaplanski, Gilles; Prévost-Blondel, Armelle; Kato, Masashi; Schultze, Joachim L; Tartour, Eric; Kroemer, Guido; Degli-Esposti, Mariapia; Chaput, Nathalie; Zitvogel, Laurence

    2012-06-01

    During cancer development, a number of regulatory cell subsets and immunosuppressive cytokines subvert adaptive immune responses. Although it has been shown that tumor-derived interleukin (IL)-18 participates in the PD-1-dependent tumor progression in NK cell-controlled cancers, the mechanistic cues underlying this immunosuppression remain unknown. Here, we show that IL-18 converts a subset of Kit(-) (CD11b(-)) into Kit(+) natural killer (NK) cells, which accumulate in all lymphoid organs of tumor bearers and mediate immunoablative functions. Kit(+) NK cells overexpressed B7-H1/PD-L1, a ligand for PD-1. The adoptive transfer of Kit(+) NK cells promoted tumor growth in two pulmonary metastases tumor models and significantly reduced the dendritic and NK cell pools residing in lymphoid organs in a B7-H1-dependent manner. Neutralization of IL-18 by RNA interference in tumors or systemically by IL-18-binding protein dramatically reduced the accumulation of Kit(+)CD11b(-) NK cells in tumor bearers. Together, our findings show that IL-18 produced by tumor cells elicits Kit(+)CD11b(-) NK cells endowed with B7-H1-dependent immunoablative functions in mice. PMID:22427351

  3. NK cell regulation of CD4 T cell-mediated graft-versus-host disease.

    PubMed

    Noval Rivas, Magali; Hazzan, Marc; Weatherly, Kathleen; Gaudray, Florence; Salmon, Isabelle; Braun, Michel Y

    2010-06-15

    CD3-negative NK cells are granular lymphocytes capable of producing inflammatory cytokines and killing malignant, infected, or stressed cells. We have recently observed a new role for NK cells in the control of the proliferation of CD4 T cells under persistent antigenic stimulation. Monoclonal anti-male CD4 T cells transferred into Rag2-/- male recipients did not expand or were rapidly eliminated. Remarkably, T cells transferred into NK cell-deficient Rag2-/- Il-2Rgammac-/- male hosts expanded extensively and mediated tissue lesions usually observed in chronic graft-versus-host disease (GVHD). T cell failure to proliferate and to induce chronic GVHD was the result of NK cell activity, because depletion of the recipient's NK1.1+ cells by Ab treatment induced T cell expansion and chronic GVHD. T cells under chronic Ag stimulation upregulated ligands of the activating receptor NKG2D, and regulatory activity of NK cells was inhibited by the injection of Abs directed to NKG2D. On the contrary, blocking NKG2A inhibitory receptors did not increase NK cell regulatory activity. Finally, we show that NK regulation of T cell expansion did not involve perforin-mediated lytic activity of NK cells, but depended on T cell surface expression of a functional Fas molecule. These results highlight the potential role played by NK cells in controlling the Ag-specific CD4+ T cells responsible for chronic GVHD. PMID:20488796

  4. NK cells activated by Interleukin-4 in cooperation with Interleukin-15 exhibit distinctive characteristics.

    PubMed

    Kiniwa, Tsuyoshi; Enomoto, Yutaka; Terazawa, Natsumi; Omi, Ai; Miyata, Naoko; Ishiwata, Kenji; Miyajima, Atsushi

    2016-09-01

    Natural killer (NK) cells are known to be activated by Th1-type cytokines, such as IL-2, -12, or -18, and they secrete a large amount of IFN-γ that accelerates Th1-type responses. However, the roles of NK cells in Th2-type responses have remained unclear. Because IL-4 acts as an initiator of Th2-type responses, we examined the characteristics of NK cells in mice overexpressing IL-4. In this study, we report that IL-4 overexpression induces distinctive characteristics of NK cells (B220(high)/CD11b(low)/IL-18Rα(low)), which are different from mature conventional NK (cNK) cells (B220(low)/CD11b(high)/IL-18Rα(high)). IL-4 overexpression induces proliferation of tissue-resident macrophages, which contributes to NK cell proliferation via production of IL-15. These IL-4-induced NK cells (IL4-NK cells) produce higher levels of IFN-γ, IL-10, and GM-CSF, and exhibit high cytotoxicity compared with cNK cells. Furthermore, incubation of cNK cells with IL-15 and IL-4 alters their phenotype to that similar to IL4-NK cells. Finally, parasitic infection, which typically causes strong Th2-type responses, induces the development of NK cells with characteristics similar to IL4-NK cells. These IL4-NK-like cells do not develop in IL-4Rα KO mice by parasitic infection. Collectively, these results suggest a novel role of IL-4 in immune responses through the induction of the unique NK cells. PMID:27551096

  5. CD3+/CD19+-depleted grafts in HLA-matched allogeneic peripheral blood stem cell transplantation lead to early NK cell cytolytic responses and reduced inhibitory activity of NKG2A.

    PubMed

    Eissens, D N; Schaap, N P M; Preijers, F W M B; Dolstra, H; van Cranenbroek, B; Schattenberg, A V M; Joosten, I; van der Meer, A

    2010-03-01

    Natural killer (NK) cells have an important function in the anti-tumor response early after stem cell transplantation (SCT). As part of a prospective randomized phase III study, directly comparing the use of CD3(+)/CD19(+)-depleted peripheral blood stem cell (PBSC) harvests with CD34(+)-selected PBSC harvests in allogeneic human leukocyte antigen-matched SCT, we here show that the use of CD3(+)/CD19(+)-depleted PBSC grafts leads to early NK cell repopulation and reconstitution of the CD56(dim) and CD56(bright) NK cell subsets, with concomitant high cytolytic capacity. In the CD34 group, this process took significantly longer. Moreover, in the CD3/19 group after reconstitution, a higher percentage of killer immunoglobulin-like receptor-positive NK cells was found. Although similar percentages of CD94-positive NK cells were found in both groups, in the CD34 group, almost all expressed the inhibitory CD94:NKG2A complex, whereas in the CD3/19 group, the inhibitory CD94:NKG2A and the activating CD94:NKG2C complex were equally distributed. This preferential development of NKG2C-expressing NK cells in the CD3/19 group was paralleled by a loss of NKG2A-mediated inhibition of NK cell degranulation. These results show that the use of CD3(+)/CD19(+)-depleted grafts facilitates strong NK cell cytolytic responses directly after SCT, and the rapid emergence of an NK cell receptor phenotype that is more prone to activation. PMID:20033055

  6. NK cells, autoantibodies, and immunologic infertility: a complex interplay.

    PubMed

    De Carolis, Caterina; Perricone, Carlo; Perricone, Roberto

    2010-12-01

    Infertility and recurrent spontaneous abortion (RSA) are heterogeneous conditions that have been frequently explained with an immunological pathomechanism. A deeper insight into apparently unexplained infertility and RSA shows increasing evidences supporting both alloimmune and autoimmune mechanisms, in which natural killer (NK) cells and autoantibodies seem to play a relevant role. Successful pregnancy is considered as Th1-Th2 cooperation phenomenon, with a predominantly Th2-type lymphocytes response, together with the emerging role of interleukin (IL)-12, IL-15, and IL-18 and of other unidentified soluble factors dependent on NK cells. Uterine NK cells comprise the largest population at implantation site, and their activity, characteristics, and abundance suggest that they participate at the "decidualization" process that, vice versa, induces NK activation and recruitment in each menstrual cycle. However, NK cell alteration may be associated with impaired pregnancy, and the modulation in the number of circulating NK cells is most likely to be a primary event rather than an active inflammation/drug administration consequence during an inflammatory/autoimmune process, thus playing an important role in the pathogenesis of immunological infertility. Relationships within immunological infertility, recurrent spontaneous abortion, autoantibodies, and NK cells will be reviewed herein. PMID:19908167

  7. Mobilization of NK cells by exercise: downmodulation of adhesion molecules on NK cells by catecholamines.

    PubMed

    Nagao, F; Suzui, M; Takeda, K; Yagita, H; Okumura, K

    2000-10-01

    The change of plasma catecholamine concentration correlates with the change of natural killer (NK) activity and NK cell number in peripheral blood mononuclear cells (PBMC) during and after moderate exercise. We studied the causal relation between exercise-induced catecholamine and expression of adhesion molecules on NK cells during and after exercise. The expression of CD44 and CD18 on CD3(-)CD56(+) NK cells was significantly reduced during exercise (P < 0.01). When PBMC were stimulated with 10(-8)M norepinephrine in vitro, the expression of these adhesion molecules on CD3(-)CD56(+) NK cells was downmodulated within 30 min. The binding capacity of NK cells to a CD44 ligand, hyaluronate, was reduced by the stimulation with norepinephrine (P < 0.01). The intravenous injection of norepinephrine in mice decreased the expression of CD44 and CD18 on CD3(-)NK1.1(+) cells (P < 0.01) and increased the number of CD3(-)NK1.1(+) cells in PBMC (P < 0.01). These findings suggest that exercise-induced catecholamines modulate the expression of adhesion molecules on NK cells, resulting in the mobilization of NK cells into the circulation. PMID:11003990

  8. A Modified NK Cell Degranulation Assay Applicable for Routine Evaluation of NK Cell Function

    PubMed Central

    Shabrish, Snehal; Gupta, Maya; Madkaikar, Manisha

    2016-01-01

    Natural killer (NK) cells play important role in innate immunity against tumors and viral infections. Studies show that lysosome-associated membrane protein-1 (LAMP-1, CD107a) is a marker for degranulation of NK and cytotoxic T cells and its expression is a sensitive marker for the cytotoxic activity determination. The conventional methods of determination of CD107a on NK cells involve use of peripheral blood mononuclear cells (PBMC) or pure NK cells and K562 cells as stimulants. Thus, it requires large volume of blood sample which is usually difficult to obtain in pediatric patients and patients with cytopenia and also requires specialized laboratory for maintaining cell line. We have designed a flow cytometric assay to determine CD107a on NK cells using whole blood, eliminating the need for isolation of PBMC or isolate NK cells. This assay uses phorbol-12-myristate-13-acetate (PMA) and calcium ionophore (Ca2+-ionophore) instead of K562 cells for stimulation and thus does not require specialized cell culture laboratory. CD107a expression on NK cells using modified NK cell degranulation assay compared to the conventional assay was significantly elevated (p < 0.0001). It was also validated by testing patients diagnosed with familial hemophagocytic lymphohistiocytosis (FHL) with defect in exocytosis. This assay is rapid, cost effective, and reproducible and requires significantly less volume of blood which is important for clinical evaluation of NK cells. PMID:27413758

  9. NK cells: tuned by peptide?

    PubMed

    Das, Jayajit; Khakoo, Salim I

    2015-09-01

    Natural killer cells express multiple receptors for major histocompatibility complex (MHC) class I, including the killer cell immunoglobulin-like receptors (KIRs) and the C-type lectin-like CD94:NKG2 receptors. The KIR locus is extremely polymorphic, paralleling the diversity of its classical MHC class I ligands. Similarly, the conservation of the NKG2 family of receptors parallels the conservation of MHC-E, the ligand for CD94:NKG2A/C/E. Binding of both CD94:NKG2 heterodimers and KIR to their respective MHC class I ligand is peptide dependent, and despite the evolution of these receptors, they have retained the property of peptide selectivity. Such peptide selectivity affects these two systems in different ways. HLA-E binding non-inhibitory peptides augment inhibition at CD94:NKG2A, while HLA-C binding non-inhibitory peptides antagonize inhibition at KIR2DL2/3, implying that KIRs are specialized to respond positively to changes in peptide repertoire. Thus, while specific KIRs, such as KIR2DL3, are associated with beneficial outcomes from viral infections, viral peptides augment inhibition at CD94:NKGA. Conversely, NKG2A-positive NK cells sense MHC class I downregulation more efficiently than KIRs. Thus, these two receptor:ligand systems appear to have complementary functions in recognizing changes in MHC class I. PMID:26284480

  10. NK Cells during Dengue Disease and Their Recognition of Dengue Virus-Infected cells

    PubMed Central

    Beltrán, Davis; López-Vergès, Sandra

    2014-01-01

    The innate immune response, in addition to the B- and T-cell response, plays a role in protection against dengue virus (DENV) infection and the degree of disease severity. Early activation of natural killer (NK) cells and type-I interferon-dependent immunity may be important in limiting viral replication during the early stages of DENV infection and thus reducing subsequent pathogenesis. NK cells may also produce cytokines that reduce inflammation and tissue injury. On the other hand, NK cells are also capable of inducing liver injury at early-time points of DENV infection. In vitro, NK cells can kill antibody-coated DENV-infected cells through antibody-dependent cell-mediated cytotoxicity. In addition, NK cells may directly recognize DENV-infected cells through their activating receptors, although the increase in HLA class I expression may allow infected cells to escape the NK response. Recently, genome-wide association studies have shown an association between MICB and MICA, which encode ligands of the activating NK receptor NKG2D, and dengue disease outcome. This review focuses on recognition of DENV-infected cells by NK cells and on the regulation of expression of NK cell ligands by DENV. PMID:24829565

  11. Acute response of peripheral CCr5 chemoreceptor and NK cells in individuals submitted to a single session of low-intensity strength exercise with blood flow restriction.

    PubMed

    Dorneles, Gilson Pires; Colato, Alana Schraiber; Galvão, Simone Lunelli; Ramis, Thiago Rozales; Ribeiro, Jerri Luiz; Romão, Pedro Roosevelt; Peres, Alessandra

    2016-07-01

    The purpose of this study was to compare the peripheral expression of natural killers and CCR5 in a session of low-intensity strength training with vascular occlusion and in high-intensity training. Young males were randomized into session groups of a high-intensity strength training (HI) and a session group of low-intensity strength training with vascular occlusion (LI-BFR). The exercise session consisted in knee extension and bicep curl in 80% 1RM (HI) and 30% 1RM (LI-BFR) with equalized volumes. Blood collection was made before, immediately after and 24 h after each training session. Immunophenotyping was carried out through CD195+ (CCR5) e CD3-CD16+CD56+ (NK) in peripheral blood and analysed by flow cytometry and presented in frequency (%). Peripheral frequency of NK cells showed no significant difference in LI-BFR group in time effect, while a gradual reduction of NK cells was identified in HI group in before-24 h postexercise and after-24 h postexercise comparison. However, significant differences have been found in relative change of NK cells immediately after exercise between sessions. In addition, HI and LI-BFR groups showed a significant reduction in the cells expressed CCR5 during 24 h postsession compared to the postsession, but CCR5 also differed when comparing before-24 h after session in the HI group. No differences were observed amongst the groups. LIO induced CCR5 response similar to the HI session, while the NK cells remained in similar frequency during the studied moments in LI-BFR, but not in HI group, suggesting that local hypoxia created by the blood flow restriction was able to prevent a change in the frequency of peripheral cells and a possible immunosuppression. PMID:25643617

  12. Type I IFN promotes NK cell expansion during viral infection by protecting NK cells against fratricide.

    PubMed

    Madera, Sharline; Rapp, Moritz; Firth, Matthew A; Beilke, Joshua N; Lanier, Lewis L; Sun, Joseph C

    2016-02-01

    Type I interferon (IFN) is crucial in host antiviral defense. Previous studies have described the pleiotropic role of type I IFNs on innate and adaptive immune cells during viral infection. Here, we demonstrate that natural killer (NK) cells from mice lacking the type I IFN-α receptor (Ifnar(-/-)) or STAT1 (which signals downstream of IFNAR) are defective in expansion and memory cell formation after mouse cytomegalovirus (MCMV) infection. Despite comparable proliferation, Ifnar(-/-) NK cells showed diminished protection against MCMV infection and exhibited more apoptosis compared with wild-type NK cells. Furthermore, we show that Ifnar(-/-) NK cells express increased levels of NK group 2 member D (NKG2D) ligands during viral infection and are susceptible to NK cell-mediated fratricide in a perforin- and NKG2D-dependent manner. Adoptive transfer of Ifnar(-/-) NK cells into NK cell-deficient mice reverses the defect in survival and expansion. Our study reveals a novel type I IFN-dependent mechanism by which NK cells evade mechanisms of cell death after viral infection. PMID:26755706

  13. Microchip Screening Platform for Single Cell Assessment of NK Cell Cytotoxicity

    PubMed Central

    Guldevall, Karolin; Brandt, Ludwig; Forslund, Elin; Olofsson, Karl; Frisk, Thomas W.; Olofsson, Per E.; Gustafsson, Karin; Manneberg, Otto; Vanherberghen, Bruno; Brismar, Hjalmar; Kärre, Klas; Uhlin, Michael; Önfelt, Björn

    2016-01-01

    Here, we report a screening platform for assessment of the cytotoxic potential of individual natural killer (NK) cells within larger populations. Human primary NK cells were distributed across a silicon–glass microchip containing 32,400 individual microwells loaded with target cells. Through fluorescence screening and automated image analysis, the numbers of NK and live or dead target cells in each well could be assessed at different time points after initial mixing. Cytotoxicity was also studied by time-lapse live-cell imaging in microwells quantifying the killing potential of individual NK cells. Although most resting NK cells (≈75%) were non-cytotoxic against the leukemia cell line K562, some NK cells were able to kill several (≥3) target cells within the 12-h long experiment. In addition, the screening approach was adapted to increase the chance to find and evaluate serial killing NK cells. Even if the cytotoxic potential varied between donors, it was evident that a small fraction of highly cytotoxic NK cells were responsible for a substantial portion of the killing. We demonstrate multiple assays where our platform can be used to enumerate and characterize cytotoxic cells, such as NK or T cells. This approach could find use in clinical applications, e.g., in the selection of donors for stem cell transplantation or generation of highly specific and cytotoxic cells for adoptive immunotherapy. PMID:27092139

  14. Uterine NK cells: active regulators at the maternal-fetal interface

    PubMed Central

    Moffett, Ashley; Colucci, Francesco

    2014-01-01

    Pregnancy presents an immunological conundrum because two genetically different individuals coexist. The maternal lymphocytes at the uterine maternal-fetal interface that can recognize mismatched placental cells are T cells and abundant distinctive uterine NK (uNK) cells. Multiple mechanisms exist that avoid damaging T cell responses to the fetus, whereas activation of uNK cells is probably physiological. Indeed, genetic epidemiological data suggest that the variability of NK cell receptors and their MHC ligands define pregnancy success; however, exactly how uNK cells function in normal and pathological pregnancy is still unclear, and any therapies aimed at suppressing NK cells must be viewed with caution. Allorecognition of fetal placental cells by uNK cells is emerging as the key maternal-fetal immune mechanism that regulates placentation. PMID:24789879

  15. NK Cells Help Induce Anti-Hepatitis B Virus CD8+ T Cell Immunity in Mice.

    PubMed

    Zheng, Meijuan; Sun, Rui; Wei, Haiming; Tian, Zhigang

    2016-05-15

    Although recent clinical studies demonstrate that NK cell function is impaired in hepatitis B virus (HBV)-persistent patients, whether or how NK cells play a role in anti-HBV adaptive immunity remains to be explored. Using a mouse model mimicking acute HBV infection by hydrodynamic injection of an HBV plasmid, we observed that although serum hepatitis B surface Ag and hepatitis B envelope Ag were eliminated within 3 to 4 wk, HBV might persist for >8 wk in CD8(-/-) mice and that adoptive transfer of anti-HBV CD8(+) T cells restored the ability to clear HBV in HBV-carrier Rag1(-/-) mice. These results indicate that CD8(+) T cells are critical in HBV elimination. Furthermore, NK cells increased IFN-γ production after HBV plasmid injection, and NK cell depletion led to significantly increased HBV persistence along with reduced frequency of hepatitis B core Ag-specific CD8(+) T cells. Adoptive transfer of IFN-γ-sufficient NK cells restored donor CD8(+) T cell function, indicating that NK cells positively regulated CD8(+) T cells via secreting IFN-γ. We also observed that NK cell depletion correlated with decreased effector memory CD8(+) T cell frequencies. Importantly, adoptive transfer experiments showed that NK cells were involved in anti-HBV CD8(+) T cell recall responses. Moreover, DX5(+)CD49a(-) conventional, but not DX5(-)CD49a(+) liver-resident, NK cells were involved in improving CD8(+) T cell responses against HBV. Overall, the current study reveals that NK cells, especially DX5(+)CD49a(-) conventional NK cells, promote the antiviral activity of CD8(+) T cell responses via secreting IFN-γ in a mouse model mimicking acute HBV infection. PMID:27183639

  16. Functional Assessment of NK and LAK Cells Following Space Flight

    NASA Technical Reports Server (NTRS)

    Kaur, Indreshpal; Pierson, Duane L.; Paloski, W. H. (Technical Monitor)

    1999-01-01

    Space flight associated stress alters some aspects of the human immune response. In this study, we determined the effects of 10 days aboard the Space Shuttle on the cytotoxic activity of NK and LAK cells. PBMCs were collected from 10-ml blood specimens from 5 astronauts 10 days before launch, immediately after landing, and again at 3 days after landing and stored at -80 C. All PBMCs were thawed simultaneously, and the cytotoxic activities of NK and LAK cells were measured by a 4 hour Cr-51 release assay. K562 cells were used to assess NK cell cytotoxicity. Following 4 days of IL-2 activation, the LAK cell cytotoxic activity was determined using K562 cells and Daudi cells as the target cells. NK cell cytotoxicity decreased at landing (p<.05) in 3/5 astronauts, and recovered to preflight levels by 3 days following landing; NK cell cytotoxicity was increased (p=0.1) in the remaining 2 astronauts at landing. In 4/5 astronauts, LAK cytotoxic activity was decreased at landing against K562 cells (p = 0.13) and Daudi cells (p = 0.08). Phenotyping of PBMC's and LAK cells showed alterations in some surface markers and adhesion molecules (CD11b, CD11c, CD11a, CD16, L-selectin, and CD3).

  17. Murine peripheral NK-cell populations originate from site-specific immature NK cells more than from BM-derived NK cells.

    PubMed

    Pinhas, Nissim; Sternberg-Simon, Michal; Chiossone, Laura; Shahaf, Gitit; Walzer, Thierry; Vivier, Eric; Mehr, Ramit

    2016-05-01

    Murine NK cells can be divided by the expression of two cell surface markers, CD27 and Mac-1 (a.k.a. CD11b), into four separate subsets. These subsets suggest a linear development model: CD27(-) Mac-1(-) → CD27(+) Mac-1(-) → CD27(+) Mac-1(+) → CD27(-) Mac-1(+) . Here, we used a combination of BrdU labeling experiments and mathematical modeling to gain insights regarding NK-cell development in mouse bone marrow (BM), spleen and liver. The modeling results that best fit the experimental data show that the majority of NK cells already express CD27 upon entering the NK-cell developmental pathway. Additionally, only a small fraction of NK cells exit the BM to other sites, suggesting that peripheral NK-cell populations originate from site-specific immature NK cells more than from BM-derived mature NK cells. PMID:26919267

  18. Licensed and Unlicensed NK Cells: Differential Roles in Cancer and Viral Control

    PubMed Central

    Tu, Megan M.; Mahmoud, Ahmad Bakur; Makrigiannis, Andrew P.

    2016-01-01

    Natural killer (NK) cells are known for their well characterized ability to control viral infections and eliminate tumor cells. Through their repertoire of activating and inhibitory receptors, NK cells are able to survey different potential target cells for various surface markers, such as MHC-I – which signals to the NK cell that the target is healthy – as well as stress ligands or viral proteins, which alert the NK cell to the aberrant state of the target and initiate a response. According to the “licensing” hypothesis, interactions between self-specific MHC-I receptors – Ly49 in mice and KIR in humans – and self-MHC-I molecules during NK cell development is crucial for NK cell functionality. However, there also exists a large proportion of NK cells in mice and humans, which lack self-specific MHC-I receptors and are consequentially “unlicensed.” While the licensed NK cell subset plays a major role in the control of MHC-I-deficient tumors, this review will go on to highlight the important role of the unlicensed NK cell subset in the control of MHC-I-expressing tumors, as well as in viral control. Unlike the licensed NK cells, unlicensed NK cells seem to benefit from the lack of self-specific inhibitory receptors, which could otherwise be exploited by some aberrant cells for immunoevasion by upregulating the expression of ligands or mimic ligands for these receptors. PMID:27199990

  19. Licensed and Unlicensed NK Cells: Differential Roles in Cancer and Viral Control.

    PubMed

    Tu, Megan M; Mahmoud, Ahmad Bakur; Makrigiannis, Andrew P

    2016-01-01

    Natural killer (NK) cells are known for their well characterized ability to control viral infections and eliminate tumor cells. Through their repertoire of activating and inhibitory receptors, NK cells are able to survey different potential target cells for various surface markers, such as MHC-I - which signals to the NK cell that the target is healthy - as well as stress ligands or viral proteins, which alert the NK cell to the aberrant state of the target and initiate a response. According to the "licensing" hypothesis, interactions between self-specific MHC-I receptors - Ly49 in mice and KIR in humans - and self-MHC-I molecules during NK cell development is crucial for NK cell functionality. However, there also exists a large proportion of NK cells in mice and humans, which lack self-specific MHC-I receptors and are consequentially "unlicensed." While the licensed NK cell subset plays a major role in the control of MHC-I-deficient tumors, this review will go on to highlight the important role of the unlicensed NK cell subset in the control of MHC-I-expressing tumors, as well as in viral control. Unlike the licensed NK cells, unlicensed NK cells seem to benefit from the lack of self-specific inhibitory receptors, which could otherwise be exploited by some aberrant cells for immunoevasion by upregulating the expression of ligands or mimic ligands for these receptors. PMID:27199990

  20. Polyclonal Expansion of NKG2C+ NK Cells in TAP-Deficient Patients

    PubMed Central

    Béziat, Vivien; Sleiman, Marwan; Goodridge, Jodie P.; Kaarbø, Mari; Liu, Lisa L.; Rollag, Halvor; Ljunggren, Hans-Gustaf; Zimmer, Jacques; Malmberg, Karl-Johan

    2015-01-01

    Adaptive natural killer (NK) cell responses to human cytomegalovirus infection are characterized by the expansion of NKG2C+ NK cells expressing self-specific inhibitory killer-cell immunoglobulin-like receptors (KIRs). Here, we set out to study the HLA class I dependency of such NKG2C+ NK cell expansions. We demonstrate the expansion of NKG2C+ NK cells in patients with transporter associated with antigen presentation (TAP) deficiency, who express less than 10% of normal HLA class I levels. In contrast to normal individuals, expanded NKG2C+ NK cell populations in TAP-deficient patients display a polyclonal KIR profile and remain hyporesponsive to HLA class I-negative target cells. Nonetheless, agonistic stimulation of NKG2C on NK cells from TAP-deficient patients yielded significant responses in terms of degranulation and cytokine production. Thus, while interactions with self-HLA class I molecules likely shape the KIR repertoire of expanding NKG2C+ NK cells during adaptive NK cell responses in normal individuals, they are not a prerequisite for NKG2C+ NK cell expansions to occur. The emergence of NKG2C-responsive adaptive NK cells in TAP-deficient patients may contribute to antiviral immunity and potentially explain these patients’ low incidence of severe viral infections. PMID:26500647

  1. Loss of STAT3 in murine NK cells enhances NK cell-dependent tumor surveillance.

    PubMed

    Gotthardt, Dagmar; Putz, Eva M; Straka, Elisabeth; Kudweis, Petra; Biaggio, Mario; Poli, Valeria; Strobl, Birgit; Müller, Mathias; Sexl, Veronika

    2014-10-01

    The members of the signal transducer and activator of transcription (STAT) family of transcription factors modulate the development and function of natural killer (NK) cells. NK cell-mediated tumor surveillance is particularly important in the body's defense against hematological malignancies such as leukemia. STAT3 inhibitors are currently being developed, although their potential effects on NK cells are not clear. We have investigated the function of STAT3 in NK cells with Stat3(Δ/Δ)Ncr1-iCreTg mice, whose NK cells lack STAT3. In the absence of STAT3, NK cells develop normally and in normal numbers, but display alterations in the kinetics of interferon-γ (IFN-γ) production. We report that STAT3 directly binds the IFN-γ promoter. In various in vivo models of hematological diseases, loss of STAT3 in NK cells enhances tumor surveillance. The reduced tumor burden is paralleled by increased expression of the activating receptor DNAM-1 and the lytic enzymes perforin and granzyme B. Our findings imply that STAT3 inhibitors will stimulate the cytolytic activity of NK cells against leukemia, thereby providing an additional therapeutic benefit. PMID:25185262

  2. NK cells modulate the inflammatory response to corneal epithelial abrasion and thereby support wound healing

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Natural killer cells are lymphocytes of the innate immune system that have crucial cytotoxic and regulatory roles in adaptive immunity and inflammation. Herein, we consider a role for these cells in corneal wound healing. After a 2-mm central epithelial abrasion of the mouse cornea, a subset of clas...

  3. Inflammasome-Dependent Induction of Adaptive NK Cell Memory.

    PubMed

    van den Boorn, Jasper G; Jakobs, Christopher; Hagen, Christian; Renn, Marcel; Luiten, Rosalie M; Melief, Cornelis J M; Tüting, Thomas; Garbi, Natalio; Hartmann, Gunther; Hornung, Veit

    2016-06-21

    Monobenzone is a pro-hapten that is exclusively metabolized by melanocytes, thereby haptenizing melanocyte-specific antigens, which results in cytotoxic autoimmunity specifically against pigmented cells. Studying monobenzone in a setting of contact hypersensitivity (CHS), we observed that monobenzone induced a long-lasting, melanocyte-specific immune response that was dependent on NK cells, yet fully intact in the absence of T- and B cells. Consistent with the concept of "memory NK cells," monobenzone-induced NK cells resided in the liver and transfer of these cells conferred melanocyte-specific immunity to naive animals. Monobenzone-exposed skin displayed macrophage infiltration and cutaneous lymph nodes showed an inflammasome-dependent influx of macrophages with a tissue-resident phenotype, coinciding with local NK cell activation. Indeed, macrophage depletion or the absence of the NLRP3 inflammasome, the adaptor protein ASC or interleukin-18 (IL-18) abolished monobenzone CHS, thereby establishing a non-redundant role for the NLRP3 inflammasome as a critical proinflammatory checkpoint in the induction of hapten-dependent memory NK cells. PMID:27287410

  4. Epirubicin pretreatment enhances NK cell-mediated cytotoxicity against breast cancer cells in vitro

    PubMed Central

    Feng, Hui; Dong, Ying; Wu, Jing; Qiao, Yuan; Zhu, Ge; Jin, Haofan; Cui, Jiuwei; Li, Wei; Liu, Yong-Jun; Chen, Jingtao; Song, Yanqiu

    2016-01-01

    Anthracycline-based chemotherapy is a conventional treatment for breast cancer. However, it can negatively affect host immune function and thereby impair patients’ quality of life. Boosting the host immune system and reducing the adverse effect of chemotherapy are important for effective cancer treatment. Natural killer (NK) cells stimulate immune responses against cancer; autologous immune enhancement therapy with NK cells prolongs patient survival without significant adverse effects. This study investigated the effects of combined treatment with the anthracycline agent epirubicin (EPI) and NK cells on human breast cancer cells. NK cells were obtained by autologous adoptive cell transfer from breast cancer patients and amplified for 14 days in vitro. The cytotoxicity of NK cells against breast cancer cells was higher following EPI (5.0 μg/ml) pretreatment than without EPI pretreatment or application of EPI alone. The expression of NKG2D ligands [unique long 16-binding protein (ULBP) 1, ULBP2, and major histocompatibility complex class I-related chain A] in breast cancer cells was upregulated by pretreatment with EPI, which also increased the secretion of interferon-γ and tumor necrosis factor-α and expression of perforin and granzyme B in NK cells. These results indicate that EPI-NK cell treatment has synergistic cytotoxic effects against breast cancer cells, and suggest that anthracycline-based chemotherapy and NK cell-based immunotherapy can be combined for more effective breast cancer treatment. PMID:27158340

  5. KIR2DL2 inhibitory pathway enhances Th17 cytokine secretion by NK cells in response to herpesvirus infection in multiple sclerosis patients.

    PubMed

    Rizzo, Roberta; Bortolotti, Daria; Fainardi, Enrico; Gentili, Valentina; Bolzani, Silvia; Baldi, Eleonora; Casetta, Ilaria; Granieri, Enrico; Rotola, Antonella; Furlan, Roberto; Di Luca, Dario

    2016-05-15

    We have previously demonstrated that multiple sclerosis (MS) patients with KIR2DL2 expression on Natural killer (NK) cells are more susceptible to herpes simplex virus 1 (HSV-1) infection. We explored cytokine expression by NK cells during HSV-1 infection in association with KIR2DL2 expression. MS KIR2DL2(+) NK cells failed to control HSV-1 infection and secreted high levels of Th17 cytokines, while MS KIR2DL2(-) NK cells released Th1 cytokines, mainly IFN-gamma. Our data showed, for the first time, a peculiar Th17 cytokine secretion by MS KIR2DL2(+) NK cells in the presence of HSV-1 infection, that could be implicated in MS pathogenesis. PMID:27138091

  6. NKG2D is a Key Receptor for Recognition of Bladder Cancer Cells by IL-2-Activated NK Cells and BCG Promotes NK Cell Activation

    PubMed Central

    García-Cuesta, Eva María; López-Cobo, Sheila; Álvarez-Maestro, Mario; Esteso, Gloria; Romera-Cárdenas, Gema; Rey, Mercedes; Cassady-Cain, Robin L.; Linares, Ana; Valés-Gómez, Alejandro; Reyburn, Hugh Thomson; Martínez-Piñeiro, Luis; Valés-Gómez, Mar

    2015-01-01

    Intravesical instillation of bacillus Calmette–Guérin (BCG) is used to treat superficial bladder cancer, either papillary tumors (after transurethral resection) or high-grade flat carcinomas (carcinoma in situ), reducing recurrence in about 70% of patients. Initially, BCG was proposed to work through an inflammatory response, mediated by phagocytic uptake of mycobacterial antigens and cytokine release. More recently, other immune effectors such as monocytes, natural killer (NK), and NKT cells have been suggested to play a role in this immune response. Here, we provide a comprehensive study of multiple bladder cancer cell lines as putative targets for immune cells and evaluated their recognition by NK cells in the presence and absence of BCG. We describe that different bladder cancer cells can express multiple activating and inhibitory ligands for NK cells. Recognition of bladder cancer cells depended mainly on NKG2D, with a contribution from NKp46. Surprisingly, exposure to BCG did not affect the immune phenotype of bladder cells nor increased NK cell recognition of purified IL-2-activated cell lines. However, NK cells were activated efficiently when BCG was included in mixed lymphocyte cultures, suggesting that NK activation after mycobacteria treatment requires the collaboration of various immune cells. We also analyzed the percentage of NK cells in peripheral blood of a cohort of bladder cancer patients treated with BCG. The total numbers of NK cells did not vary during treatment, indicating that a more detailed study of NK cell activation in the tumor site will be required to evaluate the response in each patient. PMID:26106390

  7. Adenovirus type 35, but not type 5, stimulates NK cell activation via plasmacytoid dendritic cells and TLR9 signaling.

    PubMed

    Pahl, Jens H W; Verhoeven, Dirk H J; Kwappenberg, Kitty M C; Vellinga, Jort; Lankester, Arjan C; van Tol, Maarten J D; Schilham, Marco W

    2012-05-01

    In hematopoietic stem cell transplant (HSCT) recipients, disseminated adenoviral infections during the first two months after HSCT can lead to severe complications and fatal outcome. Since NK cells are usually the first lymphocytes to reconstitute after HSCT and have been implicated in the clearance of adenovirus-infected cells, it was investigated whether NK cells are activated by adenovirus in vitro. Exposure of PBMC to human adenovirus type 5 (HAdV5) or HAdV35 resulted in the up-regulation of the activation marker CD69 on NK cells and enhanced the cytolytic activity of NK cells. HAdV5-induced NK cell activation relied on the contribution of T cells as the depletion of T cells from PBMC abolished NK cell activation. In contrast, NK cell activation in response to HAdV35 occurred in the absence of T cells. Plasmacytoid dendritic cells (pDC) were necessary and sufficient to mediate NK cell activation. HAdV35 induced significantly more interferon-α (IFN-α) production by pDC than HAdV5. The increased IFN-α production and NK cell activation correlated with a higher infection efficiency of viruses with the type 35 fiber. The IFN-α response of pDC was enhanced by the presence of NK cells, suggesting a reciprocal interaction between pDC and NK cells. Incubation with a TLR9 antagonist impaired the IFN-α production by pDC as well as NK cell activation, implying that TLR9 signaling is critically involved in the IFN-α response of pDC and NK cell activation after HAdV35 exposure. In conclusion, two human adenovirus serotypes from two different species differ considerably in their capacity to stimulate pDC and NK cells. PMID:22424784

  8. CD19-CAR engineered NK-92 cells are sufficient to overcome NK cell resistance in B-cell malignancies.

    PubMed

    Romanski, Annette; Uherek, Christoph; Bug, Gesine; Seifried, Erhard; Klingemann, Hans; Wels, Winfried S; Ottmann, Oliver G; Tonn, Torsten

    2016-07-01

    Many B-cell acute and chronic leukaemias tend to be resistant to killing by natural killer (NK) cells. The introduction of chimeric antigen receptors (CAR) into T cells or NK cells could potentially overcome this resistance. Here, we extend our previous observations on the resistance of malignant lymphoblasts to NK-92 cells, a continuously growing NK cell line, showing that anti-CD19-CAR (αCD19-CAR) engineered NK-92 cells can regain significant cytotoxicity against CD19 positive leukaemic cell lines and primary leukaemia cells that are resistant to cytolytic activity of parental NK-92 cells. The 'first generation' CAR was generated from a scFv (CD19) antibody fragment, coupled to a flexible hinge region, the CD3ζ chain and a Myc-tag and cloned into a retrovirus backbone. No difference in cytotoxic activity of NK-92 and transduced αCD19-CAR NK-92 cells towards CD19 negative targets was found. However, αCD19-CAR NK-92 cells specifically and efficiently lysed CD19 expressing B-precursor leukaemia cell lines as well as lymphoblasts from leukaemia patients. Since NK-92 cells can be easily expanded to clinical grade numbers under current Good Manufactoring Practice (cGMP) conditions and its safety has been documented in several phase I clinical studies, treatment with CAR modified NK-92 should be considered a treatment option for patients with lymphoid malignancies. PMID:27008316

  9. Pseudorabies Virus US3 Protein Kinase Protects Infected Cells from NK Cell-Mediated Lysis via Increased Binding of the Inhibitory NK Cell Receptor CD300a

    PubMed Central

    Grauwet, K.; Vitale, M.; De Pelsmaeker, S.; Jacob, T.; Laval, K.; Moretta, L.; Parodi, M.; Parolini, S.; Cantoni, C.

    2015-01-01

    ABSTRACT Several reports have indicated that natural killer (NK) cells are of particular importance in the innate response against herpesvirus infections. As a consequence, herpesviruses have developed diverse mechanisms for evading NK cells, although few such mechanisms have been identified for the largest herpesvirus subfamily, the alphaherpesviruses. The antiviral activity of NK cells is regulated by a complex array of interactions between activating/inhibitory receptors on the NK cell surface and the corresponding ligands on the surfaces of virus-infected cells. Here we report that the US3 protein kinase of the alphaherpesvirus pseudorabies virus (PRV) displays previously uncharacterized immune evasion properties: it triggers the binding of the inhibitory NK cell receptor CD300a to the surface of the infected cell, thereby providing increased CD300a-mediated protection of infected cells against NK cell-mediated lysis. US3-mediated CD300a binding was found to depend on aminophospholipid ligands of CD300a and on group I p21-activated kinases. These data identify a novel alphaherpesvirus strategy for evading NK cells and demonstrate, for the first time, a role for CD300a in regulating NK cell activity upon contact with virus-infected target cells. IMPORTANCE Herpesviruses have developed fascinating mechanisms to evade elimination by key elements of the host immune system, contributing to their ability to cause lifelong infections with recurrent reactivation events. Natural killer (NK) cells are central in the innate antiviral response. Here we report that the US3 protein kinase of the alphaherpesvirus pseudorabies virus displays a previously uncharacterized capacity for evasion of NK cells. Expression of US3 protects infected cells from NK cell-mediated lysis via increased binding of the inhibitory NK cell receptor CD300a. We show that this US3-mediated increase in CD300a binding depends on aminophospholipids and on cellular p21-activated kinases (PAKs). The

  10. Transcription factor KLF2 regulates homeostatic NK cell proliferation and survival.

    PubMed

    Rabacal, Whitney; Pabbisetty, Sudheer K; Hoek, Kristen L; Cendron, Delphine; Guo, Yin; Maseda, Damian; Sebzda, Eric

    2016-05-10

    Natural killer (NK) cells are innate lymphocytes that recognize and lyse virally infected or transformed cells. This latter property is being pursued in clinics to treat leukemia with the hope that further breakthroughs in NK cell biology can extend treatments to other cancers. At issue is the ability to expand transferred NK cells and prolong their functionality within the context of a tumor. In terms of NK cell expansion and survival, we now report that Kruppel-like factor 2 (KLF2) is a key transcription factor that underpins both of these events. Excision of Klf2 using gene-targeted mouse models promotes spontaneous proliferation of immature NK cells in peripheral tissues, a phenotype that is replicated under ex vivo conditions. Moreover, KLF2 imprints a homeostatic migration pattern on mature NK cells that allows these cells to access IL-15-rich microenvironments. KLF2 accomplishes this feat within the mature NK cell lineage via regulation of a subset of homing receptors that respond to homeostatic ligands while leaving constitutively expressed receptors that recognize inflammatory cytokines unperturbed. Under steady-state conditions, KLF2-deficient NK cells alter their expression of homeostatic homing receptors and subsequently undergo apoptosis due to IL-15 starvation. This novel mechanism has implications regarding NK cell contraction following the termination of immune responses including the possibility that retention of an IL-15 transpresenting support system is key to extending NK cell activity in a tumor environment. PMID:27114551

  11. CRACC-targeting Fc-fusion protein induces activation of NK cells and DCs and improves T cell immune responses to antigenic targets.

    PubMed

    Aldhamen, Yasser A; Rastall, David P W; Chen, Weimin; Seregin, Sergey S; Pereira-Hicks, Cristiane; Godbehere, Sarah; Kaminski, Norbert E; Amalfitano, Andrea

    2016-06-01

    The CD2-like receptor activating cytotoxic cell (CRACC) receptor is a member of the SLAM family of receptors that are found on several types of immune cells. We previously demonstrated that increasing the abundance of the adaptor protein EAT-2 during vaccination enhanced innate and adaptive immune responses to vaccine antigens. Engagement of the CRACC receptor in the presence of the EAT-2 adaptor generally results in immune cell activation, while activating CRACC signaling in cells that lack EAT-2 adaptor inhibits their effector and regulatory functions. As EAT-2 is the only SAP adaptor that interacts with the CRACC receptor, we hypothesized that technologies that specifically modulate CRACC signaling during vaccination may also improve antigen specific adaptive immune responses. To test this hypothesis, we constructed a CRACC-targeting Fc fusion protein and included it in vaccination attempts. Indeed, mice co-vaccinated with the CRACC-Fc fusion protein and an adenovirus vaccine expressing the HIV-Gag protein had improved Gag-specific T cell responses, as compared to control mice. These responses are characterized by increased numbers of Gag-specific tetramer+ CD8+ T cells and increases in production of IFNγ, TNFα, and IL2, by Gag-specific CD8+ T cells. Moreover, our results revealed that use of the CRACC-Fc fusion protein enhances vaccine-elicited innate immune responses, as characterized by increased dendritic cells (DCs) maturation and IFNγ production from NK cells. This study highlights the importance of CRACC signaling during the induction of an immune response generally, and during vaccinations specifically, and also lends insight into the mechanisms underlying our prior results noting EAT-2-dependent improvements in vaccine efficacy. PMID:27151882

  12. The transcription factor E4BP4 is not required for extramedullary pathways of NK cell development.

    PubMed

    Crotta, Stefania; Gkioka, Annita; Male, Victoria; Duarte, João H; Davidson, Sophia; Nisoli, Ilaria; Brady, Hugh J M; Wack, Andreas

    2014-03-15

    NK cells contribute to antitumor and antiviral immunosurveillance. Their development in the bone marrow (BM) requires the transcription factor E4BP4/NFIL3, but requirements in other organs are less well defined. In this study, we show that CD3(-)NK1.1(+)NKp46(+)CD122(+) NK cells of immature phenotype and expressing low eomesodermin levels are found in thymus, spleen, and liver of E4BP4-deficient mice, whereas numbers of mature, eomesodermin(high) conventional NK cells are drastically reduced. E4BP4-deficient CD44(+)CD25(-) double-negative 1 thymocytes efficiently develop in vitro into NK cells with kinetics, phenotype, and functionality similar to wild-type controls, whereas no NK cells develop from E4BP4-deficient BM precursors. In E4BP4/Rag-1 double-deficient (DKO) mice, NK cells resembling those in Rag-1-deficient controls are found in similar numbers in the thymus and liver. However, NK precursors are reduced in DKO BM, and no NK cells develop from DKO BM progenitors in vitro. DKO thymocyte precursors readily develop into NK cells, but DKO BM transfers into nude recipients and NK cells in E4BP4/Rag-1/IL-7 triple-KO mice indicated thymus-independent NK cell development. In the presence of T cells or E4BP4-sufficient NK cells, DKO NK cells have a selective disadvantage, and thymic and hepatic DKO NK cells show reduced survival when adoptively transferred into lymphopenic hosts. This correlates with higher apoptosis rates and lower responsiveness to IL-15 in vitro. In conclusion, we demonstrate E4BP4-independent development of NK cells of immature phenotype, reduced fitness, short t1/2, and potential extramedullary origin. Our data identify E4BP4-independent NK cell developmental pathways and a role for E4BP4 in NK cell homeostasis. PMID:24534532

  13. Proapoptotic Bim regulates antigen-specific NK cell contraction and the generation of the memory NK cell pool after cytomegalovirus infection.

    PubMed

    Min-Oo, Gundula; Bezman, Natalie A; Madera, Sharline; Sun, Joseph C; Lanier, Lewis L

    2014-06-30

    Apoptosis is critical for the elimination of activated lymphocytes after viral infection. Proapoptotic factor Bim (Bcl2l11) controls T lymphocyte contraction and the formation of memory T cells after infection. Natural killer (NK) cells also undergo antigen-driven expansion to become long-lived memory cells after mouse cytomegalovirus (MCMV) infection; therefore, we examined the role of Bim in regulating the MCMV-driven memory NK cell pool. Despite responding similarly early after infection, Bcl2l11(-/-) Ly49H(+) NK cells show impaired contraction and significantly outnumber wild-type (WT) cells after the expansion phase. The inability to reduce the effector pool leads to a larger Bcl2l11(-/-) NK memory subset, which displays a less mature phenotype (CD11b(lo), CD27(+)) and lower levels of NK cell memory-associated markers KLRG1 and Ly6C. Bcl2l11(-/-) memory NK cells demonstrate a reduced response to m157-mediated stimulation and do not protect as effectively as WT memory NK cells in an MCMV challenge model. Thus, Bim-mediated apoptosis drives selective contraction of effector NK cells to generate a pool of mature, MCMV-specific memory cells. PMID:24958849

  14. MicroRNA transcriptomes of distinct human NK cell populations identify miR-362-5p as an essential regulator of NK cell function

    PubMed Central

    Ni, Fang; Guo, Chuang; Sun, Rui; Fu, Binqing; Yang, Yue; Wu, Lele; Ren, Sitong; Tian, Zhigang; Wei, Haiming

    2015-01-01

    Natural killer (NK) cells are critical effectors in the immune response against malignancy and infection, and microRNAs (miRNAs) play important roles in NK cell biology. Here we examined miRNA profiles of human NK cells from different cell compartments (peripheral blood, cord blood, and uterine deciduas) and of NKT and T cells from peripheral blood, and we identified a novel miRNA, miR-362-5p, that is highly expressed in human peripheral blood NK (pNK) cells. We also demonstrated that CYLD, a negative regulator of NF-κB signaling, was a target of miR-362-5p in NK cells. Furthermore, we showed that the over-expression of miR-362-5p enhanced the expression of IFN-γ, perforin, granzyme-B, and CD107a in human primary NK cells, and we found that silencing CYLD with a small interfering RNA (siRNA) mirrored the effect of miR-362-5p over-expression. In contrast, the inhibition of miR-362-5p had the opposite effect in NK cells, which was abrogated by CYLD siRNA, suggesting that miR-362-5p promotes NK-cell function, at least in part, by the down-regulation of CYLD. These results provide a resource for studying the roles of miRNAs in human NK cell biology and contribute to a better understanding of the physiologic significance of miRNAs in the regulation of NK cell function. PMID:25909817

  15. Influenza Vaccination Generates Cytokine-Induced Memory-like NK Cells: Impact of Human Cytomegalovirus Infection.

    PubMed

    Goodier, Martin R; Rodriguez-Galan, Ana; Lusa, Chiara; Nielsen, Carolyn M; Darboe, Alansana; Moldoveanu, Ana L; White, Matthew J; Behrens, Ron; Riley, Eleanor M

    2016-07-01

    Human NK cells are activated by cytokines, immune complexes, and signals transduced via activating ligands on other host cells. After vaccination, or during secondary infection, adaptive immune responses can enhance both cytokine-driven and Ab-dependent NK cell responses. However, induction of NK cells for enhanced function after in vitro exposure to innate inflammatory cytokines has also been reported and may synergize with adaptive signals to potentiate NK cell activity during infection or vaccination. To test this hypothesis, we examined the effect of seasonal influenza vaccination on NK cell function and phenotype in 52 previously unvaccinated individuals. Enhanced, IL-2-dependent, NK cell IFN-γ responses to Influenza A/California/7/2009 virus were detected up to 4 wk postvaccination and higher in human CMV (HCMV)-seronegative (HCMV(-)) individuals than in HCMV-seropositive (HCMV(+)) individuals. By comparison, robust NK cell degranulation responses were observed both before and after vaccination, due to high titers of naturally occurring anti-influenza Abs in human plasma, and did not differ between HCMV(+) and HCMV(-) subjects. In addition to these IL-2-dependent and Ab-dependent responses, NK cell responses to innate cytokines were also enhanced after influenza vaccination; this was associated with proliferation of CD57(-) NK cells and was most evident in HCMV(+) subjects. Similar enhancement of cytokine responsiveness was observed when NK cells were cocultured in vitro with Influenza A/California/7/2009 virus, and this was at least partially dependent upon IFN-αβR2. In summary, our data indicate that attenuated or live viral vaccines promote cytokine-induced memory-like NK cells and that this process is influenced by HCMV infection. PMID:27233958

  16. Influenza Vaccination Generates Cytokine-Induced Memory-like NK Cells: Impact of Human Cytomegalovirus Infection

    PubMed Central

    Goodier, Martin R.; Rodriguez-Galan, Ana; Lusa, Chiara; Nielsen, Carolyn M.; Darboe, Alansana; Moldoveanu, Ana L.; White, Matthew J.; Behrens, Ron

    2016-01-01

    Human NK cells are activated by cytokines, immune complexes, and signals transduced via activating ligands on other host cells. After vaccination, or during secondary infection, adaptive immune responses can enhance both cytokine-driven and Ab-dependent NK cell responses. However, induction of NK cells for enhanced function after in vitro exposure to innate inflammatory cytokines has also been reported and may synergize with adaptive signals to potentiate NK cell activity during infection or vaccination. To test this hypothesis, we examined the effect of seasonal influenza vaccination on NK cell function and phenotype in 52 previously unvaccinated individuals. Enhanced, IL-2–dependent, NK cell IFN-γ responses to Influenza A/California/7/2009 virus were detected up to 4 wk postvaccination and higher in human CMV (HCMV)-seronegative (HCMV−) individuals than in HCMV-seropositive (HCMV+) individuals. By comparison, robust NK cell degranulation responses were observed both before and after vaccination, due to high titers of naturally occurring anti-influenza Abs in human plasma, and did not differ between HCMV+ and HCMV− subjects. In addition to these IL-2–dependent and Ab-dependent responses, NK cell responses to innate cytokines were also enhanced after influenza vaccination; this was associated with proliferation of CD57− NK cells and was most evident in HCMV+ subjects. Similar enhancement of cytokine responsiveness was observed when NK cells were cocultured in vitro with Influenza A/California/7/2009 virus, and this was at least partially dependent upon IFN-αβR2. In summary, our data indicate that attenuated or live viral vaccines promote cytokine-induced memory-like NK cells and that this process is influenced by HCMV infection. PMID:27233958

  17. Differential requirement for Nfil3 during NK cell development.

    PubMed

    Seillet, Cyril; Huntington, Nicholas D; Gangatirkar, Pradnya; Axelsson, Elin; Minnich, Martina; Brady, Hugh J M; Busslinger, Meinrad; Smyth, Mark J; Belz, Gabrielle T; Carotta, Sebastian

    2014-03-15

    NK cells can be grouped into distinct subsets that are localized to different organs and exhibit a different capacity to secrete cytokines and mediate cytotoxicity. Despite these hallmarks that reflect tissue-specific specialization in NK cells, little is known about the factors that control the development of these distinct subsets. The basic leucine zipper transcription factor Nfil3 (E4bp4) is essential for bone marrow-derived NK cell development, but it is not clear whether Nfil3 is equally important for all NK cell subsets or how it induces NK lineage commitment. In this article, we show that Nfil3 is required for the formation of Eomes-expressing NK cells, including conventional medullary and thymic NK cells, whereas TRAIL(+) Eomes(-) NK cells develop independently of Nfil3. Loss of Nfil3 during the development of bone marrow-derived NK cells resulted in reduced expression of Eomes and, conversely, restoration of Eomes expression in Nfil3(-/-) progenitors rescued NK cell development and maturation. Collectively, these findings demonstrate that Nfil3 drives the formation of mature NK cells by inducing Eomes expression and reveal the differential requirements of NK cell subsets for Nfil3. PMID:24532575

  18. Phenotypic profile of expanded NK cells in chronic lymphoproliferative disorders: a surrogate marker for NK-cell clonality

    PubMed Central

    Bárcena, Paloma; Jara-Acevedo, María; Tabernero, María Dolores; López, Antonio; Sánchez, María Luz; García-Montero, Andrés C.; Muñoz-García, Noemí; Vidriales, María Belén; Paiva, Artur; Lecrevisse, Quentin; Lima, Margarida; Langerak, Anton W.; Böttcher, Sebastian; van Dongen, Jacques J.M.

    2015-01-01

    Currently, the lack of a universal and specific marker of clonality hampers the diagnosis and classification of chronic expansions of natural killer (NK) cells. Here we investigated the utility of flow cytometric detection of aberrant/altered NK-cell phenotypes as a surrogate marker for clonality, in the diagnostic work-up of chronic lymphoproliferative disorders of NK cells (CLPD-NK). For this purpose, a large panel of markers was evaluated by multiparametric flow cytometry on peripheral blood (PB) CD56low NK cells from 60 patients, including 23 subjects with predefined clonal (n = 9) and polyclonal (n = 14) CD56low NK-cell expansions, and 37 with CLPD-NK of undetermined clonality; also, PB samples from 10 healthy adults were included. Clonality was established using the human androgen receptor (HUMARA) assay. Clonal NK cells were found to show decreased expression of CD7, CD11b and CD38, and higher CD2, CD94 and HLADR levels vs. normal NK cells, together with a restricted repertoire of expression of the CD158a, CD158b and CD161 killer-associated receptors. In turn, NK cells from both clonal and polyclonal CLPD-NK showed similar/overlapping phenotypic profiles, except for high and more homogeneous expression of CD94 and HLADR, which was restricted to clonal CLPD-NK. We conclude that the CD94hi/HLADR+ phenotypic profile proved to be a useful surrogate marker for NK-cell clonality. PMID:26556869

  19. Genotypes of NK cell KIR Receptors, Their Ligands, and Fcγ Receptors in the Response of Neuroblastoma Patients to Hu14.18-IL2 Immunotherapy

    PubMed Central

    Delgado, DC; Hank, JA; Kolesar, J; Lorentzen, D; Gan, J; Seo, S; Kim, KM; Shusterman, S; Gillies, SD; Reisfeld, RA; Yang, R; Gadbaw, B; DeSantes, KD; London, WB; Seeger, RC; Maris, JM; Sondel, PM

    2010-01-01

    Response to immunocytokine (IC) therapy is dependent on natural killer (NK) cells in murine neuroblastoma (NBL) models. Furthermore, killer immunoglobulin-like receptor (KIR) / KIR-ligand mismatch is associated with improved outcome to autologous stem cell transplant for NBL. Additionally, clinical anti-tumor response to monoclonal antibodies has been associated with specific polymorphic-FcγR alleles. Relapsed/refractory NBL patients received the hu14.18-IL2 IC (humanized anti-GD2 monoclonal antibody linked to human IL2) in a Children’s Oncology Group (COG) Phase II trial. In this report, these patients were genotyped for KIR, HLA and FcR alleles to determine if KIR receptor-ligand mismatch or specific FcγR alleles were associated with anti-tumor response. Thirty-eight of 39 patients enrolled had DNA available for analysis; 24 were found to have autologous KIR/KIR -ligand mismatch; 14 were matched. Seven of 24 mismatched patients experienced either complete response or improvement of their disease after IC therapy. There was no response or comparable improvement of disease in patients who were matched. Thus KIR/KIR-ligand mismatch was associated with response/improvement to IC (p = 0.03). There was a trend towards patients with the FcγR2A 131-H/H genotype showing a higher response rate compared to other FcγR2A genotypes (p = 0.06). These analyses indicate that response or improvement of relapsed/refractory neuroblastoma patients after IC treatment is associated with autologous KIR/KIR-ligand mismatch, consistent with a role for NK cells in this clinical response. PMID:20935224

  20. The nature of the natural killer (NK) cell of human intestinal mucosa and mesenteric lymph node.

    PubMed Central

    Gibson, P R; Jewell, D P

    1985-01-01

    The relationship of the mononuclear cell (MNC) from human intestinal mucosa and mesenteric lymph node mediating anti-K-562 activity with that of peripheral blood has been assessed. Depletion of macrophages did not alter the measured cytotoxicity confirming that the effector cells were lymphocytes. Complement lysis of Leu 7 and Leu 11b coated cells reduced intestinal natural killer (NK) activity by a similar degree to that of peripheral blood but mesenteric lymph node NK activity was affected to a lesser extent. The response in NK activity of mucosal and nodal MNC to short incubation with lymphoblastoid interferon was similar to that for peripheral blood MNC. Twenty-four hours incubation of MNC with low concentrations of purified interleukin-2 (IL-2) consistently augmented intestinal and nodal NK activity but failed to augment that of peripheral blood MNC. No differences between the inhibitory effects of cAMP and prostaglandin E2 on NK activity from the three sites were seen. In addition, inhibition of cyclo-oxygenase activity with indomethacin had no effect on NK activity of intestinal and peripheral blood MNC while the lipoxygenase inhibitor, nordihydroguaiaretic acid, suppressed intestinal and peripheral blood NK activity similarly. In conclusion, anti-K-562 activity by intestinal MNC is mediated by NK cells with similar phenotypic and functional properties to those of peripheral blood. However, the increased sensitivity of mucosal NK cells to IL-2 suggests that higher proportions of NK cell precursors may be present in intestinal MNC populations. PMID:2412737

  1. Dysregulation of regulatory CD56(bright) NK cells/T cells interactions in multiple sclerosis.

    PubMed

    Laroni, Alice; Armentani, Eric; Kerlero de Rosbo, Nicole; Ivaldi, Federico; Marcenaro, Emanuela; Sivori, Simona; Gandhi, Roopali; Weiner, Howard L; Moretta, Alessandro; Mancardi, Giovanni L; Uccelli, Antonio

    2016-08-01

    Recent evidence has shown that CD56(bright) NK cells, a subset of NK cells abundant in lymph nodes, may have an immunoregulatory function. In multiple sclerosis (MS), expansion of CD56(bright) NK cells has been associated to successful response to different treatments and to remission of disease during pregnancy; how whether they exert immunoregulation in physiologic conditions and whether this is impaired in MS is not known. We dissected the immunoregulatory role of CD56(bright) NK cells function in healthy subjects (HS) and compared it with that of untreated MS subjects or patients with clinically isolated syndrome suggestive of MS (CIS). We found that CD56(bright) NK cells from HS acquire, upon inflammatory cues, the capability of suppressing autologous CD4+T cell proliferation through direct cytotoxicity requiring engagement of natural cytotoxicity receptors (NCRs) and secretion of granzyme B. CD56(bright) NK cells from patients with MS/CIS did not differ in frequency and share a similar phenotype but displayed a significantly lower ability to inhibit autologous T cell proliferation. This impairment was not related to deficient expression of NCRs or granzyme B by CD56(bright) NK cells, but to increased HLA-E expression on T cells from MS/CIS subjects, which could enhance the inhibitory effect mediated by NKG2A that is homogeneously expressed on CD56(bright) NK cells. The defect in controlling autologous T cells by CD56(bright) NK cells in MS/CIS might contribute to the excess of autoimmune response that is associated to disease development. PMID:27157273

  2. Role of alloreactive KIR2DS1(+) NK cells in haploidentical hematopoietic stem cell transplantation.

    PubMed

    Marcenaro, Emanuela; Carlomagno, Simona; Pesce, Silvia; Della Chiesa, Mariella; Moretta, Alessandro; Sivori, Simona

    2011-10-01

    In allo-HSCT, donor-derived, "alloreactive" NK cells have been shown to play a crucial role in the treatment of acute leukemia, contributing to eradication of leukemic blasts (GvL effect) and to clearance of residual recipient DCs and T lymphocytes (thus, preventing GvHD and graft rejection, respectively). Such alloreactive NK cells do not express CD94/NKG2A but express inhibitory KIRs, specific for HLA class I allotypes, present in the donor but lacking in the recipient. This review is focused on the role of the activating KIR2DS1 receptor (specific for the C2-epitope of HLA-C) in haplo-HSCT. Recent data indicate that KIR2DS1 expression in HSC donors may represent a remarkable advantage in alloreactive NK responses. This is a result of a substantial increase in the NK-mediated capability to kill, not only recipients' leukemic cells but also DCs and T cell blasts. The beneficial effects mediated by alloreactive KIR2DS1(+) NK cells may occur after de novo expression of CCR7 upon interaction with allogeneic, KIR ligand-mismatched CCR7(+) cells. As a consequence, they can be redirected to LNs, where they can prevent priming of donor T cells and induction of GvHD. Finally, KIR2DS1 expression may also significantly amplify the size of the alloreactive NK cell subset by switching a subset of "not alloreactive" NK cells into potent alloreactive cells. PMID:21791599

  3. Repression of GSK3 restores NK cell cytotoxicity in AML patients

    PubMed Central

    Parameswaran, Reshmi; Ramakrishnan, Parameswaran; Moreton, Stephen A.; Xia, Zhiqiang; Hou, Yongchun; Lee, Dean A.; Gupta, Kalpana; deLima, Marcos; Beck, Rose C.; Wald, David N.

    2016-01-01

    Natural killer cells from acute myeloid leukaemia patients (AML-NK) show a dramatic impairment in cytotoxic activity. The exact reasons for this dysfunction are not fully understood. Here we show that the glycogen synthase kinase beta (GSK3β) expression is elevated in AML-NK cells. Interestingly, GSK3 overexpression in normal NK cells impairs their ability to kill AML cells, while genetic or pharmacological GSK3 inactivation enhances their cytotoxic activity. Mechanistic studies reveal that the increased cytotoxic activity correlates with an increase in AML-NK cell conjugates. GSK3 inhibition promotes the conjugate formation by upregulating LFA expression on NK cells and by inducing ICAM-1 expression on AML cells. The latter is mediated by increased NF-κB activation in response to TNF-α production by NK cells. Finally, GSK3-inhibited NK cells show significant efficacy in human AML mouse models. Overall, our work provides mechanistic insights into the AML-NK dysfunction and a potential NK cell therapy strategy. PMID:27040177

  4. Formaldehyde exposure impairs the function and differentiation of NK cells.

    PubMed

    Kim, Eun-Mi; Lee, Hwa-Youn; Lee, Eun-Hee; Lee, Ki-Mo; Park, Min; Ji, Kon-Young; Jang, Ji-Hun; Jeong, Yun-Hwa; Lee, Kwang-Ho; Yoon, Il-Joo; Kim, Su-Man; Jeong, Moon-Jin; Kim, Kwang Dong; Kang, Hyung-Sik

    2013-11-25

    We investigated the cytotoxic effects of formaldehyde (FA) on lymphocytes. FA-exposed mice showed a profound reduction not only in the number of natural killer (NK) cells but also in the expression of NK cell-specific receptors, but these mice did not exhibit decreases in the numbers of T or B lymphocytes. FA exposure also induced decreases in NK cytolytic activity and in the expression of NK cell-associated genes, such as IFN-γ, perforin and CD122. To determine the effect of FA on tumorigenicity, C57BL/6 mice were subcutaneously injected with B16F10 melanoma cells after FA exposure. The mass of the B16F10 tumor and the concentration of extravascular polymorphonuclear leukocytes were greater than those in unexposed tumor-bearing control mice. The number and cytolytic activity of NK cells were also reduced in B16F10 tumor-bearing mice exposed to FA. To determine how FA reduces the NK cell number, NK precursor (pNK) cells were treated with FA, and the differentiation status of the NK cells was analyzed. NK cell differentiation was impaired by FA treatment in a concentration-dependent manner. These findings indicate that FA exposure may promote tumor progression by impairing NK cell function and differentiation. PMID:24060340

  5. Conversion of peripheral blood NK cells to a decidual NK-like phenotype by a cocktail of defined factors

    PubMed Central

    Cerdeira, Ana Sofia; Rajakumar, Augustine; Royle, Caroline M.; Lo, Agnes; Husain, Zaheed; Thadhani, Ravi I.; Sukhatme, Vikas P.; Karumanchi, S. Ananth; Kopcow, Hernan D.

    2013-01-01

    Natural killer (NK) cells that populate the decidua are important regulators of normal placentation. In contrast to peripheral blood NK (pNK) cells, decidual NK cells (dNK) lack cytotoxicity, secrete pro-angiogenic factors and regulate trophoblast invasion. Here we show that exposure to a combination of hypoxia, transforming growth factor beta 1, and a demethylating agent, results in NK cells that express Killer cell Immunoglobulin like Receptors, the dNK cell markers CD9 and CD49a, and dNK pattern of chemokine receptors. These cells secrete vascular endothelial growth factor, a potent pro-angiogenic molecule, display reduced cytotoxicity and promote invasion of human trophoblast cell lines. These findings have potential therapeutic applications for placental disorders associated with altered NK cell biology. PMID:23487420

  6. FoxO1-mediated autophagy is required for NK cell development and innate immunity.

    PubMed

    Wang, Shuo; Xia, Pengyan; Huang, Guanling; Zhu, Pingping; Liu, Jing; Ye, Buqing; Du, Ying; Fan, Zusen

    2016-01-01

    Natural killer (NK) cells exert a crucial role in early immune responses as a major innate effector component. However, the underlying mechanisms of NK cell development remain largely elusive. Here we show that robust autophagy appears in the stage of immature NK cells (iNKs), which is required for NK cell development. Autophagy defects result in damaged mitochondria and accumulation of reactive oxygen species (ROS) that leads to apoptosis of NK cells. Autophagy protects NK cell viability during development through removal of damaged mitochondria and intracellular ROS. Phosphorylated Forkhead box O (FoxO)1 is located to the cytoplasm of iNKs and interacts with Atg7, leading to induction of autophagy. FoxO1 deficiency or an inactive FoxO1(AAA) mutant abrogates autophagy initiation in iNKs and impairs NK cell development and viral clearance. Therefore we conclude that FoxO1-mediated autophagy is required for NK cell development and NK cell-induced innate immunity. PMID:27010363

  7. FoxO1-mediated autophagy is required for NK cell development and innate immunity

    PubMed Central

    Wang, Shuo; Xia, Pengyan; Huang, Guanling; Zhu, Pingping; Liu, Jing; Ye, Buqing; Du, Ying; Fan, Zusen

    2016-01-01

    Natural killer (NK) cells exert a crucial role in early immune responses as a major innate effector component. However, the underlying mechanisms of NK cell development remain largely elusive. Here we show that robust autophagy appears in the stage of immature NK cells (iNKs), which is required for NK cell development. Autophagy defects result in damaged mitochondria and accumulation of reactive oxygen species (ROS) that leads to apoptosis of NK cells. Autophagy protects NK cell viability during development through removal of damaged mitochondria and intracellular ROS. Phosphorylated Forkhead box O (FoxO)1 is located to the cytoplasm of iNKs and interacts with Atg7, leading to induction of autophagy. FoxO1 deficiency or an inactive FoxO1AAA mutant abrogates autophagy initiation in iNKs and impairs NK cell development and viral clearance. Therefore we conclude that FoxO1-mediated autophagy is required for NK cell development and NK cell-induced innate immunity. PMID:27010363

  8. NK Cell Phenotypic Modulation in Lung Cancer Environment

    PubMed Central

    Hao, Jun-Wei; Li, Yang; Liu, Bin; Yu, Yan; Shi, Fu-Dong; Zhou, Qing-Hua

    2014-01-01

    Background Nature killer (NK) cells play an important role in anti-tumor immunotherapy. But it indicated that tumor cells impacted possibly on NK cell normal functions through some molecules mechanisms in tumor microenvironment. Materials and methods Our study analyzed the change about NK cells surface markers (NK cells receptors) through immunofluorescence, flow cytometry and real-time PCR, the killed function from mouse spleen NK cell and human high/low lung cancer cell line by co-culture. Furthermore we certificated the above result on the lung cancer model of SCID mouse. Results We showed that the infiltration of NK cells in tumor periphery was related with lung cancer patients' prognosis. And the number of NK cell infiltrating in lung cancer tissue is closely related to the pathological types, size of the primary cancer, smoking history and prognosis of the patients with lung cancer. The expression of NK cells inhibitor receptors increased remarkably in tumor micro-environment, in opposite, the expression of NK cells activated receptors decrease magnificently. Conclusions The survival time of lung cancer patient was positively related to NK cell infiltration degree in lung cancer. Thus, the down-regulation of NKG2D, Ly49I and the up-regulation of NKG2A may indicate immune tolerance mechanism and facilitate metastasis in tumor environment. Our research will offer more theory for clinical strategy about tumor immunotherapy. PMID:25299645

  9. Teach Your NK Cells Well.

    PubMed

    Cooper, Megan A

    2016-08-16

    Natural killer cells readily kill target cells, and education ensures tolerance to self. In this issue of Immunity, Boudreau et al. (2016) and Chen et al. (2016) report new mechanisms of human and mouse natural killer cell education by inhibitory and activating receptors. PMID:27533007

  10. Coordinated expression of DNAM-1 and LFA-1 in educated NK cells.

    PubMed

    Enqvist, Monika; Ask, Eivind Heggernes; Forslund, Elin; Carlsten, Mattias; Abrahamsen, Greger; Béziat, Vivien; Andersson, Sandra; Schaffer, Marie; Spurkland, Anne; Bryceson, Yenan; Önfelt, Björn; Malmberg, Karl-Johan

    2015-05-01

    The functional capacity of NK cells is dynamically tuned by integrated signals from inhibitory and activating cell surface receptors in a process termed NK cell education. However, the understanding of the cellular and molecular mechanisms behind this functional tuning is limited. In this study, we show that the expression of the adhesion molecule and activation receptor DNAX accessory molecule 1 (DNAM-1) correlates with the quantity and quality of the inhibitory input by HLA class I-specific killer cell Ig-like receptors and CD94/NKG2A as well as with the magnitude of functional responses. Upon target cell recognition, the conformational state of LFA-1 changed in educated NK cells, associated with rapid colocalization of both active LFA-1 and DNAM-1 at the immune synapse. Thus, the coordinated expression of LFA-1 and DNAM-1 is a central component of NK cell education and provides a potential mechanism for controlling cytotoxicity by functionally mature NK cells. PMID:25825444

  11. Intrinsic Contribution of Perforin to NK-Cell Homeostasis during Mouse Cytomegalovirus Infection

    PubMed Central

    Arapović, Maja; Brizić, Ilija; Popović, Branka; Jurković, Slaven; Jordan, Stefan; Krmpotić, Astrid; Arapović, Jurica; Jonjić, Stipan

    2016-01-01

    In addition to their role as effector cells in virus control, natural killer (NK) cells have an immunoregulatory function in shaping the antiviral T-cell response. This function is further pronounced in perforin-deficient mice that show the enhanced NK-cell proliferation and cytokine secretion upon mouse cytomegalovirus (MCMV) infection. Here, we confirmed that stronger activation and maturation of NK cells in perforin-deficient mice correlates with higher MCMV load. To further characterize the immunoregulatory potential of perforin, we compared the response of NK cells that express or do not express perforin using bone-marrow chimeras. Our results demonstrated that the enhanced proliferation and maturation of NK cells in MCMV-infected bone-marrow chimeras is an intrinsic property of perforin-deficient NK cells. Thus, in addition to confirming that NK-cell proliferation is virus load dependent, our data extend this notion demonstrating that perforin plays an intrinsic role as a feedback mechanism in the regulation of NK-cell proliferation during viral infections. PMID:27092144

  12. miR-150 regulates the development of NK and iNKT cells

    PubMed Central

    Bezman, Natalie A.; Chakraborty, Tirtha; Bender, Timothy

    2011-01-01

    Natural killer (NK) and invariant NK T (iNKT) cells are critical in host defense against pathogens and for the initiation of adaptive immune responses. miRNAs play important roles in NK and iNKT cell development, maturation, and function, but the roles of specific miRNAs are unclear. We show that modulation of miR-150 expression levels has a differential effect on NK and iNKT cell development. Mice with a targeted deletion of miR-150 have an impaired, cell lineage–intrinsic defect in their ability to generate mature NK cells. Conversely, a gain-of-function miR-150 transgene promotes the development of NK cells, which display a more mature phenotype and are more responsive to activation. In contrast, overexpression of miR-150 results in a substantial reduction of iNKT cells in the thymus and in the peripheral lymphoid organs. The transcription factor c-Myb has been shown to be a direct target of miR-150. Our finding of increased NK cell and decreased iNKT cell frequencies in Myb heterozygous bone marrow chimeras suggests that miR-150 differentially controls the development of NK and iNKT cell lineages by targeting c-Myb. PMID:22124110

  13. Intrinsic Contribution of Perforin to NK-Cell Homeostasis during Mouse Cytomegalovirus Infection.

    PubMed

    Arapović, Maja; Brizić, Ilija; Popović, Branka; Jurković, Slaven; Jordan, Stefan; Krmpotić, Astrid; Arapović, Jurica; Jonjić, Stipan

    2016-01-01

    In addition to their role as effector cells in virus control, natural killer (NK) cells have an immunoregulatory function in shaping the antiviral T-cell response. This function is further pronounced in perforin-deficient mice that show the enhanced NK-cell proliferation and cytokine secretion upon mouse cytomegalovirus (MCMV) infection. Here, we confirmed that stronger activation and maturation of NK cells in perforin-deficient mice correlates with higher MCMV load. To further characterize the immunoregulatory potential of perforin, we compared the response of NK cells that express or do not express perforin using bone-marrow chimeras. Our results demonstrated that the enhanced proliferation and maturation of NK cells in MCMV-infected bone-marrow chimeras is an intrinsic property of perforin-deficient NK cells. Thus, in addition to confirming that NK-cell proliferation is virus load dependent, our data extend this notion demonstrating that perforin plays an intrinsic role as a feedback mechanism in the regulation of NK-cell proliferation during viral infections. PMID:27092144

  14. Suppression of NK cell-mediated cytotoxicity against PRRSV-infected porcine alveolar macrophages in vitro.

    PubMed

    Cao, Jun; Grauwet, Korneel; Vermeulen, Ben; Devriendt, Bert; Jiang, Ping; Favoreel, Herman; Nauwynck, Hans

    2013-06-28

    The adaptive immunity against PRRSV has already been studied in depth, but only limited data are available on the innate immune responses against this pathogen. In the present study, we analyzed the interaction between porcine natural killer (NK) cells and PRRSV-infected primary porcine alveolar macrophages (PAMs), since NK cells are one of the most important components of innate immunity and PAMs are primary target cells of PRRSV infection. NK cytotoxicity assays were performed using enriched NK cells as effector cells and virus-infected or mock-inoculated PAMs as target cells. The NK cytotoxicity against PRRSV-infected PAMs was decreased starting from 6h post inoculation (hpi) till the end of the experiment (12 hpi) and was significantly lower than that against pseudorabies virus (PrV)-infected PAMs. UV-inactivated PRRSV also suppressed NK activity, but much less than infectious PRRSV. Furthermore, co-incubation with PRRSV-infected PAMs inhibited degranulation of NK cells. Finally, using the supernatant of PRRSV-infected PAMs collected at 12 hpi showed that the suppressive effect of PRRSV on NK cytotoxicity was not mediated by soluble factors. In conclusion, PRRSV-infected PAMs showed a reduced susceptibility toward NK cytotoxicity, which may represent one of the multiple evasion strategies of PRRSV. PMID:23522639

  15. Ly49C Impairs NK Cell Memory in Mouse Cytomegalovirus Infection.

    PubMed

    Forbes, Catherine A; Scalzo, Anthony A; Degli-Esposti, Mariapia A; Coudert, Jerome D

    2016-07-01

    NK cells possess inhibitory receptors that are responsible for self-MHC class I recognition; beyond their inhibitory function, accumulating evidence indicates that such receptors confer NK cell functional competence through an unclear process termed "licensing." Ly49C is the main self-specific inhibitory Ly49 receptor in H-2(b) C57BL/6 (B6) mice. We used B6 Ly49C-transgenic and B6 β2 microglobulin (β2m)-knockout Ly49C-transgenic mice to investigate the impact of licensing through this inhibitory receptor in precursor and mature NK cells. We found that self-specific inhibitory receptors affected NK cell precursor survival and proliferation at particular developmental stages in an MHC class I-dependent manner. The presence of Ly49C impacted the NK cell repertoire in a β2m-dependent manner, with reduced Ly49A(+), Ly49G2(+), and Ly49D(+) subsets, an increased DNAM-1(+) subset, and higher NKG2D expression. Licensed NK cells displayed a skewed distribution of the maturation stages, which was characterized by differential CD27 and CD11b expression, toward the mature phenotypes. We found that Ly49C-mediated licensing induced a split effect on NK cell functions, with increased cytokine-production capabilities following engagement of various activating receptors while cytotoxicity remained unchanged. Analysis of licensed NK cell functions in vivo, in a system of mouse CMV infection, indicated that licensing did not play a major role in the NK cell antiviral response during acute infection, but it strongly impaired the generation and/or persistence of memory NK cells. This study unravels multifaceted effects of licensing on NK cell populations and their functions. PMID:27233959

  16. Cutting edge: stage-specific requirement of IL-18 for antiviral NK cell expansion.

    PubMed

    Madera, Sharline; Sun, Joseph C

    2015-02-15

    Although NK cells are considered part of the innate immune system, recent studies have demonstrated the ability of Ag-experienced NK cells to become long-lived and contribute to potent recall responses similar to T and B cells. The precise signals that promote the generation of a long-lived NK cell response are largely undefined. In this article, we demonstrate that NK cells require IL-18 signaling to generate a robust primary response during mouse CMV (MCMV) infection but do not require this signal for memory cell maintenance or recall responses. IL-12 signaling and STAT4 in activated NK cells increased the expression of the adaptor protein MyD88, which mediates signaling downstream of the IL-18 and IL-1 receptors. During MCMV infection, NK cells required MyD88, but not IL-1R, for optimal expansion. Thus, an IL-18-MyD88 signaling axis facilitates the prolific expansion of NK cells in response to primary viral infection, but not recall responses. PMID:25589075

  17. Stage-specific requirement of IL-18 for antiviral NK cell expansion

    PubMed Central

    Madera, Sharline; Sun, Joseph C.

    2014-01-01

    Although natural killer (NK) cells are considered part of the innate immune system, recent studies have demonstrated the ability of antigen-experienced NK cells to become long-lived and contribute to potent recall responses similar to T and B cells. The precise signals that promote the generation of a long-lived NK cell response are largely undefined. Here, we demonstrate that NK cells require interleukin (IL)-18 signaling to generate a robust primary response during mouse cytomegalovirus (MCMV) infection, but do not require this signal for memory cell maintenance or recall responses. IL-12 signaling and STAT4 in activated NK cells increased the expression of the adaptor protein MyD88, which mediates signaling downstream of the IL-18 and IL-1 receptors. During MCMV infection, NK cells required MyD88 but not IL-1 receptor for optimal expansion. Thus, an IL-18-MyD88 signaling axis facilitates the prolific expansion of NK cells in response to primary viral infection, but not recall responses. PMID:25589075

  18. NK cells expressing inhibitory KIR for non-self-ligands remain tolerant in HLA-matched sibling stem cell transplantation.

    PubMed

    Björklund, Andreas T; Schaffer, Marie; Fauriat, Cyril; Ringdén, Olle; Remberger, Mats; Hammarstedt, Christina; Barrett, A John; Ljungman, Per; Ljunggren, Hans-Gustaf; Malmberg, Karl-Johan

    2010-04-01

    Natural killer (NK)-cell alloreactivity in recipients of hematopoietic stem cell grafts from HLA-identical siblings is intriguing and has suggested breaking of NK-cell tolerance during the posttransplantation period. To examine this possibility, we analyzed clinical outcomes in a cohort of 105 patients with myeloid malignancies who received T cell-replete grafts from HLA-matched sibling donors. Presence of inhibitory killer cell immunoglobulin-like receptors (KIRs) for nonself HLA class I ligands had no effect on disease-free survival, incidence of relapse, or graft-versus-host disease. A longitudinal analysis of the NK-cell repertoire and function revealed a global hyporesponsiveness of NK cells early after transplantation. Functional responses recovered at approximately 6 months after transplantation. Importantly, NKG2A(-) NK cells expressing KIRs for nonself HLA class I ligands remained tolerant at all time points. Furthermore, a direct comparison of NK-cell reconstitution in T cell-replete and T cell-depleted HLA-matched sibling stem cell transplantation (SCT) revealed that NKG2A(+) NK cells dominated the functional repertoire early after transplantation, with intact tolerance of NKG2A(-) NK cells expressing KIRs for nonself ligands in both settings. Our results provide evidence against the emergence of alloreactive NK cells in HLA-identical allogeneic SCT. PMID:20097883

  19. NK Cell-Mediated Antibody-Dependent Cellular Cytotoxicity in Cancer Immunotherapy

    PubMed Central

    Wang, Wei; Erbe, Amy K.; Hank, Jacquelyn A.; Morris, Zachary S.; Sondel, Paul M.

    2015-01-01

    Natural killer (NK) cells play a major role in cancer immunotherapies that involve tumor-antigen targeting by monoclonal antibodies (mAbs). NK cells express a variety of activating and inhibitory receptors that serve to regulate the function and activity of the cells. In the context of targeting cells, NK cells can be “specifically activated” through certain Fc receptors that are expressed on their cell surface. NK cells can express FcγRIIIA and/or FcγRIIC, which can bind to the Fc portion of immunoglobulins, transmitting activating signals within NK cells. Once activated through Fc receptors by antibodies bound to target cells, NK cells are able to lyse target cells without priming, and secrete cytokines like interferon gamma to recruit adaptive immune cells. This antibody-dependent cell-mediated cytotoxicity (ADCC) of tumor cells is utilized in the treatment of various cancers overexpressing unique antigens, such as neuroblastoma, breast cancer, B cell lymphoma, and others. NK cells also express a family of receptors called killer immunoglobulin-like receptors (KIRs), which regulate the function and response of NK cells toward target cells through their interaction with their cognate ligands that are expressed on tumor cells. Genetic polymorphisms in KIR and KIR-ligands, as well as FcγRs may influence NK cell responsiveness in conjunction with mAb immunotherapies. This review focuses on current therapeutic mAbs, different strategies to augment the anti-tumor efficacy of ADCC, and genotypic factors that may influence patient responses to antibody-dependent immunotherapies. PMID:26284063

  20. Altered effector functions of NK cells in chronic hepatitis C are associated with IFNL3 polymorphism.

    PubMed

    Rogalska-Taranta, Magdalena; Markova, Antoaneta A; Taranta, Andrzej; Lunemann, Sebastian; Schlaphoff, Verena; Flisiak, Robert; Manns, Michael P; Cornberg, Markus; Kraft, Anke R M; Wedemeyer, Heiner

    2015-08-01

    Interferon α-mediated effector functions of NK cells may contribute to the control of HCV replication and the pathogenesis of liver disease. The single-nucleotide polymorphism rs12979860 near IFNL3 (previously known as IL28B) is important in response to IFN-α treatment and in spontaneous resolution of acute hepatitis C. The role of the IFNL3 polymorphism in NK cell function is unclear. Thus, we investigated the role of IFNL3 polymorphism in type I IFN-dependent regulation of NK cell functions in patients with cHC and healthy control subjects. We demonstrated a marked polarization of NK cells toward cytotoxicity in response to IFN-α stimulation in patients with hepatitis C. That TRAIL up-regulation was present, particularly in patients with the IFNL3-TT allele, was supported by a shift in the pSTAT-1:pSTAT-4 ratios toward pSTAT-1. In patients bearing the IFNL3-TT allele, NK cell effector function correlated with liver disease activity. In contrast, higher cytokine production of NK cells was observed in healthy individuals with the IFNL3-CC genotype, which may support spontaneous HCV clearance in acute infection. Overall, these findings show that the role of NK cells may differ in chronic infection vs. early antiviral defense and that the IFNL3 genotype differentially influences NK cell function. PMID:26034208

  1. NK Cell-based Immunotherapies in Pediatric Oncology

    PubMed Central

    McDowell, Kimberly A.; Hank, Jacquelyn A.; DeSantes, Kenneth B.; Capitini, Christian M.; Otto, Mario; Sondel, Paul M.

    2015-01-01

    The past decade has seen several anti-cancer immunotherapeutic strategies transition from “promising preclinical models” to treatments with proven clinical activity or benefit. In 2013, the journal Science selected the field of Cancer Immunotherapy as the overall number-1 breakthrough for the year in all of scientific research. In the setting of cancer immunotherapy for adult malignancies, many of these immunotherapy strategies have relied on the cancer patient’s endogenous anti-tumor T cell response. While much promising research in pediatric oncology is similarly focused on T cell reactivity, several pediatric malignancies themselves, or the chemo-radiotherapy used to achieve initial responses, can be associated with profound immune suppression, particularly of the T cell system. A separate component of the immune system, also able to mediate anti-tumor effects and less suppressed by conventional cancer treatment, is the NK cell system. In recent years, several distinct immunotherapeutic approaches that rely on the activity of NK cells have moved from preclinical development into clinical testing, and some have shown clear antitumor benefit. This review provides an overview of NK cell-based immunotherapy efforts that are directed towards childhood malignancies, with an emphasis on protocols that are already in clinical testing. PMID:25590232

  2. NK Cells Respond to Haptens by the Activation of Calcium Permeable Plasma Membrane Channels

    PubMed Central

    Grandclément, Camille; Pick, Horst; Vogel, Horst; Held, Werner

    2016-01-01

    Natural Killer (NK) cells mediate innate immunity to infected and transformed cells. Yet, NK cells can also mount hapten-specific recall responses thereby contributing to contact hypersensitivity (CHS). However, since NK cells lack antigen receptors that are used by the adaptive immune system to recognize haptens, it is not clear if NK cells respond directly to haptens and, if so, what mediates these responses. Here we show that among four haptens the two that are known to induce NK cell-dependent CHS trigger the rapid influx of extracellular Ca2+ into NK cells and lymphocyte cell lines. Thus lymphocytes can respond to haptens independent of antigen presentation and antigen receptors. We identify the Ca2+-permeable cation channel TRPC3 as a component of the lymphocyte response to one of these haptens. These data suggest that the response to the second hapten is based on a distinct mechanism, consistent with the capacity of NK cells to discriminate haptens. These findings raise the possibility that antigen-receptor independent activation of immune cells contributes to CHS. PMID:26963818

  3. Folate-conjugated immunoglobulin targets melanoma tumor cells for NK cell effector functions

    PubMed Central

    Skinner, Cassandra C.; McMichael, Elizabeth L.; Jaime-Ramirez, Alena C.; Abrams, Zachary B.; Lee, Robert J.; Carson, William E.

    2016-01-01

    The folate receptor (FR) is over-expressed on the vascular side of cancerous cells including those of the breast, ovaries, testes, and cervix. We hypothesized that a folate-conjugated immunoglobulin (F-IgG) would bind to the FR that is over-expressed on melanoma tumor cells to target these cells for lysis by natural killer (NK) cells. Folate receptor expression was confirmed in the Mel-39 (human melanoma) cell line by flow cytometry and immunoblot analysis, using KB (human oral epithelial) and F01 (human melanoma) as a positive and negative control, respectively. FR-positive and negative cell lines were treated with F-IgG or control immunoglobulin G (C-IgG) in the presence or absence of cytokines in order to determine NK cell ability to lyse FR-positive cell lines. NK cell activation was significantly upregulated and lysis of Mel 39 tumor cells enhanced following treatment with F-IgG, as compared to C-IgG at all effector:target (E:T) ratios (p<0.01). This trend was further enhanced by NK cell stimulation with the activating cytokine interleukin-12 (IL-12). NK cell production of cytokines such as interferon-gamma (IFN-γ), macrophage inflammatory protein 1 alpha (MIP-1α), and regulated on activation normal T-cell expressed and secreted (RANTES) were also significantly increased in response to co-stimulation with IL-12 stimulation and F-IgG-coated Mel 39 target cells, as compared to controls (p<0.01). In contrast, F-IgG did not bind to the FR-negative cell line F01 and had no significant effect on NK cell lysis or cytokine production. This research indicates the potential use of F-IgG for its ability to induce an immune response from NK cells against FR-positive melanoma tumor cells which can be further enhanced by the addition of cytokines. PMID:27035691

  4. Folate-conjugated immunoglobulin targets melanoma tumor cells for NK cell effector functions.

    PubMed

    Skinner, Cassandra C; McMichael, Elizabeth L; Jaime-Ramirez, Alena C; Abrams, Zachary B; Lee, Robert J; Carson, William E

    2016-08-01

    The folate receptor (FR) is overexpressed on the vascular side of cancerous cells including those of the breast, ovaries, testes, and cervix. We hypothesized that a folate-conjugated immunoglobulin (F-IgG) would bind to the FR that is overexpressed on melanoma tumor cells to target these cells for lysis by natural killer (NK) cells. Folate receptor expression was confirmed in the Mel-39 (human melanoma) cell line by flow cytometry and immunoblot analysis using KB (human oral epithelial) and F01 (human melanoma) as a positive and a negative control, respectively. FR-positive and FR-negative cell lines were treated with F-IgG or control immunoglobulin G in the presence or absence of cytokines to determine NK cell ability to lyse FR-positive cell lines. NK cell activation was significantly upregulated and lysis of Mel 39 tumor cells increased following treatment with F-IgG compared with control immunoglobulin G at all effector : target (E : T) ratios (P<0.01). This trend further increased by NK cell stimulation with the activating cytokine interleukin-12. NK cell production of cytokines such as interferon-gamma, macrophage inflammatory protein 1α, and regulated on activation normal T-cell expressed and secreted (RANTES) was also significantly increased in response to costimulation with interleukin-12 stimulation and F-IgG-coated Mel 39 target cells compared with controls (P<0.01). In contrast, F-IgG did not bind to the FR-negative cell line F01 and had no significant effect on NK cell lysis or cytokine production. This research indicates the potential use of F-IgG for its ability to induce an immune response from NK cells against FR-positive melanoma tumor cells, which can be further increased by the addition of cytokines. PMID:27035691

  5. Influenza Virus Targets Class I MHC-Educated NK Cells for Immunoevasion

    PubMed Central

    Mahmoud, Ahmad Bakur; Tu, Megan M.; Wight, Andrew; Zein, Haggag S.; Rahim, Mir Munir A.; Lee, Seung-Hwan; Sekhon, Harman S.; Brown, Earl G.; Makrigiannis, Andrew P.

    2016-01-01

    The immune response to influenza virus infection comprises both innate and adaptive defenses. NK cells play an early role in the destruction of tumors and virally-infected cells. NK cells express a variety of inhibitory receptors, including those of the Ly49 family, which are functional homologs of human killer-cell immunoglobulin-like receptors (KIR). Like human KIR, Ly49 receptors inhibit NK cell-mediated lysis by binding to major histocompatibility complex class I (MHC-I) molecules that are expressed on normal cells. During NK cell maturation, the interaction of NK cell inhibitory Ly49 receptors with their MHC-I ligands results in two types of NK cells: licensed (“functional”), or unlicensed (“hypofunctional”). Despite being completely dysfunctional with regard to rejecting MHC-I-deficient cells, unlicensed NK cells represent up to half of the mature NK cell pool in rodents and humans, suggesting an alternative role for these cells in host defense. Here, we demonstrate that after influenza infection, MHC-I expression on lung epithelial cells is upregulated, and mice bearing unlicensed NK cells (Ly49-deficient NKCKD and MHC-I-deficient B2m-/- mice) survive the infection better than WT mice. Importantly, transgenic expression of an inhibitory self-MHC-I-specific Ly49 receptor in NKCKD mice restores WT influenza susceptibility, confirming a direct role for Ly49. Conversely, F(ab’)2-mediated blockade of self-MHC-I-specific Ly49 inhibitory receptors protects WT mice from influenza virus infection. Mechanistically, perforin-deficient NKCKD mice succumb to influenza infection rapidly, indicating that direct cytotoxicity is necessary for unlicensed NK cell-mediated protection. Our findings demonstrate that Ly49:MHC-I interactions play a critical role in influenza virus pathogenesis. We suggest a similar role may be conserved in human KIR, and their blockade may be protective in humans. PMID:26928844

  6. Recombinant LPG3 Stimulates IFN-Γ and TNF-A Secretion by Human NK Cells

    PubMed Central

    RASOLZADEH, Sanaz; HAJI FATAHALIHA, Mostafa; HOSSEINI, Maryam; JAFARI, Reza; MIAHIPOUR, Abolfazl; MOVASSAGHPOUR, Ali Akbar; BABALO, Zohreh; RAFATI, Sima; YOUSEFI, Mehdi

    2015-01-01

    Background: Natural killer (NK) cells play an important role in early stages of innate immune responses against viral and tumoral attacks. Activation of NK cells by leishmaniasis results in secretion of cytokines such as interferon (IFN)-γ and tumor necrosis factor (TNF)-α, which enhances the phagocytosis and clearance of parasite. Lipophosphoglycan 3 (LPG3), the Leishmania homologous with GRP94 (glucose regulated protein 94), a member of HSP90 family, contributes to LPG assembly as the most abundant macromolecule on the surface of Leishmania promastigotes. Methods: We purified NK cells from healthy individuals (n=10) using magnetic-activated cell sorting (MACS) technology. Purified NK cells were co-incubated with different concentrations of recombinant LPG3 (rLPG3), and its N-terminal (NT) and C-terminal (CT) fragments. Finally, the production of IFN-γ and TNF-α by NK cells were measured by ELISA. Results: Recombinant LPG3 but not its fragments (CT and NT), can significantly enhance the production of TNF-α by NK cells (P<0.05). Moreover, rLPG3, CT, and NT fragments were markedly stimulated the secretion of IFN-γ by NK cells (P<0.001). Conclusion: The Leishmania LPG3 antigen can effectively activate NK cells, in vitro. Leishmania LPG3 participates in the innate immunity against leishmaniasis and thereby improves the effective parasite destruction. However, its efficiency should be tested in vivo. PMID:26811721

  7. Recombinant LPG3 Stimulates IFN-Γ and TNF-Α Secretion by Human NK Cells

    PubMed Central

    RASOLZADEH, Sanaz; HAJI FATAHALIHA, Mostafa; HOSSEINI, Maryam; JAFARI, Reza; MIAHIPOUR, Abolfazl; SADREDDINI, Sevil; BABALO, Zohreh; SAMADI KAFIL, Hossein; YOUSEFI, Mehdi

    2015-01-01

    Background: Natural killer (NK) cells play an important role in early stages of innate immune responses against viral and tumoral attacks. Activation of NK cells by leishmaniasis results in secretion of cytokines such as interferon (IFN)-γ and tumor necrosis factor (TNF)-α, which enhance the phagocytosis and clearance of parasite. Lipophosphoglycan 3 (LPG3), the Leishmania homologous with GRP94 (glucose regulated protein 94), a member of HSP90 family, contributes to LPG assembly as the most abundant macromolecule on the surface of Leishmania promastigotes. Methods: We purified NK cells from healthy individuals (n=10) using magnetic-activated cell sorting (MACS) technology. Purified NK cells were co-incubated with different concentrations of recombinant LPG3 (rLPG3), and its N-terminal (NT) and C-terminal (CT) fragments. Finally, the production of IFN-γ and TNF-α by NK cells were measured by ELISA. Results: Recombinant LPG3 but not its fragments (CT and NT), could significantly enhance the production of TNF-α by NK cells (P<0.05). Moreover, rLPG3, CT, and NT fragments were markedly stimulated the secretion of IFN-γ by NK cells (P<0.001). Conclusion: The Leishmania LPG3 antigen could effectively activate NK cells, in vitro. Leishmania LPG3 participates in the innate immunity against leishmaniasis and thereby improves the effective parasite destruction. However, its efficiency should be tested in vivo. PMID:26622301

  8. NK Cell-Specific Gata3 Ablation Identifies the Maturation Program Required for Bone Marrow Exit and Control of Proliferation.

    PubMed

    Ali, Alaa Kassim; Oh, Jun Seok; Vivier, Eric; Busslinger, Meinrad; Lee, Seung-Hwan

    2016-02-15

    NK cells are innate lymphocytes capable of eliciting an innate immune response to pathogens. NK cells develop and become mature in the bone marrow (BM) before they migrate out to peripheral organs. Although the developmental program leading to mature NK cells has been studied in the context of several transcription factors, the stage-specific role of GATA3 in NK cell development has been incompletely understood. Using NKp46-Cre-Gata3(fl/fl) mice in which Gata3 deficiency was induced as early as the immature stage of NK cell differentiation, we demonstrated that GATA3 is required for the NK cell maturation beyond the CD27 single-positive stage and is indispensable for the maintenance of liver-resident NK cells. The frequencies of NK cells from NKp46-Cre-Gata3(fl/fl) mice were found higher in the BM but lower in peripheral organs compared with control littermates, indicating that GATA3 controls the maturation program required for BM egress. Despite the defect in maturation, upon murine CMV infection, NK cells from NKp46-Cre-Gata3(fl/fl) mice expanded vigorously, achieving NK cell frequencies surpassing those in controls and therefore provided comparable protection. The heightened proliferation of NK cells from NKp46-Cre-Gata3(fl/fl) mice was cell intrinsic and associated with enhanced upregulation of CD25 expression. Taken together, our results demonstrate that GATA3 is a critical regulator for NK cell terminal maturation and egress out of the BM and that immature NK cells present in the periphery of NKp46-Cre-Gata3(fl/fl) mice can rapidly expand and provide a reservoir of NK cells capable of mounting an efficient cytotoxic response upon virus infection. PMID:26773150

  9. NK/T cell lymphoma associated with peripheral eosinophilia.

    PubMed

    Yap, E; Wan Jamaluddin, W F; Tumian, N R; Mashuri, F; Mohammed, F; Tan, G C; Masir, N; Abdul Wahid, F S

    2014-12-01

    NK/T cell lymphoma, nasal type is an aggressive and uncommon malignancy. Disease that occurs outside of the aerodigestive tract exhibits an even more aggressive clinical behaviour and does not respond as well to conventional therapy compared to its nasal counterpart. We report such a case of NK/T cell lymphoma, nasal type, that presented as an anterior chest wall mass, arising from the left pectoralis muscle. An interesting feature we wish to highlight is the associated eosinophilia that corresponded to disease activity, exhibiting fluctuations with surgical resection and chemotherapy. To the best of our knowledge this is the third reported case of NK/T cell lymphoma that is associated with peripheral eosinophilia. Our case highlights the role of certain NK cell subsets that play a major role in eosinophilic activation in NK/T lymphomas and calls for more research into further classification of this disease by virtue of its NK cell subsets. PMID:25500520

  10. Cytomegalovirus generates long-lived antigen-specific NK cells with diminished bystander activation to heterologous infection

    PubMed Central

    Min-Oo, Gundula

    2014-01-01

    Natural killer (NK) cells play a key role in the host response to cytomegalovirus (CMV) and can mediate an enhanced response to secondary challenge with CMV. We assessed the ability of mouse CMV (MCMV)–induced memory Ly49H+ NK cells to respond to challenges with influenza, an acute viral infection localized to the lung, and Listeria monocytogenes, a systemic bacterial infection. MCMV-memory NK cells did not display enhanced activation or proliferation after infection with influenza or Listeria, as compared with naive Ly49H+ or Ly49H− NK cells. Memory NK cells also showed impaired activation compared with naive cells when challenged with a mutant MCMV lacking m157, highlighting their antigen-specific response. Ex vivo, MCMV-memory NK cells displayed reduced phosphorylation of STAT4 and STAT1 in response to stimulation by IL-12 and type I interferon (IFN), respectively, and IFN-γ production was reduced in response to IL-12 + IL-18 compared with naive NK cells. However, costimulation of MCMV-memory NK cells with IL-12 and m157 antigen rescues their impaired response compared with cytokines alone. These findings reveal that MCMV-primed memory NK cells are diminished in their response to cytokine-driven bystander responses to heterologous infections as they become specialized and antigen-specific for the control of MCMV upon rechallenge. PMID:25422494

  11. Natural Killer Cells for Immunotherapy – Advantages of the NK-92 Cell Line over Blood NK Cells

    PubMed Central

    Klingemann, Hans; Boissel, Laurent; Toneguzzo, Frances

    2016-01-01

    Natural killer (NK) cells are potent cytotoxic effector cells for cancer therapy and potentially for severe viral infections. However, there are technical challenges to obtain sufficient numbers of functionally active NK cells from a patient’s blood since they represent only 10% of the lymphocytes and are often dysfunctional. The alternative is to obtain cells from a healthy donor, which requires depletion of the allogeneic T cells to prevent graft-versus-host reactions. Cytotoxic cell lines have been established from patients with clonal NK-cell lymphoma. Those cells can be expanded in culture in the presence of IL-2. Except for the NK-92 cell line, though, none of the other six known NK cell lines has consistently and reproducibly shown high antitumor cytotoxicity. Only NK-92 cells can easily be genetically manipulated to recognize specific tumor antigens or to augment monoclonal antibody activity through antibody-dependent cellular cytotoxicity. NK-92 is also the only cell line product that has been infused into patients with advanced cancer with clinical benefit and minimal side effects. PMID:27014270

  12. Chimeric NK-receptor–bearing T cells mediate antitumor immunotherapy

    PubMed Central

    Zhang, Tong; Lemoi, Bethany A.; Sentman, Charles L.

    2005-01-01

    NKG2D is an activating cell-surface receptor expressed on natural killer (NK) cells and some T-cell subsets. Its ligands are primarily expressed on tumor cells. The aim of this study was to determine whether chimeric NK-receptor—bearing T cells would directly kill tumor cells and lead to induction of host immunity against tumors. Chimeric NK receptors were produced by linking NKG2D or DNAX activating protein of 10 kDa (Dap10) to the cytoplasmic portion of the CD3ζ chain. Our results showed that chimeric (ch) NKG2D-bearing T cells responded to NKG2D-ligand–bearing tumor cells (RMA/Rae-1β, EG7) but not to wild-type tumor cells (RMA). This response was dependent upon ligand expression on the target cells but not on expression of major histocompatibility complex (MHC) molecules, and the response could be blocked by anti-NKG2D antibodies. These T cells produced large amounts of T-helper 1 (Th1) cytokines and proinflammatory chemokines and killed ligand–expressing tumor cells. Adoptive transfer of chNKG2D-bearing T cells inhibited RMA/Rae-1β tumor growth in vivo. Moreover, mice that had remained tumor-free were resistant to subsequent challenge with the wild-type RMA tumor cells, suggesting the generation of immunity against other tumor antigens. Taken together, our findings indicate that modification of T cells with chimeric NKG2D receptors represents a promising approach for immunotherapy against cancer. PMID:15890688

  13. Flt3 permits survival during infection by rendering dendritic cells competent to activate NK cells.

    PubMed

    Eidenschenk, Céline; Crozat, Karine; Krebs, Philippe; Arens, Ramon; Popkin, Daniel; Arnold, Carrie N; Blasius, Amanda L; Benedict, Chris A; Moresco, Eva Marie Y; Xia, Yu; Beutler, Bruce

    2010-05-25

    A previously unappreciated signal necessary for dendritic cell (DC)-mediated activation of natural killer (NK) cells during viral infection was revealed by a recessive N-ethyl-N-nitrosourea-induced mutation called warmflash (wmfl). Wmfl homozygotes displayed increased susceptibility to mouse cytomegalovirus (MCMV) infection. In response to MCMV infection in vivo, delayed NK cell activation was observed, but no intrinsic defects in NK cell activation or function were identified. Rather, coculture experiments demonstrated that NK cells are suboptimally activated by wmfl DCs, which showed impaired cytokine production in response to MCMV or synthetic TLR7 and TLR9 ligands. The wmfl mutation was identified in the gene encoding the Fms-like tyrosine kinase 3 (Flt3). Flt3 ligand (Flt3L) is transiently induced in the serum upon infection or TLR activation. However, antibody blockade reveals no acute requirement for Flt3L, suggesting that the Flt3L --> Flt3 axis programs the development of DCs, making them competent to support NK effector function. In the absence of Flt3 signaling, NK cell activation is delayed and survival during MCMV infection is markedly compromised. PMID:20457904

  14. Flt3 permits survival during infection by rendering dendritic cells competent to activate NK cells

    PubMed Central

    Eidenschenk, Céline; Crozat, Karine; Krebs, Philippe; Arens, Ramon; Popkin, Daniel; Arnold, Carrie N.; Blasius, Amanda L.; Benedict, Chris A.; Moresco, Eva Marie Y.; Xia, Yu; Beutler, Bruce

    2010-01-01

    A previously unappreciated signal necessary for dendritic cell (DC)-mediated activation of natural killer (NK) cells during viral infection was revealed by a recessive N-ethyl-N-nitrosourea-induced mutation called warmflash (wmfl). Wmfl homozygotes displayed increased susceptibility to mouse cytomegalovirus (MCMV) infection. In response to MCMV infection in vivo, delayed NK cell activation was observed, but no intrinsic defects in NK cell activation or function were identified. Rather, coculture experiments demonstrated that NK cells are suboptimally activated by wmfl DCs, which showed impaired cytokine production in response to MCMV or synthetic TLR7 and TLR9 ligands. The wmfl mutation was identified in the gene encoding the Fms-like tyrosine kinase 3 (Flt3). Flt3 ligand (Flt3L) is transiently induced in the serum upon infection or TLR activation. However, antibody blockade reveals no acute requirement for Flt3L, suggesting that the Flt3L → Flt3 axis programs the development of DCs, making them competent to support NK effector function. In the absence of Flt3 signaling, NK cell activation is delayed and survival during MCMV infection is markedly compromised. PMID:20457904

  15. A high-throughput assay of NK cell activity in whole blood and its clinical application

    SciTech Connect

    Lee, Saet-byul; Cha, Junhoe; Kim, Im-kyung; Yoon, Joo Chun; Lee, Hyo Joon; Park, Sang Woo; Cho, Sunjung; Youn, Dong-Ye; Lee, Heyja; Lee, Choong Hwan; Lee, Jae Myun; Lee, Kang Young; Kim, Jongsun

    2014-03-14

    Graphical abstract: - Highlights: • We demonstrated a simple assay of NK cell activity from whole blood. • The measurement of secreted IFN-γ from NK cell enables high-throughput screening. • The NKA assay was validated by clinical results of colorectal cancer patients. - Abstract: Natural killer (NK) cells are lymphocytes of the innate immune system and have the ability to kill tumor cells and virus-infected cells without prior sensitization. Malignant tumors and viruses have developed, however, strategies to suppress NK cells to escape from their responses. Thus, the evaluation of NK cell activity (NKA) could be invaluable to estimate the status and the outcome of cancers, viral infections, and immune-mediated diseases. Established methods that measure NKA, such as {sup 51}Cr release assay and CD107a degranulation assay, may be used to determine NK cell function, but they are complicated and time-consuming because they require isolation of peripheral blood mononuclear cells (PBMC) or NK cells. In some cases these assays require hazardous material such as radioactive isotopes. To overcome these difficulties, we developed a simple assay that uses whole blood instead of PBMC or isolated NK cells. This novel assay is suitable for high-throughput screening and the monitoring of diseases, because it employs serum of ex vivo stimulated whole blood to detect interferon (IFN)-γ secreted from NK cells as an indicator of NKA. After the stimulation of NK cells, the determination of IFNγ concentration in serum samples by enzyme-linked immunosorbent assay (ELISA) provided a swift, uncomplicated, and high-throughput assay of NKA ex vivo. The NKA results microsatellite stable (MSS) colorectal cancer patients was showed significantly lower NKA, 263.6 ± 54.5 pg/mL compared with healthy subjects, 867.5 ± 50.2 pg/mL (p value <0.0001). Therefore, the NKA could be utilized as a supportive diagnostic marker for microsatellite stable (MSS) colorectal cancer.

  16. KIR genotype predicts the capacity of human KIR+ CD56dim NK cells to respond to pathogen-associated signals1

    PubMed Central

    Korbel, Daniel S.; Norman, Paul J.; Newman, Kirsty C.; Horowitz, Amir; Gendzekhadze, Ketevan; Parham, Peter; Riley, Eleanor M.

    2010-01-01

    IFN-γ emanating from natural killer (NK) cells is an important component of innate defence against infection. Here we demonstrate that, following in vitro stimulation of human peripheral blood NK cells with a variety of microbial ligands, CD56dim as well as CD56bright NK cells contribute to the overall NK cell IFN-γ response with, for most cell donors, IFN-γ+ CD56dim NK cells outnumbering IFN-γ+ CD56bright NK cells. We also observe that the magnitude of the human NK IFN-γ response to microbial ligands varies between individuals; that the antimicrobial response of CD56bright, but not CD56dim, NK cells is highly correlated with that of myeloid accessory cells; and that the ratio of IFN-γ+ CD56dim to IFN-γ+ CD56bright NK cells following microbial stimulation differs between individuals but remains constant for a given donor over time. Furthermore, ratios of IFN-γ+ CD56dim to IFN-γ+ CD56bright NK cells for different microbial stimuli are highly correlated and the relative response of CD56dim and CD56bright NK cells is highly significantly associated with killer immunoglobulin-like receptor (KIR) genotype. These data reveal an influence of KIR genotype, possibly mediated via NK cell licensing, on the ability of NK cells to respond to non-viral infections and have implications for genetic regulation of susceptibility to infection in humans. PMID:19414796

  17. Trypanosoma brucei Co-opts NK Cells to Kill Splenic B2 B Cells

    PubMed Central

    Frenkel, Deborah; Guirnalda, Patrick; Haynes, Carole; Bockstal, Viki; Magez, Stefan; Black, Samuel J.

    2016-01-01

    After infection with T. brucei AnTat 1.1, C57BL/6 mice lost splenic B2 B cells and lymphoid follicles, developed poor parasite-specific antibody responses, lost weight, became anemic and died with fulminating parasitemia within 35 days. In contrast, infected C57BL/6 mice lacking the cytotoxic granule pore-forming protein perforin (Prf1-/-) retained splenic B2 B cells and lymphoid follicles, developed high-titer antibody responses against many trypanosome polypeptides, rapidly suppressed parasitemia and did not develop anemia or lose weight for at least 60 days. Several lines of evidence show that T. brucei infection-induced splenic B cell depletion results from natural killer (NK) cell-mediated cytotoxicity: i) B2 B cells were depleted from the spleens of infected intact, T cell deficient (TCR-/-) and FcγRIIIa deficient (CD16-/-) C57BL/6 mice excluding a requirement for T cells, NKT cell, or antibody-dependent cell-mediated cytotoxicity; ii) administration of NK1.1 specific IgG2a (mAb PK136) but not irrelevant IgG2a (myeloma M9144) prevented infection-induced B cell depletion consistent with a requirement for NK cells; iii) splenic NK cells but not T cells or NKT cells degranulated in infected C57BL/6 mice co-incident with B cell depletion evidenced by increased surface expression of CD107a; iv) purified NK cells from naïve C57BL/6 mice killed purified splenic B cells from T. brucei infected but not uninfected mice in vitro indicating acquisition of an NK cell activating phenotype by the post-infection B cells; v) adoptively transferred C57BL/6 NK cells prevented infection-induced B cell population growth in infected Prf1-/- mice consistent with in vivo B cell killing; vi) degranulated NK cells in infected mice had altered gene and differentiation antigen expression and lost cytotoxic activity consistent with functional exhaustion, but increased in number as infection progressed indicating continued generation. We conclude that NK cells in T. brucei infected mice

  18. The Self-Specific Activation Receptor SLAM Family Is Critical for NK Cell Education.

    PubMed

    Chen, Shasha; Yang, Meixiang; Du, Juan; Li, Dan; Li, Zehua; Cai, Chenxu; Ma, Yuanwu; Zhang, Lianfeng; Tian, Zhigang; Dong, Zhongjun

    2016-08-16

    NK cell education, a term describing a process for NK cell acquisition of functional competence, is primarily achieved by self-MHC-I-specific inhibitory receptors. In this study, we have demonstrated that SLAM family receptors (SFRs) redundantly expressed on hematopoietic cells function as self-specific activation receptors critical for NK cell education. To overcome gene redundancy, we generated mice simultaneously lacking seven SFRs, revealing that NK-cell-mediated rejection of semi-allogeneic hematopoietic cells largely depended on the presence of SFRs on target cells. This stimulatory effect was determined by the presence of SFR-coupled adaptors; however, SFR-deficient mice displayed enhanced reactivity to hematopoietic cells. These findings demonstrate that SFRs endow NK cells with an ability to kill hematopoietic cells during the effector phase; however, the sustained engagement of SFRs can desensitize NK cell responses during an education process. Therefore, self-specific activating ligands may be "tolerogens" for NK cells, akin to self-antigens that induce T cell tolerance. PMID:27521267

  19. Cytokine secretion and NK cell activity in human ADAM17 deficiency

    PubMed Central

    Chavkin, Maor; Schmiedel, Dominik; Wong, Eitan; Werner, Marion; Yaacov, Barak; Averbuch, Diana; Molho-Pessach, Vered; Stepensky, Polina; Kaynan, Noa; Bar-On, Yotam; Seidel, Einat; Yamin, Rachel; Sagi, Irit; Elpeleg, Orly; Mandelboim, Ofer

    2015-01-01

    Genetic deficiencies provide insights into gene function in humans. Here we describe a patient with a very rare genetic deficiency of ADAM17. We show that the patient's PBMCs had impaired cytokine secretion in response to LPS stimulation, correlating with the clinical picture of severe bacteremia from which the patient suffered. ADAM17 was shown to cleave CD16, a major NK killer receptor. Functional analysis of patient's NK cells demonstrated that his NK cells express normal levels of activating receptors and maintain high surface levels of CD16 following mAb stimulation. Activation of individual NK cell receptors showed that the patient's NK cells are more potent when activated directly by CD16, albeit no difference was observed in Antibody Depedent Cytotoxicity (ADCC) assays. Our data suggest that ADAM17 inhibitors currently considered for clinical use to boost CD16 activity should be cautiously applied, as they might have severe side effects resulting from impaired cytokine secretion. PMID:26683521

  20. Potential of autologous NK cell therapy to eradicate leukemia

    PubMed Central

    Abdel-Azim, Hisham; Heisterkamp, Nora

    2015-01-01

    B-precursor acute lymphoblastic leukemia (BP-ALL) patients are immunocompromised. We recently reported that functional natural killer (NK) cells can be grown from patient bone marrow and blood samples at diagnosis. Surprisingly, such NK cells exhibit cytotoxicity against autologous BP-ALL cells. Here, we outline unanswered questions, challenges and possible applications associated with these findings. PMID:25949882

  1. Qa-1(b)-dependent modulation of dendritic cell and NK cell cross-talk in vivo.

    PubMed

    Colmenero, Paula; Zhang, Angela L; Qian, Ting; Lu, Linrong; Cantor, Harvey; Söderström, Kalle; Engleman, Edgar G

    2007-10-01

    Dendritic cells (DC) trigger activation and IFN-gamma release by NK cells in lymphoid tissues, a process important for the polarization of Th1 responses. Little is known about the molecular signals that regulate DC-induced NK cell IFN-gamma synthesis. In this study, we analyzed whether the interaction between Qa-1(b) expressed on DC and its CD94/NKG2A receptor on NK cells affects this process. Activation of DC using CpG-oligodeoxynucleotides in Qa-1(b)-deficient mice, or transfer of CpG-oligodeoxynucleotide-activated Qa-1(b)-deficient DC into wild-type mice, resulted in dramatically increased IFN-gamma production by NK cells, as compared with that induced by Qa-1(b)-expressing DC. Masking the CD94/NKG2A inhibitory receptor on NK cells in wild-type mice similarly enhanced the IFN-gamma response of these cells to Qa-1(b)-expressing DC. Furthermore, NK cells from CD94/NKG2A-deficient mice displayed higher IFN-gamma production upon DC stimulation. These results demonstrate that Qa-1(b) is critically involved in regulating IFN-gamma synthesis by NK cells in vivo through its interaction with CD94/NKG2A inhibitory receptors. This receptor-ligand interaction may be essential to prevent unabated cytokine production by NK cells during an inflammatory response. PMID:17878358

  2. In vivo tumor surveillance by NK cells requires TYK2 but not TYK2 kinase activity

    PubMed Central

    Prchal-Murphy, Michaela; Witalisz-Siepracka, Agnieszka; Bednarik, Karoline T; Putz, Eva Maria; Gotthardt, Dagmar; Meissl, Katrin; Sexl, Veronika; Müller, Mathias; Strobl, Birgit

    2015-01-01

    Tyrosine kinase 2 (TYK2) is a Janus kinase (JAK) that is crucially involved in inflammation, carcinogenesis and defense against infection. The cytotoxic activity of natural killer (NK) cells in TYK2-deficient (Tyk2−/−) mice is severely reduced, although the underlying mechanisms are largely unknown. Using Tyk2−/− mice and mice expressing a kinase-inactive version of TYK2 (Tyk2K923E), we show that NK cell function is partly independent of the enzymatic activity of TYK2. Tyk2−/− and Tyk2K923E NK cells develop normally in the bone marrow, but the maturation of splenic Tyk2−/− NK cells (and to a lesser extent of Tyk2K923E NK cells) is impaired. In contrast, the production of interferon γ (IFNγ) in response to interleukin 12 (IL-12) or to stimulation through NK cell-activating receptors strictly depends on the presence of enzymatically active TYK2. The cytotoxic activity of Tyk2K923E NK cells against a range of target cells in vitro is higher than that of Tyk2−/− NK cells. Consistently, Tyk2K923E mice control the growth of NK cell-targeted tumors significantly better than TYK2-deficient mice, showing the physiological relevance of the finding. Inhibitors of TYK2's kinase activity are being developed for the treatment of inflammatory diseases and cancers, but their effects on tumor immune surveillance have not been investigated. Our finding that TYK2 has kinase-independent functions in vivo suggests that such inhibitors will leave NK cell mediated tumor surveillance largely intact and that they will be suitable for use in cancer therapy. PMID:26451322

  3. Mechanisms by Which Interleukin-12 Corrects Defective NK Cell Anticryptococcal Activity in HIV-Infected Patients

    PubMed Central

    Kyei, Stephen K.; Ogbomo, Henry; Li, ShuShun; Timm-McCann, Martina; Xiang, Richard F.; Huston, Shaunna M.; Ganguly, Anutosh; Colarusso, Pina; Gill, M. John

    2016-01-01

    ABSTRACT Cryptococcus neoformans is a pathogenic yeast and a leading cause of life-threatening meningitis in AIDS patients. Natural killer (NK) cells are important immune effector cells that directly recognize and kill C. neoformans via a perforin-dependent cytotoxic mechanism. We previously showed that NK cells from HIV-infected patients have aberrant anticryptococcal killing and that interleukin-12 (IL-12) restores the activity at least partially through restoration of NKp30. However, the mechanisms causing this defect or how IL-12 restores the function was unknown. By examining the sequential steps in NK cell killing of Cryptococcus, we found that NK cells from HIV-infected patients had defective binding of NK cells to C. neoformans. Moreover, those NK cells that bound to C. neoformans failed to polarize perforin-containing granules to the microbial synapse compared to healthy controls, suggesting that binding was insufficient to restore a defect in perforin polarization. We also identified lower expression of intracellular perforin and defective perforin release from NK cells of HIV-infected patients in response to C. neoformans. Importantly, treatment of NK cells from HIV-infected patients with IL-12 reversed the multiple defects in binding, granule polarization, perforin content, and perforin release and restored anticryptococcal activity. Thus, there are multiple defects in the cytolytic machinery of NK cells from HIV-infected patients, which cumulatively result in defective NK cell anticryptococcal activity, and each of these defects can be reversed with IL-12. PMID:27555306

  4. Clinical Cancer Therapy by NK Cells via Antibody-Dependent Cell-Mediated Cytotoxicity

    PubMed Central

    Alderson, Kory L.; Sondel, Paul M.

    2011-01-01

    Natural killer (NK) cells are powerful effector cells that can be directed to eliminate tumor cells through tumor-targeted monoclonal antibodies (mAbs). Some tumor-targeted mAbs have been successfully applied in the clinic and are included in the standard of care for certain malignancies. Strategies to augment the antitumor response by NK cells have led to an increased understanding of how to improve their effector responses. Next-generation reagents, such as molecularly modified mAbs and mAb-cytokine fusion proteins (immunocytokines, ICs) designed to augment NK-mediated killing, are showing promise in preclinical and some clinical settings. Continued research into the antitumor effects induced by NK cells and tumor-targeted mAbs suggests that additional intrinsic and extrinsic factors may influence the antitumor response. Therefore more research is needed that focuses on evaluating which NK cell and tumor criteria are best predictive of a clinical response and which combination immunotherapy regimens to pursue for distinct clinical settings. PMID:21660134

  5. Tethering of ICAM on target cells is required for LFA-1-dependent NK cell adhesion and granule polarization

    PubMed Central

    Gross, Catharina C.; Brzostowski, Joseph A.; Liu, Dongfang; Long, Eric O.

    2013-01-01

    αLβ2 integrin (LFA-1) has an important role in the formation of T cell and NK cell cytotoxic immunological synapses and in target cell killing. Binding of LFA-1 to ICAM on target cells promotes not only adhesion, but also polarization of cytolytic granules in NK cells. Here we tested whether LFA-1-dependent NK cell responses are regulated by the distribution and mobility of ICAM at the surface of target cells. We show that depolymerization of F-actin in NK-sensitive target cells abrogated LFA-1-dependent conjugate formation and granule polarization in primary NK cells. Degranulation, which is not controlled by LFA-1, was not impaired. Fluorescence recovery after photobleaching experiments and particle tracking by total internal reflection fluorescence microscopy revealed that ICAM-1 and ICAM-2 were distributed in largely immobile clusters. ICAM clusters were maintained and became highly mobile after actin depolymerization. Moreover, reducing ICAM-2 mobility on an NK-resistant target cell through expression of ezrin, an adapter molecule that tethers proteins to the actin cytoskeleton, enhanced LFA-1-dependent adhesion and granule polarization. Finally, while NK cells kept moving over freely diffusible ICAM-1 on a lipid bilayer, they bound and spread over solid-phase ICAM-1. We conclude that tethering, rather than clustering of ICAM promotes proper signaling by LFA-1 in NK cells. Our findings suggest that the lateral diffusion of integrin ligands on cells may be an important determinant of susceptibility to lysis by cytotoxic lymphocytes. PMID:20675589

  6. NK cell development in bone marrow and liver: site matters.

    PubMed

    Gotthardt, D; Prchal-Murphy, M; Seillet, C; Glasner, A; Mandelboim, O; Carotta, S; Sexl, V; Putz, E M

    2014-12-01

    The NKp46 protein is found on resting and activated natural killer (NK) cells and is involved in the recognition of malignant and infected cells. The expression of NKp46 is believed to precede that of DX5 in early NK cell development. We show that this is not the case in the bone marrow (BM). Here, NKp46 is predominantly expressed after DX5, whereas the liver harbors a subpopulation that expresses NKp46 but not DX5. NK cell precursors in the liver show much lower levels of Eomesodermin than NK cell precursors in the BM, although they express higher levels of granzymes and unlike the NK cell precursors in the BM are fully able to degranulate and produce interferon gamma (IFN-γ). The development of NK cells thus differs between the two organs. This needs to be considered when using NKp46 and DX5 as NK cell markers and when performing NK cell-specific gene deletion in Ncr1 transgenic mice. PMID:25319498

  7. Human Liver Stem Cells Suppress T-Cell Proliferation, NK Activity, and Dendritic Cell Differentiation

    PubMed Central

    Bruno, Stefania; Grange, Cristina; Tapparo, Marta; Pasquino, Chiara; Romagnoli, Renato; Dametto, Ennia; Amoroso, Antonio; Tetta, Ciro; Camussi, Giovanni

    2016-01-01

    Human liver stem cells (HLSCs) are a mesenchymal stromal cell-like population resident in the adult liver. Preclinical studies indicate that HLSCs could be a good candidate for cell therapy. The aim of the present study was to evaluate the immunogenicity and the immunomodulatory properties of HLSCs on T-lymphocytes, natural killer cells (NKs), and dendritic cells (DCs) in allogeneic experimental settings. We found that HLSCs inhibited T-cell proliferation by a mechanism independent of cell contact and dependent on the release of prostaglandin E2 (PGE2) and on indoleamine 2,3-dioxygenase activity. When compared with mesenchymal stromal cells (MSCs), HLSCs were more efficient in inhibiting T-cell proliferation. At variance with MSCs, HLSCs did not elicit NK degranulation. Moreover, HLSCs inhibited NK degranulation against K562, a NK-sensitive target, by a mechanism dependent on HLA-G release. When tested on DC generation from monocytes, HLSCs were found to impair DC differentiation and DCs ability to induce T-cell proliferation through PGE2. This study shows that HLSCs have immunomodulatory properties similar to MSCs, but, at variance with MSCs, they do not elicit a NK response. PMID:27127520

  8. Fully functional NK cells after unrelated cord blood transplantation.

    PubMed

    Beziat, V; Nguyen, S; Lapusan, S; Hervier, B; Dhedin, N; Bories, D; Uzunov, M; Boudifa, A; Trebeden-Negre, H; Norol, F; Marjanovic, Z; Marie, J-P; Vernant, J-P; Debre, P; Rio, B; Vieillard, V

    2009-04-01

    Promising results of umbilical cord blood transplantation (UCBT) from unrelated donors have been reported in patients with hematologic disorders. These transplants, having potential to trigger beneficial donor-versus-recipient natural killer (NK) cell-mediated alloreaction, we have conducted the first extensive analysis of the phenotypic and functional properties of NK cells after UCBT. NK cells from 25 patients with high-risk hematologic malignancies were compared with cells derived from both healthy adult and CB cells. We found that following UCBT, NK cells display not only some phenotypic features associated with maturity but also unique characteristics that make them fully functional against leukemic blasts. We propose that this full functionality of alloreactive donor-derived NK may drive graft-versus-leukemia reactions after UCBT. PMID:19151772

  9. Increased NK Cell Maturation in Patients with Acute Myeloid Leukemia

    PubMed Central

    Chretien, Anne-Sophie; Granjeaud, Samuel; Gondois-Rey, Françoise; Harbi, Samia; Orlanducci, Florence; Blaise, Didier; Vey, Norbert; Arnoulet, Christine; Fauriat, Cyril; Olive, Daniel

    2015-01-01

    Understanding immune alterations in cancer patients is a major challenge and requires precise phenotypic study of immune subsets. Improvement of knowledge regarding the biology of natural killer (NK) cells and technical advances leads to the generation of high dimensional dataset. High dimensional flow cytometry requires tools adapted to complex dataset analyses. This study presents an example of NK cell maturation analysis in Healthy Volunteers (HV) and patients with Acute Myeloid Leukemia (AML) with an automated procedure using the FLOCK algorithm. This procedure enabled to automatically identify NK cell subsets according to maturation profiles, with 2D mapping of a four-dimensional dataset. Differences were highlighted in AML patients compared to HV, with an overall increase of NK maturation. Among patients, a strong heterogeneity in NK cell maturation defined three distinct profiles. Overall, automatic gating with FLOCK algorithm is a recent procedure, which enables fast and reliable identification of cell populations from high-dimensional cytometry data. Such tools are necessary for immune subset characterization and standardization of data analyses. This tool is adapted to new immune cell subsets discovery, and may lead to a better knowledge of NK cell defects in cancer patients. Overall, 2D mapping of NK maturation profiles enabled fast and reliable identification of NK cell subsets. PMID:26594214

  10. Phenotypically distinct helper NK cells are required for gp96-mediated anti-tumor immunity

    PubMed Central

    Sedlacek, Abigail L.; Kinner-Bibeau, Lauren B.; Binder, Robert J.

    2016-01-01

    A number of Heat Shock Proteins (HSPs), in the extracellular environment, are immunogenic. Following cross-presentation of HSP-chaperoned peptides by CD91+ antigen presenting cells (APCs), T cells are primed with specificity for the derivative antigen-bearing cell. Accordingly, tumor-derived HSPs are in clinical trials for cancer immunotherapy. We investigate the role of NK cells in gp96-mediated anti-tumor immune responses given their propensity to lyse tumor cells. We show that gp96-mediated rejection of tumors requires a unique and necessary helper role in NK cells. This helper role occurs during the effector phase of the anti-tumor immune response and is required for T cell and APC function. Gp96 activates NK cells indirectly via APCs to a phenotype distinct from NK cells activated by other mechanisms such as IL-2. While NK cells have both lytic and cytokine producing properties, we show that gp96 selectively activates cytokine production in NK cells, which is important in the HSP anti-tumor immune response, and leaves their cytotoxic capacity unchanged. PMID:27431727

  11. Hepatitis C Virus Sensing by Human Trophoblasts Induces Innate Immune Responses and Recruitment of Maternal NK Cells: Potential Implications for Limiting Vertical Transmission.

    PubMed

    Giugliano, Silvia; Petroff, Margaret G; Warren, Bryce D; Jasti, Susmita; Linscheid, Caitlin; Ward, Ashley; Kramer, Anita; Dobrinskikh, Evgenia; Sheiko, Melissa A; Gale, Michael; Golden-Mason, Lucy; Winn, Virginia D; Rosen, Hugo R

    2015-10-15

    Hepatitis C virus (HCV) is the world's most common blood-borne viral infection for which there is no vaccine. The rates of vertical transmission range between 3 and 6% with odds 90% higher in the presence of HIV coinfection. Prevention of vertical transmission is not possible because of lack of an approved therapy for use in pregnancy or an effective vaccine. Recently, HCV has been identified as an independent risk factor for preterm delivery, perinatal mortality, and other complications. In this study, we characterized the immune responses that contribute to the control of viral infection at the maternal-fetal interface (MFI) in the early gestational stages. In this study, we show that primary human trophoblast cells and an extravillous trophoblast cell line (HTR8), from first and second trimester of pregnancy, express receptors relevant for HCV binding/entry and are permissive for HCV uptake. We found that HCV-RNA sensing by human trophoblast cells induces robust upregulation of type I/III IFNs and secretion of multiple chemokines that elicit recruitment and activation of decidual NK cells. Furthermore, we observed that HCV-RNA transfection induces a proapoptotic response within HTR8 that could affect the morphology of the placenta. To our knowledge, for the first time, we demonstrate that HCV-RNA sensing by human trophoblast cells elicits a strong antiviral response that alters the recruitment and activation of innate immune cells at the MFI. This work provides a paradigm shift in our understanding of HCV-specific immunity at the MFI as well as novel insights into mechanisms that limit vertical transmission but may paradoxically lead to virus-related pregnancy complications. PMID:26342030

  12. NK Cell Subtypes as Regulators of Autoimmune Liver Disease

    PubMed Central

    2016-01-01

    As major components of innate immunity, NK cells not only exert cell-mediated cytotoxicity to destroy tumors or infected cells, but also act to regulate the functions of other cells in the immune system by secreting cytokines and chemokines. Thus, NK cells provide surveillance in the early defense against viruses, intracellular bacteria, and cancer cells. However, the effecter function of NK cells must be exquisitely controlled to prevent inadvertent attack against normal “self” cells. In an organ such as the liver, where the distinction between immunotolerance and immune defense against routinely processed pathogens is critical, the plethora of NK cells has a unique role in the maintenance of homeostasis. Once self-tolerance is broken, autoimmune liver disease resulted. NK cells act as a “two-edged weapon” and even play opposite roles with both regulatory and inducer activities in the hepatic environment. That is, NK cells act not only to produce inflammatory cytokines and chemokines, but also to alter the proliferation and activation of associated lymphocytes. However, the precise regulatory mechanisms at work in autoimmune liver diseases remain to be identified. In this review, we focus on recent research with NK cells and their potential role in the development of autoimmune liver disease. PMID:27462349

  13. NK Cell Subtypes as Regulators of Autoimmune Liver Disease.

    PubMed

    Jiao, Guohui; Wang, Bangmao

    2016-01-01

    As major components of innate immunity, NK cells not only exert cell-mediated cytotoxicity to destroy tumors or infected cells, but also act to regulate the functions of other cells in the immune system by secreting cytokines and chemokines. Thus, NK cells provide surveillance in the early defense against viruses, intracellular bacteria, and cancer cells. However, the effecter function of NK cells must be exquisitely controlled to prevent inadvertent attack against normal "self" cells. In an organ such as the liver, where the distinction between immunotolerance and immune defense against routinely processed pathogens is critical, the plethora of NK cells has a unique role in the maintenance of homeostasis. Once self-tolerance is broken, autoimmune liver disease resulted. NK cells act as a "two-edged weapon" and even play opposite roles with both regulatory and inducer activities in the hepatic environment. That is, NK cells act not only to produce inflammatory cytokines and chemokines, but also to alter the proliferation and activation of associated lymphocytes. However, the precise regulatory mechanisms at work in autoimmune liver diseases remain to be identified. In this review, we focus on recent research with NK cells and their potential role in the development of autoimmune liver disease. PMID:27462349

  14. Mechanism of NK cell activation induced by coculture with dendritic cells derived from peripheral blood monocytes

    PubMed Central

    Amakata, Y; Fujiyama, Y; Andoh, A; Hodohara, K; Bamba, T

    2001-01-01

    Dendritic cells (DCs) have been regarded as one of the effective antigen-presenting cells, but the relationship between DCs and lymphocytes, in particular natural killer (NK) cells, remains unclear. In this study, we evaluated how DCs interact with both lymphocytes and NK cells using a coculture system. The number of lymphocytes increased significantly when cocultured with DCs (1·8-fold increase). In particular, the proliferation of NK cells was prominent. Furthermore, the coculture of DCs with lymphocytes induced a marked increase in IL-12 and IFN-γ secretion. When contact between the DCs and lymphocytes was prevented, the secretion of both IL-12 and IFN-γ was markedly reduced. IFN-γ production was completely blocked by an anti-IL-12 antibody, indicating that IFN-γ secretion was dependent on IL-12 secretion. The stimulating effect of the DCs on the proliferation of the lymphocytes was partially suppressed by anti-IL-12 antibodies, and was completely attenuated when cellular contact was prevented. Furthermore, the NK cell proliferation induced by coculture with DCs was significantly blocked by the inhibition of the interaction of either CD40–CD40L or CD28–B7 molecule. The coculture with DCs enhanced NK activity by 40%, and this was partially suppressed by anti-IL-12 antibodies and was completely blocked by the inhibition of cell-to-cell contact. These results indicate that the activation of NK cells by DCs is partially mediated by IL-12 secretion, and that direct contact between DCs and NK cells play a major role in this response. PMID:11422197

  15. Cryptococcus neoformans Directly Stimulates Perforin Production and Rearms NK Cells for Enhanced Anticryptococcal Microbicidal Activity▿

    PubMed Central

    Marr, Kaleb J.; Jones, Gareth J.; Zheng, Chunfu; Huston, Shaunna M.; Timm-McCann, Martina; Islam, Anowara; Berenger, Byron M.; Ma, Ling Ling; Wiseman, Jeremy C. D.; Mody, Christopher H.

    2009-01-01

    NK cells, in addition to possessing antitumor and antiviral activity, exhibit perforin-dependent microbicidal activity against the opportunistic pathogen Cryptococcus neoformans. However, the factors controlling this response, particularly whether the pathogen itself provides an activation or rearming signal, are largely unknown. The current studies were performed to determine whether exposure to this fungus alters subsequent NK cell anticryptococcal activity. NK cells lost perforin and mobilized lysosome-associated membrane protein 1 to the cell surface following incubation with the fungus, indicating that degranulation had occurred. Despite a reduced perforin content during killing, NK cells acquired an enhanced ability to kill C. neoformans, as demonstrated using auxotrophs that allowed independent assessment of the killing of two strains. De novo protein synthesis was required for optimal killing; however, there was no evidence that a soluble factor contributed to the enhanced anticryptococcal activity. Exposure of NK cells to C. neoformans caused the cells to rearm, as demonstrated by increased perforin mRNA levels and enhanced loss of perforin when transcription was blocked. Degranulation alone was insufficient to provide the activation signal as NK cells lost anticryptococcal activity following treatment with strontium chloride. However, NK cells regained the activity upon prolonged exposure to C. neoformans, which is consistent with activation by the microbe. The enhanced cytotoxicity did not extend to tumor killing since NK cells exposed to C. neoformans failed to kill NK-sensitive tumor targets (K562 cells). These studies demonstrate that there is contact-mediated microbe-specific rearming and activation of microbicidal activity that are necessary for optimal killing of C. neoformans. PMID:19307209

  16. Histone Deacetylase Inhibitors Enhance CD4 T Cell Susceptibility to NK Cell Killing but Reduce NK Cell Function

    PubMed Central

    Pace, Matthew; Williams, James; Kurioka, Ayako; Gerry, Andrew B.; Jakobsen, Bent; Klenerman, Paul; Nwokolo, Nneka; Fox, Julie

    2016-01-01

    In the search for a cure for HIV-1 infection, histone deacetylase inhibitors (HDACi) are being investigated as activators of latently infected CD4 T cells to promote their targeting by cytotoxic T-lymphocytes (CTL). However, HDACi may also inhibit CTL function, suggesting different immunotherapy approaches may need to be explored. Here, we study the impact of different HDACi on both Natural Killer (NK) and CTL targeting of HIV-1 infected cells. We found HDACi down-regulated HLA class I expression independently of HIV-1 Nef which, without significantly compromising CTL function, led to enhanced targeting by NK cells. HDACi-treated HIV-1-infected CD4 T cells were also more effectively cleared than untreated controls during NK co-culture. However, HDACi impaired NK function, reducing degranulation and killing capacity. Depending on the HDACi and dose, this impairment could counteract the benefit gained by treating infected target cells. These data suggest that following HDACi-induced HLA class I down-regulation NK cells kill HIV-1-infected cells, although HDACi-mediated NK cell inhibition may negate this effect. Our data emphasize the importance of studying the effects of potential interventions on both targets and effectors. PMID:27529554

  17. Targeting NK Cells for Anticancer Immunotherapy: Clinical and Preclinical Approaches

    PubMed Central

    Carotta, Sebastian

    2016-01-01

    The recent success of checkpoint blockade has highlighted the potential of immunotherapy approaches for cancer treatment. Although the majority of approved immunotherapy drugs target T cell subsets, it is appreciated that other components of the immune system have important roles in tumor immune surveillance as well and thus represent promising additional targets for immunotherapy. Natural killer (NK) cells are the body’s first line of defense against infected or transformed cells, as they kill target cells in an antigen-independent manner. Although several studies have clearly demonstrated the active role of NK cells in cancer immune surveillance, only few clinically approved therapies currently exist that harness their potential. Our increased understanding of NK cell biology over the past few years has renewed the interest in NK cell-based anticancer therapies, which has lead to a steady increase of NK cell-based clinical and preclinical trials. Here, the role of NK cells in cancer immune surveillance is summarized, and several novel approaches to enhance NK cell cytotoxicity against cancer are discussed. PMID:27148271

  18. Impaired NK Cell Activation and Chemotaxis toward Dendritic Cells Exposed to Complement-Opsonized HIV-1

    PubMed Central

    Ellegård, Rada; Crisci, Elisa; Andersson, Jonas; Shankar, Esaki M.; Nyström, Sofia; Hinkula, Jorma

    2015-01-01

    Mucosa resident dendritic cells (DCs) may represent one of the first immune cells that HIV-1 encounters during sexual transmission. The virions in body fluids can be opsonized with complement factors because of HIV-mediated triggering of the complement cascade, and this appears to influence numerous aspects of the immune defense targeting the virus. One key attribute of host defense is the ability to attract immune cells to the site of infection. In this study, we investigated whether the opsonization of HIV with complement (C-HIV) or a mixture of complement and Abs (CI-HIV) affected the cytokine and chemokine responses generated by DCs, as well as their ability to attract other immune cells. We found that the expression levels of CXCL8, CXCL10, CCL3, and CCL17 were lowered after exposure to either C-HIV or CI-HIV relative to free HIV (F-HIV). DCs exposed to F-HIV induced higher cell migration, consisting mainly of NK cells, compared with opsonized virus, and the chemotaxis of NK cells was dependent on CCL3 and CXCL10. NK cell exposure to supernatants derived from HIV-exposed DCs showed that F-HIV induced phenotypic activation (e.g., increased levels of TIM3, CD69, and CD25) and effector function (e.g., production of IFNγ and killing of target cells) in NK cells, whereas C-HIV and CI-HIV did not. The impairment of NK cell recruitment by DCs exposed to complement-opsonized HIV and the lack of NK activation may contribute to the failure of innate immune responses to control HIV at the site of initial mucosa infection. PMID:26157174

  19. Impaired NK Cell Activation and Chemotaxis toward Dendritic Cells Exposed to Complement-Opsonized HIV-1.

    PubMed

    Ellegård, Rada; Crisci, Elisa; Andersson, Jonas; Shankar, Esaki M; Nyström, Sofia; Hinkula, Jorma; Larsson, Marie

    2015-08-15

    Mucosa resident dendritic cells (DCs) may represent one of the first immune cells that HIV-1 encounters during sexual transmission. The virions in body fluids can be opsonized with complement factors because of HIV-mediated triggering of the complement cascade, and this appears to influence numerous aspects of the immune defense targeting the virus. One key attribute of host defense is the ability to attract immune cells to the site of infection. In this study, we investigated whether the opsonization of HIV with complement (C-HIV) or a mixture of complement and Abs (CI-HIV) affected the cytokine and chemokine responses generated by DCs, as well as their ability to attract other immune cells. We found that the expression levels of CXCL8, CXCL10, CCL3, and CCL17 were lowered after exposure to either C-HIV or CI-HIV relative to free HIV (F-HIV). DCs exposed to F-HIV induced higher cell migration, consisting mainly of NK cells, compared with opsonized virus, and the chemotaxis of NK cells was dependent on CCL3 and CXCL10. NK cell exposure to supernatants derived from HIV-exposed DCs showed that F-HIV induced phenotypic activation (e.g., increased levels of TIM3, CD69, and CD25) and effector function (e.g., production of IFNγ and killing of target cells) in NK cells, whereas C-HIV and CI-HIV did not. The impairment of NK cell recruitment by DCs exposed to complement-opsonized HIV and the lack of NK activation may contribute to the failure of innate immune responses to control HIV at the site of initial mucosa infection. PMID:26157174

  20. Activating KIRs and NKG2C in Viral Infections: Toward NK Cell Memory?

    PubMed Central

    Della Chiesa, Mariella; Sivori, Simona; Carlomagno, Simona; Moretta, Lorenzo; Moretta, Alessandro

    2015-01-01

    Natural killer (NK) cells are important players in the immune defense against viral infections. The contribution of activating killer immunoglobulin-like receptors (KIRs) and CD94/NKG2C in regulating anti-viral responses has recently emerged. Thus, in the hematopoietic stem cell transplantation setting, the presence of donor activating KIRs (aKIRs) may protect against viral infections, while in HIV-infected individuals, KIR3DS1, in combination with HLA-Bw4-I80, results in reduction of viral progression. Since, studies have been performed mainly at the genetic or transcriptional level, the effective size, the function, and the “licensing” status of NK cells expressing aKIRs, as well as the nature of their viral ligands, require further investigation. Certain viral infections, mainly due to Human cytomegalovirus (HCMV), can deeply influence the NK cell development and function by inducing a marked expansion of mature NKG2C+ NK cells expressing self-activating KIRs. This suggests that NKG2C and/or aKIRs are involved in the selective proliferation of this subset. The persistent, HCMV-induced, imprinting suggests that NK cells may display unexpected adaptive immune traits. The role of aKIRs and NKG2C in regulating NK cell responses and promoting a memory-like response to certain viruses is discussed. PMID:26617607

  1. Microchip-Based Single-Cell Imaging Reveals That CD56dimCD57-KIR-NKG2A+ NK Cells Have More Dynamic Migration Associated with Increased Target Cell Conjugation and Probability of Killing Compared to CD56dimCD57-KIR-NKG2A- NK Cells.

    PubMed

    Forslund, Elin; Sohlberg, Ebba; Enqvist, Monika; Olofsson, Per E; Malmberg, Karl-Johan; Önfelt, Björn

    2015-10-01

    NK cells are functionally educated by self-MHC specific receptors, including the inhibitory killer cell Ig-like receptors (KIRs) and the lectin-like CD94/NKG2A heterodimer. Little is known about how NK cell education influences qualitative aspects of cytotoxicity such as migration behavior and efficacy of activation and killing at the single-cell level. In this study, we have compared the behavior of FACS-sorted CD56(dim)CD57(-)KIR(-)NKG2A(+) (NKG2A(+)) and CD56(dim)CD57(-)KIR(-)NKG2A(-) (lacking inhibitory receptors; IR(-)) human NK cells by quantifying migration, cytotoxicity, and contact dynamics using microchip-based live cell imaging. NKG2A(+) NK cells displayed a more dynamic migration behavior and made more contacts with target cells than IR(-) NK cells. NKG2A(+) NK cells also more frequently killed the target cells once a conjugate had been formed. NK cells with serial killing capacity were primarily found among NKG2A(+) NK cells. Conjugates involving IR(-) NK cells were generally more short-lived and IR(-) NK cells did not become activated to the same extent as NKG2A(+) NK cells when in contact with target cells, as evident by their reduced spreading response. In contrast, NKG2A(+) and IR(-) NK cells showed similar dynamics in terms of duration of conjugation periods and NK cell spreading response in conjugates that led to killing. Taken together, these observations suggest that the high killing capacity of NKG2A(+) NK cells is linked to processes regulating events in the recognition phase of NK-target cell contact rather than events after cytotoxicity has been triggered. PMID:26320254

  2. The lytic potential of human liver NK cells is restricted by their limited expression of inhibitory killer Ig-like receptors.

    PubMed

    Burt, Bryan M; Plitas, George; Zhao, Zeguo; Bamboat, Zubin M; Nguyen, Hoang M; Dupont, Bo; DeMatteo, Ronald P

    2009-08-01

    The human liver is enriched in NK cells which are potent effectors of the innate immune system. We have determined that liver NK cells freshly isolated from surgical specimens from patients with hepatic malignancy have less cytolytic activity than autologous blood NK cells. This difference was due to a higher proportion of CD16(-) NK cells in the liver and reduced cytotoxicity by CD16(+) liver NK cells compared with their blood counterparts. CD16(+) liver NK cells had similar expression of activating NK receptors and had similar intracellular granzyme B and perforin content compared with CD16(+) blood NK cells. CD16(+) liver NK cells contained a reduced fraction of cells with inhibitory killer Ig-like receptors specific for self-MHC class I (self-killer Ig-related receptor (KIR)) and an increased fraction of self-KIR(neg)NKG2A(pos) and self-KIR(neg)NKG2A(neg) cells. Using single-cell analysis of intracellular IFN-gamma production and cytotoxicity assays, we determined that CD16(+) liver NK cells expressing self-KIR were more responsive to target cells than those cells that did not express self-KIR molecules. CD16(+) liver NK cells gained cytolytic function when stimulated with IL-2 or cultured with LPS or poly(I:C)-activated autologous liver Kupffer cells. Thus, the human liver contains NK cell subsets which have reduced effector function, but under appropriate inflammatory conditions become potent killers. PMID:19587011

  3. NK cell tolerance as the final endorsement of prenatal tolerance after in utero hematopoietic cellular transplantation

    PubMed Central

    Alhajjat, Amir M.; Lee, Amanda E.; Strong, Beverly S.; Shaaban, Aimen F.

    2015-01-01

    The primary benefits of in utero hematopoietic cellular transplantation (IUHCT) arise from transplanting curative cells prior to the immunologic maturation of the fetus. However, this approach has been routinely successful only in the treatment of congenital immunodeficiency diseases that include an inherent NK cell deficiency despite the existence of normal maternal immunity in either setting. These observations raise the possibility that fetal NK cells function as an early barrier to allogeneic IUHCT. Herein, we summarize the findings of previous studies of prenatal NK cell allospecific tolerance in mice and in humans. Cumulatively, this new information reveals the complexity of the fetal immune response in the setting of rejection or tolerance and illustrates the role for fetal NK cells in the final endorsement of allospecific prenatal tolerance. PMID:25852555

  4. Shared alterations in NK cell frequency, phenotype, and function in chronic human immunodeficiency virus and hepatitis C virus infections.

    PubMed

    Meier, Ute-Christiane; Owen, Rachel E; Taylor, Elizabeth; Worth, Andrew; Naoumov, Nikolai; Willberg, Christian; Tang, Kwok; Newton, Phillipa; Pellegrino, Pierre; Williams, Ian; Klenerman, Paul; Borrow, Persephone

    2005-10-01

    Human immunodeficiency virus (HIV) and hepatitis C virus (HCV) cause clinically important persistent infections. The effects of virus persistence on innate immunity, including NK cell responses, and the underlying mechanisms are not fully understood. We examined the frequency, phenotype, and function of peripheral blood CD3- CD56+ NK subsets in HIV+ and HCV+ patients and identified significantly reduced numbers of total NK cells and a striking shift in NK subsets, with a marked decrease in the CD56(dim) cell fraction compared to CD56(bright) cells, in both infections. This shift influenced the phenotype and functional capacity (gamma interferon production, killing) of the total NK pool. In addition, abnormalities in the functional capacity of the CD56(dim) NK subset were observed in HIV+ patients. The shared NK alterations were found to be associated with a significant reduction in serum levels of the innate cytokine interleukin 15 (IL-15). In vitro stimulation with IL-15 rescued NK cells of HIV+ and HCV+ patients from apoptosis and enhanced proliferation and functional activity. We hypothesize that the reduced levels of IL-15 present in the serum during HIV and HCV infections might impact NK cell homeostasis, contributing to the common alterations of the NK pool observed in these unrelated infections. PMID:16160163

  5. Natural killer cell dysfunction in hepatocellular carcinoma and NK cell-based immunotherapy.

    PubMed

    Sun, Cheng; Sun, Hao-yu; Xiao, Wei-hua; Zhang, Cai; Tian, Zhi-gang

    2015-10-01

    The mechanisms linking hepatitis B virus (HBV) and hepatitis C virus (HCV) infection to hepatocellular carcinoma (HCC) remain largely unknown. Natural killer (NK) cells account for 25%-50% of the total number of liver lymphocytes, suggesting that NK cells play an important role in liver immunity. The number of NK cells in the blood and tumor tissues of HCC patients is positively correlated with their survival and prognosis. Furthermore, a group of NK cell-associated genes in HCC tissues is positively associated with the prolonged survival. These facts suggest that NK cells and HCC progression are strongly associated. In this review, we describe the abnormal NK cells and their functional impairment in patients with chronic HBV and HCV infection, which contribute to the progression of HCC. Then, we summarize the association of NK cells with HCC based on the abnormalities in the numbers and phenotypes of blood and liver NK cells in HCC patients. In particular, the exhaustion of NK cells that represents lower cytotoxicity and impaired cytokine production may serve as a predictor for the occurrence of HCC. Finally, we present the current achievements in NK cell immunotherapy conducted in mouse models of liver cancer and in clinical trials, highlighting how chemoimmunotherapy, NK cell transfer, gene therapy, cytokine therapy and mAb therapy improve NK cell function in HCC treatment. It is conceivable that NK cell-based anti-HCC therapeutic strategies alone or in combination with other therapies will be great promise for HCC treatment. PMID:26073325

  6. NAP-2 Secreted by Human NK Cells Can Stimulate Mesenchymal Stem/Stromal Cell Recruitment

    PubMed Central

    Almeida, Catarina R.; Caires, Hugo R.; Vasconcelos, Daniela P.; Barbosa, Mário A.

    2016-01-01

    Summary Strategies for improved homing of mesenchymal stem cells (MSCs) to a place of injury are being sought and it has been shown that natural killer (NK) cells can stimulate MSC recruitment. Here, we studied the chemokines behind this recruitment. Assays were performed with bone marrow human MSCs and NK cells freshly isolated from healthy donor buffy coats. Supernatants from MSC-NK cell co-cultures can induce MSC recruitment but not to the same extent as when NK cells are present. Antibody arrays and ELISA assays confirmed that NK cells secrete RANTES (CCL5) and revealed that human NK cells secrete NAP-2 (CXCL7), a chemokine that can induce MSC migration. Inhibition with specific antagonists of CXCR2, a receptor that recognizes NAP-2, abolished NK cell-mediated MSC recruitment. This capacity of NK cells to produce chemokines that stimulate MSC recruitment points toward a role for this immune cell population in regulating tissue repair/regeneration. PMID:27052313

  7. [Commemorative lecture of receiving Imamura Memorial Prize. NK cell in pulmonary tuberculosis from basic and clinical point of view].

    PubMed

    Yoneda, T

    1996-11-01

    . Purified NK cells inhibit intracellular replication within monocytes in dose response manner. PMID:8958676

  8. Human NK cell development requires CD56-mediated motility and formation of the developmental synapse.

    PubMed

    Mace, Emily M; Gunesch, Justin T; Dixon, Amera; Orange, Jordan S

    2016-01-01

    While distinct stages of natural killer (NK) cell development have been defined, the molecular interactions that shape human NK cell maturation are poorly understood. Here we define intercellular interactions between developing NK cells and stromal cells which, through contact-dependent mechanisms, promote the generation of mature, functional human NK cells from CD34(+) precursors. We show that developing NK cells undergo unique, developmental stage-specific sustained and transient interactions with developmentally supportive stromal cells, and that the relative motility of NK cells increases as they move through development in vitro and ex vivo. These interactions include the formation of a synapse between developing NK cells and stromal cells, which we term the developmental synapse. Finally, we identify a role for CD56 in developmental synapse structure, NK cell motility and NK cell development. Thus, we define the developmental synapse leading to human NK cell functional maturation. PMID:27435370

  9. Human NK cell development requires CD56-mediated motility and formation of the developmental synapse

    PubMed Central

    Mace, Emily M.; Gunesch, Justin T.; Dixon, Amera; Orange, Jordan S.

    2016-01-01

    While distinct stages of natural killer (NK) cell development have been defined, the molecular interactions that shape human NK cell maturation are poorly understood. Here we define intercellular interactions between developing NK cells and stromal cells which, through contact-dependent mechanisms, promote the generation of mature, functional human NK cells from CD34+ precursors. We show that developing NK cells undergo unique, developmental stage-specific sustained and transient interactions with developmentally supportive stromal cells, and that the relative motility of NK cells increases as they move through development in vitro and ex vivo. These interactions include the formation of a synapse between developing NK cells and stromal cells, which we term the developmental synapse. Finally, we identify a role for CD56 in developmental synapse structure, NK cell motility and NK cell development. Thus, we define the developmental synapse leading to human NK cell functional maturation. PMID:27435370

  10. Variable NK cell receptors exemplified by human KIR3DL1/S1.

    PubMed

    Parham, Peter; Norman, Paul J; Abi-Rached, Laurent; Guethlein, Lisbeth A

    2011-07-01

    Variegated expression of variable NK cell receptors for polymorphic MHC class I broadens the range of an individual's NK cell response and the capacity for populations and species to survive disease epidemics and population bottlenecks. On evolutionary time scales, this component of immunity is exceptionally dynamic, unstable, and short-lived, being dependent on coevolution of ligands and receptors subject to varying, competing selection pressures. Consequently these systems of variable NK cell receptors are largely species specific and have recruited different classes of glycoprotein, even within the primate order of mammals. Such disparity helps to explain substantial differences in NK cell biology between humans and animal models, for which the population genetics is largely ignored. KIR3DL1/S1, which recognizes the Bw4 epitope of HLA-A and -B and is the most extensively studied of the variable NK cell receptors, exemplifies how variation in all possible parameters of function is recruited to diversify the human NK cell response. PMID:21690332

  11. NLRC5 shields T lymphocytes from NK-cell-mediated elimination under inflammatory conditions

    PubMed Central

    Ludigs, Kristina; Jandus, Camilla; Utzschneider, Daniel T.; Staehli, Francesco; Bessoles, Stéphanie; Dang, Anh Thu; Rota, Giorgia; Castro, Wilson; Zehn, Dietmar; Vivier, Eric; Held, Werner; Romero, Pedro; Guarda, Greta

    2016-01-01

    NLRC5 is a transcriptional regulator of MHC class I (MHCI), which maintains high MHCI expression particularly in T cells. Recent evidence highlights an important NK–T-cell crosstalk, raising the question on whether NLRC5 specifically modulates this interaction. Here we show that NK cells from Nlrc5-deficient mice exhibit moderate alterations in inhibitory receptor expression and responsiveness. Interestingly, NLRC5 expression in T cells is required to protect them from NK-cell-mediated elimination upon inflammation. Using T-cell-specific Nlrc5-deficient mice, we show that NK cells surprisingly break tolerance even towards ‘self' Nlrc5-deficient T cells under inflammatory conditions. Furthermore, during chronic LCMV infection, the total CD8+ T-cell population is severely decreased in these mice, a phenotype reverted by NK-cell depletion. These findings strongly suggest that endogenous T cells with low MHCI expression become NK-cell targets, having thus important implications for T-cell responses in naturally or therapeutically induced inflammatory conditions. PMID:26861112

  12. Hematological Malignancies Escape from NK Cell Innate Immune Surveillance: Mechanisms and Therapeutic Implications

    PubMed Central

    Farnault, Laure; Sanchez, Carole; Baier, Céline; Le Treut, Thérèse; Costello, Régis T.

    2012-01-01

    Hematological malignancies treatment improved over the last years resulting in increased achievement of complete or partial remission, but unfortunately high relapse rates are still observed. Therefore, sustainment of long-term remission is crucial. Immune system has a key role in tumor surveillance. Natural killer (NK) cells, at the frontier of innate and adaptive immune system, have a central role in tumor cells surveillance as demonstrated in the setting of allogenic stem cell transplantation. Nevertheless, tumor cells develop various mechanisms to escape from NK cells innate immune pressure. Abnormal NK cytolytic functions have been described in nearly all hematological malignancies. We present here various mechanisms involved in the escape of hematological malignancies from NK cells surveillance: NK cells quantitative deficiency and NK cell qualitative deficiency by increased inhibition signaling or decreased activating stimuli. A challenge of immunotherapy is to restore an efficient antitumor response. A combination of classical therapy plus immune modulation strategies will soon become a standard of care for hematological malignancies. PMID:22899948

  13. Isolation and (111)In-Oxine Labeling of Murine NK Cells for Assessment of Cell Trafficking in Orthotopic Lung Tumor Model.

    PubMed

    Malviya, Gaurav; Nayak, Tapan; Gerdes, Christian; Dierckx, Rudi A J O; Signore, Alberto; de Vries, Erik F J

    2016-04-01

    A noninvasive in vivo imaging method for NK cell trafficking is essential to gain further understanding of the pathogenesis of NK cell mediated immune response to the novel cancer treatment strategies, and to discover the homing sites and physiological distribution of NK cells. Although human NK cells can be labeled for in vivo imaging, little is known about the murine NK cell labeling and its application in animal models. This study describes the isolation and ex vivo radiolabeling of murine NK cells for the evaluation of cell trafficking in an orthotopic model of human lung cancer in mice. Scid-Tg(FCGR3A)Blt transgenic SCID mice were used to isolate NK cells from mouse splenocytes using the CD49b (DX5) MicroBeads positive selection method. The purity and viability of the isolated NK cells were confirmed by FACS analysis. Different labeling buffers and incubation times were evaluated to optimize (111)In-oxine labeling conditions. Functionality of the radiolabeled NK cell was assessed by (51)Cr-release assay. We evaluated physiological distribution of (111)In-oxine labeled murine NK cells in normal SCID mice and biodistribution in irradiated and nonirradiated SCID mice with orthotopic A549 human lung tumor lesions. Imaging findings were confirmed by histology. Results showed that incubation with 0.011 MBq of (111)In-oxine per million murine NK cells in PBS (pH 7.4) for 20 min is the best condition that provides optimum labeling efficiency without affecting cell viability and functionality. Physiological distribution in normal SCID mice demonstrated NK cells homing mainly in the spleen, while (111)In released from NK cells was excreted via kidneys into urine. Biodistribution studies demonstrated a higher lung uptake in orthotopic lung tumor-bearing mice than control mice. In irradiated mice, lung tumor uptake of radiolabeled murine NK cells decreased between 24 h and 72 h postinjection (p.i.), which was accompanied by tumor regression, while in nonirradiated mice

  14. Role of the NK Cell-Activating Receptor CRACC in Periodontitis

    PubMed Central

    Krämer, Benjamin; Kebschull, Moritz; Nowak, Michael; Demmer, Ryan T.; Haupt, Manuela; Körner, Christian; Perner, Sven; Jepsen, Søren

    2013-01-01

    Periodontitis is a highly prevalent, biofilm-mediated chronic inflammatory disease that results in the loss of the tooth-supporting tissues. It features two major clinical entities: chronic periodontitis, which is more common, and aggressive periodontitis, which usually has an early onset and a rapid progression. Natural killer (NK) cells are a distinct subgroup of lymphocytes that play a major role in the ability of the innate immune system to steer immune responses. NK cells are abundant in periodontitis lesions, and NK cell activation has been causally linked to periodontal tissue destruction. However, the exact mechanisms of their activation and their role in the pathophysiology of periodontitis are elusive. Here, we show that the predominant NK cell-activating molecule in periodontitis is CD2-like receptor activating cytotoxic cells (CRACC). We show that CRACC induction was significantly more pronounced in aggressive than chronic periodontitis and correlated positively with periodontal disease severity, subgingival levels of specific periodontal pathogens, and NK cell activation in vivo. We delineate how Aggregatibacter actinomycetemcomitans, an oral pathogen that is causally associated with aggressive periodontitis, indirectly induces CRACC on NK cells via activation of dendritic cells and subsequent interleukin 12 (IL-12) signaling. In contrast, we demonstrate that fimbriae from Porphyromonas gingivalis, a principal pathogen in chronic periodontitis, actively attenuate CRACC induction on NK cells. Our data suggest an involvement of CRACC-mediated NK cell activation in periodontal tissue destruction and point to a plausible distinction in the pathobiology of aggressive and chronic periodontitis that may help explain the accelerated tissue destruction in aggressive periodontitis. PMID:23250953

  15. Bacterial Manipulation of NK Cell Regulatory Activity Increases Susceptibility to Listeria monocytogenes Infection.

    PubMed

    Clark, Sarah E; Filak, Holly C; Guthrie, Brandon S; Schmidt, Rebecca L; Jamieson, Amanda; Merkel, Patricia; Knight, Vijaya; Cole, Caroline M; Raulet, David H; Lenz, Laurel L

    2016-06-01

    Natural killer (NK) cells produce interferon (IFN)-γ and thus have been suggested to promote type I immunity during bacterial infections. Yet, Listeria monocytogenes (Lm) and some other pathogens encode proteins that cause increased NK cell activation. Here, we show that stimulation of NK cell activation increases susceptibility during Lm infection despite and independent from robust NK cell production of IFNγ. The increased susceptibility correlated with IL-10 production by responding NK cells. NK cells produced IL-10 as their IFNγ production waned and the Lm virulence protein p60 promoted induction of IL-10 production by mouse and human NK cells. NK cells consequently exerted regulatory effects to suppress accumulation and activation of inflammatory myeloid cells. Our results reveal new dimensions of the role played by NK cells during Lm infection and demonstrate the ability of this bacterial pathogen to exploit the induction of regulatory NK cell activity to increase host susceptibility. PMID:27295349

  16. Bacterial Manipulation of NK Cell Regulatory Activity Increases Susceptibility to Listeria monocytogenes Infection

    PubMed Central

    Guthrie, Brandon S.; Schmidt, Rebecca L.; Jamieson, Amanda; Merkel, Patricia; Knight, Vijaya; Cole, Caroline M.; Raulet, David H.; Lenz, Laurel L.

    2016-01-01

    Natural killer (NK) cells produce interferon (IFN)-γ and thus have been suggested to promote type I immunity during bacterial infections. Yet, Listeria monocytogenes (Lm) and some other pathogens encode proteins that cause increased NK cell activation. Here, we show that stimulation of NK cell activation increases susceptibility during Lm infection despite and independent from robust NK cell production of IFNγ. The increased susceptibility correlated with IL-10 production by responding NK cells. NK cells produced IL-10 as their IFNγ production waned and the Lm virulence protein p60 promoted induction of IL-10 production by mouse and human NK cells. NK cells consequently exerted regulatory effects to suppress accumulation and activation of inflammatory myeloid cells. Our results reveal new dimensions of the role played by NK cells during Lm infection and demonstrate the ability of this bacterial pathogen to exploit the induction of regulatory NK cell activity to increase host susceptibility. PMID:27295349

  17. Human NK cell repertoire diversity reflects immune experience and correlates with viral susceptibility

    PubMed Central

    Strauss-Albee, Dara M.; Fukuyama, Julia; Liang, Emily C.; Yao, Yi; Jarrell, Justin A.; Drake, Alison L.; Kinuthia, John; Montgomery, Ruth R.; John-Stewart, Grace; Holmes, Susan; Blish, Catherine A.

    2015-01-01

    Innate natural killer (NK) cells are diverse at the single-cell level because of variegated expressions of activating and inhibitory receptors, yet the developmental roots and functional consequences of this diversity remain unknown. Because NK cells are critical for antiviral and antitumor responses, a better understanding of their diversity could lead to an improved ability to harness them therapeutically. We found that NK diversity is lower at birth than in adults. During an antiviral response to either HIV-1 or West Nile virus, NK diversity increases, resulting in terminal differentiation and cytokine production at the cost of cell division and degranulation. In African women matched for HIV-1 exposure risk, high NK diversity is associated with increased risk of HIV-1 acquisition. Existing diversity may therefore decrease the flexibility of the antiviral response. Collectively, the data reveal that human NK diversity is a previously undefined metric of immune history and function that may be clinically useful in forecasting the outcomes of infection and malignancy. PMID:26203083

  18. Contribution of Thy1+ NK cells to protective IFN-γ production during Salmonella Typhimurium infections

    PubMed Central

    Kupz, Andreas; Scott, Timothy A.; Belz, Gabrielle T.; Andrews, Daniel M.; Greyer, Marie; Lew, Andrew M.; Brooks, Andrew G.; Smyth, Mark J.; Curtiss, Roy; Bedoui, Sammy; Strugnell, Richard A.

    2013-01-01

    IFN-γ is critical for immunity against infections with intracellular pathogens, such as Salmonella enterica. However, which of the many cell types capable of producing IFN-γ controls Salmonella infections remains unclear. Using a mouse model of systemic Salmonella infection, we observed that only a lack of all lymphocytes or CD90 (Thy1)+ cells, but not the absence of T cells, Retinoic acid-related orphan receptor (ROR)-γt–dependent lymphocytes, (NK)1.1+ cells, natural killer T (NKT), and/or B cells alone, replicated the highly susceptible phenotype of IFN-γ–deficient mice to Salmonella infection. A combination of antibody depletions and adoptive transfer experiments revealed that early protective IFN-γ was provided by Thy1-expressing natural killer (NK) cells and that these cells improved antibacterial immunity through the provision of IFN-γ. Further analysis of NK cells producing IFN-γ in response to Salmonella indicated that less mature NK cells were more efficient at mediating antibacterial effector function than terminally differentiated NK cells. Inspired by recent reports of Thy1+ NK cells contributing to immune memory, we analyzed their role in secondary protection against otherwise lethal WT Salmonella infections. Notably, we observed that a newly generated Salmonella vaccine strain not only conferred superior protection compared with conventional regimens but that this enhanced efficiency of recall immunity was afforded by incorporating CD4−CD8−Thy1+ cells into the secondary response. Taken together, these findings demonstrate that Thy1-expressing NK cells play an important role in antibacterial immunity. PMID:23345426

  19. CIS is a potent checkpoint in NK cell-mediated tumor immunity.

    PubMed

    Delconte, Rebecca B; Kolesnik, Tatiana B; Dagley, Laura F; Rautela, Jai; Shi, Wei; Putz, Eva M; Stannard, Kimberley; Zhang, Jian-Guo; Teh, Charis; Firth, Matt; Ushiki, Takashi; Andoniou, Christopher E; Degli-Esposti, Mariapia A; Sharp, Phillip P; Sanvitale, Caroline E; Infusini, Giuseppe; Liau, Nicholas P D; Linossi, Edmond M; Burns, Christopher J; Carotta, Sebastian; Gray, Daniel H D; Seillet, Cyril; Hutchinson, Dana S; Belz, Gabrielle T; Webb, Andrew I; Alexander, Warren S; Li, Shawn S; Bullock, Alex N; Babon, Jeffery J; Smyth, Mark J; Nicholson, Sandra E; Huntington, Nicholas D

    2016-07-01

    The detection of aberrant cells by natural killer (NK) cells is controlled by the integration of signals from activating and inhibitory ligands and from cytokines such as IL-15. We identified cytokine-inducible SH2-containing protein (CIS, encoded by Cish) as a critical negative regulator of IL-15 signaling in NK cells. Cish was rapidly induced in response to IL-15, and deletion of Cish rendered NK cells hypersensitive to IL-15, as evidenced by enhanced proliferation, survival, IFN-γ production and cytotoxicity toward tumors. This was associated with increased JAK-STAT signaling in NK cells in which Cish was deleted. Correspondingly, CIS interacted with the tyrosine kinase JAK1, inhibiting its enzymatic activity and targeting JAK for proteasomal degradation. Cish(-/-) mice were resistant to melanoma, prostate and breast cancer metastasis in vivo, and this was intrinsic to NK cell activity. Our data uncover a potent intracellular checkpoint in NK cell-mediated tumor immunity and suggest possibilities for new cancer immunotherapies directed at blocking CIS function. PMID:27213690

  20. Resveratrol Induces Cell Cycle Arrest and Apoptosis in Malignant NK Cells via JAK2/STAT3 Pathway Inhibition

    PubMed Central

    Quoc Trung, Ly; Espinoza, J. Luis; Takami, Akiyoshi; Nakao, Shinji

    2013-01-01

    Natural killer (NK) cell malignancies, particularly aggressive NK cell leukaemias and lymphomas, have poor prognoses. Although recent regimens with L-asparaginase substantially improved outcomes, novel therapeutic approaches are still needed to enhance clinical response. Resveratrol, a naturally occurring polyphenol, has been extensively studied for its anti-inflammatory, cardioprotective and anti-cancer activities. In this study, we investigated the potential anti-tumour activities of resveratrol against the NK cell lines KHYG-1, NKL, NK-92 and NK-YS. Resveratrol induced robust G0/G1 cell cycle arrest, significantly suppressed cell proliferation and induced apoptosis in a dose- and time-dependent manner for all four cell lines. In addition, resveratrol suppressed constitutively active STAT3 in all the cell lines and inhibited JAK2 phosphorylation but had no effect on other upstream mediators of STAT3 activation, such as PTEN, TYK2, and JAK1. Resveratrol also induced downregulation of the anti-apoptotic proteins MCL1 and survivin, two downstream effectors of the STAT3 pathway. Finally, resveratrol induced synergistic effect on the apoptotic and antiproliferative activities of L-asparaginase against KHYG-1, NKL and NK-92 cells. These results suggest that resveratrol may have therapeutic potential against NK cell malignancies. Furthermore, our finding that resveratrol is a bonafide JAK2 inhibitor extends its potential benefits to other diseases with dysregulated JAK2 signaling. PMID:23372833

  1. Expansion of NK Cells Using Genetically Engineered K562 Feeder Cells.

    PubMed

    Phan, Minh-Trang Thi; Lee, Seung-Hwan; Kim, Sang-Ki; Cho, Duck

    2016-01-01

    Natural killer (NK) cells can be expanded upon activation by proliferative cytokines (such as IL-2 and IL-15). The NK cell expansion can be greatly enhanced by proteins from feeder cells such as tumor cell lines or PBMCs. Therefore, coculture systems of irradiated feeder cells and NK cells in media containing IL-2 and IL-15 have been developed to generate large numbers of NK cells, although NK cell expansion protocol using anti-CD3 antibody (OKT-3) without feeder cells has also been developed. Commonly used feeder cell lines are RPMI8866, Epstein-Barr lymphoblastoid cell line (EBV-LCL), and K562. Stimulation with NK-sensitive K562 cells is known to augment NK cell proliferation to IL-2, IL-15, and IL-21 in combination.Recently, remarkable NK cell-expansion rates are achieved when genetically engineered (GE) feeder cells are used. Dr. Dario Campana's group found that membrane-bound IL-15 and 4-1BBL, coexpressed by K562 cells, acted synergistically to augment K562-specific NK stimulatory capacity, resulting in vigorous expansion of peripheral blood CD56(+) CD3(-) NK cells without concomitant growth of T lymphocytes. Here, we describe an in vitro expansion method of human NK cells among PBMCs by coculturing with GE_K562 cells. PMID:27177665

  2. NCR1 is an activating receptor expressed on a subset of canine NK cells.

    PubMed

    Grøndahl-Rosado, Christine; Boysen, Preben; Johansen, Grethe M; Brun-Hansen, Hege; Storset, Anne K

    2016-09-01

    Defining NK cells has been challenging in many veterinary species. Although several groups have described putative NK cell populations, there is still no consensus on a definition of NK cells in the dog. In the present study, canine NK cells are characterized as CD3(-)GranzymeB(+) cells, further divided into a NCR1(+) and a NCR1(-) subset. All dogs examined displayed both subsets in blood, although of quite variable magnitude. Following vaccination an increase was observed in the CD3(-) NCR1(-) cell population in blood, but not in the CD3(-) NCR1(+) population. Non-B non-T cell cultures stimulated with IL-2 and IL-15 were dominated by CD3(-)GranzymeB(+) cells after approximately 2 weeks and a large proportion of the CD3(-)GranzymeB(+) cells expressed NCR1. IL-12 stimulation lead to a further upregulation resulting in an almost uniform expression of NCR1. The cultured cells expressed MHC class II, showed a variable expression of CD8 and were negative for CD4 and CD21. The cultures were able to kill known NK cell targets, and NCR1 was shown to be a major activating receptor. A large proportion of the NCR1(+) cells, but none of the NCR1(-) cells, produced IFNγ in response to IL-12 stimulation. These results show that NCR1 defines two subsets of canine NK cells, likely to represent different activation stages, and that NCR1 acts as an activating receptor on canine NK cells. PMID:27436439

  3. Data correlations between gender, cytomegalovirus infection and T cells, NK cells, and soluble immune mediators in elderly humans.

    PubMed

    Al-Attar, Ahmad; Presnell, Steven R; Peterson, Charlotte A; Thomas, D Travis; Lutz, Charles T

    2016-09-01

    We describe a cohort of 50 elderly subjects, age at least 70 years. We present gender-specific findings in T lymphocyte markers and soluble immune mediators. We show the correlation between cytomegalovirus infection status with CD56(dim) NK cell responses to a variety of stimuli and with CD56(bright)/CD56(dim) NK cell ratio. We also present the correlation of retinol binding protein (RBP)-4 plasma levels with NK cell responses and we explore the relationship between gender and adiponectin, 25(OH)D (vitamin D), and RBP4 in affecting CD56(dim) NK cell responses. These data are discussed in Al-Attar et al. (2016) [1]. PMID:27508213

  4. Bispecific and trispecific killer cell engagers directly activate human NK cells through CD16 signaling and induce cytotoxicity and cytokine production

    PubMed Central

    Gleason, Michelle K.; Verneris, Michael R.; Todhunter, Deborah A.; Zhang, Bin; McCullar, Valarie; Zhou, Sophia X.; Panoskaltsis-Mortari, Angela; Weiner, Louis M.; Vallera, Daniel A.; Miller, Jeffrey S.

    2012-01-01

    This study evaluates the mechanism by which bispecific and trispecific killer cell engagers (BiKEs and TriKEs) act to trigger human NK cell effector function and investigates their ability to induce NK cell cytokine and chemokine production against human B-cell leukemia. We examined the ability of BiKEs and TriKEs to trigger NK cell activation through direct CD16 signaling, measuring intracellular Ca2+ mobilization, secretion of lytic granules, induction of target cell apoptosis and production of cytokine and chemokines in response to the Raji cell line and primary leukemia targets. Resting NK cells triggered by the recombinant reagents led to intracellular Ca2+ mobilization through direct CD16 signaling. Co-culture of reagent-treated resting NK cells with Raji targets resulted in significant increases in NK cell degranulation and target cell death. BiKEs and TriKEs effectively mediated NK cytotoxicity of Raji targets at high and low effector-to-target (E:T) ratios and maintained functional stability after 24 and 48 hours of culture in human serum. NK cell production of IFN-γ, TNF-α, GM-CSF, IL-8, MIP-1α and RANTES was differentially induced in the presence of recombinant reagents and Raji targets. Moreover, significant increases in NK cell degranulation and enhancement of IFN-γ production against primary ALL and CLL targets were induced with reagent treatment of resting NK cells. In conclusion, BiKEs and TriKEs directly trigger NK cell activation through CD16, significantly increasing NK cell cytolytic activity and cytokine production against tumor targets, demonstrating their therapeutic potential for enhancing NK cell immunotherapies for leukemias and lymphomas. PMID:23075808

  5. Human NK cell lytic granules and regulation of their exocytosis

    PubMed Central

    Krzewski, Konrad; Coligan, John E.

    2012-01-01

    Natural killer (NK) cells form a subset of lymphocytes that play a key role in immuno-surveillance and host defense against cancer and viral infections. They recognize stressed cells through a variety of germline-encoded activating cell surface receptors and utilize their cytotoxic ability to eliminate abnormal cells. Killing of target cells is a complex, multi-stage process that concludes in the directed secretion of lytic granules, containing perforin and granzymes, at the immunological synapse. Upon delivery to a target cell, perforin mediates generation of pores in membranes of target cells, allowing granzymes to access target cell cytoplasm and induce apoptosis. Therefore, lytic granules of NK cells are indispensable for normal NK cell cytolytic function. Indeed, defects in lytic granule secretion lead or are related to serious and often fatal diseases, such as familial hemophagocytic lymphohistiocytosis (FHL) type 2–5 or Griscelli syndrome type 2. A number of reports highlight the role of several proteins involved in lytic granule release and NK cell-mediated killing of tumor cells. This review focuses on lytic granules of human NK cells and the advancements in understanding the mechanisms controlling their exocytosis. PMID:23162553

  6. Selection and expansion of natural killer cells for NK cell-based immunotherapy.

    PubMed

    Becker, Petra S A; Suck, Garnet; Nowakowska, Paulina; Ullrich, Evelyn; Seifried, Erhard; Bader, Peter; Tonn, Torsten; Seidl, Christian

    2016-04-01

    Natural killer (NK) cells have been used in several clinical trials as adaptive immunotherapy. The low numbers of these cells in peripheral blood mononuclear cells (PBMC) have resulted in various approaches to preferentially expand primary NK cells from PBMC. While some clinical trials have used the addition of interleukin 2 (IL-2) to co-stimulate the expansion of purified NK cells from allogeneic donors, recent studies have shown promising results in achieving in vitro expansion of NK cells to large numbers for adoptive immunotherapy. NK cell expansion requires multiple cell signals for survival, proliferation and activation. Thus, expansion strategies have been focused either to substitute these factors using autologous feeder cells or to use genetically modified allogeneic feeder cells. Recent developments in the clinical use of genetically modified NK cell lines with chimeric antigen receptors, the development of expansion protocols for the clinical use of NK cell from human embryonic stem cells and induced pluripotent stem cells are challenging improvements for NK cell-based immunotherapy. Transfer of several of these protocols to clinical-grade production of NK cells necessitates adaptation of good manufacturing practice conditions, and the development of freezing conditions to establish NK cell stocks will require some effort and, however, should enhance the therapeutic options of NK cells in clinical medicine. PMID:26810567

  7. Human Lymphoid Tissues Harbor a Distinct CD69+CXCR6+ NK Cell Population.

    PubMed

    Lugthart, Gertjan; Melsen, Janine E; Vervat, Carly; van Ostaijen-Ten Dam, Monique M; Corver, Willem E; Roelen, Dave L; van Bergen, Jeroen; van Tol, Maarten J D; Lankester, Arjan C; Schilham, Marco W

    2016-07-01

    Knowledge of human NK cells is based primarily on conventional CD56(bright) and CD56(dim) NK cells from blood. However, most cellular immune interactions occur in lymphoid organs. Based on the coexpression of CD69 and CXCR6, we identified a third major NK cell subset in lymphoid tissues. This population represents 30-60% of NK cells in marrow, spleen, and lymph node but is absent from blood. CD69(+)CXCR6(+) lymphoid tissue NK cells have an intermediate expression of CD56 and high expression of NKp46 and ICAM-1. In contrast to circulating NK cells, they have a bimodal expression of the activating receptor DNAX accessory molecule 1. CD69(+)CXCR6(+) NK cells do not express the early markers c-kit and IL-7Rα, nor killer cell Ig-like receptors or other late-differentiation markers. After cytokine stimulation, CD69(+)CXCR6(+) NK cells produce IFN-γ at levels comparable to CD56(dim) NK cells. They constitutively express perforin but require preactivation to express granzyme B and exert cytotoxicity. After hematopoietic stem cell transplantation, CD69(+)CXCR6(+) lymphoid tissue NK cells do not exhibit the hyperexpansion observed for both conventional NK cell populations. CD69(+)CXCR6(+) NK cells constitute a separate NK cell population with a distinct phenotype and function. The identification of this NK cell population in lymphoid tissues provides tools to further evaluate the cellular interactions and role of NK cells in human immunity. PMID:27226093

  8. The early intestinal immune response in experimental neonatal ovine cryptosporidiosis is characterized by an increased frequency of perforin expressing NCR1(+) NK cells and by NCR1(-) CD8(+) cell recruitment.

    PubMed

    Olsen, Line; Åkesson, Caroline Piercey; Storset, Anne K; Lacroix-Lamandé, Sonia; Boysen, Preben; Metton, Coralie; Connelley, Timothy; Espenes, Arild; Laurent, Fabrice; Drouet, Françoise

    2015-01-01

    Cryptosporidium parvum, a zoonotic protozoan parasite, causes important losses in neonatal ruminants. Innate immunity plays a key role in controlling the acute phase of this infection. The participation of NCR1+ Natural Killer (NK) cells in the early intestinal innate immune response to the parasite was investigated in neonatal lambs inoculated at birth. The observed increase in the lymphocyte infiltration was further studied by immunohistology and flow cytometry with focus on distribution, density, cellular phenotype related to cytotoxic function and activation status. The frequency of NCR1+ cells did not change with infection, while their absolute number slightly increased in the jejunum and the CD8+/NCR1- T cell density increased markedly. The frequency of perforin+ cells increased significantly with infection in the NCR1+ population (in both NCR1+/CD16+ and NCR1+/CD16- populations) but not in the NCR1-/CD8+ population. The proportion of NCR1+ cells co-expressing CD16+ also increased. The fraction of cells expressing IL2 receptor (CD25), higher in the NCR1+/CD8+ population than among the CD8+/NCR1- cells in jejunal Peyer's patches, remained unchanged during infection. However, contrary to CD8+/NCR1- lymphocytes, the intensity of CD25 expressed by NCR1+ lymphocytes increased in infected lambs. Altogether, the data demonstrating that NK cells are highly activated and possess a high cytotoxic potential very early during infection, concomitant with an up-regulation of the interferon gamma gene in the gut segments, support the hypothesis that they are involved in the innate immune response against C. parvum. The early significant recruitment of CD8+/NCR1- T cells in the small intestine suggests that they could rapidly drive the establishment of the acquired immune response. PMID:25890354

  9. Tissue-resident and memory properties of human T-cell and NK-cell subsets.

    PubMed

    Lugli, Enrico; Hudspeth, Kelly; Roberto, Alessandra; Mavilio, Domenico

    2016-08-01

    Efficient immune responses to invading pathogens are the result of the complex but coordinated synergy between a variety of cell types from both the innate and adaptive arms of the immune system. While adaptive and innate immune responses are highly complementary, some cells types within these two systems perform similar functions, underscoring the need for redundancy and increased flexibility. In this review, we will discuss the striking shared features of immunological memory and tissue residency recently discovered between T cells, a component of the adaptive immune system, and natural killer (NK) cells, members generally assigned to the innate compartment. Specifically, we will focus on the T-cell and NK-cell diversity at the single-cell level, on the discrete function of specific subsets, and on their anatomical location. Finally, we will discuss the implication of such diversity in the generation of long-term memory. PMID:27431095

  10. NK cells promote neutrophil recruitment in the brain during sepsis-induced neuroinflammation

    PubMed Central

    He, Hao; Geng, Tingting; Chen, Piyun; Wang, Meixiang; Hu, Jingxia; Kang, Li; Song, Wengang; Tang, Hua

    2016-01-01

    Sepsis could affect the central nervous system and thus induces neuroinflammation, which subsequently leads to brain damage or dysfunction. However, the mechanisms of generation of neuroinflammation during sepsis remain poorly understood. By administration of lipopolysaccharides (LPS) in mice to mimic sepsis, we found that shortly after opening the blood–brain barrier, conventional CD11b+CD27+ NK subset migrated into the brain followed by subsequent neutrophil infiltration. Interestingly, depletion of NK cells prior to LPS treatment severely impaired neutrophil recruitment in the inflamed brain. By in vivo recruitment assay, we found that brain-infiltrated NK cells displayed chemotactic activity to neutrophils, which depended on the higher expression of chemokines such as CXCL2. Moreover, microglia were also responsible for neutrophil recruitment, and their chemotactic activity was significantly impaired by ablation of NK cells. Furthermore, depletion of NK cells could significantly ameliorate depression-like behavior in LPS-treated mice. These data indicated a NK cell-regulated neutrophil recruitment in the blamed brain, which also could be seen on another sepsis model, cecal ligation and puncture. So, our findings revealed an important scenario in the generation of sepsis-induced neuroinflammation. PMID:27270556

  11. NK cells promote neutrophil recruitment in the brain during sepsis-induced neuroinflammation.

    PubMed

    He, Hao; Geng, Tingting; Chen, Piyun; Wang, Meixiang; Hu, Jingxia; Kang, Li; Song, Wengang; Tang, Hua

    2016-01-01

    Sepsis could affect the central nervous system and thus induces neuroinflammation, which subsequently leads to brain damage or dysfunction. However, the mechanisms of generation of neuroinflammation during sepsis remain poorly understood. By administration of lipopolysaccharides (LPS) in mice to mimic sepsis, we found that shortly after opening the blood-brain barrier, conventional CD11b(+)CD27(+) NK subset migrated into the brain followed by subsequent neutrophil infiltration. Interestingly, depletion of NK cells prior to LPS treatment severely impaired neutrophil recruitment in the inflamed brain. By in vivo recruitment assay, we found that brain-infiltrated NK cells displayed chemotactic activity to neutrophils, which depended on the higher expression of chemokines such as CXCL2. Moreover, microglia were also responsible for neutrophil recruitment, and their chemotactic activity was significantly impaired by ablation of NK cells. Furthermore, depletion of NK cells could significantly ameliorate depression-like behavior in LPS-treated mice. These data indicated a NK cell-regulated neutrophil recruitment in the blamed brain, which also could be seen on another sepsis model, cecal ligation and puncture. So, our findings revealed an important scenario in the generation of sepsis-induced neuroinflammation. PMID:27270556

  12. TGF-β inhibits the activation and functions of NK cells by repressing the mTOR pathway.

    PubMed

    Viel, Sébastien; Marçais, Antoine; Guimaraes, Fernando Souza-Fonseca; Loftus, Roisin; Rabilloud, Jessica; Grau, Morgan; Degouve, Sophie; Djebali, Sophia; Sanlaville, Amélien; Charrier, Emily; Bienvenu, Jacques; Marie, Julien C; Caux, Christophe; Marvel, Jacqueline; Town, Liam; Huntington, Nicholas D; Bartholin, Laurent; Finlay, David; Smyth, Mark J; Walzer, Thierry

    2016-02-16

    Transforming growth factor-β (TGF-β) is a major immunosuppressive cytokine that maintains immune homeostasis and prevents autoimmunity through its antiproliferative and anti-inflammatory properties in various immune cell types. We provide genetic, pharmacologic, and biochemical evidence that a critical target of TGF-β signaling in mouse and human natural killer (NK) cells is the serine and threonine kinase mTOR (mammalian target of rapamycin). Treatment of mouse or human NK cells with TGF-β in vitro blocked interleukin-15 (IL-15)-induced activation of mTOR. TGF-β and the mTOR inhibitor rapamycin both reduced the metabolic activity and proliferation of NK cells and reduced the abundances of various NK cell receptors and the cytotoxic activity of NK cells. In vivo, constitutive TGF-β signaling or depletion of mTOR arrested NK cell development, whereas deletion of the TGF-β receptor subunit TGF-βRII enhanced mTOR activity and the cytotoxic activity of the NK cells in response to IL-15. Suppression of TGF-β signaling in NK cells did not affect either NK cell development or homeostasis; however, it enhanced the ability of NK cells to limit metastases in two different tumor models in mice. Together, these results suggest that the kinase mTOR is a crucial signaling integrator of pro- and anti-inflammatory cytokines in NK cells. Moreover, we propose that boosting the metabolic activity of antitumor lymphocytes could be an effective strategy to promote immune-mediated tumor suppression. PMID:26884601

  13. SHP-2 expression negatively regulates NK cell function1,2

    PubMed Central

    Purdy, Amanda K.; Campbell, Kerry S.

    2009-01-01

    Src homology region 2-containing protein tyrosine phosphatase-2 (SHP-2)4 is required for full activation of Ras/ERK in many cytokine and growth factor receptor signaling pathways. In contrast, SHP-2 inhibits activation of human natural killer (NK) cells upon recruitment to killer cell Ig-like receptors (KIR)4. To determine how SHP-2 impacts NK cell activation in KIR-dependent or KIR-independent signaling pathways, we employed knockdown and overexpression strategies in NK-like cell lines and analyzed the consequences on functional responses. In response to stimulation with susceptible target cells, SHP-2-silenced NK cells had elevated cytolytic activity and IFN-γ production, whereas cells overexpressing wild type or gain-of-function mutants of SHP-2 exhibited dampened activities. Increased levels of SHP-2 expression over this range significantly suppressed microtubule organizing center (MTOC)4 polarization and granzyme B release in response to target cells. Interestingly, NK-target cell conjugation was only reduced by overexpressing SHP-2, but not potentiated in SHP-2-silenced cells, indicating that conjugation is not influenced by physiological levels of SHP-2 expression. KIR-dependent inhibition of cytotoxicity was unaffected by significant reductions in SHP-2 levels, presumably because KIR were still capable of recruiting the phosphatase under these limiting conditions. In contrast, the general suppressive effect of SHP-2 on cytotoxicity and cytokine release was much more sensitive to changes in cellular SHP-2 levels. In summary, our studies have identified a new, KIR-independent role for SHP-2 in dampening NK cell activation in response to tumor target cells in a concentration-dependent manner. This suppression of activation impacts MTOC-based cytoskeletal rearrangement and granule release. PMID:19915046

  14. Novel immune modulators used in hematology: impact on NK cells.

    PubMed

    Krieg, Stephanie; Ullrich, Evelyn

    2012-01-01

    There is a wide range of important pharmaceuticals used in treatment of cancer. Besides their known effects on tumor cells, there is growing evidence for modulation of the immune system. Immunomodulatory drugs (IMiDs(®)) play an important role in the treatment of patients with multiple myeloma or myelodysplastic syndrome and have already demonstrated antitumor, anti-angiogenic, and immunostimulating effects, in particular on natural killer (NK) cells. Tyrosine kinase inhibitors are directly targeting different kinases and are known to regulate effector NK cells and expression of NKG2D ligands (NKG2DLs) on tumor cells. Demethylating agents, histone deacetylases, and proteasome inhibitors interfere with the epigenetic regulation and protein degradation of malignant cells. There are first hints that these drugs also sensitize tumor cells to chemotherapy, radiation, and NK cell-mediated cytotoxicity by enhanced expression of TRAIL and NKG2DLs. However, these pharmaceuticals may also impair NK cell function in a dose- and time-dependent manner. In summary, this review provides an update on the effects of different novel molecules on the immune system focusing NK cells. PMID:23316191

  15. Novel immune modulators used in hematology: impact on NK cells

    PubMed Central

    Krieg, Stephanie; Ullrich, Evelyn

    2013-01-01

    There is a wide range of important pharmaceuticals used in treatment of cancer. Besides their known effects on tumor cells, there is growing evidence for modulation of the immune system. Immunomodulatory drugs (IMiDs®) play an important role in the treatment of patients with multiple myeloma or myelodysplastic syndrome and have already demonstrated antitumor, anti-angiogenic, and immunostimulating effects, in particular on natural killer (NK) cells. Tyrosine kinase inhibitors are directly targeting different kinases and are known to regulate effector NK cells and expression of NKG2D ligands (NKG2DLs) on tumor cells. Demethylating agents, histone deacetylases, and proteasome inhibitors interfere with the epigenetic regulation and protein degradation of malignant cells. There are first hints that these drugs also sensitize tumor cells to chemotherapy, radiation, and NK cell-mediated cytotoxicity by enhanced expression of TRAIL and NKG2DLs. However, these pharmaceuticals may also impair NK cell function in a dose- and time-dependent manner. In summary, this review provides an update on the effects of different novel molecules on the immune system focusing NK cells. PMID:23316191

  16. Expansions of NK-like αβT cells with chronologic aging: novel lymphocyte effectors that compensate for functional deficits of conventional NK cells and T cells.

    PubMed

    Vallejo, Abbe N; Mueller, Robert G; Hamel, David L; Way, Amanda; Dvergsten, Jeffrey A; Griffin, Patricia; Newman, Anne B

    2011-07-01

    As the repertoire of αβT cell receptors (TCR) contracts with advancing age, there is an associated age-dependent accumulation of oligoclonal T cells expressing of a variety of receptors (NKR), normally expressed on natural killer (NK) cells. Evidences for differential regulation of expression of particular NKRs between T cells and NK cells suggest that NKR expression on T cells is physiologically programmed rather than a random event of the aging process. Experimental studies show NKRs on aged αβT cells may function either as independent receptors, and/or as costimulatory receptors to the TCR. Considering the reported deficits of conventional αβTCR-driven activation and also functional deficits of classical NK cells, NKR(+) αβT cells likely represent novel immune effectors that are capable of combining innate and adaptive functions. Inasmuch as immunity is a determinant of individual fitness, the type and density of NKRs could be important contributing factors to the wide heterogeneity of health characteristics of older adults, ranging from institutionalized frail elders who are unable to mount immune responses to functionally independent community-dwelling elders who exhibit protective immunity. Understanding the biology of NKR(+) αβT cells could lead to new avenues for age-specific intervention to improve protective immunity. PMID:20932941

  17. Maternal uterine NK cell-activating receptor KIR2DS1 enhances placentation.

    PubMed

    Xiong, Shiqiu; Sharkey, Andrew M; Kennedy, Philippa R; Gardner, Lucy; Farrell, Lydia E; Chazara, Olympe; Bauer, Julien; Hiby, Susan E; Colucci, Francesco; Moffett, Ashley

    2013-10-01

    Reduced trophoblast invasion and vascular conversion in decidua are thought to be the primary defect of common pregnancy disorders including preeclampsia and fetal growth restriction. Genetic studies suggest these conditions are linked to combinations of polymorphic killer cell Ig-like receptor (KIR) genes expressed by maternal decidual NK cells (dNK) and HLA-C genes expressed by fetal trophoblast. Inhibitory KIR2DL1 and activating KIR2DS1 both bind HLA-C2, but confer increased risk or protection from pregnancy disorders, respectively. The mechanisms underlying these genetic associations with opposing outcomes are unknown. We show that KIR2DS1 is highly expressed in dNK, stimulating strong activation of KIR2DS1+ dNK. We used microarrays to identify additional responses triggered by binding of KIR2DS1 or KIR2DL1 to HLA-C2 and found different responses in dNK coexpressing KIR2DS1 with KIR2DL1 compared with dNK only expressing KIR2DL1. Activation of KIR2DS1+ dNK by HLA-C2 stimulated production of soluble products including GM-CSF, detected by intracellular FACS and ELISA. We demonstrated that GM-CSF enhanced migration of primary trophoblast and JEG-3 trophoblast cells in vitro. These findings provide a molecular mechanism explaining how recognition of HLA class I molecules on fetal trophoblast by an activating KIR on maternal dNK may be beneficial for placentation. PMID:24091323

  18. TLR7/8 agonists promote NK-DC cross-talk to enhance NK cell anti-tumor effects in hepatocellular carcinoma.

    PubMed

    Zhou, Zhixia; Yu, Xin; Zhang, Jian; Tian, Zhigang; Zhang, Cai

    2015-12-28

    Hepatocellular carcinoma (HCC) is a common cancer worldwide and the third leading cause of cancer death. Immunotherapy is considered a promising treatment with the aim to boost or arouse HCC-specific immune responses. TLR7 and TLR8 agonists are effective immunomodulators and have been applied topically for the treatment of certain skin tumors and viral infections. Here, we explored the role of TLR7 and TLR8 agonists on the activation of dendritic cells (DCs) and natural killer (NK) cells. We demonstrated that these agonists could directly activate NK cells, promoting the maturation of immature DCs. Meanwhile, DCs also assisted in the function of NK cells, resulting in enhanced anti-tumor immune responses to HCC. Importantly, the combination therapy with NK cells stimulated with DCs and TLR7/8 agonist Gardiquimod (GDQ) significantly suppresses the growth of human HepG2 liver carcinoma xenografts. This study provides a new immunotherapeutic approach for human HCC based on DC-NK cross-talk and also suggests that TLR7 and/or TLR8 agonists, particularly GDQ, may serve as potent innate and adaptive immune response immunomodulators in tumor therapy. PMID:26433159

  19. PEG-IFN Alpha but Not Ribavirin Alters NK Cell Phenotype and Function in Patients with Chronic Hepatitis C

    PubMed Central

    Markova, Antoaneta A.; Mihm, Ulrike; Schlaphoff, Verena; Lunemann, Sebastian; Filmann, Natalie; Bremer, Birgit; Berg, Thomas; Sarrazin, Christoph; Zeuzem, Stefan; Manns, Michael P.; Cornberg, Markus; Herrmann, Eva; Wedemeyer, Heiner

    2014-01-01

    Background Ribavirin (RBV) remains part of several interferon-free treatment strategies even though its mechanisms of action are still not fully understood. One hypothesis is that RBV increases responsiveness to type I interferons. Pegylated Interferon alpha (PEG-IFNa) has recently been shown to alter natural killer (NK) cell function possibly contributing to control of hepatitis C virus (HCV) infection. However, the effects of ribavirin alone or in combination with IFNa on NK cells are unknown. Methods Extensive ex vivo phenotyping and functional analysis of NK cells from hepatitis C patients was performed during antiviral therapy. Patients were treated for 6 weeks with RBV monotherapy (n = 11), placebo (n = 13) or PEG-IFNa-2a alone (n = 6) followed by PEG-IFNa/RBV combination therapy. The effects of RBV and PEG-IFNa-2a on NK cells were also studied in vitro after co-culture with K562 or Huh7.5 cells. Results Ribavirin monotherapy had no obvious effects on NK cell phenotype or function, neither ex vivo in patients nor in vitro. In contrast, PEG-IFNa-2a therapy was associated with an increase of CD56bright cells and distinct changes in expression profiles leading to an activated NK cell phenotype, increased functionality and decline of terminally differentiated NK cells. Ribavirin combination therapy reduced some of the IFN effects. An activated NK cell phenotype during therapy was inversely correlated with HCV viral load. Conclusions PEG-IFNa activates NK cells possibly contributing to virological responses independently of RBV. The role of NK cells during future IFN-free combination therapies including RBV remains to be determined. PMID:24751903

  20. Inflammatory cytokine-mediated evasion of virus-induced tumors from NK cell control

    PubMed Central

    Mishra, Rabinarayan; Polic, Bojan; Welsh, Raymond M.; Szomolanyi-Tsuda, Eva

    2013-01-01

    Infections with DNA tumor viruses, including members of the polyomavirus family, often result in tumor formation in immune-deficient hosts. The complex control involved in antiviral and antitumor immune responses during these infections can be studied in murine polyomavirus (PyV)-infected mice as a model. We found that NK cells efficiently kill cells derived from PyV-induced salivary gland tumors in vitro in an NKG2D (effector cell) -RAE-1 (target cell) - dependent manner, but in T cell-deficient mice NK cells only delay but do not prevent the development of PyV-induced tumors. Here we show that the PyV-induced tumors have infiltrating functional NK cells. The freshly removed tumors, however, lack surface RAE-1 expression, and the tumor tissues produce soluble factors that down-regulate RAE-1. These factors include the pro-inflammatory cytokines IL-1α, IL-1β, IL-33, and TNF. Each of these cytokines down-regulate RAE-1 expression and susceptibility to NK cell mediated cytotoxicity. CD11b+F4/80+ macrophages infiltrating the PyV-induced tumors produce high amounts of IL-1β and TNF. Thus, our data suggest a new mechanism whereby inflammatory cytokines generated in the tumor environment lead to evasion of NK cell-mediated control of virus-induced tumors. PMID:23772039

  1. NK cells inhibit humoral immunity by reducing the abundance of CD4+ T follicular helper cells during a chronic virus infection

    PubMed Central

    Cook, Kevin D.; Kline, Hannah C.; Whitmire, Jason K.

    2015-01-01

    There is a need to understand better how to improve B cell responses and immunity to persisting virus infections, which often cause debilitating illness or death. People with chronic virus infection show evidence of improved virus control when there is a strong neutralizing antibody response, and conversely, B cell dysfunction is associated with higher viral loads. We showed previously that NK cells inhibit CD4+ and CD8+ T cell responses to disseminating LCMV infection and that depletion of NK cells attenuates chronic infection. Here, we examined the effect of NK cell depletion on B cell responses to LCMV infection in mice. Whereas mice infected acutely generated a peak level of antibody soon after the infection was resolved, mice infected chronically showed a continued increase in antibody levels that exceeded those after acute infection. We found that early NK cell depletion rapidly increased virus-specific antibody levels to chronic infection, and this effect depended on CD4+ T cells and was associated with elevated numbers of CXCR5+CD4+ TFH cells. However, the NK cell-depleted mice controlled the infection and by 1 mo pi, had lower TFH cell numbers and antibody levels compared with mice with sustained infection. Finally, we show that NK cell depletion improved antiviral CD8+ T cell responses only when B cells and virus-specific antibody were present. Our data indicate that NK cells diminish immunity to chronic infection, in part, by suppressing TFH cell and antibody responses. PMID:25986014

  2. IL-18-induced expression of high-affinity IL-2R on murine NK cells is essential for NK-cell IFN-γ production during murine Plasmodium yoelii infection.

    PubMed

    Stegmann, Kerstin A; De Souza, J Brian; Riley, Eleanor M

    2015-12-01

    Early production of pro-inflammatory cytokines, including IFN-γ, is essential for control of blood-stage malaria infections. We have shown that IFN-γ production can be induced among human natural killer (NK) cells by coculture with Plasmodium falciparum infected erythrocytes, but the importance of this response is unclear. To further explore the role of NK cells during malaria infection, we have characterized the NK-cell response of C57BL/6 mice during lethal (PyYM) or nonlethal (Py17XNL) P. yoelii infection. Ex vivo flow cytometry revealed that NK cells are activated within 24 h of Py17XNL blood-stage infection, expressing CD25 and producing IFN-γ; this response was blunted and delayed during PyYM infection. CD25 expression and IFN-γ production were highly correlated, suggesting a causal relationship between the two responses. Subsequent in vitro experiments revealed that IL-18 signaling is essential for induction of CD25 and synergizes with IL-12 to enhance CD25 expression on splenic NK cells. In accordance with this, Py17XNL-infected erythrocytes induced NK-cell CD25 expression and IFN-γ production in a manner that is completely IL-18- and partially IL-12-dependent, and IFN-γ production is enhanced by IL-2. These data suggest that IL-2 signaling via CD25 amplifies IL-18- and IL-12-mediated NK-cell activation during malaria infection. PMID:26420375

  3. Impaired NK-mediated regulation of T-cell activity in multiple sclerosis is reconstituted by IL-2 receptor modulation.

    PubMed

    Gross, Catharina C; Schulte-Mecklenbeck, Andreas; Rünzi, Anna; Kuhlmann, Tanja; Posevitz-Fejfár, Anita; Schwab, Nicholas; Schneider-Hohendorf, Tilman; Herich, Sebastian; Held, Kathrin; Konjević, Matea; Hartwig, Marvin; Dornmair, Klaus; Hohlfeld, Reinhard; Ziemssen, Tjalf; Klotz, Luisa; Meuth, Sven G; Wiendl, Heinz

    2016-05-24

    Multiple sclerosis (MS) is a chronic inflammatory autoimmune disease of the central nervous system (CNS) resulting from a breakdown in peripheral immune tolerance. Although a beneficial role of natural killer (NK)-cell immune-regulatory function has been proposed, it still needs to be elucidated whether NK cells are functionally impaired as part of the disease. We observed NK cells in active MS lesions in close proximity to T cells. In accordance with a higher migratory capacity across the blood-brain barrier, CD56(bright) NK cells represent the major intrathecal NK-cell subset in both MS patients and healthy individuals. Investigating the peripheral blood and cerebrospinal fluid of MS patients treated with natalizumab revealed that transmigration of this subset depends on the α4β1 integrin very late antigen (VLA)-4. Although no MS-related changes in the migratory capacity of NK cells were observed, NK cells derived from patients with MS exhibit a reduced cytolytic activity in response to antigen-activated CD4(+) T cells. Defective NK-mediated immune regulation in MS is mainly attributable to a CD4(+) T-cell evasion caused by an impaired DNAX accessory molecule (DNAM)-1/CD155 interaction. Both the expression of the activating NK-cell receptor DNAM-1, a genetic alteration consistently found in MS-association studies, and up-regulation of the receptor's ligand CD155 on CD4(+) T cells are reduced in MS. Therapeutic immune modulation of IL-2 receptor restores impaired immune regulation in MS by increasing the proportion of CD155-expressing CD4(+) T cells and the cytolytic activity of NK cells. PMID:27162345

  4. Mesenchymal stem cells derived from low risk acute lymphoblastic leukemia patients promote NK cell antitumor activity.

    PubMed

    Entrena, Ana; Varas, Alberto; Vázquez, Miriam; Melen, Gustavo J; Fernández-Sevilla, Lidia M; García-Castro, Javier; Ramírez, Manuel; Zapata, Agustín G; Vicente, Ángeles

    2015-07-28

    Mesenchymal stem cells (MSCs) are key components of the bone marrow microenvironment which contribute to the maintenance of the hematopoietic stem cell niche and exert immunoregulatory functions in innate and adaptive immunity. We analyze the immunobiology of MSCs derived from acute lymphoblastic leukemia (ALL) patients and their impact on NK cell function. In contrast to the inhibitory effects on the immune response exerted by MSCs from healthy donors (Healthy-MSCs), we demonstrate that MSCs derived from low/intermediate risk ALL patients at diagnosis (ALL-MSCs) promote an efficient NK cell response including cytokine production, phenotypic activation and most importantly, cytotoxicity. Longitudinal studies indicate that these immunostimulatory effects of ALL-MSCs are progressively attenuated. Healthy-MSCs adopt ALL-MSC-like immunomodulatory features when exposed to leukemia cells, acquiring the ability to stimulate NK cell antitumor function. The mechanisms underlying to these functional changes of ALL-MSCs include reduced production of soluble inhibitory factors, differential expression of costimulatory and coinhibitory molecules, increased expression of specific TLRs and Notch pathway activation. Collectively our findings indicate that, in response to leukemia cells, ALL-MSCs could mediate a host beneficial immunomodulatory effect by stimulating the antitumor innate immune response. PMID:25917077

  5. Bovine NK cells acquire cytotoxic activity and produce IFN-gamma after stimulation by Mycobacterium bovis BCG- or Babesia bovis-exposed splenic dendritic cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Early interactions of innate immune cell populations such as DC, monocytes/macrophages and NK cells, can affect the ability of the acquired immune response to control infection of intracellular microorganisms. In this study, we investigated the activation of bovine NK cells by CD13+ splenic DC or CD...

  6. Extranodal NK/T Cell Lymphoma Causing Cardiorespiratory Failure

    PubMed Central

    2016-01-01

    Extranodal NK/T cell lymphoma is an uncommon malignancy usually involving the sinonasal area. We report an unusual case of extranodal NK/T cell lymphoma diagnosed in a 62-year-old Caucasian male who died of progressive cardiorespiratory failure but had no clinically detectable upper respiratory system lesions. The initial diagnosis was made cytologically on a sample of pericardial fluid that contained neoplastic lymphoid cells. These cells were positive for CD2, cytoplasmic CD3, and Epstein-Barr virus and negative for CD56. The diagnosis was confirmed at the autopsy, which disclosed lymphoma infiltrates in the myocardium, lungs, stomach, and pancreas. The death was caused by heart and lung failure due to uncontrollable arrhythmia and respiratory insufficiency due to the lymphoma infiltrates. To the best of our knowledge, this is the first case of extranodal NK/T cell lymphoma presenting with cardiopulmonary failure. PMID:27493813

  7. Extranodal NK/T Cell Lymphoma Causing Cardiorespiratory Failure.

    PubMed

    Li, Yiting; Damjanov, Ivan

    2016-01-01

    Extranodal NK/T cell lymphoma is an uncommon malignancy usually involving the sinonasal area. We report an unusual case of extranodal NK/T cell lymphoma diagnosed in a 62-year-old Caucasian male who died of progressive cardiorespiratory failure but had no clinically detectable upper respiratory system lesions. The initial diagnosis was made cytologically on a sample of pericardial fluid that contained neoplastic lymphoid cells. These cells were positive for CD2, cytoplasmic CD3, and Epstein-Barr virus and negative for CD56. The diagnosis was confirmed at the autopsy, which disclosed lymphoma infiltrates in the myocardium, lungs, stomach, and pancreas. The death was caused by heart and lung failure due to uncontrollable arrhythmia and respiratory insufficiency due to the lymphoma infiltrates. To the best of our knowledge, this is the first case of extranodal NK/T cell lymphoma presenting with cardiopulmonary failure. PMID:27493813

  8. Regulatory T cells inhibit CD34+ cell differentiation into NK cells by blocking their proliferation.

    PubMed

    Pedroza-Pacheco, Isabela; Shah, Divya; Domogala, Anna; Luevano, Martha; Blundell, Michael; Jackson, Nicola; Thrasher, Adrian; Madrigal, Alejandro; Saudemont, Aurore

    2016-01-01

    Graft versus Host Disease (GvHD) remains one of the main complications after hematopoietic stem cell transplantation (HSCT). Due to their ability to suppress effector cells, regulatory T cells (Tregs) have been proposed as a cellular therapy to prevent GvHD, however they also inhibit the functions of natural killer (NK) cells, key effectors of the Graft versus Leukemia effect. In this study, we have explored whether a Tregs therapy will also impact on NK cell differentiation. Using an in vitro model of hematopoietic stem cell (HSC) differentiation into NK cells, we found that activated Tregs led to a 90% reduction in NK cell numbers when added at the time of commitment to the NK cell lineage. This effect was contact dependent and was reversible upon Tregs depletion. The few NK cells that developed in these cultures were mature and exhibited normal functions. Furthermore, adoptive transfer of activated Tregs in rag(-/-) γc(-/-) mice abrogated HSC differentiation into NK cells thus confirming our in vitro findings. Collectively, these results demonstrate for the first time that activated Tregs can inhibit NK cell differentiation from HSC under specific conditions. PMID:26915707

  9. Regulatory T cells inhibit CD34+ cell differentiation into NK cells by blocking their proliferation

    PubMed Central

    Pedroza-Pacheco, Isabela; Shah, Divya; Domogala, Anna; Luevano, Martha; Blundell, Michael; Jackson, Nicola; Thrasher, Adrian; Madrigal, Alejandro; Saudemont, Aurore

    2016-01-01

    Graft versus Host Disease (GvHD) remains one of the main complications after hematopoietic stem cell transplantation (HSCT). Due to their ability to suppress effector cells, regulatory T cells (Tregs) have been proposed as a cellular therapy to prevent GvHD, however they also inhibit the functions of natural killer (NK) cells, key effectors of the Graft versus Leukemia effect. In this study, we have explored whether a Tregs therapy will also impact on NK cell differentiation. Using an in vitro model of hematopoietic stem cell (HSC) differentiation into NK cells, we found that activated Tregs led to a 90% reduction in NK cell numbers when added at the time of commitment to the NK cell lineage. This effect was contact dependent and was reversible upon Tregs depletion. The few NK cells that developed in these cultures were mature and exhibited normal functions. Furthermore, adoptive transfer of activated Tregs in rag-/- γc-/- mice abrogated HSC differentiation into NK cells thus confirming our in vitro findings. Collectively, these results demonstrate for the first time that activated Tregs can inhibit NK cell differentiation from HSC under specific conditions. PMID:26915707

  10. IL-12–producing monocytes and HLA-E control HCMV-driven NKG2C+ NK cell expansion

    PubMed Central

    Rölle, Alexander; Pollmann, Julia; Ewen, Eva-Maria; Le, Vu Thuy Khanh; Halenius, Anne; Hengel, Hartmut; Cerwenka, Adelheid

    2014-01-01

    Human cytomegalovirus (HCMV) infection is the most common cause of congenital viral infections and a major source of morbidity and mortality after organ transplantation. NK cells are pivotal effector cells in the innate defense against CMV. Recently, hallmarks of adaptive responses, such as memory-like features, have been recognized in NK cells. HCMV infection elicits the expansion of an NK cell subset carrying an activating receptor heterodimer, comprising CD94 and NKG2C (CD94/NKG2C), a response that resembles the clonal expansion of adaptive immune cells. Here, we determined that expansion of this NKG2C+ subset and general NK cell recovery rely on signals derived from CD14+ monocytes. In a coculture system, a subset of CD14+ cells with inflammatory monocyte features produced IL-12 in response to HCMV-infected fibroblasts, and neutralization of IL-12 in this model substantially reduced CD25 upregulation and NKG2C+ subset expansion. Finally, blockade of CD94/NKG2C on NK cells or silencing of the cognate ligand HLA-E in infected fibroblasts greatly impaired expansion of NKG2C+ NK cells. Together, our results reveal that IL-12, CD14+ cells, and the CD94/NKG2C/HLA-E axis are critical for the expansion of NKG2C+ NK cells in response to HCMV infection. Moreover, strategies targeting the NKG2C+ NK cell subset have the potential to be exploited in NK cell–based intervention strategies against viral infections and cancer. PMID:25384219

  11. Natural Killer Cells for Cancer Immunotherapy: Pluripotent Stem Cells-Derived NK Cells as an Immunotherapeutic Perspective

    PubMed Central

    Eguizabal, Cristina; Zenarruzabeitia, Olatz; Monge, Jorge; Santos, Silvia; Vesga, Miguel Angel; Maruri, Natalia; Arrieta, Arantza; Riñón, Marta; Tamayo-Orbegozo, Estibaliz; Amo, Laura; Larrucea, Susana; Borrego, Francisco

    2014-01-01

    Natural killer (NK) cells play an essential role in the fight against tumor development. Over the last years, the progress made in the NK-cell biology field and in deciphering how NK-cell function is regulated, is driving efforts to utilize NK-cell-based immunotherapy as a promising approach for the treatment of malignant diseases. Therapies involving NK cells may be accomplished by activating and expanding endogenous NK cells by means of cytokine treatment or by transferring exogenous cells by adoptive cell therapy and/or by hematopoietic stem cell transplantation. NK cells that are suitable for adoptive cell therapy can be derived from different sources, including ex vivo expansion of autologous NK cells, unstimulated or expanded allogeneic NK cells from peripheral blood, derived from CD34+ hematopoietic progenitors from peripheral blood and umbilical cord blood, and NK-cell lines. Besides, genetically modified NK cells expressing chimeric antigen receptors or cytokines genes may also have a relevant future as therapeutic tools. Recently, it has been described the derivation of large numbers of functional and mature NK cells from pluripotent stem cells, both embryonic stem cells and induced pluripotent stem cells, which adds another tool to the expanding NK-cell-based cancer immunotherapy arsenal. PMID:25309538

  12. NK Cell Subgroups, Phenotype, and Functions After Autologous Stem Cell Transplantation

    PubMed Central

    Jacobs, Benedikt; Tognarelli, Sara; Poller, Kerstin; Bader, Peter; Mackensen, Andreas; Ullrich, Evelyn

    2015-01-01

    High-dose chemotherapy with consecutive autologous stem cell transplantation (autoSCT) is a well-established treatment option for patients suffering from malignant lymphoma or multiple myeloma. Natural killer (NK) cells are an important part of the immune surveillance, and their cell number after autoSCT is predictive for progression-free and overall survival. To improve knowledge about the role of NK cells after autoSCT, we investigated different NK cell subgroups, their phenotype, and their functions in patients treated with autoSCT. Directly after leukocyte regeneration (>1000 leukocytes/μl) following autoSCT, CD56++ NK cells were the major NK cell subset. Surprisingly, these cells showed unusually high surface expression levels of CD57 and killer Ig-like receptors (KIRs) compared to expression levels before or at later time points after autoSCT. Moreover, these NK cells strongly upregulated KIR2DL2/3/S2 and KIR3DL1, whereas KIR2DL1/S1 remained constant, indicating that this cell population arose from more immature NK cells instead of from activated mature ones. Remarkably, NK cells were already able to degranulate and produce IFN-γ and MIP-1β upon tumor interaction early after leukocyte regeneration. In conclusion, we describe an unusual upregulation of CD57 and KIRs on CD56++ NK cells shortly after autoSCT. Importantly, these NK cells were functionally competent upon tumor interaction at this early time point. PMID:26635797

  13. IL15 promotes growth and invasion of endometrial stromal cells and inhibits killing activity of NK cells in endometriosis.

    PubMed

    Yu, Jia-Jun; Sun, Hui-Ting; Zhang, Zhong-Fang; Shi, Ru-Xia; Liu, Li-Bing; Shang, Wen-Qing; Wei, Chun-Yan; Chang, Kai-Kai; Shao, Jun; Wang, Ming-Yan; Li, Ming-Qing

    2016-08-01

    Endometriosis (EMS) is associated with an abnormal immune response to endometrial cells, which can facilitate the implantation and proliferation of ectopic endometrial tissues. It has been reported that human endometrial stromal cells (ESCs) express interleukin (IL)15. The aim of our study was to elucidate whether or not IL15 regulates the cross talk between ESCs and natural killer (NK) cells in the endometriotic milieu and, if so, how this regulation occurs. The ESC behaviors in vitro were verified by Cell Counting Kit-8 (CCK-8), Annexin/PI, and Matrigel invasion assays, respectively. To imitate the local immune microenvironment, the co-culture system between ESCs and NK cells was constructed. The effect of IL15 on NK cells in the co-culture unit was investigated by flow cytometry (FCM). In this study, we found that ectopic endometrium from patients with EMS highly expressed IL15. Rapamycin, an autophagy inducer, decreased the level of IL15 receptors (i.e. IL15Rα and IL2Rβ). IL15 inhibits apoptosis and promotes the invasiveness, viability, and proliferation of ESCs. Meanwhile, a co-culture with ESCs led to a decrease in CD16 on NK cells. In the co-culture system, IL15 treatment downregulated the levels of Granzyme B and IFN-γ in CD16(+)NK cells, NKG2D in CD56(dim)CD16(-)NK cells, and NKP44 in CD56(bright)CD16(-)NK cells. On the one hand, these results indicated that IL15 derived from ESCs directly stimulates the growth and invasion of ESCs. On the other hand, IL15 may help the immune escape of ESCs by suppressing the cytotoxic activity of NK cells in the ectopic milieu, thereby facilitating the progression of EMS. PMID:27190213

  14. NK Cell Activation in Human Hantavirus Infection Explained by Virus-Induced IL-15/IL15Rα Expression

    PubMed Central

    Braun, Monika; Björkström, Niklas K.; Gupta, Shawon; Sundström, Karin; Ahlm, Clas; Klingström, Jonas; Ljunggren, Hans-Gustaf

    2014-01-01

    Clinical infection with hantaviruses cause two severe acute diseases, hemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome (HPS). These diseases are characterized by strong immune activation, increased vascular permeability, and up to 50% case-fatality rates. One prominent feature observed in clinical hantavirus infection is rapid expansion of natural killer (NK) cells in peripheral blood of affected individuals. We here describe an unusually high state of activation of such expanding NK cells in the acute phase of clinical Puumala hantavirus infection. Expanding NK cells expressed markedly increased levels of activating NK cell receptors and cytotoxic effector molecules. In search for possible mechanisms behind this NK cell activation, we observed virus-induced IL-15 and IL-15Rα on infected endothelial and epithelial cells. Hantavirus-infected cells were shown to strongly activate NK cells in a cell-cell contact-dependent way, and this response was blocked with anti-IL-15 antibodies. Surprisingly, the strength of the IL-15-dependent NK cell response was such that it led to killing of uninfected endothelial cells despite expression of normal levels of HLA class I. In contrast, hantavirus-infected cells were resistant to NK cell lysis, due to a combination of virus-induced increase in HLA class I expression levels and hantavirus-mediated inhibition of apoptosis induction. In summary, we here describe a possible mechanism explaining the massive NK cell activation and proliferation observed in HFRS patients caused by Puumala hantavirus infection. The results add further insights into mechanisms behind the immunopathogenesis of hantavirus infections in humans and identify new possible targets for intervention. PMID:25412359

  15. A Case of Aggressive NK/T-cell Lymphoma/Leukemia with Cutaneous Involvement in Adolescence

    PubMed Central

    Kim, Soo Ho; Ko, Woo Tae; Ha, Gyoung Yim; Kim, Jung Ran

    2008-01-01

    NK/T-cell lymphoma (NKTCL) is characterized by the expression of the NK-cell antigen CD56. Non-nasal NK/T-cell lymphomas are subdivided into primary cutaneous and 4 subtypes of secondary cutaneous lymphomas; nasal type, aggressive, blastic (blastoid), and other specific NK-like cell lymphoma. Aggressive NK/T-cell lymphoma/leukemia is a rare leukemic variant of nasal type NKTCL. We herein report a rare case of aggressive NK/T-cell lymphoma/leukemia with cutaneous involvement in adolescence. PMID:27303165

  16. Advances in clinical NK cell studies: Donor selection, manufacturing and quality control

    PubMed Central

    Koehl, U.; Kalberer, C.; Spanholtz, J.; Lee, D. A.; Miller, J. S.; Cooley, S.; Lowdell, M.; Uharek, L.; Klingemann, H.; Curti, A.; Leung, W.; Alici, E.

    2016-01-01

    ABSTRACT Natural killer (NK) cells are increasingly used in clinical studies in order to treat patients with various malignancies. The following review summarizes platform lectures and 2013–2015 consortium meetings on manufacturing and clinical use of NK cells in Europe and United States. A broad overview of recent pre-clinical and clinical results in NK cell therapies is provided based on unstimulated, cytokine-activated, as well as genetically engineered NK cells using chimeric antigen receptors (CAR). Differences in donor selection, manufacturing and quality control of NK cells for cancer immunotherapies are described and basic recommendations are outlined for harmonization in future NK cell studies. PMID:27141397

  17. Reduction of the CD16−CD56bright NK Cell Subset Precedes NK Cell Dysfunction in Prostate Cancer

    PubMed Central

    Koo, Kyo Chul; Shim, Doo Hee; Yang, Chang Mo; Lee, Saet-Byul; Kim, Shi Mun; Shin, Tae Young; Kim, Kwang Hyun; Yoon, Ho Geun; Rha, Koon Ho; Lee, Jae Myun; Hong, Sung Joon

    2013-01-01

    Background Natural cytotoxicity, mediated by natural killer (NK) cells plays an important role in the inhibition and elimination of malignant tumor cells. To investigate the immunoregulatory role of NK cells and their potential as diagnostic markers, NK cell activity (NKA) was analyzed in prostate cancer (PCa) patients with particular focus on NK cell subset distribution. Methods Prospective data of NKA and NK cell subset distribution patterns were measured from 51 patients initially diagnosed with PCa and 54 healthy controls. NKA was represented by IFN-γ levels after stimulation of the peripheral blood with Promoca®. To determine the distribution of NK cell subsets, PBMCs were stained with fluorochrome-conjugated monoclonal antibodies. Then, CD16+CD56dim and CD16−CD56bright cells gated on CD56+CD3− cells were analyzed using a flow-cytometer. Results NKA and the proportion of CD56bright cells were significantly lower in PCa patients compared to controls (430.9 pg/ml vs. 975.2 pg/ml and 2.3% vs. 3.8%, respectively; p<0.001). Both tended to gradually decrease according to cancer stage progression (p for trend = 0.001). A significantly higher CD56dim-to-CD56bright cell ratio was observed in PCa patients (41.8 vs. 30.3; p<0.001) along with a gradual increase according to cancer stage progression (p for trend = 0.001), implying a significant reduction of CD56bright cells in relation to the alteration of CD56dim cells. The sensitivity and the specificity of NKA regarding PCa detection were 72% and 74%, respectively (best cut-off value at 530.9 pg/ml, AUC = 0.786). Conclusions Reduction of CD56bright cells may precede NK cell dysfunction, leading to impaired cytotoxicity against PCa cells. These observations may explain one of the mechanisms behind NK cell dysfunction observed in PCa microenvironment and lend support to the development of future cancer immunotherapeutic strategies. PMID:24223759

  18. Enhancement of anti-leukemia activity of NK cells in vitro and in vivo by inhibition of leukemia cell-induced NK cell damage

    PubMed Central

    Arriga, Roberto; Caratelli, Sara; Coppola, Andrea; Spagnoli, Giulio Cesare; Venditti, Adriano; Amadori, Sergio; Lanzilli, Giulia; Lauro, Davide; Palomba, Patrizia; Sconocchia, Tommaso; Del Principe, Maria Ilaria; Maurillo, Luca; Buccisano, Francesco; Capuani, Barbara; Ferrone, Soldano; Sconocchia, Giuseppe

    2016-01-01

    Acute myeloid leukemia (AML) cells induce, in vitro, NK cell abnormalities (NKCAs) including apoptosis and activating receptor down-regulation. The potential negative impact of AML cells on the therapeutic efficacy of NK cell-based strategies prompted us to analyze the mechanisms underlying NKCAs and to develop approaches to protect NK cells from NKCAs. NKCA induction by the AML leukemia cells target a subpopulation of peripheral blood NK cells and is interleukin-2 independent but is abrogated by a long-term culture of NK (LTNK) cells at 37°C. LTNK cells displayed a significantly enhanced ability to damage AML cells in vitro and inhibited the subcutaneous growth of ML-2 cells grafted into CB17 SCID mice. Actinomycin D restored the susceptibility of LTNK cells to NKCAs while TAPI-0, a functional analog of the tissue inhibitor of metalloproteinase (TIMP) 3, inhibits ML-2 cell-induced NKCAs suggesting that the generation of NK cell resistance to NKCAs involves RNA transcription and metalloproteinase (MPP) inactivation. This conclusion is supported by the reduced susceptibility to AML cell-induced NKCAs of LTNK cells in which TIMP3 gene and protein are over-expressed. This information may contribute to the rational design of targeted strategies to enhance the efficacy of NK cell-based-immunotherapy of AML with haploidentical NK cells. PMID:26655503

  19. Extranodal NK/T-cell lymphoma, nasal type

    PubMed Central

    Parikh, Sachin R.; Krause, John R.

    2011-01-01

    We report a case of extranodal NK/T-cell lymphoma, nasal type, which is rare in the United States and Europe. It is more prevalent in Asians and Native Americans of Mexico, Central America, and South America. A 30-year-old Southeast Asian man with facial swelling, fever, and unintentional weight loss was found to have leukopenia and thrombocytopenia. He underwent endoscopic sinus surgery, which confirmed extranodal NK/T-cell lymphoma, nasal type, and a blood and bone marrow examination, which was negative for involvement but yielded the diagnosis of alpha-E thalassemia. The patient received chemotherapy, radiotherapy, and a stem cell transplant with 100% engraftment. PMID:21738302

  20. Effect of Spaceflight on the Functions of NK and LAK Cells

    NASA Technical Reports Server (NTRS)

    Pierson, Duane L.; Grimm, Elizabeth A.; Pierson, Duane L.; Paloski, W. H. (Technical Monitor)

    1999-01-01

    Spaceflight-associated stress alters some aspects of the human immune response. In this study, we determined the effects of 10 days aboard the Space Shuttle on the cytotoxic activity of NK and LAK cells. The subjects of this study were crewmembers of two 10-day shuttle flights. Ten-ml blood specimens were obtained from ten astronauts 10 days before launch, immediately after landing, and 3 days after landing. PBMCs were separated from the blood specimens and stored at -800 C. All PBMCs were thawed simultaneously, and the cytotoxic activities of NK and LAK cells were measured by a 4-hour Cr-51 release assay. K562 cells were used to assess NK-cell cytotoxicity. After 4 days of IL-2 activation, the LAK cell cytotoxic activity was determined using K562 and Daudi cells as the target cells. NK-cell cytotoxicity was decreased at landing (p less than 0.0005) in 9/10 astronauts, and in most cases recovered to preflight levels by 3 days after landing; NK-cell cytotoxicity was increased in one astronaut at landing. LAK cytotoxic activity against K562 cells was decreased at landing in 6/10 astronauts (p=0.018), and activity against Daudi cells was decreased in 7/10 astronauts (p=0.01). Phenotyping of PBMCs and LAK cells showed alterations in some surface markers and adhesion molecules (CD1 1 b, CD1 1 c, CD1 1 a, CD1 6, L-Selectin and CD3). Thus spaceflight leads to a decrease in the functions of NK and LAK cells in most astronauts.

  1. Infection-induced regulation of NK cells by macrophages and collagen at the lymph node subcapsular sinus

    PubMed Central

    Coombes, Janine L.; Han, Seong-Ji; van Rooijen, Nico; Raulet, David H.; Robey, Ellen A.

    2012-01-01

    Summary Infection leads to heightened activation of natural killer (NK) cells, a process that likely involves direct cell-to-cell contact, but how this occurs in vivo is poorly understood. We have used two-photon laser-scanning microscopy in conjunction with Toxoplasma gondii-mouse infection models to address this question. We found that NK cells accumulated in the subcapsular region of the lymph node following infection where they formed low motility contacts with collagen fibers and CD169+ macrophages. We provide evidence that interactions with collagen regulate NK cell migration, whereas CD169+ macrophages increase the activation state of NK cells. Interestingly, a subset of CD169+ macrophages that co-express the inflammatory monocyte marker Ly6C had the most potent ability to activate NK cells. Our data reveal pathways through which NK cell migration and function are regulated following infection, and identify an important accessory cell population for activation of NK cell responses in lymph nodes. PMID:22840403

  2. NK cell immunophenotypic and genotypic analysis of infants with severe respiratory syncytial virus infection.

    PubMed

    Noyola, Daniel E; Juárez-Vega, Guillermo; Monjarás-Ávila, César; Escalante-Padrón, Francisco; Rangel-Ramírez, Verónica; Cadena-Mota, Sandra; Monsiváis-Urenda, Adriana; García-Sepúlveda, Christian A; González-Amaro, Roberto

    2015-07-01

    Respiratory syncytial virus (RSV) is the leading cause of severe lower respiratory tract infection in infants. Reduced numbers of NK cells have been reported in infants with severe RSV infection; however, the precise role of NK cells during acute RSV infection is unclear. In this study the NK and T cell phenotypes, LILRB1 gene polymorphisms and KIR genotypes of infants hospitalized with RSV infection were analyzed. Compared to controls, infants with acute RSV infection showed a higher proportion of LILRB1+ T cells; in addition, a subgroup of infants with RSV infection showed an increase in LILRB1+ NK cells. No differences in NKG2C, NKG2A, or CD161 expression between RSV infected infants and controls were observed. LILRB1 genotype distribution of the rs3760860 A>G, and rs3760861 A>G single nucleotide polymorphisms differed between infants with RSV infection and healthy donors, whereas no differences in any of the KIR genes were observed. Our results suggest that LILRB1 participates in the pathogenesis of RSV infection. Further studies are needed to define the role of LILRB1+ NK in response to RSV and to confirm an association between LILRB1 polymorphisms and the risk of severe RSV infection. PMID:25988502

  3. Absence of circulating natural killer (NK) cells in a child with erythrophagocytic lymphohistiocytosis lacking NK cell activity

    SciTech Connect

    Kawai, H.; Komiyama, A.; Aoyama, K.; Miyagawa, Y.; Akabane, T.

    1988-06-01

    A 5-year-old girl who was diagnosed as having erythrophagocytic lymphohistiocytosis died at age 9 years. Peripheral lymphocytes from the patient persistently lacked natural killer (NK) cell activity during the 4-year observation period: the percent lysis values as measured by a 4-hr /sup 51/Cr release assay at a 40:1 effector:target ratio were below 1.0% against K562 and Molt-4 cells as compared with the normal lymphocyte value (mean +/- SD) of 46.2% +/- 5.8% and 43.9% +/- 6.7%, respectively. The patient's lymphocytes never developed NK cell activity by their incubation with target cells for longer time periods or by their stimulation with interferon-alpha, interleukin-2, or polyinosinic-polycytidilic acid. Single cell-in-agarose assay showed the absence of target-binding cells (TBCs): TBC numbers were below 0.3% as compared with the normal lymphocyte value of 8.1% +/- 1.3% (mean +/- SD). Flow cytometry showed a marked decrease in Leu-7+ cells (1.7%) and the absence of Leu-11+ cells (0.4%) in the peripheral blood. These results first demonstrate a case of erythrophagocytic lymphohistiocytosis in which there is the lack of NK cell activity due to the absence of circulating NK cells.

  4. Comparative analysis of NK-cell receptor expression and function across primate species

    PubMed Central

    Ugolotti, Elisabetta; De Maria, Andrea

    2010-01-01

    Natural killer (NK) cells are lymphoid effectors that are involved in the innate immune surveillance against infected and/or tumor cells. Their function is under the fine-tuning control of cell surface receptors that display either inhibitory or activating function and in healthy condition, mediate self-tolerance. It is known that inhibitory receptors are characterized by clonal and stochastic distribution and are extremely sensible to any modification, downregulation or loss of MHC class I surface expression that are induced in autologous cells upon viral infection or cancer transformation. This alteration of the MHC class I expression weakens the strength of the inhibitory receptor-induced interaction, thus resulting in a prompt triggering of NK cell function, which ends up in the inhibition of tumor progression and proliferation of pathogen-infected cells. Thus, the inhibitory function of NK cells is only one face of the coin, since NK-cell activation is controlled by different arrays of activating receptors that finally are involved in the induction of cytolysis and/or cytokine release. Interestingly, the inhibitory NK-cell receptors that are involved in dampening NK cell-mediated responses evolved during speciation in different, often structurally unrelated surface-expressed molecules, all using a conserved signaling pathway. In detail, during evolution, the inhibitory receptors that assure the recognition of MHC class I molecules, originate in, at least, three different ways. This ended up in multigene families showing marked structural divergences that coevolved in a convergent way with the availability of appropriate MHC ligand molecules. PMID:21487512

  5. NK-DC crosstalk controls the autopathogenic Th17 response through an innate IFN-γ-IL-27 axis.

    PubMed

    Chong, Wai Po; van Panhuys, Nicholas; Chen, Jun; Silver, Phyllis B; Jittayasothorn, Yingyos; Mattapallil, Mary J; Germain, Ronald N; Caspi, Rachel R

    2015-09-21

    IFN-γ is a pathogenic cytokine involved in inflammation. Paradoxically, its deficiency exacerbates experimental autoimmune encephalomyelitis, uveitis, and arthritis. Here, we demonstrate using IFN-γ(-/-) mice repleted with IFN-γ +/+: NK cells that innate production of IFN-γ from NK cells is necessary and sufficient to trigger an endogenous regulatory circuit that limits autoimmunity. After immunization, DCs recruited IFN-γ-producing NK cells to the draining lymph node and interacted with them in a CXCR3-dependent fashion. The interaction caused DCs to produce IL-27, which in turn enhanced IFN-γ production by NK cells, forming a self-amplifying positive feedback loop. IL-10, produced by the interacting cells themselves, was able to limit this process. The NK-DC-dependent IL-27 inhibited development of the adaptive pathogenic IL-17 response and induced IL-10-producing Tr1-like cells, which ameliorated disease in an IL-10-dependent manner. Our data reveal that an early NK-DC interaction controls the adaptive Th17 response and limits tissue-specific autoimmunity through an innate IFN-γ-IL-27 axis. PMID:26347474

  6. Opportunities and limitations of NK cells as adoptive therapy for malignant disease

    PubMed Central

    Davies, James O. J.; Stringaris, Kate; Barrett, John A.; Rezvani, Katayoun

    2014-01-01

    While NK cells can be readily generated for adoptive therapy with current techniques, their optimal application to treat malignant diseases requires an appreciation of the dynamic balance between signals that either synergise with, or antagonise each other. Individuals display wide differences in NK function which determine their therapeutic efficacy. The ability of NK cells to kill target cells or produce cytokines depends on the balance between signals from activating and inhibitory cell-surface receptors. The selection of NK cells with a predominant activating profile is critical for delivering successful antitumor activity. This can be achieved through selection of KIR mismatched NK donors and by using blocking molecules against inhibitory pathways. Optimum NK cytotoxicity may require licensing or priming with tumor cells. Recent discoveries in the molecular and cellular biology of NK cells inform in the design of new strategies, including adjuvant therapies, to maximise the cytotoxic potential of NK cells for adoptive transfer to treat human malignancies. PMID:24856895

  7. UV-inactivated HSV-1 potently activates NK cell killing of leukemic cells.

    PubMed

    Samudio, Ismael; Rezvani, Katayoun; Shaim, Hila; Hofs, Elyse; Ngom, Mor; Bu, Luke; Liu, Guoyu; Lee, Jason T C; Imren, Suzan; Lam, Vivian; Poon, Grace F T; Ghaedi, Maryam; Takei, Fumio; Humphries, Keith; Jia, William; Krystal, Gerald

    2016-05-26

    Herein we demonstrate that oncolytic herpes simplex virus-1 (HSV-1) potently activates human peripheral blood mononuclear cells (PBMCs) to lyse leukemic cell lines and primary acute myeloid leukemia samples, but not healthy allogeneic lymphocytes. Intriguingly, we found that UV light-inactivated HSV-1 (UV-HSV-1) is equally effective in promoting PBMC cytolysis of leukemic cells and is 1000- to 10 000-fold more potent at stimulating innate antileukemic responses than UV-inactivated cytomegalovirus, vesicular stomatitis virus, reovirus, or adenovirus. Mechanistically, UV-HSV-1 stimulates PBMC cytolysis of leukemic cells, partly via Toll-like receptor-2/protein kinase C/nuclear factor-κB signaling, and potently stimulates expression of CD69, degranulation, migration, and cytokine production in natural killer (NK) cells, suggesting that surface components of UV-HSV-1 directly activate NK cells. Importantly, UV-HSV-1 synergizes with interleukin-15 (IL-15) and IL-2 in inducing activation and cytolytic activity of NK cells. Additionally, UV-HSV-1 stimulates glycolysis and fatty acid oxidation-dependent oxygen consumption in NK cells, but only glycolysis is required for their enhanced antileukemic activity. Last, we demonstrate that T cell-depleted human PBMCs exposed to UV-HSV-1 provide a survival benefit in a murine xenograft model of human acute myeloid leukemia (AML). Taken together, our results support the preclinical development of UV-HSV-1 as an adjuvant, alone or in combination with IL-15, for allogeneic donor mononuclear cell infusions to treat AML. PMID:26941401

  8. Functional NK cell repertoires are maintained through IL-2Rα and FasL

    PubMed Central

    Felices, Martin; Lenvik, Todd R.; Ankarlo, Dave E.M.; Foley, Bree; Curtsinger, Julie; Luo, Xianghua; Blazar, Bruce R.; Anderson, Stephen K.; Miller, Jeffrey S.

    2014-01-01

    Acquisition of a functional natural killer (NK) cell repertoire, known as education or licensing, is a complex process mediated through inhibitory receptors that recognize self. We found that NK cells containing self-killer immunoglobulin-like receptors (KIR) for cognate HLA-ligand in vivo were less susceptible to apoptosis. In vitro IL-15 withdrawal showed that uneducated NK cells upregulated Bim and Fas. Conversely, educated NK cells upregulated FasL under these conditions. Induction of cell death and Bim expression on uneducated cells correlated with increased IL-2Rα expression. Overexpression and knockdown studies showed that higher IL-2Rα limits NK cell survival in a novel manner that is independent from the role of IL-2 in activation induced cell death (AICD). To study the role of FasL in induction of IL-2Rαhi NK cell death, a co-culture assay with FasL blocking antibodies was used. IL-15 withdrawal led to FasL dependent killing of IL-2Rαhi NK cells by more educated IL-2Rαlo NK cells. Finally, CMV reactivation induces a potent long-lasting population of licensed NK cells with enhanced survival. These findings show education dependent NK cell survival advantages and killing of uneducated NK result in the maintenance of a functional repertoire, which may be manipulated to exploit NK cells for cancer immunotherapy. PMID:24634493

  9. Binding of Excreted and/or Secreted Products of Adult Hookworms to Human NK Cells in Necator americanus-Infected Individuals from Brazil▿

    PubMed Central

    Teixeira-Carvalho, Andréa; Fujiwara, Ricardo T.; Stemmy, Erik J.; Olive, Denise; Damsker, Jesse M.; Loukas, Alex; Corrêa-Oliveira, Rodrigo; Constant, Stephanie L.; Bethony, Jeffrey M.

    2008-01-01

    The impact of the interaction between excreted and/or secreted (ES) Necator americanus products and NK cells from Necator-infected individuals was analyzed. We investigated the binding of ES products to NK cells, the expression of NK cell receptors (CD56, CD159a/NKG2A, CD314/NKG2D, CD335/NKp46, and KLRF1/NKp80), the frequency of gamma interferon (IFN-γ)-producing NK cells after whole-blood in vitro stimulation, and the capacity of N. americanus ES products to induce NK cell chemotaxis. In contrast to those from noninfected individuals, NK cells from Necator-infected individuals demonstrated no binding with N. americanus ES products. This phenomenon was not due to alterations in NK cell receptor expression in infected subjects and could not be reproduced with NK cells from uninfected individuals by incubation with immunoregulatory cytokines (interleukin-10/transforming growth factor β). Further, we found that a significantly greater percentage of NK cells from infected subjects than NK cells from uninfected individuals spontaneously produced IFN-γ upon ex vivo culture. Our findings support a model whereby NK cells from Necator-infected individuals may interact with ES products, making these cells refractory to binding with exogenous ES products. During N. americanus infection, human NK cells are attracted to the site of infection by chemotactic ES products produced by adult Necator worms in the gut mucosa. Binding of ES products causes the NK cells to become activated and secrete IFN-γ locally, thereby contributing to the adult hookworm's ability to evade host immune responses. PMID:18838519

  10. UV-inactivated HSV-1 potently activates NK cell killing of leukemic cells

    PubMed Central

    Samudio, Ismael; Rezvani, Katayoun; Shaim, Hila; Hofs, Elyse; Ngom, Mor; Bu, Luke; Liu, Guoyu; Lee, Jason T. C.; Imren, Suzan; Lam, Vivian; Poon, Grace F. T.; Ghaedi, Maryam; Takei, Fumio; Humphries, Keith; Jia, William

    2016-01-01

    Herein we demonstrate that oncolytic herpes simplex virus-1 (HSV-1) potently activates human peripheral blood mononuclear cells (PBMCs) to lyse leukemic cell lines and primary acute myeloid leukemia samples, but not healthy allogeneic lymphocytes. Intriguingly, we found that UV light–inactivated HSV-1 (UV-HSV-1) is equally effective in promoting PBMC cytolysis of leukemic cells and is 1000- to 10 000-fold more potent at stimulating innate antileukemic responses than UV-inactivated cytomegalovirus, vesicular stomatitis virus, reovirus, or adenovirus. Mechanistically, UV-HSV-1 stimulates PBMC cytolysis of leukemic cells, partly via Toll-like receptor-2/protein kinase C/nuclear factor-κB signaling, and potently stimulates expression of CD69, degranulation, migration, and cytokine production in natural killer (NK) cells, suggesting that surface components of UV-HSV-1 directly activate NK cells. Importantly, UV-HSV-1 synergizes with interleukin-15 (IL-15) and IL-2 in inducing activation and cytolytic activity of NK cells. Additionally, UV-HSV-1 stimulates glycolysis and fatty acid oxidation–dependent oxygen consumption in NK cells, but only glycolysis is required for their enhanced antileukemic activity. Last, we demonstrate that T cell–depleted human PBMCs exposed to UV-HSV-1 provide a survival benefit in a murine xenograft model of human acute myeloid leukemia (AML). Taken together, our results support the preclinical development of UV-HSV-1 as an adjuvant, alone or in combination with IL-15, for allogeneic donor mononuclear cell infusions to treat AML. PMID:26941401