Viral Antigens Trigger Granzyme B Secretion by Human CD5+ B1 Cells in the Presence of Interleukin 21.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4729-4729
Author(s):  
Kai Sontheimer ◽  
Sue Blackwell ◽  
Magdalena Hagn ◽  
Verena Ebel ◽  
Thamara Beyer ◽  
...  
Keyword(s):  
Nk Cells ◽  
Conflicts Of Interest ◽  
Granzyme B ◽  
Elevated Serum ◽  
Immune Defense ◽  
Innate Immune ◽  
Viral Antigens ◽  
Wide Range ◽  
Interleukin 21 ◽  
B1 Cells

Abstract Abstract 4729 CD5+ B1 cells are thought to play a key role in innate immune responses by producing and secreting natural immunoglobulins, which bind to a wide range of bacterial, viral antigens as well as auto-antigens. Here we demonstrate that B1 cells from human cord blood are able to express the pro-apoptotic serine protease granzyme B (GrB) in response to viruses including tick-borne encephalitis virus, rabies virus, and hepatitis B virus. Using pharmacological tools we reveal that this response depends on B cell receptor stimulation, toll-like receptor 9 engagement and interleukin 21 (IL-21), a cytokine primarily found in the acute phase of viral infections. While up to 35% of freshly isolated B1 cells directly respond with GrB expression to stimulation with inactivated viruses in the presence of IL-21, similar, though lower reactions in CD5- B2 cells are only found after, but not before vaccination of the donor against the respective virus. Of note, GrB-expressing B1 cells feature a homogeneous CD19+CD5+GrB+CD20+CD27-CD38-IgD- phenotype and B1 cell-derived GrB is secreted in an enzymatically active state, reaching levels comparable to those secreted by activated cytotoxic cells. GrB induction requires activation of similar signaling pathways as in CTL and NK cells including members of the JAK/STAT pathway. Our findings suggest GrB secretion by B1 cells represents a novel innate immune response mechanism. GrB-secreting B1 cells may play a role in early anti-viral immune defense, and may contribute to elevated serum GrB levels found in various viral diseases. Further studies will elucidate whether CD5+ B1 cells, possibly in co-operation with NK cells, exhibit cytotoxicity towards virus-infected as well as tumor cells. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Viral Antigens Trigger Granzyme B Secretion by Human CD5+ B1 Cells in the Presence of Interleukin 21

2010 ◽  
Vol 24 (S1) ◽  
Author(s):  
Kai Sontheimer ◽  
Magdalena Hagn ◽  
Verena Ebel ◽  
Thamara Beyer ◽  
Thomas Simmet ◽  
...  
Keyword(s):  
Granzyme B ◽  
Viral Antigens ◽  
Interleukin 21 ◽  
B1 Cells

Enhancement of Natural Killer Cell Interferon-Gamma Production By the Derivatives of the Natural Product Phyllanthusmins Via TLR-Mediated NF-Kb and STAT3 Signaling Pathways

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 1031-1031
Author(s):  
Luxi Chen ◽  
Long Yi ◽  
Yuning Ren ◽  
Jianying Zhang ◽  
A. Douglas Kinghorn ◽  
...  
Keyword(s):  
Nk Cells ◽  
Signaling Pathways ◽  
Natural Product ◽  
Natural Killer ◽  
Nk Cell ◽  
Conflicts Of Interest ◽  
Killer Cell ◽  
Immune Defense ◽  
Ifn Gamma ◽  
Stat3 Signaling

Abstract Natural killer (NK) cells are known to play a pivotal role in regulating immune defense against tumors and viruses. However, very few studies describe the effects of natural products or their derivatives on NK cell function. We previously showed that the natural product lignan phyllanthusmin C enhances human NK cell IFN-gamma production. In this study, we successfully synthesized dozens of phyllanthusmin derivatives and screened their activities to enhance IFN-gamma production by NK cells. We found that three of these derivatives possessed capacities to induce IFN-gamma production by NK cells. Among them, derivative No. 17 had the highest efficacy and was significantly superior (P < 0.05) to that of the original phyllanthusmin C. These three phyllanthusmin derivatives had no adverse effect on human NK cell survival or proliferation. All derivatives alone or in combination with IL-12 or IL-15 induced human NK cell interferon (IFN)-gamma in comparison with the corresponding vehicle control or cytokine alone (P < 0.05). These derivatives stimulated IFN-gamma production in both CD56bright and CD56dim human NK cell subsets (P < 0.01). Mechanistically, immunoblotting assays and chromatin immunoprecipitation (ChIP) analyses combined with promoter-reporter luciferase assays revealed that these phyllanthusmin derivatives induced the phosphorylation of NF-κB and STAT3, resulting in their increased binding on theIFN G promoter, which was dependent on the Toll-like receptor (TLR) 1 and the TLR3 signaling pathways, respectively. STAT3 knockdown with lentivirus shRNA and inhibition of NF-kB signaling with a specific inhibitor (TPCK) significantly attenuated IFN-gamma production induced by these phyllanthusmin derivatives in human NK cells (P < 0.05). Blockade of TLR1 or TLR3 with their corresponding neutralizing antibodies nearly abolished activation of NF-κB or STAT3 as well as IFN-γ induction by the phyllanthusmin derivatives (P < 0.05). In conclusion, we have successfully synthesized and screened phyllanthusmin derivatives and identified three compounds that induce IFN-gamma production in human NK cells, one of which had optimal potential. The induction of human NK cell IFN-gamma production by phyllanthusmin derivatives is mediated by TLR1-NF-κB and TLR3-STAT3 signaling pathways, rendering their potential for use as agents for cancer prevention or treatment. Disclosures No relevant conflicts of interest to declare.

scholarly journals PATTERN-RECOGNIZING RECEPTORS AND THE INNATE IMMUNE RESPONSE TO VIRAL INFECTION

2018 ◽  
Keyword(s):  
Immune Response ◽  
Innate Immunity ◽  
Viral Infection ◽  
Innate Immune Response ◽  
English Literature ◽  
Innate Immune ◽  
Viral Pathogens ◽  
Antiviral Immune Response ◽  
Viral Antigens ◽  
Wide Range

The innate immune response to viral pathogens is crucial in mobilizing defensive reactions of an organism during the development of an acute viral infection. Cells of the innate immunity system detect viral antigens due to genetically programmed pattern-recognition receptors (PRRs), which are located either on the cell surface or inside the certain intracellular components. These image-recognizing receptors include Toll-like receptors (TLRs), retinoic acid-inducible gene I-like receptors (RIG-I-like receptors), nucleotide oligomerization domain-like receptors (NOD-like receptors), also known as NACHT, LRR and PYD domains of the protein, and cytosolic DNA sensors. The trigger mechanisms for these receptors are viral proteins, and nucleic acids serve as activators. The presence of PRRs that are responsible for the determination of viral antigens in cellular components allows the cells of innate immunity to recognize a wide range of viral agents that replicate in various cellular structures, and develop an immune response to them. This article summarizes the disparate data presented in modern English literature on the role of PRRs and the associated signaling pathways. Understanding the recognition of viral pathogens required triggering a cascade of cytokine and interferon production provides insights into how viruses activate the signal paths of PRRs and the effect of the interaction of viral antigens and these receptors on the formation of the antiviral immune response.

scholarly journals Plant Antimicrobial Peptides as Potential Anticancer Agents

2015 ◽  
Vol 2015 ◽  
pp. 1-11 ◽  
Author(s):  
Jaquelina Julia Guzmán-Rodríguez ◽  
Alejandra Ochoa-Zarzosa ◽  
Rodolfo López-Gómez ◽  
Joel E. López-Meza
Keyword(s):  
Antimicrobial Peptides ◽  
Anticancer Agents ◽  
Pathogenic Bacteria ◽  
Defense Mechanism ◽  
Immune Defense ◽  
Innate Immune ◽  
High Morbidity ◽  
Anticancer Activities ◽  
Innate Immune Defense ◽  
Wide Range

Antimicrobial peptides (AMPs) are part of the innate immune defense mechanism of many organisms and are promising candidates to treat infections caused by pathogenic bacteria to animals and humans. AMPs also display anticancer activities because of their ability to inactivate a wide range of cancer cells. Cancer remains a cause of high morbidity and mortality worldwide. Therefore, the development of methods for its control is desirable. Attractive alternatives include plant AMP thionins, defensins, and cyclotides, which have anticancer activities. Here, we provide an overview of plant AMPs anticancer activities, with an emphasis on their mode of action, their selectivity, and their efficacy.

Validation of Flow Cytometric Measurements of NK Cytotoxicity Against Chromium Release Assay for Use in Clinical Monitoring of Immune Function.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 5287-5287
Author(s):  
Katherine Smith ◽  
Glenn Heller ◽  
Peter Maslak
Keyword(s):  
Nk Cells ◽  
Normal Control ◽  
Granzyme B ◽  
Clinical Samples ◽  
Target Cells ◽  
51Cr Release ◽  
Effector Cells ◽  
Flow Cytometric ◽  
Wide Range ◽  
Nk Cytotoxicity

Abstract Chromium (51Cr) release assays have long been a standard technique for measuring the cytolytic activity of various immune effector cells. Although relatively easy to perform, these assays suffer from problems with sensitivity and require special radiation precautions limiting their routine use in clinical laboratories. Several flow cytometric techniques measuring cytotoxicity have been introduced and used as a measure of immune response. These assays are more rapid than the 51Cr based assay and have the added advantage of freeing the laboratory from the precautions required when working with radioactivity. Such testing has however largely been restricted to the research setting. In an attempt to extend these benefits to a clinical laboratory, we attempted to validate two commercially available flow cytometry kits (CyToxiLux Plus® & GranToxiLux®, OncoImmunin, Gathersburg, MD) measuring either caspase or granzyme B activity inside K562 target cells induced by interaction with NK cells. These assays were then compared with a standard NK cytotoxicity assay using 51Cr release technique. With K562 as a target, we parallel tested three CD56+CD3-KIR- NK clones; both the 51Cr and caspase based cytotoxicity values were highly positive. With WH autologous EBV cell line as target, we parallel tested five NK clones and five normal control donors as effectors. As expected, all cytotoxicity values for both assays were <10%. The caspase and granzyme assays were then compared and the degree of separation in the flow cytometric plots of the granzyme based assay were found to be marginally superior facilitating analysis of the data. The granzyme B assay therefore was chosen for investigation in the clinical samples. The granzyme and 51Cr assays were then compared both in 49 normal control donors and in 10 patients. There was a high level of concordance between the two assays. Comparing all results using linear correlation yielded a squared correlation (R2) value of 0.6878. However, given the wide range of NK activity in normal control donors the degree of positivity may not be clinically significant. Therefore, the data was examined in the context of a positive 51Cr response. Using a 2×2 table to exam the level of agreement, the kappa statistic was 0.63, indicating good agreement between the assays. When progressively fewer effector cells were added to a fixed number of target cells, the % cytotoxicity decreased from 39% to 0% with the flow cytometric assay and from 26% to 1% with the 51Cr assay. In the normal control donors the positive values ranged from 17% to 65% cytotoxicity using the flow assay compared with the previous range of 15% to 72% established with 51Cr. Patients having cytotoxicity values below 17% were deemed as having an unreactive or abnormal low result. One of the patients with few CD56+CD3- NK cells had consistently low cytotoxicity each time tested. For the patients who have either undergone allogeneic stem cell transplantation or have a known immunodeficiency, further clinical follow-up will serve to establish the clinical relevance of these measurements. These preliminary data suggest that the flow cytometric technique can be used in the clinical setting for immune monitoring.

Interferon-γ (IFN-γ) Mediates a Direct Fungicidal Effect of Human Natural Killer (NK) Cells Against Aspergillus Fumigatus

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2779-2779
Author(s):  
Maria Bouzani ◽  
Michael Ok ◽  
Allison Mc Cormick ◽  
Frank Ebel ◽  
Hermann Einsele ◽  
...  
Keyword(s):  
Nk Cells ◽  
Fungicidal Activity ◽  
Conflicts Of Interest ◽  
Interleukin 2 ◽  
Innate Immune ◽  
Physical Contact ◽  
Soluble Factor ◽  
Fungicidal Effect ◽  
Cytotoxic Proteins ◽  
Ifn Γ

Abstract Abstract 2779 NK cells are innate immune effector cells, which are well recognized for their role against cells transformed by malignant processes or by an intracellular infection. Although, there is increasing evidence of an interaction of NK cells also with extracellular pathogens, less is known about their interplay with moulds and especially with A. fumigatus. The interest for this interaction is mainly driven by the high mortality characterizing invasive aspergillosis (IA), an infection of immunosuppressed patients caused mainly by this fungus. In our study, we investigated the in vitro interaction of human NK cells with A. fumigatus. Methods. NK cells were magnetically depleted from the peripheral blood of healthy volunteers followed by pulsing for 24h with 500 U/ml recombinant interleukin–2 (rhIL-2). The induction of IFN-γ and Tumor Necrosis Factor-α (TNF-α) by the fungus was measured by real time RT-PCR and ELISA assays after 0, 3, 6 and 12h of co-culture. The lethal impact of NK cells and culture supernatants on A. fumigatus was evaluated by plate killing and 2,3-bis[2-methoxy-4-nitro-5-sulpho-phenyl]2H-tetrazolium-5-carboxanilide (XTT) assays. Transwell permeable membranes with pores of 0.4 μm, prohibiting the direct contact of cells placed on opposite sides but allowing the free circulation of molecules were used to evaluate the relevance of a direct cell – fungal contact on cytokine induction and fungal damage. The degranulation markers CD107a and CD107b were measured to assess the involvement of cytotoxic proteins on the anti-Aspergillus effect. The supernatant was magnetically depleted of IFN-γ. Recombinant human IFN-γ (rhIFN-γ) was used to mimic the effect of IFN-γ released by NK cells. Results. Co-cultivating human NK cells with resting (conidia) and germinated (germlings) fungal morphologies, only germlings induce a Th1 like cytokine response of NK cells, with the major representative IFN-γ (p<.05). Consistently, NK cells demonstrate a strong fungicidal effect only towards germlings (p<.05). Using transwell membranes to investigate the mechanism of this interaction, we found that physical contact was a prerequisite for the induction of cytokines (p<.05), but also for the release of a soluble factor, exhibiting fungicidal properties. Once the soluble factor was released, germlings were killed without the presence of NK cells (p<.05). To confirm this finding, germlings were incubated with supernatant obtained from the culture of NK cells with A. fumigatus. After 3 h of culture, 40% killing (p<.001) was demonstrated. To determine the soluble factor(s) causing the fungicidal effect, we explored the degranulation of cytotoxic proteins like perforin, granzymes and granulysin. FITC-labeled antibodies against the degranulation markers CD 107a, b were co-incubated with unstimulated NK cells, NK cells challenging germlings and NK cells challenging the cell line K562. In the presence of A. fumigatus, there was no expression of CD 107a, b, while the expression of these markers was significant in the culture with K562 cells. This result suggests that the granule proteins are not the means of the NK cell cytotoxicity against A. fumigatus. Moreover, the protective effect of IFN- γ for the host defense against IA is well known. So far, this effect was attributed to the influence of IFN- γ on innate immune cells, increasing their antifungal activity. In our study, we hypothesized that IFN- γ might demonstrate direct antifungal properties, mediating the fungicidal activity of NK cells. Thus, we compared the fungal killing caused by IFN- γ partially depleted supernatant and un-depleted supernatant and we found that the depletion of IFN- γ diminished the fungicidal effect (p<.05). Moreover, we exposed germlings to culture medium enriched with rhIFN- γ at concentrations equivalent to those detected in the supernatants and we observed a similar killing rates. To summarize, our study demonstrates that human NK cells interact with A. fumigatus germlings, which induce the release of Th1 like cytokines and cause significant fungal killing. Our data show for the first time that IFN-γ, released by NK cells, is a soluble molecule mediating direct fungicidal activity. These results attribute new characteristics to human NK cells, suggesting them as a potential alternative therapeutic tool against IA. Disclosures: No relevant conflicts of interest to declare.

Natural Killer Cells Display an Activated Phenotype but Reduced Effector Functions in Obese Patients

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3430-3430
Author(s):  
Sebastien Viel ◽  
Laurie Besson ◽  
Emily Charrier ◽  
Jacques Bienvenu ◽  
Emmanuel Disse ◽  
...  
Keyword(s):  
Nk Cells ◽  
Natural Killer ◽  
Nk Cell ◽  
Conflicts Of Interest ◽  
Granzyme B ◽  
K562 Cells ◽  
Obese Patients ◽  
Activation Markers ◽  
Significant Difference ◽  
The Impact

Abstract The impact of adiposity on the immune system remains largely unexplored. While obesity has been suggested to be a predisposing or adverse prognostic factor in certain neoplastic diseases it is not yet clear to what extent this may involve the innate or adaptative immune systems. Adipose tissue produces a large number of secreted molecules, or adipocytokines, which may have immunomodulatory functions. This project aimed to determine whether phenotypical and/or functional properties of circulating natural killer (NK) cells were influenced by body mass index (BMI). In a preliminary study, 47 patients with no history of hematological malignancy were included, including 14 healthy volunteers with a normal BMI (18.5-25), 10 patients considered to be overweight (25 < BMI < 30), 11 patients considered as obese (BMI > 30) and 12 patients who were previously obese and had lost weight. Peripheral blood was analyzed by flow cytometry for the following markers: activating receptors (CD16, C161, DNAM-1, 2B4, NKG2C, NKG2D, NKp46, NKp30), inhibitor receptors (NKG2A, KIR2DL1, KIR2DL2, KIR3DL1), activation markers (CD69, granzyme B, NKG2C), maturation markers (CD56, CD57, CD94, CX3CR1) and cytotoxicity markers (perforin, NKG7). Moreover the capacity of NK cells to degranulate and to produce several cytokines (TNF, IFN-g) or chemokines (MIP1-b) in response to stimulation by K562 cells or Rituximab coated -tumor B cells was evaluated. Results showed a positive correlation between BMI and total number of circulating NK cells, with a significant difference between lean patients and obese patients. Immunophenotypic analyses showed that NKp46 and CD94 expression (measured by Mean Fluorescence Intensity) were both significantly reduced with increased BMI. NK cells from obese patients also show signs of activation, characterized by an elevation of the expression of CD69 and granzyme B and a reduction of the expression of CD16. The ability of NK cells to be activated in the presence of cell lines was also reduced in obese patients: NK cell secretion of IFN-g and MIP-1b in the presence of Granta cells or MIP-1b in the presence of K562 decreased linearly with increasing BMI. NK cell degranulation upon co-culture with K562 cells was also negatively correlated with BMI. In these different assays pre-obese and ex-obese patients scored intermediate between lean and obese patients. Overall these results suggest in vivo activation and exhaustion of NK cells in obese patients. These cells are thus potentially less likely to participate as effector cells in immunotherapeutic regimens. Disclosures No relevant conflicts of interest to declare.

scholarly journals Primary adrenal insufficiency is associated with impaired natural killer cell function: a potential link to increased mortality

2017 ◽  
Vol 176 (4) ◽  
pp. 471-480 ◽  
Author(s):  
Irina Bancos ◽  
Jon Hazeldine ◽  
Vasileios Chortis ◽  
Peter Hampson ◽  
Angela E Taylor ◽  
...  
Keyword(s):  
Nk Cells ◽  
Adrenal Insufficiency ◽  
Natural Killer ◽  
Nk Cell ◽  
Respiratory Infections ◽  
Immune Defense ◽  
Innate Immune ◽  
Receptor Expression ◽  
Cell Surface Receptor ◽  
Primary Adrenal Insufficiency

Objective Mortality in patients with primary adrenal insufficiency (PAI) is significantly increased, with respiratory infections as a major cause of death. Moreover, patients with PAI report an increased rate of non-fatal infections. Neutrophils and natural killer (NK) cells are innate immune cells that provide frontline protection against invading pathogens. Thus, we compared the function and phenotype of NK cells and neutrophils isolated from PAI patients and healthy controls to ascertain whether altered innate immune responses could be a contributory factor for the increased susceptibility of PAI patients to infection. Design and methods We undertook a cross-sectional study of 42 patients with PAI due to autoimmune adrenalitis (n = 37) or bilateral adrenalectomy (n = 5) and 58 sex- and age-matched controls. A comprehensive screen of innate immune function, consisting of measurements of neutrophil phagocytosis, reactive oxygen species production, NK cell cytotoxicity (NKCC) and NK cell surface receptor expression, was performed on all subjects. Results Neutrophil function did not differ between PAI and controls. However, NKCC was significantly reduced in PAI (12.0 ± 1.5% vs 21.1 ± 2.6%, P < 0.0001). Phenotypically, the percentage of NK cells expressing the activating receptors NKG2D and NKp46 was significantly lower in PAI, as was the surface density of NKG2D (all P < 0.0001). Intracellular granzyme B expression was significantly increased in NK cells from PAI patients (P < 0.01). Conclusions Adrenal insufficiency is associated with significantly decreased NKCC, thereby potentially compromising early recognition and elimination of virally infected cells. This potential impairment in anti-viral immune defense may contribute to the increased rate of respiratory infections and ultimately mortality in PAI.

scholarly journals Inhibition of Human Natural Killer Cell Activity by Influenza Virions and Hemagglutinin

2010 ◽  
Vol 84 (9) ◽  
pp. 4148-4157 ◽  
Author(s):  
Huawei Mao ◽  
Wenwei Tu ◽  
Yinping Liu ◽  
Gang Qin ◽  
Jian Zheng ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Nk Cells ◽  
Natural Killer ◽  
Nk Cell ◽  
Cell Activity ◽  
Immune Defense ◽  
Innate Immune ◽  
Human Natural Killer Cell ◽  
Viral Pathogen ◽  
Infected Cells

ABSTRACT Natural killer (NK) cells keep viral infections under control at the early phase by directly killing infected cells. Influenza is an acute contagious respiratory viral disease transmitted from host-to-host in the first few days of infection. The evasion of host innate immune defenses including NK cells is important for its success as a viral pathogen of humans and animals. NK cells encounter influenza virus within the microenvironment of infected cells. It therefore is important to investigate the direct effects of influenza virus on NK cell activity. Recently we demonstrated that influenza virus directly infects human NK cells and induces cell apoptosis to counter their function (H. Mao, W. Tu, G. Qin, H. K. W. Law, S. F. Sia, P.-L. Chan, Y. Liu, K.-T. Lam, J. Zheng, M. Peiris, and Y.-L. Lau, J. Virol. 83:9215-9222, 2009). Here, we further demonstrated that both the intact influenza virion and free hemagglutinin protein inhibited the cytotoxicity of fresh and interleukin-2 (IL-2)-activated primary human NK cells. Hemagglutinin bound and internalized into NK cells via the sialic acids. This interaction did not decrease NKp46 expression but caused the downregulation of the ζ chain through the lysosomal pathway, which caused the decrease of NK cell cytotoxicity mediated by NKp46 and NKp30. The underlying dysregulation of the signaling pathway involved ζ chain downregulation, leading to decreased Syk and ERK activation and granule exocytosis upon target cell stimulation, finally causing reduced cytotoxicity. These findings suggest that influenza virus developed a novel strategy to evade NK cell innate immune defense that is likely to facilitate viral transmission and also contribute to virus pathogenesis.

Natural killer cell in the developing life

2015 ◽  
Vol 43 (1) ◽  
Author(s):  
Yen-Chang Lee ◽  
Syh-Jae Lin
Keyword(s):  
Nk Cells ◽  
Natural Killer ◽  
Immune Cells ◽  
Killer Cell ◽  
Immune Defense ◽  
Innate Immune ◽  
First Line ◽  
Successful Pregnancy ◽  
Neonatal Stage

AbstractNatural killer (NK) cells that provide first-line innate immune reactions against virus-infected and tumor cells have different roles in different body sites and in different stages. From the beginning of life, NK cells participate in many aspects of development, especially in a successful pregnancy and a healthy neonatal stage. This article reviews recent advances regarding the role of NK cells in implantation, placentation and immune tolerance during pregnancy as well as in the neonatal immune defense. The interactions between NK cells and other immune cells in each developmental stage are discussed.

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