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Sample records for activated killer cells

  1. Viral Evasion of Natural Killer Cell Activation

    PubMed Central

    Ma, Yi; Li, Xiaojuan; Kuang, Ersheng

    2016-01-01

    Natural killer (NK) cells play a key role in antiviral innate defenses because of their abilities to kill infected cells and secrete regulatory cytokines. Additionally, NK cells exhibit adaptive memory-like antigen-specific responses, which represent a novel antiviral NK cell defense mechanism. Viruses have evolved various strategies to evade the recognition and destruction by NK cells through the downregulation of the NK cell activating receptors. Here, we review the recent findings on viral evasion of NK cells via the impairment of NK cell-activating receptors and ligands, which provide new insights on the relationship between NK cells and viral actions during persistent viral infections. PMID:27077876

  2. Viral Evasion of Natural Killer Cell Activation.

    PubMed

    Ma, Yi; Li, Xiaojuan; Kuang, Ersheng

    2016-04-12

    Natural killer (NK) cells play a key role in antiviral innate defenses because of their abilities to kill infected cells and secrete regulatory cytokines. Additionally, NK cells exhibit adaptive memory-like antigen-specific responses, which represent a novel antiviral NK cell defense mechanism. Viruses have evolved various strategies to evade the recognition and destruction by NK cells through the downregulation of the NK cell activating receptors. Here, we review the recent findings on viral evasion of NK cells via the impairment of NK cell-activating receptors and ligands, which provide new insights on the relationship between NK cells and viral actions during persistent viral infections.

  3. Suppression of newborn natural killer cell activity by prostaglandin E2

    SciTech Connect

    Milch, P.O.; Salvatore, W.; Luft, B.; Baker, D.A.

    1988-10-01

    The effect of prostaglandin E2 on natural killer cell activity of cord blood was examined. Natural killer cell activity, determined by chromium 51 release, was significantly reduced after prostaglandin E2 (1 microgram/ml) treatment. Prostaglandin E2 has been found to enhance the cellular spread of herpesvirus. Thus prostaglandins may enhance viral infections indirectly by suppressing natural killer cell activity.

  4. Antibacterial activity of human natural killer cells

    PubMed Central

    1989-01-01

    The in vitro effects of human NK cells on viability of Gram-negative and Gram-positive bacteria was investigated. PBLs depleted of glass- adherent cells showed a significant antibacterial activity that was increased as the concentration of NK cells became higher. Leu-11- enriched cells exhibited the most efficient bactericidal activity. Stimulation of NK cells with staphylococcal enterotoxin B for 16 h produced a significant increase in the antibacterial activity of all NK cells tested. The antibacterial activity of monocyte-depleted cells and Leu-11-enriched cells was also enhanced after culturing in vitro for 16- 24 h without exogenous cytokines. Dependence of the antibacterial activity on the presence of serum in the culture medium was not found. Ultrastructural studies revealed close contact between NK cell membranes and bacteria, no evidence of phagocytosis, and extracellular bacterial ghosts, after incubation at 37 degrees C. Supernatants from purified NK cells exhibited potent bactericidal activity with kinetics and target specificity similar to that of effector cells. These results document the potent antibacterial activity of purified NK cells and suggest an extracellular mechanism of killing. PMID:2642532

  5. Radiosensitivity of human natural killer cells: Binding and cytotoxic activities of natural killer cell subsets

    SciTech Connect

    Rana, R.; Vitale, M.; Mazzotti, G.; Manzoli, L.; Papa, S. )

    1990-10-01

    The sensitivity of human natural killer (NK) cell activities (both binding and killing) after exposure of peripheral blood mononuclear cells to different doses of gamma radiation was studied. A panel of monoclonal antibodies was used to identify the NK and T-lymphocyte subsets and to evaluate their radiosensitivity. Peripheral blood mononuclear cells were irradiated with low (2-6 Gy) and high (10-30 Gy) doses and NK cell binding and cytotoxic activity against K562 target cells were studied after 3 h and 48 h in culture. The primary damage to NK cell activity was identified at the postbinding level and affected mainly the lytic machinery. After 48 h culture postirradiation, an overall depression of cytotoxic activity was observed, but ionizing radiation produced either a selection of the more cytotoxic NK cell subsets, which therefore might be considered more resistant to radiation damage than the less cytotoxic NK cells, or a long-term stimulation of cytotoxic activity in surviving cells.

  6. Effect of different levels of alcohol consumption on natural killer and lymphokine activated killer cells

    SciTech Connect

    Klassen, L.W.; DeVasure, J.M.; Lemley-Gillespie, S.D.; Thiele, G.M. Omaha VA Hospital, NE )

    1991-03-11

    The effect of alcohol consumption on natural killer (NK) cell activity is controversial as both increased and decreased levels have been reported. It was the purpose of this study to determine the effects of feeding BDF1 mice different levels of alcohol on NK and lymphokine activated killer (LAK) cell activity. After four-six weeks of chronic alcohol feeding, mice were sacrificed, spleen cells obtained and assayed for NK and IL-2 boosted NK activity against YAC-1 cells in a traditional {sup 51}chromium release assay. Cells were also cultured in the presence of IL-2 for five days and tested for cytolytic activity using P815 cells as targets. Cells from each group were passed over a nylon wool column and the adherent (AD) and nonadherent (NAD) populations collected and tested as above. Increased NK, 24 hour IL-2 boosted NK and 5 day LAK activity were observed only in the spleen cells obtained from mice on 20% alcohol. Also, NAD populations had a 2-4 fold higher lytic unit values (LU{sub 20}) at all levels of alcohol consumption and in all assays, as compared with the unseparated spleen cells. Analysis of cell surface markers on these three populations of cells show that there were differences in MAC-2, Asialo GM-1, Thy 1.2, B220 and NK 1.1 that may correlate with the differences observed in the cytolytic assays. These data suggest that different levels of alcohol affect the cytolytic activity of NK and LAK cells and may result from alterations in the cell subset populations.

  7. Reduced killer cell activity of lymphocytes from patients with asbestosis.

    PubMed Central

    Kubota, M; Kagamimori, S; Yokoyama, K; Okada, A

    1985-01-01

    Immunological abnormalities in 30 patients with asbestosis were investigated by examining the cytoxicity of natural killer (NK) cells and antibody dependent cellular cytotoxicity by killer (K) cells from peripheral blood lymphocytes; the effects of interferon on NK activity was also examined. Fifteen men and 15 women (mean age 58; range 40-72) with asbestosis but who were free of complications such as tuberculosis, carcinoma, or steroid treatment were the subjects for study. There were nine cases of type 1, 19 cases of type 2, and two cases of type 3 disease as described in the ILO classification of pneumoconiosis. They were all textile workers with a mean duration of 18 years (3-40 years) since first exposure to chrysotile. Controls matched for age and sex were selected from a population without occupational exposure to asbestos. The activity of the NK and K cells in patients with asbestosis was significantly lower than in the control group, but the populations of NK and K cells in the peripheral blood lymphocytes were not significantly different in the two groups. An in vitro experiment showed that the increase in the cytotoxicity of the NK cell after treatment with interferon-alpha was significantly lower in the subjects than in the controls. These results indicate that one of the defence mechanisms in relation to cancer is deficient in patients with asbestosis. PMID:3978049

  8. The DNA methylation profile of activated human natural killer cells.

    PubMed

    Wiencke, John K; Butler, Rondi; Hsuang, George; Eliot, Melissa; Kim, Stephanie; Sepulveda, Manuel A; Siegel, Derick; Houseman, E Andres; Kelsey, Karl T

    2016-05-01

    Natural killer (NK) cells are now recognized to exhibit characteristics akin to cells of the adaptive immune system. The generation of adaptive memory is linked to epigenetic reprogramming including alterations in DNA methylation. The study herein found reproducible genome wide DNA methylation changes associated with human NK cell activation. Activation led predominately to CpG hypomethylation (81% of significant loci). Bioinformatics analysis confirmed that non-coding and gene-associated differentially methylated sites (DMS) are enriched for immune related functions (i.e., immune cell activation). Known DNA methylation-regulated immune loci were also identified in activated NK cells (e.g., TNFA, LTA, IL13, CSF2). Twenty-one loci were designated high priority and further investigated as potential markers of NK activation. BHLHE40 was identified as a viable candidate for which a droplet digital PCR assay for demethylation was developed. The assay revealed high demethylation in activated NK cells and low demethylation in naïve NK, T- and B-cells. We conclude the NK cell methylome is plastic with potential for remodeling. The differentially methylated region signature of activated NKs revealed similarities with T cell activation, but also provided unique biomarker candidates of NK activation, which could be useful in epigenome-wide association studies to interrogate the role of NK subtypes in global methylation changes associated with exposures and/or disease states. PMID:26967308

  9. Utilizing Chimeric Antigen Receptors to Direct Natural Killer Cell Activity

    PubMed Central

    Hermanson, David L.; Kaufman, Dan S.

    2015-01-01

    Natural killer (NK) cells represent an attractive lymphocyte population for cancer immunotherapy due to their ability to lyse tumor targets without prior sensitization and without need for human leukocyte antigens-matching. Chimeric antigen receptors (CARs) are able to enhance lymphocyte targeting and activation toward diverse malignancies. CARs consist of an external recognition domain (typically a small chain variable fragment) directed at a specific tumor antigen that is linked with one or more intracellular signaling domains that mediate lymphocyte activation. Most CAR studies have focused on their expression in T cells. However, use of CARs in NK cells is starting to gain traction because they provide a method to redirect these cells more specifically to target refractory cancers. CAR-mediated anti-tumor activity has been demonstrated using NK cell lines, as well as NK cells isolated from peripheral blood, and NK cells produced from human pluripotent stem cells. This review will outline the CAR constructs that have been reported in NK cells with a focus on comparing the use of different signaling domains in combination with other co-activating domains. PMID:25972867

  10. Transcription factor Runx3 regulates interleukin-15-dependent natural killer cell activation.

    PubMed

    Levanon, Ditsa; Negreanu, Varda; Lotem, Joseph; Bone, Karen Rae; Brenner, Ori; Leshkowitz, Dena; Groner, Yoram

    2014-03-01

    Natural killer cells belong to the family of innate lymphoid cells comprising the frontline defense against infected and transformed cells. Development and activation of natural killer cells is highly dependent on interleukin-15 signaling. However, very little is known about the transcription program driving this process. The transcription factor Runx3 is highly expressed in natural killer cells, but its function in these cells is largely unknown. We show that loss of Runx3 impaired interleukin-15-dependent accumulation of mature natural killer cells in vivo and under culture conditions and pregnant Runx3(-/-) mice completely lack the unique population of interleukin-15-dependent uterine natural killer cells. Combined chromatin immunoprecipitation sequencing and differential gene expression analysis of wild-type versus Runx3-deficient in vivo activated splenic natural killer cells revealed that Runx3 cooperates with ETS and T-box transcription factors to drive the interleukin-15-mediated transcription program during activation of these cells. Runx3 functions as a nuclear regulator during interleukin-15-dependent activation of natural killer cells by regulating the expression of genes involved in proliferation, maturation, and migration. Similar studies with additional transcription factors will allow the construction of a more detailed transcriptional network that controls natural killer cell development and function.

  11. Stressin and natural killer cell activity in professional soldiers.

    PubMed

    Lauc, G; Dabelić; Dumić, J; Flögel, M

    1998-06-30

    Chronic stress causes multiple biochemical and physiological changes in the human organism. Recently we have identified stressin, a human serum glycoprotein that was significantly increased in sera of prisoners released from Serbian concentration camps. To eliminate malnutrition and maltreatment as possible causes for the increased stressin concentration, we have analyzed stressin in sera of 40 professional soldiers after involvement in major military activity and compared it to stressin in sera of 20 control individuals. As expected, the sera of professional soldiers contained more than 2.2 times higher concentrations of stressin than control sera. It is interesting that, contrary to expectations, the natural killer cell activity of professional soldiers was normal or even increased. We hypothesize that this might be an effect of winning the war that could have, at least temporarily, erased the immunosuppressive effects of stress.

  12. Effect of spaceflight on natural killer cell activity

    NASA Technical Reports Server (NTRS)

    Rykova, Marina P.; Sonnenfeld, Gerald; Lesniak, A. T.; Taylor, Gerald R.; Meshkov, Dimitrii O.; Mandel, Adrian D.; Medvedev, Andrei E.; Berry, Wallace D.; Fuchs, Boris B.; Konstantinova, Irina V.

    1992-01-01

    The effects of spaceflight on immune cell function were determined in rats flown on Cosmos 2044. Control groups included vivarium, synchronous, and antiorthostatically suspended rats. The ability of natural killer cells to lyse two different target cell lines was determined. Spleen and bone marrow cells obtained from flight rats showed significantly inhibited cytotoxicity for YAC-1 target cells compared with cells from synchronous control rats. This could have been due to exposure of the rats to microgravity. Antiorthostatic suspension did not affect the level of cytotoxicity from spleen cells of suspended rats for YAC-1 cells. On the other hand, cells from rats flown in space showed no significant differences from vivarium and synchronous control rats in cytotoxicity for K-562 target cells. Binding of natural killer cells to K-562 target cells was unaffected by spaceflight. Antiorthostatic suspension resulted in higher levels of cytotoxicity from spleen cells for Cr-51-labeled K-562 cells. The results indicate differential effects of spaceflight on function of natural killer cells. This shows that spaceflight has selective effects on the immune response.

  13. HPV vaccine stimulates cytotoxic activity of killer dendritic cells and natural killer cells against HPV-positive tumour cells

    PubMed Central

    Van den Bergh, Johan M J; Guerti, Khadija; Willemen, Yannick; Lion, Eva; Cools, Nathalie; Goossens, Herman; Vorsters, Alex; Van Tendeloo, Viggo F I; Anguille, Sébastien; Van Damme, Pierre; Smits, Evelien L J M

    2014-01-01

    Cervarix™ is approved as a preventive vaccine against infection with the human papillomavirus (HPV) strains 16 and 18, which are causally related to the development of cervical cancer. We are the first to investigate in vitro the effects of this HPV vaccine on interleukin (IL)-15 dendritic cells (DC) as proxy of a naturally occurring subset of blood DC, and natural killer (NK) cells, two innate immune cell types that play an important role in antitumour immunity. Our results show that exposure of IL-15 DC to the HPV vaccine results in increased expression of phenotypic maturation markers, pro-inflammatory cytokine production and cytotoxic activity against HPV-positive tumour cells. These effects are mediated by the vaccine adjuvant, partly through Toll-like receptor 4 activation. Next, we demonstrate that vaccine-exposed IL-15 DC in turn induce phenotypic activation of NK cells, resulting in a synergistic cytotoxic action against HPV-infected tumour cells. Our study thus identifies a novel mode of action of the HPV vaccine in boosting innate immunity, including killing of HPV-infected cells by DC and NK cells. PMID:24979331

  14. Acquisition of enhanced natural killer cell activity under anesthesia.

    PubMed

    Hsueh, C M; Lorden, J F; Hiramoto, R N; Ghanta, V K

    1992-01-01

    An increase in natural killer (NK) cell activity can be conditioned with a one trial learning paradigm to demonstrate the interaction between the central nervous system (CNS) and the immune system. In order to demonstrate learning possibilities during 'non-conscious' state, mice were anesthetized with a ketamin/rompun mixture and underwent one trial learning with odor cue as the conditioned stimulus (CS) preceding the unconditioned stimulus (US). The results indicated that mice that were exposed to camphor odor cue under the influence of anesthesia can associate the signal with the poly I:C unconditioned stimulus and were able to recall the conditioned response upon reexposure to the CS. Secondly, the conditioned association made in a conscious state can be recalled by exposure to the same olfactory odor cue in a 'non-conscious' state. The increase in the conditioned change in NK cell activity of both situations was significantly higher than the control group. The results demonstrate that learning can take place and the learned response can be recalled under the reduced awareness caused by anesthesia. The findings we report are unusual and novel in that they demonstrate that the CNS can learn new associations under conditions where the host is apparently unaware of the signals being linked. Anesthesia combined with the long interstimulus interval indicates that certain neuronal pathways in the CNS are receptive to second signals (elicited by the US) even when the second signal is separated by one day. This means the conditioned learning of a physiological response can take place unconsciously at a separate level and under situations where the host is totally unaware of the events which the brain is processing and linking as incoming information.

  15. Immunobiology of natural killer cells

    SciTech Connect

    Lotzova, E.; Herberman, R.B.

    1986-01-01

    This book combines research from many disciplines into a review of natural killer (NK) cell-mediated immunity in humans and experimental animal system. Topics for the volumes include: Volume I: Assays for NK Cell Cytotoxicity; Their Values and Pitfalls. Separation and Characterization of Phenotypically Distinct Subsets of NK Cells. Ultrastructure and Cytochemistry of the Human Large Granular Lymphocytes. Phylogeny and Ontogeny of NK Cells. Tissue and Organ distribution of NK Cells. Genetic Control of NK Cell Activity in Rodents. Phenotype, Functional Heterogeneity, and Lineage of Natural Killer Cells. Target Cell Structures, Recognition Sites, and the Mechanism of NK Cytotoxicity. Natural Killer Cytotoxic Factors (NKCF) Role in Cell-Mediated Cytotoxicity. Characteristics of Cultured NK Cells. Lectin-Dependent Killer Cells. MLC-Induced Cytotoxicity as a Model for the Development and Regulation of NK Cytotoxicity. LGL Lymphoproliferative Diseases in Man and Experimental Animals: The Characteristics of These Cells and Their Potential Experimental Uses. Index.

  16. Methylprednisolone pulse therapy induced fall in natural killer cell activity in rheumatoid arthritis.

    PubMed

    Pedersen, B K; Beyer, J M; Rasmussen, A; Klarlund, K; Pedersen, B N; Helin, P

    1984-10-01

    Natural killer (NK) cell activity was studied in 8 patients with classic or definite rheumatoid arthritis (RA) by investigating the killing of K 562 cells by peripheral blood lymphocytes before, during, and after intravenous methylprednisolone pulse therapy (MPPT). MPPT produced a considerable fall in NK activity and after 3 months NK activity was less than half that before MPPT. PMID:6516854

  17. Cytotoxic activity of allogeneic natural killer cells on U251 glioma cells in vitro.

    PubMed

    Guo, Meng; Wu, Tingting; Wan, Lixin

    2016-07-01

    The present study aimed to observe the cytotoxic activity of allogeneic natural killer (NK) cells on U251 glioma cells and to investigate their mechanism of action to establish an effective treatment strategy for neuroglioma. Cell survival curves, colony formation assays and karyotype analysis were performed to investigate the characteristics of U251 glioma cells. The present study demonstrated that natural killer group 2, member D (NKG2D)‑major histocompatibility complex class I‑related chain A/B (MICA/B) interactions contributed to the cytotoxic effect of NK cells on K562 and U251 cells. In antibody‑blocking assays to inhibit NKG2D ligands, the cytotoxic activity was not completely attenuated, which suggested that other signaling pathways contribute to the cytotoxic activity of NK cells on tumor cells in addition to the NKG2D‑mediated activity. The present study identified that the expression levels of NKG2D ligands on the surface of target cells influenced the strength of the NK cell immune response. Furthermore, allogeneic NK cells were observed to kill glioma cells in vitro, and this anticancer activity is associated with the rate of NKG2D expression on the surface of glioma cells.

  18. Natural killer cell activity in cigarette smokers and asbestos workers

    SciTech Connect

    Ginns, L.C.; Ryu, J.H.; Rogol, P.R.; Sprince, N.L.; Oliver, L.C.; Larsson, C.J.

    1985-06-01

    In order to evaluate the effects of cigarette smoking and asbestos exposure on cellular immunity, the authors tested a group of cigarette smokers and asbestos workers for natural killer (NK) activity in the peripheral blood. The mean NK activity in cigarette smokers was lower than in normal subjects (13.7 +/- 1.6 versus 29.0 +/- 3%; p less than 0.05). As a group, the mean NK activity for the asbestos-exposed group was also reduced compared with that of the nonsmoking control group (22.6 +/- 3.2%; p less than 0.05). When divided according to the smoking status, the asbestos workers who were nonsmokers or ex-smokers showed similar decreases in NK activity compared with normal subjects (19.5 +/- 6.2 and 21.2 +/- 4.5%, respectively; p less than 0.05). A subgroup of asbestos-exposed subjects who currently smoked showed no decrease in NK activity. The data show that NK activity is reduced in the peripheral blood of cigarette smokers and asbestos workers. The relatively normal NK activity found in asbestos workers who also smoked is unexplained. Impairment of NK activity is a potential mechanism for the increased incidence of infection and cancer in smokers and neoplasia in asbestos workers.

  19. Enhanced lymphokine-activated killer cell activity by an immunomodulator, Roquinimex.

    PubMed Central

    Vaz, F.; Silva, M. R.; Ascensao, J. L.

    1995-01-01

    Roquinimex (Roq) is an immunomodulator known to stimulate cellular immune responses. It is currently used for immunotherapy after bone marrow transplantation (BMT). One of the major features of this compound is an enhancement of natural killer (NK) cell activity and numbers. We studied the in vitro effect of Roq on human peripheral blood NK and adherent lymphokine-activated killer cell (ALAK) activities. In cultures supplemented with recombinant interleukin 2 (rIL-2) (1000 U ml-1) and Roq a significant increase in NK and LAK function was observed without a parallel increase in cell numbers. We also examined the generation of NK cells from human bone marrow (BM) immature progenitors, obtained by purging with 4-hydroperoxycyclophosphamide (4HC). NK cell numbers and activity were both increased when cultures with rIL-2 (10 U ml-1) were supplemented with Roq. These results confirm findings obtained in vivo and in vitro in the murine system and suggest that Roq is an active agent on these lymphoid populations. These properties and good tolerability make Roq an attractive tool for immunotherapy. PMID:8519666

  20. Bone marrow mesenchymal stem cells suppressing activation of allogeneic cytokine-induced killer/natural killer cells either by direct or indirect interaction.

    PubMed

    Li, Yang; Qu, Yu H; Wu, Yan F; Liu, Ling; Lin, Xiang H; Huang, Ke; Wei, Jing

    2015-04-01

    Bone marrow mesenchymal stem cells (MSC) were recently found to be associated with some special immunological characteristics, the immunoregulatory effect of MSC was dose-dependent. Low amount of MSC was associated with mild immunosuppression or even immune activation, while the high amount of that was associated with significant immunosuppressive effect. In this study, by using a transwell system, we explored the effect of MSC on the cell cycle, apoptosis rate and the expression of CD69, an activation marker, on the allogeneic cord blood derived cytokine-induced killer(CIK)/natural killer(NK) cells. The results showed that either by transwell or mixed cell-cell co-culture, the MSC can effect CIK/NK cells on the cell cycle, such as arrested in the G0/G1 phase, diminished the ratio of cells in S, G2/M phase, and increased the apoptosis of them. MSC can also depress the expression of CD69 on these killer cells, as well as increased the ratio of CD4(+) CD25(+) CD127(low) T regulatory (Treg) cells in the CIK/NK cell culture system. We draw conclusions that either by transwell or mixed co-culture, the MSC can suppress activation of allogeneic CB-CIK/NK cells in a dose-dependent manner.

  1. Dendritic cell exosomes directly kill tumor cells and activate natural killer cells via TNF superfamily ligands

    PubMed Central

    Munich, Stephan; Sobo-Vujanovic, Andrea; Buchser, William J.; Beer-Stolz, Donna; Vujanovic, Nikola L.

    2012-01-01

    Autocrine and paracrine cell communication can be conveyed by multiple mediators, including membrane-associate proteins, secreted proteins and exosomes. Exosomes are 30–100 nm endosome-derived vesicles consisting in cytosolic material surrounded by a lipid bilayer containing transmembrane proteins. We have previously shown that dendritic cells (DCs) express on their surface multiple TNF superfamily ligands (TNFSFLs), by which they can induce the apoptotic demise of tumor cells as well as the activation of natural killer (NK) cells. In the present study, we demonstrate that, similar to DCs, DC-derived exosomes (DCex) express on their surface TNF, FasL and TRAIL, by which they can trigger caspase activation and apoptosis in tumor cells. We also show that DCex activate NK cells and stimulate them to secrete interferonγ (IFNγ) upon the interaction of DCex TNF with NK-cell TNF receptors. These data demonstrate that DCex can mediate essential innate immune functions that were previously ascribed to DCs. PMID:23170255

  2. CD94 expression and natural killer cell activity after acute exercise.

    PubMed

    Roberts, C; Pyne, D B; Horn, P L

    2004-06-01

    This study examined the effects of acute exercise on natural killer (NK) cell numbers, their expression of CD94 and cytotoxic capacity in triathletes over a 10-week training period. Nine highly trained male triathletes (age 25.9+/-4.1 yrs, VO2max 5.14+/-0.33 L.min(-1)) attended the laboratory on weeks 0, 2, 5 and 10 for incremental submaximal and maximal cycle ergometry. Peripheral blood was analysed for white blood cell counts, lymphocyte phenotype and cytolytic activity (51Cr release from K562 cells). Maximum oxygen consumption increased from week 2 (5.14+/-0.33 L.min(-1)) to week 10 (5.28+/-0.32 L.min(-1)). Resting NK cell numbers and their expression of CD94 were not altered over the 10-week study period. Natural killer cells expressing CD94+ were not differentially recruited into the circulation and cytolytic activity of exercise-recruited NKs did not differ from those present at rest. There was longitudinal stability (over the 10 weeks of the study) in CD94 expression on NK cells, exercise recruitment of CD94+ NK cells and cytolytic capacity of NK cells. The distribution and functional activity of NK cells are not markedly influenced by 10 weeks of training in competitive triathletes. Natural killer cytotoxic activity after exercise reflects numbers of NK cells and not a changed activation state of these cells per se.

  3. Cystatin F regulates proteinase activity in IL-2-activated natural killer cells.

    PubMed

    Maher, Katarina; Konjar, Spela; Watts, Colin; Turk, Boris; Kopitar-Jerala, Natasa

    2014-01-01

    Cystatin F is a unique member of the cystatin family of cysteine protease inhibitors, which is synthesized as an inactive dimer and it is activated by N-terminal cleavage in the endolysosomes. It is expressed in the cells of the immune system: myeloid cells and the cells involved in target cell killing: natural killer (NK) cells and cytotoxic T cells (CTLs). Upon activation of the NK cells with interleukin 2 (IL-2), cystatin F was found upregulated and co-localized in cytotoxic granules with cathepsin C (CatC) and CatV. However, cystatin F inhibits the CatC in cells only when its N-terminal part is processed. Although cystatin F could inhibit both CatV and CatC, the IL-2 stimulation of the YT cells resulted in an increased CatV activity, while the CatC activity was unchanged. The incubation of IL-2 activated NK cells with a cysteine proteinase inhibitor E-64d increased the cystatin F dimer formation. Our results suggest that cystatin F not only inhibits CatV, but it is processed by the CatV in order to inhibit the CatC activity in cytotoxic granules. The regulation of the CatC activity in the cytotoxic granules of the NK cells by the cystatin F could be important for the processing and activation of granule-associated serine proteases - granzymes.

  4. Artificial antigen presenting cell (aAPC) mediated activation and expansion of natural killer T cells.

    PubMed

    East, James E; Sun, Wenji; Webb, Tonya J

    2012-01-01

    Natural killer T (NKT) cells are a unique subset of T cells that display markers characteristic of both natural killer (NK) cells and T cells(1). Unlike classical T cells, NKT cells recognize lipid antigen in the context of CD1 molecules(2). NKT cells express an invariant TCRα chain rearrangement: Vα14Jα18 in mice and Vα24Jα18 in humans, which is associated with Vβ chains of limited diversity(3-6), and are referred to as canonical or invariant NKT (iNKT) cells. Similar to conventional T cells, NKT cells develop from CD4-CD8- thymic precursor T cells following the appropriate signaling by CD1d (7). The potential to utilize NKT cells for therapeutic purposes has significantly increased with the ability to stimulate and expand human NKT cells with α-Galactosylceramide (α-GalCer) and a variety of cytokines(8). Importantly, these cells retained their original phenotype, secreted cytokines, and displayed cytotoxic function against tumor cell lines. Thus, ex vivo expanded NKT cells remain functional and can be used for adoptive immunotherapy. However, NKT cell based-immunotherapy has been limited by the use of autologous antigen presenting cells and the quantity and quality of these stimulator cells can vary substantially. Monocyte-derived DC from cancer patients have been reported to express reduced levels of costimulatory molecules and produce less inflammatory cytokines(9,10). In fact, murine DC rather than autologous APC have been used to test the function of NKT cells from CML patients(11). However, this system can only be used for in vitro testing since NKT cells cannot be expanded by murine DC and then used for adoptive immunotherapy. Thus, a standardized system that relies on artificial Antigen Presenting Cells (aAPC) could produce the stimulating effects of DC without the pitfalls of allo- or xenogeneic cells(12, 13). Herein, we describe a method for generating CD1d-based aAPC. Since the engagement of the T cell receptor (TCR) by CD1d-antigen complexes is

  5. Antibody-dependent cellular cytotoxicity toward neuroblastoma enhanced by activated invariant natural killer T cells.

    PubMed

    Mise, Naoko; Takami, Mariko; Suzuki, Akane; Kamata, Toshiko; Harada, Kazuaki; Hishiki, Tomoro; Saito, Takeshi; Terui, Keita; Mitsunaga, Tetsuya; Nakata, Mitsuyuki; Ikeuchi, Takayuki; Nakayama, Toshinori; Yoshida, Hideo; Motohashi, Shinichiro

    2016-03-01

    Anti-ganglioside GD2 antibodies mainly work through antibody-dependent cellular cytotoxicity (ADCC) and have demonstrated clinical benefit for children with neuroblastoma. However, high-risk neuroblastoma still has a high recurrence rate. For further improvement in patient outcomes, ways to maximize the cytotoxic effects of anti-GD2 therapies with minimal toxicity are required. Activated invariant natural killer T (iNKT) cells enhance both innate and type I acquired anti-tumor immunity by producing several kinds of cytokines. In this report, we investigated the feasibility of combination therapy using iNKT cells and an anti-GD2 antibody. Although some of the expanded iNKT cells expressed natural killer (NK) cell markers, including FcγR, iNKT cells were not directly associated with ADCC. When co-cultured with activated iNKT cells, granzyme A, granzyme B and interferon gamma (IFNγ) production from NK cells were upregulated, and the cytotoxicity of NK cells treated with anti-GD2 antibodies was increased. Not only cytokines produced by activated iNKT cells, but also NK-NKT cell contact or NK cell-dendritic cell contact contributed to the increase in NK cell cytotoxicity and further IFNγ production by iNKT cells and NK cells. In conclusion, iNKT cell-based immunotherapy could be an appropriate candidate for anti-GD2 antibody therapy for neuroblastoma.

  6. Indomethacin augments lymphokine-activated killer cell generation by patients with malignant mesothelioma

    SciTech Connect

    Manning, L.S.; Bowman, R.V.; Davis, M.R.; Musk, A.W.; Robinson, B.W. )

    1989-10-01

    Human malignant mesothelioma (MM) cells are resistant to natural killer (NK) cell lysis but susceptible to lysis by lymphokine-activated killer (LAK) cells from control individuals. The present study was performed to determine the capacity of patients with MM (n = 22) and individuals occupationally exposed to asbestos (the major population at risk of developing this disease, n = 52) to generate LAK cells capable of effectively lysing human mesothelioma cells. Compared to controls (n = 20), both patient groups demonstrated significantly depressed LAK cell activity against mesothelioma tumor cell targets (55 +/- 3% lysis by controls vs 34 +/- 3% lysis by patients with MM, P less than 0.005; and 45 +/- 3% lysis by asbestos-exposed individuals, P less than 0.025). Addition of 10 micrograms/ml indomethacin during LAK cell generation restored normal LAK cell activity for patients with MM (52 +/- 6% lysis of cultured human MM cells, P = NS compared to controls), suggesting that the defective cytolytic cell function observed in some patients with MM is a result of prostaglandin-induced immunosuppression. The ability of indomethacin to restore suppressed LAK cell activity in patients with MM suggests that the concomitant use of this agent in ex vivo LAK cell generation and in patients undergoing interleukin/LAK cell therapy may be beneficial.

  7. Identification of a novel gene expressed in activated natural killer cells and T cells

    SciTech Connect

    Dahl, C.A.; Schall, R.P.; He, H.; Cairns, J.S. )

    1992-01-15

    The authors have isolated a cDNA clone from a human activated NK cell-derived cDNA library that identifies a transcript [NK4] that is selectively expressed in lymphocytes. The expression of this transcript is increased after activation of T cells by mitogens or activation of NK cells by IL-2 (lymphokine-activated killer cells). The transcript levels demonstrated by Northern blot analysis increase by 12 h after activation, remain high for at least 48 h, and require protein synthesis for expression. Southern blot analysis of B lymphoblastoid lines derived from 18 unrelated individuals reveal variable banding patterns suggestive of polymorphism within the NK4 gene. No homology was found between the sequence of the coding region of this transcript and any sequences in the GenBank data base. Sequence homology to the U1 small nuclear RNA was found within the 3[prime] untranslated region immediately upstream of the site of polyadenylation, suggesting a possible role for U1 in the polyadenylation process. Sequence analysis indicates the transcript would encode a protein having a mass of 27 kDa. The presence of a signal sequence and lack of a transmembrane region suggests that the protein is secreted. In addition, the protein contains an RGD sequence that may be involved in cellular adhesion. This transcript appears to encode a novel product common to the activation pathways of both NK cells and T cells. 50 refs., 8 figs.

  8. Combination chemo-immunotherapy: kinetics of in vivo and in vitro generation of natural killer cells and lymphokine-activated killer cells in the rat.

    PubMed Central

    Stewart, L S; Sewell, H F; Thomson, A W

    1990-01-01

    Rats received a single high dose of cyclophosphamide (Cy) (150 mg/kg), followed 48 h later (on day 0) by immunization with a T cell-dependent soluble antigen, ovalbumin in Freund's complete adjuvant (FCA). The effect of this treatment on lymphoid cell subpopulations in the spleen, natural killer (NK) cell and interleukin-2 (IL-2) induced lymphokine-activated killer (LAK) cell activity was examined. Cy (with and without ovalbumin) caused a large relative increase (by day 14) in splenic OX8+, OX19- cells with NK morphology. A marked relative increase in fresh NK cell activity was noted after Cy + ovalbumin, but not consistently after Cy alone. Elevated NK activity was Cy dose- and time-dependent, was evident within 7 days post Cy/ovalbumin and persisted for at least 28 days. Pooled splenic mononuclear cells (MNC), obtained 14 days after Cy/ovalbumin, lost all cytolytic activity against YAC-1 cells when cultured in the absence of human recombinant IL-2 (rIL-2). In contrast, similarly maintained cells from normal rats displayed NK activity higher than normal 'fresh' levels. Upon culture in medium containing 500 U/ml rIL-2, however, 'augmented' NK activity was equivalent, on a per-cell basis, in both normal and Cy/ovalbumin-pretreated groups. LAK activity generated in vitro (i.e. against NK-resistant target cells) was significantly lower in the latter group, and the overall yield of cells was reduced. By day 21 after Cy/ovalbumin, augmented NK activity was significantly greater than controls, on a per-cell and total culture yield basis. Moreover, LAK activity was now similar between groups. It is concluded that the chemotherapy/immunization protocol which we have used can greatly enhance NK activity in vivo and that these cells are responsive to induction of LAK activity by IL-2 in vitro. Images Fig. 1 PMID:2317946

  9. Expression of different lipoprotein receptors in natural killer cells and their effect on natural killer proliferative and cytotoxic activity.

    PubMed Central

    De Sanctis, J B; Blanca, I; Bianco, N E

    1995-01-01

    Natural killer (NK) cells take up chylomicrons (CM), very low density (VLDL), low density (LDL), high density (HDL) and acetyl-modified low density (AcLDL) lipoproteins through different receptors, VLDL being the lipoprotein with the highest uptake and HDL the lowest. The uptake of LDL can be selectively blocked by the anti-LDL receptor, which does not affect the uptake of CM, VLDL, HDL and AcLDL. Although the uptake of lipoproteins assessed by flow cytometry using DiI is not very high, the lipoproteins are able to induce an increase in proliferative responses, VLDL, AcLDL and HDL being the most important ones with 12- and 17-fold increments, respectively. CM, VLDL and LDL at low concentrations increase NK cytotoxic activity, while HDL and AcLDL inhibit, in a dose-dependent fashion, the killing of NK cells against K562. These results suggest the presence of four different receptors that are responsible for the cytotoxic and proliferative responses observed. PMID:8550077

  10. A shed NKG2D ligand that promotes natural killer cell activation and tumor rejection

    PubMed Central

    Deng, Weiwen; Gowen, Benjamin G.; Zhang, Li; Wang, Lin; Lau, Stephanie; Iannello, Alexandre; Xu, Jianfeng; Rovis, Tihana L.; Xiong, Na; Raulet, David H.

    2016-01-01

    Immune cells, including natural killer (NK) cells, recognize transformed cells and eliminate them in a process termed immunosurveillance. It is thought that tumor cells evade immunosurveillance by shedding membrane ligands that bind to the NKG2D activating receptor on NK cells and/or T cells, and desensitize these cells. In contrast, we show that in mice, shedding of MULT1, a high affinity NKG2D ligand, causes NK cell activation and tumor rejection. Recombinant soluble MULT1 stimulated tumor rejection in mice. Soluble MULT1 functions, at least in part, by competitively reversing a global desensitization of NK cells imposed by engagement of membrane NKG2D ligands on tumor-associated cells, such as myeloid cells. The results overturn conventional wisdom that soluble ligands are inhibitory, and suggest a new approach for cancer immunotherapy. PMID:25745066

  11. Natural Killer Cell Memory.

    PubMed

    O'Sullivan, Timothy E; Sun, Joseph C; Lanier, Lewis L

    2015-10-20

    Natural killer (NK) cells have historically been considered short-lived cytolytic cells that can rapidly respond against pathogens and tumors in an antigen-independent manner and then undergo cell death. Recently, however, NK cells have been shown to possess traits of adaptive immunity and can acquire immunological memory in a manner similar to that of T and B cells. In this review, we discuss evidence of NK cell memory and the mechanisms involved in the generation and survival of these innate lymphocytes.

  12. Anger expression and natural killer cell activity in family caregivers participating in a physical activity trial.

    PubMed

    Wilcox, S; King, A C; Vitaliano, P P; Brassington, G S

    2000-07-01

    Associations between psychological functioning and natural killer cell activity (NKA) were examined in 23 older (62.2 ± 7.5 years) family caregivers randomized to a moderate intensity four-month exercise program or to a wait-list control condition. At baseline, although NKA was related to anger-control (r = -.42; trend p < .06) and anger-out (r = .50; p < .03), it was not related to depression, anxiety, perceived stress, or caregiver burden. After controlling for baseline NKA, changes in anger-control explained 14 percent of the variance in NKA four months later. Decreases in anger-control predicted increases in NKA. Group assignment (exercise vs control) was unrelated to changes in NKA over the four-month period; however, the study was not powered to detect this effect. These results are consistent with reported relationships of anger expression with other physiological measures, and extend the importance of anger expression to immune functioning in older family caregivers.

  13. Regulation of natural killer activity of lymphocytes from normal subjects and patients with chronic lymphatic leukemia by interaction between T and non-T cells

    SciTech Connect

    Khonina, N.A.; Shubinskii, G.Z.; Lozovoi, V.P.

    1987-08-01

    The authors study the effect of culture of human cells on functional activity of natural killer cells and investigate the possible mechanisms of regulation of natural killer activity by acting on cytodifferentiation of lymphocytes in normal subjects and in patients with the B-cell variant of chromic lymphatic leukemia. To estimate natural killer cell function, a membranotoxic test was carried out, using cells of the transplantable line K-562, labeled with /sup 3/H-uridine as the targets.

  14. Reduced spleen natural killer cell activity in virally challenged iron-deficient rat pups

    SciTech Connect

    Lockwood, J.F.; Sherman, A.R.

    1986-03-01

    Neonatal iron deficiency has been shown to alter several aspects of immunity in rats. This study determined the effects of iron deficiency on cytotoxicity of virally-induced natural killer cells (NK) in spleen. Rats (n = 8-12/grp) were fed 6 (severe deficiency, SID), 10 (moderate deficiency, MID), or 250 (adequate iron status, AIS) ppm iron d 1 pregnancy through d 21 lactation. Litters were adjusted to 7 on d 2. On d 17 pups were challenged intraperitoneally with 5 x 10/sup 5/ plaque forming units of vaccinia virus. Spleens were collected 4 d later and cell suspensions prepared and pooled within each litter. After isolation of mononuclear cells on a Ficoll-Hypaque gradient, macrophages were removed, and the resulting lymphocytes were incubated with Cr/sup 51/ labelled Yac-1 target cells at effector:target ratios (E:T) of 10:1 and 50:1. Cytotoxicity of NK cells were measured after 4 and 16 hrs by the Cr/sup 51/ release assay. In SID and MID groups body weights, spleen weights, and hemoglobin levels were significantly lower than in AIS pups (p < 0.001). Spleen NK cell cytotoxicity was significantly impaired in SID and MID pups. Depending on the E:T and incubation time, SID and MID cells had activities 30-50% of AIS cells (p < 0.001). Both severe and moderate iron deficiency markedly impair the cytotoxic activity of spleen natural killer cells in suckling rats.

  15. Inhibition of natural killer cell activity by eicosapentaenoic acid in vivo and in vitro

    SciTech Connect

    Yamashita, N.; Sugiyama, E.; Hamazaki, T.; Yano, S.

    1988-01-15

    To examine the effects of in vivo eicosapentaenoic acid (EPA) on natural killer (NK) cell activity, C3H/He mice each received a single intraperitoneal bolus of an emulsion of trieicosapentaenoyl-glycerol (EPA-TG). Spleen cells were tested for NK activity using /sup 51/Chromium-release assays against YAC-1 target cells. Forty eight hours after injection, NK activity was inhibited in a dose-dependent manner. EPA-TG emulsion also inhibited the NK activity of NK-enriched effector cells. Decreased cytotoxicity was first noted 24 hr after injection; it resumed the baseline by 7 days. The addition of EPA-TG emulsion to a cytotoxicity assay system resulted in moderate depression of NK activity. These results demonstrate that EPA has significant immunomodulatory effects on NK activity.

  16. Deficient natural killer cell function in preeclampsia

    SciTech Connect

    Alanen, A.; Lassila, O.

    1982-11-01

    Natural killer cell activity of peripheral blood lymphocytes was measured against K-562 target cells with a 4-hour /sup 51/Cr release assay in 15 primigravid women with preeclamptic symptoms. Nineteen primigravid women with an uncomplicated pregnancy and 18 nonpregnant women served as controls. The natural killer cell activity of preeclamptic women was observed to be significantly lower than that of both control groups. Natural killer cells in preeclamptic women responded normally to augmentation caused by interferon. These findings give further evidence for the participation of the maternal immune system in this pregnancy disorder.

  17. Mechanisms of selective killing of neuroblastoma cells by natural killer cells and lymphokine activated killer cells. Potential for residual disease eradication.

    PubMed

    Foreman, N K; Rill, D R; Coustan-Smith, E; Douglass, E C; Brenner, M K

    1993-05-01

    Widely disseminated neuroblastoma in children older than infancy remains a very poor prognosis disease. Even the introduction of marrow ablative chemotherapy with autologous rescue has not significantly improved the outlook for these children, presumably because of a failure to eradicate minimal residual disease. One additional approach which may hold promise is the use of immunomodulation with cytokines such as IL2 in the setting of minimal residual disease (MDR), for example after intensive chemotherapy and ABMT. However, considerable variability in the susceptibility of neuroblastoma cells to natural killer (NK) and lymphokine-activated (LAK) killing has been observed, and it is presently unclear how NK and LAK cells recognise neuroblastoma cells. In this paper we examine expression of cell adhesion molecules on neuroblastoma to determine which of these modify interaction with NK and LAK cells. We find that LFA-3 (CD58), the ligand for CD2 is of predominant importance in predicting susceptibility of neuroblastoma to the cytotoxic actions of NK and LAK cells, while expression of ICAM-1 (CD54) may also modify susceptibility. These findings were confirmed by blocking experiments in which co-culture of target cells with ICAM-1 and LFA-3 reduced LAK and NK cytotoxicity. Study of the immunophenotypic features of each patient's neuroblastoma cells before induction of MRD may be valuable in determining the likely effect of IL2 in predicting disease reactivation.

  18. Critical roles of co-activation receptor DNAX accessory molecule-1 in natural killer cell immunity

    PubMed Central

    Xiong, Peng; Sang, Hai-Wei; Zhu, Min

    2015-01-01

    Natural killer (NK) cells, which can exert early and powerful anti-tumour and anti-viral responses, are important components of the innate immune system. DNAX accessory molecule-1 (DNAM-1) is an activating receptor molecule expressed on the surface of NK cells. Recent findings suggest that DNAM-1 is a critical regulator of NK cell biology. DNAM-1 is involved in NK cell education and differentiation, and also plays a pivotal role in the development of cancer, viral infections and immune-related diseases. However, tumours and viruses have developed multiple mechanisms to evade the immune system. They are able to impair DNAM-1 activity by targeting the DNAM-1 receptor–ligand system. We have reviewed the roles of DNAM-1, and its biological functions, with respect to NK cell biology and DNAM-1 chimeric antigen receptor-based immunotherapy. PMID:26235210

  19. Biological Character of RetroNectin Activated Cytokine-Induced Killer Cells

    PubMed Central

    2016-01-01

    Adoptive cell therapy (ACT) using autologous cytokine-induced killer (CIK) cells is a promising treatment for metastatic carcinomas. In this study, we investigated the impact of RetroNectin on the proliferation, phenotype alternation, cytokine secretion, and cytotoxic activity of CIK cells from pancreatic cancer patients. Furthermore, we treated 13 patients with metastatic or locally advanced pancreatic cancer using autologous RetroNectin-activated CIK cells (R-CIK cells) alone or in combination with chemotherapy. Compared with only CD3 activated CIK cells (OKT-CIK cells), R-CIK cells showed stronger and faster proliferative ability, with a lower ratio of spontaneous apoptosis. Moreover, this ability continued after IL-2 was withdrawn from the culture system. R-CIK cells could also secrete higher levels of IL-2 and lower levels of IL-4 and IL-5 versus OKT-CIK cells. There was no difference between OKT-CIK and R-CIK cells in cytotoxic ability against lymphoma cell line K562. In patients who received auto-R-CIK cell infusion therapy, the overall objective response rate was 23.1%. Median survival time (mOS) after first R-CIK cell infusion was 10.57 months; the 1-year survival rate was 38.5%. No serious toxicity was associated with R-CIK cell infusion. In conclusion, RetroNectin may enhance antitumor activity of CIK cells: it is safe for use in treating pancreatic cancer. PMID:27433478

  20. Natural Killer Cell Receptors and Cytotoxic Activity in Phosphomannomutase 2 Deficiency (PMM2-CDG)

    PubMed Central

    García-López, Roberto; de la Morena-Barrio, María Eugenia; Alsina, Laia; Pérez-Dueñas, Belén; Jaeken, Jaak; Serrano, Mercedes; Casado, Mercedes; Hernández-Caselles, Trinidad

    2016-01-01

    Background PMM2-CDG is the most common N-glycosylation defect and shows an increased risk of recurrent and/or severe, sometimes fatal, infections in early life. We hypothesized that natural killer (NK) cells, as important mediators of the immune response against microbial pathogens and regulators of adaptive immunity, might be affected in this genetic disorder. Objective To evaluate possible defects on PMM2-CDG NK peripheral blood cell number, killing activity and expression of membrane receptors. Methods We studied fresh and activated NK cells from twelve PMM2-CDG cells. The number and expression of lymphoid surface receptors were studied by flow cytometry. The NK responsiveness (frequency of degranulated NK cells) and killing activity against K562 target cells was determined in the NK cytotoxicity assay. Results We found an increase of blood NK cells in three patients with a severe phenotype. Two of them, who had suffered from moderate/severe viral infections during their first year of life, also had reduced T lymphocyte numbers. Patient activated NK cells showed increased expression of CD54 adhesion molecule and NKG2D and NKp46 activating receptors. NKp46 and 2B4 expression was inversely correlated with the expression of NKG2D in activated PMM2-CDG cells. Maximal NK activity against K562 target cells was similar in control and PMM2-CDG cells. Interestingly, the NK cell responsiveness was higher in patient cells. NKG2D and specially CD54 increased surface expression significantly correlated with the increased NK cell cytolytic activity according to the modulation of the killer activity by expression of triggering receptors and adhesion molecules. Conclusions Our results indicate that hypoglycosylation in PMM2-CDG altered NK cell reactivity against target cells and the expression of CD54 and NKG2D, NKp46 and 2B4 activating receptors during NK cell activation. This suggests a defective control of NK cell killing activity and the overall anti-viral immune response

  1. Natural killer cell activity in elderly men is enhanced by beta-carotene supplementation.

    PubMed

    Santos, M S; Meydani, S N; Leka, L; Wu, D; Fotouhi, N; Meydani, M; Hennekens, C H; Gaziano, J M

    1996-11-01

    Natural killer (NK) cell activity has been postulated to be an immunologic link between beta-carotene and cancer prevention. In a cross-sectional, placebo-controlled, double-blind study we examined the effect of 10-12 y of beta-carotene supplementation (50 mg on alternate days) on NK cell activity in 59 (38 middle-aged men, 51-64 y; 21 elderly men, 65-86 y) Boston area participants in the Physicians' Health Study. No significant difference was seen in NK cell activity due to beta-carotene supplementation in the middle-aged group. The elderly men had significantly lower NK cell activity than the middle-aged men; however, there was no age-associated difference in NK cell activity in men supplemented with beta-carotene. beta-carotene-supplemented elderly men had significantly greater NK cell activity than elderly men receiving placebo. The reason for this is unknown; however, it was not due to an increase in the percentage of NK cells, nor to an increase in interleukin 2 (IL-2) receptor expression, nor to IL-2 production. beta-carotene may be acting directly on one or more of the lytic stages of NK cell cytotoxicity, or on NK cell activity-enhancing cytokines other than IL-2, such as IL-12. Our results show that long-term beta-carotene supplementation enhances NK cell activity in elderly men, which may be beneficial for viral and tumoral surveillance.

  2. Marijuana effects on immunity: suppression of human natural killer cell activity of delta-9-tetrahydrocannabinol.

    PubMed

    Specter, S C; Klein, T W; Newton, C; Mondragon, M; Widen, R; Friedman, H

    1986-01-01

    Delta-9-tetrahydrocannabinol (THC), the major psychoactive component of marijuana, was tested for its ability to modulate human natural killer (NK) cell function. THC was toxic for peripheral blood lymphocytes at 20 micrograms/ml but not at 10 micrograms/ml or less. This component of marijuana also was inhibitory for NK activity against K562, a human tumor cell line at concentrations down to 5 micrograms/ml when pre-incubated with the effector cells. Suppression of NK function was dependent upon the concentration of THC and the length of time of pre-incubation but was independent of the ratio of effector to target cells. Prostaglandins were not involved in suppression of NK activity.

  3. Cellular redox status influences both cytotoxic and NF-kappa B activation in natural killer cells.

    PubMed Central

    Valle Blázquez, M; Luque, I; Collantes, E; Aranda, E; Solana, R; Peña, J; Muñoz, E

    1997-01-01

    The role of cellular redox status in both cytotoxic activity and NF-kappa B activation in natural killer (NK) cells was investigated. The results indicate that stimulation of NK cells, either freshly isolated from peripheral blood lymphocytes (PBL) or long-term cultured NK clones, with specific cell targets results in an increased binding activity of NF-kappa B and AP-1 transcription factors measured by gel retardation. Pretreatment of NK cells with the antioxidant pyrrolidine dithiocarbarmate (PDTC) leads to the inhibition of NF-kappa B activation but the AP-1 binding to DNA was superinduced. The inhibition of NF-kappa B by PDTC paralleled with an inhibition of spontaneous cytotoxicity mediated by NK cells. Moreover, the inhibitors of serine proteases, N-alpha-tosyl-L-lysine chloromethyl ketone and N-alpha-tosyl-L-phenylalanine chloromethyl ketone, also blocked the cytolytic activity of NK cells against the sensitive target K562. In contrast, NK activity was not affected by pretreatment of the effector cells with the proteasome inhibitor N-acetyl-leu-leu-norleucinal which selectively inhibits NF-kappa B activation. Altogether, these results support the hypothesis that the activation of NK cells involved transcriptional and post-transcriptional events, and that reactive intermediates may play an important role in the molecular processes related with the generation of a cytotoxic response by NK cells. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:9155655

  4. Differential effects of stimulatory factors on natural killer cell activities of young and aged mice.

    PubMed

    Nogusa, Shoko; Murasko, Donna M; Gardner, Elizabeth M

    2012-09-01

    Age-associated influences on natural killer (NK) cell functions following cytokine stimulation were examined in splenocytes from C57BL/6 mice. NK cells of both young and aged mice exhibited significantly increased: interferon-γ production after interleukin (IL)-12 or IL-15 alone or any combination of IL-12, IL-18, and IL-2; cytotoxicity after IL-2 or IL-15; and granzyme B expression after IL-15. The only significant age-associated differences were observed in interferon-γ production after IL-15 or IL-12 + 18 + 2 and in granzyme B expression following IL-2 or IL-15. Perforin expression did not increase following stimulation; however, NK cells from aged mice expressed significantly higher levels than young mice. These results underscore the complexity of the cytokine-induced functional activities of NK cells and illustrate the differential response of NK cells from young and aged mice to cytokine stimulation.

  5. What Lies Beneath: Antibody Dependent Natural Killer Cell Activation by Antibodies to Internal Influenza Virus Proteins.

    PubMed

    Vanderven, Hillary A; Ana-Sosa-Batiz, Fernanda; Jegaskanda, Sinthujan; Rockman, Steven; Laurie, Karen; Barr, Ian; Chen, Weisan; Wines, Bruce; Hogarth, P Mark; Lambe, Teresa; Gilbert, Sarah C; Parsons, Matthew S; Kent, Stephen J

    2016-06-01

    The conserved internal influenza proteins nucleoprotein (NP) and matrix 1 (M1) are well characterised for T cell immunity, but whether they also elicit functional antibodies capable of activating natural killer (NK) cells has not been explored. We studied NP and M1-specific ADCC activity using biochemical, NK cell activation and killing assays with plasma from healthy and influenza-infected subjects. Healthy adults had antibodies to M1 and NP capable of binding dimeric FcγRIIIa and activating NK cells. Natural symptomatic and experimental influenza infections resulted in a rise in antibody dependent NK cell activation post-infection to the hemagglutinin of the infecting strain, but changes in NK cell activation to M1 and NP were variable. Although antibody dependent killing of target cells infected with vaccinia viruses expressing internal influenza proteins was not detected, opsonising antibodies to NP and M1 likely contribute to an antiviral microenvironment by stimulating innate immune cells to secrete cytokines early in infection. We conclude that effector cell activating antibodies to conserved internal influenza proteins are common in healthy and influenza-infected adults. Given the significance of such antibodies in animal models of heterologous influenza infection, the definition of their importance and mechanism of action in human immunity to influenza is essential. PMID:27428437

  6. Activated natural killer cells accelerate liver damage in patients with chronic hepatitis B virus infection.

    PubMed

    Zheng, Q; Zhu, Y Y; Chen, J; Ye, Y B; Li, J Y; Liu, Y R; Hu, M L; Zheng, Y C; Jiang, J J

    2015-06-01

    Emerging evidence indicates that natural killer (NK) cells may contribute to liver injury in patients with hepatitis B virus (HBV) infection. Because HBV infection progresses through various disease phases, the cytolytic profiles of peripheral and intrahepatic NK cells in HBV-infected patients remain to be defined. In this study, we comprehensively characterized intrahepatic and peripheral NK cells in a cohort of HBV-infected individuals, and investigated their impact on liver pathogenesis during chronic HBV infection. The study population included 34 immune-clearance (IC) patients, 36 immune-tolerant (IT) carriers and 10 healthy subjects. We found that the activity of peripheral NK cells from IC patients was functionally elevated compared to IT carriers and controls, and NK cell activation was indicated by an increased expression of CD69, CD107a, interferon (IFN)-γ and tumour necrosis factor (TNF)-α. Further analysis showed that the increased activity of both peripheral and hepatic NK cells was correlated positively with liver injury, which was assessed by serum alanine aminotransferase levels (ALT) and the liver histological activity index (HAI). Interestingly, the frequency of peripheral NK cells was reduced in IC patients (especially those with higher HAI scores of 3-4), but there was a concomitant increase in hepatic NK cells. The functionally activated NK cells are enriched preferentially in the livers of IC patients and skew towards cytolytic activity that accelerates liver injury in chronic hepatitis B (CHB) patients.

  7. Porphyromonas gingivalis Lipopolysaccharide Induced Proliferation and Activation of Natural Killer Cells in Vivo.

    PubMed

    Wang, Yuhua; Zhang, Wei; Xu, Li; Jin, Jun-O

    2016-01-01

    Porphyromonas gingivalis (P. gingivalis) lipopolysaccharide (LPS) promoted different innate immune activation than that promoted by Escherichia coli (E. coli) LPS. In this study, we examined the effect of P. gingivalis LPS on the proliferation and activation of natural killer (NK) cells in vivo and compared that function with that of E. coli LPS. Administration of P. gingivalis LPS to C57BL/6 mice induced stronger proliferation of NK cells in the spleen and submandibular lymph nodes (sLNs) and increased the number of circulating NK cells in blood compared to those treated with E. coli LPS. However, P. gingivalis LPS did not induce interferon-gamma (IFN-γ) production and CD69 expression in the spleen and sLN NK cells in vivo, and this was attributed to the minimal activation of the spleen and sLN dendritic cells (DCs), including low levels of co-stimulatory molecule expression and pro-inflammatory cytokine production. Furthermore, P. gingivalis LPS-treated NK cells showed less cytotoxic activity against Yac-1 target cells than E. coli LPS-treated NK cells. Hence, these data demonstrated that P. gingivalis LPS promoted limited activation of spleen and sLN NK cells in vivo, and this may play a role in the chronic inflammatory state observed in periodontal disease. PMID:27548133

  8. Cell cycle arrest and apoptosis, two alternative mechanisms for PMKT2 killer activity.

    PubMed

    Santos, Antonio; Alonso, Alejandro; Belda, Ignacio; Marquina, Domingo

    2013-01-01

    Pichia membranifaciens CYC 1086 secretes a unique 30kDa killer toxin (PMKT2) that inhibits a variety of spoilage yeasts and fungi of agronomical interest. The cytocidal effect of PMKT2 on Saccharomyces cerevisiae cells was studied. Metabolic events associated with the loss of S. cerevisiae viability caused by PMKT2 were qualitatively identical to those reported for K28 killer toxin activity, but different to those reported for PMKT. At higher doses, none of the cellular events accounting for the action of PMKT, the killer toxin secreted by P. membranifaciens CYC 1106, was observed for PMKT2. Potassium leakage, sodium influx and the decrease of intracellular pH were not among the primary effects of PMKT2. We report here that this protein is unable to form ion-permeable channels in liposome membranes, suggesting that channel formation is not the mechanism of cytotoxic action of PMKT2. Nevertheless, flow cytometry studies have revealed a cell cycle arrest at an early S-phase with an immature bud and pre-replicated 1n DNA content. By testing the sensitivity of cells arrested at different stages in the cell cycle, we hoped to identify the execution point for lethality more precisely. Cells arrested at the G1-phase by α-factor or arrested at G2-phase by the spindle poison methyl benzimidazol-2-yl-carbamate (MBC) were protected against the toxin. Cells released from the arrest in both cases were killed by PMKT2 at a similar rate. Nevertheless, cells released from MBC-arrest were able to grow for a short time, and then viability dropped rapidly. These findings suggest that cells released from G2-phase are initially able to divide, but die in the presence of PMKT2 after initiating the S-phase in a new cycle, adopting a terminal phenotype within that cycle. By contrast, low doses of PMKT and PMKT2 were able to generate the same cellular response. The evidence presented here shows that treating yeast with low doses of PMKT2 leads to the typical membranous, cytoplasmic

  9. Safety and Efficacy of Activated Transfected Killer Cells for Neutropenic Fungal Infections

    PubMed Central

    Lin, Lin; Ibrahim, Ashraf S.; Baquir, Beverlie; Fu, Yue; Applebaum, David; Schwartz, Julie; Wang, Amy; Avanesian, Valentina; Spellberg, Brad

    2010-01-01

    Background Invasive fungal infections cause considerable morbidity and mortality in neutropenic patients. White blood cell transfusions are a promising treatment for such infections, but technical barriers have prevented their widespread use. Methods To recapitulate white blood cell transfusions, we are developing a cell-based immunotherapy using a phagocytic cell line, HL-60. We sought to stably transfect HL-60 cells with a suicide trap (herpes simplex virus thymidine kinase), to enable purging of the cells when desired, and a bioluminescence marker, to track the cells in vivo in mice. Results Transfection was stable despite 20 months of continuous culture or storage in liquid nitrogen. Activation of these transfected cells with retinoic acid and dimethyl sulfamethoxazole enhanced their microbicidal effects. Activated transfected killer (ATAK) cells were completely eliminated after exposure to ganciclovir, confirming function of the suicide trap. ATAK cells improved the survival of neutropenic mice with lethal disseminated candidiasis and inhalational aspergillosis. Bioluminescence and histopathologic analysis confirmed that the cells were purged from surviving mice after ganciclovir treatment. Comprehensive necropsy, histopathology, and metabolomic analysis revealed no toxicity of the cells. Conclusions These results lay the groundwork for continued translational development of this promising, novel technology for the treatment of refractory infections in neutropenic hosts. PMID:20397927

  10. Inhibition of human natural killer cell functional activity by human aspartyl β-hydroxylase.

    PubMed

    Huyan, Ting; Li, Qi; Ye, Lin-Jie; Yang, Hui; Xue, Xiao-Ping; Zhang, Ming-Jie; Huang, Qing-Sheng; Yin, Da-Chuan; Shang, Peng

    2014-12-01

    Natural killer (NK) cells are a key component of the innate immune system and play pivotal roles as inflammatory regulators and in tumor surveillance. Human aspartyl β-hydroxylase (HAAH) is a plasma membrane and endoplasmic reticulum protein with hydroxylation activity, which is over-expressed in many malignant neoplasms and can be detected from the sera of tumor patients. HAAH is involved in regulating tumor cell infiltration and metastasis. Escaping from immune surveillance may help tumor cell infiltration and metastasis. However, the effects of HAAH on tumor immune surveillance have not yet been investigated carefully. The present study investigated the potential use of HAAH as an immune regulator of human NK cells. We assessed the effects of recombinant HAAH (r-HAAH) on primary human NK cell morphology, viability, cytotoxicity, apoptosis, receptors expression and cytokine/cytolytic proteins production. Our results demonstrated that r-HAAH negatively affects NK cell activity in a time and dose-dependent manner. It noticeably reduces the viability of the NK cells by increasing apoptosis and necrosis via caspase signaling pathways. Moreover, r-HAAH reduces the NK cell cytotoxicity by inhibiting surface expression of NKG2D, NKp44 and IFN-γ secretion. These findings suggest that one of the ways by which HAAH actively promotes tumor formation and proliferation is by inhibiting NK cell-surveillance activity.

  11. Lysis of neuroblastoma cell lines by human natural killer cells activated by interleukin-2 and interleukin-12.

    PubMed

    Rossi, A R; Pericle, F; Rashleigh, S; Janiec, J; Djeu, J Y

    1994-03-01

    Neuroblastoma is the most common extracranial, solid tumor in children. Despite intensive chemotherapy and bone marrow transplantation, the 5-year projected survival rate is 20% to 25%. In vitro studies have shown enhanced natural killer cell (NK) lysis of tumor cells after exposure of NK cells to interleukin-2 (IL-2). In vivo studies have demonstrated similar immunologic effects but have also revealed severe toxicities associated with the use of IL-2. IL-12 is a newly described cytokine that has several properties, including the ability to act synergistically with IL-2 in generating lymphokine-activated killer cells (LAK) against known tumor targets. We investigated the role of IL-12 in the generation of peripheral blood mononuclear cell (PBMC) lysis of neuroblastoma cell lines. PBMC were activated with IL-12 alone and in combination with IL-2. Whereas IL-12 alone produced only modest enhancement of NK cell cytotoxicity, the combination of IL-2 and IL-12 was most effective in activating NK cell lysis of neuroblastoma cell lines. Further, we showed that large granular lymphocytes were the effector cells involved in target cell lysis. Finally, the CD18 molecule was shown to be critical in the lysis of neuroblastoma cells by activated PBMC.

  12. Low natural-killer-cell activity in familial melanoma patients and their relatives.

    PubMed

    Hersey, P; Edwards, A; Honeyman, M; McCarthy, W H

    1979-07-01

    Patients with melanoma who had one or more close relatives with melanoma were studied for their natural-killer-cell (NK) activity against cultured melanoma cells and Chang cells. A high proportion of the patients and their relatives were found to have low NK activity against these target cells. In most of the patients this could not be attributed to general depression of their immune function, since B- and T-cell numbers and the mitogenic response to PHA were within normal limits. The levels of NK activity of the patients and their relatives were found to be significantly correlated, suggesting that the NK activity in these families may have been genetically (or environmentally) determined. Several genetic markers were examined in the patients and their relatives for association with the disease state and NK activity. No association with HLA antigens or ABO blood groups was detected, but there was a low incidence of the Rhesus negative phenotype in the patients (the Rh phenotype had previously been associated with high NK activity). The present results indicate that NK activity has a familial association in families with a high incidence of melanoma, and raise the question whether low NK activity may be one of the predisposing factors in the development of familial melanoma.

  13. Retargeting cytokine-induced killer cell activity by CD16 engagement with clinical-grade antibodies

    PubMed Central

    Cappuzzello, Elisa; Tosi, Anna; Zanovello, Paola; Sommaggio, Roberta; Rosato, Antonio

    2016-01-01

    ABSTRACT Cytokine-induced Killer (CIK) cells are a heterogeneous population of ex vivo expanded T lymphocytes capable of MHC-unrestricted antitumor activity, which share phenotypic and functional features with both NK and T cells. Preclinical data and initial clinical studies demonstrated their high tolerability in vivo, supporting CIK cells as a promising cell population for adoptive cell immunotherapy. In this study, we report for the first time that CIK cells display a donor-dependent expression of CD16, which can be engaged by trastuzumab or cetuximab to exert a potent antibody-dependent cell-mediated cytotoxicity (ADCC) against ovarian and breast cancer cell lines, leading to an increased lytic activity in vitro, and an enhanced therapeutic efficacy in vivo. Thus, an efficient tumor antigen-specific retargeting can be achieved by a combination therapy with clinical-grade monoclonal antibodies already widely used in cancer therapy, and CIK cell populations that are easily expandable in very large numbers, inexpensive, safe and do not require genetic manipulations. Overall, these data provide a new therapeutic strategy for the treatment of Her2 and EGFR expressing tumors by adoptive cell therapy, which could find wide implementation and application, and could also be expanded to the use of additional therapeutic antibodies. PMID:27622068

  14. Retargeting cytokine-induced killer cell activity by CD16 engagement with clinical-grade antibodies.

    PubMed

    Cappuzzello, Elisa; Tosi, Anna; Zanovello, Paola; Sommaggio, Roberta; Rosato, Antonio

    2016-08-01

    Cytokine-induced Killer (CIK) cells are a heterogeneous population of ex vivo expanded T lymphocytes capable of MHC-unrestricted antitumor activity, which share phenotypic and functional features with both NK and T cells. Preclinical data and initial clinical studies demonstrated their high tolerability in vivo, supporting CIK cells as a promising cell population for adoptive cell immunotherapy. In this study, we report for the first time that CIK cells display a donor-dependent expression of CD16, which can be engaged by trastuzumab or cetuximab to exert a potent antibody-dependent cell-mediated cytotoxicity (ADCC) against ovarian and breast cancer cell lines, leading to an increased lytic activity in vitro, and an enhanced therapeutic efficacy in vivo. Thus, an efficient tumor antigen-specific retargeting can be achieved by a combination therapy with clinical-grade monoclonal antibodies already widely used in cancer therapy, and CIK cell populations that are easily expandable in very large numbers, inexpensive, safe and do not require genetic manipulations. Overall, these data provide a new therapeutic strategy for the treatment of Her2 and EGFR expressing tumors by adoptive cell therapy, which could find wide implementation and application, and could also be expanded to the use of additional therapeutic antibodies. PMID:27622068

  15. Synthetic glycolipid activators of natural killer T cells as immunotherapeutic agents

    PubMed Central

    Carreño, Leandro J; Saavedra-Ávila, Noemí A; Porcelli, Steven A

    2016-01-01

    Certain types of glycolipids have been found to have remarkable immunomodulatory properties as a result of their ability to activate specific T lymphocyte populations with an extremely wide range of immune effector properties. The most extensively studied glycolipid reactive T cells are known as invariant natural killer T (iNKT) cells. The antigen receptors of these cells specifically recognize certain glycolipids, most notably glycosphingolipids with α-anomeric monosaccharides, presented by the major histocompatibility complex class I-like molecule CD1d. Once activated, iNKT cells can secrete a very diverse array of pro- and anti-inflammatory cytokines to modulate innate and adaptive immune responses. Thus, glycolipid-mediated activation of iNKT cells has been explored for immunotherapy in a variety of disease states, including cancer and a range of infections. In this review, we discuss the design of synthetic glycolipid activators for iNKT cells, their impact on adaptive immune responses and their use to modulate iNKT cell responses to improve immunity against infections and cancer. Current challenges in translating results from preclinical animal studies to humans are also discussed. PMID:27195112

  16. Turned on by danger: activation of CD1d-restricted invariant natural killer T cells.

    PubMed

    Lawson, Victoria

    2012-09-01

    CD1d-restricted invariant natural killer T (iNKT) cells bear characteristics of innate and adaptive lymphocytes, which allow them to bridge the two halves of the immune response and play roles in many disease settings. Recent work has characterized precisely how their activation is initiated and regulated. Novel antigens from important pathogens have been identified, as has an abundant self-antigen, β-glucopyranosylcaramide, capable of mediating an iNKT-cell response. Studies of the iNKT T-cell receptor (TCR)-antigen-CD1d complex show how docking between CD1d-antigen and iNKT TCR is highly conserved, and how small sequence differences in the TCR establish intrinsic variation in iNKT TCR affinity. The sequence of the TCR CDR3β loop determines iNKT TCR affinity for ligand-CD1d, independent of ligand identity. CD1d ligands can promote T helper type 1 (Th1) or Th2 biased cytokine responses, depending on the composition of their lipid tails. Ligands loaded into CD1d on the cell surface promote Th2 responses, whereas ligands with long hydrophobic tails are loaded endosomally and promote Th1 responses. This information is informing the design of synthetic iNKT-cell antigens. The iNKT cells may be activated by exogenous antigen, or by a combination of dendritic cell-derived interleukin-12 and iNKT TCR-self-antigen-CD1d engagement. The iNKT-cell activation is further modulated by recent foreign or self-antigen encounter. Activation of dendritic cells through pattern recognition receptors alters their antigen presentation and cytokine production, strongly influencing iNKT-cell activation. In a range of bacterial infections, dendritic cell-dependent innate activation of iNKT cells through interleukin-12 is the dominant influence on their activity.

  17. Aspergillus fumigatus contamination of lymphokine-activated killer cells infused into cancer patients.

    PubMed

    Arnow, P M; Houchins, S G; Richards, J M; Chudy, R

    1991-05-01

    Lymphokine-activated killer (LAK) cells, prepared by incubating autologous lymphocytes in cell culture medium with interleukin-2, selectively lyse tumor cells and are effective immunotherapy of some cancers. During a 3-month period, two patients at our center were infused with LAK cells subsequently found to have been contaminated by Aspergillus fumigatus. Each case was investigated by obtaining environmental cultures and assessing aseptic practices during LAK cell preparation. Investigation of the first case demonstrated a malfunction of the laminar air flow hood, under which interleukin-2 and the patient's lymphocytes had been added to cell culture medium, and showed heavy A. fumigatus contamination of the hood, adjacent countertop, and cell culture incubator. Despite repair of the laminar air flow hood and cleaning of the laboratory, a second case occurred, and cultures at that time implicated the humidified cell culture incubators as the source of A. fumigatus. Following incubator sterilization and removal of the humidification apparatus from the incubators, weekly environmental cultures in the LAK cell laboratory were negative, and none of the LAK cell cultures from the 20 patients treated during the ensuing 15 months grew A. fumigatus. Our findings show that growth of fungi in humidified incubators, which previously has caused contamination problems in tissue culture and clinical microbiology laboratories, can result in patient infections when humidified incubators are used to prepare cells for reinfusion.

  18. Surface aminopeptidase activity of rat natural killer cells. I. Biochemical and biological properties.

    PubMed

    Amoscato, A A; Spiess, R R; Brumfield, A M; Herberman, R B; Chambers, W H

    1994-04-28

    Aminopeptidase (AP) activity on rat natural killer (NK) cells was found to have the following characteristics: (1) the activity was surface associated and not secreted, as determined by extracellular location of product and by the cessation of hydrolysis of substrate upon removal of the cells from the medium. (2) The activity was linear with respect to time and cell number. (3) The enzymatic activity on splenocytes and on the NK leukemia cell line CRNK-16, but not on IL-2 activated NK (A-NK) cells, was sensitive to trypsin treatment. (4) The AP activity on intact cells had a broad pH dependency with optimal activity at slightly alkaline pH but lower activity at acidic pH. (5) There was a preference for neutral substrates and essentially no activity towards acidic substrates. (6) Enzymatic activity was inhibited in the presence of the AP inhibitors bestatin and amastatin, and in the presence of the chelator, 1,10 phenanthroline, indicating the involvement of a metalloprotease. (7) Culture of A-NK cells with bestatin resulted in a decrease in cytotoxicity against YAC-1 and P815 targets. Amastatin treatment caused only a slight decrease in cytotoxicity against YAC-1 targets, but a significant decrease in cytotoxicity against P815 targets. (8) Treatment of A-NK cultures with specific inhibitors of APases caused an increase in expression of CD2 (an increase from 20-80% with bestatin and an increase from 25-35% in the presence of amastatin). These results provide the first evidence for the existence of APases on the surface of NK cells and suggest a role for these enzymes in the regulation of cytotoxic activity and of CD2 surface expression.

  19. Activation of decidual invariant natural killer T cells promotes lipopolysaccharide-induced preterm birth.

    PubMed

    Li, Liping; Yang, Jing; Jiang, Yao; Tu, Jiaoqin; Schust, Danny J

    2015-04-01

    Invariant natural killer T (iNKT) cells are crucial for host defense against a variety of microbial pathogens, but the underlying mechanisms of iNKT cells activation by microbes are not fully explained. In this study, we investigated the molecular mechanisms of iNKT cell activation in lipopolysaccharide (LPS)-stimulated preterm birth using an adoptive transfer system and diverse neutralizing antibodies (Abs) and inhibitors. We found that adoptive transfer of decidual iNKT cells to LPS-stimulated iNKT cell deficient Jα18(-/-) mice that lack invariant Vα14Jα281T cell receptor (TCR) expression significantly decreased the time to delivery and increased the percentage of decidual iNKT cells. Neutralizing Abs against Toll-like receptor 4 (TLR-4), CD1d, interleukin (IL)-12 and IL-18, and inhibitors blocking the activation of nuclear factor κB (NF-κB), mitogen-activated protein kinase (MAPK) p38 and extracellular signal-regulated kinase (ERK) significantly reduced in vivo percentages of decidual iNKT cells, their intracellular interferon (IFN)-γ production and surface CD69 expression. In vitro, in the presence of the same Abs and inhibitors used as in vivo, decidual iNKT cells co-cultured with LPS-pulsed dendritic cells (DCs) showed significantly decreased extracellular and intracellular IFN-γ secretion and surface CD69 expression. Our data demonstrate that the activation of decidual iNKT cells plays an important role in inflammation-induced preterm birth. Activation of decidual iNKT cells also requires TLR4-mediated NF-κB, MAPK p38 and ERK pathways, the proinflammatory cytokines IL-12 and IL-18, and endogenous glycolipid antigens presented by CD1d.

  20. All-trans retinoic acid negatively regulates cytotoxic activities of nature killer cell line 92

    SciTech Connect

    Li Ang . E-mail: liang3829@sina.com.cn; He Meilan; Wang Hui; Qiao Bin; Chen Ping; Gu Hua; Zhang Mengjie; He Shengxiang

    2007-01-05

    NK cells are key components of innate immune systems and their activities are regulated by cytokines and hormones. All-trans retinoic acid (ATRA), as a metabolite of vitamin A and an immunomodulatory hormone, plays an important role in regulating immune responses. In the present study, we investigated the effect of ATRA on human NK cell line NK92. We found that ATRA dose-dependently suppressed cytotoxic activities of NK92 cells without affecting their proliferation. To explore the mechanisms underlying the ATRA influence on NK92 cells, we examined the production of cytokines (TNF-{alpha}, IFN-{gamma}), gene expression of cytotoxic-associated molecules (perforin, granzyme B, nature killer receptors (NCRs), and NKG2D), and the activation of NF-{kappa}B pathways related with immune response. Our results demonstrated that ATRA suppressed NF-{kappa}B activity and prevented I{kappa}B{alpha} degradation in a dose-dependent way, inhibited IFN-{gamma} production and gene expression of granzyme B and NKp46. Our findings suggest that ATRA is a negative regulator of NK92 cell activation and may act as a potential regulator of anti-inflammatory functions in vivo.

  1. Interleukin 2 enhances natural killer cell activity through induction of gamma interferon.

    PubMed Central

    Weigent, D A; Stanton, G J; Johnson, H M

    1983-01-01

    Highly purified interleukin 2 (IL 2), free of interferon activity, enhanced natural killer (NK) cell activity against tumor cells in mouse spleen cell cultures and in human peripheral lymphocyte cultures in a manner similar to that of interferon (IFN). We determined that IL 2 enhanced NK activity indirectly in a cascade manner by the induction of gamma IFN (IFN-gamma) in the cultures, which actually mediated the enhanced killing. Accordingly, lymphocyte cultures treated with IL 2 alone produced 10 to 100 U of IFN per ml in 6 to 24 h of culture. The IFN was typed as IFN-gamma by specific antibodies. Specific antibodies either to natural IFN-gamma or to a synthetic peptide corresponding to the human IFN-gamma N-terminal amino acids, when added to cultures treated with IL 2, completely blocked IL 2 enhancement of NK cell activity for both the mouse and human systems. IL 2-induced proliferation was not affected by the antibodies. Thus, the enhancement of NK cell activity by IL 2 is completely mediated by IL 2-induced IFN-gamma. The findings clearly indicate a cascade effect whereby one lymphokine (IL 2) induces the production of another. The latter lymphokine (IFN-gamma) then mediates an important biological effect (natural killing). PMID:6411624

  2. Green Tea Catechin Metabolites Exert Immunoregulatory Effects on CD4(+) T Cell and Natural Killer Cell Activities.

    PubMed

    Kim, Yoon Hee; Won, Yeong-Seon; Yang, Xue; Kumazoe, Motofumi; Yamashita, Shuya; Hara, Aya; Takagaki, Akiko; Goto, Keiichi; Nanjo, Fumio; Tachibana, Hirofumi

    2016-05-11

    Tea catechins, such as (-)-epigallocatechin-3-O-gallate (EGCG), have been shown to effectively enhance immune activity and prevent cancer, although the underlying mechanism is unclear. Green tea catechins are instead converted to catechin metabolites in the intestine. Here, we show that these green tea catechin metabolites enhance CD4(+) T cell activity as well as natural killer (NK) cell activity. Our data suggest that the absence of a 4'-hydroxyl on this phenyl group (B ring) is important for the effect on immune activity. In particular, 5-(3',5'-dihydroxyphenyl)-γ-valerolactone (EGC-M5), a major metabolite of EGCG, not only increased the activity of CD4(+) T cells but also enhanced the cytotoxic activity of NK cells in vivo. These data suggest that EGC-M5 might show immunostimulatory activity. PMID:27112424

  3. Natural Killer T Cell Activation Protects Mice Against Experimental Autoimmune Encephalomyelitis

    PubMed Central

    Singh, Avneesh K.; Wilson, Michael T.; Hong, Seokmann; Olivares-Villagómez, Danyvid; Du, Caigan; Stanic, Aleksandar K.; Joyce, Sebastian; Sriram, Subramaniam; Koezuka, Yasuhiko; Van Kaer, Luc

    2001-01-01

    Experimental autoimmune encephalomyelitis (EAE) serves as a prototypic model for T cell–mediated autoimmunity. Vα14 natural killer T (NKT) cells are a subset of T lymphocytes that recognize glycolipid antigens presented by the nonpolymorphic major histocompatibility complex (MHC) class I–like protein CD1d. Here, we show that activation of Vα14 NKT cells by the glycosphingolipid α-galactosylceramide (α-GalCer) protects susceptible mice against EAE. β-GalCer, which binds CD1d but is not recognized by NKT cells, failed to protect mice against EAE. Furthermore, α-GalCer was unable to protect CD1d knockout (KO) mice against EAE, indicating the requirement for an intact CD1d antigen presentation pathway. Protection of disease conferred by α-GalCer correlated with its ability to suppress myelin antigen-specific Th1 responses and/or to promote myelin antigen-specific Th2 cell responses. α-GalCer was unable to protect IL-4 KO and IL-10 KO mice against EAE, indicating a critical role for both of these cytokines. Because recognition of α-GalCer by NKT cells is phylogenetically conserved, our findings have identified NKT cells as novel target cells for treatment of inflammatory diseases of the central nervous system. PMID:11748281

  4. Activation of human natural killer cells by the soluble form of cellular prion protein

    SciTech Connect

    Seong, Yeon-Jae; Sung, Pil Soo; Jang, Young-Soon; Choi, Young Joon; Park, Bum-Chan; Park, Su-Hyung; Park, Young Woo; Shin, Eui-Cheol

    2015-08-21

    Cellular prion protein (PrP{sup C}) is widely expressed in various cell types, including cells of the immune system. However, the specific roles of PrP{sup C} in the immune system have not been clearly elucidated. In the present study, we investigated the effects of a soluble form of recombinant PrP{sup C} protein on human natural killer (NK) cells. Recombinant soluble PrP{sup C} protein was generated by fusion of human PrP{sup C} with the Fc portion of human IgG{sub 1} (PrP{sup C}-Fc). PrP{sup C}-Fc binds to the surface of human NK cells, particularly to CD56{sup dim} NK cells. PrP{sup C}-Fc induced the production of cytokines and chemokines and the degranulation of granzyme B from NK cells. In addition, PrP{sup C}-Fc facilitated the IL-15-induced proliferation of NK cells. PrP{sup C}-Fc induced phosphorylation of ERK-1/2 and JNK in NK cells, and inhibitors of the ERK or the JNK pathways abrogated PrP{sup C}-Fc-induced cytokine production in NK cells. In conclusion, the soluble form of recombinant PrP{sup C}-Fc protein activates human NK cells via the ERK and JNK signaling pathways. - Highlights: • Recombinant soluble PrP{sup C} (PrP{sup C}-Fc) was generated by fusion of human PrP{sup C} with IgG1 Fc portion. • PrP{sup C}-Fc protein induces the production of cytokines and degranulation from human NK cells. • PrP{sup C}-Fc protein enhances the IL-15-induced proliferation of human NK cells. • PrP{sup C}-Fc protein activates human NK cells via the ERK and JNK signaling pathways.

  5. Human lymphokine-activated killer (LAK) cells: III. Effect of L-phenylalanine methyl ester on LAK cell activation from human peripheral blood mononuclear cells: possible protease involvement of monocytes, natural killer cells and LAK cells.

    PubMed

    Leung, K H

    1991-01-01

    We have shown that depletion of monocytes from human peripheral blood mononuclear cells (PBMC) by L-phenylalanine methyl ester (PheOMe) enhanced lymphokine-activated killer cell (LAK) generation by recombinant interleukin-2 (rIL-2) at high cell density. In this study, we have investigated the mechanism of action of PheOMe on LAK activation by using trypsin, chymotrypsin, tosylphenylalaninechloromethanol (TPCK, a chymotrypsin inhibitor), tosyl-L-lysinechloromethane (TLCK, a trypsin inhibitor), phenylalaninol (PheOH), and benzamidine. PBMC were treated with 1-5 mM PheOMe for 40 min at room temperature in combination with the various agents, washed and assessed for their effects on natural killer (NK) activity against K562 cells and monocyte depletion. The treated cells were then cultured with or without rIL-2 for 3 days. LAK cytotoxicity was assayed against 51Cr-labeled K562 and Raji tumor target cells. TPCK at 10 micrograms/ml partially inhibited depletion of monocytes by PheOMe. TLCK did not prevent depletion of monocytes nor inhibition of NK activity induced by PheOMe. TPCK and TLCK inhibited NK activity by themselves. TPCK but not TLCK inhibited rIL-2 induction of LAK cells. On the other hand, PheOH and benzamidine (analogs of PheOMe) lacked any effect on monocyte depletion but abrogated the inhibitory effect of PheOMe on NK activity. They had no effect on rIL-2 activation of LAK activity enhanced by PheOMe. Trypsin potentiated the inhibitory effect of PheOMe on NK activity and monocyte depletion. Trypsin partially inhibited IL-2 activation of LAK activity enhanced by PheOMe. Chymotrypsin had little effect on NK activity but prevented the inhibitory effect of PheOMe on NK activity. It had little effect on monocyte depletion induced by PheOMe. PheOMe was hydrolysed by monocytes and chymotrypsin to Phe and methanol as determined by HPLC. TPCK inhibited hydrolysis of PheOMe by monocytes. Our data suggest that the effects of PheOMe on monocytes, NK cells and LAK

  6. Lymphokine-activated killer cytotoxicity in neonatal mononuclear cells: in vitro responses to tumor cell lines from pediatric solid tumors.

    PubMed

    Chin, T; Toy, C; Vandeven, C; Cairo, M S

    1989-02-01

    The presence of neonatal (cord) lymphokine-activated killer (LAK) cell activity toward natural killer cell resistant Raji and Daudi cell lines has recently been reported from our laboratory. We investigated the future therapeutic use of LAK adoptive immunotherapy by examining LAK in vitro cytotoxicity from both neonatal and adult mononuclear cells against solid tumor cell lines of relevance to pediatric oncology: SH-SY5Y (neuroblastoma), SK-NM-C (neuroblastoma-neuroepithelioma), NEP-1 (Wilms' tumor), SK-ES-1 (Ewing's sarcoma), and A-204 (rhabdomyosarcoma). Cord and adult mononuclear cells were activated by recombinant IL-2 (100 mu/ml) for 5-7 days and added in an effector:target ratio of 40:1 to 51Cr-labeled target cells. Specific cell lysis was determined after a 4-h incubation. There was a significantly high level of cord and adult LAK cytotoxicity against Wilms' (76.4 +/- 9.8 versus 77.3 +/- 6.8%) and Ewing's (84.2 +/- 5.5 versus 71.1 +/- 6.5%) cell lines and significant but moderate LAK activity against neuroepithelioma (52.0 +/- 6.6 versus 55.4 +/- 4.5%) and rhabdomyosarcoma (46.6 +/- 5.7 versus 43.9 +/- 5.2%) cell lines. There was no difference between cord and adult LAK activity toward these targets. However, a differential response toward the more classical neuroblastoma cell line, SH-SY5Y, was noted with significantly more LAK cytotoxicity from cord mononuclear cells than adult mononuclear cells (51.2 +/- 6.9 versus 28.5 +/- 8.2%) (p less than or equal to 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

  7. beta. -endorphin augments the cytolytic activity and interferon production of natural killer cells

    SciTech Connect

    Mandler, R.N.; Biddison, W.E.; Mandler, R.; Serrate, S.A.

    1986-02-01

    The in vitro effects of the neurohormone ..beta..-endorphin (b-end) on natural killer (NK) activity and interferon (IFN) production mediated by large granular lymphocytes (LGL) were investigated. LGL-enriched fractions from peripheral blood mononuclear cells (PBMC) from normal human volunteers were obtained by fractionation over discontinuous Percoll gradients. LGL were preincubated with or without various concentrations of b-end or the closely related peptides ..cap alpha..-endorphin (a-end), ..gamma..-endorphin (g-end), or D-ALA/sub 2/-..beta..-endorphin (D-ALA/sub 2/-b-end), a synthetic b-end analogue. NK activity was assayed on /sup 51/Cr-labeled K562 target cells. Preincubation of LGL effectors (but not K562 targets) for 2 to 18 hr with concentrations of b-end between 10/sup -7/ M and 10/sup -10/ M produced significant augmentation of NK cytolytic activity (mean percentage increase: 63%). The classic opiate antagonist naloxone blocked the enhancing effect when used at a 100-fold molar excess relative to b-end. These findings demonstrate that b-end enhances NK activity and IFN production of purified LGL, and suggests that b-end might bind to an opioid receptor on LGL that can be blocked by naloxone. These results lend support to the concepts of regulation of the immune response by neurohormones and the functional relationship between the nervous and immune systems.

  8. Staphylococcus-mediated T-cell activation and spontaneous natural killer cell activity in the absence of major histocompatibility complex class II molecules

    NASA Technical Reports Server (NTRS)

    Chapes, S. K.; Hoynowski, S. M.; Woods, K. M.; Armstrong, J. W.; Beharka, A. A.; Iandolo, J. J.; Spooner, B. S. (Principal Investigator)

    1993-01-01

    We used major histocompatibility complex class II antigen-deficient transgenic mice to show that in vitro natural killer cell cytotoxicity and T-cell activation by staphylococcal exotoxins (superantigens) are not dependent upon the presence of major histocompatibility complex class II molecules. T cells can be activated by exotoxins in the presence of exogenously added interleukin 1 or 2 or in the presence of specific antibody without exogenously added cytokines.

  9. Staphylococcus-mediated T-cell activation and spontaneous natural killer cell activity in the absence of major histocompatibility complex class II molecules.

    PubMed Central

    Chapes, S K; Hoynowski, S M; Woods, K M; Armstrong, J W; Beharka, A A; Iandolo, J J

    1993-01-01

    We used major histocompatibility complex class II antigen-deficient transgenic mice to show that in vitro natural killer cell cytotoxicity and T-cell activation by staphylococcal exotoxins (superantigens) are not dependent upon the presence of major histocompatibility complex class II molecules. T cells can be activated by exotoxins in the presence of exogenously added interleukin 1 or 2 or in the presence of specific antibody without exogenously added cytokines. PMID:8359928

  10. Analysis of sphingosine kinase activity in single natural killer cells from peripheral blood.

    PubMed

    Dickinson, Alexandra J; Meyer, Megan; Pawlak, Erica A; Gomez, Shawn; Jaspers, Ilona; Allbritton, Nancy L

    2015-04-01

    Sphingosine-1-phosphate (S1P), a lipid second messenger formed upon phosphorylation of sphingosine by sphingosine kinase (SK), plays a crucial role in natural killer (NK) cell proliferation, migration, and cytotoxicity. Dysregulation of the S1P pathway has been linked to a number of immune system disorders and therapeutic manipulation of the pathway has been proposed as a method of disease intervention. However, peripheral blood NK cells, as identified by surface markers (CD56(+)CD45(+)CD3(-)CD16) consist of a highly diverse population with distinct phenotypes and functions and it is unknown whether the S1P pathway is similarly diverse across peripheral blood NK cells. In this work, we measured the phosphorylation of sphingosine-fluorescein (SF) and subsequent metabolism of S1P fluorescein (S1PF) to form hexadecanoic acid fluorescein (HAF) in 111 single NK cells obtained from the peripheral blood of four healthy human subjects. The percentage of SF converted to S1PF or HAF was highly variable amongst the cells ranging from 0% to 100% (S1PF) and 0% to 97% (HAF). Subpopulations of cells with varying levels of S1PF formation and metabolism were readily identified. Across all subjects, the average percentage of SF converted to S1PF or HAF was 37 ± 36% and 12 ± 19%, respectively. NK cell metabolism of SF by the different subjects was also distinct with hierarchical clustering suggesting two possible phenotypes: low (<20%) or high (>50%) producers of S1PF. The heterogeneity of SK and downstream enzyme activity in NK cells may enable NK cells to respond effectively to a diverse array of pathogens as well as incipient tumor cells. NK cells from two subjects were also loaded with S1PF to assess the activity of S1P phosphatase (S1PP), which converts S1P to sphingosine. No NK cells (n = 41) formed sphingosine, suggesting that S1PP was minimally active in peripheral blood NK cells. In contrast to the SK activity, S1PP activity was homogeneous across the peripheral blood NK

  11. Analysis of sphingosine kinase activity in single natural killer cells from peripheral blood†

    PubMed Central

    Dickinson, Alexandra J.; Meyer, Megan; Pawlak, Erica A.; Gomez, Shawn; Jaspers, Ilona; Allbritton, Nancy L.

    2015-01-01

    Sphingosine-1-phosphate (S1P), a lipid second messenger formed upon phosphorylation of sphingosine by sphingosine kinase (SK), plays a crucial role in natural killer (NK) cell proliferation, migration, and cytotoxicity. Dysregulation of the S1P pathway has been linked to a number of immune system disorders and therapeutic manipulation of the pathway has been proposed as a method of disease intervention. However, peripheral blood NK cells, as identified by surface markers (CD56+CD45+CD3−CD16) consist of a highly diverse population with distinct phenotypes and functions and it is unknown whether the S1P pathway is similarly diverse across peripheral blood NK cells. In this work, we measured the phosphorylation of sphingosine–fluorescein (SF) and subsequent metabolism of S1P fluorescein (S1PF) to form hexadecanoic acid fluorescein (HAF) in 111 single NK cells obtained from the peripheral blood of four healthy human subjects. The percentage of SF converted to S1PF or HAF was highly variable amongst the cells ranging from 0% to 100% (S1PF) and 0% to 97% (HAF). Subpopulations of cells with varying levels of S1PF formation and metabolism were readily identified. Across all subjects, the average percentage of SF converted to S1PF or HAF was 37 ± 36% and 12 ± 19%, respectively. NK cell metabolism of SF by the different subjects was also distinct with hierarchical clustering suggesting two possible phenotypes: low (<20%) or high (>50%) producers of S1PF. The heterogeneity of SK and downstream enzyme activity in NK cells may enable NK cells to respond effectively to a diverse array of pathogens as well as incipient tumor cells. NK cells from two subjects were also loaded with S1PF to assess the activity of S1P phosphatase (S1PP), which converts S1P to sphingosine. No NK cells (n = 41) formed sphingosine, suggesting that S1PP was minimally active in peripheral blood NK cells. In contrast to the SK activity, S1PP activity was homogeneous across the peripheral blood NK

  12. Killer cell lines against Shope carcinoma cells in rabbits.

    PubMed

    Takahashi, M; Yamade, I; Seto, A

    1991-09-01

    Killer cell activity against Shope carcinoma cells was not detected in PBL nor in spleen cells from tumor-bearing B/J rabbits, but was induced by in vitro culture of these cells in the presence of IL-2 and X-irradiated carcinoma cells. HTLV-I-transformed killer cell lines were successfully obtained by the culturing of PBL from an HTLV-I-infected and tumor-bearing Chbb:HM rabbit. These killer cells included large cells with azurophilic granules in the cytoplasm and with a reniform nucleus, thus resembling large granular lymphocytes. The killer activity was similar against the Vx2K cell line from a random-bred rabbit and SCB cell lines from an B/J rabbit, suggesting the absence of MHC restriction. PMID:1655241

  13. Up‐modulation of interferon‐γ mediates the enhancement of spontanous cytotoxicity in prolactin‐activated natural killer cells

    PubMed Central

    Matera, L; Contarini, M; Bellone, G; Forno, B; Biglino, A

    1999-01-01

    Prolactin (PRL) has been shown to participate in lymphocyte activation. In particular, the constitutive natural killer (NK) and the lymphokine‐activated killer (LAK) cytotoxicity of CD56+ CD16+ cells is increased by its physiological to supraphysiological concentrations. As PRL has been shown to up‐regulate the production of interferon‐γ (IFN‐γ) by peripheral blood mononuclear cells, we studied its effect on IFN‐γ production by NK cells as a possible mechanism of autocrine activation of cytotoxicity. Released and intracellular IFN‐γ, as well as IFN‐γ mRNA expression, were increased by pituitary and recombinant human PRL, which stimulated optimal NK and LAK cytotoxicity. Treatment with blocking anti‐IFN‐γ monoclonal antibody (mAb) selectively affected PRL‐increased killing of K562 targets, demonstrating that PRL‐mediated enhancement of spontaneous cytotoxicity depends, at least in part, on up‐regulation of IFN‐γ. PMID:10583598

  14. Augmentation by transferrin of IL-2-inducible killer activity and perforin production of human CD8+ T cells.

    PubMed Central

    Nakamura, A; Sone, S; Nabioullin, R; Sugihara, K; Munekata, M; Nishioka, Y; Nii, A; Ogura, T

    1993-01-01

    The effects of human transferrin (Tf) on lymphokine (IL-2)-activated killer (LAK) induction from blood lymphocytes of healthy donors was examined. LAK cells were induced by 6-day incubation in medium with recombinant human IL-2 of lymphocytes, and their cytotoxic activity was assessed by measuring 51Cr release from NK-resistant Daudi cells. Tf alone did not induce any LAK activity, but in combination with IL-2, it augmented LAK induction dose- and time-dependently. This augmenting effect was completely abolished by pretreatment with anti-Tf antiserum. Tf augmented the proliferative response of lymphocytes to IL-2 and their expressions of receptors for IL-2 and Tf. CD8+ T cells were isolated from purified blood lymphocytes using antibody-bound magnetic beads. Addition of Tf to cultures of CD8+ cells resulted in significant augmentation of killer cell induction and perforin (PFP) production after 4 days stimulation with IL-2. These results indicate that Tf is important in generation of IL-2-inducible killer properties and PFP activity of human CD8+ T cells. PMID:8467561

  15. Effects of microwave exposure on the hamster immune system. I. Natural killer cell activity

    SciTech Connect

    Yang, H.K.; Cain, C.A.; Lockwood, J.; Tompkins, W.A.

    1983-01-01

    Hamsters were exposed to repeated or single doses of microwave energy and monitored for changes in core body temperature, circulating leukocyte profiles, serum corticosteroid levels, and natural killer (NK) cell activity in various tissues. NK cytotoxicity was measured in a /sup 51/Cr-release assay employing baby hamster kidney (BHK) targets or BHK infected with herpes simplex virus. Repeated exposure of hamsters at 15 mW/cm2 for 60 min/day had no significant effect on natural levels of spleen-cell NK activity against BHK targets. Similarly, repeated exposure at 15 mW/cm2 over a 5-day period had no demonstrable effect on the induction of spleen NK activity by vaccinia virus immunization, that is, comparable levels of NK were induced in untreated and microwave-treated animals. In contrast, treatment of hamsters with a single 60-min microwave exposure at 25 mW/cm2 caused a significant suppression in induced spleen NK activity. A similar but less marked decrease in spleen NK activity was observed in sham-exposed animals. Moreover, the sham effects on NK activity were not predictable and appeared to represent large individual animal variations in the response to stress factors. Depressed spleen NK activity was evident as early as 4 h postmicrowave treatment and returned to normal levels by 8 h. Hamsters exposed at 25 mW/cm2 showed an elevated temperature of 3.0-3.5 degrees C that returned to normal within 60 min after termination of microwave exposure. These animals also showed a marked lymphopenia and neutrophilia by 1 h posttreatment that returned to normal by 8-10 h. Serum glucocorticosteroids were elevated between 1 aNd 8 h after microwave treatment. Sham-exposed animals did not demonstrate significant changes in core body temperature, peripheral blood leukocyte (PBL) profile, or glucocorticosteroid levels as compared to minimum-handling controls.

  16. Influence of autologous dendritic cells on cytokine-induced killer cell proliferation, cell phenotype and antitumor activity in vitro

    PubMed Central

    Cao, Jingsong; Chen, Cong; Wang, Yuhuan; Chen, Xuecheng; Chen, Zeying; Luo, Xiaoling

    2016-01-01

    Dendritic cell (DCs) are essential antigen processing and presentation cells that play a key role in the immune response. In this study, DCs were co-cultured with cytokine-induced killer cells (DC-CIKs) in vitro to detect changes in cell proliferation, cell phenotype and cell cytotoxicity. The results revealed that the DCs were suitable for co-culture with CIKs at day 7, and that cell quantity of DC-CIKs was lower than that of CIKs until day 11, but it was significantly improved to 1.17-fold that of CIKs at day 13. Flow cytometry was used to detect the cell phenotype of CIKs and DC-CIKs. Compared with CIKs at day 13, the percentage of CD3+, CD3+CD4+, CD3+CD8+ and CD3+CD56+ T cells in DC-CIKs was significantly improved 1.02, 1.79, 1.26 and 2.44-fold, respectively. In addition, trypan blue staining analysis demonstrated that the cell viability of CIKs and DC-CIKs was 96% and 98%, respectively. Furthermore, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) analysis verified that CIK and DC-CIK cytotoxicity in Hela cells was 58% and 80%, respectively, with a significant difference. Taken together, our results indicate that the cell proliferation, cell phenotype and antitumor activity of CIKs were all enhanced following co-culture with DCs in vitro. These results are likely to be useful for DC-CIK application in antitumor therapies. PMID:27602134

  17. Influence of autologous dendritic cells on cytokine-induced killer cell proliferation, cell phenotype and antitumor activity in vitro

    PubMed Central

    Cao, Jingsong; Chen, Cong; Wang, Yuhuan; Chen, Xuecheng; Chen, Zeying; Luo, Xiaoling

    2016-01-01

    Dendritic cell (DCs) are essential antigen processing and presentation cells that play a key role in the immune response. In this study, DCs were co-cultured with cytokine-induced killer cells (DC-CIKs) in vitro to detect changes in cell proliferation, cell phenotype and cell cytotoxicity. The results revealed that the DCs were suitable for co-culture with CIKs at day 7, and that cell quantity of DC-CIKs was lower than that of CIKs until day 11, but it was significantly improved to 1.17-fold that of CIKs at day 13. Flow cytometry was used to detect the cell phenotype of CIKs and DC-CIKs. Compared with CIKs at day 13, the percentage of CD3+, CD3+CD4+, CD3+CD8+ and CD3+CD56+ T cells in DC-CIKs was significantly improved 1.02, 1.79, 1.26 and 2.44-fold, respectively. In addition, trypan blue staining analysis demonstrated that the cell viability of CIKs and DC-CIKs was 96% and 98%, respectively. Furthermore, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) analysis verified that CIK and DC-CIK cytotoxicity in Hela cells was 58% and 80%, respectively, with a significant difference. Taken together, our results indicate that the cell proliferation, cell phenotype and antitumor activity of CIKs were all enhanced following co-culture with DCs in vitro. These results are likely to be useful for DC-CIK application in antitumor therapies.

  18. Enhancing effects of agelasphin-11 on natural killer cell activities of normal and tumor-bearing mice.

    PubMed

    Kobayashi, E; Motoki, K; Natori, T; Uchida, T; Fukushima, H; Koezuka, Y

    1996-03-01

    Agelasphin-11 (AGL-11), a novel alpha-galactosylceramide isolated from an extract of a marine sponge, Agelas mauritianus, markedly prolonged the life span of mice intraperitoneally inoculated with B16 cells. Since AGL-11 did not show any direct cytotoxic activity against B16 cells, this compound is considered to be a biological response modifier (BRM). We focused on the enhancing effect of this compound on in vivo natural killer (NK) cell activity because several BRMs have already been determined to enhance the in vivo natural killer (NK) cell activity. When we evaluated the enhancing activity of AGL-11 using normal mice, AGL-11 enhanced in vivo NK cell activity more potently than Poly I:C, which is a positive control. In addition, we examined the effect of this compound on the NK cell activity of tumor-bearing mice, and found that AGL-11 recovers the reduced NK cell activity in a tumor-bearing condition to a higher level than that of normal mice. These results suggest that AGL-11 shows antitumor activity by the activation of antitumor effector cells such as NK cells.

  19. Effect of Helixor A on Natural Killer Cell Activity in Endometriosis

    PubMed Central

    Jeung, In-Cheul; Chung, Youn-Jee; Chae, Boah; Kang, So-Yeon; Song, Jae-Yen; Jo, Hyun-Hee; Lew, Young-Ok; Kim, Jang-Heub; Kim, Mee-Ran

    2015-01-01

    Background and Aim: NK cells are one of the major immune cells in endometriosis pathogenesis. While previous clinical studies have shown that helixor A to be an effective treatment for endometriosis, little is known about its mechanism of action, or its relationship with immune cells. The aim of this study is to investigate the effects of helixor A on Natural killer cell (NK cell) cytotoxicity in endometriosis Materials and Methods: We performed an experimental study. Samples of peritoneal fluid were obtained from January 2011 to December 2011 from 50 women with endometriosis and 50 women with other benign ovarian cysts (control). Peritoneal fluid of normal control group and endometriosis group was collected during laparoscopy. Baseline cytotoxicity levels of NK cells were measured with the peritoneal fluid of control group and endometriosis group. Next, cytotoxicity of NK cells was evaluated before and after treatment with helixor A. NK-cell activity was determined based upon the expression of CD107a, as an activation marker. Results: NK cells cytotoxicity was 79.38±2.13% in control cells, 75.55±2.89% in the control peritoneal fluid, 69.59±4.96% in endometriosis stage I/II endometriosis, and 63.88±5.75% in stage III/IV endometriosis. A significant difference in cytotoxicity was observed between the control cells and stage III/IV endometriosis, consistent with a significant decrease in the cytotoxicity of NK cells in advanced stages of endometriosis; these levels increased significantly after treatment with helixor A; 78.30% vs. 86.40% (p = 0.003) in stage I/II endometriosis, and 73.67% vs. 84.54% (p = 0.024) in stage III/IV. The percentage of cells expressing CD107a was increased significantly in each group after helixor A treatment; 0.59% vs. 1.10% (p = 0.002) in stage I/II endometriosis, and 0.79% vs. 1.40% (p = 0.014) in stage III/IV. Conclusions: Helixor A directly influenced NK-cell cytotoxicity through direct induction of CD107a expression. Our results

  20. Posttransplant adoptive immunotherapy with activated natural killer cells in patients with metastatic breast cancer.

    PubMed

    deMagalhaes-Silverman, M; Donnenberg, A; Lembersky, B; Elder, E; Lister, J; Rybka, W; Whiteside, T; Ball, E

    2000-01-01

    Relapse after high-dose chemotherapy is the main cause of therapeutic failure in patients with metastatic breast cancer. Adoptive immunotherapy with interleukin-2 (IL-2) plus activated natural killer cells may eliminate residual disease without excessive toxicity. The authors sought to determine if immunotherapy immediately after transplantation would affect engraftment and the toxicity associated with transplantation. Fifteen consecutive patients with metastatic breast cancer were allocated to three cohorts. Cohort 1 (five patients) received high-dose cyclophosphamide, thiotepa, and carboplatin (CTCb) followed by peripheral blood stem cell infusion and granulocyte colony-stimulating factor at 10 micrograms/kg. Cohort 2 (five patients) received in addition rhIL-2 (2 x 10(6) IU/m2/day) for 4 days intravenously via continuous infusion after peripheral blood stem cell infusion. In cohort 3 (five patients), peripheral blood stem cell transplant was followed by infusion of autologous activated NK cells and rhIL-2 (2 x 10(6) IU/m2/day) for 4 days (via continuous intravenous infusion). Generation of activated NK cells was possible in all patients in cohort 3. All patients has successful engraftment. Median time to absolute neutrophil count more than 0.5 x 10(9)/L was 8 days (range, 8 to 11 days) in cohort 1, 9 days (range, 7 to 11 days) in cohort 2, and 9 days (range, 8 to 9 days) in cohort 3. Median time until the platelet count was more than 20 x 10(9)/L was 14 days (range, 9 to 22 days) in cohort 1, 11 days (range, 6 to 14 days) in cohort 2, and 12 days (range, 11 to 21 days) in cohort 3. All patients developed neutropenic fevers, but the overall toxicity associated with the infusion of IL-2 (cohort 2) or IL-2 plus activated NK cells (cohort 3) did not differ from that observed in cohort 1. Complete responses were achieved in one patient in cohort 1, in two patients in cohort 2, and in one patient in cohort 3. In conclusion, post-transplant adoptive immunotherapy with

  1. IL-2-Activated Killer Cells and Native Cytokines in Treatment of Patients with Advanced Cancer.

    PubMed

    Ostanin, Alexander A.; Chernykh, Helena R.; Leplina, Olga Y.; Shevela, Ekaterina Ya.; Niconov, Sergey D.; Kozlov, Vladimir A.

    1997-12-01

    We evaluated the efficiency/tolerability of and the immunological changes induced by the adoptive immunotherapy (AIT) with IL-2-activated killer cells, and preparation of native cytokines from swine spleen (PSS) in treatment of 20 patients with advanced cancer (10 patients with primary lung cancer; 3 with metastatic melanoma; 2 with advanced neuroblastoma; 2 with ovarian cancer; renal cancer; gastric adenocarcinoma; and colorectal cancer). The partial/minor response of duration period 2-10 months was observed in 20% of patients. 2/4 patients, who underwent partial surgical tumor resection and following AIT course, sustained the event-free survival for more than 24 months. The response to the therapy was revealed in 4/10 patients with lung cancer, 2/2 patients with neuroblastoma, of whom each had ovarian and colorectal cancers. The evaluation of a dose of infused LAKcells as well as combined i.v./local (endobronchial or endoperitoneal) LAK administration were necessary to assure positive response in patients. The cytokine and/or side effects were moderate and the combined LAK-PSS infusions were generally well tolerated by the patients. The treatment was followed by activation of the patient immune system that included: (i) rebound in amount of peripheral blood lymphocytes; (ii) gain in amount of CD3(+) T cells and those CD4(+) helper/inducer; (iii) enchantment of lymphocyte proliferation and cytokine production (IL-2, IL-1, TNF-alpha). Being injected to patients in combination with LAK cells, cytokines related to PSS action and/or those, either exogenous or secondary, and released by in vitro and in vivo, activated lymphocytes and could cause the therapeutic effects.

  2. Enhancement of anti-tumor activity of natural killer cells by BALL-1, a B cell lymphoma line.

    PubMed

    Hirashima, M; Yoshida, N; Seki, M; Okada, H; Takamura, S; Mihara, Y

    1998-04-01

    The anti-tumor activity of human peripheral blood mononuclear cells (PBMC) against various tumor cell line cells (K562, Daudi, KMG-2, and KATOIII) was enhanced by coculture with irradiated BALL-1, but not with other irradiated B cell line cells (NALM-1, Namalwa, and Daudi). PBMC cocultured with BALL-1, however, failed to exhibit evident cytotoxicity against autologous concanavalin A-induced lymphoblasts. The enhancement of the anti-tumor activity seemed not to be correlated with EBNA and HLA-DR expression on B cell line cells. Monoclonal antibodies (mAbs) against interleukin (IL)-2, interferon-gamma, IL-12, IL-15, tumor necrosis factor-alpha and lymphotoxin showed little or no suppression of the anti-tumor activity of PBMC treated with irradiated BALL-1. Furthermore, the culture supernatants of BALL-1 failed to enhance the anti-tumor activity of PBMC, suggesting no involvement of soluble factors in the induction of the anti-tumor activity. The anti-tumor activity of PBMC treated with BALL-1 was synergistically enhanced by an additional IL-2 stimulation. Periodate-lysine-paraformaldehyde-fixed, but not ethanol- or acetone-fixed, BALL-1 could significantly enhance the anti-tumor activity. Furthermore, BALL-1-derived membrane fraction, but not that of Daudi, enhances the anti-tumor activity. It was thus suggested that some membrane glycoproteins on the cell surface of BALL-1 play a crucial role in the induction of the anti-tumor activity. By analysis using mAbs against human leukocytes, we found that depletion of CD11b, CD16, and CD56-positive cells resulted in decreased anti-tumor activity, suggesting that the main effector cells in the BALL-1-induced anti-tumor activity were natural killer (NK) cells. The present results thus raise the possibility that BALL-1, probably via membrane glycoproteins, modulates NK cell-mediated anti-tumor activity. PMID:9617349

  3. The Human Natural Killer Cell Immune Synapse

    NASA Astrophysics Data System (ADS)

    Davis, Daniel M.; Chiu, Isaac; Fassett, Marlys; Cohen, George B.; Mandelboim, Ofer; Strominger, Jack L.

    1999-12-01

    Inhibitory killer Ig-like receptors (KIR) at the surface of natural killer (NK) cells induced clustering of HLA-C at the contacting surface of target cells. In this manner, inhibitory immune synapses were formed as human NK cells surveyed target cells. At target/NK cell synapses, HLA-C/KIR distributed into rings around central patches of intercellular adhesion molecule-1/lymphocyte function-associated antigen-1, the opposite orientation to mature murine T cell-activating synapses. This organization of protein was stable for at least 20 min. Cells could support multiple synapses simultaneously, and clusters of HLA-C moved as NK cells crawled over target cells. Clustering required a divalent metal cation, explaining how metal chelators inhibit KIR function. Surprisingly, however, formation of inhibitory synapses was unaffected by ATP depletion and the cytoskeletal inhibitors, colchicine and cytochalsins B and D. Clearly, supramolecular organization within plasma membranes is critical for NK cell immunosurveillance.

  4. Effect of elevated serum prolactin concentrations on cytokine production and natural killer cell activity.

    PubMed

    Clodi, M; Svoboda, T; Kotzmann, H; Deyssig, R; Woloszczuk, W; Zielinski, C C; Luger, A

    1992-12-01

    In vitro and in vivo studies in rodents and human suggested an immunostimulatory effect of prolactin. The aim of the present study was to determine the impact of chronically elevated serum prolactin concentrations on the immune system in patients with prolactinomas. For this purpose parameters of the humoral and cellular immune system were studied in seven patients with prolactinomas on two occasions (1) when their serum prolactin concentration had been normalized through treatment with dopamine agonists and (2) when their serum prolactin concentration was high. Serum concentrations of immunoglobulines, interleukin 1, 3 and 6, TNF-alpha, interferon-gamma and the soluble interleukin 2 receptor, leukocyte subsets and the natural killer cell activity were found to be within the normal range on both occasions, i.e. at normal and at high serum prolactin concentrations. The assumption could be made that long-lasting elevation of serum prolactin concentration induces adaptive changes when the acute stimulatory effects of prolactin on several parameters of the immune system have subsided.

  5. Alzheimer caregiver stress: basal natural killer cell activity, pituitary-adrenal cortical function, and sympathetic tone.

    PubMed

    Irwin, M; Hauger, R; Patterson, T L; Semple, S; Ziegler, M; Grant, I

    1997-01-01

    The association between Alzheimer caregiving and natural killer (NK) cell activity and basal plasma levels of adrenocorticotropic hormone (ACTH), cortisol, beta-endorphin, prolactin, epinephrine, norepinephrine, and neuropeptide Y was determined in 100 spousal Alzheimer caregivers and 33 age- and gender-comparable control volunteers upon intake into a study of the psychological and physiologic impact of caregiving. The relationship between these physiologic measures and individual characteristics such as age, gender, medical status, severity of stress, severity of depressive symptoms, and caregiver burden was tested. In addition, the association between NK activity and alterations of the neuroendocrine measures was investigated. As compared to controls, the Alzheimer caregivers had similar levels of NK activity and of basal plasma neuroendocrine hormones and sympathetic measures. While older age and male gender status were associated with increased levels of ACTH, neither medical caseness, severity of life stress, nor severity of depressive symptoms was associated with alterations in any of the multiple physiologic domains. Classification of Alzheimer caregiver burden identified caregivers who were mismatched in terms of the amount of care they were required to provide and the amount of respite time received. The mismatched caregivers had significantly higher basal plasma ACTH but no change in other physiological measures, as compared to non-mismatched caregivers. NK activity was negatively correlated with plasma levels of neuropeptide Y but not with any of the other neuroendocrine measures. Based on this cross-sectional evaluation of NK activity and neuroendocrine and sympathetic measures, we conclude that most Alzheimer caregivers do not show evidence of altered basal physiology.

  6. 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.

  7. Immunobiology of natural killer cells. Volume II

    SciTech Connect

    Lotzova, E.; Herberman, R.B.

    1986-01-01

    This book provides a review of natural killer (NK) cell-mediated immunity in humans and experimental animal system. Topics for the volume include: In vivo activities of NK cells against primary and metastatic tumors in experimental animals; involvement of NK cells in human malignant disease; impaired NK cell profile in leukemia patients; in vivo modulation of NK activity in cancer patients; implications of aberrant NK cell activity in nonmalignant, chronic diseases; NK cell role in regulation of the growth and functions of hemopoietic and lymphoid cells; NK cells active against viral, bacterial, protozoan, and fungal infections; cytokine secretion and noncytotoxic functions of human large granular lymphocytes; augmentation of NK activity; regulation of NK cell activity by suppressor cells; NK cell cloning technology and characteristics of NK cell clones; comparison of antibody-dependent cellular cytotoxicity (ADCC) and NK activity, and index.

  8. Bovine CD2-/NKp46+ cells are fully functional natural killer cells with a high activation status

    PubMed Central

    Boysen, Preben; Olsen, Ingrid; Berg, Ingvild; Kulberg, Siri; Johansen, Grethe M; Storset, Anne K

    2006-01-01

    Background Natural killer (NK) cells in the cow have been elusive due to the lack of specific NK cell markers, and various criteria including a CD3-/CD2+ phenotype have been used to identify such cells. The recent characterization of the NK-specific NKp46 receptor has allowed a more precise definition of bovine NK cells. NK cells are known as a heterogeneous cell group, and we here report the first functional study of bovine NK cell subsets, based on the expression of CD2. Results Bovine CD2- NK cells, a minor subset in blood, proliferated more rapidly in the presence of IL-2, dominating the cultures after a few days. Grown separately with IL-2, CD2- and CD2+ NK cell subsets did not change CD2 expression for at least two weeks. In blood, CD2- NK cells showed a higher expression of CD44 and CD25, consistent with a high activation status. A higher proportion of CD2- NK cells had intracellular interferon-gamma in the cytoplasm in response to IL-2 and IL-12 stimulation, and the CD2- subset secreted more interferon-gamma when cultured separately. Cytotoxic capacity was similar in both subsets, and both carried transcripts for the NK cell receptors KIR, CD16, CD94 and KLRJ. Ligation by one out of two tested anti-CD2 monoclonal antibodies could trigger interferon-gamma production from NK cells, but neither of them could alter cytotoxicity. Conclusion These results provide evidence that bovine CD2- as well as CD2+ cells of the NKp46+ phenotype are fully functional NK cells, the CD2- subset showing signs of being more activated in the circulation. PMID:16643649

  9. Cannabinoids increase lung cancer cell lysis by lymphokine-activated killer cells via upregulation of ICAM-1.

    PubMed

    Haustein, Maria; Ramer, Robert; Linnebacher, Michael; Manda, Katrin; Hinz, Burkhard

    2014-11-15

    Cannabinoids have been shown to promote the expression of the intercellular adhesion molecule 1 (ICAM-1) on lung cancer cells as part of their anti-invasive and antimetastatic action. Using lung cancer cell lines (A549, H460) and metastatic cells derived from a lung cancer patient, the present study addressed the impact of cannabinoid-induced ICAM-1 on cancer cell adhesion to lymphokine-activated killer (LAK) cells and LAK cell-mediated cytotoxicity. Cannabidiol (CBD), a non-psychoactive cannabinoid, enhanced the susceptibility of cancer cells to adhere to and subsequently be lysed by LAK cells, with both effects being reversed by a neutralizing ICAM-1 antibody. Increased cancer cell lysis by CBD was likewise abrogated when CBD-induced ICAM-1 expression was blocked by specific siRNA or by antagonists to cannabinoid receptors (CB1, CB2) and to transient receptor potential vanilloid 1. In addition, enhanced killing of CBD-treated cancer cells was reversed by preincubation of LAK cells with an antibody to lymphocyte function associated antigen-1 (LFA-1) suggesting intercellular ICAM-1/LFA-1 crosslink as crucial event within this process. ICAM-1-dependent pro-killing effects were further confirmed for the phytocannabinoid Δ(9)-tetrahydrocannabinol (THC) and R(+)-methanandamide (MA), a hydrolysis-stable endocannabinoid analogue. Finally, each cannabinoid elicited no significant increase of LAK cell-mediated lysis of non-tumor bronchial epithelial cells, BEAS-2B, associated with a far less pronounced (CBD, THC) or absent (MA) ICAM-1 induction as compared to cancer cells. Altogether, our data demonstrate cannabinoid-induced upregulation of ICAM-1 on lung cancer cells to be responsible for increased cancer cell lysis by LAK cells. These findings provide proof for a novel antitumorigenic mechanism of cannabinoids.

  10. Lymphokine-activated killer (LAK) cells can be focused at sites of tumor growth by products of macrophage activation

    SciTech Connect

    Migliori, R.J.; Gruber, S.A.; Sawyer, M.D.; Hoffman, R.; Ochoa, A.; Bach, F.H.; Simmons, R.L.

    1987-08-01

    Successful adoptive cancer immunotherapy presumably depends on the accumulation of tumoricidal leukocytes at the sites of tumor growth. Large numbers of lymphokine-activated killer (LAK) cells can be generated in vitro by growth in high concentrations of interleukin-2 (IL-2), but relatively few arrive at the tumor site after intravenous injection. We hypothesize that the delivery of LAK cells to tumor sites may be augmented by previously demonstrated lymphocyte-recruiting factors, including activated macrophage products such as interleukin-1 (IL-1) and tumor necrosis factor. /sup 111/Indium-labeled LAK cells were injected intravenously into syngeneic mice bearing the macrophage activator endotoxin (LPS) in one hind footpad, and saline solution was injected into the contralateral footpad. Significantly more activity was recovered from the LPS-bearing footpad at all times during a 96-hour period. Recombinant IL-1 also attracted more LAK cells after injection into tumor-free hind footpads. Furthermore, LAK cells preferentially homed to hind footpads that were bearing 3-day established sarcomas after intralesional injections of LPS, IL-1, or tumor necrosis factor when compared with contralateral tumor-bearing footpads injected with saline solution alone. In preliminary experiments, mice with hind-footpad tumors appeared to survive longer after combined systemic IL-2 and LAK therapy if intralesional LPS was administered. These studies demonstrate that macrophage activation factors that have been shown capable of attracting circulating normal lymphocytes can also effectively attract LAK cells from the circulation. By the stimulation of macrophages at the sites of tumor growth, more LAK cells can be attracted. It is hoped that by focusing the migration of LAK cells to tumors, LAK cells and IL-2 would effect tumor regression more efficiently and with less toxicity.

  11. Efficient Killing of High Risk Neuroblastoma Using Natural Killer Cells Activated by Plasmacytoid Dendritic Cells

    PubMed Central

    Cordeau, Martine; Belounis, Assila; Lelaidier, Martin; Cordeiro, Paulo; Sartelet, Hervé; Duval, Michel

    2016-01-01

    High-risk neuroblastoma (NB) remains a major therapeutic challenge despite the recent advent of disialoganglioside (GD2)-antibody treatment combined with interleukin (IL)-2 and granulocyte monocyte-colony stimulating factor (GM-CSF). Indeed, more than one third of the patients still die from this disease. Here, we developed a novel approach to improve the current anti-GD2 immunotherapy based on NK cell stimulation using toll-like receptor (TLR)-activated plasmacytoid dendritic cells (pDCs). We demonstrated that this strategy led to the efficient killing of NB cells. When the expression of GD2 was heterogeneous on NB cells, the combination of pDC-mediated NK-cell activation and anti-GD2 treatment significantly increased the cytotoxicity of NK cells against NB cells. Activation by pDCs led to a unique NK-cell phenotype characterized by increased surface expression of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), with increased expression of CD69 on CD56dim cytotoxic cells, and strong interferon-γ production. Additionally, NB-cell killing was mediated by the TRAIL death-receptor pathway, as well as by the release of cytolytic granules via the DNAX accessory molecule 1 pathway. NK-cell activation and lytic activity against NB was independent of cell contact, depended upon type I IFN produced by TLR-9-activated pDCs, but was not reproduced by IFN-α stimulation alone. Collectively, these results highlighted the therapeutic potential of activated pDCs for patients with high-risk NB. PMID:27716850

  12. Cytotoxic T lymphocytes and natural killer cells display impaired cytotoxic functions and reduced activation in patients with alcoholic hepatitis.

    PubMed

    Støy, Sidsel; Dige, Anders; Sandahl, Thomas Damgaard; Laursen, Tea Lund; Buus, Christian; Hokland, Marianne; Vilstrup, Hendrik

    2015-02-15

    The dynamics and role of cytotoxic T lymphocytes (CTLs), natural killer (NK) cells, and NKT cells in the life-threatening inflammatory disease alcoholic hepatitis is largely unknown. These cells directly kill infected and damaged cells through, e.g., degranulation and interferon-γ (IFNγ) production, but cause tissue damage if overactivated. They also assist tissue repair via IL-22 production. We, therefore, aimed to investigate the frequency, functionality, and activation state of such cells in alcoholic hepatitis. We analyzed blood samples from 24 severe alcoholic hepatitis patients followed for 30 days after diagnosis. Ten healthy abstinent volunteers and 10 stable abstinent alcoholic cirrhosis patients were controls. Using flow cytometry we assessed cell frequencies, NK cell degranulation capacity following K562 cell stimulation, activation by natural killer group 2 D (NKG2D) expression, and IL-22 and IFNγ production. In alcoholic hepatitis we found a decreased frequency of CTLs compared with healthy controls (P < 0.001) and a similar trend for NK cells (P = 0.089). The NK cell degranulation capacity was reduced by 25% compared with healthy controls (P = 0.02) and by 50% compared with cirrhosis patients (P = 0.04). Accordingly, the NKG2D receptor expression was markedly decreased on NK cells, CTLs, and NKT cells (P < 0.05, all). The frequencies of IL-22-producing CTLs and NK cells were doubled compared with healthy controls (P < 0.05, all) but not different from cirrhosis patients. This exploratory study for the first time showed impaired cellular cytotoxicity and activation in alcoholic hepatitis. This is unlikely to cause hepatocyte death but may contribute toward the severe immune incompetence. The results warrant detailed and mechanistic studies.

  13. Innate and cytokine-driven signals, rather than microbial antigens, dominate in natural killer T cell activation during microbial infection

    PubMed Central

    Tatituri, Raju V.V.; Watts, Gerald F.M.; Bhowruth, Veemal; Leadbetter, Elizabeth A.; Barton, Nathaniel; Cohen, Nadia R.; Hsu, Fong-Fu; Besra, Gurdyal S.

    2011-01-01

    Invariant natural killer T cells (iNKT cells) are critical for host defense against a variety of microbial pathogens. However, the central question of how iNKT cells are activated by microbes has not been fully explained. The example of adaptive MHC-restricted T cells, studies using synthetic pharmacological α-galactosylceramides, and the recent discovery of microbial iNKT cell ligands have all suggested that recognition of foreign lipid antigens is the main driver for iNKT cell activation during infection. However, when we compared the role of microbial antigens versus innate cytokine-driven mechanisms, we found that iNKT cell interferon-γ production after in vitro stimulation or infection with diverse bacteria overwhelmingly depended on toll-like receptor–driven IL-12. Importantly, activation of iNKT cells in vivo during infection with Sphingomonas yanoikuyae or Streptococcus pneumoniae, pathogens which are known to express iNKT cell antigens and which require iNKT cells for effective protection, also predominantly depended on IL-12. Constitutive expression of high levels of IL-12 receptor by iNKT cells enabled instant IL-12–induced STAT4 activation, demonstrating that among T cells, iNKT cells are uniquely equipped for immediate, cytokine-driven activation. These findings reveal that innate and cytokine-driven signals, rather than cognate microbial antigen, dominate in iNKT cell activation during microbial infections. PMID:21555485

  14. In vivo glucocorticoid effects on porcine natural killer cell activity and circulating leukocytes.

    PubMed

    Salak-Johnson, J L; McGlone, J J; Norman, R L

    1996-03-01

    Porcine natural killer (NK) cell cytotoxicity, plasma cortisol, total white blood cells (WBC), neutrophil:lymphocyte ratio (N:L), and circulating blood leukocytes were examined from pigs injected i.v. with either saline, ACTH, cortisol, or treated with metyrapone. Plasma cortisol increased (P < .05) after ACTH and cortisol treatments and decreased (P < .05) after metyrapone treatment; thus, treatments had the intended effects on in vivo cortisol concentrations. In Exp. 1, pigs were injected with either saline or ACTH at 0600 after the initial blood samples were taken (time 0). The ACTH had no effect (P > .10) on NK cytotoxicity. Pigs injected i.v. with ACTH had fewer lymphocytes and more neutrophils (P < .05) than control pigs. The N:L ratio was greater (P < .05) among ACTH-injected than among control pigs. In Exp. 2, pigs were injected i.v. with either saline or 40 or 400 micrograms of cortisol at 0600 after the initial blood samples were obtained (time 0). Cortisol at 40 micrograms had no effect (P > .10) on NK cytotoxicity. However, a 400-micrograms bolus of cortisol reduced (P < .05) NK cytotoxicity (control = 39.5, cortisol = 28.3% cytotoxicity, SEM = 3.7). Each dose of cortisol reduced (P < .05) circulating blood lymphocyte numbers. In Exp. 3, pigs were fed 1 g of metyrapone or no metyrapone the night before sampling. Blood samples were obtained at 0600, 0700, and 0800. Metyrapone reduced (P < .05) NK cytotoxicity (control = 28.6, metyrapone = 11.8%, SEM = 1.9). Pigs treated with metyrapone had greater (P < .05) numbers of neutrophils than control pigs. Numbers of lymphocytes were greater (P < .05) among control than among treated pigs. Pigs treated with metyrapone had a greater (P < .05) N:L ratio than control pigs. In conclusion, normal physiological concentrations or moderately increased blood cortisol concentrations did not influence NK activity, although leukocyte distributions were changed. We conclude that greatly increased or greatly decreased

  15. Natural killer T cells in adipose tissue are activated in lean mice.

    PubMed

    Kondo, Taisuke; Toyoshima, Yujiro; Ishii, Yoshiyuki; Kyuwa, Shigeru

    2013-01-01

    Adipose tissues are closely connected with the immune system. It has been suggested that metabolic syndromes such as type 2 diabetes, arteriosclerosis and liver steatosis can be attributed to adipose tissue inflammation characterized by macrophage infiltration. To understand a physiological and pathological role of natural killer T (NKT) cells on inflammation in adipose tissue, we characterized a subset of NKT cells in abdominal and subcutaneous adipose tissues in C57BL/6J mice fed normal or high-fat diets. NKT cells comprised a larger portion of lymphocytes in adipose tissues compared with the spleen and peripheral blood, with epididymal adipose tissue having the highest number of NKT cells. Furthermore, some NKT cells in adipose tissues expressed higher levels of CD69 and intracellular interferon-γ, whereas the Vβ repertoires of NKT cells in adipose tissues were similar to other cells. In obese mice fed a high-fat diet, adipose tissue inflammation had little effect on the Vβ repertoire of NKT cells in epididymal adipose tissues. We speculate that the NKT cells in adipose tissues may form an equivalent subset in other tissues and that these subsets are likely to participate in adipose tissue inflammation. Additionally, the high expression level of CD69 and intracellular IFN-γ raises the possibility that NKT cells in adipose tissue may be stimulated by some physiological mechanism.

  16. Modulation of in vitro porcine natural killer cell activity by recombinant interleukin-1 alpha, interleukin-2 and interleukin-4.

    PubMed Central

    Knoblock, K F; Canning, P C

    1992-01-01

    In order to understand better how cytokines modulate porcine lymphocyte-mediated natural cytotoxicity and to develop a rapid and reliable colorimetric assay to study that activity in young pigs, we studied inherent and cytokine induced in vitro natural killer (NK) activity. The cytokines we studied were human recombinant interleukin-1 alpha (IL-1 alpha), IL-2, IL-4 and interferon-gamma (IFN-gamma). Natural killer activity by peripheral blood mononuclear cells (PBMC), reported as per cent specific lysis (%SL), was determined by the colorimetric measurement of lactate dehydrogenase released from tumour cell targets, YAC-1 and K562. Inherent NK activity was low and remained relatively unchanged by alterations of assay length or effector cell concentration. Low NK activity was also observed in response to IL-4 and IFN-gamma. IL-2 and, to a lesser extent, IL-1 alpha induced significant NK activity with trends towards increasing %SL with increasing cytokine dose. Optimal IL-1 alpha- and IL-2-induced NK activity could be observed at 18 hr, with significant activity stimulated by IL-2 as early as 4 hr. IL-2-induced NK activity was sensitive to effector cell concentration; %SL decreased as the effector to target ratio decreased. IL-1 alpha- and IL-2-induced NK activities were decreased in the presence of IL-4. These results indicate porcine PBMC are sensitive to in vitro modulation by human recombinant IL-1 alpha, IL-2 and IL-4. The ability of IL-1 alpha and IL-2 to induce swine NK activity and the ability of IL-4 to inhibit that activity are similar to the actions of those cytokines in human NK systems. PMID:1634252

  17. Conditioned enhancement of natural killer cell activity, but not interferon, with camphor or saccharin-LiCl conditioned stimulus.

    PubMed

    Ghanta, V K; Hiramoto, N S; Solvason, H B; Tyring, S K; Spector, N H; Hiramoto, R N

    1987-01-01

    Pavlovian conditioning of the natural killer (NK) cell response has been demonstrated by pairing camphor with polyinosinic:polycytidylic acid (poly I:C) in nine association trials. The NK cell response could be conditioned also by using combined saccharin and lithium chloride (LiCl) as the conditioned stimulus. The camphor and saccharin-LiCl paradigms were tested to determine if the conditioned NK cell activity was the result of conditioning of the interferon response. Interferon levels were measured at 6 hr and NK cell activity at 24 hr after application of the conditioned stimulus. The interferon levels measured in separate experiments were not uniformly elevated in conditioned animals compared with controls.

  18. Killer Cell Lectin-like Receptor G1 Inhibits NK Cell Function through Activation of Adenosine 5'-Monophosphate-Activated Protein Kinase.

    PubMed

    Müller-Durovic, Bojana; Lanna, Alessio; Polaco Covre, Luciana; Mills, Rachel S; Henson, Sian M; Akbar, Arne N

    2016-10-01

    NK cells are the first line of defense against infected and transformed cells. Defective NK cell activity was shown to increase susceptibility for viral infections and reduce tumor immune-surveillance. With age, the incidence of infectious diseases and malignancy rises dramatically, suggesting that impaired NK cell function might contribute to disease in these individuals. We found an increased frequency of NK cells with high expression of the inhibitory killer cell lectin-like receptor G1 (KLRG1) in individuals >70 y. The role of KLRG1 in ageing is not known, and the mechanism of KLRG1-induced inhibition of NK cell function is not fully understood. We report that NK cells with high KLRG1 expression spontaneously activate the metabolic sensor AMP-activated protein kinase (AMPK) and that activation of AMPK negatively regulates NK cell function. Pre-existing AMPK activity is further amplified by ligation of KLRG1 in these cells, which leads to internalization of the receptor and allows interaction with AMPK. We show that KLRG1 activates AMPK by preventing its inhibitory dephosphorylation by protein phosphatase-2C rather than inducing de novo kinase activation. Finally, inhibition of KLRG1 or AMPK prevented KLRG1-induced activation of AMPK and reductions in NK cell cytotoxicity, cytokine secretion, proliferation, and telomerase expression. This novel signaling pathway links metabolic sensing, effector function, and cell differentiation with inhibitory receptor signaling that may be exploited to enhance NK cell activity during ageing.

  19. Targeting natural killer cells in cancer immunotherapy.

    PubMed

    Guillerey, Camille; Huntington, Nicholas D; Smyth, Mark J

    2016-08-19

    Alteration in the expression of cell-surface proteins is a common consequence of malignant transformation. Natural killer (NK) cells use an array of germline-encoded activating and inhibitory receptors that scan for altered protein-expression patterns, but tumor evasion of detection by the immune system is now recognized as one of the hallmarks of cancer. NK cells display rapid and potent immunity to metastasis or hematological cancers, and major efforts are now being undertaken to fully exploit NK cell anti-tumor properties in the clinic. Diverse approaches encompass the development of large-scale NK cell-expansion protocols for adoptive transfer, the establishment of a microenvironment favorable to NK cell activity, the redirection of NK cell activity against tumor cells and the release of inhibitory signals that limit NK cell function. In this Review we detail recent advances in NK cell-based immunotherapies and discuss the advantages and limitations of these strategies. PMID:27540992

  20. Functional advantage of educated KIR2DL1(+) natural killer cells for anti-HIV-1 antibody-dependent activation.

    PubMed

    Gooneratne, S L; Center, R J; Kent, S J; Parsons, M S

    2016-04-01

    Evidence from the RV144 HIV-1 vaccine trial implicates anti-HIV-1 antibody-dependent cellular cytotoxicity (ADCC) in vaccine-conferred protection from infection. Among effector cells that mediate ADCC are natural killer (NK) cells. The ability of NK cells to be activated in an antibody-dependent manner is reliant upon several factors. In general, NK cell-mediated antibody-dependent activation is most robust in terminally differentiated CD57(+) NK cells, as well as NK cells educated through ontological interactions between inhibitory killer immunoglobulin-like receptors (KIR) and their major histocompatibility complex class I [MHC-I or human leucocyte antigen (HLA-I)] ligands. With regard to anti-HIV-1 antibody-dependent NK cell activation, previous research has demonstrated that the epidemiologically relevant KIR3DL1/HLA-Bw4 receptor/ligand combination confers enhanced activation potential. In the present study we assessed the ability of the KIR2DL1/HLA-C2 receptor/ligand combination to confer enhanced activation upon direct stimulation with HLA-I-devoid target cells or antibody-dependent stimulation with HIV-1 gp140-pulsed CEM.NKr-CCR5 target cells in the presence of an anti-HIV-1 antibody source. Among donors carrying the HLA-C2 ligand for KIR2DL1, higher interferon (IFN)-γ production was observed within KIR2DL1(+) NK cells than in KIR2DL1(-) NK cells upon both direct and antibody-dependent stimulation. No differences in KIR2DL1(+) and KIR2DL1(-) NK cell activation were observed in HLA-C1 homozygous donors. Additionally, higher activation in KIR2DL1(+) than KIR2DL1(-) NK cells from HLA-C2 carrying donors was observed within less differentiated CD57(-) NK cells, demonstrating that the observed differences were due to education and not an overabundance of KIR2DL1(+) NK cells within differentiated CD57(+) NK cells. These observations are relevant for understanding the regulation of anti-HIV-1 antibody-dependent NK cell responses.

  1. The role of natural killer cells in chronic myeloid leukemia

    PubMed Central

    Danier, Anna Carolyna Araújo; de Melo, Ricardo Pereira; Napimoga, Marcelo Henrique; Laguna-Abreu, Maria Theresa Cerávolo

    2011-01-01

    Chronic myeloid leukemia is a neoplasia resulting from a translocation between chromosomes 9 and 22 producing the BCR-ABL hybrid known as the Philadelphia chromosome (Ph). In chronic myeloid leukemia a proliferation of malignant myeloid cells occurs in the bone marrow due to excessive tyrosine kinase activity. In order to maintain homeostasis, natural killer cells, by means of receptors, identify the major histocompatibility complex on the surface of tumor cells and subsequently induce apoptosis. The NKG2D receptor in the natural killer cells recognizes the transmembrane proteins related to major histocompatibility complex class I chain-related genes A and B (MICA and MICB), and it is by the interaction between NKG2D and MICA that natural killer cells exert cytotoxic activity against chronic myeloid leukemia tumor cells. However, in the case of chronic exposure of the NKG2D receptor, the MICA ligand releases soluble proteins called sMICA from the tumor cell surface, which negatively modulate NKG2D and enable the tumor cells to avoid lysis mediated by the natural killer cells. Blocking the formation of sMICA may be an important antitumor strategy. Treatment using tyrosine kinase inhibitors induces modulation of NKG2DL expression, which could favor the activity of the natural killer cells. However this mechanism has not been fully described in chronic myeloid leukemia. In the present study, we analyze the role of natural killer cells to reduce proliferation and in the cellular death of tumor cells in chronic myeloid leukemia. PMID:23049299

  2. Unusual Indolent Course of a Chronic Active Epstein-Barr Virus-Associated Natural Killer Cell Lymphoproliferative Disorder

    PubMed Central

    Al-Riyami, Arwa Z.; Al-Farsi, Khalil; Al-Khabori, Murtadha; Al-Huneini, Mohammed; Al-Hadabbi, Ibrahim

    2016-01-01

    Natural killer (NK) cell lymphoproliferative disorders are uncommon and the Epstein-Barr virus (EBV) plays an important aetiological role in their pathogenesis. We report a 20-year-old male with a chronic active EBV infection associated with a NK cell lymphoproliferative disorder which had an unusual indolent course. He presented to the Sultan Qaboos University Hospital in Muscat, Oman, in December 2011 with a history of intermittent fever and coughing. Examinations revealed generalised lymphadenopathy, hepatosplenomegaly, leukocytosis, transaminitis, diffuse bilateral lung infiltrates and bone marrow lymphocyte involvement. A polymerase chain reaction (PCR) test revealed a high EBV viral load in the peripheral blood cells. The patient received a course of piperacillin-tazobactam for Klebsiella pneumoniae, but no active treatment for the lymphoproliferative disorder. However, his lymphocyte count, serum lactate dehydrogenase and liver enzymes dropped spontaneously. In addition, EBV PCR copies fluctuated and then decreased significantly. He remained clinically asymptomatic over the following four years. PMID:27226916

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

    PubMed Central

    El-Sherbiny, Y M; Holmes, T D; Wetherill, L F; Black, E V I; Wilson, E B; Phillips, S L; Scott, G B; Adair, R A; Dave, R; Scott, K J; Morgan, R S M; Coffey, M; Toogood, G J; Melcher, A A; Cook, G P

    2015-01-01

    Human natural killer (NK) cells play an important role in anti-viral immunity. However, studying their activation kinetics during infection is highly problematic. A clinical trial of a therapeutic virus provided an opportunity to study human NK cell activation in vivo in a controlled manner. Ten colorectal cancer patients with liver metastases received between one and five doses of oncolytic reovirus prior to surgical resection of their tumour. NK cell surface expression of the interferon-inducible molecules CD69 and tetherin peaked 24–48 h post-infection, coincident with a peak of interferon-induced gene expression. The interferon response and NK cell activation were transient, declining by 96 h post-infection. Furthermore, neither NK cell activation nor the interferon response were sustained in patients undergoing multiple rounds of virus treatment. These results show that reovirus modulates human NK cell activity in vivo and suggest that this may contribute to any therapeutic effect of this oncolytic virus. Detection of a single, transient peak of activation, despite multiple treatment rounds, has implications for the design of reovirus-based therapy. Furthermore, our results suggest the existence of a post-infection refractory period when the interferon response and NK cell activation are blunted. This refractory period has been observed previously in animal models and may underlie the enhanced susceptibility to secondary infections that is seen following viral infection. PMID:25469725

  4. KillerOrange, a Genetically Encoded Photosensitizer Activated by Blue and Green Light

    PubMed Central

    Bozhanova, Nina G.; Sharonov, George V.; Staroverov, Dmitriy B.; Egorov, Evgeny S.; Ryabova, Anastasia V.; Solntsev, Kyril M.; Mishin, Alexander S.; Lukyanov, Konstantin A.

    2015-01-01

    Genetically encoded photosensitizers, proteins that produce reactive oxygen species when illuminated with visible light, are increasingly used as optogenetic tools. Their applications range from ablation of specific cell populations to precise optical inactivation of cellular proteins. Here, we report an orange mutant of red fluorescent protein KillerRed that becomes toxic when illuminated with blue or green light. This new protein, KillerOrange, carries a tryptophan-based chromophore that is novel for photosensitizers. We show that KillerOrange can be used simultaneously and independently from KillerRed in both bacterial and mammalian cells offering chromatic orthogonality for light-activated toxicity. PMID:26679300

  5. Gut-targeted immunonutrition boosting natural killer cell activity using Saccharomyces boulardii lysates in immuno-compromised healthy elderly subjects.

    PubMed

    Naito, Yasuhiro; Marotta, Francesco; Kantah, Makoto K; Zerbinati, Nicola; Kushugulova, Almagul; Zhumadilov, Zhaxybay; Illuzzi, Nicola; Sapienza, Chiara; Takadanohara, Hiroshi; Kobayashi, Riyichi; Catanzaro, Roberto

    2014-04-01

    The aim of this study was to assess the immunomodulatory effect of KC-1317 (a symbiotic mixture containing Saccharomyces boulardii lysate in a cranberry, colostrum-derived lactoferrin, fragaria, and lactose mixture) supplementation in immune-compromised but otherwise healthy elderly subjects. A liquid formulation of KC-1317 was administered in a randomized controlled trial (RCT) fashion to healthy volunteers (65-79 years) previously selected for low natural killer (NK) cell activity, and this parameter was checked at the completion of the study. A significant improvement in NK cell activity of KC-1317 consumers was observed as compared to placebo at the end of 2 months. Although preliminary, these beneficial immune-modulatory effects of KC-1317 in aged individuals might indicate its employment within a wider age-management strategy. PMID:24059806

  6. Superresolution microscopy reveals nanometer-scale reorganization of inhibitory natural killer cell receptors upon activation of NKG2D.

    PubMed

    Pageon, Sophie V; Cordoba, Shaun-Paul; Owen, Dylan M; Rothery, Stephen M; Oszmiana, Anna; Davis, Daniel M

    2013-07-23

    Natural killer (NK) cell responses are regulated by a dynamic equilibrium between activating and inhibitory receptor signals at the immune synapse (or interface) with target cells. Although the organization of receptors at the immune synapse is important for appropriate integration of these signals, there is little understanding of this in detail, because research has been hampered by the limited resolution of light microscopy. Through the use of superresolution single-molecule fluorescence microscopy to reveal the organization of the NK cell surface at the single-protein level, we report that the inhibitory receptor KIR2DL1 is organized in nanometer-scale clusters at the surface of human resting NK cells. Nanoclusters of KIR2DL1 became smaller and denser upon engagement of the activating receptor NKG2D, establishing an unexpected crosstalk between activating receptor signals and the positioning of inhibitory receptors. These rearrangements in the nanoscale organization of surface NK cell receptors were dependent on the actin cytoskeleton. Together, these data establish that NK cell activation involves a nanometer-scale reorganization of surface receptors, which in turn affects models for signal integration and thresholds that control NK cell effector functions and NK cell development. PMID:23882121

  7. Effects of beta-adrenergic receptor activation, cholera toxin and forskolin on human natural killer cell function.

    PubMed Central

    Whalen, M M; Bankhurst, A D

    1990-01-01

    Membranes from highly purified natural killer (NK) cells were ADP-ribosylated by treatment with cholera toxin (CTX). CTX resulted in a single band of specific 32P incorporation at Mr 43,600. CTX treatment of intact NK cells caused a 9-fold increase in cyclic AMP (cAMP) concentrations. Pretreatment of NK cells with CTX diminished their ability to lyse K562 tumour cells by up to 79%. Forskolin treatment elevated NK cell cAMP levels 8-fold and decreased lysis of K562 cells by up to 45%. Adrenaline and isoprenaline (isoproterenol) both inhibited lysis of K562 cells by approx. 35% and elevated cAMP by at least 2.5-fold, and their inhibition of lysis was reversed by propranolol. These data suggest that the stimulatory guanine-nucleotide-binding protein GS coupled to beta-adrenergic receptors is involved in transducing signals which inhibit NK cell lysis of tumour cells. CTX and forskolin also diminish the ability of NK cells to bind K562 cells (binding is necessary for lysis). This suggests that the NK-cell receptor(s) for the tumour cell may be altered as a consequence of cAMP-mediated events or by activation of GS. Images Fig. 5. PMID:2176460

  8. Metabolic regulation of natural killer cells.

    PubMed

    Finlay, David K

    2015-08-01

    Natural killer (NK) cells have key roles in anti-viral and anti-tumour immune responses. Recent research demonstrates that cellular metabolism is an important determinant for the function of pro-inflammatory immune cells, including activated NK cells. The mammalian target of rapamcyin (mTOR) complex 1 (mTORC1) has been identified as a key metabolic regulator that promotes glycolytic metabolism in multiple immune cell subsets. Glycolysis is integrally linked to pro-inflammatory immune responses such that activated NK cells and effector T-cell subsets are reliant on sufficient glucose availability for maximal effector function. This article will discuss the regulation of cellular metabolism in NK cells as compared with that of T lymphocytes and discuss the implications for NK cell responses to viral infection and cancer.

  9. Lipid and Carbohydrate Modifications of α-Galactosylceramide Differently Influence Mouse and Human Type I Natural Killer T Cell Activation*

    PubMed Central

    Birkholz, Alysia; Nemčovič, Marek; Yu, Esther Dawen; Girardi, Enrico; Wang, Jing; Khurana, Archana; Pauwels, Nora; Farber, Elisa; Chitale, Sampada; Franck, Richard W.; Tsuji, Moriya; Howell, Amy; Van Calenbergh, Serge; Kronenberg, Mitchell; Zajonc, Dirk M.

    2015-01-01

    The ability of different glycosphingolipids (GSLs) to activate type I natural killer T cells (NKT cells) has been known for 2 decades. The possible therapeutic use of these GSLs has been studied in many ways; however, studies are needed in which the efficacy of promising GSLs is compared under identical conditions. Here, we compare five unique GSLs structurally derived from α-galactosylceramide. We employed biophysical and biological assays, as well as x-ray crystallography to study the impact of the chemical modifications of the antigen on type I NKT cell activation. Although all glycolipids are bound by the T cell receptor of type I NKT cells in real time binding assays with high affinity, only a few activate type I NKT cells in in vivo or in vitro experiments. The differences in biological responses are likely a result of different pharmacokinetic properties of each lipid, which carry modifications at different parts of the molecule. Our results indicate a need to perform a variety of assays to ascertain the therapeutic potential of type I NKT cell GSL activators. PMID:26018083

  10. Lipid and Carbohydrate Modifications of α-Galactosylceramide Differently Influence Mouse and Human Type I Natural Killer T Cell Activation.

    PubMed

    Birkholz, Alysia; Nemčovič, Marek; Yu, Esther Dawen; Girardi, Enrico; Wang, Jing; Khurana, Archana; Pauwels, Nora; Farber, Elisa; Chitale, Sampada; Franck, Richard W; Tsuji, Moriya; Howell, Amy; Van Calenbergh, Serge; Kronenberg, Mitchell; Zajonc, Dirk M

    2015-07-10

    The ability of different glycosphingolipids (GSLs) to activate type I natural killer T cells (NKT cells) has been known for 2 decades. The possible therapeutic use of these GSLs has been studied in many ways; however, studies are needed in which the efficacy of promising GSLs is compared under identical conditions. Here, we compare five unique GSLs structurally derived from α-galactosylceramide. We employed biophysical and biological assays, as well as x-ray crystallography to study the impact of the chemical modifications of the antigen on type I NKT cell activation. Although all glycolipids are bound by the T cell receptor of type I NKT cells in real time binding assays with high affinity, only a few activate type I NKT cells in in vivo or in vitro experiments. The differences in biological responses are likely a result of different pharmacokinetic properties of each lipid, which carry modifications at different parts of the molecule. Our results indicate a need to perform a variety of assays to ascertain the therapeutic potential of type I NKT cell GSL activators. PMID:26018083

  11. Natural killer cells: In health and disease.

    PubMed

    Mandal, Arundhati; Viswanathan, Chandra

    2015-06-01

    Natural killer (NK) cells constitute our bodies' frontline defense system, guarding against tumors and launching attacks against infections. The activities of NK cells are regulated by the interaction of various receptors expressed on their surfaces with cell surface ligands. While the role of NK cells in controlling tumor activity is relatively clear, the fact that they are also linked to various other disease conditions is now being highlighted. Here, we present an overview of the role of NK cells during normal body state as well as under diseased state. We discuss the possible utilization of these powerful cells as immunotherapeutic agents in combating diseases such as asthma, autoimmune diseases, and HIV-AIDS. This review also outlines current challenges in NK cell therapy.

  12. G1-4A, a polysaccharide from Tinospora cordifolia induces peroxynitrite dependent killer dendritic cell (KDC) activity against tumor cells.

    PubMed

    Pandey, Vipul K; Amin, Prayag J; Shankar, Bhavani S

    2014-12-01

    Dendritic cells (DC) play a central role in the development of an adaptive immune response against tumor. In addition to its role in antigen presentation, DC also possesses cytotoxic activity against tumor cells. We have earlier shown phenotypic and functional maturation of bone marrow derived dendritic cells (BMDC) by G1-4A, an arabinogalactan derived from Tinospora cordifolia. In this study, we have investigated the killer phenotype of BMDC matured in the presence of G1-4A, [mBMDC (G1-4A)] on tumor cells. We have observed several fold increase in killing of tumor cells by mBMDC (G1-4A). The tumoricidal activity was not specific to syngeneic tumors cells but could kill xenogenic tumors also. Nitric oxide released by mBMDC (G1-4A) generates peroxynitrite in tumor cells and is responsible for killing of target cells. This killing was completely abrogated by inducible nitric oxide synthase (iNOS) inhibitor 1400W and NADPH oxidase inhibitor apocyanin. The killed target cells are phagocytosed by BMDC which further activate syngeneic cytotoxic T cells. These results thus show that G1-4A treated mBMDC acquire killer phenotype along with maturation which plays an important role in activation of cytotoxic T cells. PMID:25278461

  13. Beta-carotene-induced enhancement of natural killer cell activity in elderly men: an investigation of the role of cytokines.

    PubMed

    Santos, M S; Gaziano, J M; Leka, L S; Beharka, A A; Hennekens, C H; Meydani, S N

    1998-07-01

    We showed previously that natural killer (NK) cell activity is significantly greater in elderly men supplemented with beta-carotene than in those taking placebo. In an attempt to determine the mechanism of beta-carotene's effect, we analyzed the production of NK cell-enhancing cytokines (interferon alpha, interferon gamma, and interleukin 12). Boston-area participants in the Physicians' Health Study (men aged 65-88 y; mean age, 73 y) who had been supplemented with beta-carotene (50 mg on alternate days) for an average of 12 y were enrolled in a randomized, placebo-controlled, double-blind study. Elderly subjects taking beta-carotene supplements had significantly greater plasma beta-carotene concentrations than those taking placebo. Beta-carotene-supplemented elderly men had significantly greater NK cell activity than did elderly men receiving placebo. Percentages of NK cells (CD16+CD56+) were not significantly different between the beta-carotene and placebo groups. Production of interleukin 12, interferon alpha, or concanavalin A-stimulated interferon gamma by cultured peripheral blood mononuclear cells was not significantly different between beta-carotene-supplemented elderly and those taking placebo. Our results indicate that beta-carotene-induced enhancement of NK cell activity is not mediated by changes in percentages of CD16+CD56+ NK cells nor through up-regulation of interleukin 12 or interferon alpha.

  14. Sulfatide-Mediated Activation of Type II Natural Killer T Cells Prevents Hepatic Ischemic Reperfusion Injury In Mice

    PubMed Central

    Arrenberg, Philomena; Maricic, Igor; Kumar, Vipin

    2011-01-01

    Background & Aims Hepatic ischemic reperfusion injury (IRI) is a major complication of liver transplantation and resectional hepatic surgeries. Natural killer T (NKT) cells predominate in liver, where they recognize lipid antigens bound to CD1d molecules. Type I NKT cells utilize a semi-invariant T-cell receptor and react with α-galactosylceramide; type II NKT cells use diverse T-cell receptors. Some type II NKT cells recognize the self-glycolipid sulfatide. It is not clear whether or how these distinct NKT cell subsets mediate hepatocellular damage following IRI. Methods We examined the roles of type I and type II NKT cells in mice with partial hepatic, warm ischemia and reperfusion injury. Results Mice that lack type I NKT cells (Jα18−/−) were protected from hepatic IRI, indicated by reduced hepatocellular necrosis and serum levels of alanine aminotransferase. Sulfatide-mediated activation of type II NKT cells reduced IFN-γ secretion by type I NKT cells and prevented IRI. Protection from hepatic IRI by sulfatide-mediated inactivation of type I NKT cells was associated with significant reductions in hepatic recruitment of myeloid cell subsets, especially the CD11b+Gr-1int, Gr-1−, and NK cells. Conclusion In mice, subsets of NKT cells have opposing roles in hepatic IRI: type I NKT cells promote injury whereas sulfatide-reactive type II NKT cells protect against injury. CD1d activation of NKT cells is conserved from mice to humans, so strategies to modify these processes might be developed to treat patients with hepatic reperfusion injury. PMID:20950612

  15. Natural killer cells in viral arthritis.

    PubMed Central

    Aaskov, J G; Dalglish, D A; Harper, J J; Douglas, J F; Donaldson, M D; Hertzog, P J

    1987-01-01

    Changes in natural killer (NK) cell activity were studied in patients with polyarthritis associated with rubella or Ross River virus infections. In 30 of 32 Ross River virus patients, peripheral NK cell activity was depressed at some stage of the disease but returned to normal levels as patients recovered from arthritic symptoms. Similar changes did not occur in rubella patients and no difference was found between changes in peripheral NK activity and serum interferon (IFN) levels in rubella patients with arthritis and those without. Neither the peak of NK cell activity in peripheral blood lymphocytes (PBL) recovered early in Ross River virus and rubella infections, nor the depression of NK cell activity late in Ross River virus infections could be correlated with changes in serum IFN levels. The decrease in PBL-NK cell activity in epidemic polyarthritis (EPA) patients could not be attributed solely to loss of NK cells from the peripheral circulation because limiting-cell-dilution (LCD) analyses indicated changes in peripheral NK cell activity were due to changes in both the number and lytic activity of NK cells. Despite the association between HLA-DR7 and EPA no differences were found in levels of peripheral NK cell activity in DR7+ and DR7- EPA patients. The demonstration that peripheral NK cells could kill autologous synovial cells suggested that NK cells in joints of EPA patients may contribute to the arthritis associated with Ross River virus infection. PMID:2443284

  16. Natural Killer cell-dependent and non-dependent anti-viral activity of 2-Cys Peroxiredoxin against HIV

    PubMed Central

    Asmal, Mohammed; Letvin, Norman L.; Geiben-Lynn, Ralf

    2013-01-01

    2-cys peroxiredoxins (Prx), a group of anti-oxidative enzyme proteins, act directly on virally-infected cells to inhibit HIV-1 replication, and indirectly through destruction of HIV infected cells by stimulation of Natural Killer (NK) cell-mediated immune responses. We assayed for antibody-dependent NK cell mediated viral inhibition (ADCVI) using plasma from SIV-infected rhesus macaques. We found that Prx-1 strongly increased ADCVI in a dose-dependent manner, suggesting augmentation of NK cell killing. We also investigated the effect of Prx-1 on NK cell-independent HIV-1 and HIV-2 inhibition. We found that primary HIV isolates were potently inhibited at nM concentrations, regardless of viral clade, receptor usage or anti-retroviral drug resistance. During NK cell independent inhibition, we found that Prx-1 reversed the HIV-1 induced gene expression of Heat shock protein 90 kDa alpha (cystolic), class A member 2, (HSP90), a protein of the stress pathway. Prx-1 highly activated Cyclin-dependent kinase inhibitor 2B (CDKN2B), a gene of the TGF-β pathway, and Baculoviral IAP repeat-containing 2 (Birc-2), an anti-apoptotic gene of the NF-κB pathway. We identified gene-expression networks highly dependent on the NFκB and ERK1/2 pathways. Our findings demonstrate that Prx-1 inhibits HIV replication through NK cell-dependent and NK cell-independent mechanisms. PMID:24244928

  17. [Activity of natural killer cells of the spleen of mice exposed to low-intensity of extremely high frequency electromagnetic radiation].

    PubMed

    Ogaĭ, V B; Novoselova, E G; Cherenkov, D A; Fesenko, E E

    2003-01-01

    The dose dependence of natural killer (NK) cell activity from mouse spleen upon action of low-intensity millimeter waves in the exposure range from 5 to 96 hours was studied. It has found an increase of NK activity by 24 hours posttreatment that returned to normal level in a day after the cessation of the irradiation. Also the stimulation of isolated NK cell activity after millimeter waves treatment within 1 hour was revealed.

  18. Revving up Natural Killer Cells and Cytokine-Induced Killer Cells Against Hematological Malignancies

    PubMed Central

    Pittari, Gianfranco; Filippini, Perla; Gentilcore, Giusy; Grivel, Jean-Charles; Rutella, Sergio

    2015-01-01

    Natural killer (NK) cells belong to innate immunity and exhibit cytolytic activity against infectious pathogens and tumor cells. NK-cell function is finely tuned by receptors that transduce inhibitory or activating signals, such as killer immunoglobulin-like receptors, NK Group 2 member D (NKG2D), NKG2A/CD94, NKp46, and others, and recognize both foreign and self-antigens expressed by NK-susceptible targets. Recent insights into NK-cell developmental intermediates have translated into a more accurate definition of culture conditions for the in vitro generation and propagation of human NK cells. In this respect, interleukin (IL)-15 and IL-21 are instrumental in driving NK-cell differentiation and maturation, and hold great promise for the design of optimal NK-cell culture protocols. Cytokine-induced killer (CIK) cells possess phenotypic and functional hallmarks of both T cells and NK cells. Similar to T cells, they express CD3 and are expandable in culture, while not requiring functional priming for in vivo activity, like NK cells. CIK cells may offer some advantages over other cell therapy products, including ease of in vitro propagation and no need for exogenous administration of IL-2 for in vivo priming. NK cells and CIK cells can be expanded using a variety of clinical-grade approaches, before their infusion into patients with cancer. Herein, we discuss GMP-compliant strategies to isolate and expand human NK and CIK cells for immunotherapy purposes, focusing on clinical trials of adoptive transfer to patients with hematological malignancies. PMID:26029215

  19. NKG2D functions as an activating receptor on natural killer cells in the common marmoset (Callithrix jacchus).

    PubMed

    Watanabe, Masamichi; Kudo, Yohei; Kawano, Mitsuko; Nakayama, Masafumi; Nakamura, Kyohei; Kameda, Mai; Ebara, Masamune; Sato, Takeki; Nakamura, Marina; Omine, Kaito; Kametani, Yoshie; Suzuki, Ryuji; Ogasawara, Kouetsu

    2014-11-01

    The natural killer group 2 membrane D (NKG2D) receptor is an NK-activating receptor that plays an important role in host defense against tumors and viral infections. Although the marmoset is an important and reliable animal model, especially for the study of human-specific viral infections, functional characterization of NKG2D on marmoset NK cells has not previously been conducted. In the present study, we investigated a subpopulation of marmoset NK cells that express NKG2D and exhibit cytolytic potential. On the basis of their CD16 and CD56 expression patterns, marmoset NK cells can be classified into three subpopulations: CD16(+) CD56(-), CD16(-) CD56(+) and CD16(-) CD56(-) cells. NKG2D expression on marmoset CD16(+) CD56(-) and CD16(-) CD56(+) splenocytes was confirmed using an NKG2D ligand composed of an MHC class I chain-related molecule A (MICA)-Fc fusion protein. When marmoset splenocytes were cultured with IL-2 for 4 days, NKG2D expression was retained on CD16(+) CD56(-) and CD16(-) CD56(+). In addition, CD16(+) CD56(+) cells within the marmoset NK population appeared which expressed NKG2D after IL-2 stimulation. IL-2-activated marmoset NK cells showed strong cytolytic activity against K562 target cells and target cells stably expressing MICA. Further, the cytolytic activity of marmoset splenocytes was significantly reduced after addition of MICA-Fc fusion protein. Thus, NKG2D functions as an activating receptor on marmoset NK cells that possesses cytotoxic potential, and phenotypic profiles of marmoset NK cell subpopulations are similar to those seen in humans. PMID:24860119

  20. Inositol hexaphosphate-induced enhancement of natural killer cell activity correlates with suppression of colon carcinogenesis in rats

    PubMed Central

    Zhang, Zheng; Song, Yang; Wang, Xiu-Li

    2005-01-01

    AIM: To investigate the anti-neoplastic effect of inositol hexaphosphate (InsP6 or phytic acid) on dimethylhydrazine (DMH)-induced colon tumor in rats and its effect on blood natural killer (NK) cell activity. METHODS: Healthy Wistar rats, 4 wk old, were divided into control group (fed with common food) and InsP6 group (fed with common food+2% sodium inositol hexaphosphate in the drinking water), 15 rats in each group. Both groups were injected with 1,2-dimethylhydrazine subcutaneously (20 mg/kg body weight) once a week for 20 wk. Rats were killed after 21 wk. The whole large intestine was isolated to determine the general condition of tumors and to test blood NK cell activity by lactate-dehydrogenase-release assay. RESULTS: Administration of InsP6 significantly increased blood NK cell activity in DMH-induced colorectal tumor in rats. InsP6 group had a smaller tumor size on average and a smaller number of tumors than the control group. Its mortality was also higher than that in control. However, the variables of body weight and tumor incidence were not significantly different between the two groups. CONCLUSION: InsP6 can increase blood NK cell activity in DMH-induced colon tumor in rats and inhibit tumor growth and metastasis in rats. PMID:16124063

  1. Scorpion venom activates natural killer cells in hepatocellular carcinoma via the NKG2D-MICA pathway.

    PubMed

    Chen, Han; Zhidan, Wang; Xia, Ren; Zhaoxia, Wang; Qing, Jia; Qiang, Guo; Haipeng, Yin; Hengxiao, Wang

    2016-06-01

    Previous studies have demonstrated that polypeptides extracted from scorpion venom (PESV) inhibited cell proliferation in several tumors, however, the effect on dysfunctional and exhausted natural killer cells which contribute to tumor escape from immune surveillance remain to be elucidated. In this study, we determined the effect of PESV on NK infiltration into H22 cells orthotopic transplantation tumors and on the expression of MHC class I chain-related proteins A (MICA) in HepG2 cells. We found that tumor growth in mice was significantly inhibited by PESV and the survival time of tumor-bearing mice treated with PESV was significantly prolonged. Moreover, levels of tumor-infiltrating NK cells, NKG2D protein, perforin and granzyme B mRNA were significantly increased in the group treated with PESV compared with the tumor-bearing control group. In addition, In addition, up-regulation of MICA by PESV enhances the susceptibility of HepG2 cells to NK lysis in vitro. These results indicate that the inhibitory effects of PESV on hepatic carcinoma are likely mediated by up-regulation of NK cell activity via the MICA-NKG2D pathway. PMID:27089390

  2. Alpha-phellandrene promotes immune responses in normal mice through enhancing macrophage phagocytosis and natural killer cell activities.

    PubMed

    Lin, Jen-Jyh; Lin, Ju-Hwa; Hsu, Shu-Chun; Weng, Shu-Wen; Huang, Yi-Ping; Tang, Nou-Ying; Lin, Jaung-Geng; Chung, Jing-Gung

    2013-01-01

    α-Phellandrene, a natural compound from natural plants, has been used in the food and perfume industry. We investigated the effects of α-phellandrene on the immune responses on normal murine cells in vivo. Normal BALB/c mice were treated orally with or without α-phellandrene at 0, 1, 5 and 25 mg/kg and olive oil as a positive control for two weeks. Results indicated that α-phellandrene did not change the weight of animals when compared to olive oil (vehicle for α-phellandrene)-treated groups. After flow cytometric assay of blood samples it was shown that α-phellandrene increased the percentage of CD3 (T-cell marker), CD11b (monocytes) and MAC3 (macrophages), but reduced the percentage of CD19 (B-cell marker) cell surface markers in α-phellandrene-treated groups, compared to untreated groups. α-Phellandrene promoted the phagocytosis of macrophages from blood samples at 5 and 25 mg/kg treatment and promoted natural killer cell activity from splenocytes at 25 mg/kg. Furthermore, α-phellandrene increased B-cell proliferation at 25 mg/kg with or without stimulation but promoted cell proliferation only at 25 mg/kg treatment with stimulation. Based on these observations, 25 mg/kg with α-phellandrene seems to have promoted immune responses in this murine model.

  3. Scorpion venom activates natural killer cells in hepatocellular carcinoma via the NKG2D-MICA pathway.

    PubMed

    Chen, Han; Zhidan, Wang; Xia, Ren; Zhaoxia, Wang; Qing, Jia; Qiang, Guo; Haipeng, Yin; Hengxiao, Wang

    2016-06-01

    Previous studies have demonstrated that polypeptides extracted from scorpion venom (PESV) inhibited cell proliferation in several tumors, however, the effect on dysfunctional and exhausted natural killer cells which contribute to tumor escape from immune surveillance remain to be elucidated. In this study, we determined the effect of PESV on NK infiltration into H22 cells orthotopic transplantation tumors and on the expression of MHC class I chain-related proteins A (MICA) in HepG2 cells. We found that tumor growth in mice was significantly inhibited by PESV and the survival time of tumor-bearing mice treated with PESV was significantly prolonged. Moreover, levels of tumor-infiltrating NK cells, NKG2D protein, perforin and granzyme B mRNA were significantly increased in the group treated with PESV compared with the tumor-bearing control group. In addition, In addition, up-regulation of MICA by PESV enhances the susceptibility of HepG2 cells to NK lysis in vitro. These results indicate that the inhibitory effects of PESV on hepatic carcinoma are likely mediated by up-regulation of NK cell activity via the MICA-NKG2D pathway.

  4. A forward genetic screen reveals novel independent regulators of ULBP1, an activating ligand for natural killer cells

    PubMed Central

    Gowen, Benjamin G; Chim, Bryan; Marceau, Caleb D; Greene, Trever T; Burr, Patrick; Gonzalez, Jeanmarie R; Hesser, Charles R; Dietzen, Peter A; Russell, Teal; Iannello, Alexandre; Coscoy, Laurent; Sentman, Charles L; Carette, Jan E; Muljo, Stefan A; Raulet, David H

    2015-01-01

    Recognition and elimination of tumor cells by the immune system is crucial for limiting tumor growth. Natural killer (NK) cells become activated when the receptor NKG2D is engaged by ligands that are frequently upregulated in primary tumors and on cancer cell lines. However, the molecular mechanisms driving NKG2D ligand expression on tumor cells are not well defined. Using a forward genetic screen in a tumor-derived human cell line, we identified several novel factors supporting expression of the NKG2D ligand ULBP1. Our results show stepwise contributions of independent pathways working at multiple stages of ULBP1 biogenesis. Deeper investigation of selected hits from the screen showed that the transcription factor ATF4 drives ULBP1 gene expression in cancer cell lines, while the RNA-binding protein RBM4 supports ULBP1 expression by suppressing a novel alternatively spliced isoform of ULBP1 mRNA. These findings offer insight into the stress pathways that alert the immune system to danger. DOI: http://dx.doi.org/10.7554/eLife.08474.001 PMID:26565589

  5. A forward genetic screen reveals novel independent regulators of ULBP1, an activating ligand for natural killer cells.

    PubMed

    Gowen, Benjamin G; Chim, Bryan; Marceau, Caleb D; Greene, Trever T; Burr, Patrick; Gonzalez, Jeanmarie R; Hesser, Charles R; Dietzen, Peter A; Russell, Teal; Iannello, Alexandre; Coscoy, Laurent; Sentman, Charles L; Carette, Jan E; Muljo, Stefan A; Raulet, David H

    2015-01-01

    Recognition and elimination of tumor cells by the immune system is crucial for limiting tumor growth. Natural killer (NK) cells become activated when the receptor NKG2D is engaged by ligands that are frequently upregulated in primary tumors and on cancer cell lines. However, the molecular mechanisms driving NKG2D ligand expression on tumor cells are not well defined. Using a forward genetic screen in a tumor-derived human cell line, we identified several novel factors supporting expression of the NKG2D ligand ULBP1. Our results show stepwise contributions of independent pathways working at multiple stages of ULBP1 biogenesis. Deeper investigation of selected hits from the screen showed that the transcription factor ATF4 drives ULBP1 gene expression in cancer cell lines, while the RNA-binding protein RBM4 supports ULBP1 expression by suppressing a novel alternatively spliced isoform of ULBP1 mRNA. These findings offer insight into the stress pathways that alert the immune system to danger.

  6. The antileukemic potential of natural killer cells.

    PubMed

    Torelli, Giovanni F; Peragine, Nadia; Mariglia, Paola; Foà, Robin

    2016-01-01

    The antileukemic potential of natural killer (NK) cells has over the years raised considerable interest and new immune-based treatment protocols characterized by the infusion of freshly isolated or ex vivo activated and expanded effectors have been designed. Several aspects still need to be addressed, including the optimal timing of NK infusion during the course of the disease, the best preparative regimen, the origin of NK cells and the possible need of ex vivo NK cell manipulation before the infusion. The aims of this review are to discuss the experimental and clinical data available on the role played by NK cells for leukemia patients and to revise the different good manufacturing practice protocols for ex vivo manipulation of these effector cells.

  7. Effects of exercise and training on natural killer cell counts and cytolytic activity: a meta-analysis.

    PubMed

    Shephard, R J; Shek, P N

    1999-09-01

    Meta-analysis techniques have been used to accumulate data from 94 studies describing the natural killer (NK) cell response of some 900 volunteers to acute and chronic exercise. NK cell numbers have been indicated in terms of CD3-CD16+CD56+, CD16+ or CD56+ phenotypes, and cytolytic activity has been expressed per 10,000 peripheral blood mononuclear cells or in terms of lytic units. Acute exercise has been categorised as sustained moderate (50 to 65% of aerobic power), sustained vigorous (>75% of aerobic power), brief maximal or 'supramaximal', prolonged, eccentric or resistance, and repeated exercise. In general, there was a marked increase in NK cell count at the end of exercise, probably attributable to a catecholamine-mediated demargination of cells. Following exercise, cell counts dropped to less than half of normal levels for a couple of hours but, except in unusual circumstances (e.g. prolonged, intense and stressful exercise), normal resting values are restored within 24 hours. If activity is both prolonged and vigorous, the decrease in NK cell counts and cytolytic activity may begin during the exercise session. Although the usual depression of NK cell count seems too brief to have major practical importance for health, there could be a cumulative adverse effect on immunosurveillance and health experience in athletes who induce such changes several times per week. There is a weak suggestion of an offsetting increase in resting NK cell counts and cytolytic action in trained individuals, and this merits further exploration in studies where effects of recent training sessions are carefully controlled. PMID:10541441

  8. Natural Killer Cell-Based Therapies Targeting Cancer: Possible Strategies to Gain and Sustain Anti-Tumor Activity

    PubMed Central

    Dahlberg, Carin I. M.; Sarhan, Dhifaf; Chrobok, Michael; Duru, Adil D.; Alici, Evren

    2015-01-01

    Natural killer (NK) cells were discovered 40 years ago, by their ability to recognize and kill tumor cells without the requirement of prior antigen exposure. Since then, NK cells have been seen as promising agents for cell-based cancer therapies. However, NK cells represent only a minor fraction of the human lymphocyte population. Their skewed phenotype and impaired functionality during cancer progression necessitates the development of clinical protocols to activate and expand to high numbers ex vivo to be able to infuse sufficient numbers of functional NK cells to the cancer patients. Initial NK cell-based clinical trials suggested that NK cell-infusion is safe and feasible with almost no NK cell-related toxicity, including graft-versus-host disease. Complete remission and increased disease-free survival is shown in a small number of patients with hematological malignances. Furthermore, successful adoptive NK cell-based therapies from haploidentical donors have been demonstrated. Disappointingly, only limited anti-tumor effects have been demonstrated following NK cell infusion in patients with solid tumors. While NK cells have great potential in targeting tumor cells, the efficiency of NK cell functions in the tumor microenvironment is yet unclear. The failure of immune surveillance may in part be due to sustained immunological pressure on tumor cells resulting in the development of tumor escape variants that are invisible to the immune system. Alternatively, this could be due to the complex network of immune-suppressive compartments in the tumor microenvironment, including myeloid-derived suppressor cells, tumor-associated macrophages, and regulatory T cells. Although the negative effect of the tumor microenvironment on NK cells can be transiently reverted by ex vivo expansion and long-term activation, the aforementioned NK cell/tumor microenvironment interactions upon reinfusion are not fully elucidated. Within this context, genetic modification of NK cells

  9. Natural Killer Cell-Based Therapies Targeting Cancer: Possible Strategies to Gain and Sustain Anti-Tumor Activity.

    PubMed

    Dahlberg, Carin I M; Sarhan, Dhifaf; Chrobok, Michael; Duru, Adil D; Alici, Evren

    2015-01-01

    Natural killer (NK) cells were discovered 40 years ago, by their ability to recognize and kill tumor cells without the requirement of prior antigen exposure. Since then, NK cells have been seen as promising agents for cell-based cancer therapies. However, NK cells represent only a minor fraction of the human lymphocyte population. Their skewed phenotype and impaired functionality during cancer progression necessitates the development of clinical protocols to activate and expand to high numbers ex vivo to be able to infuse sufficient numbers of functional NK cells to the cancer patients. Initial NK cell-based clinical trials suggested that NK cell-infusion is safe and feasible with almost no NK cell-related toxicity, including graft-versus-host disease. Complete remission and increased disease-free survival is shown in a small number of patients with hematological malignances. Furthermore, successful adoptive NK cell-based therapies from haploidentical donors have been demonstrated. Disappointingly, only limited anti-tumor effects have been demonstrated following NK cell infusion in patients with solid tumors. While NK cells have great potential in targeting tumor cells, the efficiency of NK cell functions in the tumor microenvironment is yet unclear. The failure of immune surveillance may in part be due to sustained immunological pressure on tumor cells resulting in the development of tumor escape variants that are invisible to the immune system. Alternatively, this could be due to the complex network of immune-suppressive compartments in the tumor microenvironment, including myeloid-derived suppressor cells, tumor-associated macrophages, and regulatory T cells. Although the negative effect of the tumor microenvironment on NK cells can be transiently reverted by ex vivo expansion and long-term activation, the aforementioned NK cell/tumor microenvironment interactions upon reinfusion are not fully elucidated. Within this context, genetic modification of NK cells

  10. Activating Receptor Signals Drive Receptor Diversity in Developing Natural Killer Cells

    PubMed Central

    Freund, Jacquelyn; May, Rebecca M.; Li, Hongchuan; McCullen, Matthew; Zhang, Bin; Lenvik, Todd; Cichocki, Frank; Anderson, Stephen K.; Kambayashi, Taku

    2016-01-01

    It has recently been appreciated that NK cells exhibit many features reminiscent of adaptive immune cells. Considerable heterogeneity exists with respect to the ligand specificity of individual NK cells and as such, a subset of NK cells can respond, expand, and differentiate into memory-like cells in a ligand-specific manner. MHC I-binding inhibitory receptors, including those belonging to the Ly49 and KIR families, are expressed in a variegated manner, which creates ligand-specific diversity within the NK cell pool. However, how NK cells determine which inhibitory receptors to express on their cell surface during a narrow window of development is largely unknown. In this manuscript, we demonstrate that signals from activating receptors are critical for induction of Ly49 and KIR receptors during NK cell development; activating receptor-derived signals increased the probability of the Ly49 bidirectional Pro1 promoter to transcribe in the forward versus the reverse direction, leading to stable expression of Ly49 receptors in mature NK cells. Our data support a model where the balance of activating and inhibitory receptor signaling in NK cells selects for the induction of appropriate inhibitory receptors during development, which NK cells use to create a diverse pool of ligand-specific NK cells. PMID:27500644

  11. Activating Receptor Signals Drive Receptor Diversity in Developing Natural Killer Cells.

    PubMed

    Freund, Jacquelyn; May, Rebecca M; Yang, Enjun; Li, Hongchuan; McCullen, Matthew; Zhang, Bin; Lenvik, Todd; Cichocki, Frank; Anderson, Stephen K; Kambayashi, Taku

    2016-08-01

    It has recently been appreciated that NK cells exhibit many features reminiscent of adaptive immune cells. Considerable heterogeneity exists with respect to the ligand specificity of individual NK cells and as such, a subset of NK cells can respond, expand, and differentiate into memory-like cells in a ligand-specific manner. MHC I-binding inhibitory receptors, including those belonging to the Ly49 and KIR families, are expressed in a variegated manner, which creates ligand-specific diversity within the NK cell pool. However, how NK cells determine which inhibitory receptors to express on their cell surface during a narrow window of development is largely unknown. In this manuscript, we demonstrate that signals from activating receptors are critical for induction of Ly49 and KIR receptors during NK cell development; activating receptor-derived signals increased the probability of the Ly49 bidirectional Pro1 promoter to transcribe in the forward versus the reverse direction, leading to stable expression of Ly49 receptors in mature NK cells. Our data support a model where the balance of activating and inhibitory receptor signaling in NK cells selects for the induction of appropriate inhibitory receptors during development, which NK cells use to create a diverse pool of ligand-specific NK cells.

  12. Impaired culture generated cytotoxicity with preservation of spontaneous natural killer-cell activity in cartilage-hair hypoplasia

    SciTech Connect

    Pierce, G.F.; Brovall, C.; Schacter, B.Z.; Polmar, S.H.

    1983-06-01

    Recent studies of cartilage-hair hypoplasia (CHH), a form of short-limbed dwarfism, have shown that all affected individuals have a cellular proliferation defect that results in a cellular immunodeficiency. However, only a minority of CHH individuals suffer from severe, life-threatening infections. For this reason, relevant immune defense mechanisms that may be responsible for maintaining intact host defenses in the majority of CHH individuals were studied. Spontaneous and allogeneic culture-induced (mixed lymphocyte response-MLR) specific and nonspecific (NK-like) cytotoxic mechanisms were analyzed and correlated with lymphocyte subpopulations present in CHH and normal individuals. Spontaneous natural-killer (NK) activity was present at or above normal levels, but culture-induced specific cytotoxicity and NK-like cytotoxicity as well as NK-like activity by T cell lines were significantly reduced in CHH individuals. The generation of radiation-resistant cytotoxicity, which normally occurs during allogeneic MLR, was markedly diminished in CHH, and was correlated with the decreased proliferation observed in CHH cultures. Preservation of spontaneous NK activity and loss of all forms of culture-induced cytotoxicity was associated with an increase in the proportion of lymphocytes bearing a thymic independent NK phenotype, and a significant decrease in thymic derived cytolytic T cell sub-populations in CHH individuals. Therefore, an intact cellular cytotoxic effector mechanism has been identified in CHH (i.e., NK activity).

  13. Effects of branched-chain amino acid-enriched nutrient mixture on natural killer cell activity in viral cirrhosis.

    PubMed

    Takegoshi, K; Nanasawa, H; Itoh, H; Yasuyama, T; Ohmoto, Y; Sugiyama, K

    1998-06-01

    A controlled study was performed in 18 viral cirrhosis patients to evaluate whether immune function, as indicated by natural killer (NK) cell activity, was improved by a branched-chain amino acid-enriched nutrient mixture (nutrient-mixture), Aminoleban EN. Five patients received the nutrient-mixture (100 g/day) for 2 to 6 weeks preceded by control periods. Five additional patients received the nutrient-mixture for 2 to 4 weeks, and the remaining 8 patients did not receive the nutrient-mixture. NK cell activity, CD16, CD8, CD11b, and amino acids were assayed before and after the administration of the drug in the nutrient-mixture-supplemented group, and two times with 1 to 6 month intervals in the control group. In the nutrient-mixture-supplemented group (n = 10), increasing NK cell activity, expressed as the ratio of values of post-treatment to that of baseline (ratio > 1.25) was detected in 7 (70%) patients, whereas in the control group (n = 13), it was detected in only 1 (7.7%) (p < 0.01). While in the affected group (NK cell activity ratio > 1.25, n = 7), all patients had compensated liver cirrhosis, in the unaffected group (NK cell activity ratio < 1.25, n = 3), 2 of 3 patients had decompensated liver cirrhosis (p < 0.02). Laboratory data, indicating severity of liver cirrhosis, such as total bilirubin and albumin, showed better values (p < 0.01, p < 0.05 respectively), and baseline NK cell activity was low (8.7 +/- 7.2% vs 33.3 +/- 13.0%, p < 0.05) in the affected group than unaffected group. NK cell subpopulations such as CD16 (%), CD11b (%) and one of the populations of T cell such as CD8 (%) showed no significant change throughout the study. As for amino acids analysis, Fischer's ratio was increased in the nutrient-mixture-supplemented group compared to the control group (p < 0.05), but none of the amino acids showed significant change. Thus the changes in NK cell activity were not explained by increase in NK cell subpopulations nor changes of amino acids

  14. Antimicrobial activity of various immunomodulators: independence from normal levels of circulating monocytes and natural killer cells. Technical report

    SciTech Connect

    Morahan, P.S.; Dempsey, W.L.; Volkman, A.; Connor, J.

    1986-01-01

    The effects of /sup 89/Sr treatment on the natural host resistance of CD-1 mice and the enhancement of resistance by immunomodulators to infection with Listeria monocytogenes or herpes simplex virus type 2 (HSV-2) were determined. In the CD-1 mouse, single-dose treatment with /sup 89/Sr caused a profound decrease in the number of circulating monocytes (Mo), lymphocytes, and polymorphonuclear leukocytes (PMN) within 1 week. There was also marked functional impairment of the Mo inflammatory response, as well as markedly decreased spontaneous and activatable cytoxicity by splenic natural killer (NK) cells. Despite this profound cellular suppression, there was no significant change in natural resistance of CD-1 mice to L. monocytogenes of HSV-2 infection. Furthermore, prophylactic treatment of mice with the biologic immunomodulator Corynebacterium parvum or the synthetic immunomodulators maleic anhydride-divinyl ether or avridine in liposomes resulted in comparable enhancement of resistance in /sup 89/Sr-treated and normal mice. These data indicate that natural and immunomodulator-enhanced resistance of CD-1 mice to microbail infections do not depend on normal levels of Mo, PMN, or NK cells. The resistance enhancement may rely on activated tissue macrophages. In contrast to the early changes in circulating leukocytes, the residenet peritoneal cell populations were not markedly altered until after day 30. There then was a distinct decline in lymphocytes and a gradual decline in activated tissue macrophages.

  15. Polyinosinic-cytidylic acid as an adjuvant on natural killer- and dendritic cell-mediated antitumor activities.

    PubMed

    Huang, Yu-Kun; Zheng, Zhi; Qiu, Fu

    2013-06-01

    Previously, we demonstrated that treatment with E7(44-62) and the adjuvant polyinosinic-cytidylic acid (poly(I:C)) in a rodent model generates antitumor immune responses, but the effect of E7(44-62) with poly(I:C) on natural killer (NK)- and dendritic cell (DC)-mediated antitumor activities is still unclear. Our goal was to examine the antitumor effects of E7(44-62) with poly(I:C). We examined the ability of E7(44-62) with poly(I:C) to induce toll-like receptor 3 (TLR3) expression, tumor necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ) mRNA expression, and tumor cell-killing activity in human NK cells as well as its ability to induce CD11c and CD86 expression and proliferation in human DCs. We found that E7(44-62) with poly(I:C) treatment markedly increased TLR3 expression and cytotoxicity against HeLa cells in human NK92 cells. Moreover, treatment with E7(44-62) and poly(I:C) markedly up-regulated IFN-γ and TNF-α mRNA expression in NK92 cells. Human patients with cervical cancer exhibited a marked decrease in the frequency of DCs; however, ex vivo treatment with E7(44-62) and poly(I:C) restored DC frequency. Stimulation of human DCs in patients with E7(44-62) and poly(I:C) led to high levels of CD11c and CD86 expression. Our data reveal the involvement of E7(44-62) combined with poly(I:C) in potentiating antitumor cytotoxicity and cytokine-producing activities in human NK92 cells and DCs.

  16. An Improved Flow Cytometry Method For Precise Quantitation Of Natural-Killer Cell Activity

    NASA Technical Reports Server (NTRS)

    Crucian, Brian; Nehlsen-Cannarella, Sandra; Sams, Clarence

    2006-01-01

    The ability to assess NK cell cytotoxicity using flow cytometry has been previously described and can serve as a powerful tool to evaluate effector immune function in the clinical setting. Previous methods used membrane permeable dyes to identify target cells. The use of these dyes requires great care to achieve optimal staining and results in a broad spectral emission that can make multicolor cytometry difficult. Previous methods have also used negative staining (the elimination of target cells) to identify effector cells. This makes a precise quantitation of effector NK cells impossible due to the interfering presence of T and B lymphocytes, and the data highly subjective to the variable levels of NK cells normally found in human peripheral blood. In this study an improved version of the standard flow cytometry assay for NK activity is described that has several advantages of previous methods. Fluorescent antibody staining (CD45FITC) is used to positively identify target cells in place of membranepermeable dyes. Fluorescent antibody staining of target cells is less labor intensive and more easily reproducible than membrane dyes. NK cells (true effector lymphocytes) are also positively identified by fluorescent antibody staining (CD56PE) allowing a simultaneous absolute count assessment of both NK cells and target cells. Dead cells are identified by membrane disruption using the DNA intercalating dye PI. Using this method, an exact NK:target ratio may be determined for each assessment, including quantitation of NK target complexes. Backimmunoscatter gating may be used to track live vs. dead Target cells via scatter properties. If desired, NK activity may then be normalized to standardized ratios for clinical comparisons between patients, making the determination of PBMC counts or NK cell percentages prior to testing unnecessary. This method provides an exact cytometric determination of NK activity that highly reproducible and may be suitable for routine use in the

  17. Human cancer cells with stem cell-like phenotype exhibit enhanced sensitivity to the cytotoxicity of IL-2 and IL-15 activated natural killer cells.

    PubMed

    Yin, Tao; Wang, Guoping; He, Sisi; Liu, Qin; Sun, Jianhong; Wang, Yongsheng

    2016-02-01

    Tumors harbor a population of cancer stem cells (CSCs) which can drive tumor progression and therapeutical resistance. Nature killer (NK) cells are best known for their ability to directly recognize and kill malignant cells. However, the susceptibility of cancer stem cells to NK cells is not fully understood. Here we demonstrated that human CD44+CD24- breast CSCs were shown enhanced sensitivity to IL-2 and IL-15 activated NK cells. CD44+CD24- CSCs expressed higher levels of NKG2D ligands ULBP1, ULBP2 and MICA. Blockade assay showed that the sensitivity of CSCs to NK cells-mediated lysis was mainly dependent on NKG2D. Furthermore, redox oxygen species (ROS)-low tumor cells were more sensitive to NK cells. The presence of antioxidant enzymes inhibitor L-S,R-buthionine sulfoximine or H2O2 retarded the cytotoxicity of NK cells to CD44+CD24- CSCs. In addition, NK cells could readily target CD133+ colonal CSCs. Our findings provide novel targets for NK cells-based immunotherapy and are of great importance for translational medicine.

  18. NKR-P1A is a target-specific receptor that activates natural killer cell cytotoxicity.

    PubMed

    Ryan, J C; Niemi, E C; Nakamura, M C; Seaman, W E

    1995-05-01

    NKR-P1A is a lectinlike surface molecule expressed on rat natural killer (NK) cells. NKR-P1A has structural and functional features of an activating NK cell receptor, but a requirement for NKR-P1A in target cell lysis has not been determined. To define the role of NKR-P1A in natural killing, we have generated a mutant of the rat NK cell line, RNK-16, lacking expression of all members of the NKR-P1 receptor family. Although these NKR-P1-deficient NK cells were able to kill many standard tumor targets, including YAC-1, they were selectively deficient in the lysis of IC-21 macrophage, B-16 melanoma, and C1498 lymphoma targets. Reexpression of a single member of the NKR-P1 family, NKR-P1A, on mutant cells restored lysis of IC-21, and killing of IC-21 targets through rat NKR-P1A was completely blocked by F(ab')2 anti-NKR-P1A. Reexpression of NKR-P1A also restored transmembrane signaling to IC-21, as assessed by the generation of inositol-1,4,5-trisphosphate. The generation of inositol-1,4,5-trisphosphate was also restored in response to B-16 targets, but both B-16 and C1498 cells remained resistant to lysis, indicating that other NK cell molecules, perhaps within the NKR-P1 family, are required for the efficient killing of these tumors. These results are the first to demonstrate that NKR-P1A is a target-specific receptor that activates natural killing. PMID:7722466

  19. NKR-P1A is a target-specific receptor that activates natural killer cell cytotoxicity

    PubMed Central

    1995-01-01

    NKR-P1A is a lectinlike surface molecule expressed on rat natural killer (NK) cells. NKR-P1A has structural and functional features of an activating NK cell receptor, but a requirement for NKR-P1A in target cell lysis has not been determined. To define the role of NKR-P1A in natural killing, we have generated a mutant of the rat NK cell line, RNK-16, lacking expression of all members of the NKR-P1 receptor family. Although these NKR-P1-deficient NK cells were able to kill many standard tumor targets, including YAC-1, they were selectively deficient in the lysis of IC-21 macrophage, B-16 melanoma, and C1498 lymphoma targets. Reexpression of a single member of the NKR-P1 family, NKR-P1A, on mutant cells restored lysis of IC-21, and killing of IC-21 targets through rat NKR-P1A was completely blocked by F(ab')2 anti-NKR- P1A. Reexpression of NKR-P1A also restored transmembrane signaling to IC-21, as assessed by the generation of inositol-1,4,5-trisphosphate. The generation of inositol-1,4,5-trisphosphate was also restored in response to B-16 targets, but both B-16 and C1498 cells remained resistant to lysis, indicating that other NK cell molecules, perhaps within the NKR-P1 family, are required for the efficient killing of these tumors. These results are the first to demonstrate that NKR-P1A is a target-specific receptor that activates natural killing. PMID:7722466

  20. Activation of natural killer T cells in NZB/W mice induces Th1-type immune responses exacerbating lupus.

    PubMed

    Zeng, Defu; Liu, Yinping; Sidobre, Stephane; Kronenberg, Mitchell; Strober, Samuel

    2003-10-01

    In vivo treatment of mice with the natural killer T (NKT) cell ligand, alpha-galactosylceramide (alphaGalCer), ameliorates autoimmune diabetes and experimental autoimmune encephalomyelitis (EAE) by shifting pathogenic Th1-type immune responses to nonpathogenic Th2-type responses. In the current study, in vivo activation of NKT cells in adult NZB/W mice by multiple injections of alphaGalCer induced an abnormal Th1-type immune response as compared with the Th2-type response observed in nonautoimmune C57BL/6 mice. This resulted in decreased serum levels of IgE, increased levels of IgG2a and IgG2a anti-double-stranded DNA (anti-dsDNA) Ab's, and exacerbated lupus. Conversely, treatment of NZB/W mice with blocking anti-CD1d mAb augmented Th2-type responses, increased serum levels of IgE, decreased levels of IgG2a and IgG2a anti-dsDNA Ab's, and ameliorated lupus. While total CD4+ T cells markedly augmented in vitro IgM anti-dsDNA Ab secretion by splenic B cells, the non-CD1d-reactive (CD1d-alphaGalCer tetramer-negative) CD4+ T cells (accounting for 95% of all CD4+ T cells) failed to augment Ab secretion. The CD1d-reactive tetramer-positive CD4+ T cells augmented anti-dsDNA Ab secretion about tenfold. In conclusion, activation of NKT cells augments Th1-type immune responses and autoantibody secretion that contribute to lupus development in adult NZB/W mice, and anti-CD1d mAb might be useful for treating lupus.

  1. Activation of Human T-Helper/Inducer Cell, T-Cytotoxic Cell, B-Cell, and Natural Killer (NK)-Cells and induction of Natural Killer Cell Activity against K562 Chronic Myeloid Leukemia Cells with Modified Citrus Pectin

    PubMed Central

    2011-01-01

    Background Modified citrus pectin (MCP) is known for its anti-cancer effects and its ability to be absorbed and circulated in the human body. In this report we tested the ability of MCP to induce the activation of human blood lymphocyte subsets like T, B and NK-cells. Methods MCP treated human blood samples were incubated with specific antibody combinations and analyzed in a flow cytometer using a 3-color protocol. To test functionality of the activated NK-cells, isolated normal lymphocytes were treated with increasing concentrations of MCP. Log-phase PKH26-labeled K562 leukemic cells were added to the lymphocytes and incubated for 4 h. The mixture was stained with FITC-labeled active form of caspase 3 antibody and analyzed by a 2-color flow cytometry protocol. The percentage of K562 cells positive for PKH26 and FITC were calculated as the dead cells induced by NK-cells. Monosaccharide analysis of the MCP was performed by high-performance anion-exchange chromatography with pulse amperometric detection (HPAEC-PAD). Results MCP activated T-cytotoxic cells and B-cell in a dose-dependent manner, and induced significant dose-dependent activation of NK-cells. MCP-activated NK-cells demonstrated functionality in inducing cancer cell death. MCP consisted of oligogalacturonic acids with some containing 4,5-unsaturated non-reducing ends. Conclusions MCP has immunostimulatory properties in human blood samples, including the activation of functional NK cells against K562 leukemic cells in culture. Unsaturated oligogalacturonic acids appear to be the immunostimulatory carbohydrates in MCP. PMID:21816083

  2. Transpresentation of interleukin-15 by IL-15/IL-15Rα mRNA-engineered human dendritic cells boosts antitumoral natural killer cell activity

    PubMed Central

    Van den Bergh, Johan; Willemen, Yannick; Lion, Eva; Van Acker, Heleen; De Reu, Hans; Anguille, Sébastien; Goossens, Herman; Berneman, Zwi

    2015-01-01

    In cancer immunotherapy, the use of dendritic cell (DC)-based vaccination strategies can improve overall survival, but until now durable clinical responses remain scarce. To date, DC vaccines are designed primarily to induce effective T-cell responses, ignoring the antitumor activity potential of natural killer (NK) cells. Aiming to further improve current DC vaccination outcome, we engineered monocyte-derived DC to produce interleukin (IL)-15 and/or IL-15 receptor alpha (IL-15Rα) using mRNA electroporation. The addition of IL-15Rα to the protocol, enabling IL-15 transpresentation to neighboring NK cells, resulted in significantly better NK-cell activation compared to IL-15 alone. Next to upregulation of NK-cell membrane activation markers, IL-15 transpresentation resulted in increased NK-cell secretion of IFN-γ, granzyme B and perforin. Moreover, IL-15-transpresenting DC/NK cell cocultures from both healthy donors and acute myeloid leukemia (AML) patients in remission showed markedly enhanced cytotoxic activity against NK cell sensitive and resistant tumor cells. Blocking IL-15 transpresentation abrogated NK cell-mediated cytotoxicity against tumor cells, pointing to a pivotal role of IL-15 transpresentation by IL-15Rα to exert its NK cell-activating effects. In conclusion, we report an attractive approach to improve antitumoral NK-cell activity in DC-based vaccine strategies through the use of IL-15/IL-15Rα mRNA-engineered designer DC. PMID:26675759

  3. Natural killer (NK) activity of pit cells perfused from livers of rats treated with ethanol

    SciTech Connect

    Albornoz, L.; Jones, J.M.; Crutchfield, C.; Veech, R.L. Univ. of Arkansas Medical Sciences, Little Rock )

    1991-03-11

    The liver is the major site of ethanol (ETOH) metabolism. Liver sinusoids contain lymphocytes with NK activity. The authors treated LEW rats for 2 weeks with i.p. injection of 1.25 ml 25% ETOH/kg 3 times/week and 5% ETOH in drinking water. Livers were perfused at 5-fold physiological pressure and cells obtained were banded on 1.077 density Ficoll. Their cytotoxicity was tested against {sup 51}Cr-labeled YAC-1 or U937 and compared to spleen and blood lymphocytes. In untreated rats, pit cell NK activity was 2-fold that of splenic lymphocytes and 4-fold that of blood lymphocytes. Compared to controls, ETOH-treated rats exhibited a 30 to 90% rise in pit cell NK activity detected with YAC-1 or U937 targets. The pit cell enhanced NK activity in ETOH-treated rats was further increased if polyinosinicpolycytidilic acid was injection i.p. 18 hours before the assay. Blood and spleen lymphocyte NK activity of ETOH-treated rats was also greater than in controls. There was no evidence that ETOH merely redistributed lymphocytes among the tissues. Although ETOH acutely inhibits NK activity in vitro, chronic ETOH increases in vivo.

  4. Cytolysis of oligodendrocytes is mediated by killer (K) cells but not by natural killer (NK) cells.

    PubMed

    Satoh, J; Kim, S U; Kastrukoff, L F

    1991-03-01

    The cytotoxic activity of killer (K) cells against enriched cultures of bovine oligodendrocytes (BOL) was investigated in multiple sclerosis (MS) and controls. Human K cells mediated cytotoxicity to primary cultures of BOL in the presence of anti-BOL antiserum in all study groups, while BOL were resistant to human natural killer (NK) cells. Cytotoxic activity was significantly reduced in MS when compared to age-matched normal controls but not when compared to other neurologic disease (OND) patients. K cell-mediated lysis of BOL could also be induced with anti-galactocerebroside antibody but not with other antibodies including those specific for OL antigens (myelin basic protein, proteolipid apoprotein, and 2',3'-cyclic nucleotide 3'-phosphodiesterase). Enrichment of the effector population indicated that antibody-dependent cell-mediated cytotoxicity (ADCC) to BOL was mediated by large granular lymphocytes, and the effector population was further characterized by flow cytometry. The effector cells mediating ADCC could be inhibited by protein A of Staphylococcus aureus, and by K562 cells in cold competition assay. These observations indicate that oligodendrocytes are resistant to NK cells but are susceptible to cytolysis mediated by K cells. This may represent a potentially important immune mechanism in the pathogenesis of MS.

  5. Natural Killer Cells for Therapy of Leukemia

    PubMed Central

    Suck, Garnet; Linn, Yeh Ching; Tonn, Torsten

    2016-01-01

    Summary Clinical application of natural killer (NK) cells against leukemia is an area of intense investigation. In human leukocyte antigen-mismatched allogeneic hematopoietic stem cell transplantations (HSCT), alloreactive NK cells exert powerful anti-leukemic activity in preventing relapse in the absence of graft-versus-host disease, particularly in acute myeloid leukemia patients. Adoptive transfer of donor NK cells post-HSCT or in non-transplant scenarios may be superior to the currently widely used unmanipulated donor lymphocyte infusion. This concept could be further improved through transfusion of activated NK cells. Significant progress has been made in good manufacturing practice (GMP)-compliant large-scale production of stimulated effectors. However, inherent limitations remain. These include differing yields and compositions of the end-product due to donor variability and inefficient means for cryopreservation. Moreover, the impact of the various novel activation strategies on NK cell biology and in vivo behavior are barely understood. In contrast, reproduction of the third-party NK-92 drug from a cryostored GMP-compliant master cell bank is straightforward and efficient. Safety for the application of this highly cytotoxic cell line was demonstrated in first clinical trials. This novel ‘off-the-shelf’ product could become a treatment option for a broad patient population. For specific tumor targeting chimeric-antigen-receptor-engineered NK-92 cells have been designed. PMID:27226791

  6. Natural killer cell dysfunction is a distinguishing feature of systemic onset juvenile rheumatoid arthritis and macrophage activation syndrome.

    PubMed

    Villanueva, Joyce; Lee, Susan; Giannini, Edward H; Graham, Thomas B; Passo, Murray H; Filipovich, Alexandra; Grom, Alexei A

    2005-01-01

    Macrophage activation syndrome (MAS) has been reported in association with many rheumatic diseases, most commonly in systemic juvenile rheumatoid arthritis (sJRA). Clinically, MAS is similar to hemophagocytic lymphohistiocytosis (HLH), a genetic disorder with absent or depressed natural killer (NK) function. We have previously reported that, as in HLH, patients with MAS have profoundly decreased NK activity, suggesting that this abnormality might be relevant to the pathogenesis of the syndrome. Here we examined the extent of NK dysfunction across the spectrum of diseases that comprise juvenile rheumatoid arthritis (JRA). Peripheral blood mononuclear cells (PBMC) were collected from patients with pauciarticular (n = 4), polyarticular (n = 16), and systemic (n = 20) forms of JRA. NK cytolytic activity was measured after co-incubation of PBMC with the NK-sensitive K562 cell line. NK cells (CD56+/T cell receptor [TCR]-alphabeta-), NK T cells (CD56+/TCR-alphabeta+), and CD8+ T cells were also assessed for perforin and granzyme B expression by flow cytometry. Overall, NK cytolytic activity was significantly lower in patients with sJRA than in other JRA patients and controls. In a subgroup of patients with predominantly sJRA, NK cell activity was profoundly decreased: in 10 of 20 patients with sJRA and in only 1 of 20 patients with other JRA, levels of NK activity were below two standard deviations of pediatric controls (P = 0.002). Some decrease in perforin expression in NK cells and cytotoxic T lymphocytes was seen in patients within each of the JRA groups with no statistically significant differences. There was a profound decrease in the proportion of circulating CD56bright NK cells in three sJRA patients, a pattern similar to that previously observed in MAS and HLH. In conclusion, a subgroup of patients with JRA who have not yet had an episode of MAS showed decreased NK function and an absence of circulating CD56bright population, similar to the abnormalities observed

  7. Opioid peptides mediate the suppressive effect of stress on natural killer cell cytotoxicity.

    PubMed

    Shavit, Y; Lewis, J W; Terman, G W; Gale, R P; Liebeskind, J C

    1984-01-13

    The cytotoxic activity of natural killer cells was investigated in rats subjected to one of two inescapable footshock stress paradigms, both of which induce analgesia, but only one via activation of opioid mechanisms. Splenic natural killer cell activity was suppressed by the opioid, but not the nonopioid, form of stress. This suppression was blocked by the opioid antagonist naltrexone. Similar suppression of natural killer activity was induced by high doses of morphine. These results suggest that endogenous opioid peptides mediate the suppressive effect of certain forms of stress on natural killer cell cytotoxicity.

  8. Influence of schedule of interleukin 2 administration on therapy with interleukin 2 and lymphokine activated killer cells.

    PubMed

    Thompson, J A; Lee, D J; Lindgren, C G; Benz, L A; Collins, C; Shuman, W P; Levitt, D; Fefer, A

    1989-01-01

    The purpose of this study was to compare the toxicity, immunomodulatory changes, and antitumor efficacy of interleukin 2 (IL-2) and lymphokine activated killer (LAK) cell therapy with two durations of IL-2 infusion. Patients with progressive melanoma, non-Hodgkin's lymphoma, renal carcinoma, or colon carcinoma received IL-2 at 3 X 10(6) units/m2/day on days 1-5 and 13-17, either by bolus injection every 8 h (q8h) or by continuous i.v. (CIV) administration. Peripheral blood mononuclear cells were harvested by leukapheresis on days 8, 9, and 10, were incubated in vitro for 5 days for generation of LAK cells, and were infused on days 13, 14, and 15. The first 11 patients were treated with IL-2 q8h, and the subsequent 13 patients were treated by CIV infusion. Toxicity consisted primarily of fever, chills, emesis, diarrhea, weight gain, and edema but did not require intensive care unit support and did not differ significantly between treatment groups. IL-2-induced lymphocytosis on day 8 was higher with CIV than with q8h administration with a mean lymphocyte count/microliter of 5610 +/- 700 (SE) versus 3300 +/- 500. Immunomodulatory changes observed on days 8 and 20 were also greater with CIV IL-2 and included an increase in peripheral blood mononuclear cell IL-2 receptor expression as well as a marked rise in the number of Leu-11+ and Leu-19+ peripheral blood mononuclear cells. The total leukapheresis yield per patient and total number of LAK cells infused per patient were higher with CIV than q8h administration, with 49.8 +/- 4.9 X 10(9) versus 39.4 +/- 5.4 X 10(9) and 42.6 +/- 5.0 X 10(9) versus 34.0 +/- 5.4 X 10(9), respectively. The cells infused displayed phenotypic evidence of activation and exhibited marked lytic reactivity to Daudi, Raji, and HT-144 targets. One complete and one minimal response were observed in 2 of 8 patients with metastatic renal cell carcinoma who received CIV IL-2 and LAK cells. The results show that IL-2 is more biologically active by CIV

  9. Activated and expanded natural killer cells target osteosarcoma tumor initiating cells in an NKG2D-NKG2DL dependent manner.

    PubMed

    Fernández, L; Valentín, J; Zalacain, M; Leung, W; Patiño-García, A; Pérez-Martínez, A

    2015-11-01

    Current therapies fail to cure most metastatic or recurrent bone cancer. We explored the efficacy and the pathways involved in natural killer (NK) cells' elimination of osteosarcoma (OS) cells, including tumor initiating cells (TICs), which are responsible for chemotherapy resistance, recurrence, and metastasis. The expression of ligands for NK cell receptors was studied in primary OS cell lines by flow cytometry. In vitro cytotoxicity of activated and expanded NK (NKAE) cells against OS was tested, and the pathways involved explored by using specific antibody blockade. NKAE cells' ability to target OS TICs was analyzed by flow cytometry and sphere formation assays. Spironolactone (SPIR) was tested for its ability to increase OS cells' susceptibility to NK cell lysis in vitro and in vivo. We found OS cells were susceptible to NKAE cells' lysis both in vivo and in vitro, and this cytolytic activity relied on interaction between NKG2D receptor and NKG2D ligands (NKG2DL). SPIR increased OS cells' susceptibility to lysis by NKAE cells, and could shrink the OS TICs. Our results show NKAE cells target OS cells including the TICs compartment, supporting the use of NK-cell based immunotherapies for OS.

  10. Enhanced natural killer cell activation by exopolysaccharides derived from yogurt fermented with Lactobacillus delbrueckii ssp. bulgaricus OLL1073R-1.

    PubMed

    Makino, Seiya; Sato, Asako; Goto, Ayako; Nakamura, Marie; Ogawa, Miho; Chiba, Yoshika; Hemmi, Jun; Kano, Hiroshi; Takeda, Kazuyoshi; Okumura, Ko; Asami, Yukio

    2016-02-01

    Yogurt is generally recognized as a beneficial food for our health, but research into its physiological effects has focused mainly on intestinal dysfunctions such as constipation and diarrhea. We previously found yogurt fermented with Lactobacillus delbrueckii ssp. bulgaricus OLL1073R-1 (hereafter OLL1073R-1) could reduce risks of catching the common cold and flu in human trials. It was assumed that immunostimulatory exopolysaccharide (EPS) produced from OLL1073R-1 play an important role in this context. However, few studies have examined the immunostimulatory effects of traditional Bulgarian yogurts fermented with different strains of lactobacilli and their metabolites. Therefore, we screened 139 L. delbrueckii ssp. bulgaricus strains and identified OLL1073R-1 as the most robust producer of EPS. This strain was also the only strain that induced the production of IFN-γ in vitro. Oral administration of the EPS or yogurt fermented with OLL1073R-1 and Streptococcus thermophilus OLS3059 (OLL1073R-1 yogurt) augmented natural killer (NK) cell activity and induced IFN-γ production in spleen cells in mice, whereas 2 other yogurts fermented with other strains had no effect on NK cell activity. Cellular preparations of the OLL1073R-1 strain also slightly augmented NK cell activity, but were less effective than EPS itself. The EPS-dependent stimulation of NK cell activity was abrogated in IFN-γ knockout mice and in myeloid differentiation factor 88 knockout mice. Furthermore, IFN-γ production from spleen cells stimulated with EPS was completely blocked with both anti-IL-12 and anti-IL-18 antibodies in vitro. These findings suggest that NK cell activation by OLL1073R-1 yogurt is EPS-dependent, occurs via IL-12- and IL-18-mediated IFN-γ production, and requires myeloid differentiation factor 88. We showed that traditional Bulgarian yogurt could exert immunostimulatory effects by selecting starter strains and part of the mechanisms depend on IFN-γ inducible EPS produced

  11. Enhanced natural killer cell activation by exopolysaccharides derived from yogurt fermented with Lactobacillus delbrueckii ssp. bulgaricus OLL1073R-1.

    PubMed

    Makino, Seiya; Sato, Asako; Goto, Ayako; Nakamura, Marie; Ogawa, Miho; Chiba, Yoshika; Hemmi, Jun; Kano, Hiroshi; Takeda, Kazuyoshi; Okumura, Ko; Asami, Yukio

    2016-02-01

    Yogurt is generally recognized as a beneficial food for our health, but research into its physiological effects has focused mainly on intestinal dysfunctions such as constipation and diarrhea. We previously found yogurt fermented with Lactobacillus delbrueckii ssp. bulgaricus OLL1073R-1 (hereafter OLL1073R-1) could reduce risks of catching the common cold and flu in human trials. It was assumed that immunostimulatory exopolysaccharide (EPS) produced from OLL1073R-1 play an important role in this context. However, few studies have examined the immunostimulatory effects of traditional Bulgarian yogurts fermented with different strains of lactobacilli and their metabolites. Therefore, we screened 139 L. delbrueckii ssp. bulgaricus strains and identified OLL1073R-1 as the most robust producer of EPS. This strain was also the only strain that induced the production of IFN-γ in vitro. Oral administration of the EPS or yogurt fermented with OLL1073R-1 and Streptococcus thermophilus OLS3059 (OLL1073R-1 yogurt) augmented natural killer (NK) cell activity and induced IFN-γ production in spleen cells in mice, whereas 2 other yogurts fermented with other strains had no effect on NK cell activity. Cellular preparations of the OLL1073R-1 strain also slightly augmented NK cell activity, but were less effective than EPS itself. The EPS-dependent stimulation of NK cell activity was abrogated in IFN-γ knockout mice and in myeloid differentiation factor 88 knockout mice. Furthermore, IFN-γ production from spleen cells stimulated with EPS was completely blocked with both anti-IL-12 and anti-IL-18 antibodies in vitro. These findings suggest that NK cell activation by OLL1073R-1 yogurt is EPS-dependent, occurs via IL-12- and IL-18-mediated IFN-γ production, and requires myeloid differentiation factor 88. We showed that traditional Bulgarian yogurt could exert immunostimulatory effects by selecting starter strains and part of the mechanisms depend on IFN-γ inducible EPS produced

  12. Natural killer cells and natural killer T cells in Lyme arthritis

    PubMed Central

    2013-01-01

    Introduction Natural killer (NK) and natural killer T (NKT) cells provide a first line of defense against infection. However, these cells have not yet been examined in patients with Lyme arthritis, a late disease manifestation. Lyme arthritis usually resolves with antibiotic treatment. However, some patients have persistent arthritis after spirochetal killing, which may result from excessive inflammation, immune dysregulation and infection-induced autoimmunity. Methods We determined the frequencies and phenotypes of NK cells and invariant NKT (iNKT) cells in paired peripheral blood (PB) and synovial fluid (SF) samples from eight patients with antibiotic-responsive arthritis and fifteen patients with antibiotic-refractory arthritis using flow cytometry and cytokine analyses. Results In antibiotic-responsive patients, who were seen during active infection, high frequencies of CD56bright NK cells were found in SF, the inflammatory site, compared with PB (P <0.001); at both sites, a high percentage of cells expressed the activation receptor NKG2D and the chaperone CD94, a low percentage expressed inhibitory killer immunoglobulin-like receptors (KIR), and a high percentage produced IFN-γ. In antibiotic-refractory patients, who were usually evaluated near the conclusion of antibiotics when few if any live spirochetes remained, the phenotype of CD56bright cells in SF was similar to that in patients with antibiotic-responsive arthritis, but the frequency of these cells was significantly less (P = 0.05), and the frequencies of CD56dim NK cells tended to be higher. However, unlike typical NKdim cells, these cells produced large amounts of IFN-γ, suggesting that they were not serving a cytotoxic function. Lastly, iNKT cell frequencies in the SF of antibiotic-responsive patients were significantly greater compared with that of antibiotic-refractory patients where these cells were often absent (P = 0.003). Conclusions In patients with antibiotic-responsive arthritis

  13. Natural Killer Cells: Key Players in Endometriosis.

    PubMed

    Thiruchelvam, Uma; Wingfield, Mary; O'Farrelly, Cliona

    2015-10-01

    Endometriosis affects more than 10% of women, causing significant pain and morbidity. It is also a significant cause of infertility. The aetiology of the disease remains an enigma, and the mechanisms responsible for the associated infertility are unclear. A role for immune cells in endometriosis has been postulated, with attention directed towards natural killer (NK) cells and macrophages. NK cells kill tumours and infected cells but also have roles in tissue remodelling in several organs including the uterus and are key to successful pregnancy. Here, we explore evidence (from peer-reviewed published articles) of phenotypic and functional abnormalities in NK cell subpopulations of women with endometriosis. It is clear that peripheral blood NK cells and peritoneal NK cells have reduced cytotoxic function in women with endometriosis. Uterine NK cells have a vital role in infertility, but very little research has been carried out in this area. We propose that abnormal u NK cell activity may contribute to the pathogenesis of endometriosis and its associated infertility and that future research should focus on this complex area.

  14. Induction of natural killer cell activity of thoracic duct lymphocytes by polyinosinic-polycytidylic acid (poly(I:C)) or interferon.

    PubMed

    Fresa, K L; Korngold, R; Murasko, D M

    1985-04-01

    Natural killer (NK) cell activity of thoracic duct lymphocytes (TDL) was examined in normal mice and in mice treated with polyinosinic-polycytidylic acid (poly(I:C) and interferon (IFN). TDL from mice treated with phosphate-buffered saline (PBS) expressed little or no NK cell activity against YAC-1 target cells at effector-to-target ratios of up to 200:1, even after in vitro treatment with murine L-cell IFN. In contrast, TDL from poly(I:C)- or IFN-treated mice expressed significant NK activity, which correlated with the significantly higher NK activity of splenocytes from these mice compared to the NK activity of splenocytes from PBS-treated mice. These data indicate that although TDL from normal mice express no detectable NK cell activity, NK cell activity can be induced in TDL by in vivo treatment with poly(I:C) or IFN. PMID:2581698

  15. Requirement of T-lymphokine-activated killer cell-originated protein kinase for TRAIL resistance of human HeLa cervical cancer cells

    SciTech Connect

    Kwon, Hyeok-Ran; Lee, Ki Won; Dong, Zigang; Lee, Kyung Bok; Oh, Sang-Muk

    2010-01-01

    T-lymphokine-activated killer cell-originated protein kinase (TOPK) appears to be highly expressed in various cancer cells and to play an important role in maintaining proliferation of cancer cells. However, the underlying mechanism by which TOPK regulates growth of cancer cells remains elusive. Here we report that upregulated endogenous TOPK augments resistance of cancer cells to apoptosis induced by tumor necrosis factor-related apoptosis inducing ligand (TRAIL). Stable knocking down of TOPK markedly increased TRAIL-mediated apoptosis of human HeLa cervical cancer cells, as compared with control cells. Caspase 8 or caspase 3 activities in response to TRAIL were greatly incremented in TOPK-depleted cells. Ablation of TOPK negatively regulated TRAIL-mediated NF-{kappa}B activity. Furthermore, expression of NF-{kappa}B-dependent genes, FLICE-inhibitory protein (FLIP), inhibitor of apoptosis protein 1 (c-IAP1), or X-linked inhibitor of apoptosis protein (XIAP) was reduced in TOPK-depleted cells. Collectively, these findings demonstrated that TOPK contributed to TRAIL resistance of cancer cells via NF-{kappa}B activity, suggesting that TOPK might be a potential molecular target for successful cancer therapy using TRAIL.

  16. Antigen Presenting Cell-Mediated Expansion of Human Umbilical Cord Blood Yields Log-Scale Expansion of Natural Killer Cells with Anti-Myeloma Activity

    PubMed Central

    Shah, Nina; Martin-Antonio, Beatriz; Yang, Hong; Ku, Stephanie; Lee, Dean A.; Cooper, Laurence J. N.; Decker, William K.; Li, Sufang; Robinson, Simon N.; Sekine, Takuya; Parmar, Simrit; Gribben, John; Wang, Michael; Rezvani, Katy; Yvon, Eric; Najjar, Amer; Burks, Jared; Kaur, Indreshpal; Champlin, Richard E.; Bollard, Catherine M.; Shpall, Elizabeth J.

    2013-01-01

    Natural killer (NK) cells are important mediators of anti-tumor immunity and are active against several hematologic malignancies, including multiple myeloma (MM). Umbilical cord blood (CB) is a promising source of allogeneic NK cells but large scale ex vivo expansion is required for generation of clinically relevant CB-derived NK (CB-NK) cell doses. Here we describe a novel strategy for expanding NK cells from cryopreserved CB units using artificial antigen presenting feeder cells (aAPC) in a gas permeable culture system. After 14 days, mean fold expansion of CB-NK cells was 1848-fold from fresh and 2389-fold from cryopreserved CB with >95% purity for NK cells (CD56+/CD3−) and less than 1% CD3+ cells. Though surface expression of some cytotoxicity receptors was decreased, aAPC-expanded CB-NK cells exhibited a phenotype similar to CB-NK cells expanded with IL-2 alone with respect to various inhibitory receptors, NKG2C and CD94 and maintained strong expression of transcription factors Eomesodermin and T-bet. Furthermore, CB-NK cells formed functional immune synapses with and demonstrated cytotoxicity against various MM targets. Finally, aAPC-expanded CB-NK cells showed significant in vivo activity against MM in a xenogenic mouse model. Our findings introduce a clinically applicable strategy for the generation of highly functional CB-NK cells which can be used to eradicate MM. PMID:24204673

  17. Bovine natural killer cells are present in Escherichia coli infected mammary gland tissue and show antimicrobial activity in vitro.

    PubMed

    Sipka, Anja; Pomeroy, Brianna; Klaessig, Suzanne; Schukken, Ynte

    2016-10-01

    Natural killer (NK) cells are early responders in bacterial infections but their role in bovine mastitis has not been characterized. For the first time, we show the presence of NK cells (NKp46(+)/CD3(-)) in bovine mammary gland tissue after an intramammary challenge with Escherichia (E.) coli. A small number of NK cells was detected in milk from quarters before and during an E. coli challenge. In vitro cultures of primary bovine mammary gland epithelial cells stimulated with UV irradiated E. coli induced significant migration of peripheral blood NK cells (pbNK) within 2h. Furthermore, pbNK cells significantly reduced counts of live E. coli in vitro within 2h of culture. The results show that bovine NK cells have the capacity to migrate to the site of infection and produce antibacterial mediators. These findings introduce NK cells as a leukocyte population in the mammary gland with potential functions in the innate immune response in bovine mastitis. PMID:27638120

  18. Use of a SCID mouse/human lymphoma model to evaluate cytokine-induced killer cells with potent antitumor cell activity

    PubMed Central

    1991-01-01

    C.B-17 severe combined immune deficient (SCID) mice, which lack functional B and T lymphocytes, allow xenografts and, therefore, can be used to study the biology of human malignancies. Two different human B cell lymphoma cell lines, SU-DHL-4 and OCI-Ly8, which both harbor the t(14;18) chromosomal translocation, were injected into C.B-17 SCID mice. Mice injected intravenously or intraperitoneally developed tumors and died in a dose-dependent manner. The presence of tumor cells in various murine tissues could be demonstrated by a clonogenic tumor assay, staining of frozen sections with a monoclonal antibody (mAb) against a human B cell antigen (CD19), and with the polymerase chain reaction technique. A protocol using cytotoxic effector cells was developed and used to selectively deplete the tumor cells from bone marrow. These cells were developed by growing peripheral blood mononuclear cells in the presence of interferon gamma (IFN-gamma), anti- CD3 mAb, and interleukin 2 (IL-2). The timing of IFN-gamma treatment was critical and optimal if IFN-gamma was added before IL-2 treatment. The cells that were stimulated by IFN-gamma, followed by IL-2, could be expanded by treatment with a mAb directed against CD3. These cells could be further activated by IL-1, but not by tumor necrosis factor alpha. With this protocol, a tumor cell kill of 3 logs was obtained as measured by a clonogenic assay. Interestingly, despite their high cytotoxic activity against lymphoma cells, these cells had little toxicity against a subset of normal human hematopoietic precursor cells (granulocyte/macrophage colony-forming units). These cells were further tested by treating murine bone marrow contaminated with the human lymphoma cell line SU-DHL-4, and injecting these cells into SCID mice to assay for tumor growth in vivo. The animals injected with bone marrow contaminated with SU-DHL-4 cells had enhanced survival if the bone marrow was treated with the cytokine-induced killer cells before

  19. Accumulation and activation of natural killer cells in local intraperitoneal HIV-1/MuLV infection results in early control of virus infected cells.

    PubMed

    Johansson, Susanne E; Brauner, Hanna; Hinkula, Jorma; Wahren, Britta; Berg, Louise; Johansson, Maria H

    2011-01-01

    Natural killer (NK) cells are important effectors in resistance to viral infections. The role of NK cells in the acute response to human immunodeficiency virus 1 (HIV-1) infected cells was investigated in a mouse model based on a HIV-1/murine leukemia virus (MuLV) pseudovirus. Splenocytes infected with HIV-1/MuLV were injected intraperitoneally and local immunologic responses and persistence of infected cells were investigated. In vivo depletion with an anti-NK1.1 antibody showed that NK cells are important in resistance to virus infected cells. Moreover, NK cell frequency in the peritoneal cavity increased in response to infected cells and these NK cells had a more mature phenotype, as determined by CD27 and Mac-1 expression. Interestingly, after injection of HIV-1/MuLV infected cells, but not MuLV infected cells, peritoneal NK cells had an increased cytotoxic activity. In conclusion, NK cells play a role in the early control of HIV-1/MuLV infected cells in vivo.

  20. Coordinated Activation of Toll-Like Receptor8 (TLR8) and NLRP3 by the TLR8 Agonist, VTX-2337, Ignites Tumoricidal Natural Killer Cell Activity

    PubMed Central

    Dietsch, Gregory N.; Lu, Hailing; Yang, Yi; Morishima, Chihiro; Chow, Laura Q.; Disis, Mary L.; Hershberg, Robert M.

    2016-01-01

    VTX-2337 (USAN: motolimod) is a selective toll-like receptor 8 (TLR8) agonist, which is in clinical development as an immunotherapy for multiple oncology indications, including squamous cell carcinoma of the head and neck (SCCHN). Activation of TLR8 enhances natural killer cell activation, increases antibody-dependent cell-mediated cytotoxicity, and induces Th1 polarizing cytokines. Here, we show that VTX-2337 stimulates the release of mature IL-1β and IL-18 from monocytic cells through coordinated actions on both TLR8 and the NOD-like receptor pyrin domain containing 3 (NLRP3) inflammasome complex. In vitro, VTX-2337 primed monocytic cells to produce pro-IL-1β, pro-IL-18, and caspase-1, and also activated the NLRP3 inflammasome, thereby mediating the release of mature IL-1β family cytokines. Inhibition of caspase-1 blocked VTX-2337-mediated NLRP3 inflammasome activation, but had little impact on production of other TLR8-induced mediators such as TNFα. IL-18 activated natural killer cells and complemented other stimulatory pathways, including FcγRIII and NKG2D, resulting in IFNγ production and expression of CD107a. NLRP3 activation in vivo was confirmed by a dose-related increase in plasma IL-1β and IL-18 levels in cynomolgus monkeys administered VTX-2337. These results are highly relevant to clinical studies of combination VTX-2337/cetuximab treatment. Cetuximab, a clinically approved, epidermal growth factor receptor-specific monoclonal antibody, activates NK cells through interactions with FcγRIII and facilitates ADCC of tumor cells. Our preliminary findings from a Phase I open-label, dose-escalation, trial that enrolled 13 patients with recurrent or metastatic SCCHN show that patient NK cells become more responsive to stimulation by NKG2D or FcγRIII following VTX-2337 treatment. Together, these results indicate that TLR8 stimulation and inflammasome activation by VTX-2337 can complement FcγRIII engagement and may augment clinical responses in SCCHN

  1. Cancer-associated fibroblast suppresses killing activity of natural killer cells through downregulation of poliovirus receptor (PVR/CD155), a ligand of activating NK receptor

    PubMed Central

    Inoue, Tomoko; Adachi, Katsuyuki; Kawana, Kei; Taguchi, Ayumi; Nagamatsu, Takeshi; Fujimoto, Asaha; Tomio, Kensuke; Yamashita, Aki; Eguchi, Satoko; Nishida, Haruka; Nakamura, Hiroe; Sato, Masakazu; Yoshida, Mitsuyo; Arimoto, Takahide; Wada-Hiraike, Osamu; Oda, Katsutoshi; Osuga, Yutaka; Fujii, Tomoyuki

    2016-01-01

    Cancer-associated fibroblasts (CAFs) play an important role in cancer expansion and progression in tumor microenvironment (TME), via both direct and indirect interactions. Natural killer (NK) cells play a crucial role in anticancer immunity. We investigated the inhibitory effects of CAFs on NK cell activity. CAFs were isolated from endometrial cancer tissue, while normal endometrial fibroblasts (NEFs) were obtained from normal endometrium with no pathological abnormality. NK cells were obtained from allogenic healthy volunteers. CAFs or NEFs were co-cultured at an NK/fibroblast ratio of 1:1 with or without inserted membrane. For NK cell activity, K562 cells were cultured as target cells. NK cell-killing activity was determined by calculating the ratio of PI-positive K562 cells in the presence of NK cells co-cultured with fibroblasts versus NK cells alone. To examine whether NK cell activity was suppressed by IDO pathway, we inhibited IDO activity using the IDO inhibitor 1-MT. We demonstrated that CAFs derived from endometrial cancer induced greater suppression of the killing activity of allogenic NK cells compared with normal endometrial fibroblasts (NEFs). The suppression of NK cell activity by CAFs was inhibited when a membrane was inserted between the CAFs and NK cells, but not by 1-MT, an inhibitor of IDO. We focused on receptor-ligand interactions between CAFs and NK cell and found that cell-surface poliovirus receptor (PVR/CD155), a ligand of activating NK receptor DNAM-1, was downregulated in the CAFs compared with NEFs. To confirm whether PVR downregulation results in the decrease of NK cell-killing activity, PVR expression in NEFs was knocked down using siRNA against PVR (PVRsi). NK cell activity was suppressed by co-culture with PVR-knockdown NEFs, to a similar extent than CAF-induced suppression. CAFs showed increased suppression of NK cell-killing activity compared with NEFs, due to decreased PVR cell surface expression, a ligand of an NK activating

  2. Psychoneuroendocrine immunology: perception of stress can alter body temperature and natural killer cell activity.

    PubMed

    Hiramoto, R N; Solvason, H B; Hsueh, C M; Rogers, C F; Demissie, S; Hiramoto, N S; Gauthier, D K; Lorden, J F; Ghanta, V K

    1999-01-01

    Psychoimmunology has been credited with using the mind as a way to alter immunity. The problem with this concept is that many of the current psychoimmunology techniques in use are aimed at alleviating stress effects on the immune system rather than at direct augmentation of immunity by the brain. Studies in animals provide a model that permits us to approach the difficulties associated with gaining an understanding of the CNS-immune system connection. A particular advantage of using animals over humans is that psychological and social contributions play a less prominent role for animals than for human subjects, since the animals are all inbred and reared under identical controlled conditions. If the insightful information provided by animal studies is correct, then psychotherapy for the treatment of diseases might be made more effective if some aspect of this knowledge is included in the design of the treatment. We emphasize conditioning as a regimen and an acceptable way to train the brain to remember an output pathway to raise immunity. We propose that a specific drug or perception (mild stress, represented by rotation, total body heating or handling) could substitute and kindle the same output pathway without the need for conditioning. If this view is correct, then instead of using conditioning, it may be possible to use an antigen to activate desired immune cells, and substitute a drug or an external environmental sensory stimulus (perception) to energize the output pathway to these cells. Alternatively, monitoring alterations of body temperature in response to a drug or perception might allow us to follow how effectively the brain is performing in altering immunity. Studies with animals suggest that there are alternative ways to use the mind to raise natural or acquired immunity in man.

  3. Medial Septal NMDA Glutamate Receptors are Involved in Modulation of Blood Natural Killer Cell Activity in Rats.

    PubMed

    Podlacha, Magdalena; Glac, Wojciech; Listowska, Magdalena; Grembecka, Beata; Majkutewicz, Irena; Myślińska, Dorota; Plucińska, Karolina; Jerzemowska, Grażyna; Grzybowska, Maria; Wrona, Danuta

    2016-03-01

    The purpose of the present study was to determine the specific role of the medial septal (MS) NMDA glutamate receptors on peripheral blood natural killer cell cytotoxicity (NKCC) and their (large granular lymphocyte, LGL) number, as well as the plasma concentration of tumor necrosis factor α (TNF-α) and corticosterone in male Wistar rats exposed to elevated plus maze (EPM) stress or non-stress conditions. The NMDA groups were injected with NMDA glutamate receptor agonist (N-methyl-D-aspartate; 0.25 μg/rat), the D-AP7 group was injected with DL-2-amino-7-phosphoheptanoate (0.1 μg/rat), an antagonist of NMDA glutamate receptors, and the control Sal group with saline (0.5 μl/rat) via previously implanted cannulae into the MS. There was an increase in the NKCC, NK/LGL number and plasma TNF-α concentration after the NMDA injections, being much stronger within the rats under non-stress conditions rather than the rats exposed to EPM stress. These parameters were decreased in the D-AP7 rats, suggesting receptor/ion channel specificity. Moreover, a lower plasma corticosterone concentration within the NMDA rather than the Sal and D-AP7 groups was found. The obtained results suggest that activation of the NMDA glutamate receptors in the MS, accompanied by changes in the corticosterone and cytokine responses, may be involved in modulation of the blood natural anti-tumor response, under EPM stress and non-stress conditions. PMID:26454750

  4. Present and Future of Allogeneic Natural Killer Cell Therapy

    PubMed Central

    Lim, Okjae; Jung, Mi Young; Hwang, Yu Kyeong; Shin, Eui-Cheol

    2015-01-01

    Natural killer (NK) cells are innate lymphocytes that are capable of eliminating tumor cells and are therefore used for cancer therapy. Although many early investigators used autologous NK cells, including lymphokine-activated killer cells, the clinical efficacies were not satisfactory. Meanwhile, human leukocyte antigen (HLA)-haploidentical hematopoietic stem cell transplantation revealed the antitumor effect of allogeneic NK cells, and HLA-haploidentical, killer cell immunoglobulin-like receptor ligand-mismatched allogeneic NK cells are currently used for many protocols requiring NK cells. Moreover, allogeneic NK cells from non-HLA-related healthy donors have been recently used in cancer therapy. The use of allogeneic NK cells from non-HLA-related healthy donors allows the selection of donor NK cells with higher flexibility and to prepare expanded, cryopreserved NK cells for instant administration without delay for ex vivo expansion. In cancer therapy with allogeneic NK cells, optimal matching of donors and recipients is important to maximize the efficacy of the therapy. In this review, we summarize the present state of allogeneic NK cell therapy and its future directions. PMID:26089823

  5. T-cell receptor beta gene rearrangements in clones derived from human CD4-8- cells expressing natural killer cell activity.

    PubMed

    Christmas, S E; Moore, M

    1988-12-01

    Clones derived from highly purified human peripheral blood Leu 19+ cells in the presence of phytohaemagglutinin (PHA) and interleukin-2 (IL-2) expressed cytotoxic activity against natural killer (NK)-resistant as well as NK-sensitive targets. All 66 clones analysed had a germ line configuration of T-cell receptor (TCR) beta genes and 38/40 also had unrearranged TCR gamma genes. The two exceptions were both CD3+ clones, but these did not have a cytotoxic repertoire noticeably different from CD3- clones without TCR gamma gene rearrangements. Clones were also obtained from highly purified CD4-8- cells, most of which were also cytotoxic for NK-resistant and NK-sensitive targets. About 90% of these clones were CD3+ but only around 50% remained negative for CD4 and CD8 while a significant number (12.7%) were positive for both CD4 and CD8. All clones analysed had rearranged TCR gamma genes and most had also rearranged TCR beta genes, including 20/25 of the clones which were CD3+4-8-. Many of the clones showed two rearrangements of TCR beta genes, and 3/4 CD3- clones had rearranged TCR beta as well as TCR gamma genes. There was no correlation between cytotoxic activity and TCR gene status or phenotype of these CD4-8- derived clones, except that clones which were Leu 19+ tended to have higher cytotoxic activity against NK-sensitive and NK-resistant targets than Leu 19-clones. The results strongly indicate that TCR beta and gamma gene products are not involved in the cytotoxicity mediated by these clones. They also suggest that some CD4-8- cells may be capable of limited differentiation in vitro.

  6. Adoptive immunotherapy of human pancreatic cancer with lymphokine-activated killer cells and interleukin-2 in a nude mouse model

    SciTech Connect

    Marincola, F.M.; Da Pozzo, L.F.; Drucker, B.J.; Holder, W.D. Jr. )

    1990-11-01

    A pancreatic cancer cell line was grown in orthotopic and heterotopic positions in young Swiss/NIH nude mice, which were tested with adoptive immunotherapy. Mice were injected with 1 x 10(7) human cancer cells in the subcutaneous tissue and duodenal lobe of the pancreas. The mice were randomly divided into four groups: group IA (LAK + IL-2) (N = 25) received 2 X 10(7) human lymphokine-activated killer (LAK) cells from normal donors by tail vein injection followed by 10,000 units of human recombinant interleukin-2 (IL-2) given intraperitoneally every 12 hours for 28 days; group IB (IL-2) (N = 27) was given the same dose of IL-2 alone; group IC (RPMI-1640) (N = 18) received a placebo consisting of 1 ml of RPMI-1640 intraperitoneally every 12 hours; and group ID (LAK) (N = 14) received 2 X 10(7) LAK cells but no IL-2. Toxicity was significantly higher in group IB, with a mortality rate of 45.5% (10/22 animals) versus a 0% mortality (0/25) in group IA. None of the group IA or IB animals died of pancreatic cancer during the experiment. The animals that did not receive IL-2 died before 28 days in 14.2% of group IC and in 16.7% of group ID. The area under the growth curve of subcutaneous tumors during the course of treatment and the pancreatic tumor weight at the end of treatment were compared in each group. Subcutaneous tumors had a reduced rate of growth in group IA animals compared to all the other treatments. Pancreatic tumor growth was slowed in group IA. The animals treated with IL-2 alone (group IB) showed some slowing of tumor growth that was intermediate between group IA, group IC, and group ID. A similar experiment was done with irradiated (375 rad) mice. Nine nude mice with tumors were treated with LAK + IL-2 (group IIA), eight received IL-2 alone (group IIB), and seven received placebo (group IIC).

  7. Human Induced Pluripotent Stem Cells Are Targets for Allogeneic and Autologous Natural Killer (NK) Cells and Killing Is Partly Mediated by the Activating NK Receptor DNAM-1

    PubMed Central

    Monecke, Sebastian; Cyganek, Lukas; Elsner, Leslie; Hübscher, Daniela; Walter, Lutz; Streckfuss-Bömeke, Katrin; Guan, Kaomei; Dressel, Ralf

    2015-01-01

    Human induced pluripotent stem cells (hiPSCs) could be used to generate autologous cells for therapeutic purposes, which are expected to be tolerated by the recipient. However, iPSC-derived grafts are at risk of giving rise to teratomas in the host, if residuals of tumorigenic cells are not rejected by the recipient. We have analyzed the susceptibility of hiPSC lines to allogeneic and autologous natural killer (NK) cells. IL-2-activated, in contrast to resting NK cells killed hiPSC lines efficiently (P=1.69x10-39). Notably, the specific lysis of the individual hiPSC lines by IL-2-activated NK cells was significantly different (P=1.72x10-6) and ranged between 46 % and 64 % in 51Cr-release assays when compared to K562 cells. The hiPSC lines were killed by both allogeneic and autologous NK cells although autologous NK cells were less efficient (P=8.63x10-6). Killing was partly dependent on the activating NK receptor DNAM-1 (P=8.22x10-7). The DNAM-1 ligands CD112 and CD155 as well as the NKG2D ligands MICA and MICB were expressed on the hiPSC lines. Low amounts of human leukocyte antigen (HLA) class I proteins, which serve as ligands for inhibitory and activating NK receptors were also detected. Thus, the susceptibility to NK cell killing appears to constitute a common feature of hiPSCs. Therefore, NK cells might reduce the risk of teratoma formation even after autologous transplantations of pluripotent stem cell-derived grafts that contain traces of pluripotent cells. PMID:25950680

  8. Copy number variations of HLA-I and activation of NKp30 pathway determine the sensitivity of gastric cancer cells to the cytotoxicity of natural killer cells.

    PubMed

    Xing, R; Li, L; Chen, L; Gao, Z; Wang, H; Li, W; Cui, J; Tian, G; Liang, Q; Yu, J; Sung, J J; Luo, G; Gao, H; Xu, X; Yang, H; Wang, J; Zhang, X; Wang, J M; Huang, J; Yu, Y; Wang, J; Lu, Y

    2016-05-19

    Nude mice are important in vivo model for characterization of cell malignancy behavior; however, many cancer cells fail to form tumors in it. Understanding this defective mechanism may provide novel insights into tumorigenesis and how tumor cells escape innate immunity. Whole-genome sequencing was conducted on two gastric cancer (GC) cells, BGC823 and AGS, which do and do not form tumors in nude mice, to identify their genomic differences relevant to natural killer (NK) cells. We found that the tumorigenic capacity of human GC cell lines was dependent on the recruitment and activation of NK cells in xenograft tumors. We used whole-genome sequence (WGS) on GC cell lines to identify potential genes controlling susceptibility to NK-mediated killing. The tumorigenic cell line BGC823 expressed high levels of HLA-I because of copy gain and was resistant to NK cell killing. In contrast, another cell line AGS expressing low levels of HLA-I with activated NKp30/MAPK/IL-12 (interleukin-12) or IL-2 (interleukin-2) pathway was susceptible to NK lysis. Treatment of tumor bearing mice with systemic administration of IL-12 in combination with intratumor injection of anti-HLA-I antibody significantly increased NK cell recruitment into xenograft tumors, which became sensitive to NK killing, resulting in reduced tumor progression. In human GC specimens, decreased HLA-I expression and increased NK cells surrounding tumor cells were correlated with decreased metastasis potential and better prognosis of patients. Our results provide a mechanistic basis for GC cells to escape NK lysis and a promising prospect of NK immunotherapy for GC cells.

  9. Renal cell carcinoma-infiltrating natural killer cells express differential repertoires of activating and inhibitory receptors and are inhibited by specific HLA class I allotypes.

    PubMed

    Schleypen, Julia S; Von Geldern, Marion; Weiss, Elisabeth H; Kotzias, Nicole; Rohrmann, Karl; Schendel, Dolores J; Falk, Christine S; Pohla, Heike

    2003-10-10

    Among tumor-infiltrating lymphocytes (TILs) directly isolated from renal cell carcinomas (RCCs), we found substantial numbers of natural killer (NK) cells in most tumor tissues. They could be identified reliably in situ with an antibody directed against the activating receptor (AR) NKp46 that is exclusively expressed by all NK cells. NK-enriched TILs (NK-TILs) showed cytotoxicity against major histocompatibility complex (MHC) class I-negative cell lines. The ability to detect lysis of target cells was dependent on the percentage of NK cells within the TILs, and cytotoxicity was only observed after overnight activation with low-dose interleukin-2 (IL-2). Infiltrating NK cells were found to express various inhibitory receptors (IRs); among these the CD94/NKG2A receptor complex was overrepresented compared to the autologous peripheral blood mononuclear cell (PBMC) population. Other IRs were underrepresented, indicating that NK subpopulations vary in their tumor-infiltrating capacity. IRs expressed by NK-TILs are functional since receptor engagement with MHC class I ligands presented by human leukocyte antigen (HLA)-transfected target cell lines was able to inhibit NK-mediated cytotoxicity. NK-TILs were also able to lyse autologous or allogeneic tumor cell lines in vitro. This activity correlated with low HLA class I surface expression since lysis could be inhibited by interferon (IFN)-gamma-expressing RCC transductants that displayed a higher surface density of HLA class I molecules. Therefore, NK cells infiltrating tumor tissues have an inherent ability to recognize transformed cells, but they require cytokine activation and are sensitive to inhibition by IR ligands.

  10. Enhancement of tumor cell susceptibility to lymphokine-activated killer cells by treatment with the streptococcal preparation OK432.

    PubMed

    Yamaue, H; Tanimura, H; Tsunoda, T; Iwahashi, M; Tani, M; Tamai, M; Noguchi, K; Hotta, T; Arii, K

    1992-01-01

    We investigated whether tumor cell lysis by LAK cells was augmented by treatment with OK432 in vitro. NK and LAK activity against K562 cells was not enhanced by their treatment with OK432. In contrast, the susceptibility of OK432-treated Daudi and KATO-III cells to lysis by LAK cells was enhanced. Succinate dehydrogenase activity and RNA synthesis were impaired in Daudi and KATO-III cells by treatment with OK432, and moreover the expression of HLA Class I antigen and beta 2-microglobulin was inhibited in OK432-treated KATO-III cells. Thus, it is suggested that the enhancement of the susceptibility of OK432-treated tumor cells with regard to succinate dehydrogenase activity, RNA synthesis, and HLA Class I antigen expression.

  11. The assessment of cytotoxic T cell and natural killer cells activity in residents of high and ordinary background radiation areas of Ramsar-Iran.

    PubMed

    Borzoueisileh, Sajad; Monfared, Ali Shabestani; Abediankenari, Saeid; Mostafazadeh, Amrollah

    2013-01-01

    The effective radiation dose of human from natural sources is about 2.4 mSv/y and the dose limit for radiation workers is 20 mSv/y. Ramsar, a city in Iran, has been the subject of concern in the last forty years for a high level of radiation measured in some spots as high as 260 mSv/y. Carcinogenesis is one of the most studied effects of radiation especially in high doses. Recent studies showed that the high level of natural radiation received by inhabitants of this area, paradoxically don't have significant health effect. Natural killer (NK) cells and cytotoxic T cells are the most important cells in tumor immune surveillance and CD107a is a widely expressed intracellular protein located in the lysosomal/endosomal membrane. CD107a transiently located on the cell membrane can be used as a marker of CD8 + T cell degranulation following stimulation. It is also expressed, to a lower extent, on activated NK cells. In this study, 60 healthy people were selected randomly and their consent obtained and confounding factors such as sex, age, life-styles was matched then the count of activated NK and CD8 + cells was compared in high and normal background radiation areas inhabitants of Ramsar. After filling the questionnaire and measurement of background radiation, blood samples of 30 healthy people from each region were analyzed immediately by means of flowcytometry. The leukocytes and their subsets were not significantly different between two groups and the count of active cells was higher in control group. The result shows that the changes in immune system occur due to radiation and maybe it is as a result of higher radiosensitivity of activated cells. PMID:23531635

  12. Antitumor activities of valproic acid on Epstein-Barr virus-associated T and natural killer lymphoma cells.

    PubMed

    Iwata, Seiko; Saito, Takashi; Ito, Yoshinori; Kamakura, Maki; Gotoh, Kensei; Kawada, Jun-Ichi; Nishiyama, Yukihiro; Kimura, Hiroshi

    2012-02-01

    Epstein-Barr virus (EBV), which infects B cells, T cells, and natural killer (NK) cells, is associated with multiple lymphoid malignancies. Recently, histone deacetylase (HDAC) inhibitors have been reported to have anticancer effects against various tumor cells. In the present study, we evaluated the killing effect of valproic acid (VPA), which acts as an HDAC inhibitor, on EBV-positive and -negative T and NK lymphoma cells. Treatment of multiple T and NK cell lines (SNT13, SNT16, Jurkat, SNK6, KAI3 and KHYG1) with 0.1-5 mM of VPA inhibited HDAC, increased acetylated histone levels and reduced cell viability. No significant differences were seen between EBV-positive and -negative cell lines. Although VPA induced apoptosis in some T and NK cell lines (SNT16, Jurkat and KHYG1) and cell cycle arrest, it did not induce lytic infection in EBV-positive T or NK cell lines. Because the killing effect of VPA was modest (1 mM VPA reduced cell viability by between 22% and 56%), we tested the effects of the combination of 1 mM of VPA and 0.01 μM of the proteasome inhibitor bortezomib. The combined treated of cells with VPA and bortezomib had an additive killing effect. Finally, we administered VPA to peripheral blood mononuclear cells from three patients with EBV-associated T or NK lymphoproliferative diseases. In these studies, VPA had a greater killing effect against EBV-infected cells than uninfected cells, and the effect was increased when VPA was combined with bortezomib. These results indicate that VPA has antitumor effects on T and NK lymphoma cells and that VPA and bortezomib may have synergistic effects, irrespective of the presence of EBV. PMID:22017376

  13. Brominated Flame Retardants, Tetrabromobisphenol A and Hexabromocyclododecane, Activate Mitogen-Activated Protein Kinases (MAPKs) in Human Natural Killer Cells

    PubMed Central

    Cato, Anita; Celada, Lindsay; Kibakaya, Esther Caroline; Simmons, Nadia; Whalen, Margaret M.

    2014-01-01

    NK cells provide a vital surveillance against virally infected cells, tumor cells, and antibody-coated cells through the release of cytolytic mediators and gamma interferon (IFN-γ). Hexabromocyclododecane (HBCD) is a brominated flame retardant used primarily in expanded (EPS) and extruded (XPS) polystyrene foams for thermal insulation in the building and construction industry. Tetrabromobisphenol A (TBBPA) is used both as a reactive and an additive flame retardant in a variety of materials. HBCD and TBBPA contaminate the environment and are found in human blood samples. In previous studies, we have shown that other environmental contaminants, such as the dibutyltin (DBT) and tributyltin (TBT), decrease NK lytic function by activating mitogen-activated protein kinases (MAPKs) in the NK cells. HBCD and TBBPA also interfere with NK cell(s) lytic function. The current study evaluates whether HBCD and/or TBBPA have the capacity to activate MAPKs and MAPK kinases (MAP2Ks). The effects of concentrations of HBCD and TBBPA that inhibited lytic function on the phosphorylation state and total levels of the MAPKs (p44/42, p38, and JNK) and the phosphorylation and total levels of the MAP2Ks (MEK1/2 and MKK3/6) were examined. Results indicate that exposure of human NK cells to 10-0.5 µM HBCD or TBBPA activate MAPKs and MAP2Ks. This HBCD and TBBPA-induced activation of MAPKs may leave them unavailable for activation by virally infected or tumor target cells and thus contributes to the observed decreases in lytic function seen in NK cells exposed to HBCD and TBBPA. PMID:25341744

  14. Natural killer cells in hepatitis B virus infection.

    PubMed

    Wu, Shao-fei; Wang, Wen-jing; Gao, Yue-qiu

    2015-01-01

    Natural killer cells are a unique type of lymphocytes with cytotoxic capacity, and play important roles against tumors and infections. Recently, natural killer cells have been increasingly valued in their effects in hepatitis B virus infection. Since hepatitis B virus is not cytopathic, the subsequent antiviral immune responses of the host are responsible for sustaining the liver injury, which may result in cirrhosis and even hepatocellular carcinoma. Many studies have confirmed that natural killer cells participate in anti-hepatitis B virus responses both in the early phase after infection and in the chronic phase via cytolysis, degranulation, and cytokine secretion. However, natural killer cells play dichotomic roles: they exert antiviral and immunoregulatory functions whilst contribute to the pathogenesis of liver injury. Here, we review the roles of natural killer cells in hepatitis B virus infection, introducing novel therapeutic strategies for controlling hepatitis B virus infection via the modulation of natural killer cells.

  15. Bringing natural killer cells to the clinic: ex vivo manipulation.

    PubMed

    Childs, Richard W; Berg, Maria

    2013-01-01

    Recently, there has been a substantial gain in our understanding of the role that natural killer (NK) cells play in mediating innate host immune responses against viruses and cancer. Although NK cells have long been known to be capable of killing cancer cells independently of antigen recognition, the full therapeutic potential of NK cell-based immunotherapy has yet to be realized. Here we review novel methods to activate and expand human NK cells ex vivo for adoptive transfer in humans, focusing on the important phenotypic and functional differences observed among freshly isolated, cytokine activated, and ex vivo-expanded NK populations. PMID:24319186

  16. Imaging Lung Clearance of Radiolabeled Tumor Cells to Study Mice with Normal, Activated or Depleted Natural Killer (NK) Cells

    SciTech Connect

    Kulkarni, P.V.; Bennett, M.; Constantinescu, A.; Arora, V.; Viguet, M.; Antich, P.; Parkey, R.W.; Mathews, D.; Mason, R.P.; Oz, O.K.

    2003-08-26

    Lung clearance of 51CR and 125I iododeoxyuridine (IUDR) labeled cancer cells assess NK cell activity. It is desirable to develop noninvasive imaging technique to assess NK activity in mice. We labeled target YAC-1 tumor cells with 125I, 111In, 99mTc, or 67Ga and injected I.V. into three groups of BALB/c mice. Animals were treated with medium (group I), 300mg/kg cyclophosmamide (CY) to kill NK cell (group II), or anti-LY49C/1) (ab')2 mAb to augment NK function (group III). Lungs were removed 15 min or 2 h later for tissue counting. Control and treated mice were imaged every 5 min with a scintillating camera for 1 h after 15 min of infusion of the 111In labeled cells. Lung clearance increased after 15 min (lodging: 60-80%) and (2 h retention: 3-7%). Similar results were obtained with all the isotopes studied. Images distinguished the control and treated mice for lung activity. Cells labeled with 111In, 99mTc or 67Ga are cleared similar to those labeled with 51Cr or 125I. NK cell destruction of tumor cells may be assessed by noninvasive imaging method either by SPECT (99mTc, 111In, 67Ga) or by PET (68Ga)

  17. Role of protein kinase C in the TBT-induced inhibition of lytic function and MAPK activation in human natural killer cells

    PubMed Central

    Abraha, Abraham B.; Rana, Krupa; Whalen, Margaret M.

    2010-01-01

    Human natural killer (NK) cells are lymphocytes that destroy tumor and virally infected cells. Previous studies have shown that exposures of NK cells to tributyltin (TBT) greatly diminish their ability to destroy tumor cells (lytic function) while activating mitogen-activated protein kinases (MAPK) (p44/42, p38, and JNK) in the NK cells. The signaling pathway that regulates NK lytic function appears to include activation of protein kinase C (PKC) as well as MAPK activity. The TBT-induced activation of MAPKs would trigger a portion of the NK lytic signaling pathway, which would then leave the NK cell unable to trigger this pathway in response to a subsequent encounter with a target cell. In the present study we evaluated the involvement of PKC in the inhibition of NK lysis of tumor cells and activation of MAPKs caused by TBT exposures. TBT caused a 2–3 fold activation of PKC at concentrations ranging from 50–300 nM (16–98 ng/mL), indicating that activation of PKC occurs in response to TBT exposures. This would then leave the NK cell unable to respond to targets. Treatment with the PKC inhibitor, bisindolylmaleimide I, caused an 85% decrease in the ability of NK cells to lyse tumor cells validating the involvement of PKC in the lytic signaling pathway. The role of PKC in the activation of MAPKs by TBT was also investigated using bisindolylmaleimide I. The results indicated that in NK cells where PKC activation was blocked there was no activation of the MAPK, p44/42 in response to TBT. However, TBT-induced activation of the MAPKs, p38 and JNK did not require PKC activation. These results indicate the pivotal role of PKC in the TBT-induced loss of NK lytic function including the activation of p44/42 by TBT in NK cells. PMID:20390410

  18. Comparison of the proliferation, cytotoxic activity and cytokine secretion function of cascade primed immune cells and cytokine-induced killer cells in vitro.

    PubMed

    Li, Gui-Xin; Zhao, Shu-Shu; Zhang, Xu-Guang; Wang, Wen-Hao; Liu, Jin; Xue, Ke-Wei; Li, Xiao-Yan; Guo, Ying-Xue; Wang, Li-Hua

    2015-08-01

    The present study aimed to compare the antitumor effects of cascade primed immune (CAPRI) cells and cytokine-induced killer (CIK) cells in vitro, through investigating cell morphology, proliferation, cytotoxic activity to tumor cells and the ability of these cells to secrete cytokines. Peripheral blood samples (50 ml) were obtained from three healthy volunteers and peripheral blood mononuclear cells (PBMCs) were obtained from each via Ficoll-Conray density gradient centrifugation. Each suspension of PBMCs (1 x 10(6)/ml) was divided into two parts; CAPRI cells were obtained from one part through a series of induction, amplification and cytokine cultures, while CIK cells were obtained from the other part through induction with different cytokines. During the culture process, the proliferation and morphological changes were observed for the two cell types using Trypan blue staining. At day 14, the cytotoxic activity of the two cell types was examined through determining lactate dehydrogenase release in the presence of K562 leukemia cells and MCF-7 breast cancer cells. In addition, secretory levels of interferon (IFN)-γ and interleukin (IL)-2 were detected using enzyme-linked immunospot (ELISPOT) technology. The results revealed that at day 5 and 14 of culture, there were significantly fewer CAPRI cells compared with CIK cells (P<0.001), although the survival rate of each cell type was >95%. The cytotoxic activity of CAPRI cells towards the K562 cell line was effector-target ratio-dependent (40:1 and 20:1) with values of 55.1 ± 3.25 and 35.0 ± 2.65%, respectively, which were significantly reduced compared with the corresponding data in CIK cells, 60.0 ± 3.03 and 39.7 ± 3.42% (P=0.004 and 0.005, respectively). Furthermore, the cytotoxic activity of CAPRI cells towards MCF-7 cells were 71.5 ± 3.06, 56.0 ± 3.76 and 40.2 ± 2.90% at effector-target ratios 40:1, 20:1 and 10:1, respectively. These data were significantly higher than the corresponding values in CIK

  19. Cloning and characterization of the 2B4 gene encoding a molecule associated with non-MHC-restricted killing mediated by activated natural killer cells and T cells

    SciTech Connect

    Mathew, P.A.; Garni-Wagner, B.A.; Land, K.; Takashima, A.; Stoneman, E.; Bennett, M.; Kumar, V. )

    1993-11-15

    The authors have recently described a signal transducing molecule, 2B4, expressed on all NK and T cells that mediate non-MHC-restricted killing. The gene encoding this molecule was cloned and its nucleotide sequence determined. The encoded protein of 398 amino acids has a leader peptide of 18 amino acids and a transmembrane region of 24 amino acids. The predicted protein has eight N-linked glycosylation sites, suggesting that it is highly glycosylated. Comparison of 2B4 with sequences in the databanks indicates that 2B4 is a member of the Ig supergene family, and it shows homology to murine and rat CD48 and human LFA-3. Northern blot analysis has shown at least three transcripts for 2B4 in adherent lymphokine-activated killer cells of several mouse strains and TCR-[gamma]/[delta] dendritic epidermal T cell lines but not in allospecific T cell clones. These three mRNA are the products of differential splicing of heterogeneous nuclear RNA. Southern blot analysis of genomic DNA from several mouse strains revealed that 2B4 belongs to a family of closely related genes. The 2B4 gene has been mapped to mouse chromosome 1 by analysis of 2B4 expression in recombinant inbred mouse strains. 48 refs., 6 figs., 2 tabs.

  20. Interleukin-15-activated natural killer cells kill autologous osteoclasts via LFA-1, DNAM-1 and TRAIL, and inhibit osteoclast-mediated bone erosion in vitro.

    PubMed

    Feng, Shan; Madsen, Suzi H; Viller, Natasja N; Neutzsky-Wulff, Anita V; Geisler, Carsten; Karlsson, Lars; Söderström, Kalle

    2015-07-01

    Osteoclasts reside on bone and are the main bone resorbing cells playing an important role in bone homeostasis, while natural killer (NK) cells are bone-marrow-derived cells known to play a crucial role in immune defence against viral infections. Although mature NK cells traffic through bone marrow as well as to inflammatory sites associated with enhanced bone erosion, including the joints of patients with rheumatoid arthritis, little is known about the impact NK cells may have on mature osteoclasts and bone erosion. We studied the interaction between human NK cells and autologous monocyte-derived osteoclasts from healthy donors in vitro. We show that osteoclasts express numerous ligands for receptors present on activated NK cells. Co-culture experiments revealed that interleukin-15-activated, but not resting, NK cells trigger osteoclast apoptosis in a dose-dependent manner, resulting in drastically decreased bone erosion. Suppression of bone erosion requires contact between NK cells and osteoclasts, but soluble factors also play a minor role. Antibodies masking leucocyte function-associated antigen-1, DNAX accessory molecule-1 or tumour necrosis factor-related apoptosis-inducing ligand enhance osteoclast survival when co-cultured with activated NK cells and restore the capacity of osteoclasts to erode bone. These results suggest that interleukin-15-activated NK cells may directly affect bone erosion under physiological and pathological conditions.

  1. Interleukin-15-activated natural killer cells kill autologous osteoclasts via LFA-1, DNAM-1 and TRAIL, and inhibit osteoclast-mediated bone erosion in vitro

    PubMed Central

    Feng, Shan; Madsen, Suzi H; Viller, Natasja N; Neutzsky-Wulff, Anita V; Geisler, Carsten; Karlsson, Lars; Söderström, Kalle

    2015-01-01

    Osteoclasts reside on bone and are the main bone resorbing cells playing an important role in bone homeostasis, while natural killer (NK) cells are bone-marrow-derived cells known to play a crucial role in immune defence against viral infections. Although mature NK cells traffic through bone marrow as well as to inflammatory sites associated with enhanced bone erosion, including the joints of patients with rheumatoid arthritis, little is known about the impact NK cells may have on mature osteoclasts and bone erosion. We studied the interaction between human NK cells and autologous monocyte-derived osteoclasts from healthy donors in vitro. We show that osteoclasts express numerous ligands for receptors present on activated NK cells. Co-culture experiments revealed that interleukin-15-activated, but not resting, NK cells trigger osteoclast apoptosis in a dose-dependent manner, resulting in drastically decreased bone erosion. Suppression of bone erosion requires contact between NK cells and osteoclasts, but soluble factors also play a minor role. Antibodies masking leucocyte function-associated antigen-1, DNAX accessory molecule-1 or tumour necrosis factor-related apoptosis-inducing ligand enhance osteoclast survival when co-cultured with activated NK cells and restore the capacity of osteoclasts to erode bone. These results suggest that interleukin-15-activated NK cells may directly affect bone erosion under physiological and pathological conditions. PMID:25684021

  2. Endogenous production of interleukin 15 by activated human monocytes is critical for optimal production of interferon-gamma by natural killer cells in vitro.

    PubMed Central

    Carson, W E; Ross, M E; Baiocchi, R A; Marien, M J; Boiani, N; Grabstein, K; Caligiuri, M A

    1995-01-01

    Natural killer (NK) cells are large granular lymphocytes that constitutively express functional IL-2 receptors. We have shown that recombinant human IL-15 uses the IL-2 receptor to activate human NK cells and can synergize with recombinant human IL-12 to stimulate NK cell production of IFN-gamma in vitro. IFN-gamma production by NK cells is critical in the prevention of overwhelming infection by obligate intracellular microbial pathogens in several experimental animal models. Herein, we demonstrate that human monocytes produce IL-15 protein within 5 h of activation with LPS. Using an IL-15-neutralizing antiserum in a coculture of LPS-activated monocytes and NK cells, we demonstrate that monocyte-derived IL-15 is critical for optimal NK cell production of IFN-gamma. Endogenous IL-15 activates NK cells through the IL-2 receptor, and with endogenous IL-12, regulates NK cell IFN-gamma after monocyte activation by LPS. These in vitro studies are the first to characterize a function for endogenous IL-15, and as such, suggest an important role for IL-15 during the innate immune response. IL-15 may be an important ligand for the NK cell IL-2 receptor in vivo. Images PMID:8675621

  3. Isolation and identification of normal killer cells from Syrian hamsters

    SciTech Connect

    Matveeva, V.A.; Klyuchareva, T.E.

    1986-09-01

    This paper gives data on isolation of normal killer cells from the blood and various tissues of Syrian hamsters in a Percoll density gradient and their identification on the basis of morphologic criteria and cytotoxic activity (CTA). CTA of the isolated cells was studied in the cytotoxic test with target cells of a human MOLT-4 thymoma cell labeled with /sup 51/Cr. Isolation of large granular lymphocytes from blood, spleen, and bone marrow of Syrian hamsters in Percoll density gradient is shown in the results of five experiments used for cells of each type.

  4. [Change in the activity of natural killer cells in normal subjects and in virus diseases on exposure to interferon in vitro].

    PubMed

    Petrov, R V; Saidov, M Z; Koval'chuk, L V; Sorokin, A M; Kaganov, B S

    1984-04-01

    The activity of natural killers was examined in peripheral blood of healthy subjects and patients with chronic hepatitis and disseminated sclerosis. An attempt was made to correct natural killer activity by human leukocyte interferon in vitro. To assess the activity of natural killers, use was made of the method of serial dilutions. An optimal effector/target ratio was employed in experiments. The patients with chronic hepatitis and disseminated sclerosis demonstrated a reduction in the activity of natural killers whatever the effector/target ratio. The action of interferon in vitro is specific immunomodulatory in nature. Administration of interferon in a dose of 250 Units/ml raises the magnitude of the cytotoxic index in healthy donors and in patients with chronic hepatitis and disseminated sclerosis, making the shape of the killer activity curve approach that of normal. Such an approach can be used for preliminary assessment of the sensitivity of natural killers to interferon in viral diseases of man. The potentialities and efficacy of interferon in clinical medicine are discussed.

  5. Natural Killer Cell Reduction and Uteroplacental Vasculopathy.

    PubMed

    Golic, Michaela; Haase, Nadine; Herse, Florian; Wehner, Anika; Vercruysse, Lisbeth; Pijnenborg, Robert; Balogh, Andras; Saether, Per Christian; Dissen, Erik; Luft, Friedrich C; Przybyl, Lukasz; Park, Joon-Keun; Alnaes-Katjavivi, Patji; Staff, Anne Cathrine; Verlohren, Stefan; Henrich, Wolfgang; Muller, Dominik N; Dechend, Ralf

    2016-10-01

    Uterine natural killer cells are important for uteroplacental development and pregnancy maintenance. Their role in pregnancy disorders, such as preeclampsia, is unknown. We reduced the number of natural killer cells by administering rabbit anti-asialo GM1 antiserum in an established rat preeclamptic model (female human angiotensinogen×male human renin) and evaluated the effects at the end of pregnancy (day 21), compared with preeclamptic control rats receiving normal rabbit serum. In 100% of the antiserum-treated, preeclamptic rats (7/7), we observed highly degenerated vessel cross sections in the mesometrial triangle at the end of pregnancy. This maternal uterine vasculopathy was characterized by a total absence of nucleated/living cells in the vessel wall and perivascularly and prominent presence of fibrosis. Furthermore, there were no endovascular trophoblast cells within the vessel lumen. In the control, normal rabbit serum-treated, preeclamptic rats, only 20% (1/5) of the animals displayed such vasculopathy. We confirmed the results in healthy pregnant wild-type rats: after anti-asialo GM1 treatment, 67% of maternal rats displayed vasculopathy at the end of pregnancy compared with 0% in rabbit serum-treated control rats. This vasculopathy was associated with a significantly lower fetal weight in wild-type rats and deterioration of fetal brain/liver weight ratio in preeclamptic rats. Anti-asialo GM1 application had no influence on maternal hypertension and albuminuria during pregnancy. Our results show a new role of natural killer cells during hypertensive pregnancy in maintaining vascular integrity. In normotensive pregnancy, this integrity seems important for fetal growth.

  6. Natural Killer Cell Reduction and Uteroplacental Vasculopathy.

    PubMed

    Golic, Michaela; Haase, Nadine; Herse, Florian; Wehner, Anika; Vercruysse, Lisbeth; Pijnenborg, Robert; Balogh, Andras; Saether, Per Christian; Dissen, Erik; Luft, Friedrich C; Przybyl, Lukasz; Park, Joon-Keun; Alnaes-Katjavivi, Patji; Staff, Anne Cathrine; Verlohren, Stefan; Henrich, Wolfgang; Muller, Dominik N; Dechend, Ralf

    2016-10-01

    Uterine natural killer cells are important for uteroplacental development and pregnancy maintenance. Their role in pregnancy disorders, such as preeclampsia, is unknown. We reduced the number of natural killer cells by administering rabbit anti-asialo GM1 antiserum in an established rat preeclamptic model (female human angiotensinogen×male human renin) and evaluated the effects at the end of pregnancy (day 21), compared with preeclamptic control rats receiving normal rabbit serum. In 100% of the antiserum-treated, preeclamptic rats (7/7), we observed highly degenerated vessel cross sections in the mesometrial triangle at the end of pregnancy. This maternal uterine vasculopathy was characterized by a total absence of nucleated/living cells in the vessel wall and perivascularly and prominent presence of fibrosis. Furthermore, there were no endovascular trophoblast cells within the vessel lumen. In the control, normal rabbit serum-treated, preeclamptic rats, only 20% (1/5) of the animals displayed such vasculopathy. We confirmed the results in healthy pregnant wild-type rats: after anti-asialo GM1 treatment, 67% of maternal rats displayed vasculopathy at the end of pregnancy compared with 0% in rabbit serum-treated control rats. This vasculopathy was associated with a significantly lower fetal weight in wild-type rats and deterioration of fetal brain/liver weight ratio in preeclamptic rats. Anti-asialo GM1 application had no influence on maternal hypertension and albuminuria during pregnancy. Our results show a new role of natural killer cells during hypertensive pregnancy in maintaining vascular integrity. In normotensive pregnancy, this integrity seems important for fetal growth. PMID:27550919

  7. Correlation of natural killer cell activity and clearance of Cryptococcus neoformans from mice after adoptive transfer of splenic nylon wool-nonadherent cells.

    PubMed Central

    Hidore, M R; Murphy, J W

    1986-01-01

    Previous reports demonstrate that natural killer (NK) cells inhibit the growth of Cryptococcus neoformans in vitro, but conclusive evidence supporting the effectiveness of NK cells in host resistance to cryptococci is not available. The objective of these studies was to assess the ability of NK cells to clear C. neoformans from the lungs, livers, and spleens of infected mice. CBA/J mice were depleted of NK cells, as well as other natural effector cells, by an intraperitoneal injection of cyclophosphamide (Cy), 240 mg/kg of body weight. One day later, 7.5 X 10(7) nylon wool-nonadherent (NWN) spleen cells, either untreated or treated with anti-asialo GM1 and complement to remove NK cells, were adoptively transferred to Cy-pretreated mice. On day 2 after Cy treatment, the mice were injected intravenously with 2 X 10(4) cryptococci. At 4 and 6 days after Cy treatment, tissues were assayed for NK reactivity, using a 4-h 51Cr-release assay, and for in vivo clearance of cryptococci as reflected by mean log10 CFU per organ. We observed that Cy treatment depleted NK activity against YAC-1 targets and reduced in vivo clearance of C. neoformans from the tissues of infected mice. Additionally, Cy treatment depleted the total lung and spleen cellularity and the total number of peripheral blood lymphocytes when compared with those in normal untreated control mice. Also, spleen weights were significantly decreased in comparison with those of untreated animals 4 days after Cy treatment. Adoptive transfer of untreated NWN spleen cells into Cy-depressed mice restored the NK cell activity which correlated with enhanced clearance of cryptococci from lungs, livers, and spleens. In contrast, treatment of NWN spleen cells with anti-asialo GM1 and complement before adoptive transfer abrogated the ability of these cells to restore NK activity or reduce the numbers of cryptococci present in tissues of infected mice. Taken together, these data indicate that NK cells are the cells effective

  8. Evidence for Natural Killer Cell Memory

    PubMed Central

    Marcus, Assaf; Raulet, David H.

    2013-01-01

    Natural killer (NK) cells are generally considered part of the innate immune system. However, over the past few years, evidence has accumulated suggesting that NK cells have certain features characteristic of the adaptive immune system. NK cells reportedly respond in an antigen-specific manner to a variety of small molecules and certain viruses, and mediate enhanced responses to these antigens upon re-challenge. In infections with mouse cytomegalovirus (MCMV), MCMV-specific NK cells undergo clonal expansion, and display increased effector function after the resolution of the infection. In addition, inflammatory conditions resulting from exposure to certain cytokines seem to cause prolonged effector function in NK cells in an antigen non-specific fashion. Taken together, these studies reveal new aspects of NK biology, and suggest that NK cells, like T and B cells, may carry out memory responses, and may also exhibit greater capacity to distinguish antigens than was previously recognized. PMID:24028966

  9. New Indole Tubulin Assembly Inhibitors Cause Stable Arrest of Mitotic Progression, Enhanced Stimulation of Natural Killer Cell Cytotoxic Activity, and Repression of Hedgehog-Dependent Cancer

    PubMed Central

    La Regina, Giuseppe; Bai, Ruoli; Coluccia, Antonio; Famiglini, Valeria; Pelliccia, Sveva; Passacantilli, Sara; Mazzoccoli, Carmela; Ruggieri, Vitalba; Verrico, Annalisa; Miele, Andrea; Monti, Ludovica; Nalli, Marianna; Alfonsi, Romina; Di Marcotullio, Lucia; Gulino, Alberto; Ricci, Biancamaria; Soriani, Alessandra; Santoni, Angela; Caraglia, Michele; Porto, Stefania; Pozzo, Eleonora Da; Martini, Claudia; Brancale, Andrea; Marinelli, Luciana; Novellino, Ettore; Vultaggio, Stefania; Varasi, Mario; Mercurio, Ciro; Bigogno, Chiara; Dondio, Giulio; Hamel, Ernest; Lavia, Patrizia; Silvestri, Romano

    2015-01-01

    We designed 39 new 2-phenylindole derivatives as potential anticancer agents bearing the 3,4,5-trimethox-yphenyl moiety with a sulfur, ketone, or methylene bridging group at position 3 of the indole and with halogen or methoxy substituent(s) at positions 4–7. Compounds 33 and 44 strongly inhibited the growth of the P-glycoprotein-overexpressing multi-drug-resistant cell lines NCI/ADR-RES and Messa/Dx5. At 10 nM, 33 and 44 stimulated the cytotoxic activity of NK cells. At 20–50 nM, 33 and 44 arrested >80% of HeLa cells in the G2/M phase of the cell cycle, with stable arrest of mitotic progression. Cell cycle arrest was followed by cell death. Indoles 33, 44, and 81 showed strong inhibition of the SAG-induced Hedgehog signaling activation in NIH3T3 Shh-Light II cells with IC50 values of 19, 72, and 38 nM, respectively. Compounds of this class potently inhibited tubulin polymerization and cancer cell growth, including stimulation of natural killer cell cytotoxic activity and repression of Hedgehog-dependent cancer. PMID:26132075

  10. [Passive smoking--active killer].

    PubMed

    Palavra, Irena Rojnić; Franelić, Iva Pejnović; Milanović, Sanja Musić; Puljić, Kresimir

    2013-01-01

    Although still not perceived in this way, passive smoking is a public health issue of great importance. World Health Organization estimates that as a result of passive exposure to tobacco smoke each year 600,000 people die, of which 165,000 children. There are 33% of men, 35% of women and 40% of children who do not smoke, but are exposed to second hand smoke, and still only 11% of the world population is protected by adequate smoke-free legislation. Scientific literature provides evidence that passive exposure to tobacco smoke can result in numerous adverse health effects: asthma and allergies, respiratory infections and (middle) ear infections, cancers of various localization, accelerated atherosclerosis and cardiovascular diseases, retardation of growth and development in children, and in pregnancy it can lead to congenital anomalies and premature birth as well as lower body weight and length of the child. Certainly, the scariest consequence of all is sudden infant death syndrome, also called "death in the crib". Smoke-free policies have proven their effectiveness, but while implementing the laws, it is necessary to raise public awareness of the hazards of, both active and passive, exposure to tobacco smoke. PMID:24490334

  11. [Additive effect of marihuana and retrovirus in the anergy of natural killer cells in mice].

    PubMed

    Ongrádi, J; Specter, S; Horváth, A; Friedman, H

    1999-01-10

    Among the immunosuppressive effects of marijuana, impairment of natural killer cell activity is significant. HIV also inhibits these cells. Friend leukemia virus complex and its helper component Rowson-Parr virus induce early immunosuppression in mice resembling human AIDS, and late leukemia, providing a small animal AIDS model. Leukemia susceptible BALB/c and resistant C57BL/6 mice were infected with these viruses. At different time points, their natural killer cells separated from spleens were treated with 0 to 10 micrograms/ml tetrahydrocannabinol, subsequently mixed with Yac-1 target cells for 4 and 18 h. The natural killer cell activity in both mouse strains infected by either virus complex or helper virus weakened on days 2 to 4 postinfection, normalized by day 8 and enhanced on days 11 to 14. Natural killer cell activity upon the effect of low concentration (1.0 to 2.5 micrograms/ml) of tetrahydrocannabinol slightly increased in BALB/c, was unaffected in C57BL/6, especially in 18 h assays. In the combined effects of marijuana and retrovirus, damages by marijuana dominated over those of retroviruses. Inhibition or reactive enhancement of natural killer cell activity on the effect of viruses are similar to those of infected but marijuana-free counterparts, but on the level of uninfected cells treated with marijuana. The effects of marijuana and retrovirus are additive resulting in anergy of natural killer cells.

  12. A Novel Saccharomyces cerevisiae Killer Strain Secreting the X Factor Related to Killer Activity and Inhibition of S. cerevisiae K1, K2 and K28 Killer Toxins.

    PubMed

    Melvydas, Vytautas; Bružauskaitė, Ieva; Gedminienė, Genovaitė; Šiekštelė, Rimantas

    2016-09-01

    It was determined that Kx strains secrete an X factor which can inhibit all known Saccharomyces cerevisiae killer toxins (K1, K2, K28) and some toxins of other yeast species-the phenomenon not yet described in the scientific literature. It was shown that Kx type yeast strains posess a killer phenotype producing small but clear lysis zones not only on the sensitive strain α'1 but also on the lawn of S. cerevisiae K1, K2 and K28 type killer strains at temperatures between 20 and 30 °C. The pH at which killer/antikiller effect of Kx strain reaches its maximum is about 5.0-5.2. The Kx yeast were identified as to belong to S. cerevisiae species. Another newly identified S. cerevisiae killer strain N1 has killer activity but shows no antikilller properties against standard K1, K2 and K28 killer toxins. The genetic basis for Kx killer/antikiller phenotype was associated with the presence of M-dsRNA which is bigger than M-dsRNA of standard S. cerevisiae K1, K2, K28 type killer strains. Killer and antikiller features should be encoded by dsRNA. The phenomenon of antikiller (inhibition) properties was observed against some killer toxins of other yeast species. The molecular weight of newly identified killer toxins which produces Kx type strains might be about 45 kDa.

  13. Killer toxin from a novel killer yeast Pichia kudriavzevii RY55 with idiosyncratic antibacterial activity.

    PubMed

    Bajaj, Bijender Kumar; Raina, Sandeepu; Singh, Satbir

    2013-08-01

    The killer phenomenon of yeast may have technological implications in many areas like beverage fermentation, food technology, biological control in agriculture, and in medicine. In the present study the killer phenomenon in Pichia kudriavzevii (P. kudriavzevii RY55) is being reported for the first time. The P. kudriavzevii RY55 toxin exhibited excellent antibacterial activity against several pathogens of human health significance such as Escherichia coli, Enterococcus faecalis, Klebsiella sp., Staphylococcus aureus, Pseudomonas aeruginosa and Pseudomonas alcaligenes. Killer toxin was purified to homogeneity by using ammonium sulphate precipitation and ion exchange chromatography and characterized for few properties. P. kudriavzevii RY55 killer toxin may be of vast significance in the development of novel antimicrobial chemotherapeutic agents, new bio-based safer candidates for food preservation and biocontrol, and starter cultures for fermentation industries. PMID:22961241

  14. Killer toxin from a novel killer yeast Pichia kudriavzevii RY55 with idiosyncratic antibacterial activity.

    PubMed

    Bajaj, Bijender Kumar; Raina, Sandeepu; Singh, Satbir

    2013-08-01

    The killer phenomenon of yeast may have technological implications in many areas like beverage fermentation, food technology, biological control in agriculture, and in medicine. In the present study the killer phenomenon in Pichia kudriavzevii (P. kudriavzevii RY55) is being reported for the first time. The P. kudriavzevii RY55 toxin exhibited excellent antibacterial activity against several pathogens of human health significance such as Escherichia coli, Enterococcus faecalis, Klebsiella sp., Staphylococcus aureus, Pseudomonas aeruginosa and Pseudomonas alcaligenes. Killer toxin was purified to homogeneity by using ammonium sulphate precipitation and ion exchange chromatography and characterized for few properties. P. kudriavzevii RY55 killer toxin may be of vast significance in the development of novel antimicrobial chemotherapeutic agents, new bio-based safer candidates for food preservation and biocontrol, and starter cultures for fermentation industries.

  15. Crude extract of Polygonum cuspidatum promotes immune responses in leukemic mice through enhancing phagocytosis of macrophage and natural killer cell activities in vivo.

    PubMed

    Chueh, Fu-Shin; Lin, Jen-Jyh; Lin, Jing-Pin; Yu, Fu-Shun; Lin, Ju-Hwa; Ma, Yi-Shih; Huang, Yi-Ping; Lien, Jin-Cherng; Chung, Jing-Gung

    2015-01-01

    Polygonum cuspidatum is a traditional Chinese herbal medicine used in the treatment of various diseases. In the present study, we investigated whether the crude extract of Polygonum cuspidatum (CEPC) could affect immune responses of murine leukemia cells in vivo. Normal BALB/c mice were i.p. injected with WEHI-3 cells to generate leukemic mice and then were treated orally with CEPC at 0, 50, 100 and 200 mg/kg for three weeks. Animals were weighed and blood, liver, spleen samples were collected for further analyses. Results indicated that CEPC did not significantly affect the body and liver weight of animals, but reduced the weight of spleen when compared to control groups. Flow cytometric assay demonstrated that CEPC increased the percentage of CD3- (T-cell marker) and CD19- (B-cell marker) positive cells, but reduced that of CD11b-positive ones (monocytes). However, it did not significantly affect the proportion of Mac-3-positive cells (macrophages), compared to control groups. Results indicated that CEPC promoted phagocytosis by macrophages from blood samples at all examined doses but did not affect that of macrophages from the peritoneal cavity. CEPC also promoted natural killer cell activity of splenocytes at 200 mg/kg of CEPC. CEPC promoted B-cell proliferation at 200 mg/kg treatment when cells were stimulated with lipopolysaccharides but did not promote T-cell proliferation at three doses of CEPC treatment on concanavalin A stimulation. PMID:25792654

  16. Low-dose ionizing radiation induces direct activation of natural killer cells and provides a novel approach for adoptive cellular immunotherapy.

    PubMed

    Yang, Guozi; Kong, Qingyu; Wang, Guanjun; Jin, Haofan; Zhou, Lei; Yu, Dehai; Niu, Chao; Han, Wei; Li, Wei; Cui, Jiuwei

    2014-12-01

    Recent evidence indicates that limited availability and cytotoxicity have restricted the development of natural killer (NK) cells in adoptive cellular immunotherapy (ACI). While it has been reported that low-dose ionizing radiation (LDIR) could enhance the immune response in animal studies, the influence of LDIR at the cellular level has been less well defined. In this study, the authors aim to investigate the direct effects of LDIR on NK cells and the potential mechanism, and explore the application of activation and expansion of NK cells by LDIR in ACI. The authors found that expansion and cytotoxicity of NK cells were markedly augmented by LDIR. The levels of IFN-γ and TNF-α in the supernatants of cultured NK cells were significantly increased after LDIR. Additionally, the effect of the P38 inhibitor (SB203580) significantly decreased the expanded NK cell cytotoxicity, cytokine levels, and expression levels of FasL and perforin. These findings indicate that LDIR induces a direct expansion and activation of NK cells through possibly the P38-MAPK pathway, which provides a potential mechanism for stimulation of NK cells by LDIR and a novel but simplified approach for ACI.

  17. Effects of dendritic cell-activated and cytokine-induced killer cell therapy on 22 children with acute myeloid leukemia after chemotherapy.

    PubMed

    Bai, Yan; Zheng, Jin-e; Wang, Nan; Cai, He-hua; Zhai, Li-na; Wu, Yao-hui; Wang, Fang; Jin, Run-ming; Zhou, Dong-feng

    2015-10-01

    The efficiency of dendritic cell-activated and cytokine-induced killer cell (DC-CIK) therapy on children with acute myeloid leukemia (AML) after chemotherapy was investigated. Mononuclear cells were collected from children achieving complete remission after chemotherapy, cultured in vitro and transfused back into the same patient. Interleukin-2 (IL-2) was injected subcutaneously every other day 10 times at the dose of 1 × 10(6) units. Peripheral blood lymphocyte subsets and minimal residual disease (MRD) were detected by flow cytometry. Function of bone marrow was monitored by methods of morphology, immunology, cytogenetics and molecular biology. The side effects were also observed during the treatment. The average follow-up period for all the 22 patients was 71 months and relapse occurred in two AML patients (9.1%). The percentage of CD3(+)/CD8(+) cells in peripheral blood of 15 patients at the 3rd month after DC-CIK treatment (36.73% ± 12.51%) was dramatically higher than that before treatment (29.20% ± 8.34%, P < 0.05). The MRD rate was >0.1% in 5 patients before the treatment, and became lower than 0.1% 3 months after the treatment. During the transfusion of DC-CIK, side effects including fever, chills and hives appeared in 7 out of 22 (31.82%) cases but disappeared quickly after symptomatic treatments. There were no changes in electrocardiography and liver-renal functions after the treatment. MRD in children with AML can be eliminated by DC-CIK therapy which is safe and has fewer side effects.

  18. Effects of dendritic cell-activated and cytokine-induced killer cell therapy on 22 children with acute myeloid leukemia after chemotherapy.

    PubMed

    Bai, Yan; Zheng, Jin-e; Wang, Nan; Cai, He-hua; Zhai, Li-na; Wu, Yao-hui; Wang, Fang; Jin, Run-ming; Zhou, Dong-feng

    2015-10-01

    The efficiency of dendritic cell-activated and cytokine-induced killer cell (DC-CIK) therapy on children with acute myeloid leukemia (AML) after chemotherapy was investigated. Mononuclear cells were collected from children achieving complete remission after chemotherapy, cultured in vitro and transfused back into the same patient. Interleukin-2 (IL-2) was injected subcutaneously every other day 10 times at the dose of 1 × 10(6) units. Peripheral blood lymphocyte subsets and minimal residual disease (MRD) were detected by flow cytometry. Function of bone marrow was monitored by methods of morphology, immunology, cytogenetics and molecular biology. The side effects were also observed during the treatment. The average follow-up period for all the 22 patients was 71 months and relapse occurred in two AML patients (9.1%). The percentage of CD3(+)/CD8(+) cells in peripheral blood of 15 patients at the 3rd month after DC-CIK treatment (36.73% ± 12.51%) was dramatically higher than that before treatment (29.20% ± 8.34%, P < 0.05). The MRD rate was >0.1% in 5 patients before the treatment, and became lower than 0.1% 3 months after the treatment. During the transfusion of DC-CIK, side effects including fever, chills and hives appeared in 7 out of 22 (31.82%) cases but disappeared quickly after symptomatic treatments. There were no changes in electrocardiography and liver-renal functions after the treatment. MRD in children with AML can be eliminated by DC-CIK therapy which is safe and has fewer side effects. PMID:26489623

  19. Endocytosis and Intracellular Trafficking of Human Natural Killer Cell Receptors

    PubMed Central

    Masilamani, Madhan; Peruzzi, Giovanna; Borrego, Francisco; Coligan, John E.

    2009-01-01

    Natural killer (NK) cells play a vital role in the defense against viral infections and tumor development. NK cell function is primarily regulated by the sum of signals from a broad array of activation and inhibitory receptors. Key to generating the input level of either activating or inhibitory signals is the maintenance of receptor expression levels on the cell surface. Although the mechanisms of endocytosis and trafficking for some cell surface receptors, such as transferrin receptor, and certain immune receptors, are very well known, that is not the situation for receptors expressed by NK cells. Recent studies have uncovered that endocytosis and trafficking routes characteristic for specific activation and inhibitory receptors can regulate the functional responses of NK cells. In this review, we summarize the current knowledge of receptor endocytosis and trafficking, and integrate this with our current understanding of NK cell receptor trafficking. PMID:19719476

  20. Ozone exposed epithelial cells modify cocultured natural killer cells

    PubMed Central

    Müller, Loretta; Brighton, Luisa E.

    2013-01-01

    Ozone (O3) causes significant adverse health effects worldwide. Nasal epithelial cells (NECs) are among the first sites within the respiratory system to be exposed to inhaled air pollutants. They recruit, activate, and interact with immune cells via soluble mediators and direct cell-cell contacts. Based on our recent observation demonstrating the presence of natural killer (NK) cells in nasal lavages, the goal of this study was to establish a coculture model of NECs and NK cells and examine how exposure to O3 modifies this interaction. Flow cytometry analysis was used to assess immunophenotypes of NK cells cocultured with either air- or O3-exposed NECs. Our data show that coculturing NK cells with O3-exposed NECs decreased intracellular interferon-γ (IFN-γ), enhanced, albeit not statistically significant, IL-4, and increased CD16 expression on NK cells compared with air controls. Additionally, the cytotoxicity potential of NK cells was reduced after coculturing with O3-exposed NECs. To determine whether soluble mediators released by O3-exposed NECs caused this shift, apical and basolateral supernatants of air- and O3-exposed NECs were used to stimulate NK cells. While the conditioned media of O3-exposed NECs alone did not reduce intracellular IFN-γ, O3 enhanced the expression of NK cell ligands ULBP3 and MICA/B on NECs. Blocking ULBP3 and MICA/B reversed the effects of O3-exposed NECs on IFN-γ production in NK cells. Taken together, these data showed that interactions between NECs and NK cells in the context of O3 exposure changes NK cell activity via direct cell-cell interactions and is dependent on ULBP3/MICA/B expressed on NECs. PMID:23241529

  1. In vitro induction of lymphokine-activated killer (LAK) activity in patients with neuroblastoma.

    PubMed

    Handgretinger, R; Bruchelt, G; Kimmig, A; Dopfer, R; Niethammer, D; Treuner, J

    1989-01-01

    Therapy of disseminated neuroblastoma remains an unsolved problem in pediatric oncology. Therefore, new therapeutic approaches have to be developed for this malignancy. In this paper, we investigated the possibility of the in vitro generation and expansion of lymphokine-activated killer (LAK) cells in patients with disseminated neuroblastoma. Although the patients had very low Natural Killer (NK) activity, it was possible to induce LAK activity in peripheral mononuclear lymphocytes (PMNC) by incubation with Interleukin-2 (IL-2). Moreover, the PMNCs could be expanded up to 50-fold in the presence of Interleukin-2 while maintaining or even increasing their LAK activity. The target cells were neuroblastoma cell lines and, in one case, autologous neuroblastoma cells. Additionally, it was possible to induce LAK cell activity against autologous neuroblastoma cells in bone marrow-derived mononuclear cells.

  2. Immunosuppression of pulmonary natural killer activity by exposure to ozone

    SciTech Connect

    Burleson, G.R.; Keyes, L.L.; Stutzman, J.D. )

    1989-01-01

    Ozone is an oxidant gas and an ubiquitous oxidant air pollutant with the potential to adversely affect pulmonary immune function with a consequent increase in disease susceptibility. Pulmonary natural killer (NK) activity was measured in order to assess the pulmonary immunotoxicity of continuous ozone exposure. Continuous ozone exposures at 1.0 ppm were performed for 23.5 hours per day for either 1, 5, 7, or 10 consecutive days. Pulmonary immune function was assessed by measuring natural killer (NK) activity from whole-lung homogenates of male Fischer-344 rats. Results of this study indicated that continuous ozone exposure for 1, 5, or 7 days resulted in a significant decrease in pulmonary NK activity. This suppressed pulmonary NK activity returned to control levels after continuous exposure to ozone for 10 days. The suppressed pulmonary NK response was thus attenuated and returned to normal values in the continued presence of ozone gas. This attenuation process is dynamic, complex, and doubtless involves several cell types and/or products of these cells. Pulmonary NK activity was also suppressed at 0.5 ppm ozone, but not at 0.1 ppm ozone, following 23.5 hours of exposure. NK activity is important for defense against viral, bacterial, and neoplastic disease. The depressed NK activity resulting from continuous ozone exposure could therefore result in a compromised ability to defend against pulmonary diseases.

  3. Activation of p44/42 MAPK Plays a Role in the TBT-induced Loss of Human Natural Killer (NK) Cell Function

    PubMed Central

    Dudimah, Fred D.; Griffey, Denisha; Wang, Xiaofei; Whalen, Margaret M.

    2009-01-01

    Natural Killer (NK) cells destroy (lyse) tumor cells, virally infected cells and antibody-coated cells. Previous studies indicated that exposure to the environmental contaminant tributyltin (TBT) decreases the lytic function of NK cells and activates mitogen activated protein kinases (MAPK), including p44/42 (Aluoch and Whalen, 2005). If activation of p44/42 is required for TBT-induced decreases of lytic function, then activation of p44/42 to similar extents by pharmacological agents such as Phorbol 12-myristate 13-acetate (PMA) should mimic to some extent changes induced in NK cells with TBT exposures. NK cells were exposed to PMA concentrations between 0.25 and 10 nM for 10 min, 1 h, and 6 h before determining the lytic function (51Cr release assay) and phosphorylation state of MAPKs (Western blot). A 1 h exposure of NK cells to 5 nM PMA resulted in a loss of lytic function of 47%. Western blot analysis showed that a 1 h exposure to 5 nM PMA caused a 6 fold increase in phospho-p44/42 levels. Previous studies showed a 5 fold increase in phospho-p44/42 in response to a 1 h exposure to 300 nM TBT. Exposure to 300 nM TBT caused about a 40% decrease in lytic function. This study supports the hypothesis that p44/42 activation (as seen with TBT exposures) can cause a loss of NK-cell lytic function. PMID:20213532

  4. A One Year Follow-Up Study of Natural Killer and Dendritic Cells Activities in Multiple Sclerosis Patients Receiving Glatiramer Acetate (GA)

    PubMed Central

    Høglund, Rune A.; Holmøy, Trygve; Harbo, Hanne F.; Maghazachi, Azzam A.

    2013-01-01

    Background Multiple sclerosis (MS) is a chronic inflammatory, demyelinating and neurodegenerative disease. It is thought to be mediated by CD4+ Th1/Th17 cells. More recently, cells of the innate immune system such as dendritic cells (DCs) and natural killer (NK) cells have been in focus. Glatiramer acetate (GA) is an approved drug for treating MS patients. Methodology/Principal Findings In the current study we examined the activities of NK and DCs in nine relapsing remitting MS patients for up to one year after initiation of GA treatment. We observed that NK cells isolated from most of these patients have increased cytotoxic activity against K562 cells. Further analysis showed that the same NK cells lysed both autologous immature (i) and mature (m) DCs. In most patients this increased activity was correlated with increased NK cell activating cytotoxicity receptors such as NKp30, NKp44, NKp46 and NKG2D, and reduced expression of the inhibitory molecule CD158 on the surface of these NK cells. The expression of HLA-DR was increased on iDCs and mDCs in the majority of the patients, but no consistency was observed for the expression of HLA-I or HLA-E. Also, the co-stimulatory receptors CD80, CD83 or CD86 expression was down-regulated on iDCs and mDCs in most cases. Further, the expression of CCR6 was increased on mDCs at later time points of therapy (between 32–48 weeks). Conclusions/Significance Our results are the first showing the effects of GA treatment on NK cells in MS patients, which may impact future use of this and other drugs to treat this disease. PMID:23614042

  5. Lack of correlation between mycoplasma induced IFN-gamma production in vitro and natural killer cell activity against FLD-3 cells

    NASA Technical Reports Server (NTRS)

    Kumar, V.; Lust, J.; Gifaldi, A.; Bennett, M.; Sonnenfeld, G.

    1983-01-01

    The role of interferon (IFN) in the normal-killer-cell (NK) mediated lysis of tumor cells in vitro is investigated experimentally. Normal mouse spleen cells and spleen cells treated with anti-Thy-1.2 serum are cultured for 24 h with Friend erythroleukemia (FLD-3) cells in RPMI 1640 medium; supernatant fluid from cultures with FLD-3 lysis are assayed for IFN-gamma, and it is found that pretreatment with anti-Thy-1.2 suppresses IFN-gamma generation without affecting the ability of NK to mediate the lysis of FLD-3. Further tests indicate that the generation of IFN-gamma is stimulated by the presence of Mycoplasma arginini in the FLD-3 cells.

  6. Molecular cloning of L-JAK, a Janus family protein-tyrosine kinase expressed in natural killer cells and activated leukocytes.

    PubMed Central

    Kawamura, M; McVicar, D W; Johnston, J A; Blake, T B; Chen, Y Q; Lal, B K; Lloyd, A R; Kelvin, D J; Staples, J E; Ortaldo, J R

    1994-01-01

    Protein-tyrosine kinases (PTKs) are critical enzymes for receptor-mediated signaling in lymphocytes. Because natural killer (NK) cells are large granular lymphocytes with specialized effector function, we set out to identify PTKs preferentially expressed in these cells. One such PTK was identified and molecularly cloned. The predicted amino acid sequence shows that this kinase lacks SH2 or SH3 domains typical of src family kinases but has tandem nonidentical catalytic domains, indicating that it is a member of the Janus family of PTKs. Immunoprecipitation using antiserum generated against a peptide corresponding to the deduced amino acid sequence of this gene revealed a kinase with a molecular weight of approximately 125,000. The pattern of expression of this kinase contrasted sharply with that of other Janus kinases, which are ubiquitously expressed. The kinase described in the present study was found to be more limited in its expression; expression was found in NK cells and an NK-like cell line but not in resting T cells or in other tissues. In contrast, stimulated and transformed T cells expressed the gene, suggesting a role in lymphoid activation. Because of its homology and tissue expression, we have tentatively termed this PTK gene L-JAK for leukocyte Janus kinase. Images PMID:8022790

  7. Expression of conformationally constrained adhesion peptide in an antibody CDR loop and inhibition of natural killer cell cytotoxic activity by an antibody antigenized with the RGD motif.

    PubMed Central

    Zanetti, M; Filaci, G; Lee, R H; del Guercio, P; Rossi, F; Bacchetta, R; Stevenson, F; Barnaba, V; Billetta, R

    1993-01-01

    We report that an antibody engineered to express three Arg-Gly-Asp (RGD) repeats in the third complementarity-determining region of the heavy chain (antigenized antibody) efficiently inhibits the lysis of human erythroleukemia K-562 cells by natural killer (NK) cells. Synthetic peptides containing RGD did not inhibit. Inhibition was specific for the (RGD)3-containing loop and required simultaneous occupancy of the Fc receptor (CD16) on effector cells. The antigenized antibody inhibited other forms of cytotoxicity mediated by NK cells but not cytotoxicity mediated by major histocompatibility complex-restricted cytotoxic T lymphocytes (CTL). A three-dimensional model of the engineered antibody loop shows the structure and physicochemical characteristics probably required for the ligand activity. The results indicate that an RGD motif is involved in the productive interaction between NK and target cells. Moreover, they show that peptide expression in the hypervariable loops of an antibody molecule is an efficient procedure for stabilizing oligopeptides within a limited spectrum of tertiary structures. This is a new approach towards imparting ligand properties to antibody molecules and can be used to study the biological function and specificity of short peptide motifs, including those involved in cell adhesion. Images PMID:8223447

  8. Analysis of GzmbCre as a Model System for Gene Deletion in the Natural Killer Cell Lineage.

    PubMed

    Xu, Yiying; Evaristo, Cesar; Alegre, Maria-Luisa; Gurbuxani, Sandeep; Kee, Barbara L

    2015-01-01

    The analysis of gene function in mature and activated natural killer cells has been hampered by the lack of model systems for Cre-mediated recombination in these cells. Here we have investigated the utility of GzmbCre for recombination of loxp sequences in these cells predicated on the observation that Gzmb mRNA is highly expressed in mature and activated natural killer cells. Using two different reporter strains we determined that gene function could be investigated in mature natural killer cells after GzmbCre mediated recombination in vitro in conditions that lead to natural killer cell activation such as in the cytokine combination of interleukin 2 and interleukin 12. We demonstrated the utility of this model by creating GzmbCre;Rosa26IKKbca mice in which Cre-mediated recombination resulted in expression of constitutively active IKKβ, which results in activation of the NFκB transcription factor. In vivo and in vitro activation of IKKβ in natural killer cells revealed that constitutive activation of this pathway leads to natural killer cell hyper-activation and altered morphology. As a caveat to the use of GzmbCre we found that this transgene can lead to recombination in all hematopoietic cells the extent of which varies with the particular loxp flanked allele under investigation. We conclude that GzmbCre can be used under some conditions to investigate gene function in mature and activated natural killer cells.

  9. Orally Administered Salacia reticulata Extract Reduces H1N1 Influenza Clinical Symptoms in Murine Lung Tissues Putatively Due to Enhanced Natural Killer Cell Activity

    PubMed Central

    Romero-Pérez, Gustavo A.; Egashira, Masayo; Harada, Yuri; Tsuruta, Takeshi; Oda, Yuriko; Ueda, Fumitaka; Tsukahara, Takamitsu; Tsukamoto, Yasuhiro; Inoue, Ryo

    2016-01-01

    Influenza is a major cause of respiratory tract infection. Although most cases do not require further hospitalization, influenza periodically causes epidemics in humans that can potentially infect and kill millions of people. To countermeasure this threat, new vaccines need to be developed annually to match emerging influenza viral strains with increased resistance to existing vaccines. Thus, there is a need for finding and developing new anti-influenza viral agents as alternatives to current treatments. Here, we tested the antiviral effects of an extract from the stems and roots of Salacia reticulata (SSRE), a plant rich in phytochemicals, such as salacinol, kotalanol, and catechins, on H1N1 influenza virus-infected mice. Following oral administration of 0.6 mg/day of SSRE, the incidence of coughing decreased in 80% of mice, and only one case of severe pulmonary inflammation was detected. Moreover, when compared with mice given Lactobacillus casei JCM1134, a strain previously shown to help increase in vitro natural killer (NK) cell activity, SSRE-administered mice showed greater and equal NK cell activity in splenocytes and pulmonary cells, respectively, at high effector cell:target cell ratios. Next, to test whether or not SSRE would exert protective effects against influenza in the absence of gut microbiota, mice were given antibiotics before being inoculated influenza virus and subsequently administered SSRE. SSRE administration induced an increase in NK cell activity in splenocytes and pulmonary cells at levels similar to those detected in mice not treated with antibiotics. Based on our results, it can be concluded that phytochemicals in the SSRE exerted protective effects against influenza infection putatively via modulation of the immune response, including enhancement of NK cell activity, although some protective effects were not necessarily through modulation of gut microbiota. Further investigation is necessary to elucidate the molecular mechanisms

  10. Orally Administered Salacia reticulata Extract Reduces H1N1 Influenza Clinical Symptoms in Murine Lung Tissues Putatively Due to Enhanced Natural Killer Cell Activity.

    PubMed

    Romero-Pérez, Gustavo A; Egashira, Masayo; Harada, Yuri; Tsuruta, Takeshi; Oda, Yuriko; Ueda, Fumitaka; Tsukahara, Takamitsu; Tsukamoto, Yasuhiro; Inoue, Ryo

    2016-01-01

    Influenza is a major cause of respiratory tract infection. Although most cases do not require further hospitalization, influenza periodically causes epidemics in humans that can potentially infect and kill millions of people. To countermeasure this threat, new vaccines need to be developed annually to match emerging influenza viral strains with increased resistance to existing vaccines. Thus, there is a need for finding and developing new anti-influenza viral agents as alternatives to current treatments. Here, we tested the antiviral effects of an extract from the stems and roots of Salacia reticulata (SSRE), a plant rich in phytochemicals, such as salacinol, kotalanol, and catechins, on H1N1 influenza virus-infected mice. Following oral administration of 0.6 mg/day of SSRE, the incidence of coughing decreased in 80% of mice, and only one case of severe pulmonary inflammation was detected. Moreover, when compared with mice given Lactobacillus casei JCM1134, a strain previously shown to help increase in vitro natural killer (NK) cell activity, SSRE-administered mice showed greater and equal NK cell activity in splenocytes and pulmonary cells, respectively, at high effector cell:target cell ratios. Next, to test whether or not SSRE would exert protective effects against influenza in the absence of gut microbiota, mice were given antibiotics before being inoculated influenza virus and subsequently administered SSRE. SSRE administration induced an increase in NK cell activity in splenocytes and pulmonary cells at levels similar to those detected in mice not treated with antibiotics. Based on our results, it can be concluded that phytochemicals in the SSRE exerted protective effects against influenza infection putatively via modulation of the immune response, including enhancement of NK cell activity, although some protective effects were not necessarily through modulation of gut microbiota. Further investigation is necessary to elucidate the molecular mechanisms

  11. Analysis of the effect of a sunscreen agent on the suppression of natural killer cell activity induced in human subjects by radiation from solarium lamps

    SciTech Connect

    Hersey, P.; MacDonald, M.; Burns, C.; Schibeci, S.; Matthews, H.; Wilkinson, F.J.

    1987-03-01

    Previous studies in rodents have shown that ultraviolet radiation (UVR) may have direct effects on the immune system in the skin and at higher doses may induce systemic suppression of immune responses. We have previously shown that UVR from sun or solarium beds may induce systemic effects in human subjects. The purpose of the present study was to examine whether these systemic effects in human subjects could be prevented by use of commercially available sunscreen agents. Groups of 12 normal subjects were exposed to radiation from solarium lamps after application of a sunscreen agent or the base used in its preparation. Twelve half-hourly exposures induced a depression of natural killer (NK) cell activity against a melanoma and the K562 target cell which was not prevented by use of the sunscreen agent. Changes in functional activity were accompanied by a reduction in NK cell numbers assessed by Leu-11 monoclonal antibodies against the labile Fc receptor. Application of the sunscreen agent also did not protect against effects of solarium exposure on recall antigen skin tests and immunoglobulin production in vitro in pokeweed mitogen-stimulated cultures of B and T cells. These results suggest that further evaluation of the wave-length spectrum of UVR and the effectiveness of sunscreen agents in prevention of UVR-induced effects on the immune system is needed.

  12. A novel bispecific protein (ULBP2-BB4) targeting the NKG2D receptor on natural killer (NK) cells and CD138 activates NK cells and has potent antitumor activity against human multiple myeloma in vitro and in vivo.

    PubMed

    von Strandmann, Elke Pogge; Hansen, Hinrich P; Reiners, Katrin S; Schnell, Roland; Borchmann, Peter; Merkert, Sabine; Simhadri, Venkateswara R; Draube, Andreas; Reiser, Marcel; Purr, Ingvill; Hallek, Michael; Engert, Andreas

    2006-03-01

    The inability of the immune system to recognize and kill malignant plasma cells in patients with multiple myeloma (MM) has been attributed in part to the ineffective activation of natural killer (NK) cells. In order to activate and target NK cells to the malignant cells in MM we designed a novel recombinant bispecific protein (ULBP2-BB4). While ULBP2 binds the activating NK receptor NKG2D, the BB4 moiety binds to CD138, which is overexpressed on a variety of malignancies, including MM. ULBP2-BB4 strongly activated primary NK cells as demonstrated by a significant increase in interferon-gamma (IFN-gamma) secretion. In vitro, ULBP2-BB4 enhanced the NK-mediated lysis of 2 CD138+ human MM cell lines, U-266 and RPMI-8226, and of primary malignant plasma cells in the allogenic and autologous setting. Moreover, in a nude mouse model with subcutaneously growing RPMI-8226 cells, the cotherapy with ULBP-BB4 and human peripheral blood lymphocytes abrogated the tumor growth. These data suggest potential clinical use of this novel construct in patients with MM. The use of recombinant NK receptor ligands that target NK cells to tumor cells might offer new approaches for other malignancies provided a tumor antigen-specific antibody is available.

  13. Conditioned tolerance to drug-induced (poly I:C) natural killer cell activation: effects of drug-dosage and context-specificity parameters.

    PubMed

    Dyck, D G; Driedger, S M; Nemeth, R; Osachuk, T A; Greenberg, A H

    1987-09-01

    Three experiments were conducted to evaluate the role of drug-dosage and stimulus-specificity parameters on the tolerance of drug-induced (poly I:C) natural killer (NK) cell activity. In the first experiment a protocol which provided mice with four weekly 20 micrograms/mouse ip injections of the immunostimulatory synthetic polynucleotide (poly I:C) following exposure to either a simple odor cue or a complex cue resulted in tolerance of NK cell activity. The identical protocol with a higher drug dose (50 micrograms/mouse) did not produce tolerance. In a second experiment, the stimulus specificity of tolerance was assessed by giving two groups of mice repeated signaled drug injections. For one of these groups the final poly I:C injection of the series was signaled, while for the other group it was not. Although both groups were tolerant relative to controls not previously exposed to the drug, indirect evidence of conditioning was obtained. Specifically, it was found that tolerance among mice receiving the signal on the test was such that they were not different from undrugged controls, while uncued mice had significantly higher levels of NK cell activity. The third experiment evaluated the role of stimulus specificity within an extinction paradigm. It was found that tolerance was reversed in mice provided with repeated nonreinforced reexposure to drug-signaling cues, while mice exposed to novel cues remained tolerant. These results further support the hypothesis that associative factors contribute to the tolerance of a drug-induced immune response.

  14. Natural killer cell lymphoma of the parotid gland.

    PubMed

    Furukawa, Masayuki; Suzuki, Hideaki; Tohmiya, Yasuo; Matsuura, Kazuto; Takahashi, Etsu; Ichinohasama, Ryo; Kobayashi, Toshimitsu

    2003-01-01

    The majority of all parotid lymphomas are of the non-Hodgkin type and of B-cell origin. Primary natural killer cell lymphomas of the parotid gland are extremely rare. We present a case of natural killer cell lymphoma in a 34-year-old woman. The disease was refractory to chemotherapy, and the patient eventually succumbed due to lymphoma-associated hemophagocytic syndrome. PMID:14564097

  15. Characterisation of the morphogenetic course and secretion of two different types of mucoid material by granulated metrial gland/lymphokine-activated killer cells.

    PubMed

    Dyugovskaya, L; Berkutski, T; Ginsburg, H

    1995-12-01

    Lymphocytes from mesenteric lymph nodes of ordinary and nude mice were grown in microtitre wells on embryonic mesenchymal-fibroblast monolayers. Human recombinant interleukin-2 (80 units ml-1) was added. Clones of lymphokine-activated killer (LAK) cells developed. The incidence of clone-forming cells was 52-136 per 10(5) cells in lymph nodes from nude mice and 4.2-8.3 per 10(5) cells in lymph nodes from ordinary mice. On a limited number of fibroblast monolayers propagated in culture, the maturing LAK cells were induced to synthesise and secrete 2 types of flowing mucoid material. After methanol fixation and Alcian blue/periodic acid-Schiff (PAS) (at pH 1) staining, the first type of material was distinctively stained turquoise, indicating a highly sulphated proteoglycan, chondroitin sulphate; the second type of material, a macromolecular neutral polysaccharide, was not stained and appeared to have been dissolved. Glycogen was stained deep brilliant purple. After treatment with PAS alone, the chondroitin sulphate was not stained and appeared as a bright area, the neutral polysaccharide mass being stained deep red. This polysaccharide material was characteristically secreted as droplets or 'streamlets' emerging from the cell surface and extending through the first type of material to coalesce with the already accumulated main extracellular mucoid layer spreading between the cells. Clones of secretory LAK cells were obtained from gravid and nongravid mouse uteri as well as from tracheal explants. Change of medium or passage with fresh medium to a new inducing batch of monolayer, at the blastoid-large granular lymphocyte stage (on d 3 to 7), was critical for high reproducibility of secretion. The course of differentiation was found ultimately to be dependent on the embryonic mesenchymal monolayer, suggesting induction by a morphogenetic signal. A correlation can be drawn between the secretory activity and the morphological profile at maturation of highly distinctive

  16. CD16-mediated p21ras activation is associated with Shc and p36 tyrosine phosphorylation and their binding with Grb2 in human natural killer cells

    PubMed Central

    1996-01-01

    The Src homology (SH) 2/SH3 domain-containing protein Grb2 and the oncoprotein Shc have been implicated in a highly conserved mechanism that regulates p21ras activation. We investigated the involvement of these adaptor proteins in the signaling pathway induced by CD16 or interleukin (IL) 2R triggering in human natural killer (NK) cells. Both p46 and p52 forms of Shc were rapidly and transiently tyrosine phosphorylated upon CD16 or IL-2 stimulation with different kinetics. Shc immunoprecipitates from lysates of CD16- or IL-2-stimulated NK cells contained Grb2 and an unidentified 145-kD tyrosine phosphoprotein. Grb2 immunoprecipitates from anti-CD16-stimulated NK cells contained not only Shc, but also a 36-kD tyrosine phosphoprotein (p36). The interaction between Grb2 and Shc or p36 occurred via the Grb2SH2 domain as indicated by in vitro binding assays using a bacteriologically synthesized glutathione S-transferase-Grb2SH2 fusion protein. We also present evidence that p21ras is activated by CD16 and IL-2R cross-linking. Accumulation of guanosine triphosphate-bound Ras was detected within 1 minute and occurred with kinetics similar to inductive protein tyrosine phosphorylation and Grb2 association of Shc and p36 adaptor proteins. PMID:8551221

  17. The effects of IL-17 upon human natural killer cells.

    PubMed

    Al Omar, Suliman; Flanagan, Brian F; Almehmadi, Mazen; Christmas, Stephen E

    2013-04-01

    These experiments were designed to investigate the effects of IL-17 upon the phenotype and function of human Natural Killer (NK) cells. Peripheral blood mononuclear cells from healthy subjects were cultured in the presence or absence of different combinations of IL-17s and changes in relative numbers and cell surface phenotype of NK cells and CD56+CD3+ cells measured by flow cytometry. Real time PCR was used to measure changes in expression of the cytotoxicity-related genes perforin A and granzymes A and B and IL-17 receptors. A chromium release assay was used to measure cytotoxic function against K562 tumour cells. IL-17D, IL-17A, IL-17F or the combination of both of the latter had little effect upon NK cell surface expression of Killer Immunoglobulin-like Receptors, although IL-17A modestly increased NK cell numbers. Slight but not significant increases in expression of perforin and granzymes were induced by IL-17A and/or IL-17F. Both IL-17A and D significantly increased cytotoxic function of NK cells at some E:T ratios. Similarly, numbers of NK cells induced to express CD107a after interaction with K562 cells were increased, but not significantly, by all combinations of IL-17s tested. IL-17RC was not found at the NK cell surface but was expressed at the message level and the protein detected intracellularly. NK cells are known to produce IL-17 but here we report that there is little response to this cytokine although some isoforms may moderately enhance cytotoxic function. There may therefore be some enhancement of NK cell function resulting from Th17 cell activation.

  18. Natural killer (NK) cell cytotoxicity in athymic (nude) rats.

    PubMed

    Grzelak, I; Olszewski, W L; Fossum, S; Engeset, A

    1984-01-01

    The in vitro and in vivo natural killer (NK) cell activity of congenitally athymic, nude (ATH) rats and of normal, euthymic (EUTH) rats was compared. We found: a) a higher level of in vitro NK cell activity in blood, spleen and lymph nodes of ATH rats compared with their heterozygous littermates, b) in the spleen the number of NK lytic units per organ was not higher in ATH compared with EUTH whereas it was significantly higher in lymph nodes, c) a lack of age-dependence of in vitro NK cell activity tested in culture with heat inactivated fetal calf serum, d) a higher rate of in vivo elimination of target tumor cells in 4-week ATH rats compared with EUTH rats, e) an age-dependent decrease in the rate of in vivo target cell elimination in both groups, and finally, f) an age-dependent increase in the inhibitory effect of autologous serum on NK cell activity in vitro in both groups. These findings show that the blood and lymphoid organs of athymic rats contain a substantially higher proportion of NK cells, active both in vitro and in vivo against K562 tumor cells, than their euthymic littermates. In the spleen this increased proportion can be attributed to the lack of T cells, whereas in the ATH rat lymph nodes there is an absolute increase in NK cell activity, and that the decrease of cytotoxicity in vivo with age reflects the increasing inhibitory properties of autologous serum both in nude and in normal rats.

  19. Multiplicity and plasticity of natural killer cell signaling pathways

    PubMed Central

    Chiesa, Sabrina; Mingueneau, Michael; Fuseri, Nicolas; Malissen, Bernard; Raulet, David H.; Malissen, Marie; Vivier, Eric; Tomasello, Elena

    2006-01-01

    Natural killer (NK) cells express an array of activating receptors that associate with DAP12 (KARAP), CD3ζ, and/or FcRγ ITAM (immunoreceptor tyrosine-based activation motif)–bearing signaling subunits. In T and mast cells, ITAM-dependent signals are integrated by critical scaffolding elements such as LAT (linker for activation of T cells) and NTAL (non–T-cell activation linker). Using mice that are deficient for ITAM-bearing molecules, LAT or NTAL, we show that NK cell cytotoxicity and interferon-γ secretion are initiated by ITAM-dependent and -independent as well as LAT/NTAL-dependent and -independent pathways. The role of these various signaling circuits depends on the target cell as well as on the activation status of the NK cell. The multiplicity and the plasticity of the pathways that initiate NK cell effector functions contrast with the situation in T cells and B cells and provide an explanation for the resiliency of NK cell effector functions to various pharmacologic inhibitors and genetic mutations in signaling molecules. PMID:16291591

  20. Determination of natural killer cell function by flow cytometry.

    PubMed Central

    Kane, K L; Ashton, F A; Schmitz, J L; Folds, J D

    1996-01-01

    Natural killer cells (NK cells) are a subset of peripheral blood lymphocytes that mediate non-major histocompatibility complex-restricted cytotoxicity of foreign target cells. The "gold standard" assay for NK cell activity has been the chromium release assay. This method is not easily performed in the clinical laboratory because of difficulties with disposal of radioactive and hazardous materials, short reagent half-lives, expense, and difficulties with assay standardization. We describe a flow cytometric assay for the clinical measurement of NK cell activity. This study compared the chromium release assay and the flow cytometric assay by using clinically relevant specimens. There were no significant differences between the two assays in the measurement of lytic activity for 17 peripheral blood specimens or in reproducibility in repeated samplings of healthy individuals. We also established a normal range of values for NK activity in healthy adults and identified a small cluster of individuals who have exceptionally high or low levels of NK activity. The flow cytometric assay was validated by testing specimens from subjects expected to have abnormally low levels of NK activity (pregnant women) and specimens from healthy individuals in whom the activity of NK cells was enhanced by exposure to interleukin-2 or alpha interferon. Treatment with these agents was associated with a significant increase in NK activity. These results confirm and extend those of others, showing that the flow cytometric assay is a viable alternative to the chromium release assay for measuring NK cell activity. PMID:8705672

  1. Lysis of pig endothelium by IL-2 activated human natural killer cells is inhibited by swine and human major histocompatibility complex (MHC) class I gene products.

    PubMed

    Itescu, S; Artrip, J H; Kwiatkowski, P A; Wang, S F; Minanov, O P; Morgenthau, A S; Michler, R E

    1997-01-01

    We have previously described a form of xenograft rejection, mediated by natural killer (NK) cells, occurring in pig-to-primate organ transplants beyond the period of antibody-mediated hyperacute rejection. In this study, two distinct NK activation pathways were identified as mechanisms of pig aortic endotheliual cell (PAEC) lysis by human NK cells. Using an antibody-dependent cellular cytotoxicity (ADCC) assay, a progressive increase in human NK lysis of PAEC was observed following incubation with human IgG at increasing serum titer. In the absence of IgG, a second mechanism of PAEC lysis by human NK cells was observed following activation with IL-2. IL-2 activation of human NK cells increased lysis of PAEC by over 3-fold compared with ADCC. These results indicate that IL-2 activation of human NK cells induces significantly higher levels of lytic activity than does conventional ADCC involving IgG and FcRIII. We next investigated the role of MHC class I molecules in the regulation of NK lysis following IL-2 activation. PAEC expression of SLA class I molecules was increased by up to 75% by treatment with human TNFa. Following treatment with TNFa at 1 u/ml, IL-2 activated human NK lysis of PAEC was inhibited at every effector:target (E:T) ratio tested. Maximal effect occurred at an E:T ratio of 10:1, with TNFa inhibiting specific lysis by 59% (p < 0.01). Incubation with an anti-SLA class I Mab, but not IgG isotype control, abrogated the protective effects of TNFa on NK lysis of PAEC, suggesting direct inhibitory effects of SLA class I molecules on human NK function. To investigate whether human MHC class I molecules might have similar effects on human NK lysis of PAEC, further experiments were performed using a soluble peptide derived from the alpha-helical region of HLA-B7. Incubation with the HLA-B7 derived peptide significantly reduced the IL-2 activated NK lytic activity against PAEC in a dose-dependent fashion. Maximal effect occurred at a concentration of 10 mg

  2. Cytokine profile and natural killer cell activity in Listeria monocytogenes infected mice treated orally with Petiveria alliacea extract.

    PubMed

    Queiroz, M L; Quadros, M R; Santos, L M

    2000-08-01

    In this work, we investigated the effects of Petiveria alliacea extract on the production of Th1-type and Th2-type cytokines and on NK cells activity in normal and Listeria monocytogenes infected mice. Our results demonstrated that in normal/non-infected mice P. alliacea administration led to increased levels of Interleukin-2 (IL-2). The infection alone enhanced INF-gamma levels and NK cell activity at 48 and 72 hours of infection. The treatment with five consecutive doses of 1000 mg/kg/day of P. alliacea extract, given previously to infection, led to further increases in IL-2 levels, in relation to normal/non-infected/P. alliacea treated controls, and in INF-gamma levels at 72 h of infection, compared to infected mice. On the other hand, the production of IL-4 and IL-10 were not altered either by the infection or by the treatment with P. alliacea extract. NK cells activity increased at 48 h and 72 h following the inoculation of the bacteria. When mice were treated with P. alliacea previously to infection, NK activity was higher than that observed at 48 h, 72 h and 120 h of infection in the infected animal. Based on these findings we suggest that P. alliacea up-regulates anti-bacterial immune response by enhancing both Th1 function and the activity of NK cells.

  3. Regulation of Natural Killer Cell Function by STAT3

    PubMed Central

    Cacalano, Nicholas A.

    2016-01-01

    Natural killer (NK) cells, key members of a distinct hematopoietic lineage, innate lymphoid cells, are not only critical effectors that mediate cytotoxicity toward tumor and virally infected cells but also regulate inflammation, antigen presentation, and the adaptive immune response. It has been shown that NK cells can regulate the development and activation of many other components of the immune response, such as dendritic cells, which in turn, modulate the function of NK cells in multiple synergistic feed back loops driven by cell–cell contact, and the secretion of cytokines and chemokines that control effector function and migration of cells to sites of immune activation. The signal transducer and activator of transcription (STAT)-3 is involved in driving almost all of the pathways that control NK cytolytic activity as well as the reciprocal regulatory interactions between NK cells and other components of the immune system. In the context of tumor immunology, NK cells are a first line of defense that eliminates pre-cancerous and transformed cells early in the process of carcinogenesis, through a mechanism of “immune surveillance.” Even after tumors become established, NK cells are critical components of anticancer immunity: dysfunctional NK cells are often found in the peripheral blood of cancer patients, and the lack of NK cells in the tumor microenvironment often correlates to poor prognosis. The pathways and soluble factors activated in tumor-associated NK cells, cancer cells, and regulatory myeloid cells, which determine the outcome of cancer immunity, are all critically regulated by STAT3. Using the tumor microenvironment as a paradigm, we present here an overview of the research that has revealed fundamental mechanisms through which STAT3 regulates all aspects of NK cell biology, including NK development, activation, target cell killing, and fine tuning of the innate and adaptive immune responses. PMID:27148255

  4. Decreased non-MHC-restricted (CD56+) killer cell cytotoxicity after spaceflight

    NASA Technical Reports Server (NTRS)

    Mehta, S. K.; Kaur, I.; Grimm, E. A.; Smid, C.; Feeback, D. L.; Pierson, D. L.

    2001-01-01

    Cytotoxic activity of non-major histocompatibility complex-restricted (CD56+) (NMHC) killer cells and cell surface marker expression of peripheral blood mononuclear cells were determined before and after spaceflight. Ten astronauts (9 men, 1 woman) from two space shuttle missions (9- and 10-day duration) participated in the study. Blood samples were collected 10 days before launch, within 3 h after landing, and 3 days after landing. All peripheral blood mononuclear cell preparations were cryopreserved and analyzed simultaneously in a 4-h cytotoxicity (51)Cr release assay using K562 target cells. NMHC killer cell lytic activity was normalized per 1,000 CD56+ cells. When all 10 subjects were considered as one study group, NMHC killer cell numbers did not change significantly during the three sampling periods, but at landing lytic activity had decreased by approximately 40% (P < 0.05) from preflight values. Nine of ten astronauts had decreased lytic activity immediately after flight. NMHC killer cell cytotoxicity of only three astronauts returned toward preflight values by 3 days after landing. Consistent with decreased NMHC killer cell cytotoxicity, urinary cortisol significantly increased after landing compared with preflight levels. Plasma cortisol and ACTH levels at landing were not significantly different from preflight values. No correlation of changes in NMHC killer cell function or hormone levels with factors such as age, gender, mission, or spaceflight experience was found. After landing, expression of the major lymphocyte surface markers (CD3, CD4, CD8, CD14, CD16, CD56), as determined by flow cytometric analysis, did not show any consistent changes from measurements made before flight.

  5. NZ28-induced inhibition of HSF1, SP1 and NF-κB triggers the loss of the natural killer cell-activating ligands MICA/B on human tumor cells.

    PubMed

    Schilling, Daniela; Kühnel, Annett; Tetzlaff, Fabian; Konrad, Sarah; Multhoff, Gabriele

    2015-05-01

    The activity of natural killer (NK) cells is regulated by activating and inhibiting receptors, whereby the C-type lectin natural killer group 2D (NKG2D) receptor serves as the major activating receptor on NK cells which recognizes major histocompatibility class I chain-related proteins A and B (MICA/B). The MICA/B expression has been described to be regulated by the transcription factor heat shock factor 1 (HSF1). Inhibition of heat shock protein 90 (Hsp90) is known to induce the heat shock response via activation of HSF1 which is associated with tumor development, metastasis and therapy resistance and also with an increased susceptibility to NK cell-mediated lysis. Therefore, we compared the effects of Hsp90 inhibitor NVP-AUY922, HSF1 inhibitor NZ28 and HSF1 knockdown on the sensitivity of lung (H1339) and breast (MDA-MB-231, T47D) cancer cells to NK cell-mediated cytotoxicity and the expression of the NKG2D ligands MICA/B. Although NVP-AUY922 activates HSF1, neither the MICA/B surface density on tumor cells nor their susceptibility to NK cell-mediated lysis was affected. A single knockdown of HSF1 by shRNA decreased the surface expression of MICB but not that of MICA, and thereby, the NK cell-mediated lysis was only partially blocked. In contrast, NZ28 completely blocked the MICA/B membrane expression on tumor cells and thereby strongly inhibited the NK cell-mediated cytotoxicity. This effect might be explained by a simultaneous inhibition of the transcription factors HSF1, Sp1 and NF-κB by NZ28. These findings suggest that new anticancer therapeutics should be investigated with respect to their effects on the innate immune system.

  6. Natural Killer Cell Mediated Cytotoxic Responses in the Tasmanian Devil

    PubMed Central

    Brown, Gabriella K.; Kreiss, Alexandre; Lyons, A. Bruce; Woods, Gregory M.

    2011-01-01

    The Tasmanian devil (Sarcophilus harrisii), the world's largest marsupial carnivore, is under threat of extinction following the emergence of an infectious cancer. Devil facial tumour disease (DFTD) is spread between Tasmanian devils during biting. The disease is consistently fatal and devils succumb without developing a protective immune response. The aim of this study was to determine if Tasmanian devils were capable of forming cytotoxic antitumour responses and develop antibodies against DFTD cells and foreign tumour cells. The two Tasmanian devils immunised with irradiated DFTD cells did not form cytotoxic or humoral responses against DFTD cells, even after multiple immunisations. However, following immunisation with xenogenic K562 cells, devils did produce cytotoxic responses and antibodies against this foreign tumour cell line. The cytotoxicity appeared to occur through the activity of natural killer (NK) cells in an antibody dependent manner. Classical NK cell responses, such as innate killing of DFTD and foreign cancer cells, were not observed. Cells with an NK-like phenotype comprised approximately 4 percent of peripheral blood mononuclear cells. The results of this study suggest that Tasmanian devils have NK cells with functional cytotoxic pathways. Although devil NK cells do not directly recognise DFTD cancer cells, the development of antibody dependent cell-mediated cytotoxicity presents a potential pathway to induce cytotoxic responses against the disease. These findings have positive implications for future DFTD vaccine research. PMID:21957452

  7. Cytotoxic and natural killer cell stimulatory constituents of Phyllanthus songboiensis

    PubMed Central

    Ren, Yulin; Yuan, Chunhua; Deng, Youcai; Kanagasabai, Ragu; Ninh, Tran Ngoc; Tu, Vuong Tan; Chai, Hee-Byung; Soejarto, Djaja D.; Fuchs, James R.; Yalowich, Jack C.; Yu, Jianhua; Kinghorn, A. Douglas

    2014-01-01

    A dichapetalin-type triterpenoid and a dibenzylbutyrolactone-type lignan, together with five known lignans, a known aromatic diterpenoid, and a known acylated phytosterol, were isolated from the aerial parts of Phyllanthus songboiensis, collected in Vietnam. Their structures were determined by interpretation of the spectroscopic data, and the inhibitory activity toward the HT-29 human colon cancer cells of all isolates was evaluated by a cytotoxicity assay. The known arylnaphthalene lignan, (+)-acutissimalignan A, was highly cytotoxic toward HT-29 cells, with an IC50 value of 19 nM, but this compound was inactive as a DNA topoisomerase IIα (topo IIα) poison. The known phytosterol, (−)-β-sitosterol-3-O-β-D-(6-O-palmitoyl)glucopyranoside, was found to stimulate natural killer (NK) cells at a concentration of 10 μM in the presence of interleukin 12 (IL-12). PMID:25596805

  8. Activation of protein tyrosine kinase: a possible requirement for fixed-bacteria and lipopolysaccharide-induced increase in human natural killer cell activity.

    PubMed

    Puente, J; Salas, M A; Canon, C; Miranda, D; Wolf, M E; Mosnaim, A D

    1996-05-01

    Preincubation of peripheral blood lymphocytes (PBL) from drug-free, healthy volunteers with either the protein tyrosine kinase inhibitor genistein (GNT, n = 10, final concentration 200 microM) or the protein kinase A activator dybutiryl-cyclic-AMP (cAMP, n = 11, final concentration 10 microM), resulted in a significant inhibition of natural killer cell activity (NKCA, expressed as percentage of specific chromium release). With the exception of 4 out of the 11 cAMP-treated samples, individual values for NKCA in the drug preincubated specimens were at least 20% below the same subject baseline activity; furthermore, NKC lytic function was non-detectable in 4 out of the 10 and in 1 out of the 11 samples pretreated with either GNT or cAMP, respectively. PBL preincubation with glutaraldehyde-fixed Gram-negative bacteria (GNB, n = 13, final GNB-to-effector cell ratio of 50 : 1) resulted in a statistically significant increase in NKCA (baseline (x +/- SD) of 21.6 +/- 16.4 and bacteria treated samples of 41.5 +/- 24.6, respectively, Student's paired t-test p < 0.05). At least a 20% increase in NKC lytic function over its own baseline value was recorded for 11 out of the 13 samples tested (Table 1). Preincubation with GNB and GNT (5 samples) not only blocked the immunostimulant effects of GNB (Student's paired t-test p < 0.05), but in most cases individual values for NKCA were similar to those recorded for GNT-only treated samples. Use of cAMP instead of GNT also blocked, but to a smaller extent, the GNB-produced increases in NKC lytic function (paired Student's t-test < 0.05). PBL preincubation with lipopolysaccharide (LPS, n = 11, final concentration 50 micrograms/ml) resulted in a statistically significant increase in NKCA (baseline (x +/- SD) of 20.7 +/- 14.1 and LPS treated samples of 39.2 +/- 18.5, respectively, Student's paired t-test < 0.05). At least a 20% increase in NKCA over its own baseline value was observed for each and everyone of the 11 samples studied

  9. Therapeutic depletion of natural killer cells controls persistent infection.

    PubMed

    Waggoner, Stephen N; Daniels, Keith A; Welsh, Raymond M

    2014-02-01

    Persistent viral infections are associated with host and viral factors that impair effective antiviral immunity. Natural killer (NK) cells contribute to establishment of persistent lymphocytic choriomeningitis virus (LCMV) infection in mice through suppression of virus-specific T cell responses during the first few days of infection, but NK cell depletion during those early time points can enable severe T cell-mediated immune pathology and death of the host. Here we show that long after their peak in cytolytic activation, NK cells continue to support viral persistence at later times of infection. Delayed depletion of NK cells, 2 to 3 weeks after infection, enhanced virus-specific T cell responses and viral control. This enhancing effect of delayed NK cell depletion on antiviral immunity, in contrast to early NK cell depletion, was not associated with increased morbidity and mortality, and mice quickly regained weight after treatment. The efficacy of the depletion depended in part upon the size of the original virus inoculum, the viral load at the time of depletion, and the presence of CD4 T cells. Each of these factors is an important contributor to the degree of CD8 T cell dysfunction during viral persistence. Thus, NK cells may continuously contribute to exhaustion of virus-specific T cells during chronic infection, possibly by depleting CD4 T cells. Targeting of NK cells could thus be considered in combination with blockade of other immunosuppressive pathways, such as the interleukin-10 (IL-10) and programmed death 1 (PD-1) pathways, as a therapy to cure chronic human infections, including those with HIV or hepatitis C virus. IMPORTANCE Persistent virus infections are a major threat to global human health. The capacity of viruses, including HIV and hepatitis C virus, to overwhelm or subvert host immune responses contributes to a prolonged state of dampened antiviral immune functionality, which in turn facilitates viral persistence. Recent efforts have focused on

  10. FLT3 ligand administration after hematopoietic cell transplantation increases circulating dendritic cell precursors that can be activated by CpG oligodeoxynucleotides to enhance T-cell and natural killer cell function.

    PubMed

    Chen, Wei; Chan, Anissa S H; Dawson, Amanda J; Liang, Xueqing; Blazar, Bruce R; Miller, Jeffrey S

    2005-01-01

    Dendritic cells (DCs) are key effectors in innate immunity and play critical roles in triggering adaptive immune responses. FLT3 ligand (FLT3-L) is essential for DC development from hematopoietic progenitors. In a phase I clinical trial, we demonstrated that immunotherapy with subcutaneous injection of FLT3-L is safe and well tolerated in cancer patients recovering from autologous hematopoietic cell transplantation (HCT). FLT3-L administration significantly increased the frequency and absolute number of blood DC precursors without affecting other mature cell lineages during the 6-week course of FLT3-L therapy. After 14 days of FLT3-L administration, the number of blood CD11c + DCs, plasmacytoid DCs (PDCs), and CD14 + monocytes increased by 5.3-, 2.9-, 3.8-fold, respectively, and was maintained at increased levels throughout FLT3-L therapy. FLT3-L-increased blood DCs in HCT patients were immature and had modest enhancing effects on in vitro T-cell proliferation to antigens and natural killer (NK) cell function. The addition of type B CpG oligodeoxynucleotides (ODNs) to peripheral blood mononuclear cells obtained from HCT patients receiving FLT3-L therapy induced rapid maturation of both CD11c + DCs and PDCs and enhanced T-cell proliferative responses. In addition, CpG ODN induced potent activation of NK cells from FLT3-L-treated patients with increased surface CD69 expression and augmented cytotoxicity. CpG ODN-induced activation of NK cells was primarily via an indirect mechanism through PDCs. These findings suggest that FLT3-L mobilization of DC precursors followed by a specific DC stimulus such as CpG ODN may provide a novel strategy to manipulate antitumor immunity in patients after HCT. PMID:15625541

  11. Importance of killer immunoglobulin-like receptors in allogeneic hematopoietic stem cell transplantation

    PubMed Central

    Franceschi, Danilo Santana Alessio; de Souza, Cármino Antonio; Aranha, Francisco José Penteado; Cardozo, Daniela Maira; Sell, Ana Maria; Visentainer, Jeane Eliete Laguila

    2011-01-01

    Hematopoietic stem cell transplantation is the treatment of choice for many hematologic diseases, such as multiple myeloma, bone marrow aplasia and leukemia. Human leukocyte antigen (HLA) compatibility is an important tool to prevent post-transplant complications such as graft rejection and graft-versus-host disease, but the high rates of relapse limit the survival of transplant patients. Natural Killer cells, a type of lymphocyte that is a key element in the defense against tumor cells, cells infected with viruses and intracellular microbes, have different receptors on their surfaces that regulate their cytotoxicity. Killer immunoglobulin-like receptors are the most important, interacting consistently with human leukocyte antigen class I molecules present in other cells and thus controlling the activation of natural killer cells. Several studies have shown that certain combinations of killer immunoglobulin-like receptors and human leukocyte antigens (in both donors and recipients) can affect the chances of survival of transplant patients, particularly in relation to the graft-versusleukemia effect, which may be associated to decreased relapse rates in certain groups. This review aims to shed light on the mechanisms and effects of killer immunoglobulin-like receptors - human leukocyte antigen associations and their implications following hematopoietic stem cell transplantation, and to critically analyze the results obtained by the studies presented herein. PMID:23284260

  12. Identification of Anti-tumor Cells Carrying Natural Killer (NK) Cell Antigens in Patients With Hematological Cancers.

    PubMed

    Krzywinska, Ewelina; Allende-Vega, Nerea; Cornillon, Amelie; Vo, Dang-Nghiem; Cayrefourcq, Laure; Panabieres, Catherine; Vilches, Carlos; Déchanet-Merville, Julie; Hicheri, Yosr; Rossi, Jean-François; Cartron, Guillaume; Villalba, Martin

    2015-10-01

    Natural killer (NK) cells, a cytotoxic lymphocyte lineage, are able to kill tumor cells in vitro and in mouse models. However, whether these cells display an anti-tumor activity in cancer patients has not been demonstrated. Here we have addressed this issue in patients with several hematological cancers. We found a population of highly activated CD56(dim)CD16(+) NK cells that have recently degranulated, evidence of killing activity, and it is absent in healthy donors. A high percentage of these cells expressed natural killer cell p46-related protein (NKp46), natural-killer group 2, member D (NKG2D) and killer inhibitory receptors (KIRs) and a low percentage expressed NKG2A and CD94. They are also characterized by a high metabolic activity and active proliferation. Notably, we found that activated NK cells from hematological cancer patients have non-NK tumor cell antigens on their surface, evidence of trogocytosis during tumor cell killing. Finally, we found that these activated NK cells are distinguished by their CD45RA(+)RO(+) phenotype, as opposed to non-activated cells in patients or in healthy donors displaying a CD45RA(+)RO(-) phenotype similar to naïve T cells. In summary, we show that CD45RA(+)RO(+) cells, which resemble a unique NK population, have recognized tumor cells and degranulate in patients with hematological neoplasias.

  13. Identification of Anti-tumor Cells Carrying Natural Killer (NK) Cell Antigens in Patients With Hematological Cancers

    PubMed Central

    Krzywinska, Ewelina; Allende-Vega, Nerea; Cornillon, Amelie; Vo, Dang-Nghiem; Cayrefourcq, Laure; Panabieres, Catherine; Vilches, Carlos; Déchanet-Merville, Julie; Hicheri, Yosr; Rossi, Jean-François; Cartron, Guillaume; Villalba, Martin

    2015-01-01

    Natural killer (NK) cells, a cytotoxic lymphocyte lineage, are able to kill tumor cells in vitro and in mouse models. However, whether these cells display an anti-tumor activity in cancer patients has not been demonstrated. Here we have addressed this issue in patients with several hematological cancers. We found a population of highly activated CD56dimCD16+ NK cells that have recently degranulated, evidence of killing activity, and it is absent in healthy donors. A high percentage of these cells expressed natural killer cell p46-related protein (NKp46), natural-killer group 2, member D (NKG2D) and killer inhibitory receptors (KIRs) and a low percentage expressed NKG2A and CD94. They are also characterized by a high metabolic activity and active proliferation. Notably, we found that activated NK cells from hematological cancer patients have non-NK tumor cell antigens on their surface, evidence of trogocytosis during tumor cell killing. Finally, we found that these activated NK cells are distinguished by their CD45RA+RO+ phenotype, as opposed to non-activated cells in patients or in healthy donors displaying a CD45RA+RO− phenotype similar to naïve T cells. In summary, we show that CD45RA+RO+ cells, which resemble a unique NK population, have recognized tumor cells and degranulate in patients with hematological neoplasias. PMID:26629531

  14. Are increased frequency of macrophage-like and natural killer (NK) cells, together with high levels of NKT and CD4+CD25high T cells balancing activated CD8+ T cells, the key to control Chagas’ disease morbidity?

    PubMed Central

    Vitelli-Avelar, D M; Sathler-Avelar, R; Massara, R L; Borges, J D; Lage, P S; Lana, M; Teixeira-Carvalho, A; Dias, J C P; Elói-Santos, S M; Martins-Filho, O A

    2006-01-01

    The immunological response during early human Trypanosoma cruzi infection is not completely understood, despite its role in driving the development of distinct clinical manifestations of chronic infection. Herein we report the results of a descriptive flow cytometric immunophenotyping investigation of major and minor peripheral blood leucocyte subpopulations in T. cruzi-infected children, characterizing the early stages of the indeterminate clinical form of Chagas’ disease. Our results indicated significant alterations by comparison with uninfected children, including increased values of pre-natural killer (NK)-cells (CD3– CD16+ CD56–), and higher values of proinflammatory monocytes (CD14+ CD16+ HLA-DR++). The higher values of activated B lymphocytes (CD19+ CD23+) contrasted with impaired T cell activation, indicated by lower values of CD4+ CD38+ and CD4+ HLA-DR+ lymphocytes, a lower frequency of CD8+ CD38+ and CD8+ HLA-DR+ cells; a decreased frequency of CD4+ CD25HIGH regulatory T cells was also observed. These findings reinforce the hypothesis that simultaneous activation of innate and adaptive immunity mechanisms in addition to suppression of adaptive cellular immune response occur during early events of Chagas’ disease. Comparative cross-sectional analysis of these immunophenotypes with those exhibited by patients with late chronic indeterminate and cardiac forms of disease suggested that a shift toward high values of macrophage-like cells extended to basal levels of proinflammatory monocytes as well as high values of mature NK cells, NKT and regulatory T cells, may account for limited tissue damage during chronic infection favouring the establishment/maintenance of a lifelong indeterminate clinical form of the disease. On the other hand, development of an adaptive cell-mediated inflammatory immunoprofile characterized by high levels of activated CD8+ cells and basal levels of mature NK cells, NKT and CD4+ CD25HIGH cells might lead to late chronic

  15. Recognition of adult and pediatric acute lymphoblastic leukemia blasts by natural killer cells.

    PubMed

    Torelli, Giovanni F; Peragine, Nadia; Raponi, Sara; Pagliara, Daria; De Propris, Maria S; Vitale, Antonella; Bertaina, Alice; Barberi, Walter; Moretta, Lorenzo; Basso, Giuseppe; Santoni, Angela; Guarini, Anna; Locatelli, Franco; Foà, Robin

    2014-07-01

    In this study, we aimed to investigate the pathways of recognition of acute lymphoblastic leukemia blasts by natural killer cells and to verify whether differences in natural killer cell activating receptor ligand expression among groups defined by age of patients, or presence of cytogenetic/molecular aberrations correlate with the susceptibility to recognition and killing. We analyzed 103 newly diagnosed acute lymphoblastic leukemia patients: 46 adults and 57 children. Pediatric blasts showed a significantly higher expression of Nec-2 (P=0.03), ULBP-1 (P=0.01) and ULBP-3 (P=0.04) compared to adult cells. The differential expression of these ligands between adults and children was confined to B-lineage acute lymphoblastic leukemia with no known molecular alterations. Within molecularly defined subgroups of patients, a high surface expression of NKG2D and DNAM1 ligands was found on BCR-ABL(+) blasts, regardless of patient age. Accordingly, BCR-ABL(+) blasts proved to be significantly more susceptible to natural killer-dependent lysis than B-lineage blasts without molecular aberrations (P=0.03). Cytotoxic tests performed in the presence of neutralizing antibodies indicated a pathway of acute lymphoblastic leukemia cell recognition in the setting of the Nec-2/DNAM-1 interaction. These data provide a biological explanation of the different roles played by alloreactive natural killer cells in pediatric versus adult acute lymphoblastic leukemia and suggest that new natural killer-based strategies targeting specific subgroups of patients, particularly those BCR-ABL(+), are worth pursuing further.

  16. Natural killer cells in patients with polycythemia vera.

    PubMed

    Sanchez, Carole; Baier, Céline; Colle, Julien G; Chelbi, Rabie; Rihet, Pascal; Le Treut, Thérèse; Imbert, Jean; Sébahoun, Gérard; Venton, Geoffroy; Costello, Régis T

    2015-09-01

    Natural killer cells (NK) are pivotal cells of innate immunity. They are potent antileukemic cytotoxic effectors. A defect in their cytotoxicity has been described in some hematopoietic malignancies such as acute myeloid leukemia, multiple myeloma and myelodysplastic syndromes. This defect is at least partially linked to a decreased or absent expression of some activating NK cells molecules, more particularly the so-called natural cytotoxicity receptors. In the present study, we more particularly focused our attention on NK cells of polycythemia vera, a myeloproliferative disease characterized by the presence of mutated JAK2 tyrosine kinase. The polymerase chain reaction analysis of NK cells from patients showed that they expressed the mutated form of JAK2. In polycythemia vera the proportion of NK was increased compared to healthy donors. The proliferative and cytotoxic abilities of NK cells from patients were similar to healthy donors. Expression of activating or inhibitory receptors was comparable in patients and donors, with nonetheless an imbalance for the inhibitory form of the CD158a,h couple of receptors in patients. Finally, the transcriptomic profile analysis clearly identified a discriminant signature between NK cells from patients and donors that could putatively be the consequence of abnormal continuous activation of mutated JAK2.

  17. Phenotypic analysis of nylon-wool-adherent suppressor cells that inhibit the effector process of tumour cell lysis by lymphokine-activated killer cells in patients with advanced gastric carcinoma.

    PubMed

    Koyama, S; Fukao, K

    1994-01-01

    The causes of down-regulation of cytotoxic immune responses in cancer patients have not been fully evaluated. We previously demonstrated that T-cell-growth-factor-activated peripheral blood lymphocytes (PBL) with the surface phenotype CD8+ CD11b-, from patients with widespread metastasis of gastric carcinoma, inhibited the effector process of lymphokine-activated-killer(LAK)-cell-mediated cytolysis. In this study, we examined suppressor cell activity in freshly prepared PBL from 18 patients with advanced gastric carcinoma, and 10 normal healthy individuals. The suppressor cell activity was assayed by recording whether or not PBL inhibited directly the effector process of LAK cell cytotoxicity. Most of the PBL suspensions from cancer patients showed that they contained a population of cells that can directly inhibit the effector phase of tumor cell lysis of the cytotoxic cells. To analyze further the PBL responsible for the suppression, the cells were passed over a nylon-wool column. Nylon-wool-adherent cells significantly augmented the suppression, while the cells passing through abrogated the suppressive effect. Most nylon-wool-adherent cells from 10 normal healthy controls did not inhibit the cytotoxic reaction. To determine further the suppressor-effector population in nylon-wool-adherent cells, negative-selection studies using CD8-, CD4- or CD11b-coated magnetic beads, and positive-selection studies using CD8- or CD4-coated magnetic beads were performed. Finally the results suggest that the suppressor-effector cells comprise at least two different surface phenotypes: CD8+ T and CD8-CD11b+ cells. The possible role of CD4+ T cells and HLA-DR+ LeuM3+ macrophages as suppressor cells was ruled out in nylon-wool-adherent cells. CD8+ T and possibly CD8-CD11b+ cells apparently suppressed the efferent limb of the antitumor immunity. The selective immune suppression mediated by these cells may partly be concerned with escape mechanisms of gastric carcinoma from the host

  18. Pit cells as liver-associated natural killer cells: morphology and function.

    PubMed

    Nakatani, Kazuki; Kaneda, Kenji; Seki, Shuichi; Nakajima, Yuji

    2004-03-01

    Pit cells are one type of hepatic sinusoidal cells, defined morphologically as large granular lymphocytes (LGLs) and functionally as liver-associated natural killer (NK) cells. They are situated inside the sinusoidal lumen, adhering to the endothelial cells and Kupffer cells. They contain multivesicular body-related dense granules and rod-cored vesicles. The number and size of granules and vesicles differ between hepatic NK cells and peripheral blood cells, suggesting possible differentiation of the latter into the former in the microenvironment of the liver. In addition to NK cells, natural killer T (NKT) cells are also abundant in the liver. They share several morphological properties with NK cells, and at least some are probably observed as pit cells under an electron microscope. NK cells recognize target cells with their surface receptors such as inhibitory and activating receptors. They exert antitumor functions by exocytosis of perforin/granzyme-containing granules, induction of death receptor-mediated apoptosis in target cells, and production of various cytokines that augment the activities of other immune cells. NKT cells play important roles in initiating and assisting the function of NK cells by producing interferon-gamma.

  19. Interferon induces natural killer cell blastogenesis in vivo

    NASA Technical Reports Server (NTRS)

    Biron, C. A.; Sonnenfeld, G.; Welsh, R. M.

    1984-01-01

    Interferon (IFN), types beta and gamma, and IFN inducers polyinosinic-polycytidylic acid and lymphocytic choriomeningitis virus, all stimulated the generation of blast-natural killer (NK) cells in mouse spleens, Blast-NK cells were characterized on the basis of size, 3H-thymidine uptake, and NK cell markers These data indicate that in addition to augmenting NK cell-mediated lysis, IFN may regulate NK cell proliferation in vivo.

  20. The skin test antigen stimulated killer (STAK) cell mediating NK like CMC is OKM1 positive and OKT3 negative.

    PubMed Central

    Tartof, D; Curran, J J; Levitt, D; Loken, M R

    1983-01-01

    Recently we demonstrated that candida antigen stimulated natural killer cell like cell-mediated cytolysis (NK like CMC) in peripheral blood mononuclear cells (PBMNC) isolated from normal individuals (Tartof et al., 1980). Utilizing monoclonal antibodies directed against human mononuclear cell subpopulations in conjunction with a fluorescence activated cell sorter (FACS) we determined that, similar to the previously described NK cell, the skin test antigen stimulated killer (STAK) cell is a larger OKM1 positive, OKT3 negative cell. We obtained similar results using two different skin test antigens. Thus, stimulation of NK like CMC in PBMNC by skin test antigens probably represents activation of NK or NK like cells. PMID:6360439

  1. Studies of natural killer cell activity in a drug-free, healthy population. Response to a challenge with taxol, estramustine and lipopolysaccharide.

    PubMed

    Puente, J; Diaz, M; Salas, M A; Miranda, D; Gaggero, A; Wolf, M E; Mosnaim, A D

    1995-08-01

    Preincubation of peripheral blood lymphocytes with the microtubule disturbing agents estramustine (ETMN; n = 7, final conc. 20 microM) or taxol (TX; n = 13, final conc. 10 microM), resulted in a statistically significant inhibition of natural killer cell activity [(NKCA); baseline (x +/- SD; expressed as percentage of specific chromium release) of 32.2 +/- 30.5 and 34.4 +/- 27.7 and drug treated samples of 13.9 +/- 19.9 and 12.5 +/- 20.8, respectively; Student's paired t-test p < 0.005]. Furthermore, most individual values for NKCA in the drug preincubated samples were at least 20% below the same subject baseline lytic function (except for TX sample No.1), and NKCA was non detectable in 4 out of 7 and 5 out of 13 samples (pretreatment with either ETMN or TX< respectively). The use of other concentrations and different preincubation times for these chemotherapeutic agents also produced NKCA inhibition, which was time and dose dependent. Preincubation with lipopolysaccharide (LPS; n = 16, final conc. 50 micrograms/ml), an endotoxin prominently involved in the etiology of septic shock, resulted in a statistically significant enhancement of NKCA [baseline (x +/- SD; expressed as percentage of specific chromium release) of 25.4 +/- 20.4 and LPS treated sample of 36.6 +/- 17.4, respectively; Student's t paired t-test p < 0.005]. At least a 20% increase in NKC lytic function over its own baseline value was recorded for each and everyone of the samples tested with LPS.2+ the pathophysiology associated with septic shock.

  2. Impaired cytotoxicity associated with defective natural killer cell differentiation in myelodysplastic syndromes.

    PubMed

    Hejazi, Maryam; Manser, Angela R; Fröbel, Julia; Kündgen, Andrea; Zhao, Xiaoyi; Schönberg, Kathrin; Germing, Ulrich; Haas, Rainer; Gattermann, Norbert; Uhrberg, Markus

    2015-05-01

    Natural killer cells are well known to mediate anti-leukemic responses in myeloid leukemia but their role in myelodysplastic syndromes is not well understood. Here, in a cohort of newly diagnosed patients (n=75), widespread structural and functional natural killer cell defects were identified. One subgroup of patients (13%) had a selective deficiency of peripheral natural killer cells (count <10/mm(3) blood) with normal frequencies of T and natural killer-like T cells. Natural killer cell-deficient patients were predominantly found in high-risk subgroups and deficiency of these cells was significantly associated with poor prognosis. In the second subgroup, comprising the majority of patients (76%), natural killer cells were present but exhibited poor cytotoxicity. The defect was strongly associated with reduced levels of perforin and granzyme B. Notably, natural killer cell function and arming of cytotoxic granules could be fully reconstituted by in vitro stimulation. Further phenotypic analysis of these patients revealed an immature natural killer cell compartment that was biased towards CD56(bright) cells. The residual CD56(dim) cells exhibited a significant increase of the unlicensed NKG2A(-)KIR(-) subset and a striking reduction in complexity of the repertoire of killer cell immunoglobulin-like receptors. Taken together, these results suggest that the widespread defects in natural killer cell function occurring in patients with myelodysplastic syndromes are mostly due to either unsuccessful or inefficient generation of mature, functionally competent natural killer cells, which might contribute to disease progression through impaired immune surveillance.

  3. Paucity of natural killer and cytotoxic T cells in human neuromyelitis optica lesions.

    PubMed

    Saadoun, Samira; Bridges, Leslie R; Verkman, A S; Papadopoulos, Marios C

    2012-12-19

    Neuromyelitis optica is a severe inflammatory demyelinating disease of the central nervous system. Most patients with neuromyelitis optica have circulating immunoglobulin G (IgG) antibodies against the astrocytic water channel protein aquaporin-4 (AQP4), which are pathogenic. Anti-AQP4 IgG-mediated complement-dependent astrocyte toxicity is a key mechanism of central nervous system damage in neuromyelitis optica, but the role of natural killer and cytotoxic T cells is unknown. Our objective was to determine whether natural killer and cytotoxic T cells play a role in human neuromyelitis optica lesions. We immunostained four actively demyelinating lesions, obtained from patients with anti-AQP4 IgG positive neuromyelitis optica, for Granzyme B and Perforin. The inflammatory cells were perivascular neutrophils, eosinophils and macrophages, with only occasional Granzyme B+ or Perforin+ cells. Greater than 95% of inflamed vessels in each lesion had no surrounding Granzyme B+ or Perforin+ cells. Granzyme B+ or Perforin+ cells were abundant in human spleen (positive control). Although natural killer cells produce central nervous system damage in mice injected with anti-AQP4 IgG, our findings here indicate that natural killer-mediated and T cell-mediated cytotoxicity are probably not involved in central nervous system damage in human neuromyelitis optica.

  4. Spectrum of diseases associated with increased proportions or absolute numbers of peripheral blood natural killer cells.

    PubMed

    Okuno, S H; Tefferi, A; Hanson, C A; Katzmann, J A; Li, C Y; Witzig, T E

    1996-06-01

    In a retrospective review of 1501 lymphoid flow cytometric studies of peripheral blood, we identified an increased proportion of natural killer cells in 125 cases (8%), 49 (3%) of which had a concomitant increase in absolute number of natural killer cells. Of the latter, the most frequent associated disorder was chronic natural killer cell lymphocytosis. Substantial quantitative increases in natural killer cells were also observed in some patients with lymphoma, leukaemia, immune thrombocytopenic purpura, or myelodysplastic syndrome. Our study provides incidence figures and clinical associations of an increased number of natural killer cells in the peripheral blood.

  5. Dietary Supplementation with White Button Mushroom Enhances Natural Killer Cell Activity in C57BL/6 Mice

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Mushrooms have been shown to possess anti-tumor, anti-viral, and anti-bacterial properties. These effects of mushrooms are suggested to be due to their ability to modulate immune cell functions. However, majority of these studies evaluated the effect of administering extracts of exotic mushrooms thr...

  6. The influence of natural killer cells in neuroblastoma.

    PubMed

    Reynolds, J V; Shou, J; Choi, H; Sigal, R; Ziegler, M M; Daly, J M

    1989-02-01

    Human neuroblastoma (NRB) cell lines are markedly sensitive to natural killer (NK) cell lysis in vitro, but patients with NRBs have low or absent NK activity. This study evaluated the NK sensitivity of murine NRBs (C1300 and TBJ) in the regulation of NRB growth and determined the effects of recombinant (r) interferon gamma and recombinant interleukin 2 (rIL-2). Both basal (8% +/- 3% specific cytotoxicity) and induced (20% +/- 3%) NK lyses of C1300-NRB were observed. In vivo depletion of NK cells with anti-asialo GM-1 significantly enhanced growth of C1300-NRB and decreased survival. Treatment with r-interferon gamma or rIL-2 on days 1 through 3 after C1300-NRB inoculation significantly prolonged the mean tumor latency period, decreased the tumor growth rate, and enhanced in vitro NK killing of C1300-NRB and YAC-1. The effects of r-interferon gamma and IL-2 were abrogated by pretreatment with anti-asialo GM-1. These results demonstrated that NK cells form one important component of regulation of a murine NRB, but immunomodulation with potent lymphokines requires cooperation of more than one cell type.

  7. Nitrogen availability of grape juice limits killer yeast growth and fermentation activity during mixed-culture fermentation with sensitive commercial yeast strains.

    PubMed Central

    Medina, K; Carrau, F M; Gioia, O; Bracesco, N

    1997-01-01

    The competition between selected or commercial killer strains of type K2 and sensitive commercial strains of Saccharomyces cerevisiae was studied under various conditions in sterile grape juice fermentations. The focus of this study was the effect of yeast inoculation levels and the role of assimilable nitrogen nutrition on killer activity. A study of the consumption of free amino nitrogen (FAN) by pure and mixed cultures of killer and sensitive cells showed no differences between the profiles of nitrogen assimilation in all cases, and FAN was practically depleted in the first 2 days of fermentation. The effect of the addition of assimilable nitrogen and the size of inoculum was examined in mixed killer and sensitive strain competitions. Stuck and sluggish wine fermentations were observed to depend on nitrogen availability when the ratio of killer to sensitive cells was low (1:10 to 1:100). A relationship between the initial assimilable nitrogen content of must and the proportion of killer cells during fermentation was shown. An indirect relationship was found between inoculum size and the percentage of killer cells: a smaller inoculum resulted in a higher proportion of killer cells in grape juice fermentations. In all cases, wines obtained with pure-culture fermentations were preferred to mixed-culture fermentations by sensory analysis. The reasons why killer cells do not finish fermentation under competitive conditions with sensitive cells are discussed. PMID:9212430

  8. Natural killer cells: the secret weapon in dendritic cell vaccination strategies.

    PubMed

    Van Elssen, Catharina H M J; Oth, Tammy; Germeraad, Wilfred T V; Bos, Gerard M J; Vanderlocht, Joris

    2014-03-01

    In cancer therapy, dendritic cell (DC) vaccination is still being explored. Clinical responses, however, are diverse and there is a lack of immunologic readout systems that correspond with clinical outcome. Only in the minority of patients, T-cell responses correlate with clinical outcome, indicating that other immune cells also gain anticancer activity. We still have limited knowledge of the effect of DC vaccination on different immune effector cells. However, it has been shown that bidirectional cross-talk between natural killer (NK) cells and DCs is responsible for enhanced activation of both cell types and increases their antitumor activity. In this review, we postulate the possibility that NK cells are the secret weapons in DC vaccination and studying their behavior together with T-cell activation in vaccinated individuals might predict clinical outcome. PMID:24590885

  9. Cardif (MAVS) Regulates the Maturation of Natural Killer Cells

    PubMed Central

    Haynes, LaTeira D.; Verma, Shilpi; McDonald, Bryan; Wu, Runpei; Tacke, Robert; Ekstein, Jennifer; Feuvrier, Ariana; Benedict, Chris A.; Hedrick, Catherine C.

    2015-01-01

    Cardif, also known as IPS-1, VISA and, MAVS, is an intracellular adaptor protein that functions downstream of the RIG-I family of pattern recognition receptors. Cardif is required for the production of type I-IFNs and other inflammatory cytokines after RIG-I like receptors recognize intracellular antigenic RNA. Studies have recently shown that Cardif may have other roles in the immune system in addition to its role in viral immunity. In this study, we find that the absence of Cardif alters normal natural killer cell development and maturation. Cardif−/− mice have a 35% loss of mature CD27−CD11b+ NK cells in the periphery. Additionally, Cardif−/− NK cells have altered surface marker expression, lower cytoxicity, decreased intracellular STAT1 levels, increased apoptosis and decreased proliferation compared to wild-type NK cells. Mixed chimeric mice revealed that the defective maturation and increased apoptotic rate of peripheral Cardif−/− NK cells is cell-intrinsic. However, Cardif−/− mice showed enhanced control of mouse cytomegalovirus (MCMV, a DNA β-herpesvirus) by NK cells, commensurate with increased activation and IFNγ production by these immature NK cell subsets. These results indicate that the skewed differentiation and altered STAT expression of Cardif−/− NK cells can result in their hyper-responsiveness in some settings, and support recent findings that Cardif-dependent signaling can regulate aspects of immune cell development and/or function distinct from its well characterized role in mediating cell-intrinsic defense to RNA viruses. PMID:26232430

  10. Evolution of non-cytotoxic uterine natural killer cells.

    PubMed

    Kalkunte, Satyan; Chichester, Clinton O; Gotsch, Francesca; Sentman, Charles L; Romero, Roberto; Sharma, Surendra

    2008-05-01

    The immune tolerance and de novo vascularization are two highly intriguing processes at the maternal-fetal interface that appear to be central to normal pregnancy outcome. Immune tolerance occurs despite the local presence of an active maternal immune system including macrophages, dendritic cells and specialized CD56(bright)CD16(-) uterine natural killer (uNK) cells (65-70%). Recent observations indicate that the phenotypic and functional repertoire of uNK cells is distinct from peripheral blood NK and endometrial NK cells, challenging the understanding of their temporal occurrence and function. Origin and specialized programming of uNK cells continue to be debated. uNK cells, replete with an armamentarium to kill the foreign, tolerate the conceptus and facilitate pregnancy. Why do these uNK cells remain non-cytotoxic? Are these NK cells 'multitasking' in nature harboring beneficial and detrimental roles in pregnancy? Are there distinct subpopulations of NK cells that may populate the decidua? We propose that the endometrium/decidua functions as an 'inducible tertiary lymphoid tissue' that supports the recruitment and expansion of CD56(bright)CD16(-) NK cells and induces transcriptional up-regulation of angiogenic machinery in response to exposure to local hormonal factors, cytokine milieu and perhaps hypoxia. The angiogenic features of uNK cells could further result in a 'multitasking' phenotype that still remains to be characterized. This article discusses the factors and pathways that bridge the angiogenic and non-cytotoxic response machineries at the maternal-fetal interface. PMID:18405313

  11. Newtonian cell interactions shape natural killer cell education.

    PubMed

    Goodridge, Jodie P; Önfelt, Björn; Malmberg, Karl-Johan

    2015-09-01

    Newton's third law of motion states that for every action on a physical object there is an equal and opposite reaction. The dynamic change in functional potential of natural killer (NK) cells during education bears many features of such classical mechanics. Cumulative physical interactions between cells, under a constant influence of homeostatic drivers of differentiation, lead to a reactive spectrum that ultimately shapes the functionality of each NK cell. Inhibitory signaling from an array of self-specific receptors appear not only to suppress self-reactivity but also aid in the persistence of effector functions over time, thereby allowing the cell to gradually build up a functional potential. Conversely, the frequent non-cytolytic interactions between normal cells in the absence of such inhibitory signaling result in continuous stimulation of the cells and attenuation of effector function. Although an innate cell, the degree to which the fate of the NK cell is predetermined versus its ability to adapt to its own environment can be revealed through a Newtonian view of NK cell education, one which is both chronological and dynamic. As such, the development of NK cell functional diversity is the product of qualitatively different physical interactions with host cells, rather than simply the sum of their signals or an imprint based on intrinsically different transcriptional programs.

  12. Newtonian cell interactions shape natural killer cell education.

    PubMed

    Goodridge, Jodie P; Önfelt, Björn; Malmberg, Karl-Johan

    2015-09-01

    Newton's third law of motion states that for every action on a physical object there is an equal and opposite reaction. The dynamic change in functional potential of natural killer (NK) cells during education bears many features of such classical mechanics. Cumulative physical interactions between cells, under a constant influence of homeostatic drivers of differentiation, lead to a reactive spectrum that ultimately shapes the functionality of each NK cell. Inhibitory signaling from an array of self-specific receptors appear not only to suppress self-reactivity but also aid in the persistence of effector functions over time, thereby allowing the cell to gradually build up a functional potential. Conversely, the frequent non-cytolytic interactions between normal cells in the absence of such inhibitory signaling result in continuous stimulation of the cells and attenuation of effector function. Although an innate cell, the degree to which the fate of the NK cell is predetermined versus its ability to adapt to its own environment can be revealed through a Newtonian view of NK cell education, one which is both chronological and dynamic. As such, the development of NK cell functional diversity is the product of qualitatively different physical interactions with host cells, rather than simply the sum of their signals or an imprint based on intrinsically different transcriptional programs. PMID:26284479

  13. Functional heterogeneity among cytotoxic clones derived from natural killer cells.

    PubMed

    Christmas, S E; Moore, M

    1987-01-01

    Clones were obtained from highly purified populations of human peripheral blood natural killer (NK) cells propagated in the presence of interleukin-2 and phytohaemagglutinin. Almost all clones were cytotoxic against standard NK targets and many were also able to kill the B lymphoblastoid cell line BSM. This latter property was not necessarily a result of the incorporation of this cell line into the feeder mixture used to derive the clones. In most cloning experiments there was a high degree of concordance between the killing of the NK targets K562 and Molt 4 by panels of clones. In some cases this extended to the killing of BSM targets but in other instances there was no relationship or even an inverse correlation between killing of BSM and other targets. In a single cloning experiment there was no relationship between killing of BSM and Raji targets. In some cases a panel of clones could be divided into two or more distinct groups based on their differential activity towards BSM and K562. Such differences were not solely due to inter-donor variation. These findings were extended by cold target inhibition experiments in which at least three types of clone were identified. In one group of clones, which was nonreactive towards BSM, cold BSM significantly enhanced the killing of K562 in a dose-dependent fashion. These experiments provide evidence for a limited degree of functional heterogeneity among clones derived from human peripheral blood NK cells.

  14. Killer-cell immunoglobulin-like receptor genes and ligands and their role in hematologic malignancies.

    PubMed

    Varbanova, Viktoria; Naumova, Elissaveta; Mihaylova, Anastasiya

    2016-04-01

    Natural killer (NK) cells are considered crucial for the elimination of emerging tumor cells. Effector NK-cell functions are controlled by interactions of inhibitory and activating killer-cell immunoglobulin-like receptors (KIRs) on NK cells with human leukocyte antigen (HLA) class I ligands on target cells. KIR and HLA are highly polymorphic genetic systems segregating independently, creating a great diversity in KIR/HLA gene profiles in different individuals. There is an increasing evidence supporting the relevance of KIR and HLA ligand gene background for the occurrence and outcome of certain cancers. However, the data are still controversial and the mechanisms of receptor-ligand mediated NK-cell action remain unclear. Here, the main characteristics and functions of KIRs and their HLA class I ligands are reviewed. In addition, we review the HLA and KIR correlations with different hematological malignancies and discuss our current understanding of the biological significance and mechanisms underlying these associations.

  15. Recognition of Microbial Glycolipids by Natural Killer T Cells.

    PubMed

    Zajonc, Dirk M; Girardi, Enrico

    2015-01-01

    T cells can recognize microbial antigens when presented by dedicated antigen-presenting molecules. While peptides are presented by classical members of the major histocompatibility complex (MHC) family (MHC I and II), lipids, glycolipids, and lipopeptides can be presented by the non-classical MHC member, CD1. The best studied subset of lipid-reactive T cells are type I natural killer T (iNKT) cells that recognize a variety of different antigens when presented by the non-classical MHCI homolog CD1d. iNKT cells have been shown to be important for the protection against various microbial pathogens, including B. burgdorferi, the causative agents of Lyme disease, and S. pneumoniae, which causes pneumococcal meningitis and community-acquired pneumonia. Both pathogens carry microbial glycolipids that can trigger the T cell antigen receptor (TCR), leading to iNKT cell activation. iNKT cells have an evolutionary conserved TCR alpha chain, yet retain the ability to recognize structurally diverse glycolipids. They do so using a conserved recognition mode, in which the TCR enforces a conserved binding orientation on CD1d. TCR binding is accompanied by structural changes within the TCR binding site of CD1d, as well as the glycolipid antigen itself. In addition to direct recognition of microbial antigens, iNKT cells can also be activated by a combination of cytokines (IL-12/IL-18) and TCR stimulation. Many microbes carry TLR antigens, and microbial infections can lead to TLR activation. The subsequent cytokine response in turn lower the threshold of TCR-mediated iNKT cell activation, especially when weak microbial or even self-antigens are presented during the cause of the infection. In summary, iNKT cells can be directly activated through TCR triggering of strong antigens, while cytokines produced by the innate immune response may be necessary for TCR triggering and iNKT cell activation in the presence of weak antigens. Here, we will review the molecular basis of iNKT cell

  16. Recognition of Microbial Glycolipids by Natural Killer T Cells

    PubMed Central

    Zajonc, Dirk M.; Girardi, Enrico

    2015-01-01

    T cells can recognize microbial antigens when presented by dedicated antigen-presenting molecules. While peptides are presented by classical members of the major histocompatibility complex (MHC) family (MHC I and II), lipids, glycolipids, and lipopeptides can be presented by the non-classical MHC member, CD1. The best studied subset of lipid-reactive T cells are type I natural killer T (iNKT) cells that recognize a variety of different antigens when presented by the non-classical MHCI homolog CD1d. iNKT cells have been shown to be important for the protection against various microbial pathogens, including B. burgdorferi, the causative agents of Lyme disease, and S. pneumoniae, which causes pneumococcal meningitis and community-acquired pneumonia. Both pathogens carry microbial glycolipids that can trigger the T cell antigen receptor (TCR), leading to iNKT cell activation. iNKT cells have an evolutionary conserved TCR alpha chain, yet retain the ability to recognize structurally diverse glycolipids. They do so using a conserved recognition mode, in which the TCR enforces a conserved binding orientation on CD1d. TCR binding is accompanied by structural changes within the TCR binding site of CD1d, as well as the glycolipid antigen itself. In addition to direct recognition of microbial antigens, iNKT cells can also be activated by a combination of cytokines (IL-12/IL-18) and TCR stimulation. Many microbes carry TLR antigens, and microbial infections can lead to TLR activation. The subsequent cytokine response in turn lower the threshold of TCR-mediated iNKT cell activation, especially when weak microbial or even self-antigens are presented during the cause of the infection. In summary, iNKT cells can be directly activated through TCR triggering of strong antigens, while cytokines produced by the innate immune response may be necessary for TCR triggering and iNKT cell activation in the presence of weak antigens. Here, we will review the molecular basis of iNKT cell

  17. Effects of OK-432 on murine bone marrow and the production of natural killer cells

    SciTech Connect

    Pollack, S.B.; Rosse, C.

    1985-01-01

    The streptococcal preparation, OK-432, which augments anti-tumor responses in humans and mice, has been shown to be a potent immunomodulator. Among its effects is a pronounced augmentation of natural killer (NK) activity. The hypothesis that OK-432 alters the rates of production and maturation of NK cells in the bone marrow was tested. Studies to determine the kinetic parameters of NK cell production in normal C57BL/6J mice using tritiated thymidine, /sup 3/H-TdR, as a DNA marker are described. We are now extending those studies to determine the effect of OK-432 on the bone marrow and on the production of NK cells in the marrow. Initial observations are reported which indicate that OK-432 has profound effects on the cellularity and mitotic activity of the bone marrow, and in particular, on cells with the characteristics of natural killer cells within the marrow. 17 refs., 3 figs., 4 tabs.

  18. Natural Killer Cell Diversity in Viral Infection: Why and How Much?

    PubMed Central

    Blish, Catherine A.

    2016-01-01

    Natural killer cells are a diverse group of innate lymphocytes that are specialized to rapidly respond to cancerous or virus-infected cells. NK cell function is controlled by the integration of signals from activating and inhibitory receptors expressed at the cell surface. Variegated expression patterns of these activating and inhibitory receptors at the single cell level leads to a highly diverse NK cell repertoire. Here I review the factors that influence NK cell repertoire diversity and its functional consequences for our ability to fight viruses. PMID:27635417

  19. Crude extract of Polygonum cuspidatum stimulates immune responses in normal mice by increasing the percentage of Mac-3-positive cells and enhancing macrophage phagocytic activity and natural killer cell cytotoxicity

    PubMed Central

    CHUEH, FU-SHIN; LIN, JEN-JYH; LIN, JU-HWA; WENG, SHU-WEN; HUANG, YI-PING; CHUNG, JING-GUNG

    2015-01-01

    Polygonum cuspidatum is a natural plant that is used in traditional Chinese herbal medicine. The crude extract of Polygonum cuspidatum (CEPC) has numerous biological effects; however, there is a lack of studies on the effects of CEPC on immune responses in normal mice. The aim of the present study was to determine the in vivo effects of CEPC on immune responses in normal mice. CEPC (0, 50, 100, 150 and 200 mg/kg) was orally administered to BALB/c mice for three weeks, following which blood, liver, and spleen samples were collected. CEPC did not significantly affect the total body weight, or tissue weights of the liver or spleen, as compared with the control mice. CEPC increased the percentages of CD3 (T-cell marker), 11b (monocytes) and Mac-3 (macrophages) positive-cells, and reduced the percentage of CD19-positive cells (B-cell marker), as compared with the control mice. CEPC (100 mg/kg) stimulated macrophage phagocytosis of blood samples but did not affect macrophage phagocytosis in the peritoneum. Activity of the splenic natural killer cells was increased in response to CEPC (50 mg/kg) treatment. Furthermore, CEPC inhibited T- and B-cell proliferation when the cells were stimulated with concanavalin A and lipopolysaccharide, respectively. PMID:25338846

  20. Crude extract of Polygonum cuspidatum stimulates immune responses in normal mice by increasing the percentage of Mac-3-positive cells and enhancing macrophage phagocytic activity and natural killer cell cytotoxicity.

    PubMed

    Chueh, Fu-Shin; Lin, Jen-Jyh; Lin, Ju-Hwa; Weng, Shu-Wen; Huang, Yi-Ping; Chung, Jing-Gung

    2015-01-01

    Polygonum cuspidatum is a natural plant that is used in traditional Chinese herbal medicine. The crude extract of Polygonum cuspidatum (CEPC) has numerous biological effects; however, there is a lack of studies on the effects of CEPC on immune responses in normal mice. The aim of the present study was to determine the in vivo effects of CEPC on immune responses in normal mice. CEPC (0, 50, 100, 150 and 200 mg/kg) was orally administered to BALB/c mice for three weeks, following which blood, liver, and spleen samples were collected. CEPC did not significantly affect the total body weight, or tissue weights of the liver or spleen, as compared with the control mice. CEPC increased the percentages of CD3 (T-cell marker), 11b (monocytes) and Mac-3 (macrophages) positive-cells, and reduced the percentage of CD19-positive cells (B-cell marker), as compared with the control mice. CEPC (100 mg/kg) stimulated macrophage phagocytosis of blood samples but did not affect macrophage phagocytosis in the peritoneum. Activity of the splenic natural killer cells was increased in response to CEPC (50 mg/kg) treatment. Furthermore, CEPC inhibited T- and B-cell proliferation when the cells were stimulated with concanavalin A and lipopolysaccharide, respectively. PMID:25338846

  1. Effect of chaetocin on renal cell carcinoma cells and cytokine-induced killer cells

    PubMed Central

    Rombo, Roman; Weiher, Hans; Schmidt-Wolf, Ingo G.H.

    2016-01-01

    We examined the cytotoxic effects of chaetocin on clear cell renal cell carcinoma (ccRCC) cells and the possibility to combine the effects of chaetocin with the effects of cytokine-induced killer cells (CIK) assayed by MTT assay and FACS analysis. Chaetocin is a thiodioxopiperazine produced by fungi belonging to the chaetomiaceae family. In 2007, it was first reported that chaetocin shows potent and selective ex vivo anti-cancer activity by inducing reactive oxygen species. CIK cells are generated from CD3+/CD56- T lymphocytes with double negative CD4-/CD8- phenotype that are isolated from human blood. The addition of distinct interleukins and antibodies results in the generation of CIK cells that are able to specifically target and destroy renal carcinoma cells. The results of this research state that the anti-ccRCC activity of chaetocin is weak and does not show a high grade of selectivity on clear cell renal cell carcinoma cells. Although the CIK cells show a high grade of selective anti-ccRCC activity, this effect could not be improved by the addition of chaetocin. So chaetocin seems to be no suitable agent for specific targeting ccRCC cells or for the combination therapy with CIK cells in renal cancer. PMID:27141211

  2. Interleukin-15-transferred cytokine-induced killer cells elevated anti-tumor activity in a gastric tumor-bearing nude mice model.

    PubMed

    Peng, Zheng; Liang, Wentao; Li, Zexue; Xu, Yingxin; Chen, Lin

    2016-02-01

    Gastric cancer is the second leading cause of cancer-related mortality worldwide. Adoptive cell therapy (ACT) for gastric cancer is a novel therapy modality. However, the therapeutic effectiveness in vivo is still limited. The objective of this study was to assess the value of interleukin-15 (IL-15)-transferred cytokine-induced killer (CIK) cells in ACT for gastric cancer. IL-15-IRES-TK retroviral vector was constructed and transferred into the CIK cells. A gastric tumor-bearing nude mice model was constructed by subcutaneously injecting gastric cancer cells, BGC-823. Gastric tumor-bearing nude mice were randomly divided into three groups (five mice each group) and injected with physiological saline, CIK cells, and IL-15-IRES-TK-transfected CIK cells for 2 weeks, respectively. IL-15-IRES-TK-transferred CIK cells were prepared successfully and flow cytometry (FCM) analysis indicated that the transfection rate reached 85.7% after 5 days culture. In vivo experiment, we found that CIK cells retarded tumor growth by reducing tumor volume and tumor weight, as well as increasing tumor inhibition rate. Furthermore, IL-15-IRES-TK-transferred CIK cells showed a much stronger inhibition on tumor growth than CIK cells alone. Tumor morphology observation and growth indexes also showed that IL-15-transfected CIK cells had stronger cytotoxicity to tumor tissue than CIK cells. IL-15-IRES-TK transfection could elevate the effects of CIK cells to gastric carcinoma. The engineered CIK cells carrying IL-15-IRES-TK may be used in the ACT for gastric carcinoma, but prudent clinical trial is still indispensable.

  3. Asbestos fibres inhibit the in vitro activity of lymphokine-activated killer (LAK) cells from healthy individuals and patients with malignant mesothelioma.

    PubMed Central

    Manning, L S; Davis, M R; Robinson, B W

    1991-01-01

    Asbestos exposure is associated with an increased incidence of several malignancies, including malignant mesothelioma (MM). This study evaluates the relationship between asbestos exposure and the in vitro generation and function of LAK cells, an immune effector cell population with powerful lytic activity against MM cells. Both serpentine (chrysotile) and amphibole (amosite and crocidolite) forms of asbestos fibres suppress LAK cell generation, viability (by 5-11%, P less than 0.02) and cell recovery (by 13-15%, P less than 0.02). However, the LAK cells generated in the presence of the amphiboles were as effective as unexposed cells in lysing both standard tumour cell targets (K562, 56.4% lysis versus 61.5%, respectively, P greater than 0.5; NS; Daudi, 60.5% lysis versus 64.5% P greater than 0.5; NS), and MM tumour cell targets (mean of three MM cell lines 48.3% versus 46.3%, P greater than 0.5; NS), whereas the function of LAK cells generated in the presence of chrysotile was significantly reduced against three out of the five tumour cell targets tested (P less than 0.03). In the presence of asbestos fibres, LAK cell function was reduced against all five tumour cell targets (P less than 0.01), irrespective of whether the cell donors were healthy individuals or patients with MM. NK cell activity was also suppressed (P less than 0.01). The serpentine form of asbestos, chrysotile, was significantly more suppressive of both effector cell functions than either of the amphiboles (P less than 0.01). These findings suggest that asbestos exposure may suppress the function and in some instances the generation of immune effector cell mechanisms, thereby increasing the risk of disease and malignancy. PMID:1846329

  4. IMPAIRED NATURAL KILLER CELL LYSIS IN BREAST CANCER PATIENTS WITH HIGH LEVELS OF PSYCHOLOGICAL STRESS IS ASSOCIATED WITH ALTERED EXPRESSION OF KILLER IMMUNOGLOBULIN-LIKE RECEPTORS

    PubMed Central

    Varker, Kimberly A.; Terrell, Catherine E.; Welt, Marilyn; Suleiman, Samer; Thornton, Lisa; Andersen, Barbara L.; Carson, William E.

    2007-01-01

    Background We previously reported that cancer-related psychological stress is associated with reduced natural killer (NK) cell lysis. We hypothesized that reduced NK cell cytotoxicity in patients with increased levels of stress would correlate with alterations in the expression of inhibitory NK cell receptors (killer immunoglobulin-like receptors, or KIRs). The specific aim of this study was to examine KIR expression in patients with high or low levels of psychologic stress and correlate alterations in KIR expression with NK cell function. Materials and Methods 227 patients underwent baseline evaluation of cancer-related psychological stress and were randomized to psychosocial intervention versus observation. From this population, two groups were defined based on pre-treatment measurements of NK lytic activity, stress levels, and the availability of cryopreserved peripheral blood mononuclear cells (PBMC). Group I (n = 9) had low stress by the Impact of Events Scale (IES), and high NK cell lysis at the 50:1 effector: target ratio (NK50 = 52–89%). Group II (n = 8) had high stress and low NK50 (27–52%). Lymphokine activated killer (LAK) activity, antibody dependent cellular cytotoxicity (ADCC), and expression of cytokine receptors, adhesion molecules, and killer immunoglobulin-like receptors (KIRs) were assessed in PBMC. Results Incubation of PBMC with NK-stimulatory cytokines (IL-2, IL-12, or IL-15) led to significant increases in cytotoxic activity regardless of IES/NK50 scores. There were no significant group differences in NK cell surface expression of the IL-2 receptor components CD25 and CD122, antibody-dependent lysis of HER2/neu-positive SKBr3 cells treated with an anti-HER2/neu monoclonal antibody, expression of adhesion molecules (CD2, CD11a, CD18) and markers of activation (CD69), or expression of the KIRs CD158a, NKG2a, NKB1, and CD161. However, levels of CD158b were significantly higher in Group I after incubation in media alone or with IL-2, and CD94

  5. The occurrence of killer activity in yeasts isolated from natural habitats.

    PubMed

    Wójcik, Monika; Kordowska-Wiater, Monika

    2015-01-01

    Yeast's ability to restrict the growth and kill other yeasts, fungi and bacteria has been known for over 50 years. Killer activity was detected in yeasts deposited in the world collections or isolated from natural habitats. In this study, isolates from the forest environment, leaves of fruit trees, flower petals, cereals and frozen fruit have been screened in terms of their killer activities. Killer activity was tested on strains belonging to six yeast species: Candida, Rhodotorula, Pichia, Pachysolen, Yarrowia, Trichosporon. The reference strains were Kluyveromyces lactis Y-6682 and Kluyveromyces marxinanus Y-8281, well-known to be sensitive to yeast killer toxins. Among one hundred and two tested strains, 24 (23.5% of isolates) showed positive killer action, and 10 (9.8% of the isolates) a weak killer action against at least one sensitive reference strain. The highest killer activity was observed among isolates from forest soil and flowers. PMID:26636138

  6. Activation of human T-helper/inducer cell, T-cytotoxic/suppressor cell, B-cell, and natural killer (NK)-cells and induction of NK cell activity against K562 chronic myeloid leukemia cells with modified citrus pectin

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background Modified citrus pectin (MCP) is known for its anti-cancer effects and its ability to be absorbed and circulated in the human body. In this report we tested the ability of MCP to induce the activation of human blood lymphocyte subsets including T-helper/inducer cell, Tcytotoxic/suppres...

  7. Emotional stability, anxiety, and natural killer activity under examination stress.

    PubMed

    Borella, P; Bargellini, A; Rovesti, S; Pinelli, M; Vivoli, R; Solfrini, V; Vivoli, G

    1999-08-01

    This study was performed to evaluate the relation between a stable personality trait, a mood state and immune response to an examination stress. A self-reported measure of emotional stability (BFQ-ES scale) was obtained in a sample (n = 39) randomly selected from 277 cadets; this personality trait was also investigated by completing a neuroticism scale (Eysenck personality inventory) and a trait-anxiety scale (STAI). Natural killer (NK) cell activity was measured at baseline, long before the examination time and the examination day. The state-anxiety scale evaluated the response to the stressful stimulus. Taking subjects all together, the academic task did not result in significant modification over baseline in NK cell activity. Subjects were then divided into three groups based on emotional stability and state-anxiety scores: high emotional stability/low anxiety, medium, and low emotional stability/high anxiety. Examination stress induced significant increases in NK cell activity in the high emotional stability/low anxiety group, no effect in the medium group, and significant decreases in the low emotional stability/high anxiety group. The repeated-measure ANOVA revealed a significant interaction of group x period (baseline vs. examination) for both lytic units and percent cytolysis. The results did not change after introducing coffee and smoking habits as covariates. Our findings suggest that the state-anxiety acts in concert with a stable personality trait to modulate NK response in healthy subjects exposed to a psychological naturalistic stress. The relation between anxiety and poor immune control has been already described, whereas the ability of emotional stability to associate with an immunoenhancement has not yet reported. The peculiarity of our population, a very homogeneous and healthy group for life style and habits, can have highlighted the role of emotional stability, and may account for the difference with other studies.

  8. Suppression of natural killer cell activity by rabbit antibody to human beta 2-microglobulin (beta 2m) is an Fc-mediated phenomenon and is not beta 2m specific.

    PubMed

    Jones, R A; Richards, S J; Patel, D; Scott, C S

    1991-07-01

    Natural killer (NK) cell cytotoxicity constitutes an important component of the host immune defence system. The NK effector cell has been relatively well defined in terms of immunophenotypic characteristics, but in contrast to the functional T-cell receptor molecule associated with major histocompatibility complex (MHC)-restricted cytotoxic activity, the NK cell receptor has not to date been defined. However, several studies have suggested that the beta 2-microglobulin (beta 2m) molecule is functionally associated with NK cell activity. Using various heterospecific and monoclonal antibodies, this study has shown that intact rabbit IgG antibody bound either directly or indirectly to peripheral mononuclear cell (PMNC) effector populations significantly reduced their lytic activity against K562 targets. Substitution of F(ab)2 fragments for rabbit IgG antibodies, or the use of monoclonal antibodies alone, failed to reduce peripheral blood mononuclear cell (PMNC) lytic activity. Addition of non-NK cell components labelled with rabbit anti-beta 2m to purified NK-enriched effector cell populations also suppressed K562 lysis. In contrast, pre-treatment of a NK-enriched PMNC fraction with rabbit anti-beta 2m enhanced target lysis. These results strongly suggest that antibody-induced suppression of PMNC NK activity is mediated via rabbit Fc attached to co-existing non-NK cells in the mononuclear fraction, and are inconsistent with the previously suggested functional association between NK activity and membrane beta 2m.

  9. Studies of lymphokine-activated killer (LAK) cells. I. Evidence using novel monoclonal antibodies that most human LAK precursor cells share a common surface marker

    PubMed Central

    1989-01-01

    Separation of LAK precursor (LAKp) cells (as defined by LAK effector generation after incubation with IL-2 for 7 d) from cells with NK activity/LGL morphology was achieved on Percoll gradients using a longer, slower centrifugation than that used for optimal NK enrichment. mAb were generated using the various Percoll fractions as the immunizing cells and used for separation and depletion studies. Two mAbs DM-1 (IgM,k) and DM-2 (IgM,k) recognizing 2-15% and 15-30% of PBL, respectively, abrogated a large proportion of LAK generative potential after complement depletion, but had little effect on NK or LAK effector activity. Cell sorting experiments indicated that the majority of LAKp cells are found within the DM-1+ population and that DM-1+ cells are not simply an accessory cell required for LAKp generation. Further, these two mAbs do not recognize cells that are responsible for generating cytotoxicity during MLC or co-culture with the PR-1 EBV lymphoblastoid cell line. Western blot analysis indicated that DM-1 and DM-2 recognize a 38,000 and 44,000 dalton moiety, respectively. The frequency of cells bearing these antigens and the intensity of cell surface staining decreased during the 7-d culture period, suggesting that these antibodies recognize determinants found only at the precursor level. These findings indicate that cells other than NK effectors or mature T cells are capable of generating a LAK cell response. These LAK precursor cells share a common differentiation surface antigen and are different from AK or antigen-specific CTL precursors. The possibility exists that these cells are identical to, or include, the NK precursor cell. PMID:2784480

  10. Induction of BCL2-Interacting Killer, BIK, is Mediated for Anti-Cancer Activity of Curcumin in Human Head and Neck Squamous Cell Carcinoma Cells

    PubMed Central

    Xi, Yue; Gao, Hang; Callaghan, Michael U.; Fribley, Andrew M.; Garshott, Danielle M.; Xu, Zhi-Xiang; Zeng, Qinghua; Li, Yu-Lin

    2015-01-01

    Naturally occurring diarylheptanoid curcumin (CUR), a principal component of the Asian spice turmeric, has been shown to have anti-cancer effects in many tumor types. However, a detailed mechanism regarding CUR induced tumor cell killing remain to be comprehensively explored. Using two head neck squamous cell carcinoma (HNSCC) cell lines FaDu (hypopharyngeal) and Cal27 (tongue), we demonstrated a novel mechanism by which CUR levies the cytotoxic effect. We found that CUR induced upregulation of pro-apoptotic Bik, down-regulation of survival signaling by AKT and NF-κB prior to the induction of the caspase-cascade reduction of cell proliferation, are primary mechanisms of CUR-induced cell death, thus providing insights into the anti-tumor activity of CUR in HNSCC cells. PMID:25767602

  11. Influence of malignant cell clonogenic capacities and position along the maturation pathway on their susceptibility to lymphokine-activated killer cell cytotoxicity.

    PubMed

    Thomas, X; Anglaret, B; Adeleine, P; Maritaz, O; Bailly, M; Fiere, D; Archimbaud, E

    1998-01-01

    In order to investigate the sensitivity of malignant target cells to lysis by LAK cells according to their clonogenic capacities and their position along the maturation pathway, we compared clonogenic and chromium release cytotoxicity assays performed on human hematopoietic cell lines using Effector: Target ratios of 1:1, 3:1, 6:1, 12:1, 24:1, 48:1 and 96:1, and studied the sensitivity of HL-60 and U937 human cell lines after exposure to different factors including GM-CSF, SCF, IFN, Retinoic acid (RA), DMSO, and TPA which are able to recruit cells into the cell cycle or to induce cell differentiation. There was a good correlation between the lysis of the target cells using 51Cr release and the growth inhibition in semisolid medium. The degree of inhibition was significantly higher using the colony growth assay (p = 0.006). Regarding the effects of culturing cell lines with proliferating and differentiating agents on the sensitivity of these cell lines to LAK cytolysis, a correlation was noted between the proliferative response of the U937 cell line and susceptibility to LAK cell lysis (p = 0.01), while results appeared close to significance with HL-60. The most significant effects were a decreased sensitivity of HL-60 to LAK lysis with RA (p < 0.001) and TPA (p < 0.001), and an increased susceptibility of U937 to LAK lysis with GM-CSF (p < 0.0001). In studies planned to investigate whether susceptibility of treated cells to LAK activity was a consequence of a downregulation of adhesion molecules expressed on target cell surface, the proportion of cells expressing adhesion molecules was not significantly changed, except for CD54 expression on HL-60 cells which showed a higher expression, after cells were treated with RA or DMSO. We conclude that clonogenic cells are more sensitive to LAK cell lysis and that cell line sensitivity to LAK cytolysis can be modulated by a variety of agents of potential therapeutic use. The poor correlation between adhesion molecules

  12. Skin TLR7 triggering promotes accumulation of respiratory dendritic cells and natural killer cells.

    PubMed

    Hackstein, Holger; Hagel, Nicole; Knoche, Angela; Kranz, Sabine; Lohmeyer, Jürgen; von Wulffen, Werner; Kershaw, Olivia; Gruber, Achim D; Bein, Gregor; Baal, Nelli

    2012-01-01

    The TLR7 agonist imiquimod has been used successfully as adjuvant for skin treatment of virus-associated warts and basal cell carcinoma. The effects of skin TLR7 triggering on respiratory leukocyte populations are unknown. In a placebo-controlled experimental animal study we have used multicolour flow cytometry to systematically analyze the modulation of respiratory leukocyte subsets after skin administration of imiquimod. Compared to placebo, skin administration of imiquimod significantly increased respiratory dendritic cells (DC) and natural killer cells, whereas total respiratory leukocyte, alveolar macrophages, classical CD4+ T helper and CD8+ T killer cell numbers were not or only moderately affected. DC subpopulation analyses revealed that elevation of respiratory DC was caused by an increase of respiratory monocytic DC and CD11b(hi) DC subsets. Lymphocyte subpopulation analyses indicated a marked elevation of respiratory natural killer cells and a significant reduction of B lymphocytes. Analysis of cytokine responses of respiratory leukocytes after stimulation with Klebsiella pneumonia indicated reduced IFN-γ and TNF-α expression and increased IL-10 and IL-12p70 production after 7 day low dose skin TLR7 triggering. Additionally, respiratory NK cytotoxic activity was increased after 7d skin TLR7 triggering. In contrast, lung histology and bronchoalveolar cell counts were not affected suggesting that skin TLR7 stimulation modulated respiratory leukocyte composition without inducing overt pulmonary inflammation. These data suggest the possibility to modulate respiratory leukocyte composition and respiratory cytokine responses against pathogens like Klebsiella pneumonia through skin administration of a clinically approved TLR7 ligand. Skin administration of synthetic TLR7 ligands may represent a novel, noninvasive means to modulate respiratory immunity. PMID:22927956

  13. Skin TLR7 Triggering Promotes Accumulation of Respiratory Dendritic Cells and Natural Killer Cells

    PubMed Central

    Hackstein, Holger; Hagel, Nicole; Knoche, Angela; Kranz, Sabine; Lohmeyer, Jürgen; von Wulffen, Werner; Kershaw, Olivia; Gruber, Achim D.; Bein, Gregor; Baal, Nelli

    2012-01-01

    The TLR7 agonist imiquimod has been used successfully as adjuvant for skin treatment of virus-associated warts and basal cell carcinoma. The effects of skin TLR7 triggering on respiratory leukocyte populations are unknown. In a placebo-controlled experimental animal study we have used multicolour flow cytometry to systematically analyze the modulation of respiratory leukocyte subsets after skin administration of imiquimod. Compared to placebo, skin administration of imiquimod significantly increased respiratory dendritic cells (DC) and natural killer cells, whereas total respiratory leukocyte, alveolar macrophages, classical CD4+ T helper and CD8+ T killer cell numbers were not or only moderately affected. DC subpopulation analyses revealed that elevation of respiratory DC was caused by an increase of respiratory monocytic DC and CD11bhi DC subsets. Lymphocyte subpopulation analyses indicated a marked elevation of respiratory natural killer cells and a significant reduction of B lymphocytes. Analysis of cytokine responses of respiratory leukocytes after stimulation with Klebsiella pneumonia indicated reduced IFN-γ and TNF-α expression and increased IL-10 and IL-12p70 production after 7 day low dose skin TLR7 triggering. Additionally, respiratory NK cytotoxic activity was increased after 7d skin TLR7 triggering. In contrast, lung histology and bronchoalveolar cell counts were not affected suggesting that skin TLR7 stimulation modulated respiratory leukocyte composition without inducing overt pulmonary inflammation. These data suggest the possibility to modulate respiratory leukocyte composition and respiratory cytokine responses against pathogens like Klebsiella pneumonia through skin administration of a clinically approved TLR7 ligand. Skin administration of synthetic TLR7 ligands may represent a novel, noninvasive means to modulate respiratory immunity. PMID:22927956

  14. Membrane nanotubes facilitate long-distance interactions between natural killer cells and target cells.

    PubMed

    Chauveau, Anne; Aucher, Anne; Eissmann, Philipp; Vivier, Eric; Davis, Daniel M

    2010-03-23

    Membrane nanotubes are membranous tethers that physically link cell bodies over long distances. Here, we present evidence that nanotubes allow human natural killer (NK) cells to interact functionally with target cells over long distances. Nanotubes were formed when NK cells contacted target cells and moved apart. The frequency of nanotube formation was dependent on the number of receptor/ligand interactions and increased on NK cell activation. Most importantly, NK cell nanotubes contained a submicron scale junction where proteins accumulated, including DAP10, the signaling adaptor that associates with the activating receptor NKG2D, and MHC class I chain-related protein A (MICA), a cognate ligand for NKG2D, as occurs at close intercellular synapses between NK cells and target cells. Quantitative live-cell fluorescence imaging suggested that MICA accumulated at small nanotube synapses in sufficient numbers to trigger cell activation. In addition, tyrosine-phosphorylated proteins and Vav-1 accumulated at such junctions. Functionally, nanotubes could aid the lysis of distant target cells either directly or by moving target cells along the nanotube path into close contact for lysis via a conventional immune synapse. Target cells moving along the nanotube path were commonly polarized such that their uropods faced the direction of movement. This is the opposite polarization than for normal cell migration, implying that nanotubes can specifically drive target cell movement. Finally, target cells that remained connected to an NK cell by a nanotube were frequently lysed, whereas removing the nanotube using a micromanipulator reduced lysis of these target cells.

  15. Molecular identification and expression analysis of a natural killer cell enhancing factor (NKEF) from rock bream Oplegnathus fasciatus and the biological activity of its recombinant protein

    PubMed Central

    Kim, Ju-Won; Choi, Hye-Sung; Kwon, Mun-Gyeong; Park, Myoung-Ae; Hwang, Jee-Youn; Kim, Do-Hyung; Park, Chan-Il

    2011-01-01

    Natural killer cell enhancing factor (NKEF) belongs to the defined peroxiredoxin (Prx) family. Rock bream NKEF cDNA was identified by expressed sequence tag (EST) analysis of rock bream liver that was stimulated with the LPS. The full-length RbNKEF cDNA (1062 bp) contained an open reading frame (ORF) of 594 bp encoding 198 amino acids. RbNKEF was significantly expressed in the gill, liver, and intestine. mRNA expression of NKEF in the head kidney was examined under viral and bacterial challenge via real-time RT-PCR. Experimental challenge of rock bream with Edwardsiella tarda, Streptococcus iniae, and RSIV resulted in significant increases in RbNKEF mRNA in the head kidney. To obtain a recombinant NKEF, the RbNKEF ORF was expressed in Escherichia coli BL21 (DE3), and the purified soluble protein exhibited a single band corresponding to the predicted molecular mass. When kidney leucocytes were treated with a high concentration of rRbNKEF (10 μg/mL), they exhibited significantly enhanced cell proliferation and viability under oxidative stress. PMID:24371552

  16. Estrogen Therapy Delays Autoimmune Diabetes and Promotes the Protective Efficiency of Natural Killer T-Cell Activation in Female Nonobese Diabetic Mice.

    PubMed

    Gourdy, Pierre; Bourgeois, Elvire A; Levescot, Anaïs; Pham, Linh; Riant, Elodie; Ahui, Marie-Louise; Damotte, Diane; Gombert, Jean-Marc; Bayard, Francis; Ohlsson, Claes; Arnal, Jean-François; Herbelin, André

    2016-01-01

    Therapeutic strategies focused on restoring immune tolerance remain the main avenue to prevent type 1 diabetes (T1D). Because estrogens potentiate FoxP3+ regulatory T cells (Treg) and invariant natural killer T (iNKT) cells, two regulatory lymphocyte populations that are functionally deficient in nonobese diabetic (NOD) mice, we investigated whether estradiol (E2) therapy influences the course of T1D in this model. To this end, female NOD mice were sc implanted with E2- or placebo-delivering pellets to explore the course of spontaneous and cyclophosphamide-induced diabetes. Treg-depleted and iNKT-cell-deficient (Jα18(-/-)) NOD mice were used to assess the respective involvement of these lymphocyte populations in E2 effects. Early E2 administration (from 4 wk of age) was found to preserve NOD mice from both spontaneous and cyclophosphamide-induced diabetes, and a complete protection was also observed throughout treatment when E2 treatment was initiated after the onset of insulitis (from 12 wk of age). This delayed E2 treatment remained fully effective in Treg-depleted mice but failed to entirely protect Jα18(-/-) mice. Accordingly, E2 administration was shown to restore the cytokine production of iNKT cells in response to in vivo challenge with the cognate ligand α-galactosylceramide. Finally, transient E2 administration potentiated the previously described protective action of α-galactosylceramide treatment in NOD females. This study provides original evidence that E2 therapy strongly protects NOD mice from T1D and reveals the estrogen/iNKT cell axis as a new effective target to counteract diabetes onset at the stage of insulitis. Estrogen-based therapy should thus be considered for T1D prevention. PMID:26485613

  17. Estrogen Therapy Delays Autoimmune Diabetes and Promotes the Protective Efficiency of Natural Killer T-Cell Activation in Female Nonobese Diabetic Mice.

    PubMed

    Gourdy, Pierre; Bourgeois, Elvire A; Levescot, Anaïs; Pham, Linh; Riant, Elodie; Ahui, Marie-Louise; Damotte, Diane; Gombert, Jean-Marc; Bayard, Francis; Ohlsson, Claes; Arnal, Jean-François; Herbelin, André

    2016-01-01

    Therapeutic strategies focused on restoring immune tolerance remain the main avenue to prevent type 1 diabetes (T1D). Because estrogens potentiate FoxP3+ regulatory T cells (Treg) and invariant natural killer T (iNKT) cells, two regulatory lymphocyte populations that are functionally deficient in nonobese diabetic (NOD) mice, we investigated whether estradiol (E2) therapy influences the course of T1D in this model. To this end, female NOD mice were sc implanted with E2- or placebo-delivering pellets to explore the course of spontaneous and cyclophosphamide-induced diabetes. Treg-depleted and iNKT-cell-deficient (Jα18(-/-)) NOD mice were used to assess the respective involvement of these lymphocyte populations in E2 effects. Early E2 administration (from 4 wk of age) was found to preserve NOD mice from both spontaneous and cyclophosphamide-induced diabetes, and a complete protection was also observed throughout treatment when E2 treatment was initiated after the onset of insulitis (from 12 wk of age). This delayed E2 treatment remained fully effective in Treg-depleted mice but failed to entirely protect Jα18(-/-) mice. Accordingly, E2 administration was shown to restore the cytokine production of iNKT cells in response to in vivo challenge with the cognate ligand α-galactosylceramide. Finally, transient E2 administration potentiated the previously described protective action of α-galactosylceramide treatment in NOD females. This study provides original evidence that E2 therapy strongly protects NOD mice from T1D and reveals the estrogen/iNKT cell axis as a new effective target to counteract diabetes onset at the stage of insulitis. Estrogen-based therapy should thus be considered for T1D prevention.

  18. Natural killer cells and regulatory T cells in early pregnancy loss

    PubMed Central

    SHARMA, SURENDRA

    2015-01-01

    Survival of the allogeneic embryo in the uterus depends on the maintenance of immune tolerance at the maternal-fetal interface. The pregnant uterus is replete with activated maternal immune cells. How this immune tolerance is acquired and maintained has been a topic of intense investigation. The key immune cells that predominantly populate the pregnant uterus are natural killer (NK) cells. In normal pregnancy, these cells are not killers, but rather provide a microenvironment that is pregnancy compatible and supports healthy placentation. In placental mammals, an array of highly orchestrated immune elements to support successful pregnancy outcome has been incorporated. This includes active cooperation between maternal immune cells, particularly NK cells, and trophoblast cells. This intricate process is required for placentation, immune regulation and to remodel the blood supply to the fetus. During the past decade, various types of maternal immune cells have been thought to be involved in cross-talk with trophoblasts and in programming immune tolerance. RegulatoryT cells (Tregs) have attracted a great deal of attention in promoting implantation and immune tolerance beyond implantation. However, what has not been fully addressed is how this immune-trophoblast axis breaks down during adverse pregnancy outcomes, particularly early pregnancy loss, and in response to unscheduled inflammation. Intense research efforts have begun to shed light on the roles of NK cells and Tregs in early pregnancy loss, although much remains to be unraveled in order to fully characterize the mechanisms underlying their detrimental activity. An increased understanding of host-environment interactions that lead to the cytotoxic phenotype of these otherwise pregnancy compatible maternal immune cells is important for prediction, prevention and treatment of pregnancy maladies, particularly recurrent pregnancy loss. In this review, we discuss relevant information from experimental and human

  19. Natural killer cells and regulatory T cells in early pregnancy loss.

    PubMed

    Sharma, Surendra

    2014-01-01

    Survival of the allogeneic embryo in the uterus depends on the maintenance of immune tolerance at the maternal-fetal interface. The pregnant uterus is replete with activated maternal immune cells. How this immune tolerance is acquired and maintained has been a topic of intense investigation. The key immune cells that predominantly populate the pregnant uterus are natural killer (NK) cells. In normal pregnancy, these cells are not killers, but rather provide a microenvironment that is pregnancy compatible and supports healthy placentation. In placental mammals, an array of highly orchestrated immune elements to support successful pregnancy outcome has been incorporated. This includes active cooperation between maternal immune cells, particularly NK cells, and trophoblast cells. This intricate process is required for placentation, immune regulation and to remodel the blood supply to the fetus. During the past decade, various types of maternal immune cells have been thought to be involved in cross-talk with trophoblasts and in programming immune tolerance. Regulatory T cells (Tregs) have attracted a great deal of attention in promoting implantation and immune tolerance beyond implantation. However, what has not been fully addressed is how this immune-trophoblast axis breaks down during adverse pregnancy outcomes, particularly early pregnancy loss, and in response to unscheduled inflammation. Intense research efforts have begun to shed light on the roles of NK cells and Tregs in early pregnancy loss, although much remains to be unraveled in order to fully characterize the mechanisms underlying their detrimental activity. An increased understanding of host-environment interactions that lead to the cytotoxic phenotype of these otherwise pregnancy compatible maternal immune cells is important for prediction, prevention and treatment of pregnancy maladies, particularly recurrent pregnancy loss. In this review, we discuss relevant information from experimental and human

  20. CAR-T cells are serial killers

    PubMed Central

    Davenport, Alexander J; Jenkins, Misty R; Ritchie, David S; Prince, H Miles; Trapani, Joseph A; Kershaw, Michael H; Darcy, Phillip K; Neeson, Paul J

    2015-01-01

    Chimeric antigen receptor (CAR) T cells have enjoyed unprecedented clinical success against haematological malignancies in recent years. However, several aspects of CAR T cell biology remain unknown. We recently compared CAR and T cell receptor (TCR)-based killing in the same effector cell and showed that CAR T cells can not only efficiently kill single tumor targets, they can also kill multiple tumor targets in a sequential manner. Single and serial killing events were not sustained long term due to CAR down-regulation after 20 hours. PMID:26587330

  1. Cord Blood as a Source of Natural Killer Cells.

    PubMed

    Mehta, Rohtesh S; Shpall, Elizabeth J; Rezvani, Katayoun

    2015-01-01

    Cord blood (CB) offers several unique advantages as a graft source for hematopoietic stem cell transplantation (HSCT). The risk of relapse and graft vs. host disease after cord blood transplantation (CBT) is lower than what is typically observed after other graft sources with a similar degree of human leukocyte antigen mismatch. Natural killer (NK) cells have a well-defined role in both innate and adaptive immunity and as the first lymphocytes to reconstitute after HSCT and CBT, and they play a significant role in protection against early relapse. In this article, we highlight the uses of CB NK cells in transplantation and adoptive immunotherapy. First, we will describe differences in the phenotype and functional characteristics of NK cells in CB as compared with peripheral blood. Then, we will review some of the obstacles we face in using resting CB NK cells for adoptive immunotherapy, and discuss methods to overcome them. We will review the current literature on killer-cell immunoglobulin-like receptors ligand mismatch and outcomes after CBT. Finally, we will touch on current strategies for the use of CB NK cells in cellular immunotherapy. PMID:26779484

  2. Carbamate pesticide-induced apoptosis and necrosis in human natural killer cells.

    PubMed

    Li, Q; Kobayashi, M; Kawada, T

    2014-01-01

    We previously found that ziram, a carbamate fungicide, significantly induced apoptosis and necrosis in human NK-92MI, a natural killer cell line. To investigate whether other carbamate pesticides also induce apoptosis and necrosis in human natural killer cell, we conducted further experiments with NK-92CI, a human natural killer cell line using a more sensitive assay. NK-92CI cells were treated with ziram, thiram, maneb or carbaryl at 0.031-40 microM for 2-24 h in the present study. Apoptosis and necrosis were determined by FITC-Annexin-V/PI staining. To explore the mechanism of apoptosis, intracellular levels of active caspases 3 and mitochondrial cytochrome-c release were determined by flow cytometry. We found that ziram and thiram also induced apoptosis and necrosis in a time- and dose-dependent manner; however, maneb and carbaryl induced apoptosis and necrosis only at higher doses in NK-92CI cells. The strength of the apoptosis-inducing effect differed among the pesticides, and the order was as follows: thiram > ziram greater than maneb greater than carbaryl. NK-92CI was more sensitive to ziram than NK-92MI. Moreover, ziram and thiram significantly increased the intracellular level of active caspase 3 in NK-92CI and caspase inhibitor significantly inhibited the apoptosis. Ziram and thiram significantly caused mitochondrial cytochrome-c release in NK-92CI. These findings indicate that carbamate pesticides can induce apoptosis in natural killer cells, and the apoptosis is mediated by both the caspase-cascade and mitochondrial cytochrome-c pathways.

  3. Lymphokine-activated killer cells targeted by monoclonal antibodies to the disialogangliosides GD2 and GD3 specifically lyse human tumor cells of neuroectodermal origin.

    PubMed

    Honsik, C J; Jung, G; Reisfeld, R A

    1986-10-01

    Monoclonal antibodies 14.18 (IgG3) and 11C64 (IgG3) directed against disialogangliosides GD2 and GD3, respectively, when used in conjunction with human peripheral blood mononuclear cells (PBMCs) stimulated with human recombinant interleukin (rIL-2) lyse both human melanoma and neuroblastoma cells by antibody-dependent cellular cytotoxicity. Such monoclonal antibody-"armed" effector cells are specifically directed to targets expressing the given disialoganglioside without detectable cross-reactivity. In addition, antibody-dependent cellular cytotoxicity as well as the natural killing ability of human PBMCs is augmented by a brief coincubation with rIL-2. PBMCs augmented by rIL-2 and armed with monoclonal antibodies significantly suppressed tumor growth in the xenotransplant nude mouse model. Our results suggest that once a threshold level of activation of PBMCs is achieved, additional rIL-2 (over three orders of magnitude of concentration) does not significantly enhance cytolytic augmentation. Furthermore, anti-GD3 monoclonal antibody 11C64 together with rIL-2-stimulated PBMCs from melanoma patients with widely differing tumor burdens effectively lyse melanoma tumor targets in antibody-dependent cellular cytotoxicity. Our results also suggest that GD2 and GD3 represent distinct and relevant immunotherapeutic target structures on melanoma whereas GD2 does the same for neuroblastoma tumors. Our data suggest that targeting of activated human effector cells may provide a new and effective cancer immunotherapy protocol.

  4. Lymphokine-activated killer cells targeted by monoclonal antibodies to the disialogangliosides GD2 and GD3 specifically lyse human tumor cells of neuroectodermal origin.

    PubMed Central

    Honsik, C J; Jung, G; Reisfeld, R A

    1986-01-01

    Monoclonal antibodies 14.18 (IgG3) and 11C64 (IgG3) directed against disialogangliosides GD2 and GD3, respectively, when used in conjunction with human peripheral blood mononuclear cells (PBMCs) stimulated with human recombinant interleukin (rIL-2) lyse both human melanoma and neuroblastoma cells by antibody-dependent cellular cytotoxicity. Such monoclonal antibody-"armed" effector cells are specifically directed to targets expressing the given disialoganglioside without detectable cross-reactivity. In addition, antibody-dependent cellular cytotoxicity as well as the natural killing ability of human PBMCs is augmented by a brief coincubation with rIL-2. PBMCs augmented by rIL-2 and armed with monoclonal antibodies significantly suppressed tumor growth in the xenotransplant nude mouse model. Our results suggest that once a threshold level of activation of PBMCs is achieved, additional rIL-2 (over three orders of magnitude of concentration) does not significantly enhance cytolytic augmentation. Furthermore, anti-GD3 monoclonal antibody 11C64 together with rIL-2-stimulated PBMCs from melanoma patients with widely differing tumor burdens effectively lyse melanoma tumor targets in antibody-dependent cellular cytotoxicity. Our results also suggest that GD2 and GD3 represent distinct and relevant immunotherapeutic target structures on melanoma whereas GD2 does the same for neuroblastoma tumors. Our data suggest that targeting of activated human effector cells may provide a new and effective cancer immunotherapy protocol. PMID:3094017

  5. Classification of human natural killer cells based on migration behavior and cytotoxic response.

    PubMed

    Vanherberghen, Bruno; Olofsson, Per E; Forslund, Elin; Sternberg-Simon, Michal; Khorshidi, Mohammad Ali; Pacouret, Simon; Guldevall, Karolin; Enqvist, Monika; Malmberg, Karl-Johan; Mehr, Ramit; Önfelt, Björn

    2013-02-21

    Despite intense scrutiny of the molecular interactions between natural killer (NK) and target cells, few studies have been devoted to dissection of the basic functional heterogeneity in individual NK cell behavior. Using a microchip-based, time-lapse imaging approach allowing the entire contact history of each NK cell to be recorded, in the present study, we were able to quantify how the cytotoxic response varied between individual NK cells. Strikingly, approximately half of the NK cells did not kill any target cells at all, whereas a minority of NK cells was responsible for a majority of the target cell deaths. These dynamic cytotoxicity data allowed categorization of NK cells into 5 distinct classes. A small but particularly active subclass of NK cells killed several target cells in a consecutive fashion. These "serial killers" delivered their lytic hits faster and induced faster target cell death than other NK cells. Fast, necrotic target cell death was correlated with the amount of perforin released by the NK cells. Our data are consistent with a model in which a small fraction of NK cells drives tumor elimination and inflammation.

  6. Heat Shock Protein 70 (Hsp70) Peptide Activated Natural Killer (NK) Cells for the Treatment of Patients with Non-Small Cell Lung Cancer (NSCLC) after Radiochemotherapy (RCTx) – From Preclinical Studies to a Clinical Phase II Trial

    PubMed Central

    Specht, Hanno M.; Ahrens, Norbert; Blankenstein, Christiane; Duell, Thomas; Fietkau, Rainer; Gaipl, Udo S.; Günther, Christine; Gunther, Sophie; Habl, Gregor; Hautmann, Hubert; Hautmann, Matthias; Huber, Rudolf Maria; Molls, Michael; Offner, Robert; Rödel, Claus; Rödel, Franz; Schütz, Martin; Combs, Stephanie E.; Multhoff, Gabriele

    2015-01-01

    Heat shock protein 70 (Hsp70) is frequently overexpressed in tumor cells. An unusual cell surface localization could be demonstrated on a large variety of solid tumors including lung, colorectal, breast, squamous cell carcinomas of the head and neck, prostate and pancreatic carcinomas, glioblastomas, sarcomas and hematological malignancies, but not on corresponding normal tissues. A membrane (m)Hsp70-positive phenotype can be determined either directly on single cell suspensions of tumor biopsies by flow cytometry using cmHsp70.1 monoclonal antibody or indirectly in the serum of patients using a novel lipHsp70 ELISA. A mHsp70-positive tumor phenotype has been associated with highly aggressive tumors, causing invasion and metastases and resistance to cell death. However, natural killer (NK), but not T cells were found to kill mHsp70-positive tumor cells after activation with a naturally occurring Hsp70 peptide (TKD) plus low dose IL-2 (TKD/IL-2). Safety and tolerability of ex vivo TKD/IL-2 stimulated, autologous NK cells has been demonstrated in patients with metastasized colorectal and non-small cell lung cancer (NSCLC) in a phase I clinical trial. Based on promising clinical results of the previous study, a phase II randomized clinical study was initiated in 2014. The primary objective of this multicenter proof-of-concept trial is to examine whether an adjuvant treatment of NSCLC patients after platinum-based radiochemotherapy (RCTx) with TKD/IL-2 activated, autologous NK cells is clinically effective. As a mHsp70-positive tumor phenotype is associated with poor clinical outcome only mHsp70-positive tumor patients will be recruited into the trial. The primary endpoint of this study will be the comparison of the progression-free survival of patients treated with ex vivo activated NK cells compared to patients who were treated with RCTx alone. As secondary endpoints overall survival, toxicity, quality-of-life, and biological responses will be determined in both

  7. Favorable impact of natural killer cell reconstitution on chronic graft-versus-host disease and cytomegalovirus reactivation after allogeneic hematopoietic stem cell transplantation.

    PubMed

    Kheav, Vissal David; Busson, Marc; Scieux, Catherine; Peffault de Latour, Régis; Maki, Guitta; Haas, Philippe; Mazeron, Marie-Christine; Carmagnat, Maryvonnick; Masson, Emeline; Xhaard, Aliénor; Robin, Marie; Ribaud, Patricia; Dulphy, Nicolas; Loiseau, Pascale; Charron, Dominique; Socié, Gérard; Toubert, Antoine; Moins-Teisserenc, Hélène

    2014-12-01

    Natural killer cells are the first lymphocyte subset to reconstitute, and play a major role in early immunity after allogeneic hematopoietic stem cell transplantation. Cells expressing the activating receptor NKG2C seem crucial in the resolution of cytomegalovirus episodes, even in the absence of T cells. We prospectively investigated natural killer-cell reconstitution in a cohort of 439 adult recipients who underwent non-T-cell-depleted allogeneic hematopoietic stem cell transplantation between 2005 and 2012. Freshly collected blood samples were analyzed 3, 6, 12 and 24 months after transplantation. Data were studied with respect to conditioning regimen, source of stem cells, underlying disease, occurrence of graft-versus-host disease, and profiles of cytomegalovirus reactivation. In multivariate analysis we found that the absolute numbers of CD56(bright) natural killer cells at month 3 were significantly higher after myeloablative conditioning than after reduced intensity conditioning. Acute graft-versus-host disease impaired reconstitution of total and CD56(dim) natural killer cells at month 3. In contrast, high natural killer cell count at month 3 was associated with a lower incidence of chronic graft-versus-host disease, independently of a previous episode of acute graft-versus-host disease and stem cell source. NKG2C(+)CD56(dim) and total natural killer cell counts at month 3 were lower in patients with reactivation of cytomegalovirus between month 0 and month 3, but expanded greatly afterwards. These cells were also less numerous in patients who experienced later cytomegalovirus reactivation between month 3 and month 6. Our results advocate a direct role of NKG2C-expressing natural killer cells in the early control of cytomegalovirus reactivation after allogeneic hematopoietic stem cell transplantation.

  8. Favorable impact of natural killer cell reconstitution on chronic graft-versus-host disease and cytomegalovirus reactivation after allogeneic hematopoietic stem cell transplantation

    PubMed Central

    Kheav, Vissal David; Busson, Marc; Scieux, Catherine; de Latour, Régis Peffault; Maki, Guitta; Haas, Philippe; Mazeron, Marie-Christine; Carmagnat, Maryvonnick; Masson, Emeline; Xhaard, Aliénor; Robin, Marie; Ribaud, Patricia; Dulphy, Nicolas; Loiseau, Pascale; Charron, Dominique; Socié, Gérard; Toubert, Antoine; Moins-Teisserenc, Hélène

    2014-01-01

    Natural killer cells are the first lymphocyte subset to reconstitute, and play a major role in early immunity after allogeneic hematopoietic stem cell transplantation. Cells expressing the activating receptor NKG2C seem crucial in the resolution of cytomegalovirus episodes, even in the absence of T cells. We prospectively investigated natural killer-cell reconstitution in a cohort of 439 adult recipients who underwent non-T-cell-depleted allogeneic hematopoietic stem cell transplantation between 2005 and 2012. Freshly collected blood samples were analyzed 3, 6, 12 and 24 months after transplantation. Data were studied with respect to conditioning regimen, source of stem cells, underlying disease, occurrence of graft-versus-host disease, and profiles of cytomegalovirus reactivation. In multivariate analysis we found that the absolute numbers of CD56bright natural killer cells at month 3 were significantly higher after myeloablative conditioning than after reduced intensity conditioning. Acute graft-versus-host disease impaired reconstitution of total and CD56dim natural killer cells at month 3. In contrast, high natural killer cell count at month 3 was associated with a lower incidence of chronic graft-versus-host disease, independently of a previous episode of acute graft-versus-host disease and stem cell source. NKG2C+CD56dim and total natural killer cell counts at month 3 were lower in patients with reactivation of cytomegalovirus between month 0 and month 3, but expanded greatly afterwards. These cells were also less numerous in patients who experienced later cytomegalovirus reactivation between month 3 and month 6. Our results advocate a direct role of NKG2C-expressing natural killer cells in the early control of cytomegalovirus reactivation after allogeneic hematopoietic stem cell transplantation. PMID:25085354

  9. In Vivo Proof of Concept of Adoptive Immunotherapy for Hepatocellular Carcinoma Using Allogeneic Suicide Gene-modified Killer Cells

    PubMed Central

    Leboeuf, Céline; Mailly, Laurent; Wu, Tao; Bour, Gaetan; Durand, Sarah; Brignon, Nicolas; Ferrand, Christophe; Borg, Christophe; Tiberghien, Pierre; Thimme, Robert; Pessaux, Patrick; Marescaux, Jacques; Baumert, Thomas F.; Robinet, Eric

    2014-01-01

    Cell therapy based on alloreactivity has completed clinical proof of concept against hematological malignancies. However, the efficacy of alloreactivity as a therapeutic approach to treat solid tumors is unknown. Using cell culture and animal models, we aimed to investigate the efficacy and safety of allogeneic suicide gene-modified killer cells as a cell-based therapy for hepatocellular carcinoma (HCC), for which treatment options are limited. Allogeneic killer cells from healthy donors were isolated, expanded, and phenotypically characterized. Antitumor cytotoxic activity and safety were studied using a panel of human or murine HCC cell lines engrafted in immunodeficient or immunocompetent mouse models. Human allogeneic suicide gene-modified killer cells (aSGMKCs) exhibit a high, rapid, interleukin-2–dependent, and non–major histocompatibility complex class I-restricted in vitro cytotoxicity toward human hepatoma cells, mainly mediated by natural killer (NK) and NK-like T cells. In vivo evaluation of this cell therapy product demonstrates a marked, rapid, and sustained regression of HCC. Preferential liver homing of effector cells contributed to its marked efficacy. Calcineurin inhibitors allowed preventing rejection of allogeneic lymphocytes by the host immune system without impairing their antitumor activity. Our results demonstrate proof of concept for aSGMKCs as immunotherapy for HCC and open perspectives for the clinical development of this approach. PMID:24445938

  10. Killer cell immunoglobulin-like receptor gene association with cryptorchidism.

    PubMed

    Niepiekło-Miniewska, Wanda; Kuśnierczyk, Piotr; Havrylyuk, Anna; Kamieniczna, Marzena; Nakonechnyy, Andrij; Chopyak, Valentyna; Kurpisz, Maciej

    2015-12-01

    Cryptorchidism is a condition where a testis persists in the abdominal cavity. Thus, due to elevated temperature we may expect induction of aberrant immune reactions depending on genetic constitution of individual. This may be reflected by development of anti-sperm antibodies (ASA) in cryptorchid males. Also, natural killer (NK) cells which belong to innate immunity may control adaptive immunity. Therefore, the gene system encoding polymorphic NK cell immunoglobulin receptors (KIRs) has been studied. 109 prepubertal boys with cryptorchidism and 136 ethnically matched young male donors were selected to study NK cell KIRs. DNA was isolated using automatic Maxwell(®) system from the peripheral venous blood drawn onto anticoagulant. Olerup SSP KIR Genotyping kit including Taq polymerase was used for detection of KIR genes. Human leukocyte antigen-C (HLA-C) groups, C1 and C2 were established using a Olerup SSP KIR HLA Ligand kit. KIR2DL2 (killer immunoglobulin-like receptor two-domain long 2) and KIR2DS2 (killer immunoglobulin-like receptor two-domain short 2) genes were less frequent in patients than in control individuals (corrected p values: 0.0110 and 0.0383, respectively). However, no significant differences were observed between ASA-positive and ASA-negative patients, or between bilateral or unilateral cryptorchidism. No association between KIR ligands C1 and C2, alone or together with KIR2DL2, was found. However, the results suggest that KIR2DL2+/KIR2DS2+ genotype may be, to some extent, protective against cryptorchidism.

  11. 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.

  12. Clinical utility of natural killer cells in cancer therapy and transplantation

    PubMed Central

    Knorr, David; Bachanova, Veronika; Verneris, Michael R.; Miller, Jeffrey S.

    2014-01-01

    Natural killer (NK) cells recognize deranged cells that display stress receptors or loss of major histocompatibility complex (MHC) class I. During development, NK cells become “licensed” only after they encounter cognate human leucocyte antigen (HLA) class I, leading to the acquisition of effector function. NK cells can be exploited for cancer therapy in several ways. These include targeting within monoclonal antibodies alone or combined with ex vivo and in vivo NK cell activation to facilitate adoptive immunotherapy using donor-derived NK cell products to induce graft-vs-tumor effects. In the adoptive transfer setting, persistence and in vivo expansion requires lymphodepleting chemotherapy to prevent rejection and provide homeostatic cytokines (such as IL-15) that activate NK cells. IL-15 has the advantage of avoiding regulatory T-cell expansion. Clinical applications are currently being tested. To enhance in vivo expansion, IL-2 has been used at low doses. However, low dose administration also leads to the stimulation of regulatory T cells. Monoclonal antibodies and bispecific killer engagers (BiKEs) may enhance specificity by targeting CD16 on NK cells to tumor antigens. Inhibition of CD16 shedding may also promote enhanced cytotoxicity. Future strategies include exploiting favorable donor immunogenetics or ex vivo expansion of NK cells from blood, progenitors, or pluripotent cells. Comparative clinical trials are needed to test these approaches. PMID:24618042

  13. Killer Cell Immunoglobulin-like Receptor Genotype and Haplotype Investigation of Natural Killer Cells from an Australian Population of Chronic Fatigue Syndrome/Myalgic Encephalomyelitis Patients.

    PubMed

    Huth, T K; Brenu, E W; Staines, D R; Marshall-Gradisnik, S M

    2016-01-01

    Killer cell immunoglobulin-like receptor (KIR) genes encode for activating and inhibitory surface receptors, which are correlated with the regulation of Natural Killer (NK) cell cytotoxic activity. Reduced NK cell cytotoxic activity has been consistently reported in Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME) patients, and KIR haplotypes and allelic polymorphism remain to be investigated. The aim of this article was to conduct a pilot study to examine KIR genotypes, haplotypes, and allelic polymorphism in CFS/ME patients and nonfatigued controls (NFCs). Comparison of KIR and allelic polymorphism frequencies revealed no significant differences between 20 CFS/ME patients and 20 NFCs. A lower frequency of the telomeric A/B motif (P < 0.05) was observed in CFS/ME patients compared with NFCs. This pilot study is the first to report the differences in the frequency of KIR on the telomeric A/B motif in CFS/ME patients. Further studies with a larger CFS/ME cohort are required to validate these results.

  14. Killer Cell Immunoglobulin-like Receptor Genotype and Haplotype Investigation of Natural Killer Cells from an Australian Population of Chronic Fatigue Syndrome/Myalgic Encephalomyelitis Patients

    PubMed Central

    Huth, T. K.; Brenu, E. W.; Staines, D. R.; Marshall-Gradisnik, S. M.

    2016-01-01

    Killer cell immunoglobulin-like receptor (KIR) genes encode for activating and inhibitory surface receptors, which are correlated with the regulation of Natural Killer (NK) cell cytotoxic activity. Reduced NK cell cytotoxic activity has been consistently reported in Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME) patients, and KIR haplotypes and allelic polymorphism remain to be investigated. The aim of this article was to conduct a pilot study to examine KIR genotypes, haplotypes, and allelic polymorphism in CFS/ME patients and nonfatigued controls (NFCs). Comparison of KIR and allelic polymorphism frequencies revealed no significant differences between 20 CFS/ME patients and 20 NFCs. A lower frequency of the telomeric A/B motif (P < 0.05) was observed in CFS/ME patients compared with NFCs. This pilot study is the first to report the differences in the frequency of KIR on the telomeric A/B motif in CFS/ME patients. Further studies with a larger CFS/ME cohort are required to validate these results. PMID:27346947

  15. Killer Cell Immunoglobulin-like Receptor Genotype and Haplotype Investigation of Natural Killer Cells from an Australian Population of Chronic Fatigue Syndrome/Myalgic Encephalomyelitis Patients.

    PubMed

    Huth, T K; Brenu, E W; Staines, D R; Marshall-Gradisnik, S M

    2016-01-01

    Killer cell immunoglobulin-like receptor (KIR) genes encode for activating and inhibitory surface receptors, which are correlated with the regulation of Natural Killer (NK) cell cytotoxic activity. Reduced NK cell cytotoxic activity has been consistently reported in Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME) patients, and KIR haplotypes and allelic polymorphism remain to be investigated. The aim of this article was to conduct a pilot study to examine KIR genotypes, haplotypes, and allelic polymorphism in CFS/ME patients and nonfatigued controls (NFCs). Comparison of KIR and allelic polymorphism frequencies revealed no significant differences between 20 CFS/ME patients and 20 NFCs. A lower frequency of the telomeric A/B motif (P < 0.05) was observed in CFS/ME patients compared with NFCs. This pilot study is the first to report the differences in the frequency of KIR on the telomeric A/B motif in CFS/ME patients. Further studies with a larger CFS/ME cohort are required to validate these results. PMID:27346947

  16. Bovine colostrum enhances natural killer cell activity and immune response in a mouse model of influenza infection and mediates intestinal immunity through toll-like receptors 2 and 4.

    PubMed

    Wong, Eric B; Mallet, Jean-François; Duarte, Jairo; Matar, Chantal; Ritz, Barry W

    2014-04-01

    Oral administration of bovine colostrum affects intestinal immunity, including an increased percentage of natural killer (NK) cells. However, effects on NK cell cytotoxic activity and resistance to infection as well as a potential mechanism remain unclear. Therefore, we investigated the effects of bovine colostrum (La Belle, Inc, Bellingham, WA) on the NK cytotoxic response to influenza infection and on toll-like receptor (TLR) activity in a primary intestinal epithelial cell culture. We hypothesized that colostrum would increase NK cell activity and that TLR-2 and TLR-4 blocking would reduce interleukin 6 production by epithelial cells in response to contact stimulation with colostrum. Four-month-old female C57BL/6 mice were supplemented with 1 g of colostrum per kilogram of body weight before and after infection with influenza A virus (H1N1). Animals were assessed for weight loss, splenic NK cell activity, and lung virus titers. Colostrum-supplemented mice demonstrated less reduction in body weight after influenza infection, indicating a less severe infection, increased NK cell cytotoxicity, and less virus burden in the lungs compared with controls. Colostrum supplementation enhanced NK cell cytotoxicity and improved the immune response to primary influenza virus infection in mice. To investigate a potential mechanism, a primary culture of small intestine epithelial cells was then stimulated with colostrum. Direct activation of epithelial cells resulted in increased interleukin 6 production, which was inhibited with TLR-2 and TLR-4 blocking antibodies. The interaction between colostrum and immunity may be dependent, in part, on the interaction of colostrum components with innate receptors at the intestinal epithelium, including TLR-2 and TLR-4. PMID:24774068

  17. Bovine colostrum enhances natural killer cell activity and immune response in a mouse model of influenza infection and mediates intestinal immunity through toll-like receptors 2 and 4.

    PubMed

    Wong, Eric B; Mallet, Jean-François; Duarte, Jairo; Matar, Chantal; Ritz, Barry W

    2014-04-01

    Oral administration of bovine colostrum affects intestinal immunity, including an increased percentage of natural killer (NK) cells. However, effects on NK cell cytotoxic activity and resistance to infection as well as a potential mechanism remain unclear. Therefore, we investigated the effects of bovine colostrum (La Belle, Inc, Bellingham, WA) on the NK cytotoxic response to influenza infection and on toll-like receptor (TLR) activity in a primary intestinal epithelial cell culture. We hypothesized that colostrum would increase NK cell activity and that TLR-2 and TLR-4 blocking would reduce interleukin 6 production by epithelial cells in response to contact stimulation with colostrum. Four-month-old female C57BL/6 mice were supplemented with 1 g of colostrum per kilogram of body weight before and after infection with influenza A virus (H1N1). Animals were assessed for weight loss, splenic NK cell activity, and lung virus titers. Colostrum-supplemented mice demonstrated less reduction in body weight after influenza infection, indicating a less severe infection, increased NK cell cytotoxicity, and less virus burden in the lungs compared with controls. Colostrum supplementation enhanced NK cell cytotoxicity and improved the immune response to primary influenza virus infection in mice. To investigate a potential mechanism, a primary culture of small intestine epithelial cells was then stimulated with colostrum. Direct activation of epithelial cells resulted in increased interleukin 6 production, which was inhibited with TLR-2 and TLR-4 blocking antibodies. The interaction between colostrum and immunity may be dependent, in part, on the interaction of colostrum components with innate receptors at the intestinal epithelium, including TLR-2 and TLR-4.

  18. Homeostatic regulation of marginal zone B cells by invariant natural killer T cells.

    PubMed

    Wen, Xiangshu; Yang, Jun-Qi; Kim, Peter J; Singh, Ram Raj

    2011-01-01

    Marginal zone B cells (MZB) mount a rapid antibody response, potently activate naïve T cells, and are enriched in autoreactive B cells. MZBs express high levels of CD1d, the restriction element for invariant natural killer T cells (iNKT). Here, we examined the effect of iNKT cells on MZB cell activation and numbers in vitro and in vivo in normal and autoimmune mice. Results show that iNKT cells activate MZBs, but restrict their numbers in vitro and in vivo in normal BALB/c and C57/BL6 mice. iNKT cells do so by increasing the activation-induced cell death and curtailing proliferation of MZB cells, whereas they promote the proliferation of follicular B cells. Sorted iNKT cells can directly execute this function, without help from other immune cells. Such MZB regulation by iNKTs is mediated, at least in part, via CD1d on B cells in a contact-dependent manner, whereas iNKT-induced proliferation of follicular B cells occurs in a contact- and CD1d-independent manner. Finally, we show that iNKT cells reduce 'autoreactive' MZB cells in an anti-DNA transgenic model, and limit MZB cell numbers in autoimmune-prone (NZB×NZW)F1 and non-obese diabetic mice, suggesting a potentially new mechanism whereby iNKT cells might regulate pathologic autoimmunity. Differential regulation of follicular B cells versus potentially autoreactive MZBs by iNKT cells has important implications for autoimmune diseases as well as for conditions that require a rapid innate B cell response.

  19. Cytokine-induced memory-like natural killer cells exhibit enhanced responses against myeloid leukemia.

    PubMed

    Romee, Rizwan; Rosario, Maximillian; Berrien-Elliott, Melissa M; Wagner, Julia A; Jewell, Brea A; Schappe, Timothy; Leong, Jeffrey W; Abdel-Latif, Sara; Schneider, Stephanie E; Willey, Sarah; Neal, Carly C; Yu, Liyang; Oh, Stephen T; Lee, Yi-Shan; Mulder, Arend; Claas, Frans; Cooper, Megan A; Fehniger, Todd A

    2016-09-21

    Natural killer (NK) cells are an emerging cellular immunotherapy for patients with acute myeloid leukemia (AML); however, the best approach to maximize NK cell antileukemia potential is unclear. Cytokine-induced memory-like NK cells differentiate after a brief preactivation with interleukin-12 (IL-12), IL-15, and IL-18 and exhibit enhanced responses to cytokine or activating receptor restimulation for weeks to months after preactivation. We hypothesized that memory-like NK cells exhibit enhanced antileukemia functionality. We demonstrated that human memory-like NK cells have enhanced interferon-γ production and cytotoxicity against leukemia cell lines or primary human AML blasts in vitro. Using mass cytometry, we found that memory-like NK cell functional responses were triggered against primary AML blasts, regardless of killer cell immunoglobulin-like receptor (KIR) to KIR-ligand interactions. In addition, multidimensional analyses identified distinct phenotypes of control and memory-like NK cells from the same individuals. Human memory-like NK cells xenografted into mice substantially reduced AML burden in vivo and improved overall survival. In the context of a first-in-human phase 1 clinical trial, adoptively transferred memory-like NK cells proliferated and expanded in AML patients and demonstrated robust responses against leukemia targets. Clinical responses were observed in five of nine evaluable patients, including four complete remissions. Thus, harnessing cytokine-induced memory-like NK cell responses represents a promising translational immunotherapy approach for patients with AML.

  20. Cytokine-induced memory-like natural killer cells exhibit enhanced responses against myeloid leukemia.

    PubMed

    Romee, Rizwan; Rosario, Maximillian; Berrien-Elliott, Melissa M; Wagner, Julia A; Jewell, Brea A; Schappe, Timothy; Leong, Jeffrey W; Abdel-Latif, Sara; Schneider, Stephanie E; Willey, Sarah; Neal, Carly C; Yu, Liyang; Oh, Stephen T; Lee, Yi-Shan; Mulder, Arend; Claas, Frans; Cooper, Megan A; Fehniger, Todd A

    2016-09-21

    Natural killer (NK) cells are an emerging cellular immunotherapy for patients with acute myeloid leukemia (AML); however, the best approach to maximize NK cell antileukemia potential is unclear. Cytokine-induced memory-like NK cells differentiate after a brief preactivation with interleukin-12 (IL-12), IL-15, and IL-18 and exhibit enhanced responses to cytokine or activating receptor restimulation for weeks to months after preactivation. We hypothesized that memory-like NK cells exhibit enhanced antileukemia functionality. We demonstrated that human memory-like NK cells have enhanced interferon-γ production and cytotoxicity against leukemia cell lines or primary human AML blasts in vitro. Using mass cytometry, we found that memory-like NK cell functional responses were triggered against primary AML blasts, regardless of killer cell immunoglobulin-like receptor (KIR) to KIR-ligand interactions. In addition, multidimensional analyses identified distinct phenotypes of control and memory-like NK cells from the same individuals. Human memory-like NK cells xenografted into mice substantially reduced AML burden in vivo and improved overall survival. In the context of a first-in-human phase 1 clinical trial, adoptively transferred memory-like NK cells proliferated and expanded in AML patients and demonstrated robust responses against leukemia targets. Clinical responses were observed in five of nine evaluable patients, including four complete remissions. Thus, harnessing cytokine-induced memory-like NK cell responses represents a promising translational immunotherapy approach for patients with AML. PMID:27655849

  1. Advantages and applications of CAR-expressing natural killer cells

    PubMed Central

    Glienke, Wolfgang; Esser, Ruth; Priesner, Christoph; Suerth, Julia D.; Schambach, Axel; Wels, Winfried S.; Grez, Manuel; Kloess, Stephan; Arseniev, Lubomir; Koehl, Ulrike

    2015-01-01

    In contrast to donor T cells, natural killer (NK) cells are known to mediate anti-cancer effects without the risk of inducing graft-versus-host disease (GvHD). In order to improve cytotoxicity against resistant cancer cells, auspicious efforts have been made with chimeric antigen receptor (CAR) expressing T- and NK cells. These CAR-modified cells express antigen receptors against tumor-associated surface antigens, thus redirecting the effector cells and enhancing tumor-specific immunosurveillance. However, many cancer antigens are also expressed on healthy tissues, potentially leading to off tumor/on target toxicity by CAR-engineered cells. In order to control such potentially severe side effects, the insertion of suicide genes into CAR-modified effectors can provide a means for efficient depletion of these cells. While CAR-expressing T cells have entered successfully clinical trials, experience with CAR-engineered NK cells is mainly restricted to pre-clinical investigations and predominantly to NK cell lines. In this review we summarize the data on CAR expressing NK cells focusing on the possible advantage using these short-lived effector cells and discuss the necessity of suicide switches. Furthermore, we address the compliance of such modified NK cells with regulatory requirements as a new field in cellular immunotherapy. PMID:25729364

  2. Natural killer cells in hepatitis C: Current progress

    PubMed Central

    Yoon, Joo Chun; Yang, Chang Mo; Song, Youkyong; Lee, Jae Myun

    2016-01-01

    Patients infected with the hepatitis C virus (HCV) are characterized by a high incidence of chronic infection, which results in chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma. The functional impairment of HCV-specific T cells is associated with the evolution of an acute infection to chronic hepatitis. While T cells are the important effector cells in adaptive immunity, natural killer (NK) cells are the critical effector cells in innate immunity to virus infections. The findings of recent studies on NK cells in hepatitis C suggest that NK cell responses are indeed important in each phase of HCV infection. In the early phase, NK cells are involved in protective immunity to HCV. The immune evasion strategies used by HCV may target NK cells and might contribute to the progression to chronic hepatitis C. NK cells may control HCV replication and modulate hepatic fibrosis in the chronic phase. Further investigations are, however, needed, because a considerable number of studies observed functional impairment of NK cells in chronic HCV infection. Interestingly, the enhanced NK cell responses during interferon-α-based therapy of chronic hepatitis C indicate successful treatment. In spite of the advances in research on NK cells in hepatitis C, establishment of more physiological HCV infection model systems is needed to settle unsolved controversies over the role and functional status of NK cells in HCV infection. PMID:26819513

  3. Comparative Genomics of Natural Killer Cell Receptor Gene Clusters

    PubMed Central

    Kelley, James; Walter, Lutz; Trowsdale, John

    2005-01-01

    Many receptors on natural killer (NK) cells recognize major histocompatibility complex class I molecules in order to monitor unhealthy tissues, such as cells infected with viruses, and some tumors. Genes encoding families of NK receptors and related sequences are organized into two main clusters in humans: the natural killer complex on Chromosome 12p13.1, which encodes C-type lectin molecules, and the leukocyte receptor complex on Chromosome 19q13.4, which encodes immunoglobulin superfamily molecules. The composition of these gene clusters differs markedly between closely related species, providing evidence for rapid, lineage-specific expansions or contractions of sets of loci. The choice of NK receptor genes is polarized in the two species most studied, mouse and human. In mouse, the C-type lectin-related Ly49 gene family predominates. Conversely, the single Ly49 sequence is a pseudogene in humans, and the immunoglobulin superfamily KIR gene family is extensive. These different gene sets encode proteins that are comparable in function and genetic diversity, even though they have undergone species-specific expansions. Understanding the biological significance of this curious situation may be aided by studying which NK receptor genes are used in other vertebrates, especially in relation to species-specific differences in genes for major histocompatibility complex class I molecules. PMID:16132082

  4. Consequences of the crosstalk between monocytes/macrophages and natural killer cells

    PubMed Central

    Michel, Tatiana; Hentges, François; Zimmer, Jacques

    2013-01-01

    The interaction between natural killer (NK) cells and different other immune cells like T cells and dendritic cells is well-described, but the crosstalk with monocytes or macrophages and the nature of ligands/receptors implicated are just emerging. The macrophage-NK interaction is a major first-line defense against pathogens (bacteria, viruses, fungi, and parasites). The recruitment and the activation of NK cells to perform cytotoxicity or produce cytokines at the sites of inflammation are important to fight infections. The two main mechanisms by which macrophages can prime NK cells are (1) activation through soluble mediators such as IL-12, IL-18, and (2) stimulation through direct cell-to-cell contact. We will discuss the progress in matters of modulation of NK cell functions by monocytes and macrophages, in the steady state and during diseases. PMID:23316194

  5. Natural killer cell immunosenescence in acute myeloid leukaemia patients: new targets for immunotherapeutic strategies?

    PubMed

    Sanchez-Correa, Beatriz; Campos, Carmen; Pera, Alejandra; Bergua, Juan M; Arcos, Maria Jose; Bañas, Helena; Casado, Javier G; Morgado, Sara; Duran, Esther; Solana, Rafael; Tarazona, Raquel

    2016-04-01

    Several age-associated changes in natural killer (NK) cell phenotype have been reported that contribute to the defective NK cell response observed in elderly patients. A remodelling of the NK cell compartment occurs in the elderly with a reduction in the output of immature CD56(bright) cells and an accumulation of highly differentiated CD56(dim) NK cells. Acute myeloid leukaemia (AML) is generally a disease of older adults. NK cells in AML patients show diminished expression of several activating receptors that contribute to impaired NK cell function and, in consequence, to AML blast escape from NK cell immunosurveillance. In AML patients, phenotypic changes in NK cells have been correlated with disease progression and survival. NK cell-based immunotherapy has emerged as a possibility for the treatment of AML patients. The understanding of age-associated alterations in NK cells is therefore necessary to define adequate therapeutic strategies in older AML patients.

  6. Immunotherapeutic strategies targeting natural killer T cell responses in cancer.

    PubMed

    Shissler, Susannah C; Bollino, Dominique R; Tiper, Irina V; Bates, Joshua P; Derakhshandeh, Roshanak; Webb, Tonya J

    2016-08-01

    Natural killer T (NKT) cells are a unique subset of lymphocytes that bridge the innate and adaptive immune system. NKT cells possess a classic αβ T cell receptor (TCR) that is able to recognize self and foreign glycolipid antigens presented by the nonclassical class I major histocompatibility complex (MHC) molecule, CD1d. Type I NKT cells (referred to as invariant NKT cells) express a semi-invariant Vα14Jα18 TCR in mice and Vα24Jα18 TCR in humans. Type II NKT cells are CD1d-restricted T cells that express a more diverse set of TCR α chains. The two types of NKT cells often exert opposing effects especially in tumor immunity, where type II cells generally suppress tumor immunity while type I NKT cells can enhance anti-tumor immune responses. In this review, we focus on the role of NKT cells in cancer. We discuss their effector and suppressive functions, as well as describe preclinical and clinical studies utilizing therapeutic strategies focused on harnessing their potent anti-tumor effector functions, and conclude with a discussion on potential next steps for the utilization of NKT cell-targeted therapies for the treatment of cancer. PMID:27393665

  7. Synthesis and evaluation of immunostimulant plasmalogen lysophosphatidylethanolamine and analogues for natural killer T cells.

    PubMed

    Ni, Guanghui; Li, Zhiyuan; Liang, Kangjiang; Wu, Ting; De Libero, Gennaro; Xia, Chengfeng

    2014-06-01

    Plasmalogen lysophosphatidylethanolamine (pLPE) had been identified as a self antigen for natural killer T cells (NKT cells). It is very important in the development, maturation and activation of NKT cells in thymus. Besides, pLPE is a novel type of antigen for NKT cells. To evaluate the structure-activity relationship (SAR) of this new antigen, pLPE and its analogues referred to different aliphatic chains and linkages at the sn-1 position of the glycerol backbone were synthesized, and the biological activities of these analogues was characterized. It is discovered that the linkages between phosphate and lipid moiety are not important for the antigens' activities. The pLPE analogues 1, 3, 4, 7 and 9, which have additional double bonds on lipid parts, were identified as new NKT agonists. Moreover, the analogues 4, 7 and 9 were discovered as potent Th2 activators for NKT cells.

  8. KLRG+ invariant natural killer T cells are long-lived effectors.

    PubMed

    Shimizu, Kanako; Sato, Yusuke; Shinga, Jun; Watanabe, Takashi; Endo, Takaho; Asakura, Miki; Yamasaki, Satoru; Kawahara, Kazuyoshi; Kinjo, Yuki; Kitamura, Hiroshi; Watarai, Hiroshi; Ishii, Yasuyuki; Tsuji, Moriya; Taniguchi, Masaru; Ohara, Osamu; Fujii, Shin-ichiro

    2014-08-26

    Immunological memory has been regarded as a unique feature of the adaptive immune response mediated in an antigen-specific manner by T and B lymphocytes. However, natural killer (NK) cells and γδT cells, which traditionally are classified as innate immune cells, have been shown in recent studies to have hallmark features of memory cells. Invariant NKT cell (iNKT cell)-mediated antitumor effects indicate that iNKT cells are activated in vivo by vaccination with iNKT cell ligand-loaded CD1d(+) cells, but not by vaccination with unbound NKT cell ligand. In such models, it previously was thought that the numbers of IFN-γ-producing cells in the spleen returned to the basal level around 1 wk after the vaccination. In the current study, we demonstrate the surprising presence of effector memory-like iNKT cells in the lung. We found long-term antitumor activity in the lungs of mice was enhanced after vaccination with iNKT cell ligand-loaded dendritic cells. Further analyses showed that the KLRG1(+) (Killer cell lectin-like receptor subfamily G, member 1-positive) iNKT cells coexpressing CD49d and granzyme A persisted for several months and displayed a potent secondary response to cognate antigen. Finally, analyses of CDR3β by RNA deep sequencing demonstrated that some particular KLRG1(+) iNKT-cell clones accumulated, suggesting the selection of certain T-cell receptor repertoires by an antigen. The current findings identifying effector memory-like KLRG1(+) iNKT cells in the lung could result in a paradigm shift regarding the basis of newly developed extrathymic iNKT cells and could contribute to the future development of antitumor immunotherapy by uniquely energizing iNKT cells. PMID:25118276

  9. Natural killer cells facilitate PRAME-specific T-cell reactivity against neuroblastoma.

    PubMed

    Spel, Lotte; Boelens, Jaap-Jan; van der Steen, Dirk M; Blokland, Nina J G; van Noesel, Max M; Molenaar, Jan J; Heemskerk, Mirjam H M; Boes, Marianne; Nierkens, Stefan

    2015-11-01

    Neuroblastoma is the most common solid tumor in children with an estimated 5-year progression free survival of 20-40% in stage 4 disease. Neuroblastoma actively avoids recognition by natural killer (NK) cells and cytotoxic T lymphocytes (CTLs). Although immunotherapy has gained traction for neuroblastoma treatment, these immune escape mechanisms restrain clinical results. Therefore, we aimed to improve neuroblastoma immunogenicity to further the development of antigen-specific immunotherapy against neuroblastoma. We found that neuroblastoma cells significantly increase surface expression of MHC I upon exposure to active NK cells which thereby readily sensitize neuroblastoma cells for recognition by CTLs. We show that oncoprotein PRAME serves as an immunodominant antigen for neuroblastoma as NK-modulated neuroblastoma cells are recognized by PRAMESLLQHLIGL/A2-specific CTL clones. Furthermore, NK cells induce MHC I upregulation in neuroblastoma through contact-dependent secretion of IFNγ. Our results demonstrate remarkable plasticity in the peptide/MHC I surface expression of neuroblastoma cells, which is reversed when neuroblastoma cells experience innate immune attack by sensitized NK cells. These findings support the exploration of NK cells as adjuvant therapy to enforce neuroblastoma-specific CTL responses.

  10. Natural killer cell biology: an update and future directions.

    PubMed

    Campbell, Kerry S; Hasegawa, Jun

    2013-09-01

    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 2 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 might 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 patients with cancer. 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 pathologic conditions.

  11. Interleukin-5 Supports the Expansion of Fas Ligand-Expressing Killer B Cells that Induce Antigen-Specific Apoptosis of CD4+ T Cells and Secrete Interleukin-10

    PubMed Central

    Klinker, Matthew W.; Reed, Tamra J.; Fox, David A.; Lundy, Steven K.

    2013-01-01

    Beyond their critical role in humoral immunity, B lymphocytes can employ a variety of immunomodulatory mechanisms including expression of the apoptosis-inducing molecule Fas ligand (FasL; CD178). Here, we extensively characterized the surface phenotype of FasL+ killer B cells, showing they are enriched in the IgMhighCD5+CD1dhigh B cell subset previously reported to contain a higher frequency of B cells producing interleukin-10 (IL-10). A rare population of B cells expressing IL-10 was present among FasL+ B cells, but most FasL+ B cells did not produce IL-10. We also identify interleukin-5 (IL-5) as a novel inducer of killer B cell function. Constitutively FasL+ B cells expressed higher levels of the IL-5 receptor, and treating B cells with IL-5 and CD40L resulted in the expansion of a B cell population enriched for FasL+ cells. B cells stimulated with IL-5 and CD40L were potent inducers of apoptosis in activated primary CD4+ T cells, and this killing function was antigen-specific and dependent upon FasL. IL-5 also enhanced IL-10 secretion in B cells stimulated with CD40L. Taken together these findings elucidate the relationship of FasL+ B cells and IL-10-producing B cells and demonstrate that IL-5 can induce or enhance both killer B cell activity and IL-10 secretion in B cells. Finally, we found that the killer B cell activity induced by IL-5 was completely blocked by IL-4, suggesting the existence of a previously unknown antagonistic relationship between these type-2 cytokines in modulating the activity of killer B cells. Targeting this IL-5/IL-4 signaling axis may therefore represent a novel area of drug discovery in inflammatory disorders. PMID:23940537

  12. [Music therapy induced alternations in natural killer cell count and function].

    PubMed

    Hasegawa, Y; Kubota, N; Inagaki, T; Shinagawa, N

    2001-03-01

    The effects of music therapy on natural killer (NK) cell count and activity (NKCA) were studied in 19 persons. Alzheimer's disease, cerebrovessel disease and Parkinson's disease subjects were assigned to a music therapy. Blood samples were drawn at rest and after completion of music therapy. Music therapy did not change the number of circulating lymphocytes. The percentage of NK cells increased during music therapy, along with an increase in the NK cell activity. The proportion of T cells, CD4 and CD8 did not change significantly during music therapy. One hour after the music therapy session, plasma adrenaline increased but cortisol and noradrenalin did not change. The results indicate that music therapy can significantly increase NK cell count and activity. The change in NK cell and function were independent of neuro-degenerative diseases.

  13. Type I interferon regulation of natural killer cell function in primary and secondary infections.

    PubMed

    Stackaruk, Michele L; Lee, Amanda J; Ashkar, Ali A

    2013-08-01

    The priming of natural killer (NK) cells by type I interferon (IFN) is necessary for protection against primary and secondary viral infections. However, the pathway by which type I IFN activates NK cells to elicit antiviral responses is controversial. There is evidence to suggest that type I IFN priming of NK cells occurs through both direct and indirect pathways. As with many innate mechanisms, type I IFN and NK cells also orchestrate the adaptive immune response and thus aid in protection against secondary infections. Type I IFN can shape CD4(+) T cell, B cell and humoral memory formation. In addition, long-lived NK cells can perform specific and enhanced memory-like protection in secondary infections. This review outlines the different mechanisms underlying type I IFN regulation of NK cells and how type I IFN and NK cells can be used as a therapeutic target in vaccinations.

  14. Maternal uterine natural killer cells nurture fetal growth: in medio stat virtus.

    PubMed

    Colucci, Francesco; Kieckbusch, Jens

    2015-02-01

    Much research in reproductive immunology is preoccupied with maternal tolerance of the semi-allogeneic fetus. This inevitably leads to the assumption that the maternal immune system should be suppressed, similarly to the immunosuppression needed to avoid rejection of an allograft. However, the parallels with transplantation immunology are misleading, and we discuss how interactions between variable immune system genes expressed on maternal natural killer (NK) cells and on the fetal trophoblast modulate fetal growth. Exaggerated suppression or activation of maternal NK cells associates with both extremes of birth weight.

  15. The role of p44/42 activation in tributyltin- induced inhibition of human natural killer cells: Effects of MEK inhibitors

    PubMed Central

    Abraha, Abraham B.; Whalen, Margaret M.

    2008-01-01

    Destruction of tumor cells is a key function of NK cells. Previous studies have shown that tributyltin (TBT) can significantly reduce the lytic function of the human NK cells with accompanying increases in the phosphorylation (activation) states of the mitogen activated protein kinases (MAPKs), p44/42. The current studies examine the role of p44/42 activation in the TBT-induced reduction of NK-lytic function, by using MAPK kinase (MEK) inhibitors, PD98059 and U0126. A 1 h treatment with PD98059 or U0126 or both decreased the ability of NK cells to lyse K562 tumor cells. PD98059, U0126 or a combination of both inhibitors were able to completely block TBT-induced activation of p44/42. However, when p44/42 activation was blocked by the presence of PD98059, U0126, or the combination, subsequent exposure to TBT was still able to decrease the lytic function of NK cells. These results indicate that TBT-induced activation of p44/42 occurs via the activation of its upstream activator, MEK, and not by a TBT-induced inhibition of p44/42 phosphatase activity. Additionally, as lytic function was never completely blocked by MEK inhibitors, the results indicate that activation of p44/42 pathway is not solely responsible for the activation of lytic function of freshly isolated human NK cells. Finally, the results showed that TBT-induced activation of p44/42 is not solely responsible for the loss of lytic function. PMID:18989867

  16. Enhanced targeting of stem-like solid tumor cells with radiation and natural killer cells

    PubMed Central

    Ames, Erik; Canter, Robert J; Grossenbacher, Steven K; Mac, Stephanie; Smith, Rachel C; Monjazeb, Arta M; Chen, Mingyi; Murphy, William J

    2015-01-01

    Natural killer (NK) cells are innate lymphocytes postulated to mediate resistance against primary haematopoietic but not solid tumor malignancies. Cancer stem cells (CSCs) are a small subset of malignant cells with stem-like properties which are resistant to chemo- and radiotherapies and are able to repopulate a tumor after cytoreductive treatments. We observed increased frequencies of stem-like tumor cells after irradiation, with increased expression of stress ligands on surviving stem-like cells. Ex vivo NK cells activated by low dose IL2 in vitro and IL15 in vivo displayed an increased ability to target solid tumor stem-like cells both in vitro and in vivo after irradiation. Mechanistically, both upregulation of stress-related ligands on the stem-like cells as well as debulking of non-stem populations contributed to these effects as determined by data from cell lines, primary tumor samples, and most relevant patient derived specimens. In addition, pretreatment of tumor-bearing mice with local radiation prior to NK transfer resulted in significantly longer survival indicating that radiation therapy in conjunction with NK cell adoptive immunotherapy targeting stem-like cancer cells may offer a promising novel radio-immunotherapy approach in the clinic. PMID:26405602

  17. Induced Pluripotent Stem Cell-Derived Natural Killer Cells for Treatment of Ovarian Cancer.

    PubMed

    Hermanson, David L; Bendzick, Laura; Pribyl, Lee; McCullar, Valarie; Vogel, Rachel Isaksson; Miller, Jeff S; Geller, Melissa A; Kaufman, Dan S

    2016-01-01

    Natural killer (NK) cells can provide effective immunotherapy for ovarian cancer. Here, we evaluated the ability of NK cells isolated from peripheral blood (PB) and NK cells derived from induced pluripotent stem cell (iPSC) to mediate killing of ovarian cancer cells in a mouse xenograft model. A mouse xenograft model was used to evaluate the intraperitoneal delivery of three different NK cell populations: iPSC-derived NK cells, PB-NK cells that had been activated and expanded in long-term culture, and overnight activated PB-NK cells that were isolated through CD3/CD19 depletion of PB B and T cells. Bioluminescent imaging was used to monitor tumor burden of luciferase expressing tumor lines. Tumors were allowed to establish prior to administering NK cells via intraperitoneal injection. These studies demonstrate a single dose of any of the three NK cell populations significantly reduced tumor burden. When mice were given three doses of either iPSC-NK cells or expanded PB-NK cells, the median survival improved from 73 days in mice untreated to 98 and 97 days for treated mice, respectively. From these studies, we conclude iPSC-derived NK cells mediate antiovarian cancer killing at least as well as PB-NK cells, making these cells a viable resource for immunotherapy for ovarian cancer. Due to their ability to be easily differentiated into NK cells and their long-term expansion potential, iPSCs can be used to produce large numbers of well-defined NK cells that can be banked and used to treat a large number of patients including treatment with multiple doses if necessary.

  18. Increased killer immunoglobulin-like receptor expression and functional defects in natural killer cells in lung cancer.

    PubMed

    Al Omar, Suliman Y; Marshall, Ernie; Middleton, Derek; Christmas, Stephen E

    2011-05-01

    Frequencies of natural killer (NK) cells from patients with non-small cell lung cancer (NSCLC) or small cell lung cancer (SCLC) did not differ from healthy controls. A higher proportion of NK cells from NSCLC patients expressed the killer immunoglobulin-like receptor (KIR) CD158b than in controls (P = 0.0004), in the presence or absence of its ligand, HLA-C1. A similar result was obtained for CD158e in the presence of its ligand HLA-Bw4 in NSCLC patients (P = 0.003); this was entirely attributable to the Bw4I group of alleles in the presence of which a fivefold higher percentage of CD158e(+) NK cells was found in NSCLC patients than controls. Proportions of CD158b(+) NK cells declined with advancing disease in NSCLC patients. Expression of NKp46, CD25 and perforin A, and production of interferon-γ following stimulation with interleukin-12 and interleukin-18, were all significantly lower in NK cells from NSCLC patients than in controls. Both NK cell cytotoxicity and granzyme B expression were also reduced in lung cancer patients. Increased inhibitory KIR expression would decrease NK cell cytotoxic function against tumour cells retaining class I HLA expression. Furthermore, the reduced ability to produce interferon-γ would restrict the ability of NK cells to stimulate T-cell responses in patients with lung cancer.

  19. Natural Killer Cell Recognition of Melanoma: New Clues for a More Effective Immunotherapy

    PubMed Central

    Tarazona, Raquel; Duran, Esther; Solana, Rafael

    2016-01-01

    Natural killer (NK) cells participate in the early immune response against melanoma and also contribute to the development of an adequate adaptive immune response by their crosstalk with dendritic cells and cytokine secretion. Melanoma resistance to conventional therapies together with its high immunogenicity justifies the development of novel therapies aimed to stimulate effective immune responses against melanoma. However, melanoma cells frequently escape to CD8 T cell recognition by the down-regulation of major histocompatibility complex (MHC) class I molecules. In this scenario, NK cells emerge as potential candidates for melanoma immunotherapy due to their capacity to recognize and destroy melanoma cells expressing low levels of MHC class I molecules. In addition, the possibility to combine immune checkpoint blockade with other NK cell potentiating strategies (e.g., cytokine induction of activating receptors) has opened new perspectives in the potential use of adoptive NK cell-based immunotherapy in melanoma. PMID:26779186

  20. Natural Killer Cell Recognition of Melanoma: New Clues for a More Effective Immunotherapy.

    PubMed

    Tarazona, Raquel; Duran, Esther; Solana, Rafael

    2015-01-01

    Natural killer (NK) cells participate in the early immune response against melanoma and also contribute to the development of an adequate adaptive immune response by their crosstalk with dendritic cells and cytokine secretion. Melanoma resistance to conventional therapies together with its high immunogenicity justifies the development of novel therapies aimed to stimulate effective immune responses against melanoma. However, melanoma cells frequently escape to CD8 T cell recognition by the down-regulation of major histocompatibility complex (MHC) class I molecules. In this scenario, NK cells emerge as potential candidates for melanoma immunotherapy due to their capacity to recognize and destroy melanoma cells expressing low levels of MHC class I molecules. In addition, the possibility to combine immune checkpoint blockade with other NK cell potentiating strategies (e.g., cytokine induction of activating receptors) has opened new perspectives in the potential use of adoptive NK cell-based immunotherapy in melanoma.

  1. Killer artificial antigen-presenting cells: a novel strategy to delete specific T cells.

    PubMed

    Schütz, Christian; Fleck, Martin; Mackensen, Andreas; Zoso, Alessia; Halbritter, Dagmar; Schneck, Jonathan P; Oelke, Mathias

    2008-04-01

    Several cell-based immunotherapy strategies have been developed to specifically modulate T cell-mediated immune responses. These methods frequently rely on the utilization of tolerogenic cell-based antigen-presenting cells (APCs). However, APCs are highly sensitive to cytotoxic T-cell responses, thus limiting their therapeutic capacity. Here, we describe a novel bead-based approach to modulate T-cell responses in an antigen-specific fashion. We have generated killer artificial APCs (kappaaAPCs) by coupling an apoptosis-inducing alpha-Fas (CD95) IgM mAb together with HLA-A2 Ig molecules onto beads. These kappaaAPCs deplete targeted antigen-specific T cells in a Fas/Fas ligand (FasL)-dependent fashion. T-cell depletion in cocultures is rapidly initiated (30 minutes), dependent on the amount of kappaaAPCs and independent of activation-induced cell death (AICD). kappaaAPCs represent a novel technology that can control T cell-mediated immune responses, and therefore has potential for use in treatment of autoimmune diseases and allograft rejection. PMID:18096763

  2. [Nasal type natural killer/T cell lymphoma: case series and literature review].

    PubMed

    Düzlü, Mehmet; Ant, Ayça; Tutar, Hakan; Karamert, Recep; Şahin, Melih; Sayar, Erolcan; Cesur, Nesibe

    2016-01-01

    Nasal type natural killer/T-cell lymphoma is a rare type of extranodal non-Hodgkin lymphoma which originates from nasal cavity and paranasal sinuses. Exact diagnosis of nasal natural killer/T-cell lymphoma, which is a rapidly progressive clinical condition, may be established by immunohistochemical analysis on biopsy material after clinical suspicion. In this article, we report four cases of nasal natural killer/T-cell lymphoma who were followed-up in our clinic and discuss the diagnosis and treatment of the disease in light of the literature data. PMID:27405082

  3. Novel targets for natural killer/T-cell lymphoma immunotherapy.

    PubMed

    Kumai, Takumi; Kobayashi, Hiroya; Harabuchi, Yasuaki

    2016-01-01

    Extranodal natural killer/T-cell lymphoma, nasal type (NKTL) is a rare but highly aggressive Epstein-Barr virus-related malignancy, which mainly occurs in nasopharyngeal and nasal/paranasal areas. In addition to its high prevalence in Asian, Central American and South American populations, its incidence rate has been gradually increasing in Western countries. The current mainstay of treatment is a combination of multiple chemotherapies and irradiation. Although chemoradiotherapy can cure NKTL, it often causes severe and fatal adverse events. Because a growing body of evidence suggests that immunotherapy is effective against hematological malignancies, this treatment could provide an alternative to chemoradiotherapy for treatment of NKTL. In this review, we focus on how recent findings could be used to develop efficient immunotherapies against NKTL. PMID:26642249

  4. Novel targets for natural killer/T-cell lymphoma immunotherapy.

    PubMed

    Kumai, Takumi; Kobayashi, Hiroya; Harabuchi, Yasuaki

    2016-01-01

    Extranodal natural killer/T-cell lymphoma, nasal type (NKTL) is a rare but highly aggressive Epstein-Barr virus-related malignancy, which mainly occurs in nasopharyngeal and nasal/paranasal areas. In addition to its high prevalence in Asian, Central American and South American populations, its incidence rate has been gradually increasing in Western countries. The current mainstay of treatment is a combination of multiple chemotherapies and irradiation. Although chemoradiotherapy can cure NKTL, it often causes severe and fatal adverse events. Because a growing body of evidence suggests that immunotherapy is effective against hematological malignancies, this treatment could provide an alternative to chemoradiotherapy for treatment of NKTL. In this review, we focus on how recent findings could be used to develop efficient immunotherapies against NKTL.

  5. Type I natural killer T cells: naturally born for fighting

    PubMed Central

    Tan, Jin-quan; Xiao, Wei; Wang, Lan; He, Yu-ling

    2010-01-01

    Type І natural killer T cells (NKT cells), a subset of CD1d-restricted T cells with invariant Vαβ TCR, are characterized by prompt production of large amounts of Th1 and/or Th2 cytokines upon primary stimulation through the TCR complex. The rapid release of cytokines implies that type І NKT cells may play a critical role in modulating the upcoming immune responses, such as anti-tumor response, protection against infection, and autoimmunity. As a bridge between innate and adaptive immunity, type І NKT cells differentiate and mature upon stimulations to achieve and maintain a homeostasis. Orchestrating with other arms of adaptive immunity, type І NKT cells show strong cytotoxic effects in response to various tumors in a direct and/or indirect manner(s). This review will focus primarily on type І NKT cell development, homeostasis, and effector functions, especially in anti-tumor immunity, and followed by their potential applications in treatment of cancers. PMID:20694020

  6. Natural killer cells regulate eosinophilic inflammation in chronic rhinosinusitis

    PubMed Central

    Kim, Ji Heui; Choi, Go Eun; Lee, Bong-Jae; Kwon, Seog Woon; Lee, Seung-Hyo; Kim, Hun Sik; Jang, Yong Ju

    2016-01-01

    Eosinophils play a major pathologic role in the pathogenesis of diverse inflammatory diseases including chronic rhinosinusitis (CRS). Dysregulated production of prostaglandin (PG), particularly PGD2, is considered to be an important contributing factor to eosinophilic inflammation in CRS primarily through proinflammatory and chemotactic effects on eosinophils. Here, we provide evidence that PGD2 can promote eosinophilic inflammation through a suppression of Natural killer (NK) cell effector function and NK cell-mediated eosinophil regulation. Eosinophil apoptosis mediated by NK cells was significantly decreased in CRS patients compared with healthy controls. This decrease was associated with NK cell dysfunction and eosinophilic inflammation. Tissue eosinophils were positively correlated with blood eosinophils in CRS patients. In a murine model of CRS, NK cell depletion caused an exacerbation of blood eosinophilia and eosinophilic inflammation in the sinonasal tissue. PGD2 and its metabolite, but not PGE2 and a panel of cytokines including TGF-β, were increased in CRS patients compared with controls. Effector functions of NK cells were potently suppressed by PGD2-dependent, rather than PGE2-dependent, pathway in controls and CRS patients. Thus, our results suggest decreased NK cell-mediated eosinophil regulation, possibly through an increased level of PGD2, as a previously unrecognized link between PG dysregulation and eosinophilic inflammation in CRS. PMID:27271931

  7. Human natural killer cell development in secondary lymphoid tissues.

    PubMed

    Freud, Aharon G; Yu, Jianhua; Caligiuri, Michael A

    2014-04-01

    For nearly a decade it has been appreciated that critical steps in human natural killer (NK) cell development likely occur outside of the bone marrow and potentially necessitate distinct microenvironments within extramedullary tissues. The latter include the liver and gravid uterus as well as secondary lymphoid tissues such as tonsils and lymph nodes. For as yet unknown reasons these tissues are naturally enriched with NK cell developmental intermediates (NKDI) that span a maturation continuum starting from an oligopotent CD34(+)CD45RA(+) hematopoietic precursor cell to a cytolytic mature NK cell. Indeed despite the detection of NKDI within the aforementioned tissues, relatively little is known about how, why, and when these tissues may be most suited to support NK cell maturation and how this process fits in with other components of the human immune system. With the discovery of other innate lymphoid subsets whose immunophenotypes overlap with those of NKDI, there is also need to revisit and potentially re-characterize the basic immunophenotypes of the stages of the human NK cell developmental pathway in vivo. In this review, we provide an overview of human NK cell development in secondary lymphoid tissues and discuss the many questions that remain to be answered in this exciting field.

  8. Optimization of natural killer T cell-mediated immunotherapy in cancer using cell-based and nanovector vaccines.

    PubMed

    Faveeuw, C; Trottein, F

    2014-03-15

    α-Galactosylceramide (α-GalCer) represents a new class of immune stimulators and vaccine adjuvants that activate type I natural killer T (NKT) cells to swiftly release cytokines and to exert helper functions for acquired immune responses. This unique property prompted clinicians to exploit the antitumor potential of NKT cells. Here, we review the effects of α-GalCer in (pre)clinics and discuss current and future strategies that aim to optimize NKT cell-mediated antitumor therapy, with a particular focus on cell-based and nanovector vaccines.

  9. Activation of Natural Killer T Cells by α-Galactosylceramide Rapidly Induces the Full Maturation of Dendritic Cells In Vivo and Thereby Acts as an Adjuvant for Combined CD4 and CD8 T Cell Immunity to a Coadministered Protein

    PubMed Central

    Fujii, Shin-ichiro; Shimizu, Kanako; Smith, Caroline; Bonifaz, Laura; Steinman, Ralph M.

    2003-01-01

    The maturation of dendritic cells (DCs) allows these antigen-presenting cells to initiate immunity. We pursued this concept in situ by studying the adjuvant action of α-galactosylceramide (αGalCer) in mice. A single i.v. injection of glycolipid induced the full maturation of splenic DCs, beginning within 4 h. Maturation was manifest by marked increases in costimulator and major histocompatibility complex class II expression, interferon (IFN)-γ production, and stimulation of the mixed leukocyte reaction. These changes were not induced directly by αGalCer but required natural killer T (NKT) cells acting independently of the MyD88 adaptor protein. To establish that DC maturation was responsible for the adjuvant role of αGalCer, mice were given αGalCer together with soluble or cell-associated ovalbumin antigen. Th1 type CD4+ and CD8+ T cell responses developed, and the mice became resistant to challenge with ovalbumin-expressing tumor. DCs from mice given ovalbumin plus adjuvant, but not the non-DCs, stimulated ovalbumin-specific proliferative responses and importantly, induced antigen-specific, IFN-γ producing, CD4+ and CD8+ T cells upon transfer into naive animals. In the latter instance, immune priming did not require further exposure to ovalbumin, αGalCer, NKT, or NK cells. Therefore a single dose of αGalCer i.v. rapidly stimulates the full maturation of DCs in situ, and this accounts for the induction of combined Th1 CD4+ and CD8+ T cell immunity to a coadministered protein. PMID:12874260

  10. Activation of natural killer T cells by alpha-galactosylceramide rapidly induces the full maturation of dendritic cells in vivo and thereby acts as an adjuvant for combined CD4 and CD8 T cell immunity to a coadministered protein.

    PubMed

    Fujii, Shin-Ichiro; Shimizu, Kanako; Smith, Caroline; Bonifaz, Laura; Steinman, Ralph M

    2003-07-21

    The maturation of dendritic cells (DCs) allows these antigen-presenting cells to initiate immunity. We pursued this concept in situ by studying the adjuvant action of alpha-galactosylceramide (alphaGalCer) in mice. A single i.v. injection of glycolipid induced the full maturation of splenic DCs, beginning within 4 h. Maturation was manifest by marked increases in costimulator and major histocompatibility complex class II expression, interferon (IFN)-gamma production, and stimulation of the mixed leukocyte reaction. These changes were not induced directly by alphaGalCer but required natural killer T (NKT) cells acting independently of the MyD88 adaptor protein. To establish that DC maturation was responsible for the adjuvant role of alphaGalCer, mice were given alphaGalCer together with soluble or cell-associated ovalbumin antigen. Th1 type CD4+ and CD8+ T cell responses developed, and the mice became resistant to challenge with ovalbumin-expressing tumor. DCs from mice given ovalbumin plus adjuvant, but not the non-DCs, stimulated ovalbumin-specific proliferative responses and importantly, induced antigen-specific, IFN-gamma producing, CD4+ and CD8+ T cells upon transfer into naive animals. In the latter instance, immune priming did not require further exposure to ovalbumin, alphaGalCer, NKT, or NK cells. Therefore a single dose of alphaGalCer i.v. rapidly stimulates the full maturation of DCs in situ, and this accounts for the induction of combined Th1 CD4+ and CD8+ T cell immunity to a coadministered protein.

  11. Potassium channels mediate killing by human natural killer cells

    SciTech Connect

    Schlichter, L.; Sidell N.; Hagiwara, S.

    1986-01-01

    Human natural killer (NK) cells in peripheral blood spontaneously recognize and kill a wide variety of target cells. It has been suggested that ion channels are involved in the killing process because there is a Ca-dependent stage and because killing by presensitized cytotoxic T lymphocytes, which in many respects resembles NK killing, is associated with changes in K and Na transport in the target cell. Using the whole-cell variation of the patch-clamp technique, the authors found a voltage-dependent potassium (K/sup +/) current in NK cells. The K/sup +/ current was reduced in a dose-dependent manner by the K-channel blockers 4-aminopyridine and quinidine and by the traditional Ca-channel blockers verapamil and Cd/sup 2 +/. They tested the effects of ion-channel blockers on killing of two commonly used target cell lines: K562, which is derived from a human myeloid leukemia, and U937, which is derived from a human histiocytic leukemia. Killing of K562 target cells, determined in a standard /sup 51/Cr-release assay, was inhibited in a dose-dependent manner by verapamil, quinidine, Cd/sup 2 +/, and 4-aminopyridine at concentrations comparable to those that blocked the K/sup +/ current in NK cells. In K562 target cells only a voltage-dependent Na= current was found and it was blocked by concentrations of tetrodotoxin that had no effect on killing. Killing of U937 target cells was also inhibited by the two ion-channel blockers tested, quinidine and verapamil. In this cell line only a small K/sup +/ current was found that was similar to the one in NK cells. The findings show that there are K channels in NK cells and that these channels play a necessary role in the killing process.

  12. Terminal Differentiation of CD56(dim)CD16(+) Natural Killer Cells Is Associated with Increase in Natural Killer Cell Frequencies After Antiretroviral Treatment in HIV-1 Infection.

    PubMed

    Ahmad, Fareed; Tufa, Dejene Milkessa; Mishra, Neha; Jacobs, Roland; Schmidt, Reinhold E

    2015-12-01

    HIV-1 infection results in immunological abnormalities of natural killer (NK) cells such as disturbed distribution of NK cell subsets and downmodulation of activating and upregulation of inhibitory receptors thereby diminishing NK cell killing capacity and cytokine secretion. Antiretroviral treatment (ART) is known to restore phenotype and functions of NK cells. However, the effects of ART on NK cell terminal differentiation, activation, and disturbed distribution have not been studied yet longitudinally. Here, we analyzed the effects of ART on these parameters of peripheral blood NK cells in a longitudinal as well as in a cross-sectional study. We observed that expanded CD56(-)CD16(+) NK cell frequency is inversely correlated with the frequency of CD56(dim)CD16(+) NK cells in treatment-naive HIV-1 patients. Loss of CD56(dim)CD16(+) and expansion of CD56(-)CD16(+) NK cells again restore to the levels of healthy controls after ART. Enhanced immune activation of different NK cell subsets is partially restored after ART. Terminal differentiation of CD56(dim)CD16(+) NK cells is enhanced after ART as measured by CD57 expression. Frequencies of CD57(+)CD56(dim)CD16(+) NK cells are directly correlated with the frequencies of total NK cells suggesting that an increase in the frequencies of CD57(+)CD56(dim)CD16(+) NK cells is reflected by increased frequencies of total NK cells after ART. Taken together these data demonstrate that ART has an effect on the immune restoration of NK cells and is enhanced in the terminal differentiation of CD56(dim)CD16(+) NK cells, which is associated with increased frequencies of total NK cells after ART.

  13. Tolerogen-induced interferon-producing killer dendritic cells (IKDCs) protect against EAE

    PubMed Central

    Huarte, Eduardo; Rynda-Apple, Agnieszka; Riccardi, Carol; Skyberg, Jerod A.; Golden, Sarah; Rollins, MaryClare F.; Ramstead, Andrew; Jackiw, Larissa O.; Maddaloni, Massimo; Pascual, David W.

    2011-01-01

    Natural killer (NK) cells and dendritic cells (DCs) have been shown to link the innate and adaptive immune systems. Likewise, a new innate cell subset, interferon-producing killer DCs (IKDCs), shares phenotypic and functional characteristics with both DCs and NK cells. Here, we show IKDCs play an essential role in the resolution of experimental autoimmune encephalomyelitis (EAE) upon treatment with the tolerizing agent, myelin oligodendrocyte glycoprotein (MOG), genetically fused to reovirus protein σ1 (termed MOG-pσ1). Activated IKDCs were recruited subsequent MOG-pσ1 treatment of EAE, and disease resolution was abated upon NK1.1 cell depletion. These IKDCs were able to kill activated CD4+ T cells and mature dendritic DCs, thus, contributing to EAE remission. In addition, IKDCs were responsible for MOG-pσ1-mediated MOG-specific regulatory T cell recruitment to the CNS. The IKDCs induced by MOG-pσ1 expressed elevated levels of HVEM for interactions with cognate ligand-positive cells: LIGHT+ NK and Teff cells and BTLA+ B cells. Further characterization revealed these activated IKDCs being MHC class IIhigh, and upon their adoptive transfer (CD11c+NK1.1+MHC class IIhigh), IKDCs, but not CD11c+NK1.1+MHC class IIintermediate/low (unactivated) cells, conferred protection against EAE. These activated IKDCs showed enhanced CD107a, PD-L1, and granzyme B expression and could present OVA, unlike unactivated IKDCs. Thus, these results demonstrate the interventional potency induced HVEM+ IKDCs to resolve autoimmune disease. PMID:22018711

  14. Membrane-Bound TRAIL Supplements Natural Killer Cell Cytotoxicity Against Neuroblastoma Cells

    PubMed Central

    Sheard, Michael A.; Asgharzadeh, Shahab; Liu, Yin; Lin, Tsen-Yin; Wu, Hong-Wei; Ji, Lingyun; Groshen, Susan; Lee, Dean A.; Seeger, Robert C.

    2013-01-01

    Neuroblastoma cells have been reported to be resistant to death induced by soluble, recombinant forms of TRAIL (CD253/TNFSF10) due to low or absent expression of caspase-8 and/or TRAIL-receptor 2 (TRAIL-R2/DR5/CD262/TNFRSF10b). However, their sensitivity to membrane-bound TRAIL on natural killer (NK) cells is not known. Comparing microarray gene expression and response to NK cell-mediated cytotoxicity, we observed a correlation between TRAIL-R2 expression and the sensitivity of fourteen neuroblastoma cell lines to the cytotoxicity of NK cells activated with IL-2 plus IL-15. Even though most NK cytotoxicity was dependent upon perforin, the cytotoxicity was supplemented by TRAIL in fourteen of seventeen (82%) neuroblastoma cell lines as demonstrated using an anti-TRAIL neutralizing antibody. Similarly, a recently developed NK cell expansion system employing IL-2 plus lethally irradiated K562 feeder cells constitutively expressing membrane-bound IL-21 (K562 clone 9.mbIL21) resulted in activated NK cells derived from normal healthy donors and neuroblastoma patients that also utilized TRAIL to supplement cytotoxicity. Exogenous IFNγ up-regulated expression of caspase-8 in three of four neuroblastoma cell lines and increased the contribution of TRAIL to NK cytotoxicity against two of the three lines; however, relatively little inhibition of cytotoxicity was observed when activated NK cells were treated with an anti-IFNγ neutralizing antibody. Constraining the binding of anti-TRAIL neutralizing antibody to membrane-bound TRAIL but not soluble TRAIL indicated that membrane-bound TRAIL alone was responsible for essentially all of the supplemental cytotoxicity. Together, these findings support a role for membrane-bound TRAIL in the cytotoxicity of NK cells against neuroblastoma cells. PMID:23719242

  15. Characterization of tumor infiltrating natural killer cell subset.

    PubMed

    Levi, Inbar; Amsalem, Hagai; Nissan, Aviram; Darash-Yahana, Merav; Peretz, Tamar; Mandelboim, Ofer; Rachmilewitz, Jacob

    2015-05-30

    The presence of tumor-infiltrating Natural Killer (NK) within a tumor bed may be indicative of an ongoing immune response toward the tumor. However, many studies have shown that an intense NK infiltration, is associated with advanced disease and may even facilitate cancer development. The exact role of the tumor infiltrating NK cells and the correlation between their presence and poor prognosis remains unclear. Interestingly, during pregnancy high numbers of a specific NK subset, CD56(bright)CD16(dim), are accumulated within first trimester deciduas. These decidual NK (dNK) cells are unique in their gene expression pattern secret angiogenic factors that induce vascular growth. In the present study we demonstrate a significant enrichment of a CD56(brigh)CD16(dim) NK cells within tumors. These NK cells express several dNK markers including VEGF. Hence, this study adds new insights into the identity of tumor residual NK cells, which has clear implications for the treatment of human cancer. PMID:26079948

  16. Expression of killer inhibitory receptors on cytotoxic cells from HIV-1-infected individuals

    PubMed Central

    Galiani, M D; Aguado, E; Tarazona, R; Romero, P; Molina, I; Santamaria, M; Solana, R; PeñA, J

    1999-01-01

    Dysfunction of cytotoxic activity of T and natural killer (NK) lymphocytes is a main immunological feature in patients with AIDS, but its basis are not well understood. It has been recently described that T and NK cell-mediated cytotoxicity can be regulated by HLA killer inhibitory receptors (KIR). In this work, we have determined on cytotoxic T cells and NK cells from HIV-1-infected individuals the expression of the following KIR molecules: p58, p70, and ILT2 (immunoglobulin-like family KIR) as well as CD94 and NKG2A (C-lectin-type family KIR). With some exceptions, no significant changes were found on the expression of immunoglobulin-like KIR in either CD8+ or CD56+ cells. Interestingly, the percentages of CD8+ and CD56+ cells expressing CD94 were significantly increased in these individuals. We also show that, in vitro, IL-10 up-regulates CD94 expression on CD8+ and CD56+ cells obtained from normal individuals, suggesting that the augmented expression observed in HIV-infected individuals could be related to the high levels of IL-10 previously described in HIV-1-infected individuals. PMID:10193420

  17. The dual-functional capability of cytokine-induced killer cells and application in tumor immunology.

    PubMed

    Zhang, Qiang; Liu, Xiao-yan; Zhang, Teng; Zhang, Xin-feng; Zhao, Lin; Long, Fei; Liu, Zhuang-kai; Wang, En-hua

    2015-05-01

    Cytokine-induced killer (CIK) cells represent a heterogeneous cell population, including a large majority of CD3+CD56+ cells, a relatively minor fractions of typical T cells (CD3+CD56-), and natural killer (NK) cells (CD3-CD56+). In order to elucidate the tumor killing mechanism of these three subpopulations of CIK cells, this review summarized the concordances and differences among CD3+CD56+ CIK cells, CD3-CD56+ NK cells and CD3+CD56- T cells to the following aspects: the effects of cell surface molecules, mechanisms of tumor killing, and clinical applications of these cells in immunotherapy. NK cells can be classified into CD56brightCD16- NK cells, which produce cytokines in response to monokine co-stimulation, and the CD56dimCD16+ NK cells, which contribute to lysing susceptible target. Also, the immunity of NK cells is mainly regulated by several immune-receptors, such as ACR, ICR, NCR and KIRs. T cells require TCR and co-stimulatory molecules for initiation of T cell activation. The CD3+CD56+ CIK cells co-express with T-cell marker CD3 and NK cell marker CD56 to appear the most potent cytotoxicity and high impact on adoptive cellular immunotherapy. These CIK subpopulations share some similar tumor killing mechanisms. LFA-1 not only mediates the binding of NK cells to target cells through its ligand ICAM-1 to localize actin accumulation but also acts as a co-stimulatory receptor on NK cells. LFA-1 also functions as co-stimulatory receptor for T cells to transmit intracellular signals from the TCR to LFA-1. Furthermore, cytotoxic effect of CD3+CD56+ CIK cells is blocked by antibodies directly against LFA-1 and its counter receptor, ICAM-1. Clinically, antibody-dependent cell-mediated cytotoxicity (ADCC) is shown in both NK cells and T cells for tumor killing while dendritic cells are another main regulator for the activation of three subpopulations. In summary, CD3+CD56+ CIK cells have dual-functional capability as T-cell subsets which acquire NK cells function

  18. Innate lymphoid cells and natural killer T cells in the gastrointestinal tract immune system.

    PubMed

    Montalvillo, Enrique; Garrote, José Antonio; Bernardo, David; Arranz, Eduardo

    2014-05-01

    The gastrointestinal tract is equipped with a highly specialized intrinsic immune system. However, the intestine is exposed to a high antigenic burden that requires a fast, nonspecific response -so-called innate immunity- to maintain homeostasis and protect the body from incoming pathogens. In the last decade multiple studies helped to unravel the particular developmental requirements and specific functions of the cells that play a role in innate immunity. In this review we shall focus on innate lymphoid cells, a newly discovered, heterogeneous set of cells that derive from an Id2-dependent lymphoid progenitor cell population. These cells have been categorized on the basis of the pattern of cytokines that they secrete, and the transcription factors that regulate their development and functions. Innate lymphoid cells play a role in the early response to pathogens, the anatomical contention of the commensal flora, and the maintenance of epithelial integrity.Amongst the various innate lymphoid cells we shall lay emphasis on a subpopulation with several peculiarities, namely that of natural killer T cells, a subset of T lymphocytes that express both T-cell and NK-cell receptors. The most numerous fraction of the NKT population are the so-called invariant NKT or iNKT cells. These iNKT cells have an invariant TCR and recognize the glycolipidic structures presented by the CD1d molecule, a homolog of class-I MHC molecules. Following activation they rapidly acquire cytotoxic activity and secrete both Th1 and Th2 cytokines, including IL-17. While their specific role is not yet established, iNKT cells take part in a great variety of intestinal immune responses ranging from oral tolerance to involvement in a number of gastrointestinal conditions.

  19. Relationship between uterine natural killer cells and unexplained repeated miscarriage

    PubMed Central

    Farghali, Mohamed M.; El-kholy, Abdel-Latif G.; Swidan, Khaled H.; Abdelazim, Ibrahim A.; Rashed, Ahmed R.; El-Sobky, Ezzat; Goma, Mostafa F.

    2015-01-01

    Objective To evaluate the relation between uterine killer (uK) cells and unexplained repeated miscarriage (RM). Material and Methods Eighty women with unexplained repeated miscarriage and missed miscarriage of current pregnancy were studied. Fetal viability and gestational age of the current pregnancy were confirmed by ultrasound, followed by suction evacuation to collect abortion specimens and uterine wall curettage to collect decidua specimens. Abortion specimens were collected for long-term monolayer cell culture and subsequent chromosome analysis using conventional G-banding. Decidua specimens were subjected to immunohistochemical staining using monoclonal antibodies specific to CD56+ and CD16+ expressed by uK cells. Results CD56+ CD16+ uK cells were found in 85% [68/80] of the studied decidua specimens of women with unexplained repeated miscarriage; 88.5% [54/61] had normal abortion karyotyping and 73.7% [14/19] had abnormal abortion karyotyping. Moreover, 73.75% [59/80] of the studied women with a past history of early miscarriage had CD56+ CD16+ uK cells in their decidua specimens, and 66.25% [53/80] of studied women with a past history of late miscarriage had CD56+ CD16+ uK cells in their decidua specimens; the association between early and late miscarriage and CD56+ CD16+ uK cells in decidua specimens was significant. Conclusion CD56+CD16+ uK cells were predominant in the decidua specimens of the studied women with repeated miscarriage. A significant association was found between the presence of CD56+ CD16+ uK cells in the studied decidua specimens and unexplained repeated miscarriage. PMID:26692771

  20. MHC class I target recognition, immunophenotypes and proteomic profiles of natural killer cells within the spleens of day-14 chick embryos

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Chicken natural killer (NK) cells are not well defined, so little is known about the molecular interactions controlling their activity. At day 14 of embryonic development, chick spleens are a rich source of T-cellfree CD8aa+, CD3_ cells with natural killing activity. Cell-mediated cytotoxicity assay...

  1. Potassium Channels Mediate Killing by Human Natural Killer Cells

    NASA Astrophysics Data System (ADS)

    Schlichter, Lyanne; Sidell, Neil; Hagiwara, Susumu

    1986-01-01

    Human natural killer (NK) cells in peripheral blood spontaneously recognize and kill a wide variety of target cells. It has been suggested that ion channels are involved in the killing process because there is a Ca-dependent stage and because killing by presensitized cytotoxic T lymphocytes, which in many respects resembles NK killing, is associated with changes in K and Na transport in the target cell. However, no direct evidence exists for ion channels in NK cells or in their target cells. Using the whole-cell variation of the patch-clamp technique, we found a voltage-dependent potassium (K+) current in NK cells. The K+ current was reduced in a dose-dependent manner by the K-channel blockers 4-aminopyridine and quinidine and by the traditional Ca-channel blockers verapamil and Cd2+. We tested the effects of ion-channel blockers on killing of two commonly used target cell lines: K562, which is derived from a human myeloid leukemia, and U937, which is derived from a human histiocytic leukemia. Killing of K562 target cells, determined in a standard 51Cr-release assay, was inhibited in a dose-dependent manner by verapamil, quinidine, Cd2+, and 4-aminopyridine at concentrations comparable to those that blocked the K+ current in NK cells. In K562 target cells only a voltage-dependent Na+ current was found and it was blocked by concentrations of tetrodotoxin that had no effect on killing. Killing of U937 target cells was also inhibited by the two ion-channel blockers tested, quinidine and verapamil. In this cell line only a small K+ current was found that was similar to the one in NK cells. We could not find any evidence of a Ca2+ current in target cells or in NK cells; therefore, our results cannot explain the Ca dependence of killing. Our findings show that there are K channels in NK cells and that these channels play a necessary role in the killing process. In contrast, the endogenous channel type in the target cell is probably not a factor in determining target cell

  2. Cytokine balance and cytokine-driven natural killer cell dysfunction in systemic juvenile idiopathic arthritis.

    PubMed

    Avau, Anneleen; Put, Karen; Wouters, Carine H; Matthys, Patrick

    2015-02-01

    Systemic juvenile idiopathic arthritis (sJIA) is a severe inflammatory childhood disorder, characterized by a specific pattern of systemic features and a typical cytokine profile. Patients are at risk to develop macrophage activation syndrome (MAS), an acute life-threatening condition defined by excessive proliferation and activation of macrophages and T cells. Defects of unknown cause in the natural killer (NK) cell cytotoxic capacity are presumed to underlie the pathogenesis of MAS and have been detected in sJIA patients. Here, we provide an overview of the cytokine profiles in sJIA and related mouse models. We discuss the influence of cytokines on NK cell function, and hypothesize that NK cell dysfunction in sJIA is caused by altered cytokine profiles.

  3. A PEPTIDE ANTAGONIST DISRUPTS NATURAL KILLER CELL INHIBITORY SYNAPSE FORMATION1

    PubMed Central

    Borhis, Gwenoline; Ahmed, Parvin S.; Mbiribindi, Bérénice; Naiyer, Mohammed M.; Davis, Daniel M; Purbhoo, Marco A; Khakoo, Salim I

    2013-01-01

    Productive engagement of MHC Class I by inhibitory NK cell receptors depends on the peptide bound by the MHC class I molecule. Peptide:MHC complexes that bind weakly to killer cell immunoglobulin-like receptors (KIR) can antagonize the inhibition mediated by high affinity peptide:MHC complexes and cause NK cell activation. We show that low affinity peptide:MHC complexes stall inhibitory signalling at the step of SHP-1 recruitment and do not go on to form the KIR microclusters induced by high affinity peptide:MHC, which are associated with Vav dephosphorylation and downstream signalling. Furthermore the low affinity peptide:MHC complexes prevented the formation of KIR microclusters by high affinity peptide:MHC. Thus peptide antagonism of NK cells is an active phenomenon of inhibitory synapse disruption. PMID:23382564

  4. Invariant natural killer T cells and their ligands: focus on multiple sclerosis

    PubMed Central

    O'Keeffe, Joan; Podbielska, Maria; Hogan, Edward L

    2015-01-01

    Invariant natural killer T (iNKT) cells are an innate population of T cells identified by the expression of an invariant T-cell receptor and reactivity to lipid-based antigens complexed with CD1d. They account for a small percentage of lymphocytes, but are extremely potent and play central roles in immunity to infection, in some cancers, and in autoimmunity. The list of relevant stimulatory lipids and glycolipid antigens now includes a range of endogenous self-antigens including the myelin-derived acetylated galactosylceramides. Recent progress in studies to identify the nature of lipid recognition for iNKT cells in autoimmune diseases like multiple sclerosis is likely to foster the development of therapeutic strategies aimed at harnessing iNKT cell activity. PMID:25976210

  5. Cytokine-induced killer cell therapy-associated idiopathic thrombocytopenic purpura: rare but noteworthy.

    PubMed

    Fu, Xiaomin; Zhang, Yong; Gao, Quanli; Lin, Jizhen; Zhang, Qinxian; Xu, Benling; Song, Yongping

    2016-09-01

    Idiopathic thrombocytopenic purpura (ITP) is characterized by a diminished platelet count, an autoimmune condition with antibodies against platelets and an increased tendency to bleed. The association between ITP and solid tumors is uncommon. Cytokine-induced killer (CIK) cell therapy is a well tolerated and promising cancer treatment with minimal toxicity. For the first time, CIK cell therapy was reported to be followed by ITP. The mechanism through which CIK induces ITP remains unclear. Imbalanced ratio of Th cells, decreased numbers or impaired function of Treg cells and excessive secretion of cytokines inducing abnormal activation of B cells may be among the possible reasons. Therefore, a better understanding of this rare condition will require further investigation of these cases. PMID:27485074

  6. Natural Killer cell activation distinguishes M. tuberculosis-mediated Immune reconstitution syndrome (IRIS) from chronic HIV and HIV-MTB co-infection

    PubMed Central

    Conradie, F.; Foulkes, A.S.; Ive, P.; Yin, X.; Roussos, K.; Glencross, D.K.; Lawrie, D.; Stevens, W.; Montaner, L.J.; Sanne; Azzoni, L.

    2011-01-01

    Background With increased access to antiretroviral treatment (ART), Immune Reconstitution Inflammatory Syndrome (IRIS) in Mycobacterium tuberculosis (MTB)-infected populations remains a clinical challenge. We studied a cross-sectional cohort of HIV-infected subjects in Johannesburg (South Africa) to help define the immune correlates that best distinguish IRIS from ongoing MTB cases. Methods We studied HIV+ subjects developing MTB-related unmasking IRIS (u-TB-IRIS) after ART initiation; control groups were HIV subjects and HIV-TB co-infected subjects with comparable ART treatment. Testing was conducted with whole blood-based 4-color flow cytometry and plasma-based Luminex cytokine assessment. Results NK cell activation, C-reactive protein and IL-8 serum concentration were significantly higher in u-TB-IRIS subjects as compared to both control groups. In addition, all MTB co-infected subjects, independent of clinical presentation, had higher neutrophils and T cell activation, together with lower lymphocytes, CD4+ T cell and myeloid DC counts. Using conditional inference tree analysis we show that elevated NK cell activation in combination with lymphocyte count characterizes the immunological profile of u-TB-IRIS. Conclusions Our results support a role for innate immune effectors in the immunopathogenesis of unmasking MTB-related IRIS, and identify new immune parameters defining this pathology. PMID:21826013

  7. Psychosocial resources, aging, and natural killer cell terminal maturity

    PubMed Central

    Segerstrom, Suzanne C.; Al-Attar, Ahmad; Lutz, Charles T.

    2012-01-01

    Psychosocial factors may influence aspects of immunological aging. The present study tested the hypothesis that psychosocial resources correlate with the expression of the cell surface maker CD57 on natural killer (NK) immune cells. CD57 is a marker of terminal maturation and senescence in this cell subset. The study further tested the relative contribution of specific resources in the social, psychological, financial, and status-skill domains, given the potential differential value of different resources for younger and older adults, and the contribution of relative vs. absolute resources. Younger (N=38) and older (N=34) women completed measures of relative and absolute resources and had blood drawn. Examined both between groups and within the older women, older age and fewer total relative resources were associated with more CD57 expression on NK cells. One SD in resources was the equivalent of 5 years of aging among the older women. Among the specific resource types, a preponderance of financial resources, both relative and absolute, was associated with less CD57 expression on NK cells, and these relationships did not significantly vary between younger and older women. There was no evidence that depressive symptoms mediated the effects of resources on CD57 expression on NK cells. These findings provide support for the hypothesis that the sense that one has substantial resources, particular with regard to finances and possessions, may retard age-associated aspects of the microenvironment in which NK cells develop and mature, independent of effects on distress, and this process may begin in younger adulthood. PMID:22708535

  8. Mature natural killer cells reset their responsiveness when exposed to an altered MHC environment

    PubMed Central

    Joncker, Nathalie T.; Shifrin, Nataliya; Delebecque, Frédéric

    2010-01-01

    Some mature natural killer (NK) cells cannot be inhibited by major histocompatibility complex (MHC) I molecules, either because they lack corresponding inhibitory receptors or because the host lacks the corresponding MHC I ligands for the receptors. Such NK cells nevertheless remain self-tolerant and exhibit a generalized hyporesponsiveness to stimulation through activating receptors. To address whether NK cell responsiveness is set only during the NK cell differentiation process, we transferred mature NK cells from wild-type (WT) to MHC I–deficient hosts or vice versa. Remarkably, mature responsive NK cells from WT mice became hyporesponsive after transfer to MHC I–deficient mice, whereas mature hyporesponsive NK cells from MHC I–deficient mice became responsive after transfer to WT mice. Altered responsiveness was evident among mature NK cells that had not divided in the recipient animals, indicating that the cells were mature before transfer and that alterations in activity did not require cell division. Furthermore, the percentages of NK cells expressing KLRG1, CD11b, CD27, and Ly49 receptors specific for H-2b were not markedly altered after transfer. Thus, the functional activity of mature NK cells can be reset when the cells are exposed to a changed MHC environment. These findings have important implications for how NK cell functions may be curtailed or enhanced in the context of disease. PMID:20819928

  9. Effect of in vivo activation of natural killer (NK) cells by a tilorone analogue on the survival of mice injected intravenously with different experimental murine tumours

    PubMed Central

    ALGARRA, I.; GONZÁLEZ, A.; PÉREZ, M.; GAFORIO, J J; GARRIDO, F.

    1996-01-01

    We studied the effect of a tilorone analogue (RMI 10,874DA) and anti-asialo GM1 serum on the survival of BALB/c and C57B1/6 mice after i.v. injections of different syngeneic murine tumour cells. Tumour lines used were different clones from chemically (GR9 wild type, GR9.B9, B7.1.B4, B7.1.B5, B7.2.38), and ultraviolet light (GRUV3)-induced sarcomas; B16 melanoma and LSTRA and YC8 lymphomas. Pretreatment of mice with tilorone inhibited metastatic colonization and increased survival significantly in all cases. In some tumour systems, the effect was attenuated when high numbers of cells were injected. Abrogation of NK cells with anti-asialo GM1 serum significantly decreased (in all tumours and at different cell doses) survival in comparison with untreated mice injected with tumours, regardless of cell dose used. These results clearly suggest that NK cell activation in vivo by the tilorone analogue we tested prolongs survival and inhibits metastasis formation in mice, even when pretreatment consists of a single dose of the analogue. PMID:8608652

  10. Therapeutic Potential and Challenges of Natural Killer Cells in Treatment of Solid Tumors

    PubMed Central

    Gras Navarro, Andrea; Björklund, Andreas T.; Chekenya, Martha

    2015-01-01

    Natural killer (NK) cells are innate lymphoid cells that hold tremendous potential for effective immunotherapy for a broad range of cancers. Due to the mode of NK cell killing, requiring one-to-one target engagement and site-directed release of cytolytic granules, the therapeutic potential of NK cells has been most extensively explored in hematological malignancies. However, their ability to precisely kill antibody coated cells, cancer stem cells, and genotoxically altered cells, while maintaining tolerance to healthy cells makes them appealing therapeutic effectors for all cancer forms, including metastases. Due to their release of pro-inflammatory cytokines, NK cells may potently reverse the anti-inflammatory tumor microenvironment (TME) and augment adaptive immune responses by promoting differentiation, activation, and/or recruitment of accessory immune cells to sites of malignancy. Nevertheless, integrated and coordinated mechanisms of subversion of NK cell activity against the tumor and its microenvironment exist. Although our understanding of the receptor ligand interactions that regulate NK cell functionality has evolved remarkably, the diversity of ligands and receptors is complex, as is their mechanistic foundations in regulating NK cell function. In this article, we review the literature and highlight how the TME manipulates the NK cell phenotypes, genotypes, and tropism to evade tumor recognition and elimination. We discuss counter strategies that may be adopted to augment the efficacy of NK cell anti-tumor surveillance, the clinical trials that have been undertaken so far in solid malignancies, critically weighing the challenges and opportunities with this approach. PMID:25972872

  11. CD45 Isoform Profile Identifies Natural Killer (NK) Subsets with Differential Activity

    PubMed Central

    Krzywinska, Ewelina; Cornillon, Amelie; Allende-Vega, Nerea; Vo, Dang-Nghiem; Rene, Celine; Lu, Zhao-Yang; Pasero, Christine; Olive, Daniel; Fegueux, Nathalie; Ceballos, Patrick; Hicheri, Yosr; Sobecki, Michal; Rossi, Jean-François; Cartron, Guillaume; Villalba, Martin

    2016-01-01

    The leucocyte-specific phosphatase CD45 is present in two main isoforms: the large CD45RA and the short CD45RO. We have recently shown that distinctive expression of these isoforms distinguishes natural killer (NK) populations. For example, co-expression of both isoforms identifies in vivo the anti tumor NK cells in hematological cancer patients. Here we show that low CD45 expression associates with less mature, CD56bright, NK cells. Most NK cells in healthy human donors are CD45RA+CD45RO-. The CD45RA-RO+ phenotype, CD45RO cells, is extremely uncommon in B or NK cells, in contrast to T cells. However, healthy donors possess CD45RAdimRO- (CD45RAdim cells), which show immature markers and are largely expanded in hematopoietic stem cell transplant patients. Blood borne cancer patients also have more CD45RAdim cells that carry several features of immature NK cells. However, and in opposition to their association to NK cell progenitors, they do not proliferate and show low expression of the transferrin receptor protein 1/CD71, suggesting low metabolic activity. Moreover, CD45RAdim cells properly respond to in vitro encounter with target cells by degranulating or gaining CD69 expression. In summary, they are quiescent NK cells, with low metabolic status that can, however, respond after encounter with target cells. PMID:27100180

  12. Protein Translation and Cell Death: The Role of Rare tRNAs in Biofilm Formation and in Activating Dormant Phage Killer Genes

    PubMed Central

    García-Contreras, Rodolfo; Zhang, Xue-Song; Kim, Younghoon; Wood, Thomas K.

    2008-01-01

    We discovered previously that the small Escherichia coli proteins Hha (hemolysin expression modulating protein) and the adjacent, poorly-characterized YbaJ are important for biofilm formation; however, their roles have been nebulous. Biofilms are intricate communities in which cell signaling often converts single cells into primitive tissues. Here we show that Hha decreases biofilm formation dramatically by repressing the transcription of rare codon tRNAs which serves to inhibit fimbriae production and by repressing to some extent transcription of fimbrial genes fimA and ihfA. In vivo binding studies show Hha binds to the rare codon tRNAs argU, ileX, ileY, and proL and to two prophage clusters D1P12 and CP4-57. Real-time PCR corroborated that Hha represses argU and proL, and Hha type I fimbriae repression is abolished by the addition of extra copies of argU, ileY, and proL. The repression of transcription of rare codon tRNAs by Hha also leads to cell lysis and biofilm dispersal due to activation of prophage lytic genes rzpD, yfjZ, appY, and alpA and due to induction of ClpP/ClpX proteases which activate toxins by degrading antitoxins. YbaJ serves to mediate the toxicity of Hha. Hence, we have identified that a single protein (Hha) can control biofilm formation by limiting fimbriae production as well as by controlling cell death. The mechanism used by Hha is the control of translation via the availability of rare codon tRNAs which reduces fimbriae production and activates prophage lytic genes. Therefore, Hha acts as a toxin in conjunction with co-transcribed YbaJ (TomB) that attenuates Hha toxicity. PMID:18545668

  13. Immune checkpoint inhibitors enhance cytotoxicity of cytokine-induced killer cells against human myeloid leukaemic blasts.

    PubMed

    Poh, Su Li; Linn, Yeh Ching

    2016-05-01

    We studied whether blockade of inhibitory receptors on cytokine-induced killer (CIK) cells by immune checkpoint inhibitors could increase its anti-tumour potency against haematological malignancies. CIK cultures were generated from seven normal donors and nine patients with acute myeloid leukaemia (AML), acute lymphoblastic leukaemia (ALL) or multiple myeloma (MM). The inhibitory receptors B and T lymphocyte attenuator, CD200 receptor, lymphocyte activation gene-3 (LAG-3) and T cell immunoglobulin and mucin-domain-containing-3 (TIM-3) were present at variable percentages in most CIK cultures, while cytotoxic T lymphocyte-associated protein 4 (CTLA-4), programmed death-1 (PD-1) and killer cell immunoglobulin-like receptors (KIR2DL1/2/3) were expressed at low level in most cultures. Without blockade, myeloid leukaemia cells were susceptible to autologous and allogeneic CIK-mediated cytotoxicity. Blockade of KIR, LAG-3, PD-1 and TIM-3 but not CTLA-4 resulted in remarkable increase in killing against these targets, even in those with poor baseline cytotoxicity. ALL and MM targets were resistant to CIK-mediated cytotoxicity, and blockade of receptors did not increase cytotoxicity to a meaningful extent. Combination of inhibitors against two receptors did not further increase cytotoxicity. Interestingly, potentiation of CIK killing by blocking antibodies was not predicted by expression of receptors on CIK and their respective ligands on the targets. Compared to un-activated T and NK cells, blockade potentiated the cytotoxicity of CIK cells to a greater degree and at a lower E:T ratio, but without significant increase in cytotoxicity against normal white cell. Our findings provide the basis for clinical trial combining autologous CIK cells with checkpoint inhibitors for patients with AML.

  14. Adoptive Cell Therapy of Melanoma with Cytokine-induced Killer Cells

    PubMed Central

    Kim, Ji Sung; Kim, Yong Guk; Pyo, Minji; Lee, Hong Kyung; Hong, Jin Tae; Kim, Youngsoo

    2015-01-01

    Melanoma is the most aggressive skin cancer and its incidence is gradually increasing worldwide. Patients with metastatic melanoma have a very poor prognosis (estimated 5-year survival rate of <16%). In the last few years, several drugs have been approved for malignant melanoma, such as tyrosine kinase inhibitors and immune checkpoint blockades. Although new therapeutic agents have improved progression-free and overall survival, their use is limited by drug resistance and drug-related toxicity. At the same time, adoptive cell therapy of metastatic melanoma with tumor-infiltrating lymphocytes has shown promising results in preclinical and clinical studies. In this review, we summarize the currently available drugs for treatment of malignant melanoma. In addition, we suggest cytokine-induced killer (CIK) cells as another candidate approach for adoptive cell therapy of melanoma. Our preclinical study and several previous studies have shown that CIK cells have potent anti-tumor activity against melanomas in vitro and in an in vivo human tumor xenograft model without any toxicity. PMID:25922594

  15. Murine cytomegalovirus stimulates natural killer cell function but kills genetically resistant mice treated with radioactive strontium

    SciTech Connect

    Masuda, A.; Bennett, M.

    1981-12-01

    Treatment of C3H/St mice with 100 microCi of 89Sr weakened their genetic resistance to murine cytomegalovirus (MCMV) infection. The criteria utilized to detect increased susceptibility were: (i) survival of mice; (ii) numbers of MCMV-infected cells in the spleens and liver; and (iii) serum glutamic pyruvic transaminase levels. The natural killer (NK) cell activity of spleen cells from mice treated with 89Sr is very low. However, the NK activities of spleen cells of both normal and 89Sr-treated mice were greatly augmented 3 days after infection with MCMV. These NK cells lysed a variety of tumor cells and shared several features with conventional NK cells, but were not lysed by anti-Nk-1.2 serum (specific for NK cells) plus complement. Splenic adherent cells did not lyse tumor cells themselves but were necessary for the stimulation of NK cells by MCMV. The paradox of high NK cell function and poor survival in 89Sr-treated mice infected with MCMV was a surprise. We conclude that these augmented NK cells, of themselves, cannot account for the genetic resistance of C3H/St mice to infection with MCMV.

  16. 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

  17. CXCL16-positive dendritic cells enhance invariant natural killer T cell-dependent IFNγ production and tumor control

    PubMed Central

    Veinotte, Linnea; Gebremeskel, Simon; Johnston, Brent

    2016-01-01

    ABSTRACT Crosstalk interactions between dendritic cells (DCs) and invariant natural killer T (iNKT) cells are important in regulating antitumor responses elicited by glycolipid antigens. iNKT cells constitutively express the chemokine receptor CXCR6, while cytokine-activated DCs upregulate the transmembrane chemokine ligand, CXCL16. This study examined the co-stimulatory role of CXCR6/CXCL16 interactions in glycolipid-dependent iNKT cell activation and tumor control. Spleen and liver DCs in wild-type mice, but not iNKT cell deficient (Jα18−/−) mice, transiently upregulated surface CXCL16 following in vivo administration of the glycolipid antigen α-galactosylceramide. Recombinant CXCL16 did not directly induce iNKT cell activation in vitro but enhanced interferon (IFN)-γ production when mouse or human iNKT cells were stimulated with plate-bound anti-CD3. Compared with glycolipid-loaded CXCL16neg DCs, CXCL16hi DCs induced higher levels of IFNγ production in iNKT cell cultures and following adoptive transfer in vivo. The number of IFNγ+ iNKT cells and expansion of T-bet+ iNKT cells were reduced in vivo when CXCL16−/− DCs were used to activate iNKT cells. Enhanced IFNγ production in vivo was not dependent on CXCR6 expression on natural killer (NK) cells. Adoptive transfer of glycolipid-loaded CXCL16hi DCs provided superior protection against tumor metastasis compared to CXCL16neg DC transfers. Similarly, wild-type DCs provided superior protection against metastasis compared with CXCL16−/− DCs. These experiments implicate an important role for CXCR6/CXCL16 interactions in regulating iNKT cell IFNγ production and tumor control. The selective use of CXCL16hi DCs in adoptive transfer immunotherapies may prove useful for enhancing T helper (Th) type 1 responses and clinical outcomes in cancer patients. PMID:27471636

  18. ImuVert activation of natural killer cytotoxicity and interferon gamma production via CD16 triggering.

    PubMed

    Cunningham-Rundles, S; Pearson, F C

    1990-01-01

    The effect and mechanism of action of ImuVert, a new biological response modifier consisting of ribosomes and natural membrane vesicles of Serratia marcescens, on endogenous natural killer (NK) cells and activated NK activity has been analyzed. The studies showed that endogenous NK activity of peripheral blood mononuclear cells (PBMC) from normal cell donors was significantly increased (P less than 0.03) against K562, U937, and Molt-4 target cells. PBMC from cord blood of newborn infants lacking NK activity were upregulated (1.5-4 fold over endogenous NK activity) by ImuVert. Other studies showed that the abnormal NK activity of PBMC from patients with the human immunodeficiency virus (HIV) infection was significantly augmented in vitro (P less than 0.01) by ImuVert. ImuVert strongly stimulated interferon gamma production and in combination with interleukin 2 produced synergistically enhanced interferon gamma production and greater cytotoxicity than that induced by either alone. Studies on lymphocyte differentiation antigen expression following treatment with ImuVert indicated that ImuVert triggers interferon gamma production through binding the low affinity IgG Fc receptor, type III, CD16. The studies suggest that ImuVert may trigger interferon gamma production by binding to the Fc receptor and that the amplitude of the ensuing reaction and the ability of ImuVert to induce cytotoxicity in a setting where this activity has been down regulated is based on the absence of suppressor activation or direct contra suppressor activity.

  19. An Optimized Method for Isolating and Expanding Invariant Natural Killer T Cells from Mouse Spleen.

    PubMed

    Govindarajan, Srinath; Elewaut, Dirk; Drennan, Michael

    2015-01-01

    The ability to rapidly secrete cytokines upon stimulation is a functional characteristic of the invariant natural killer T (iNKT) cell lineage. iNKT cells are therefore characterized as an innate T cell population capable of activating and steering adaptive immune responses. The development of improved techniques for the culture and expansion of murine iNKT cells facilitates the study of iNKT cell biology in in vitro and in vivo model systems. Here we describe an optimized procedure for the isolation and expansion of murine splenic iNKT cells. Spleens from C57Bl/6 mice are removed, dissected and strained and the resulting cellular suspension is layered over density gradient media. Following centrifugation, splenic mononuclear cells (MNCs) are collected and CD5-positive (CD5(+)) lymphocytes are enriched for using magnetic beads. iNKT cells within the CD5(+) fraction are subsequently stained with αGalCer-loaded CD1d tetramer and purified by fluorescence activated cell sorting (FACS). FACS sorted iNKT cells are then initially cultured in vitro using a combination of recombinant murine cytokines and plate-bound T cell receptor (TCR) stimuli before being expanded in the presence of murine recombinant IL-7. Using this technique, approximately 10(8) iNKT cells can be generated within 18-20 days of culture, after which they can be used for functional assays in vitro, or for in vivo transfer experiments in mice. PMID:26555769

  20. Super natural killer cells that target metastases in the tumor draining lymph nodes.

    PubMed

    Chandrasekaran, Siddarth; Chan, Maxine F; Li, Jiahe; King, Michael R

    2016-01-01

    Tumor draining lymph nodes are the first site of metastasis in most types of cancer. The extent of metastasis in the lymph nodes is often used in staging cancer progression. We previously showed that nanoscale TRAIL liposomes conjugated to human natural killer cells enhance their endogenous therapeutic potential in killing cancer cells cultured in engineered lymph node microenvironments. In this work, it is shown that liposomes decorated with apoptosis-inducing ligand TRAIL and an antibody against a mouse natural killer cell marker are carried to the tumor draining inguinal lymph nodes and prevent the lymphatic spread of a subcutaneous tumor in mice. It is shown that targeting natural killer cells with TRAIL liposomes enhances their retention time within the tumor draining lymph nodes to induce apoptosis in cancer cells. It is concluded that this approach can be used to kill cancer cells within the tumor draining lymph nodes to prevent the lymphatic spread of cancer.

  1. Attenuation of natural killer cell functions by capsaicin through a direct and TRPV1-independent mechanism.

    PubMed

    Kim, Hun Sik; Kwon, Hyung-Joon; Kim, Gye Eun; Cho, Mi-Hyang; Yoon, Seung-Yong; Davies, Alexander J; Oh, Seog Bae; Lee, Heuiran; Cho, Young Keol; Joo, Chul Hyun; Kwon, Seog Woon; Kim, Sun Chang; Kim, Yoo Kyum

    2014-07-01

    The assessment of the biological activity of capsaicin, the compound responsible for the spicy flavor of chili pepper, produced controversial results, showing either carcinogenicity or cancer prevention. The innate immune system plays a pivotal role in cancer pathology and prevention; yet, the effect of capsaicin on natural killer (NK) cells, which function in cancer surveillance, is unclear. This study found that capsaicin inhibited NK cell-mediated cytotoxicity and cytokine production (interferon-γ and tumor necrosis factor-α). Capsaicin impaired the cytotoxicity of NK cells, thereby inhibiting lysis of standard target cells and gastric cancer cells by modulating calcium mobilization in NK cells. Capsaicin also induced apoptosis in gastric cancer cells, but that effect required higher concentrations and longer exposure times than those required to trigger NK cell dysfunction. Furthermore, capsaicin inhibited the cytotoxicity of isolated NK cells and of an NK cell line, suggesting a direct effect on NK cells. Antagonists of transient receptor potential vanilloid subfamily member 1 (TRPV1), a cognate capsaicin receptor, or deficiency in TRPV1 expression failed to prevent the defects induced by capsaicin in NK cells expressing functional TRPV1. Thus, the mechanism of action of capsaicin on NK cells is largely independent of TRPV1. Taken together, capsaicin may have chemotherapeutic potential but may impair NK cell function, which plays a central role in tumor surveillance. PMID:24743513

  2. Natural killer cell subsets in cerebrospinal fluid of patients with multiple sclerosis

    PubMed Central

    Rodríguez-Martín, E; Picón, C; Costa-Frossard, L; Alenda, R; Sainz de la Maza, S; Roldán, E; Espiño, M; Villar, L M; Álvarez-Cermeño, J C

    2015-01-01

    Changes in blood natural killer (NK) cells, important players of the immune innate system, have been described in multiple sclerosis (MS). We studied percentages and total cell counts of different effector and regulatory NK cells in cerebrospinal fluid (CSF) of MS patients and other neurological diseases to gain clearer knowledge of the role of these cells in neuroinflammation. NK cell subsets were assessed by flow cytometry in CSF of 85 consecutive MS patients (33 with active disease and 52 with stable MS), 16 with other inflammatory diseases of the central nervous system (IND) and 17 with non-inflammatory neurological diseases (NIND). MS patients showed a decrease in percentages of different CSF NK subpopulations compared to the NIND group. However, absolute cell counts showed a significant increase of all NK subsets in MS and IND patients, revealing that the decrease in percentages does not reflect a real reduction of these immune cells. Remarkably, MS patients showed a significant increase of regulatory/effector (CD56bright/CD56dim) NK ratio compared to IND and NIND groups. In addition, MS activity associated with an expansion of NK T cells. These data show that NK cell subsets do not increase uniformly in all inflammatory neurological disease and suggest strongly that regulatory CD56bright and NK T cells may arise in CSF of MS patients as an attempt to counteract the CNS immune activation characteristic of the disease. PMID:25565222

  3. Natural killer cell subsets in cerebrospinal fluid of patients with multiple sclerosis.

    PubMed

    Rodríguez-Martín, E; Picón, C; Costa-Frossard, L; Alenda, R; Sainz de la Maza, S; Roldán, E; Espiño, M; Villar, L M; Álvarez-Cermeño, J C

    2015-05-01

    Changes in blood natural killer (NK) cells, important players of the immune innate system, have been described in multiple sclerosis (MS). We studied percentages and total cell counts of different effector and regulatory NK cells in cerebrospinal fluid (CSF) of MS patients and other neurological diseases to gain clearer knowledge of the role of these cells in neuroinflammation. NK cell subsets were assessed by flow cytometry in CSF of 85 consecutive MS patients (33 with active disease and 52 with stable MS), 16 with other inflammatory diseases of the central nervous system (IND) and 17 with non-inflammatory neurological diseases (NIND). MS patients showed a decrease in percentages of different CSF NK subpopulations compared to the NIND group. However, absolute cell counts showed a significant increase of all NK subsets in MS and IND patients, revealing that the decrease in percentages does not reflect a real reduction of these immune cells. Remarkably, MS patients showed a significant increase of regulatory/effector (CD56(bright) /CD56(dim) ) NK ratio compared to IND and NIND groups. In addition, MS activity associated with an expansion of NK T cells. These data show that NK cell subsets do not increase uniformly in all inflammatory neurological disease and suggest strongly that regulatory CD56(bright) and NK T cells may arise in CSF of MS patients as an attempt to counteract the CNS immune activation characteristic of the disease. PMID:25565222

  4. Natural killer cell subsets in cerebrospinal fluid of patients with multiple sclerosis.

    PubMed

    Rodríguez-Martín, E; Picón, C; Costa-Frossard, L; Alenda, R; Sainz de la Maza, S; Roldán, E; Espiño, M; Villar, L M; Álvarez-Cermeño, J C

    2015-05-01

    Changes in blood natural killer (NK) cells, important players of the immune innate system, have been described in multiple sclerosis (MS). We studied percentages and total cell counts of different effector and regulatory NK cells in cerebrospinal fluid (CSF) of MS patients and other neurological diseases to gain clearer knowledge of the role of these cells in neuroinflammation. NK cell subsets were assessed by flow cytometry in CSF of 85 consecutive MS patients (33 with active disease and 52 with stable MS), 16 with other inflammatory diseases of the central nervous system (IND) and 17 with non-inflammatory neurological diseases (NIND). MS patients showed a decrease in percentages of different CSF NK subpopulations compared to the NIND group. However, absolute cell counts showed a significant increase of all NK subsets in MS and IND patients, revealing that the decrease in percentages does not reflect a real reduction of these immune cells. Remarkably, MS patients showed a significant increase of regulatory/effector (CD56(bright) /CD56(dim) ) NK ratio compared to IND and NIND groups. In addition, MS activity associated with an expansion of NK T cells. These data show that NK cell subsets do not increase uniformly in all inflammatory neurological disease and suggest strongly that regulatory CD56(bright) and NK T cells may arise in CSF of MS patients as an attempt to counteract the CNS immune activation characteristic of the disease.

  5. A species of human alpha interferon that lacks the ability to boost human natural killer activity.

    PubMed Central

    Ortaldo, J R; Herberman, R B; Harvey, C; Osheroff, P; Pan, Y C; Kelder, B; Pestka, S

    1984-01-01

    Most species of recombinant leukocyte interferons (IFN-alpha A, -alpha B, -alpha C, -alpha D, -alpha F, -alpha I, and -alpha K) were capable of boosting human natural killer (NK) activity after a 2-hr treatment of cells at a concentration of 1-80 units/ml. In contrast, recombinant human IFN-alpha J was found to be incapable of augmenting NK activity after exposure of cells for 2 hr to concentrations as high as 10,000 units/ml. This inability of IFN-alpha J to boost NK activity was not complete because, after exposure of cells to a high concentration of IFN-alpha J (10,000 units/ml) for 18 hr, boosting of cytolysis was observed. IFN-alpha J appeared to interact with receptors for IFN on NK cells since it was found to interfere with the boosting of NK activity by other species of IFN-alpha. In contrast to its deficient ability to augment NK activity, IFN-alpha J has potent antiviral and antiproliferative activities. Such extensive dissociation of these biological activities has not been observed previously with any other natural or recombinant IFN species. Thus, this IFN species may be useful for evaluating the relative importance of various biological activities on the therapeutic effects of IFN, for understanding structure-function relationships, and for determining the biochemical pathways related to the various biological effects of IFN. PMID:6589637

  6. Natural Killer Cells Mediate Protection against Yersinia pseudotuberculosis in the Mesenteric Lymph Nodes

    PubMed Central

    Rosenheinrich, Maik; Heine, Wiebke; Schmühl, Carina M.; Pisano, Fabio; Dersch, Petra

    2015-01-01

    Natural killer cells play a crucial role in the initial defense against bacterial pathogens. The crosstalk between host cells infected with intracellular pathogens and NK cells has been studied intensively, but not much attention has been given to characterize the role of NK cells in the response to extracellular bacterial pathogens such as yersiniae. In this study we used antibody-mediated NK cell depletion to address the importance of this immune cell type in controlling a Y. pseudotuberculosis infection. Analysis of the bacterial counts was used to follow the infection and flow cytometry was performed to characterize the composition and dynamic of immune cells. Depletion of NK cells led to higher bacterial loads within the mesenteric lymph nodes. We further show that in particular CD11b+ CD27+ NK cells which express higher levels of the activation marker CD69 increase within the mesenteric lymph nodes during a Y. pseudotuberculosis infection. Moreover, in response to the activation NK cells secrete higher levels of IFNy, which in turn triggers the production of the proinflammatory cytokine TNFα. These results suggest, that NK cells aid in the clearance of Y. pseudotuberculosis infections mainly by triggering the expression of proinflammatory cytokines manipulating the host immune response. PMID:26296209

  7. INCREASES IN CYTOSOLIC CALCIUM ION LEVELS IN HUMAN NATURAL KILLER CELLS IN RESPONSE TO BUTYLTIN EXPOSURE

    PubMed Central

    Lane, Rhonda; Ghazi, Sabah O.; Whalen, Margaret M.

    2009-01-01

    This study investigated whether exposures to butyltins (BTs), tributylin (TBT) and dibutyltin (DBT) were able to alter cytosolic calcium levels in human natural killer (NK) cells. Additionally, the effects of cytosolic calcium ion increases on the activation state of mitogen activated protein kinases (MAPKs) in NK cells were also investigated. NK cells are an intital immune defense against the development of tumors or viral infections. TBT and DBT are widespread environmental contaminants, due to their various industrial applications. Both TBT and DBT have been shown to decrease the ability of NK cells to lyse tumor cells (lytic function). TBT has also been shown to activate MAPKs in NK cells. The results of this study indicated that TBT increased cytosolic calcium levels by as much as 100% after a 60 min exposure to 500 nM TBT while DBT increased cytosolic calcium levels to a much smaller extent (and required higher concentrations). The results also indicated that increases in cytosolic calcium could activate MAPKs but only for a short period of time (5 min), while previous studies showed that activation of MAPKs by TBT last for at least 6 hours. Thus, it appears that TBT stimulated increases in cytosolic calcium may contribute to, but are not fully responsible for, TBT-induced activation of MAPKs. PMID:19365649

  8. Regulation of human natural killer cell migration and proliferation by the exodus subfamily of CC chemokines.

    PubMed

    Robertson, M J; Williams, B T; Christopherson, K; Brahmi, Z; Hromas, R

    2000-01-10

    Natural killer (NK) cells play an important role in innate and adaptive immune responses to obligate intracellular pathogens. Nevertheless, the regulation of NK cell trafficking and migration to inflammatory sites is poorly understood. Exodus-1/MIP-3alpha/LARC, Exodus-2/6Ckine/SLC, and Exodus-3/MIP-3beta/ELC/CKbeta-11 are CC chemokines that share a unique aspartate-cysteine-cysteine-leucine motif near their amino terminus and preferentially stimulate the migration of T lymphocytes. The effects of Exodus chemokines on human NK cells were examined. Exodus-1, -2, and -3 did not induce detectable chemotaxis of resting peripheral blood NK cells. In contrast, Exodus-2 and -3 stimulated migration of polyclonal activated peripheral blood NK cells in a dose-dependent fashion. Exodus-2 and -3 also induced dose-dependent chemotaxis of NKL, an IL-2-dependent human NK cell line. Results of modified checkerboard assays indicate that migration of NKL cells in response to Exodus-2 and -3 represents true chemotaxis and not simply chemokinesis. Exodus-1, -2, and -3 did not induce NK cell proliferation in the absence of other stimuli. Nevertheless, Exodus-2 and -3 significantly augmented IL-2-induced proliferation of normal human CD56(dim) NK cells. In contrast, Exodus-1, -2, and -3 did not affect the cytolytic activity of resting or activated peripheral blood NK cells. Expression of message for CCR7, a shared receptor for Exodus-2 and -3, was detected in activated polyclonal NK cells and NKL cells but not resting NK cells. Taken together, these results indicate that Exodus-2 and -3 can participate in the recruitment and proliferation of activated NK cells. Exodus-2 and -3 may regulate interactions between T cells and NK cells that are crucial for the generation of optimal immune responses.

  9. Gene Modification of Human Natural Killer Cells Using a Retroviral Vector.

    PubMed

    Kellner, Joshua N; Cruz, Conrad R; Bollard, Catherine M; Yvon, Eric S

    2016-01-01

    As part of the innate immune system, natural killer (NK) cells are regarded as promising effector cells for adoptive cell therapy approaches to treat patients with cancer. In some cases, genetic modification of the NK cells may be considered but such manipulation has to be integrated into the expansion method to allow the generation of clinically relevant numbers of gene-modified NK cells. Therefore, an efficient gene transfer procedure is needed.Our group developed a retrovirus-based transduction protocol capable of robust expansion of gene-modified NK cells with a high rate of transgene expression. Actively dividing cells is a prerequisite for efficient gene transfer when using a retroviral vector. In the procedure presented here, strong activation of the NK cells was provided by a combination of IL-15 and the K-562 feeder cells. Beside the interest in developing a simple procedure compliant with good manufacturing practice (GMP) for the production of therapeutic products, this approach also provides a valuable means of generating genetically modified primary NK cells for future preclinical studies. PMID:27177668

  10. Regulation and antimetastatic functions of liver-associated natural killer cells.

    PubMed

    Wiltrout, R H

    2000-04-01

    The liver is a complex organ composed of hepatic parenchymal cells and a variety of non-parenchymal cells that consist of endothelial cells, Kupffer cells, and several subsets of resident lymphocytes, including natural killer (NK), T, and NK1.1+/CD3+ (NK/T) cells. The regulation of these various lymphoid subpopulations and their relative contributions to antiviral, antitumor and pathogenic inflammatory responses in the liver remain topics of much interest. Studies from our laboratory have shown that various immune stimulants and cytokines can augment liver-associated NK activity at least partially through the mobilization of NK cells from the bone marrow to the liver. The mobilization process can be dependent on the induction of interferon (IFN)-gamma and/or tumor necrosis factor-alpha and on very late activation antigen-4/vascular cell adhesion molecule-1 interaction. The induction of IFN-gamma by cytokines such as interleukin (IL)-12 also rapidly triggers the induction of chemokine genes in parenchymal cells that may contribute to the localization of NK and T cells. Both IL-2 and IL-12 trigger changes in the number and functions of liver-associated leukocyte subsets, and induce antimetastatic effects that are likely mediated through several direct and indirect mechanisms. The overall goal of these studies is to understand the interactions and functions of liver-associated NK1.1+ cells in the context of innate and adaptive immune responses to neoplasia.

  11. Cytotoxic function of umbilical cord blood natural killer cells: relevance to adoptive immunotherapy.

    PubMed

    Lin, Syh-Jae; Kuo, Ming-Ling

    2011-11-01

    Decreased graft-versus-host disease (GVHD), ease of accessibility, and sustained engraftment encourage the use of umbilical cord blood (UCB) as an alternative source to bone marrow for immune reconstitution in children with leukemia. Natural killer (NK) cells rapidly expand after stem cell transplantation and are important for regulating GVHD and providing graft-versus-leukemia (GVL) effects. This review highlights the phenotypic and functional differences between UCB NK cells and adult peripheral blood (APB) NK cells, and discusses the possible therapeutic benefit of using UCB NK cells for adoptive immunotherapy in leukemia. Alloreactive NK cells show potent cytotoxic activities against human leukocyte antigen (HLA)-nonidentical leukemic cells and reduce leukemia relapses. The higher numbers of NK progenitors in UCB makes it a convenient source for ex vivo expansion of UCB NK cells for posttransplant treatment. UCB NK cells readily respond to interleukin-15, which may greatly enhance their antitumor effect. Activation and expansion protocols for UCB NK cells are currently being developed.

  12. Rapid reuptake of granzyme B leads to emperitosis: an apoptotic cell-in-cell death of immune killer cells inside tumor cells.

    PubMed

    Wang, S; He, M-f; Chen, Y-h; Wang, M-y; Yu, X-M; Bai, J; Zhu, H-y; Wang, Y-y; Zhao, H; Mei, Q; Nie, J; Ma, J; Wang, J-f; Wen, Q; Ma, L; Wang, Y; Wang, X-n

    2013-01-01

    A cell-in-cell process refers to the invasion of one living cell into another homotypic or heterotypic cell. Different from non-apoptotic death processes of internalized cells termed entosis or cannibalism, we previously reported an apoptotic cell-in-cell death occurring during heterotypic cell-in-cell formation. In this study, we further demonstrated that the apoptotic cell-in-cell death occurred only in internalized immune killer cells expressing granzyme B (GzmB). Vacuole wrapping around the internalized cells inside the target cells was the common hallmark during the early stage of all cell-in-cell processes, which resulted in the accumulation of reactive oxygen species and subsequent mitochondrial injury of encapsulated killer or non-cytotoxic immune cells. However, internalized killer cells mediated rapid bubbling of the vacuoles with the subsequent degranulation of GzmB inside the vacuole of the target cells and underwent the reuptake of GzmB by killer cells themselves. The confinement of GzmB inside the vacuole surpassed the lysosome-mediated cell death occurring in heterotypic or homotypic entosis processes, resulting in a GzmB-triggered caspase-dependent apoptotic cell-in-cell death of internalized killer cells. On the contrary, internalized killer cells from GzmB-deficient mice underwent a typical non-apoptotic entotic cell-in-cell death similar to that of non-cytotoxic immune cells or tumor cells. Our results thus demonstrated the critical involvement of immune cells with cytotoxic property in apoptotic cell-in-cell death, which we termed as emperitosis taken from emperipolesis and apoptosis. Whereas entosis or cannibalism may serve as a feed-on mechanism to exacerbate and nourish tumor cells, emperitosis of immune killer cells inside tumor cells may serve as an in-cell danger sensation model to prevent the killing of target cells from inside, implying a unique mechanism for tumor cells to escape from immune surveillance. PMID:24113190

  13. Interaction of natural killer cells with neutrophils exerts a significant antitumor immunity in hematopoietic stem cell transplantation recipients.

    PubMed

    Ueda, Ryosuke; Narumi, Kenta; Hashimoto, Hisayoshi; Miyakawa, Reina; Okusaka, Takuji; Aoki, Kazunori

    2016-01-01

    Autologous hematopoietic stem cell transplantation (HSCT) can induce a strong antitumor immunity by homeostatic proliferation (HP) of T cells and suppression of regulatory T cells following preconditioning-induced lymphopenia. However, the role of innate immunity including natural killer (NK) cells is still not understood. Here, first, we examined whether NK cells exert an antitumor effect after syngeneic HSCT in a murine colon cancer model. Flow cytometry showed that NK cells as well as T cells rapidly proliferated after HSCT, and the frequency of mature NK cells was increased in tumor during HP. Furthermore, NK cells undergoing HP were highly activated, which contributed to substantial tumor suppression. Then, we found that a large number of neutrophils accumulated in tumor early after syngeneic HSCT. It was recently reported that neutrophil-derived mediators modulate NK cell effector functions, and so we examined whether the neutrophils infiltrated in tumor are associated with NK cell-mediated antitumor effect. The depletion of neutrophils significantly impaired an activation of NK cells in tumor and increased the fraction of proliferative NK cells accompanied by a decrease in NK cell survival. The results suggested that neutrophils in tumor prevent NK cells from activation-induced cell death during HP, thus leading to a significant antitumor effect by NK cells. This study revealed a novel aspect of antitumor immunity induced by HSCT and may contribute to the development of an effective therapeutic strategy for cancer using HSCT.

  14. mTORC1-dependent metabolic reprogramming is a prerequisite for Natural Killer cell effector function

    PubMed Central

    Donnelly, Raymond P.; Loftus, Róisín M.; Keating, Sinéad E.; Liou, Kevin T.; Biron, Christine A.; Gardiner, Clair M.; Finlay, David K.

    2014-01-01

    The mammalian target of rapamcyin complex 1 (mTORC1) is a key regulator of cellular metabolism and also has fundamental roles in controlling immune responses. Emerging evidence suggests that these two functions of mTORC1 are integrally linked. However, little is known regarding mTORC1 function in controlling the metabolism and function of natural killer (NK) cells, lymphocytes that play key roles in anti-viral and anti-tumour immunity. This study investigated the hypothesis that mTORC1-controlled metabolism underpins normal NK cell pro-inflammatory function. We demonstrate that mTORC1 is robustly stimulated in NK cells activated in vivo and in vitro. This mTORC1 activity is required for the production of the key NK cell effector molecules IFNγ, important in delivering antimicrobial and immunoregulatory functions, and granzyme B, a critical component of NK cell cytotoxic granules. The data reveal that NK cells undergo dramatic metabolic reprogramming upon activation, up-regulating rates of glucose uptake and glycolysis, and that mTORC1 activity is essential for attaining this elevated glycolytic state. Directly limiting the rate of glycolysis is sufficient to inhibit IFNγ production and granzyme B expression. This study provides the highly novel insight that mTORC1-mediated metabolic reprogramming of NK cells is a prerequisite for the acquisition of normal effector functions. PMID:25261477

  15. Leukocytosis and natural killer cell function parallel neurobehavioral fatigue induced by 64 hours of sleep deprivation.

    PubMed Central

    Dinges, D F; Douglas, S D; Zaugg, L; Campbell, D E; McMann, J M; Whitehouse, W G; Orne, E C; Kapoor, S C; Icaza, E; Orne, M T

    1994-01-01

    The hypothesis that sleep deprivation depresses immune function was tested in 20 adults, selected on the basis of their normal blood chemistry, monitored in a laboratory for 7 d, and kept awake for 64 h. At 2200 h each day measurements were taken of total leukocytes (WBC), monocytes, granulocytes, lymphocytes, eosinophils, erythrocytes (RBC), B and T lymphocyte subsets, activated T cells, and natural killer (NK) subpopulations (CD56/CD8 dual-positive cells, CD16-positive cells, CD57-positive cells). Functional tests included NK cytotoxicity, lymphocyte stimulation with mitogens, and DNA analysis of cell cycle. Sleep loss was associated with leukocytosis and increased NK cell activity. At the maximum sleep deprivation, increases were observed in counts of WBC, granulocytes, monocytes, NK activity, and the proportion of lymphocytes in the S phase of the cell cycle. Changes in monocyte counts correlated with changes in other immune parameters. Counts of CD4, CD16, CD56, and CD57 lymphocytes declined after one night without sleep, whereas CD56 and CD57 counts increased after two nights. No changes were observed in other lymphocyte counts, in proliferative responses to mitogens, or in plasma levels of cortisol or adrenocorticotropin hormone. The physiologic leukocytosis and NK activity increases during deprivation were eliminated by recovery sleep in a manner parallel to neurobehavioral function, suggesting that the immune alterations may be associated with biological pressure for sleep. PMID:7910171

  16. HIV Latency-Reversing Agents Have Diverse Effects on Natural Killer Cell Function

    PubMed Central

    Garrido, Carolina; Spivak, Adam M.; Soriano-Sarabia, Natalia; Checkley, Mary Ann; Barker, Edward; Karn, Jonathan; Planelles, Vicente; Margolis, David M.

    2016-01-01

    In an effort to clear persistent HIV infection and achieve a durable therapy-free remission of HIV disease, extensive pre-clinical studies and early pilot clinical trials are underway to develop and test agents that can reverse latent HIV infection and present viral antigen to the immune system for clearance. It is, therefore, critical to understand the impact of latency-reversing agents (LRAs) on the function of immune effectors needed to clear infected cells. We assessed the impact of LRAs on the function of natural killer (NK) cells, the main effector cells of the innate immune system. We studied the effects of three histone deacetylase inhibitors [SAHA or vorinostat (VOR), romidepsin, and panobinostat (PNB)] and two protein kinase C agonists [prostratin (PROST) and ingenol] on the antiviral activity, cytotoxicity, cytokine secretion, phenotype, and viability of primary NK cells. We found that ex vivo exposure to VOR had minimal impact on all parameters assessed, while PNB caused a decrease in NK cell viability, antiviral activity, and cytotoxicity. PROST caused non-specific NK cell activation and, interestingly, improved antiviral activity. Overall, we found that LRAs can alter the function and fate of NK cells, and these effects must be carefully considered as strategies are developed to clear persistent HIV infection. PMID:27708642

  17. Natural Killer Cell Evasion Is Essential for Infection by Rhesus Cytomegalovirus.

    PubMed

    Sturgill, Elizabeth R; Malouli, Daniel; Hansen, Scott G; Burwitz, Benjamin J; Seo, Seongkyung; Schneider, Christine L; Womack, Jennie L; Verweij, Marieke C; Ventura, Abigail B; Bhusari, Amruta; Jeffries, Krystal M; Legasse, Alfred W; Axthelm, Michael K; Hudson, Amy W; Sacha, Jonah B; Picker, Louis J; Früh, Klaus

    2016-08-01

    The natural killer cell receptor NKG2D activates NK cells by engaging one of several ligands (NKG2DLs) belonging to either the MIC or ULBP families. Human cytomegalovirus (HCMV) UL16 and UL142 counteract this activation by retaining NKG2DLs and US18 and US20 act via lysomal degradation but the importance of NK cell evasion for infection is unknown. Since NKG2DLs are highly conserved in rhesus macaques, we characterized how NKG2DL interception by rhesus cytomegalovirus (RhCMV) impacts infection in vivo. Interestingly, RhCMV lacks homologs of UL16 and UL142 but instead employs Rh159, the homolog of UL148, to prevent NKG2DL surface expression. Rh159 resides in the endoplasmic reticulum and retains several NKG2DLs whereas UL148 does not interfere with NKG2DL expression. Deletion of Rh159 releases human and rhesus MIC proteins, but not ULBPs, from retention while increasing NK cell stimulation by infected cells. Importantly, RhCMV lacking Rh159 cannot infect CMV-naïve animals unless CD8+ cells, including NK cells, are depleted. However, infection can be rescued by replacing Rh159 with HCMV UL16 suggesting that Rh159 and UL16 perform similar functions in vivo. We therefore conclude that cytomegaloviral interference with NK cell activation is essential to establish but not to maintain chronic infection. PMID:27580123

  18. The proinflammatory cytokine interleukin-18 alters multiple signaling pathways to inhibit natural killer cell death

    USGS Publications Warehouse

    Hodge, D.L.; Subleski, J.J.; Reynolds, D.A.; Buschman, M.D.; Schill, W.B.; Burkett, M.W.; Malyguine, A.M.; Young, H.A.

    2006-01-01

    The proinflammatory cytokine, interleukin-18 (IL-18), is a natural killer (NK) cell activator that induces NK cell cytotoxicity and interferon-?? (IFN-??) expression. In this report, we define a novel role for IL-18 as an NK cell protective agent. Specifically, IL-18 prevents NK cell death initiated by different and distinct stress mechanisms. IL-18 reduces NK cell self-destruction during NK-targeted cell killing, and in the presence of staurosporin, a potent apoptotic inducer, IL-18 reduces caspase-3 activity. The critical regulatory step in this process is downstream of the mitochondrion and involves reduced cleavage and activation of caspase-9 and caspase-3. The ability of IL-18 to regulate cell survival is not limited to a caspase death pathway in that IL-18 augments tumor necrosis factor (TNF) signaling, resulting in increased and prolonged mRNA expression of c-apoptosis inhibitor 2 (cIAP2), a prosurvival factor and caspase-3 inhibitor, and TNF receptor-associated factor 1 (TRAF1), a prosurvival protein. The cumulative effects of IL-18 define a novel role for this cytokine as a molecular survival switch that functions to both decrease cell death through inhibition of the mitochondrial apoptotic pathway and enhance TNF induction of prosurvival factors. ?? Mary Ann Liebert, Inc.

  19. Natural Killer Cell Evasion Is Essential for Infection by Rhesus Cytomegalovirus.

    PubMed

    Sturgill, Elizabeth R; Malouli, Daniel; Hansen, Scott G; Burwitz, Benjamin J; Seo, Seongkyung; Schneider, Christine L; Womack, Jennie L; Verweij, Marieke C; Ventura, Abigail B; Bhusari, Amruta; Jeffries, Krystal M; Legasse, Alfred W; Axthelm, Michael K; Hudson, Amy W; Sacha, Jonah B; Picker, Louis J; Früh, Klaus

    2016-08-01

    The natural killer cell receptor NKG2D activates NK cells by engaging one of several ligands (NKG2DLs) belonging to either the MIC or ULBP families. Human cytomegalovirus (HCMV) UL16 and UL142 counteract this activation by retaining NKG2DLs and US18 and US20 act via lysomal degradation but the importance of NK cell evasion for infection is unknown. Since NKG2DLs are highly conserved in rhesus macaques, we characterized how NKG2DL interception by rhesus cytomegalovirus (RhCMV) impacts infection in vivo. Interestingly, RhCMV lacks homologs of UL16 and UL142 but instead employs Rh159, the homolog of UL148, to prevent NKG2DL surface expression. Rh159 resides in the endoplasmic reticulum and retains several NKG2DLs whereas UL148 does not interfere with NKG2DL expression. Deletion of Rh159 releases human and rhesus MIC proteins, but not ULBPs, from retention while increasing NK cell stimulation by infected cells. Importantly, RhCMV lacking Rh159 cannot infect CMV-naïve animals unless CD8+ cells, including NK cells, are depleted. However, infection can be rescued by replacing Rh159 with HCMV UL16 suggesting that Rh159 and UL16 perform similar functions in vivo. We therefore conclude that cytomegaloviral interference with NK cell activation is essential to establish but not to maintain chronic infection.

  20. Natural Killer Cell Evasion Is Essential for Infection by Rhesus Cytomegalovirus

    PubMed Central

    Sturgill, Elizabeth R.; Malouli, Daniel; Hansen, Scott G.; Burwitz, Benjamin J.; Schneider, Christine L.; Womack, Jennie L.; Verweij, Marieke C.; Ventura, Abigail B.; Bhusari, Amruta; Jeffries, Krystal M.; Legasse, Alfred W.; Axthelm, Michael K.; Hudson, Amy W.; Sacha, Jonah B.; Picker, Louis J.; Früh, Klaus

    2016-01-01

    The natural killer cell receptor NKG2D activates NK cells by engaging one of several ligands (NKG2DLs) belonging to either the MIC or ULBP families. Human cytomegalovirus (HCMV) UL16 and UL142 counteract this activation by retaining NKG2DLs and US18 and US20 act via lysomal degradation but the importance of NK cell evasion for infection is unknown. Since NKG2DLs are highly conserved in rhesus macaques, we characterized how NKG2DL interception by rhesus cytomegalovirus (RhCMV) impacts infection in vivo. Interestingly, RhCMV lacks homologs of UL16 and UL142 but instead employs Rh159, the homolog of UL148, to prevent NKG2DL surface expression. Rh159 resides in the endoplasmic reticulum and retains several NKG2DLs whereas UL148 does not interfere with NKG2DL expression. Deletion of Rh159 releases human and rhesus MIC proteins, but not ULBPs, from retention while increasing NK cell stimulation by infected cells. Importantly, RhCMV lacking Rh159 cannot infect CMV-naïve animals unless CD8+ cells, including NK cells, are depleted. However, infection can be rescued by replacing Rh159 with HCMV UL16 suggesting that Rh159 and UL16 perform similar functions in vivo. We therefore conclude that cytomegaloviral interference with NK cell activation is essential to establish but not to maintain chronic infection. PMID:27580123

  1. Killing of naive T cells by CD95L-transfected dendritic cells (DC): in vivo study using killer DC-DC hybrids and CD4(+) T cells from DO11.10 mice.

    PubMed

    Kusuhara, Masahiro; Matsue, Keiko; Edelbaum, Dale; Loftus, Julie; Takashima, Akira; Matsue, Hiroyuki

    2002-04-01

    Dendritic cells (DC) play the dual task of initiating cellular immunity against potentially harmful foreign antigens (Ag), while maintaining immunological tolerance to self-Ag and environmental Ag. As an approach to induce Ag-specific suppression, we and others introduced CD95 ligand (L) cDNA into DC. The resulting "killer" DC delivered apoptotic signals, instead of activation signals, to primed CD4(+) T cells in vitro and induced Ag-specific immunosuppression in vivo. To study the impact of killer DC on naive T cells, the fate of Ag-reactive T cells and the extent of their depletion after killer DC treatment, we performed in vitro and in vivo reconstitution experiments using: (a) killer DC-DC hybrids created between CD95L-transduced XS106 DC clone (A/J origin) and splenic DC from BALB/c mice, (b) CD4(+) T cells isolated from DO11.10 transgenic mice (BALB/c background), and (c) OVA(323-339) peptide as relevant Ag. Ovalbumin (OVA)-pulsed killer DC-DC hybrids inhibited DO11.10 T cell activation triggered by conventional DC, instead of inducing their activation. Rapid apoptosis of T cells was observed after co-culture with OVA-pulsed killer DC-DC hybrids, but not with non-pulsed killer DC-DC hybrids or OVA-pulsed control DC-DC hybrids. For in vivo reconstitution, (BALB/cxA/J)F1 mice received subcutaneous administration of killer DC-DC hybrids, followed by intravenous inoculation of DO11.10 T cells. Killer DC-DC hybrids migrated preferentially to draining lymph nodes albeit with relatively low efficiency (0.5-1% recovery) and they induced significant, but incomplete (30-40%) killing of DO11.10 T cells in this location. These results document the abilities of CD95L-transduced DC to trigger apoptosis of naive T cells in an Ag-specific manner, to overrule T cell activation signals delivered by conventional DC, and to reduce local frequencies of Ag-reactive T cells in vivo. Our data also uncover two major limitations (relatively low homing efficiency and incomplete

  2. Tumor necrosis factor-{alpha} enhances IL-15-induced natural killer cell differentiation

    SciTech Connect

    Lee, Jiwon; Lee, Suk Hyung; Shin, Nara; Jeong, Mira; Kim, Mi Sun; Kim, Mi Jeong; Yoon, Suk Ran; Chung, Jin Woong; Kim, Tae-Don; Choi, Inpyo

    2009-09-04

    The differentiation of natural killer (NK) cells is regulated by various factors including soluble growth factors and transcription factors. Here, we have demonstrated that tumor necrosis factor-{alpha} (TNF-{alpha}) is a positive regulator of NK cell differentiation. TNF-{alpha} augmented the IL-15-induced expression of NK1.1 and CD122 in mature NK cells, and TNF-{alpha} alone also induced NK cell maturation as well as IL-15. TNF-{alpha} also increased IFN-{gamma} production in NK cells in the presence of IL-15. Meanwhile, mRNA expression of several transcription factors, including T-bet and GATA-3, was increased by the addition of TNF-{alpha} and IL-15. In addition, TNF-{alpha} increased nuclear factor-kappa B (NF-{kappa}B) activity in NK cells and inhibition of NF-{kappa}B impeded TNF-{alpha}-enhanced NK cell maturation. Overall, these data suggest that TNF-{alpha} significantly increased IL-15-driven NK cell differentiation by increasing the expression of transcription factors that play crucial roles in NK cell maturation and inducing the NF-{kappa}B activity.

  3. Characterization of the natural killer T-cell response in an adoptive transfer model of atherosclerosis.

    PubMed

    VanderLaan, Paul A; Reardon, Catherine A; Sagiv, Yuval; Blachowicz, Lydia; Lukens, John; Nissenbaum, Michael; Wang, Chyung-Ru; Getz, Godfrey S

    2007-03-01

    Natural killer T (NKT) cells have recently been implicated in atherogenesis, primarily for their ability to recognize and respond to lipid antigens. Because the atherosclerotic lesion is characterized by the retention and modification of lipids in the vascular wall, NKT cells may be involved in promoting the local vascular inflammatory response. Here, we investigate the proatherogenic role of NKT cells in an adoptive transfer model of atherosclerosis, using as recipients immune-deficient, atherosclerosis-susceptible RAG1(-/-)LDLR(-/-) mice. The adoptive transfer of an NKT cell-enriched splenocyte population from Valpha14Jalpha18 T-cell receptor transgenic mice resulted in a 73% increase in aortic root lesion area compared with recipients of NKT cell-deficient splenocytes derived from CD1d(-/-) mice after 12 weeks of Western-type diet feeding. The total serum from hypercholesterolemic mice leads to a small but significant activation of Valpha14Jalpha18 T-cell receptor-expressing hybridoma line by dendritic cells that is CD1d-dependent. Therefore, these studies demonstrate that NKT cells are proatherogenic in the absence of exogenous stimulation, and this activity is likely associated with endogenous lipid antigens carried by lipoproteins in the circulation and perhaps also in the atherosclerotic plaque.

  4. Characterization of the Natural Killer T-Cell Response in an Adoptive Transfer Model of Atherosclerosis

    PubMed Central

    VanderLaan, Paul A.; Reardon, Catherine A.; Sagiv, Yuval; Blachowicz, Lydia; Lukens, John; Nissenbaum, Michael; Wang, Chyung-Ru; Getz, Godfrey S.

    2007-01-01

    Natural killer T (NKT) cells have recently been implicated in atherogenesis, primarily for their ability to recognize and respond to lipid antigens. Because the atherosclerotic lesion is characterized by the retention and modification of lipids in the vascular wall, NKT cells may be involved in promoting the local vascular inflammatory response. Here, we investigate the proatherogenic role of NKT cells in an adoptive transfer model of atherosclerosis, using as recipients immune-deficient, atherosclerosis-susceptible RAG1−/−LDLR−/− mice. The adoptive transfer of an NKT cell-enriched splenocyte population from Vα14Jα18 T-cell receptor transgenic mice resulted in a 73% increase in aortic root lesion area compared with recipients of NKT cell-deficient splenocytes derived from CD1d−/− mice after 12 weeks of Western-type diet feeding. The total serum from hypercholesterolemic mice leads to a small but significant activation of Vα14Jα18 T-cell receptor-expressing hybridoma line by dendritic cells that is CD1d-dependent. Therefore, these studies demonstrate that NKT cells are proatherogenic in the absence of exogenous stimulation, and this activity is likely associated with endogenous lipid antigens carried by lipoproteins in the circulation and perhaps also in the atherosclerotic plaque. PMID:17322392

  5. Physiology of natural killer cells. In vivo regulation of progenitors by interleukin 3

    SciTech Connect

    Kalland, T.

    1987-12-01

    Adoptive transfer of bone marrow cells to syngeneic lethally irradiated C57BL/6 mice was used to study the maturation of natural killer (NK) cells from their progenitors. The NK progenitor cell was found to be asialomonoganglioside-negative, (aGM1-) Thy-1-, NK-1-, Ly-1-, Ly-2-, and L3T4-. The NK cells emerging from the bone marrow grafts were aGM1+, NK-1+, Thy-1+/-, Ly-1-, Ly-2-, and L3T4- and to have a target specter similar to that of NK cells isolated from the spleen of normal mice. The regulatory role of interleukin 2 (IL-2) and interleukin 3 (IL-3) for the maturation of NK cells was examined by exposure of the bone marrow cells to the lymphokines in vitro before bone marrow grafting or by treatment of bone marrow-grafted mice with lymphokines through s.c. implanted miniosmotic pumps. IL-3 antagonized the IL-2-induced maturation of NK cells in vitro and strongly inhibited the generation of NK cells after adoptive transfer of bone marrow cells in vivo. The suppressive effect of IL-3 was evident throughout the treatment period (8 or 16 days) but was apparently reversible because NK activity returned to control levels within 8 days after cessation of treatment. The inhibition of cytotoxic activity was accompanied by a reduced appearance of cells with the NK phenotypic markers aGM1 or NK-1, indicating that not only the cytotoxic activity of NK cells but also their actual formation was inhibited. Concomitantly, a moderate increase in cells expressing the T cell marker L3T4 and an increased proliferative response to the T cell mitogen concanavalin A was observed. A direct estimate of the effect of IL-3 on the frequency of NK cell progenitors was obtained by limiting dilution analysis of bone marrow cells at day 8 after bone marrow transplantation.

  6. Suppressing the killer instinct.

    PubMed

    Campbell, Kerry S

    2016-05-24

    Natural killer (NK) cells are innate lymphoid cells that have adopted activating and inhibitory signaling mechanisms enabling them to be tolerant of normal cells but to distinguish and eliminate tumor cells and virus-infected cells. In this issue of Science Signaling, Matalon et al show how inhibitory receptors disrupt NK cell activation by stimulating dephosphorylation of the adaptor protein LAT (linker of activated T cells) and phospholipase C-γ by the phosphatase SHP-1 [Src homology 2 (SH2) domain-containing protein tyrosine phosphatase 1], as well as ubiquitylation of LAT by Cbl family E3 ubiquitin ligases.

  7. Autologous Stem Cell Transplant Recipients Tolerate Haploidentical Related-Donor Natural Killer Cell Enriched Infusions

    PubMed Central

    Klingemann, Hans; Grodman, Carrie; Cutler, Elliott; Duque, Marvin; Kadidlo, Diane; Klein, Andreas K.; Sprague, Kellie A.; Miller, Kenneth B.; Comenzo, Raymond L.; Kewalramani, Tarun; Yu, Neng; Van Etten, Richard A.; McKenna, David H.

    2012-01-01

    BACKGROUND In the setting of allogeneic stem cell transplantation (alloSCT), infusing natural killer (NK) cells from a major histocompatibility complex (MHC) mismatched donor can mediate an anti-leukemic effect. Graft versus tumor (GvT) effect following autologous stem cell transplantation (ASCT) may result in less disease relapse. STUDY DESIGN AND METHODS We performed a phase I clinical trial to assess the safety and feasibility of infusing distantly processed donor NK enriched mononuclear cell (NK-MC) infusions from a MHC haplotype mismatched (haploidentical) donor to patients who recently underwent ASCT for a hematologic malignancy. On day 1, peripheral blood mononuclear cells (MC) were obtained by steady-state leukapheresis and sent from Boston to the Production Assistance for Cellular Therapies (PACT) facility at the University of Minnesota, where immunomagnetic depletion of CD3 cells was performed on day 2. NK-MC product were then returned to Boston on day 2 for infusion on day 3. Toxicity, cellular product characteristics and logistic events were monitored. RESULTS At a median of 90 days (range, 49–191) following ASCT, thirteen patients were treated with escalating doses of NK-MC per kg from 105 to 2 ×107. Adverse effects included grade 2 rigors and muscle aches, but no grade 3 or 4 events, and no GvHD or marrow suppression. One air courier delay occurred. NK-MC products were viable with cytotoxic activity after transport. CONCLUSION CD3-depleted, MHC mismatched allogeneic NK-MC infusions can be safely and feasibly administered to patients after ASCT following distant processing and transport, justifying further development of this approach. PMID:22738379

  8. Natural killer cells in highly exposed hepatitis C-seronegative injecting drug users.

    PubMed

    Mina, M M; Cameron, B; Luciani, F; Vollmer-Conna, U; Lloyd, A R

    2016-06-01

    Injecting drug use remains the major risk factor for hepatitis C (HCV) transmission. A minority of long-term injecting drug users remain seronegative and aviraemic, despite prolonged exposure to HCV - termed highly exposed seronegative subjects. Natural killer (NK) cells have been implicated in this apparent protection. A longitudinal nested, three group case-control series of subjects was selected from a prospective cohort of seronegative injecting drug users who became incident cases (n = 11), remained seronegative (n = 11) or reported transient high-risk behaviour and remained uninfected (n = 11). The groups were matched by age, sex and initial risk behaviour characteristics. Stored peripheral blood mononuclear cells were assayed in multicolour flow cytometry to enumerate natural killer cell subpopulations and to assess functional activity using Toll-like receptor ligands before measurement of activation, cytokine production and natural cytotoxicity receptor expression. Principal components were derived to describe the detailed phenotypic characteristics of the major NK subpopulations (based on CD56 and CD16 co-expression), before logistic regression analysis to identify associations with exposed, seronegative individuals. The CD56(dim) CD16(+) (P = 0.05, OR 6.92) and CD56(dim) CD16(-) (P = 0.05, OR 6.07) principal components differed between exposed, seronegative individuals and pre-infection samples of the other two groups. These included CD56(dim) CD16(+) and CD56(dim) CD16(-) subsets with CD56(dim) CD16(+) IFN-γ and TNF-α on unstimulated cells, and CD56(dim) CD16(-) CD69(+) , CD107a(+) , IFN-γ and TNF-α following TLR stimulation. The cytotoxic CD56(dim) NK subset thus distinguished highly exposed, seronegative subjects, suggesting NK cytotoxicity may contribute to protection from HCV acquisition. Further investigation of the determinants of this association and prospective assessment of protection against HCV infection are warranted.

  9. Killer activity of Saccharomyces cerevisiae strains: partial characterization and strategies to improve the biocontrol efficacy in winemaking.

    PubMed

    de Ullivarri, Miguel Fernández; Mendoza, Lucía M; Raya, Raúl R

    2014-11-01

    Killer yeasts are considered potential biocontrol agents to avoid or reduce wine spoilage by undesirable species. In this study two Saccharomyces cerevisiae strains (Cf8 and M12) producing killer toxin were partially characterized and new strategies to improve their activity in winemaking were evaluated. Killer toxins were characterized by biochemical tests and growth inhibition of sensitive yeasts. Also genes encoding killer toxin were detected in the chromosomes of both strains by PCR. Both toxins showed optimal activity and production at conditions used during the wine-making process (pH 3.5 and temperatures of 15-25 °C). In addition, production of both toxins was higher when a nitrogen source was added. To improve killer activity different strategies of inoculation were studied, with the sequential inoculation of killer strains the best combination to control the growth of undesired yeasts. Sequential inoculation of Cf8-M12 showed a 45 % increase of killer activity on sensitive S. cerevisiae and spoilage yeasts. In the presence of ethanol (5-12 %) and SO2 (50 mg/L) the killer activity of both toxins was increased, especially for toxin Cf8. Characteristics of both killer strains support their future application as starter cultures and biocontrol agents to produce wines of controlled quality.

  10. Natural cytotoxicity in immunodeficiency diseases: preservation of natural killer activity and the in vivo appearance of radioresistant killing

    SciTech Connect

    Pierce, G.F.; Polmar, S.H.; Schacter, B.Z.; Brovall, C.; Hornick, D.L.; Sorensen, R.U.

    1986-01-01

    We studied spontaneous natural killer (NK) cell activity and radiation-resistant NK mediated cytotoxicity in four patients with clinically documented severe combined immune deficiency disease (SCID), and in one subject each with intestinal lymphangiectasia and cartilage-hair hypoplasia. We observed the preservation of spontaneous NK activity in all patients despite the presence of profound B- and T-lymphocytopenia and clinical immunodeficiency. NK activity was associated with relatively normal circulating numbers of OKM1+ lymphocytes, a population known to contain NK effectors. Spontaneous NK activity resistant to 3000 rad was increased in all patients, indicating the presence of activated natural killer cells in vivo. The concept of a chronically activated immune system in these patients was further supported by the presence of increased Ia positive T cells in all subjects tested, suggesting that radioresistant NK activity may be a useful parameter to measure when assessing in vivo immune activation. Our data, as well as that of others, supports the hypothesis that at least one population of NK cells is a distinct lineage arising at the differentiation level of myeloid and lymphoid stem cells in the bone marrow.

  11. The breast tumor microenvironment alters the phenotype and function of natural killer cells

    PubMed Central

    Krneta, Tamara; Gillgrass, Amy; Chew, Marianne; Ashkar, Ali A.

    2016-01-01

    Natural killer (NK) cells are innate immune cells with the ability to identify and eliminate transformed cells. However, within tumors, many studies have described NK cells as non-functional. The developmental stage of tumor-associated NK cells and how this may relate to functionality has not been explored. We examined the developmental state of NK cells from polyoma middle T antigen (pyMT) transgenic mouse (MMTV-pMT) breast tumors. In pyMT tumors, NK cells were immature as evidenced by their decreased expression of DX5 and their CD27lowCD11blow phenotype. These immature NK cells also had increased expression of NKG2A and expressed low levels of NKp46, perforin, and granzyme B. In contrast, splenic NK cells isolated from the same mice maintained their maturity and their expression of activation markers. To delineate whether the tumor microenvironment directly alters NK cells, we adoptively transferred labeled NK cells and followed their activation status in both the spleen and the tumor. NK cells that arrived at the tumor had half the expression of NKp46 within three days of transfer in comparison to those which arrived at the spleen. In an effort to modify the tumor microenvironment and assess the plasticity of intratumoral NK cells, we treated pyMT tumors with IL-12 and anti-TGF-β. After one week of treatment, the maturity of tumor-associated NK cells was increased; thus, indicating that these cells possess the ability to mature and become activated. A better understanding of how NK cells are modified by the tumor microenvironment will help to develop strategies aimed at bolstering immune responses against tumors. PMID:26277898

  12. Natural Killer Activity: Early Days, Advances, and Seminal Observations

    PubMed Central

    Ortaldo, John R.; Wiltrout, Robert H.; Reynolds, Craig W.

    2014-01-01

    This manuscript describes the early history of NK cell discovery, with emphasis on the events in the first decade of NK cell studies, 1972–1982. The authors highlight some of the earliest and most important observations that would later prove to be milestones in the study of NK cells and their activity. PMID:24941370

  13. Delineation of Natural Killer Cell Differentiation from Myeloid Progenitors in Human.

    PubMed

    Chen, Qingfeng; Ye, Weijian; Jian Tan, Wei; Mei Yong, Kylie Su; Liu, Min; Qi Tan, Shu; Loh, Eva; Te Chang, Kenneth; Chye Tan, Thiam; Preiser, Peter R; Chen, Jianzhu

    2015-01-01

    Understanding of natural killer (NK) cell development in human is incomplete partly because of limited access to appropriate human tissues. We have developed a cytokine-enhanced humanized mouse model with greatly improved reconstitution and function of human NK cells. Here we report the presence of a cell population in the bone marrow of the cytokine-treated humanized mice that express both NK cell marker CD56 and myeloid markers such as CD36 and CD33. The CD56(+)CD33(+)CD36(+) cells are also found in human cord blood, fetal and adult bone marrow. Although the CD56(+)CD33(+)CD36(+) cells do not express the common NK cell functional receptors and exhibit little cytotoxic and cytokine-producing activities, they readily differentiate into mature NK cells by acquiring expression of NK cell receptors and losing expression of the myeloid markers. Further studies show that CD33(+)CD36(+) myeloid NK precursors are derived from granulo-myelomonocytic progenitors. These results delineate the pathway of human NK cell differentiation from myeloid progenitors in the bone marrow and suggest the utility of humanized mice for studying human hematopoiesis. PMID:26456148

  14. Age-related decline of perforin expression in human cytotoxic T lymphocytes and natural killer cells.

    PubMed

    Rukavina, D; Laskarin, G; Rubesa, G; Strbo, N; Bedenicki, I; Manestar, D; Glavas, M; Christmas, S E; Podack, E R

    1998-10-01

    In this study a flow cytometric technique for detecting cytoplasmic perforin (P) has been used to quantify age-related changes in perforin expression in human peripheral blood lymphocytes (PBL). Proportions of P+ lymphocytes increased after birth, but declined rapidly after the age of 70 years. This was true for both T cells and CD16(+) and CD56(+) natural killer (NK) cells. Children showed in addition to high levels of perforin positive CD8(+) cells a much higher proportion of CD4(+)P+ cells than the other age groups. In elderly individuals there was also a highly significant reduction in mean levels of perforin per cell as compared with all other groups (P < .05 to .001). Adult women had consistently higher mean levels of perforin per cell than adult men for all P+ cell phenotypes. Functional tests clearly showed the deficiency in early spontaneous cytotoxic potential of PBL from elderly persons due to relative P deficiency, which can be corrected by stimulation of cytolytic cells with target cells and interleukin-2 (IL-2). The deficiency in cytolytic activity on the contact with target cells may have implications for antiviral and antitumor immunity in elderly persons.

  15. Design and operation specifications of an active monitoring system for detecting southern resident killer whales

    SciTech Connect

    Deng, Zhiqun; Carlson, Thomas J.; Xu, Jinshan; Martinez, Jayson J.; Weiland, Mark A.; Mueller, Robert P.; Myers, Joshua R.; Jones, Mark E.

    2011-09-30

    Before final approval is given to the Snohomish County Public Utility District No. 1 for deploying the first tidal power devices in the United States in an open water environment, a system to manage the potential risk of injury to killer whales due to collision with moving turbine blades must be demonstrated. The Pacific Northwest National Laboratory (PNNL) is tasked with establishing the performance requirements for, constructing, and testing a prototype marine animal alert system for triggering temporary turbine shutdown when there is risk of collision with a killer whale. To develop a system that relies on active sonar two critical areas must be investigated - the target strength of killer whales and the frequency content of commercially available active sonar units. PNNL studied three target strength models: a simple model, the Fourier matching model, and the Kirchoff-ray mode model. Using target strength measurements of bottlenose dolphins obtained by previous researchers and assuming killer whales share similar morphology and structure, PNNL extrapolated the target strength of an adult killer whale 7.5 m in length at a frequency of 67 kHz. To study the frequency content of a commercially available sonar unit, direct measurements of the signal transmitted by the sonar were obtained by using a hydrophone connected to a data acquisition system in both laboratory and field conditions. The measurements revealed that in addition to the primary frequency of 200 kHz, there is a secondary frequency component at 90 kHz, which is within the hearing range of killer whales. The amplitude of the 90-kHz frequency component is above the hearing threshold of killer whales but below the threshold for potential injuries.

  16. Human Invariant Natural Killer T cells possess immune-modulating functions during Aspergillus infection.

    PubMed

    Beitzen-Heineke, Antonia; Bouzani, Maria; Schmitt, Anna-Lena; Kurzai, Oliver; Hünniger, Kerstin; Einsele, Hermann; Loeffler, Juergen

    2016-02-01

    Aspergillus fumigatus is the most common cause for invasive fungal infections, a disease associated with high mortality in immune-compromised patients. CD1d-restricted invariant natural killer T (iNKT) cells compose a small subset of T cells known to impact the immune response toward various infectious pathogens. To investigate the role of human iNKT cells during A. fumigatus infection, we studied their activation as determined by CD69 expression and cytokine production in response to distinct fungal morphotypes in the presence of different CD1d(+) antigen presenting cells using flow cytometry and multiplex enzyme-linked immunosorbent assay (ELISA). Among CD1d(+) subpopulations, CD1d(+)CD1c(+) mDCs showed the highest potential to activate iNKT cells on a per cell basis. The presence of A. fumigatus decreased this effect of CD1d(+)CD1c(+) mDCs on iNKT cells and led to reduced secretion of TNF-α, G-CSF and RANTES. Production of other Th1 and Th2 cytokines was not affected by the fungus, suggesting an immune-modulating function for human iNKT cells during A. fumigatus infection.

  17. Identification of natural killer cell receptor genes in the genome of the marsupial Tasmanian devil (Sarcophilus harrisii).

    PubMed

    van der Kraan, Lauren E; Wong, Emily S W; Lo, Nathan; Ujvari, Beata; Belov, Katherine

    2013-01-01

    Within the mammalian immune system, natural killer (NK) cells contribute to the first line of defence against infectious agents and tumours. Their activity is regulated, in part, by cell surface NK cell receptors. NK receptors can be divided into two unrelated, but functionally analogous superfamilies based on the structure of their extracellular ligand-binding domains. Receptors belonging to the C-type lectin superfamily are predominantly encoded in the natural killer complex (NKC), while receptors belonging to the immunoglobulin superfamily are predominantly encoded in the leukocyte receptor complex (LRC). Natural killer cell receptors are emerging as a rapidly evolving gene family which can display significant intra- and interspecific variation. To date, most studies have focused on eutherian mammals, with significantly less known about the evolution of these receptors in marsupials. Here, we describe the identification of 43 immunoglobulin domain-containing LRC genes in the genome of the Tasmanian devil (Sarcophilus harrisii), the largest remaining marsupial carnivore and only the second marsupial species to be studied. We also identify orthologs of NKC genes KLRK1, CD69, CLEC4E, CLEC1B, CLEC1A and an ortholog of an opossum NKC receptor. Characterisation of these regions in a second, distantly related marsupial provides new insights into the dynamic evolutionary histories of these receptors in mammals. Understanding the functional role of these genes is also important for the development of therapeutic agents against Devil Facial Tumour Disease, a contagious cancer that threatens the Tasmanian devil with extinction.

  18. Cytokine-induced killer cells interact with tumor lysate-pulsed dendritic cells via CCR5 signaling.

    PubMed

    Lee, Hong Kyung; Kim, Yong Guk; Kim, Ji Sung; Park, Eun Jae; Kim, Boyeong; Park, Ki Hwan; Kang, Jong Soon; Hong, Jin Tae; Kim, Youngsoo; Han, Sang-Bae

    2016-08-10

    The antitumor activity of cytokine-induced killer (CIK) cells can be increased by co-culturing them with tumor lysate-pulsed dendritic cells (tDCs); this phenomenon has been studied mainly at the population level. Using time-lapse imaging, we examined how CIK cells gather information from tDCs at the single-cell level. tDCs highly expressed CCL5, which bound CCR5 expressed on CIK cells. tDCs strongly induced migration of Ccr5(+/+) CIK cells, but not that of Ccr5(-/-) CIK cells or Ccr5(+/+) CIK cells treated with the CCR5 antagonist Maraviroc. Individual tDCs contacted Ccr5(+/+) CIK cells more frequently and lengthily than with Ccr5(-/-) CIK cells. Consequently, tDCs increased the antitumor activity of Ccr5(+/+) CIK cells in vitro and in vivo, but did not increase that of Ccr5(-/-) CIK cells. Taken together, our data provide insight into the mechanism of CIK cell activation by tDCs at the single-cell level.

  19. Modulation of invariant natural killer T cell cytokine responses by indoleamine 2,3-dioxygenase

    PubMed Central

    Molano, Alberto; Illarionov, Petr A.; Besra, Gurdyal S.; Putterman, Chaim; Porcelli, Steven A.

    2008-01-01

    1. SUMMARY The intracellular enzyme indoleamine 2,3-dioxygenase (IDO), which degrades the rare and essential aminoacid tryptophan and converts it into a series of biologically active catabolites, has been linked to the regulation of immune tolerance by specific dendritic cell subsets, and to the downmodulation of exacerbated immune responses. Although the immunoregulatory effects of IDO may be in part due to generalized suppression of cell proliferation caused by tryptophan starvation, there is also evidence that tryptophan catabolites could be directly responsible for some of the observed effects. In this report, we investigated the consequences of IDO activity, particularly with regard to the effects of tryptophan-derived catabolites, on the cytokine responses of activated invariant natural killer T (iNKT) cells, a specialized T cell subset known to have immunoregulatory properties. Our results showed that pharmacologic inhibition of IDO skewed cytokine responses of iNKT cells towards a Th1 profile. In contrast, the presence at low micromolar concentrations of the tryptophan catabolites L-kynurenine, 3-hydroxy-kynurenine, or 3-hydroxy-anthranilic acid shifted the cytokine balance towards a Th2 pattern. These findings have implications for our current understanding of immunoregulation, and the mechanisms by which iNKT cells participate in the modulation of immune responses. PMID:18272236

  20. Contribution of Invariant Natural Killer T Cells to Skin Wound Healing.

    PubMed

    Tanno, Hiromasa; Kawakami, Kazuyoshi; Ritsu, Masae; Kanno, Emi; Suzuki, Aiko; Kamimatsuno, Rina; Takagi, Naoyuki; Miyasaka, Tomomitsu; Ishii, Keiko; Imai, Yoshimichi; Maruyama, Ryoko; Tachi, Masahiro

    2015-12-01

    In the present study, we determined the contribution of invariant natural killer T (iNKT) cells to the skin wound healing process. In iNKT cell-deficient (Jα18KO) mice lacking iNKT cells, wound closure was significantly delayed compared with wild-type mice. Collagen deposition, expression of α-smooth muscle actin and CD31, and wound breaking strength were significantly attenuated in Jα18KO mice. The adoptive transfer of liver mononuclear cells from wild-type but not from Jα18KO or interferon (IFN)-γ gene-disrupted (IFN-γKO) mice resulted in the reversal of this impaired wound healing in Jα18KO mice. IFN-γ expression was induced in the wounded tissues, which was significantly decreased at 6, 12, and 24 hours, but increased on day 3 after wounding in Jα18KO mice. The main source of the late-phase IFN-γ production in Jα18KO mice were neutrophils rather than NK cells and T cells. Administration of α-galactosylceramide, an activator of iNKT cells, resulted in the acceleration of wound healing on day 3 in wild-type mice. This effect was not observed in IFN-γKO mice. These results indicate that iNKT cells play important roles in wound healing. The iNKT cell-induced IFN-γ production may regulate the wound healing process in the early phase.

  1. Contribution of Invariant Natural Killer T Cells to Skin Wound Healing.

    PubMed

    Tanno, Hiromasa; Kawakami, Kazuyoshi; Ritsu, Masae; Kanno, Emi; Suzuki, Aiko; Kamimatsuno, Rina; Takagi, Naoyuki; Miyasaka, Tomomitsu; Ishii, Keiko; Imai, Yoshimichi; Maruyama, Ryoko; Tachi, Masahiro

    2015-12-01

    In the present study, we determined the contribution of invariant natural killer T (iNKT) cells to the skin wound healing process. In iNKT cell-deficient (Jα18KO) mice lacking iNKT cells, wound closure was significantly delayed compared with wild-type mice. Collagen deposition, expression of α-smooth muscle actin and CD31, and wound breaking strength were significantly attenuated in Jα18KO mice. The adoptive transfer of liver mononuclear cells from wild-type but not from Jα18KO or interferon (IFN)-γ gene-disrupted (IFN-γKO) mice resulted in the reversal of this impaired wound healing in Jα18KO mice. IFN-γ expression was induced in the wounded tissues, which was significantly decreased at 6, 12, and 24 hours, but increased on day 3 after wounding in Jα18KO mice. The main source of the late-phase IFN-γ production in Jα18KO mice were neutrophils rather than NK cells and T cells. Administration of α-galactosylceramide, an activator of iNKT cells, resulted in the acceleration of wound healing on day 3 in wild-type mice. This effect was not observed in IFN-γKO mice. These results indicate that iNKT cells play important roles in wound healing. The iNKT cell-induced IFN-γ production may regulate the wound healing process in the early phase. PMID:26468976

  2. NKp80 Defines a Critical Step during Human Natural Killer Cell Development.

    PubMed

    Freud, Aharon G; Keller, Karen A; Scoville, Steven D; Mundy-Bosse, Bethany L; Cheng, Stephanie; Youssef, Youssef; Hughes, Tiffany; Zhang, Xiaoli; Mo, Xiaokui; Porcu, Pierluigi; Baiocchi, Robert A; Yu, Jianhua; Carson, William E; Caligiuri, Michael A

    2016-07-12

    Human natural killer (NK) cells develop in secondary lymphoid tissues (SLTs) through distinct stages. We identified two SLT lineage (Lin)(-)CD34(-)CD117(+/-)CD94(+)CD16(-) "stage 4" subsets according to expression of the C-type lectin-like surface-activating receptor, NKp80: NKp80(-) (stage "4a") and NKp80(+) (stage "4b"). Whereas stage 4b cells expressed more of the transcription factors T-BET and EOMES, produced interferon-gamma, and were cytotoxic, stage 4a cells expressed more of the transcription factors RORγt and AHR and produced interleukin-22, similar to SLT Lin(-)CD34(-)CD117(+)CD94(-)CD16(-) "stage 3" cells, whose phenotype overlaps with that of group 3 innate lymphoid cells (ILC3s). Co-culture with dendritic cells or transplantation into immunodeficient mice produced mature NK cells from stage 3 and stage 4a populations. These data identify NKp80 as a marker of NK cell maturity in SLTs and support a model of human NK cell development through a stage 4a intermediate with ILC3-associated features.

  3. Identification and Analysis of Natural Killer Cells in Murine Nasal Passages

    PubMed Central

    Okada, Kazunari; Sato, Shintaro; Sato, Ayuko; Mandelboim, Ofer; Yamasoba, Tatsuya; Kiyono, Hiroshi

    2015-01-01

    Background Natural killer (NK) cells in the upper respiratory airways are not well characterized. In the current study, we sought to characterize and functionally assess murine nasal NK cells. Methods Using immunohistochemistry and flow cytometry, we compared the nasal NK cells of Ncr1GFP/+ knock-in mice, whose NK cells produced green fluorescent protein, with their splenic and pulmonary counterparts. In addition, we functionally analyzed the nasal NK cells of these mice in vitro. To assess the in vivo functions of nasal NK cells, C57BL/6 mice depleted of NK cells after treatment with PK136 antibody were nasally infected with influenza virus PR8. Results Immunohistochemical analysis confirmed the presence of NK cells in the lamina propria of nasal mucosa, and flow cytometry showed that these cells were of NK cell lineage. The expression patterns of Ly49 receptor, CD11b/CD27, CD62L and CD69 revealed that nasal NK cells had an immature and activated phenotype compared with that of their splenic and pulmonary counterparts. Effector functions including degranulation and IFN(interferon)-γ production after in vitro stimulation with phorbol 12-myristate-13-acetate plus ionomycin or IL(interleukin)-12 plus IL-18 were dampened in nasal NK cells, and the depletion of NK cells led to an increased influenza virus titer in nasal passages. Conclusions The NK cells of the murine nasal passage belong to the conventional NK cell linage and characteristically demonstrate an immature and activated phenotype. Despite their hyporesponsiveness in vitro, nasal NK cells play important roles in the host defense against nasal influenza virus infection. PMID:26575399

  4. Expression of complement regulatory proteins on human natural killer cell subsets.

    PubMed

    Wang, Lin; Halliday, Deborah; Johnson, Peter M; Christmas, Stephen E

    2007-10-15

    The cell surface complement regulatory (CReg) proteins CD46, CD55 and CD59 are widely distributed on human leucocytes and protect against complement-mediated damage. To investigate heterogeneity in CReg protein expression by human natural killer (NK) cells, levels were assessed on resting and activated NK cell subsets identified phenotypically on the basis of expression of CD56 and CD158 markers. Levels of all three CReg proteins on CD56+ cells were lower than on T cells (p<0.05). Freshly isolated CD56(bright) cells expressed higher levels of CD55 than CD56dim cells (p<0.05). CD158a+ cells expressed significantly lower levels of both CD46 and CD59, and CD158e+ cells expressed significantly lower levels of CD46, than CD158a(-) CD158e(-) cells, respectively (both p<0.05). Stimulation with PHA did not significantly alter NK cell surface CReg protein levels whereas, following culture with IL-2, CD46 and CD59 were decreased on both CD56bright and CD56dim subsets (p<0.05). In the case of CD59, this was independent of T cells. Only CD46 was significantly downregulated on CD158b+ (GL183+) and CD158e (NKB1+) subsets (p<0.05). However, culture in IL-15 significantly increased levels of all three CReg proteins. These observations that CReg proteins are downregulated on certain NK cell subsets following activation with IL-2 are opposite to previous findings for other leucocyte subpopulations. Activated NK cells may instead use other strategies for protection against complement-mediated damage in a local inflammatory response.

  5. Imaging burst kinetics and spatial coordination during serial killing by single natural killer cells.

    PubMed

    Choi, Paul J; Mitchison, Timothy J

    2013-04-16

    Cytotoxic lymphocytes eliminate virus-infected and cancerous cells by immune recognition and killing through the perforin-granzyme pathway. Traditional killing assays measure average target cell lysis at fixed times and high effector:target ratios. Such assays obscure kinetic details that might reveal novel physiology. We engineered target cells to report on granzyme activity, used very low effector:target ratios to observe potential serial killing, and performed low magnification time-lapse imaging to reveal time-dependent statistics of natural killer (NK) killing at the single-cell level. Most kills occurred during serial killing, and a single NK cell killed up to 10 targets over a 6-h assay. The first kill was slower than subsequent kills, especially on poor targets, or when NK signaling pathways were partially inhibited. Spatial analysis showed that sequential kills were usually adjacent. We propose that NK cells integrate signals from the previous and current target, possibly by simultaneous contact. The resulting burst kinetics and spatial coordination may control the activity of NK cells in tissues.

  6. Suppression of a Natural Killer Cell Response by Simian Immunodeficiency Virus Peptides

    PubMed Central

    Schafer, Jamie L.; Ries, Moritz; Guha, Natasha; Connole, Michelle; Colantonio, Arnaud D.; Wiertz, Emmanuel J.; Wilson, Nancy A.; Kaur, Amitinder; Evans, David T.

    2015-01-01

    Natural killer (NK) cell responses in primates are regulated in part through interactions between two highly polymorphic molecules, the killer-cell immunoglobulin-like receptors (KIRs) on NK cells and their major histocompatibility complex (MHC) class I ligands on target cells. We previously reported that the binding of a common MHC class I molecule in the rhesus macaque, Mamu-A1*002, to the inhibitory receptor Mamu-KIR3DL05 is stabilized by certain simian immunodeficiency virus (SIV) peptides, but not by others. Here we investigated the functional implications of these interactions by testing SIV peptides bound by Mamu-A1*002 for the ability to modulate Mamu-KIR3DL05+ NK cell responses. Twenty-eight of 75 SIV peptides bound by Mamu-A1*002 suppressed the cytolytic activity of primary Mamu-KIR3DL05+ NK cells, including three immunodominant CD8+ T cell epitopes previously shown to stabilize Mamu-A1*002 tetramer binding to Mamu-KIR3DL05. Substitutions at C-terminal positions changed inhibitory peptides into disinhibitory peptides, and vice versa, without altering binding to Mamu-A1*002. The functional effects of these peptide variants on NK cell responses also corresponded to their effects on Mamu-A1*002 tetramer binding to Mamu-KIR3DL05. In assays with mixtures of inhibitory and disinhibitory peptides, low concentrations of inhibitory peptides dominated to suppress NK cell responses. Consistent with the inhibition of Mamu-KIR3DL05+ NK cells by viral epitopes presented by Mamu-A1*002, SIV replication was significantly higher in Mamu-A1*002+ CD4+ lymphocytes co-cultured with Mamu-KIR3DL05+ NK cells than with Mamu-KIR3DL05- NK cells. These results demonstrate that viral peptides can differentially affect NK cell responses by modulating MHC class I interactions with inhibitory KIRs, and provide a mechanism by which immunodeficiency viruses may evade NK cell responses. PMID:26333068

  7. Suppression of a Natural Killer Cell Response by Simian Immunodeficiency Virus Peptides.

    PubMed

    Schafer, Jamie L; Ries, Moritz; Guha, Natasha; Connole, Michelle; Colantonio, Arnaud D; Wiertz, Emmanuel J; Wilson, Nancy A; Kaur, Amitinder; Evans, David T

    2015-09-01

    Natural killer (NK) cell responses in primates are regulated in part through interactions between two highly polymorphic molecules, the killer-cell immunoglobulin-like receptors (KIRs) on NK cells and their major histocompatibility complex (MHC) class I ligands on target cells. We previously reported that the binding of a common MHC class I molecule in the rhesus macaque, Mamu-A1*002, to the inhibitory receptor Mamu-KIR3DL05 is stabilized by certain simian immunodeficiency virus (SIV) peptides, but not by others. Here we investigated the functional implications of these interactions by testing SIV peptides bound by Mamu-A1*002 for the ability to modulate Mamu-KIR3DL05+ NK cell responses. Twenty-eight of 75 SIV peptides bound by Mamu-A1*002 suppressed the cytolytic activity of primary Mamu-KIR3DL05+ NK cells, including three immunodominant CD8+ T cell epitopes previously shown to stabilize Mamu-A1*002 tetramer binding to Mamu-KIR3DL05. Substitutions at C-terminal positions changed inhibitory peptides into disinhibitory peptides, and vice versa, without altering binding to Mamu-A1*002. The functional effects of these peptide variants on NK cell responses also corresponded to their effects on Mamu-A1*002 tetramer binding to Mamu-KIR3DL05. In assays with mixtures of inhibitory and disinhibitory peptides, low concentrations of inhibitory peptides dominated to suppress NK cell responses. Consistent with the inhibition of Mamu-KIR3DL05+ NK cells by viral epitopes presented by Mamu-A1*002, SIV replication was significantly higher in Mamu-A1*002+ CD4+ lymphocytes co-cultured with Mamu-KIR3DL05+ NK cells than with Mamu-KIR3DL05- NK cells. These results demonstrate that viral peptides can differentially affect NK cell responses by modulating MHC class I interactions with inhibitory KIRs, and provide a mechanism by which immunodeficiency viruses may evade NK cell responses.

  8. hIFN-α gene modification augments human natural killer cell line anti-human hepatocellular carcinoma function.

    PubMed

    Jiang, W; Zhang, C; Tian, Z; Zhang, J

    2013-11-01

    Natural killer (NK) cells are characterized by an efficient antitumor activity, and this activity has been exploited as the basis of cancer immunotherapy strategies. Interferon-α (IFN-α) is an important cytokine required for induction of the durable antitumor immune response and is an important stimulator of NK cells. In this study, to augment the efficiency of NK cell cytotoxicity to tumor cells, human IFN-α gene-modified natural killer cell line (NKL) (NKL-IFNα) cells, which could stably secrete IFN-α, were established. We investigated the natural cytotoxicity of NKL-IFNα cells against human hepatocarcinoma cells (HCCs) in vitro and in vivo. NKL-IFNα cells displayed a significantly stronger cytolytic activity against both human HCC cell lines and primary human hepatoma cancer cells compared with parental NKL cells. The increased cytolytic activity of NKL-IFNα cells was associated with the upregulation of cytotoxicity-related genes, such as perforin, granzyme B and Fas ligand, in the NK cells. Moreover, cytokines secreted by NKL-IFNα cells, such as tumor necrosis factor-α and IFN-γ, induced increased expression of Fas on the target HCC cells, and resulted in increased susceptibility of the HCC cells to NK-mediated cytolysis. Encouragingly, NKL-IFNα cells could significantly inhibit HCC tumor growth in a xenograft model and prolonged the survival of tumor-bearing nude mice. These results suggest that IFN-α gene-modified NKL cells could be suitable for the future development of cell-based immunotherapeutic strategies for hepatocellular carcinoma.

  9. Structural Basis for Phototoxicity of the Genetically Encoded Photosensitizer KillerRed

    SciTech Connect

    Pletnev, Sergei; Gurskaya, Nadya G.; Pletneva, Nadya V.; Lukyanov, Konstantin A.; Chudakov, Dmitri M.; Martynov, Vladimir I.; Popov, Vladimir O.; Kovalchuk, Mikhail V.; Wlodawer, Alexander; Dauter, Zbigniew; Pletnev, Vladimir

    2009-11-23

    KillerRed is the only known fluorescent protein that demonstrates notable phototoxicity, exceeding that of the other green and red fluorescent proteins by at least 1,000-fold. KillerRed could serve as an instrument to inactivate target proteins or to kill cell populations in photodynamic therapy. However, the nature of KillerRed phototoxicity has remained unclear, impeding the development of more phototoxic variants. Here we present the results of a high resolution crystallographic study of KillerRed in the active fluorescent and in the photobleached non-fluorescent states. A unique and striking feature of the structure is a water-filled channel reaching the chromophore area from the end cap of the {beta}-barrel that is probably one of the key structural features responsible for phototoxicity. A study of the structure-function relationship of KillerRed, supported by structure-based, site-directed mutagenesis, has also revealed the key residues most likely responsible for the phototoxic effect. In particular, Glu68 and Ser119, located adjacent to the chromophore, have been assigned as the primary trigger of the reaction chain.

  10. Intravital Imaging – Dynamic Insights into Natural Killer T Cell Biology

    PubMed Central

    Liew, Pei Xiong; Kubes, Paul

    2015-01-01

    Natural killer T (NKT) cells were first recognized more than two decades ago as a separate and distinct lymphocyte lineage that modulates an expansive range of immune responses. As innate immune cells, NKT cells are activated early during inflammation and infection, and can subsequently stimulate or suppress the ensuing immune response. As a result, researchers hope to harness the immunomodulatory properties of NKT cells to treat a variety of diseases. However, many questions still remain unanswered regarding the biology of NKT cells, including how these cells traffic from the thymus to peripheral organs and how they play such contrasting roles in different immune responses and diseases. In this new era of intravital fluorescence microscopy, we are now able to employ this powerful tool to provide quantitative and dynamic insights into NKT cell biology including cellular dynamics, patrolling, and immunoregulatory functions with exquisite resolution. This review will highlight and discuss recent studies that use intravital imaging to understand the spectrum of NKT cell behavior in a variety of animal models. PMID:26042123

  11. Acute stress reduces intraparenchymal lung natural killer cells via beta-adrenergic stimulation

    PubMed Central

    Kanemi, O; Zhang, X; Sakamoto, Y; Ebina, M; Nagatomi, R

    2005-01-01

    There are lines of evidence that natural killer (NK) cells are sensitive to physical and psychological stress. Alterations in the immune system including NK cells are known to differ among tissues and organs. The effect of stress on the lung immune system, however, has not been well documented in spite of the fact that the lungs always confront viral or bacterial attacks as well as tumour cell metastasis. In this study, we intended to investigate the effect of restraint stress on lung lymphocytes including NK cells. C57BL/6 mice were exposed to 2 h restraint stress. The concentration of plasma epinephrine significantly rose immediately after the release from restraint as compared to home-cage control mice. Flow cytometric analysis revealed that the numbers of most lymphocyte subsets including NK cells were decreased in the lungs and blood but not in the spleen, immediately after restraint stress. Immunohistochemical examination revealed that the number of NK cells was decreased in the intraparenchymal region of the lungs, while the number of alveolar macrophages did not change. The decrease in the number of NK cells in the lungs and blood was reversed by the administration of propranolol, a nonselective beta adrenergic antagonist. Taken together, our findings suggest that acute stress reduces the number of intraparenchymal lung NK cells via activation of beta adrenergic receptors. PMID:15606610

  12. The Application of Natural Killer Cell Immunotherapy for the Treatment of Cancer

    PubMed Central

    Rezvani, Katayoun; Rouce, Rayne H.

    2015-01-01

    Natural killer (NK) cells are essential components of the innate immune system and play a critical role in host immunity against cancer. Recent progress in our understanding of NK cell immunobiology has paved the way for novel NK cell-based therapeutic strategies for the treatment of cancer. In this review, we will focus on recent advances in the field of NK cell immunotherapy, including augmentation of antibody-dependent cellular cytotoxicity, manipulation of receptor-mediated activation, and adoptive immunotherapy with ex vivo-expanded, chimeric antigen receptor (CAR)-engineered, or engager-modified NK cells. In contrast to T lymphocytes, donor NK cells do not attack non-hematopoietic tissues, suggesting that an NK-mediated antitumor effect can be achieved in the absence of graft-vs.-host disease. Despite reports of clinical efficacy, a number of factors limit the application of NK cell immunotherapy for the treatment of cancer, such as the failure of infused NK cells to expand and persist in vivo. Therefore, efforts to enhance the therapeutic benefit of NK cell-based immunotherapy by developing strategies to manipulate the NK cell product, host factors, and tumor targets are the subject of intense research. In the preclinical setting, genetic engineering of NK cells to express CARs to redirect their antitumor specificity has shown significant promise. Given the short lifespan and potent cytolytic function of mature NK cells, they are attractive candidate effector cells to express CARs for adoptive immunotherapies. Another innovative approach to redirect NK cytotoxicity towards tumor cells is to create either bispecific or trispecific antibodies, thus augmenting cytotoxicity against tumor-associated antigens. These are exciting times for the study of NK cells; with recent advances in the field of NK cell biology and translational research, it is likely that NK cell immunotherapy will move to the forefront of cancer immunotherapy over the next few years. PMID

  13. Oncogenicity of human papillomavirus- or adenovirus-transformed cells correlates with resistance to lysis by natural killer cells.

    PubMed Central

    Routes, J M; Ryan, S

    1995-01-01

    The reasons for the dissimilar oncogenicities of human adenoviruses and human papillomaviruses (HPV) in humans are unknown but may relate to differences in the capacities of the E1A and E7 proteins to target cells for rejection by the host natural killer (NK) cell response. As one test of this hypothesis, we compared the abilities of E1A- and E7-expressing human fibroblastic or keratinocyte-derived human cells to be selectively killed by either unstimulated or interferon (IFN)-activated NK cells. Cells expressing the E1A oncoprotein were selectively killed by unstimulated NK cells, while the same parental cells but expressing the HPV type 16 (HPV-16) or HPV-18 E7 oncoprotein were resistant to NK cell lysis. The ability of IFN-activated NK cells to selectively kill virally transformed cells depends on IFN's ability to induce resistance to NK cell lysis in normal (i.e., non-viral oncogene-expressing) but not virally transformed cells. E1A blocked IFN's induction of cytolytic resistance, resulting in the selective lysis of adenovirus-transformed cells by IFN-activated NK cells. The extent of IFN-induced NK cell killing of E1A-expressing cells was proportional to the level of E1A expression and correlated with the ability of E1A to block IFN-stimulated gene expression in target cells. In contrast, E7 blocked neither IFN-stimulated gene expression nor IFN's induction of cytolytic resistance, thereby precluding the selective lysis of HPV-transformed cells by IFN-activated NK cells. In conclusion, E1A expression marks cells for destruction by the host NK cell response, whereas the E7 oncoprotein lacks this activity. PMID:7494272

  14. Natural killer cell immunotherapy for cancer: a new hope.

    PubMed

    Srivastava, S; Lundqvist, A; Childs, R W

    2008-01-01

    Recently there has been a substantial gain in our understanding of the role NK-cells play in mediating innate host immune responses. Although NK cells have long been known to mediate antigen independent tumor cytotoxicity, the therapeutic potential of NK cell-based immunotherapy has yet to be realized. Manipulating the balance between inhibitory and activating NK receptor signals, sensitization of tumor target cells to NK cell-mediated apoptosis, and recent discoveries in NK-cell receptor biology have fueled translational research that has led to clinical trials investigating a number of novel methods to potentiate NK cytotoxicity against human malignancies. PMID:19089686

  15. Natural killer T cell strategies to combat Epstein–Barr virus infection

    PubMed Central

    Priatel, John J; Chung, Brian K; Tsai, Kevin; Tan, Rusung

    2014-01-01

    Epstein–Barr virus (EBV) infection results in rapid loss of CD1d expression from the surface of infected B cells, thus enabling the virus to evade immune recognition by natural killer T (NKT) cells. Using pharmacologic means to boost CD1d expression, potent NKT cell effector functions can be elicited toward EBV-infected B cells, suggesting the promise of novel strategies to target EBV-associated diseases such as some B-cell malignancies. PMID:25050206

  16. Third-party CD4+ invariant natural killer T cells protect from murine GVHD lethality.

    PubMed

    Schneidawind, Dominik; Baker, Jeanette; Pierini, Antonio; Buechele, Corina; Luong, Richard H; Meyer, Everett H; Negrin, Robert S

    2015-05-28

    Graft-versus-host disease (GVHD) is driven by extensive activation and proliferation of alloreactive donor T cells causing significant morbidity and mortality following allogeneic hematopoietic cell transplantation (HCT). Invariant natural killer T (iNKT) cells are a potent immunoregulatory T-cell subset in both humans and mice. Here, we explored the role of adoptively transferred third-party CD4(+) iNKT cells for protection from lethal GVHD in a murine model of allogeneic HCT across major histocompatibility barriers. We found that low numbers of CD4(+) iNKT cells from third-party mice resulted in a significant survival benefit with retained graft-versus-tumor effects. In vivo expansion of alloreactive T cells was diminished while displaying a T helper cell 2-biased phenotype. Notably, CD4(+) iNKT cells from third-party mice were as protective as CD4(+) iNKT cells from donor mice although third-party CD4(+) iNKT cells were rejected early after allogeneic HCT. Adoptive transfer of third-party CD4(+) iNKT cells resulted in a robust expansion of donor CD4(+)CD25(+)FoxP3(+) regulatory T cells (Tregs) that were required for protection from lethal GVHD. However, in vivo depletion of myeloid-derived suppressor cells abrogated both Treg expansion and protection from lethal GVHD. Despite the fact that iNKT cells are a rare cell population, the almost unlimited third-party availability and feasibility of in vitro expansion provide the basis for clinical translation.

  17. Decreased Cytotoxicity of Peripheral and Peritoneal Natural Killer Cell in Endometriosis

    PubMed Central

    Jeung, InCheul; Cheon, Keunyoung; Kim, Mee-Ran

    2016-01-01

    Endometriosis causes significant chronic pelvic pain, dysmenorrhea, and infertility and affects 10% of all women. In endometriosis, ectopic endometrium surviving after retrograde menstruation exhibits an abnormal immune response characterized by increased levels of activated macrophages and inflammatory cytokines. Particularly, dysfunctional natural killer (NK) cells play an important role in the pathogenesis of the disease by either facilitating or inhibiting the survival, implantation, and proliferation of endometrial cells. NK cells in the peritoneum and peritoneal fluid exhibit reduced levels of cytotoxicity in women with endometriosis. Several cytokines and inhibitory factors in the serum and peritoneal fluid also dysregulate NK cell cytotoxicity. Additionally, increased numbers of immature peripheral NK cells and induction of NK cell apoptosis are evident in the peritoneal fluid of women with endometriosis. The high rate of endometriosis recurrence after pharmaceutical or surgical treatment, which is associated with dysfunctional NK cells, indicates that new immunomodulatory management strategies are required. A good understanding of immune dysfunction would enable improvement of current treatments for endometriosis. PMID:27294113

  18. Underground Adaptation to a Hostile Environment: Acute Myeloid Leukemia vs. Natural Killer Cells

    PubMed Central

    Dulphy, Nicolas; Chrétien, Anne-Sophie; Khaznadar, Zena; Fauriat, Cyril; Nanbakhsh, Arash; Caignard, Anne; Chouaib, Salem; Olive, Daniel; Toubert, Antoine

    2016-01-01

    Acute myeloid leukemia (AML) is a heterogeneous group of malignancies which incidence increases with age. The disease affects the differentiation of hematopoietic stem or precursor cells in the bone marrow and can be related to abnormal cytogenetic and/or specific mutational patterns. AML blasts can be sensitive to natural killer (NK) cell antitumor response. However, NK cells are frequently defective in AML patients leading to tumor escape. NK cell defects affect not only the expression of the activating NK receptors, including the natural cytotoxicity receptors, the NK group 2, member D, and the DNAX accessory molecule-1, but also cytotoxicity and IFN-γ release. Such perturbations in NK cell physiology could be related to the adaptation of the AML to the immune pressure and more generally to patient’s clinical features. Various mechanisms are potentially involved in the inhibition of NK-cell functions in AML, including defects in the normal lymphopoiesis, reduced expression of activating receptors through cell-to-cell contacts, and production of immunosuppressive soluble agents by leukemic blasts. Therefore, the continuous cross-talk between AML and NK cells participates to the leukemia immune escape and eventually to patient’s relapse. Methods to restore or stimulate NK cells seem to be attractive strategies to treat patients once the complete remission is achieved. Moreover, our capacity in stimulating the NK cell functions could lead to the development of preemptive strategies to eliminate leukemia-initiating cells before the emergence of the disease in elderly individuals presenting preleukemic mutations in hematopoietic stem cells. PMID:27014273

  19. Serglycin determines secretory granule repertoire and regulates natural killer cell and cytotoxic T lymphocyte cytotoxicity.

    PubMed

    Sutton, Vivien R; Brennan, Amelia J; Ellis, Sarah; Danne, Jill; Thia, Kevin; Jenkins, Misty R; Voskoboinik, Ilia; Pejler, Gunnar; Johnstone, Ricky W; Andrews, Daniel M; Trapani, Joseph A

    2016-03-01

    The anionic proteoglycan serglycin is a major constituent of secretory granules in cytotoxic T lymphocyte (CTL)/natural killer (NK) cells, and is proposed to promote the safe storage of the mostly cationic granule toxins, granzymes and perforin. Despite the extensive defects of mast cell function reported in serglycin gene-disrupted mice, no comprehensive study of physiologically relevant CTL/NK cell populations has been reported. We show that the cytotoxicity of serglycin-deficient CTL and NK cells is severely compromised but can be partly compensated in both cell types when they become activated. Reduced intracellular granzyme B levels were noted, particularly in CD27(+) CD11b(+) mature NK cells, whereas serglycin(-/-) TCR-transgenic (OTI) CD8 T cells also had reduced perforin stores. Culture supernatants from serglycin(-/-) OTI T cells and interleukin-2-activated NK contained increased granzyme B, linking reduced storage with heightened export. By contrast, granzyme A was not significantly reduced in cells lacking serglycin, indicating differentially regulated trafficking and/or storage for the two granzymes. A quantitative analysis of different granule classes by transmission electronmicroscopy showed a selective loss of dense-core granules in serglycin(-/-) CD8(+) CTLs, although other granule types were maintained quantitatively. The findings of the present study show that serglycin plays a critical role in the maturation of dense-core cytotoxic granules in cytotoxic lymphocytes and the trafficking and storage of perforin and granzyme B, whereas granzyme A is unaffected. The skewed retention of cytotoxic effector molecules markedly reduces CTL/NK cell cytotoxicity, although this is partly compensated for as a result of activating the cells by physiological means.

  20. Natural killer-cell function in hemodialysis patients: effect of the dialysis membrane.

    PubMed

    Zaoui, P; Hakim, R M

    1993-06-01

    Natural killer (NK) cells are specific peripheral blood lymphocytes which are involved in the lysis of malignant and virally transformed