scholarly journals Modulation of Secretory Lysosomes During NK Cell Education Leads to Accumulation of Granzyme B and Enhanced Functional Potential

2018 ◽  
Author(s):  
Jodie P. Goodridge ◽  
Benedikt Jacobs ◽  
Michelle L. Saetersmoen ◽  
Dennis Clement ◽  
Trevor Clancy ◽  
...  
Keyword(s):  
Nk Cells ◽  
Nk Cell ◽  
Granzyme B ◽  
Killer Cell ◽  
Cell Types ◽  
Functional Potential ◽  
Intracellular Mechanism ◽  
Inhibitory Signaling ◽  
Secretory Lysosomes ◽  
Morphological Phenotype

AbstractInhibitory signaling during natural killer (NK) cell education translates into increased responsiveness to activation; however the intracellular mechanism for functional tuning by inhibitory receptors remains unclear. We found that educated NK cells expressing self-MHC specific inhibitory killer cell immunoglobulin-like receptors (KIR) show accumulation of granzyme B, localized in dense-core secretory lysosomes, converged close to the centrosome. This discrete morphological phenotype persists in self-KIR+ NK cells independently of transcriptional programs that regulate effector function, metabolism and lysosomal biogenesis. The granzymeB dense, large secretory lysosomes in self-KIR+ NK cells were efficiently released upon target cell recognition, contributing to their enhanced cytotoxic capacity. Secretory lysosomes are part of the acidic lysosomal compartment, which has been shown to channel calcium and mediate intracellular signalling in several cell types. Interference of signaling from acidic Ca2+ stores in primary NK cells reduced both target-specific Ca2+-flux, degranulation and cytokine production. Furthermore, inhibition of PI(3,5)P2 synthesis or genetic silencing of the PI(3,5)P2-regulated lysosomal Ca2+-channel TRPML1 led to increased levels of granzyme B and enhanced functional potential. These results indicate an intrinsic role for lysosomal homeostasis in NK cell education.

scholarly journals Natural Killer Cell Derived Exosomes As a Novel Therapeutic for Acute Myeloid Leukemia

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 5226-5226
Author(s):  
Michael Boyiadzis ◽  
Chang Sook Hong ◽  
Theresa L Whiteside
Keyword(s):  
Flow Cytometry ◽  
Acute Myeloid Leukemia ◽  
Nk Cells ◽  
Natural Killer ◽  
Myeloid Leukemia ◽  
Nk Cell ◽  
Granzyme B ◽  
Killer Cell ◽  
Primary Leukemia ◽  
Acute Myeloid

Abstract Introduction: Natural killer (NK) cells play a critical role in the innate immune response through their capacity to lyse malignant cells without prior antigen-specific priming. NK cells have been evaluated for safety and efficacy in acute myeloid leukemia (AML), both in the transplant and non-transplant settings. Exosomes, small, 30-150 nm-sized extracellular vesicles originating from the endocytic compartment of parent cells, have recently emerged as a universal intercellular communication system. Exosomes are released by virtually all cells and carry proteins, lipids, and nucleic acids from the parent to recipient cells at short and long distances. The exosome molecular cargo reflects the content of the parent cell and is delivered to recipient cells in a membrane-protected vesicle. We hypothesize that exosomes produced by human activated NK cells carry the machinery necessary for the killing of leukemic cells. Methods: Venous blood (20-50 mL) was obtained from healthy donors (n=10). NK cells were isolated using Ab-based immunomagnetic selection from the recovered peripheral blood mononuclear cells. NK cells were cultured in the presence of interleukin-2 and interleukin-15, and NK-cell supernatants were used for exosome isolation by size exclusion chromatography. Protein levels, numbers and size (qNano), and exosome morphology (transmission electron microscopy) were determined. Exosome cargos were studied by Western blots and/or flow cytometry for NK cell activating and inhibitory receptors, immune inhibitory molecules and for perforin and granzyme B. Cytotoxicity of the NK cell-derived exosomes for K562 targets, AML cell lines (Kasumi, MLL-1) and primary leukemia blasts was measured using flow cytometry-based assays. Results: Activated human NK cells produced large quantities of exosomes. PKH-26-labeled NK cell-derived exosomes were avidly taken up by leukemic blasts. NK cell derived exosomes carried activating NK cell receptor NKG2D, natural cytotoxicity receptors, perforin, granzyme B, transforming growth factor beta (TGF-β), killer-cell immunoglobulin-like receptors and PD-1. NK cell derived exosomes mediated anti-leukemia activity against K562 targets, AML cell lines and primary leukemia blasts. Lysis of leukemic blasts by NK cell-derived exosomes was exosome concentration dependent. Conclusion: We report that NK cell derived exosomes have anti leukemia activity in vitro. These data provide a foundation for the future development of new therapeutic strategies using NK cell-derived exosomes for the elimination of leukemia. Disclosures No relevant conflicts of interest to declare.

scholarly journals The Human Cytomegalovirus Protein UL16 Mediates Increased Resistance to Natural Killer Cell Cytotoxicity through Resistance to Cytolytic Proteins

2003 ◽  
Vol 77 (8) ◽  
pp. 4539-4545 ◽  
Author(s):  
Jenny Odeberg ◽  
Helena Browne ◽  
Sunil Metkar ◽  
Christopher J. Froelich ◽  
Lars Brandén ◽  
...  
Keyword(s):  
Nk Cells ◽  
Human Cytomegalovirus ◽  
Deletion Mutant ◽  
Nk Cell ◽  
Granzyme B ◽  
Killer Cell ◽  
Target Cells ◽  
Natural Killer Cell Cytotoxicity ◽  
Hcmv Strain ◽  
Infected Cells

ABSTRACT Several reports have shown that human cytomegalovirus (HCMV)-infected cells are resistant to NK lysis. These studies have focused on receptor-ligand interactions, and different HCMV proteins have been indicated to mediate inhibitory NK signals. Here, we report that the HCMV protein UL16 is of major importance for the ability of HCMV-infected cells to resist NK cell-mediated cytotoxicity. Fibroblasts infected with the UL16 deletion mutant HCMV strain exhibited a 70% increased sensitivity to NK killing at 7 days postinfection compared to AD169-infected cells. Interestingly, HCMV-infected cells did not appear to engage inhibitory molecules on NK cells, since the levels of granzyme B were not reduced in supernatants obtained from NK cell cocultures with infected target cells compared to uninfected target cells. Furthermore, HCMV-infected cells, but not cells infected with the UL16 deletion mutant HCMV strain, exhibited a significantly increased resistance to the action of cytolytic proteins, including perforin, granzyme B, streptolysin O, and porcine NK lysin. In addition, fluorescence-activated cell sorting for UL16-positive transfected cells resulted in protection levels of 90% compared to control cells carrying the green fluorescent protein vector. Thus, the UL16 protein mediates an increased protection against the action of cytolytic proteins released by activated NK cells, possibly by a membrane-stabilizing mechanisms, rather than by delivering negative signals to NK cells.

scholarly journals The m15 Locus of Murine Cytomegalovirus Modulates Natural Killer Cell Responses to Promote Dissemination to the Salivary Glands and Viral Shedding

Pathogens ◽  
2021 ◽  
Vol 10 (7) ◽  
pp. 866
Author(s):  
Baca Chan ◽  
Maja Arapović ◽  
Laura Masters ◽  
Francois Rwandamuiye ◽  
Stipan Jonjić ◽  
...  
Keyword(s):  
Nk Cells ◽  
Salivary Glands ◽  
Natural Killer ◽  
Nk Cell ◽  
Acute Infection ◽  
Killer Cell ◽  
Viral Escape ◽  
Murine Cytomegalovirus ◽  
Viral Shedding ◽  
Cell Responses

As the largest herpesviruses, the 230 kb genomes of cytomegaloviruses (CMVs) have increased our understanding of host immunity and viral escape mechanisms, although many of the annotated genes remain as yet uncharacterised. Here we identify the m15 locus of murine CMV (MCMV) as a viral modulator of natural killer (NK) cell immunity. We show that, rather than discrete transcripts from the m14, m15 and m16 genes as annotated, there are five 3′-coterminal transcripts expressed over this region, all utilising a consensus polyA tail at the end of the m16 gene. Functional inactivation of any one of these genes had no measurable impact on viral replication. However, disruption of all five transcripts led to significantly attenuated dissemination to, and replication in, the salivary glands of multiple strains of mice, but normal growth during acute infection. Disruption of the m15 locus was associated with heightened NK cell responses, including enhanced proliferation and IFNγ production. Depletion of NK cells, but not T cells, rescued salivary gland replication and viral shedding. These data demonstrate the identification of multiple transcripts expressed by a single locus which modulate, perhaps in a concerted fashion, the function of anti-viral NK cells.

scholarly journals α-Pinene Enhances the Anticancer Activity of Natural Killer Cells via ERK/AKT Pathway

2021 ◽  
Vol 22 (2) ◽  
pp. 656
Author(s):  
Hantae Jo ◽  
Byungsun Cha ◽  
Haneul Kim ◽  
Sofia Brito ◽  
Byeong Mun Kwak ◽  
...  
Keyword(s):  
Nk Cells ◽  
Cancer Cells ◽  
Natural Killer ◽  
Nk Cell ◽  
Granzyme B ◽  
Akt Pathway ◽  
Cell Cytotoxicity ◽  
Anticancer Effects ◽  
Nk Cell Cytotoxicity

Natural killer (NK) cells are lymphocytes that can directly destroy cancer cells. When NK cells are activated, CD56 and CD107a markers are able to recognize cancer cells and release perforin and granzyme B proteins that induce apoptosis in the targeted cells. In this study, we focused on the role of phytoncides in activating NK cells and promoting anticancer effects. We tested the effects of several phytoncide compounds on NK-92mi cells and demonstrated that α-pinene treatment exhibited higher anticancer effects, as observed by the increased levels of perforin, granzyme B, CD56 and CD107a. Furthermore, α-pinene treatment in NK-92mi cells increased NK cell cytotoxicity in two different cell lines, and immunoblot assays revealed that the ERK/AKT pathway is involved in NK cell cytotoxicity in response to phytoncides. Furthermore, CT-26 colon cancer cells were allografted subcutaneously into BALB/c mice, and α-pinene treatment then inhibited allografted tumor growth. Our findings demonstrate that α-pinene activates NK cells and increases NK cell cytotoxicity, suggesting it is a potential compound for cancer immunotherapy.

scholarly journals Natural Killer Cell Mobilization in Breast and Prostate Cancer Survivors: The Implications of Altered Stress Hormones Following Acute Exercise

Endocrines ◽  
2021 ◽  
Vol 2 (2) ◽  
pp. 121-132
Author(s):  
Erik D. Hanson ◽  
Lauren C. Bates ◽  
Kaileigh Moertl ◽  
Elizabeth S. Evans
Keyword(s):  
Prostate Cancer ◽  
Immune System ◽  
Cancer Survivors ◽  
Nk Cells ◽  
Natural Killer ◽  
Acute Exercise ◽  
Nk Cell ◽  
Killer Cell ◽  
Current Evidence ◽  
Cell Mobilization

Natural killer (NK) cells from the innate immune system are integral to overall immunity and also in managing the tumor burden during cancer. Breast (BCa) and prostate cancer (PCa) are the most common tumors in U.S. adults. Both BCa and PCa are frequently treated with hormone suppression therapies that are associated with numerous adverse effects including direct effects on the immune system. Regular exercise is recommended for cancer survivors to reduce side effects and improve quality of life. Acute exercise is a potent stimulus for NK cells in healthy individuals with current evidence indicating that NK mobilization in individuals with BCa and PCa is comparable. NK cell mobilization results from elevations in shear stress and catecholamine levels. Despite a normal NK cell response to exercise, increases in epinephrine are attenuated in BCa and PCa. The significance of this potential discrepancy still needs to be determined. However, alterations in adrenal hormone signaling are hypothesized to be due to chronic stress during cancer treatment. Additional compensatory factors induced by exercise are reviewed along with recommendations on standardized approaches to be used in exercise immunology studies involving oncology populations.

scholarly journals Approaches to Enhance Natural Killer Cell-Based Immunotherapy for Pediatric Solid Tumors

Cancers ◽  
2021 ◽  
Vol 13 (11) ◽  
pp. 2796
Author(s):  
Aicha E. Quamine ◽  
Mallery R. Olsen ◽  
Monica M. Cho ◽  
Christian M. Capitini
Keyword(s):  
Nk Cells ◽  
Natural Killer ◽  
Solid Tumors ◽  
Nk Cell ◽  
Therapeutic Option ◽  
Treatment Options ◽  
Intensive Therapy ◽  
Killer Cell ◽  
Cell Therapies ◽  
Pediatric Solid Tumors

Treatment of metastatic pediatric solid tumors remain a significant challenge, particularly in relapsed and refractory settings. Standard treatment has included surgical resection, radiation, chemotherapy, and, in the case of neuroblastoma, immunotherapy. Despite such intensive therapy, cancer recurrence is common, and most tumors become refractory to prior therapy, leaving patients with few conventional treatment options. Natural killer (NK) cells are non-major histocompatibility complex (MHC)-restricted lymphocytes that boast several complex killing mechanisms but at an added advantage of not causing graft-versus-host disease, making use of allogeneic NK cells a potential therapeutic option. On top of their killing capacity, NK cells also produce several cytokines and growth factors that act as key regulators of the adaptive immune system, positioning themselves as ideal effector cells for stimulating heavily pretreated immune systems. Despite this promise, clinical efficacy of adoptive NK cell therapy to date has been inconsistent, prompting a detailed understanding of the biological pathways within NK cells that can be leveraged to develop “next generation” NK cell therapies. Here, we review advances in current approaches to optimizing the NK cell antitumor response including combination with other immunotherapies, cytokines, checkpoint inhibition, and engineering NK cells with chimeric antigen receptors (CARs) for the treatment of pediatric solid tumors.

scholarly journals Role and Modulation of NK Cells in Multiple Myeloma

Hemato ◽  
2021 ◽  
Vol 2 (2) ◽  
pp. 167-181
Author(s):  
Marie Thérèse Rubio ◽  
Adèle Dhuyser ◽  
Stéphanie Nguyen
Keyword(s):  
Multiple Myeloma ◽  
Monoclonal Antibodies ◽  
Nk Cells ◽  
Tumor Cells ◽  
Nk Cell ◽  
Ex Vivo ◽  
Killer Cell ◽  
Proteasome Inhibitors ◽  
Disease Evolution ◽  
Cell Functions

Myeloma tumor cells are particularly dependent on their microenvironment and sensitive to cellular antitumor immune response, including natural killer (NK) cells. These later are essential innate lymphocytes implicated in the control of viral infections and cancers. Their cytotoxic activity is regulated by a balance between activating and inhibitory signals resulting from the complex interaction of surface receptors and their respective ligands. Myeloma disease evolution is associated with a progressive alteration of NK cell number, phenotype and cytotoxic functions. We review here the different therapeutic approaches that could restore or enhance NK cell functions in multiple myeloma. First, conventional treatments (immunomodulatory drugs-IMids and proteasome inhibitors) can enhance NK killing of tumor cells by modulating the expression of NK receptors and their corresponding ligands on NK and myeloma cells, respectively. Because of their ability to kill by antibody-dependent cell cytotoxicity, NK cells are important effectors involved in the efficacy of anti-myeloma monoclonal antibodies targeting the tumor antigens CD38, CS1 or BCMA. These complementary mechanisms support the more recent therapeutic combination of IMids or proteasome inhibitors to monoclonal antibodies. We finally discuss the ongoing development of new NK cell-based immunotherapies, such as ex vivo expanded killer cell immunoglobulin-like receptors (KIR)-mismatched NK cells, chimeric antigen receptors (CAR)-NK cells, check point and KIR inhibitors.

Abstract WP319: miRNA Screening of Natural Killer Cell Identifies miR-1224 as a Post-Transcriptional Regulator of NK Cell Function After Ischemic Stroke

Stroke ◽  
2020 ◽  
Vol 51 (Suppl_1) ◽  
Author(s):  
Yan Feng ◽  
Hui Zhao ◽  
Fu-Dong Shi ◽  
Weina Jin
Keyword(s):  
Ischemic Stroke ◽  
Cerebral Ischemia ◽  
Nk Cells ◽  
Expression Profile ◽  
Mirna Expression ◽  
Cell Function ◽  
Nk Cell ◽  
Killer Cell ◽  
Mirna Expression Profile ◽  
Control Group

Objectives: To screen miRNA profile of peripheral NK cells in ischemic stroke mouse model and investigate a most promising candidate (miR-1224) for post-transcriptional regulation of NK cell function after ischemic stroke. Methods: Mice were subjected to a 60 min focal cerebral ischemia produced by transient intraluminal occlusion of MCAO. For NK cell isolation, cell suspensions from the spleens after reperfusion were enriched for NK cells using magnetic-bead sorting system after staining with anti-NK1.1 microbeads. The nCounter Mouse miRNA array was used to analyze miRNA expression profile in splenic NK cells over the time course of experimental ischemic stroke. Based on the miRNA data, we further in vitro modulated miR-1224 in NK cells using mimics or inhibitor, then injected i.v into Rag2-/-γc-/- recipient mice. Neurological function score was compared and spontaneous infection was assessed by pulmonary bacteria colony culture, and changes in potential signaling pathway (SP1/TNF-α) were verified by rt-PCR and western blot. Results: Through miRNA expression profile analysis, we have identified significant changes at each time point in peripheral NK cells after cerebral ischemia. Among all screened miRNA, miR-1224 remarkably increased in MCAO group, which was verified by PCR. Then isolated NK cells treated with mimics or inhibitors, were transferred to Rag2-/-γc-/- recipient mice. Compared with WT mice, Rag2-/-γc-/- mice with miR-1224 inhibitor exhibited increased NK cell number, enhanced NK cell activation/cytotoxicity feature, as well as better neurological behaviors and reduced pulmonary infection after MCAO. Moreover, compared with the control group, NK cells with miR-1224 inhibitor showed significantly increased SP1 gene and protein phosphorylation. As SP1 gene is one of the potential targets of miR-1224, this study suggests that miR-1224 may regulate NK cell function after MCAO, which is associated with SP1 pathway. Conclusion: The miRNA profiling of splenic NK cells provided insight into the functional mechanism and signaling pathways underlying the distinct organ-specific NK cell properties, which will contribute to the better understanding of NK cell mediated immune-response in relation to different stages of stroke.

scholarly journals The Potential for Cancer Immunotherapy in Targeting Surgery-Induced Natural Killer Cell Dysfunction

Cancers ◽  
2018 ◽  
Vol 11 (1) ◽  
pp. 2 ◽  
Author(s):  
Marisa Market ◽  
Katherine Baxter ◽  
Leonard Angka ◽  
Michael Kennedy ◽  
Rebecca Auer
Keyword(s):  
Nk Cells ◽  
Natural Killer ◽  
Cell Biology ◽  
Nk Cell ◽  
Clonal Selection ◽  
Killer Cell ◽  
Cancer Recurrence ◽  
Effector Functions ◽  
Cell Dysfunction ◽  
Cell Functions

Natural Killer (NK) cells are granular lymphocytes of the innate immune system that are able to recognize and kill tumor cells without undergoing clonal selection. Discovered over 40 years ago, they have since been recognized to possess both cytotoxic and cytokine-producing effector functions. Following trauma, NK cells are suppressed and their effector functions are impaired. This is especially important for cancer patients undergoing the removal of solid tumors, as surgery has shown to contribute to the development of metastasis and cancer recurrence postoperatively. We have recently shown that NK cells are critical mediators in the formation of metastasis after surgery. While research into the mechanism(s) responsible for NK cell dysfunction is ongoing, knowledge of these mechanisms will pave the way for perioperative therapeutics with the potential to improve cancer outcomes by reversing NK cell dysfunction. This review will discuss mechanisms of suppression in the postoperative environment, including hypercoagulability, suppressive soluble factors, the expansion of suppressive cell populations, and how this affects NK cell biology, including modulation of cell surface receptors, the potential for anergy, and immunosuppressive NK cell functions. This review will also outline potential immunotherapies to reverse postoperative NK dysfunction, with the goal of preventing surgery-induced metastasis.

scholarly journals Prognostic significance of natural killer cell-associated markers in gastric cancer: quantitative analysis using multiplex immunohistochemistry

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Hee Young Na ◽  
Yujun Park ◽  
Soo Kyung Nam ◽  
Jiwon Koh ◽  
Yoonjin Kwak ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
T Cells ◽  
B Cells ◽  
Nk Cells ◽  
Natural Killer ◽  
Immune Cells ◽  
Nk Cell ◽  
Critical Role ◽  
Killer Cell ◽  
Prognostic Significance

Abstract Background Natural killer (NK) cells mediate the anti-tumoral immune response as an important component of innate immunity. The aim of this study was to investigate the prognostic significance and functional implication of NK cell-associated surface receptors in gastric cancer (GC) by using multiplex immunohistochemistry (mIHC). Methods We performed an mIHC on tissue microarray slides, including 55 GC tissue samples. A total of 11 antibodies including CD57, NKG2A, CD16, HLA-E, CD3, CD20, CD45, CD68, CK, SMA, and ki-67 were used. CD45 + CD3-CD57 + cells were considered as CD57 + NK cells. Results Among CD45 + immune cells, the proportion of CD57 + NK cell was the lowest (3.8%), whereas that of CD57 + and CD57- T cells (65.5%) was the highest, followed by macrophages (25.4%), and B cells (5.3%). CD57 + NK cells constituted 20% of CD45 + CD57 + immune cells while the remaining 80% were CD57 + T cells. The expression of HLA-E in tumor cells correlated with that in tumoral T cells, B cells, and macrophages, but not CD57 + NK cells. The higher density of tumoral CD57 + NK cells and tumoral CD57 + NKG2A + NK cells was associated with inferior survival. Conclusions Although the number of CD57 + NK cells was lower than that of other immune cells, CD57 + NK cells and CD57 + NKG2A + NK cells were significantly associated with poor outcomes, suggesting that NK cell subsets play a critical role in GC progression. NK cells and their inhibitory receptor, NKG2A, may be potential targets in GC.

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