Publications by authors named "Heinrich Schlums"

26 Publications

  • Page 1 of 1

LIR-1 educates expanded human NK cells and defines a unique antitumor NK cell subset with potent antibody-dependent cellular cytotoxicity.

Clin Transl Immunology 2021 5;10(10):e1346. Epub 2021 Oct 5.

Department of Medicine Center for Hematology and Regenerative Medicine Karolinska Institutet Stockholm Sweden.

Objective: KIR and NKG2A receptors educate human NK cells to stay responsive to cells with diminished HLA class I. Here, we addressed whether the HLA class I-binding receptor LIR-1 (LILRB1/ILT2/CD85j), which is widely expressed on human NK cells, can mediate education and contribute to antitumor functions of NK cells.

Methods: Healthy donor NK cells either unstimulated, overnight cytokine-activated or -expanded were used to target human cell lines. Phenotype and function were analysed using flow cytometry and Cr-release assays.

Results: We found that the inhibitory receptor LIR-1 can mediate NK cell education under specific conditions. This novel finding was exclusive to expanded NK cells and further characterisation of the cells revealed high expression of granzyme B and DNAM-1, which both previously have been linked to NK cell education. Corroborating the rheostat education model, LIR-1 co-expression with an educating KIR further increased the responsiveness of expanded NK cells. Inversely, antibody masking of LIR-1 decreased the responsiveness. LIR-1 expanded NK cells displayed high intrinsic ADCC that, in contrast to KIR and NKG2A, was not inhibited by HLA class I.

Conclusion: These findings identify a unique NK cell subset attractive for adoptive cell therapy to treat cancer. Given that LIR-1 binds most HLA class I molecules, this subset may be explored in both autologous and allogeneic settings to innately reject HLA class I tumor cells as well as HLA class I target cells when combined with antitumor antibodies. Further studies are warranted to address the potential of this subset .
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http://dx.doi.org/10.1002/cti2.1346DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8491220PMC
October 2021

The transcription factor Bcl11b promotes both canonical and adaptive NK cell differentiation.

Sci Immunol 2021 03;6(57)

Broegelmann Research Laboratory, Department of Clinical Sciences, University of Bergen, N-5021 Bergen, Norway.

Epigenetic landscapes can provide insight into regulation of gene expression and cellular diversity. Here, we examined the transcriptional and epigenetic profiles of seven human blood natural killer (NK) cell populations, including adaptive NK cells. The gene, encoding a transcription factor (TF) essential for T cell development and function, was the most extensively regulated, with expression increasing throughout NK cell differentiation. Several Bcl11b-regulated genes associated with T cell signaling were specifically expressed in adaptive NK cell subsets. Regulatory networks revealed reciprocal regulation at distinct stages of NK cell differentiation, with Bcl11b repressing and in canonical and adaptive NK cells, respectively. A critical role for Bcl11b in driving NK cell differentiation was corroborated in -mutated patients and by ectopic Bcl11b expression. Moreover, was required for adaptive NK cell responses in a murine cytomegalovirus model, supporting expansion of these cells. Together, we define the TF regulatory circuitry of human NK cells and uncover a critical role for Bcl11b in promoting NK cell differentiation and function.
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http://dx.doi.org/10.1126/sciimmunol.abc9801DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8274449PMC
March 2021

Cytokines regulate the antigen-presenting characteristics of human circulating and tissue-resident intestinal ILCs.

Nat Commun 2020 04 27;11(1):2049. Epub 2020 Apr 27.

Center for Infectious Medicine, Department of Medicine, Karolinska Institutet, Karolinska University Hospital Huddinge, Stockholm, Sweden.

ILCs and T helper cells have been shown to exert bi-directional regulation in mice. However, how crosstalk between ILCs and CD4 T cells influences immune function in humans is unknown. Here we show that human intestinal ILCs co-localize with T cells in healthy and colorectal cancer tissue and display elevated HLA-DR expression in tumor and tumor-adjacent areas. Although mostly lacking co-stimulatory molecules ex vivo, intestinal and peripheral blood (PB) ILCs acquire antigen-presenting characteristics triggered by inflammasome-associated cytokines IL-1β and IL-18. IL-1β drives the expression of HLA-DR and co-stimulatory molecules on PB ILCs in an NF-κB-dependent manner, priming them as efficient inducers of cytomegalovirus-specific memory CD4 T-cell responses. This effect is strongly inhibited by the anti-inflammatory cytokine TGF-β. Our results suggest that circulating and tissue-resident ILCs have the intrinsic capacity to respond to the immediate cytokine milieu and regulate local CD4 T-cell responses, with potential implications for anti-tumor immunity and inflammation.
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http://dx.doi.org/10.1038/s41467-020-15695-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7184749PMC
April 2020

Diagnostic challenges for a novel SH2D1A mutation associated with X-linked lymphoproliferative disease.

Pediatr Blood Cancer 2020 04 29;67(4):e28184. Epub 2020 Jan 29.

Department of Medicine, Centre for Hematology and Regenerative Medicine, Karolinska Institutet, Karolinska University Hospital Huddinge, Stockholm, Sweden.

Mutations in SH2D1A, encoding the intracellular adaptor signaling lymphocyte activation molecule associated protein (SAP), are associated with X-linked lymphoproliferative disease type 1 (XLP1). We identified a novel hemizygous SH2D1A c.49G > A (p.E17K) variant in a 21-year-old patient with fatal Epstein-Barr virus infection-associated hemophagocytic lymphohistiocytosis. Cellular and biochemical assays revealed normal expression of the SAP variant protein, yet binding to phosphorylated CD244 receptor was reduced by >95%. Three healthy brothers carried the SH2D1A c.49G > A variant. Thus, data suggest that this variant represents a pathogenic mutation, but with variable expressivity. Importantly, our results highlight challenges in the clinical interpretation of SH2D1A variants and caution in using functional flow cytometry assays for the diagnosis of XLP1.
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http://dx.doi.org/10.1002/pbc.28184DOI Listing
April 2020

Dynamic Changes in Natural Killer Cell Subset Frequencies in the Absence of Cytomegalovirus Infection.

Front Immunol 2019 22;10:2728. Epub 2019 Nov 22.

Center for Autoimmune Genomics and Etiology, Cincinnati Children's Hospital Medical Center, Cincinnati, OH, United States.

Individuals lacking functional natural killer (NK) cells suffer severe, recurrent infections with cytomegalovirus (CMV), highlighting the critical role of NK cells in antiviral defense. Therefore, ongoing attempts to develop an efficacious vaccine to prevent CMV infection should potentially aim to elicit NK-cell antiviral responses as an accessory to conventional T- and B-cell based approaches. In this regard, CMV infection provokes marked phenotypic and functional differentiation of the NK-cell compartment, including development of adaptive NK cells that exhibit enhanced antiviral activity. We examined longitudinal blood samples collected from 40 CMV-seronegative adolescents to ascertain whether a CMV glycoprotein B (gB) vaccine in the absence of CMV infection can stimulate differentiation or expansion of CMV-associated subsets of NK cells. Study participants uniformly lacked the CMV-dependent NKG2C subset of NK cells, suggesting that an adjuvanted CMV gB vaccine alone is an inadequate stimulus for sustained expansion of these cells. In contrast, we observed unexpected dynamic fluctuations in the frequency of NK cells lacking FcRγ, EAT-2, and SYK, which were independent of vaccination or CMV infection. Whereas, FcRγ NK cells in CMV infection are reported to express increased levels of the maturation marker CD57, the FcRγ NK cells observed in our CMV-negative vaccine cohort express less CD57 than their FcRγ counterparts. The FcRγ NK cells in CMV-negative individuals were also functionally distinct from this subset in CMV infection, exhibiting comparable IFN-γ production and degranulation as FcRγ NK cells in response to cytokine or antibody-dependent stimuli. These results suggest that frequencies of some NK cell subsets may increase in response to unknown environmental or inflammatory cues distinct from that which occurs after CMV infection. Greater understanding of the nature of the signals driving CMV-independent accumulation of these subsets should permit development of mechanisms to facilitate vaccine-driven expansion of CMV-reactive NK cells.
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http://dx.doi.org/10.3389/fimmu.2019.02728DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6882915PMC
November 2020

Adaptive NK cells in people exposed to correlate with protection from malaria.

J Exp Med 2019 06 12;216(6):1280-1290. Epub 2019 Apr 12.

Laboratory of Immunogenetics, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Rockville, MD

How antibodies naturally acquired during infection provide clinical immunity to blood-stage malaria is unclear. We studied the function of natural killer (NK) cells in people living in a malaria-endemic region of Mali. Multi-parameter flow cytometry revealed a high proportion of adaptive NK cells, which are defined by the loss of transcription factor PLZF and Fc receptor γ-chain. Adaptive NK cells dominated antibody-dependent cellular cytotoxicity responses, and their frequency within total NK cells correlated with lower parasitemia and resistance to malaria. -infected RBCs induced NK cell degranulation after addition of plasma from malaria-resistant individuals. Malaria-susceptible subjects with the largest increase in PLZF-negative NK cells during the transmission season had improved odds of resistance during the subsequent season. Thus, antibody-dependent lysis of -infected RBCs by NK cells may be a mechanism of acquired immunity to malaria. Consideration of antibody-dependent NK cell responses to antigens is therefore warranted in the design of malaria vaccines.
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http://dx.doi.org/10.1084/jem.20181681DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6547858PMC
June 2019

Clonal expansion and compartmentalized maintenance of rhesus macaque NK cell subsets.

Sci Immunol 2018 11;3(29)

Division of Intramural Research, National Heart, Lung, and Blood Institute, NIH, Bethesda, MD, USA.

Natural killer (NK) cells recognize and eliminate infected and malignant cells. Their life histories are poorly understood, particularly in humans, due to lack of informative models and endogenous clonal markers. Here, we apply transplantation of barcoded rhesus macaque hematopoietic cells to interrogate the landscape of NK cell production, expansion, and life histories at a clonal level long term and after proliferative challenge. We identify oligoclonal populations of rhesus CD56CD16 NK cells that are characterized by marked expansions and contractions over time yet remained long-term clonally uncoupled from other hematopoietic lineages, including CD56CD16 NK cells. Individual or groups of CD56CD16 expanded clones segregated with surface expression of specific killer immunoglobulin-like receptors. These clonally distinct NK cell subpopulation patterns persisted for more than 4 years, including after transient in vivo anti-CD16-mediated depletion and subsequent regeneration. Profound and sustained interleukin-15-mediated depletion was required to generate new oligoclonal CD56CD16 NK cells. Together, our results indicate that linear NK cell production from multipotent hematopoietic progenitors or less mature CD56CD16 cells is negligible during homeostasis and moderate proliferative stress. In such settings, peripheral compartmentalized self-renewal can maintain the composition of distinct, differentiated NK cell subpopulations.
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http://dx.doi.org/10.1126/sciimmunol.aat9781DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7393805PMC
November 2018

High mTOR activity is a hallmark of reactive natural killer cells and amplifies early signaling through activating receptors.

Elife 2017 09 6;6. Epub 2017 Sep 6.

CIRI, Centre International de Recherche en Infectiologie - International Center for Infectiology Research, Lyon, France.

NK cell education is the process through which chronic engagement of inhibitory NK cell receptors by self MHC-I molecules preserves cellular responsiveness. The molecular mechanisms responsible for NK cell education remain unclear. Here, we show that mouse NK cell education is associated with a higher basal activity of the mTOR/Akt pathway, commensurate to the number of educating receptors. This higher activity was dependent on the SHP-1 phosphatase and essential for the improved responsiveness of reactive NK cells. Upon stimulation, the mTOR/Akt pathway amplified signaling through activating NK cell receptors by enhancing calcium flux and LFA-1 integrin activation. Pharmacological inhibition of mTOR resulted in a proportional decrease in NK cell reactivity. Reciprocally, acute cytokine stimulation restored reactivity of hyporesponsive NK cells through mTOR activation. These results demonstrate that mTOR acts as a molecular rheostat of NK cell reactivity controlled by educating receptors and uncover how cytokine stimulation overcomes NK cell education.
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http://dx.doi.org/10.7554/eLife.26423DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5628014PMC
September 2017

GSK3 Inhibition Drives Maturation of NK Cells and Enhances Their Antitumor Activity.

Cancer Res 2017 10 8;77(20):5664-5675. Epub 2017 Aug 8.

Division of Hematology, Oncology and Transplantation, Department of Medicine, University of Minnesota Cancer Center, Minneapolis, Minnesota.

Maturation of human natural killer (NK) cells as defined by accumulation of cell-surface expression of CD57 is associated with increased cytotoxic character and TNF and IFNγ production upon target-cell recognition. Notably, multiple studies point to a unique role for CD57 NK cells in cancer immunosurveillance, yet there is scant information about how they mature. In this study, we show that pharmacologic inhibition of GSK3 kinase in peripheral blood NK cells expanded with IL15 greatly enhances CD57 upregulation and late-stage maturation. GSK3 inhibition elevated the expression of several transcription factors associated with late-stage NK-cell maturation including T-BET, ZEB2, and BLIMP-1 without affecting viability or proliferation. When exposed to human cancer cells, NK cell expanded in the presence of a GSK3 inhibitor exhibited significantly higher production of TNF and IFNγ, elevated natural cytotoxicity, and increased antibody-dependent cellular cytotoxicity. In an established mouse xenograft model of ovarian cancer, adoptive transfer of NK cells conditioned in the same way also displayed more robust and durable tumor control. Our findings show how GSK3 kinase inhibition can greatly enhance the mature character of NK cells most desired for effective cancer immunotherapy. .
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http://dx.doi.org/10.1158/0008-5472.CAN-17-0799DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5645243PMC
October 2017

Unperturbed Cytotoxic Lymphocyte Phenotype and Function in Myalgic Encephalomyelitis/Chronic Fatigue Syndrome Patients.

Front Immunol 2017 26;8:723. Epub 2017 Jun 26.

Department of Medicine, Huddinge, Karolinska Institutet, Stockholm, Sweden.

Myalgic encephalomyelitis or chronic fatigue syndrome (ME/CFS) is a debilitating disorder linked to diverse intracellular infections as well as physiological stress. Cytotoxic lymphocytes combat intracellular infections. Their function is attenuated by stress. Despite numerous studies, the role of cytotoxic lymphocytes in ME/CFS remains unclear. Prompted by advances in the understanding of defects in lymphocyte cytotoxicity, the discovery of adaptive natural killer (NK) cell subsets associated with certain viral infections, and compelling links between stress, adrenaline, and cytotoxic lymphocyte function, we reassessed the role of cytotoxic lymphocytes in ME/CFS. Forty-eight patients from two independent cohorts fulfilling the Canada 2003 criteria for ME/CFS were evaluated with respect to cytotoxic lymphocyte phenotype and function. Results were compared to values from matched healthy controls. Reproducible differences between patients and controls were not found in cytotoxic lymphocyte numbers, cytotoxic granule content, activation status, exocytotic capacity, target cell killing, or cytokine production. One patient expressed low levels of perforin, explained by homozygosity for the p.A91V variant. However, overall, this variant was present in a heterozygous state at the expected population frequency among ME/CFS patients. No single patient displayed any pathological patterns of cellular responses. Increased expansions of adaptive NK cells or deviant cytotoxic lymphocyte adrenaline-mediated inhibition were not observed. In addition, supervised dimensionality reduction analyses of the full, multidimensional datasets did not reveal any reproducible patient/control discriminators. In summary, employing sensitive assays and analyses for quantification of cytotoxic lymphocyte differentiation and function, cytotoxicity lymphocyte aberrances were not found among ME/CFS patients. These assessments of cytotoxic lymphocytes therefore do not provide useful biomarkers for the diagnosis of ME/CFS.
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http://dx.doi.org/10.3389/fimmu.2017.00723DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5483846PMC
June 2017

CD49a Expression Defines Tissue-Resident CD8 T Cells Poised for Cytotoxic Function in Human Skin.

Immunity 2017 02 14;46(2):287-300. Epub 2017 Feb 14.

Department of Medicine Solna, Karolinska Institutet, Stockholm 171 77, Sweden; Dermatology Department, Karolinska University Hospital, Stockholm 141 86, Sweden. Electronic address:

Tissue-resident memory T (Trm) cells form a heterogeneous population that provides localized protection against pathogens. Here, we identify CD49a as a marker that differentiates CD8 Trm cells on a compartmental and functional basis. In human skin epithelia, CD8CD49a Trm cells produced interferon-γ, whereas CD8CD49a Trm cells produced interleukin-17 (IL-17). In addition, CD8CD49a Trm cells from healthy skin rapidly induced the expression of the effector molecules perforin and granzyme B when stimulated with IL-15, thereby promoting a strong cytotoxic response. In skin from patients with vitiligo, where melanocytes are eradicated locally, CD8CD49a Trm cells that constitutively expressed perforin and granzyme B accumulated both in the epidermis and dermis. Conversely, CD8CD49a Trm cells from psoriasis lesions predominantly generated IL-17 responses that promote local inflammation in this skin disease. Overall, CD49a expression delineates CD8 Trm cell specialization in human epithelial barriers and correlates with the effector cell balance found in distinct inflammatory skin diseases.
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http://dx.doi.org/10.1016/j.immuni.2017.01.009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5337619PMC
February 2017

Adaptive NK cells can persist in patients with mutation depleted of stem and progenitor cells.

Blood 2017 04 16;129(14):1927-1939. Epub 2017 Feb 16.

Center for Hematology and Regenerative Medicine, Department of Medicine, Karolinska Institutet, Karolinska University Hospital Huddinge, Stockholm, Sweden.

Heterozygous mutation is associated with immunodeficiency, lymphedema, and myelodysplastic syndrome. Disease presentation is variable, often coinciding with loss of circulating dendritic cells, monocytes, B cells, and natural killer (NK) cells. Nonetheless, in a proportion of patients carrying mutation, NK cells persist. We found that peripheral blood NK cells in symptomatic patients uniformly lacked expression of the transcription factor promyelocytic leukemia zinc finger (PLZF), as well as expression of intracellular signaling proteins FcεRγ, spleen tyrosine kinase (SYK), and EWS/FLI1-Activated Transcript 2 (EAT-2) in a variegated manner. Moreover, consistent with an adaptive identity, NK cells from patients with mutation displayed altered expression of cytotoxic granule constituents and produced interferon-γ upon Fc-receptor engagement but not following combined interleukin-12 (IL-12) and IL-18 stimulation. Canonical, PLZF-expressing NK cells were retained in asymptomatic carriers of mutation. Developmentally, GATA-binding protein-2 (GATA-2) was expressed in hematopoietic stem cells, but not in NK-cell progenitors, CD3CD56, canonical, or adaptive CD3CD56 NK cells. Peripheral blood NK cells from individuals with mutation proliferated normally in vitro, whereas lineage-negative progenitors displayed impaired NK-cell differentiation. In summary, adaptive NK cells can persist in patients with mutation, even after NK-cell progenitors expire. Moreover, our data suggest that adaptive NK cells are more long-lived than canonical, immunoregulatory NK cells.
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http://dx.doi.org/10.1182/blood-2016-08-734236DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5383869PMC
April 2017

Acquired somatic mutations in PNH reveal long-term maintenance of adaptive NK cells independent of HSPCs.

Blood 2017 04 30;129(14):1940-1946. Epub 2016 Nov 30.

Hematology Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD.

Natural killer (NK) cells have long been considered short-lived effectors of innate immunity. However, recent animal models and human studies suggest that subsets of NK cells have adaptive features. We investigate clonal relationships of various NK-cell subsets, including the adaptive population, by taking advantage of naturally occurring X-linked somatic mutations in hematopoietic stem and progenitor cells (HSPCs) from patients with paroxysmal nocturnal hemoglobinuria (PNH). The affected HSPCs and their progeny lack expression of glycosylphosphatidylinositol (GPI) anchors on their cell surface, allowing quantification of -mutant (GPI-negative) HSPC-derived peripheral blood cell populations. The fraction of GPI-negative cells within the CD56 NK cells was markedly lower than that of neutrophils and the CD56 NK-cell compartments. This discrepancy was most prominent within the adaptive CD56 NK-cell population lacking PLZF expression. The functional properties of these adaptive NK cells were similar in PNH patients and healthy individuals. Our findings support the existence of a long-lived, adaptive NK-cell population maintained independently from GPICD56.
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http://dx.doi.org/10.1182/blood-2016-08-734285DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5383870PMC
April 2017

Epigenetic Regulation of Adaptive NK Cell Diversification.

Trends Immunol 2016 07 6;37(7):451-461. Epub 2016 May 6.

Centre for Hematology and Regenerative Medicine, Department of Medicine, Karolinska Institutet, Karolinska University Hospital Huddinge, Stockholm, Sweden; Broegelmann Research Laboratory, Department of Clinical Sciences, University of Bergen, Bergen, Norway. Electronic address:

Natural killer (NK) cells were previously considered to represent short-lived, innate lymphocytes. However, mouse models have revealed expansion and persistence of differentiated NK cell subsets in response to cytomegalovirus (CMV) infection, paralleling antigen-specific T cell differentiation. Congruently, analyses of humans have uncovered CMV-associated NK cell subsets characterized by epigenetic diversification processes that lead to altered target cell specificities and functional capacities. Here, focusing on responses to viruses, we review similarities and differences between mouse and human adaptive NK cells, identifying molecular analogies that may be key to transcriptional reprogramming and functional alterations. We discuss possible molecular mechanisms underlying epigenetic diversification and hypothesize that processes driving epigenetic diversification may represent a more widespread mechanism for fine-tuning and optimization of cellular immunity.
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http://dx.doi.org/10.1016/j.it.2016.04.006DOI Listing
July 2016

Diversification and Functional Specialization of Human NK Cell Subsets.

Curr Top Microbiol Immunol 2016 ;395:63-94

Natural killer (NK) cells are lymphocytes that participate in different facets of immunity. They can act as innate sentinels through recognition and eradication of infected or transformed target cells, so-called immunosurveillance. In addition, they can contain immune responses through the killing of other activated immune cells, so-called immunoregulation. Furthermore, they instruct and regulate immune responses by producing pro-inflammatory cytokines such as IFN-γ, either upon direct target cell recognition or by relaying cytokine cues from various cell types. Recent studies in mouse and man have uncovered infection-associated expansions of NK cell subsets with specific receptor repertoires and diverse patterns of intracellular signaling molecule expression. Moreover, distinct attributes of NK cells in tissues, including tissue-resident subsets, are being further elucidated. Findings support an emerging theme of ever-increasing diversification and functional specialization among different NK cell subsets, with a functional dichotomy between subsets involved in immunoregulation or immunosurveillance. The epigenetic landscapes and transcriptional profiles of different NK cell subsets are providing insights into the molecular regulation of effector functions. Here, we review phenotypic, functional, and developmental characteristics of a spectrum of human NK cell subsets. We also discuss the molecular underpinnings of different NK cell subsets and their potential contributions to immunity as well as disease susceptibility.
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http://dx.doi.org/10.1007/82_2015_487DOI Listing
June 2016

The Past, Present, and Future of NK Cells in Hematopoietic Cell Transplantation and Adoptive Transfer.

Curr Top Microbiol Immunol 2016 ;395:225-43

Department of Medicine, University of Minnesota, Minneapolis, MN, USA.

Hematopoietic cell transplantation (HCT) has been used as a part of cancer therapy for over half a decade. Beyond the necessity for donor-derived cells to reconstitute hematopoiesis after radiation and chemotherapy, immunologic reconstitution from allogeneic cells is important for the elimination of residual tumor cells. Natural killer (NK) cells are first among lymphocytes to reconstitute post-transplant and protect against cancer relapse. In this review, we provide a historical perspective on the role of NK cells in cancer control in the transplant setting and focus on current research aimed at improving NK cell responses for therapeutic benefit.
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http://dx.doi.org/10.1007/82_2015_445DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5870762PMC
June 2016

Cytomegalovirus infection drives adaptive epigenetic diversification of NK cells with altered signaling and effector function.

Immunity 2015 Mar;42(3):443-56

Centre for Infectious Medicine, Department of Medicine, Karolinska Institutet, Karolinska University Hospital Huddinge, 14186 Stockholm, Sweden; Broegelmann Research Laboratory, Department of Clinical Sciences, University of Bergen, 5021 Bergen, Norway. Electronic address:

The mechanisms underlying human natural killer (NK) cell phenotypic and functional heterogeneity are unknown. Here, we describe the emergence of diverse subsets of human NK cells selectively lacking expression of signaling proteins after human cytomegalovirus (HCMV) infection. The absence of B and myeloid cell-related signaling protein expression in these NK cell subsets correlated with promoter DNA hypermethylation. Genome-wide DNA methylation patterns were strikingly similar between HCMV-associated adaptive NK cells and cytotoxic effector T cells but differed from those of canonical NK cells. Functional interrogation demonstrated altered cytokine responsiveness in adaptive NK cells that was linked to reduced expression of the transcription factor PLZF. Furthermore, subsets of adaptive NK cells demonstrated significantly reduced functional responses to activated autologous T cells. The present results uncover a spectrum of epigenetically unique adaptive NK cell subsets that diversify in response to viral infection and have distinct functional capabilities compared to canonical NK cell subsets.
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http://dx.doi.org/10.1016/j.immuni.2015.02.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4612277PMC
March 2015

Hemophagocytic lymphohistiocytosis in 2 patients with underlying IFN-γ receptor deficiency.

J Allergy Clin Immunol 2015 Jun 13;135(6):1638-41. Epub 2015 Jan 13.

Centre for Infectious Medicine, Department of Medicine, Karolinska Institute, Karolinska University Hospital Huddinge, Stockholm, Sweden; Broegelmann Research Laboratory, Department of Clinical Sciences, University of Bergen, Norway. Electronic address:

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http://dx.doi.org/10.1016/j.jaci.2014.11.030DOI Listing
June 2015

Transcriptional regulation of Munc13-4 expression in cytotoxic lymphocytes is disrupted by an intronic mutation associated with a primary immunodeficiency.

J Exp Med 2014 Jun 19;211(6):1079-91. Epub 2014 May 19.

Centre for Infectious Medicine, Department of Medicine; Clinical Genetics Unit, Department of Molecular Medicine and Surgery, and Center for Molecular Medicine, Karolinska Institutet, Karolinska University Hospital Huddinge, 141 86 Stockholm, Sweden Broegelmann Research Laboratory, Clinical Institute, University of Bergen, N-5021 Bergen, Norway

Autosomal recessive mutations in UNC13D, the gene that encodes Munc13-4, are associated with familial hemophagocytic lymphohistiocytosis type 3 (FHL3). Munc13-4 expression is obligatory for exocytosis of lytic granules, facilitating cytotoxicity by T cells and natural killer (NK) cells. The mechanisms regulating Munc13-4 expression are unknown. Here, we report that Munc13-4 is highly expressed in differentiated human NK cells and effector CD8(+) T lymphocytes. A UNC13D c.118-308C>T mutation, causative of FHL3, disrupted binding of the ETS family member ELF1 to a conserved intronic sequence. This mutation impairs UNC13D intron 1 recruitment of STAT4 and the chromatin remodeling complex component BRG1, diminishing active histone modifications at the locus. The intronic sequence acted as an overall enhancer of Munc13-4 expression in cytotoxic lymphocytes in addition to representing an alternative promoter encoding a novel Munc13-4 isoform. Mechanistically, T cell receptor engagement facilitated STAT4-dependent Munc13-4 expression in naive CD8(+) T lymphocytes. Collectively, our data demonstrates how chromatin remodeling within an evolutionarily conserved regulatory element in intron 1 of UNC13D regulates the induction of Munc13-4 expression in cytotoxic lymphocytes and suggests that an alternative Munc13-4 isoform is required for lymphocyte cytotoxicity. Thus, mutations associated with primary immunodeficiencies may cause disease by disrupting transcription factor binding.
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http://dx.doi.org/10.1084/jem.20131131DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4042637PMC
June 2014

Systemic lupus erythematosus immune complexes increase the expression of SLAM family members CD319 (CRACC) and CD229 (LY-9) on plasmacytoid dendritic cells and CD319 on CD56(dim) NK cells.

J Immunol 2013 Sep 16;191(6):2989-98. Epub 2013 Aug 16.

Section of Rheumatology, Department of Medical Sciences, Uppsala University, S-751 85 Uppsala, Sweden.

Patients with systemic lupus erythematosus (SLE) display an activated type I IFN system due to unceasing IFN-α release from plasmacytoid dendritic cells (pDCs) stimulated by nucleic acid-containing immune complexes (ICs). NK cells strongly promote the IFN-α production by pDCs; therefore, we investigated surface molecules that could be involved in the pDC-NK cell cross-talk. In human PBMCs stimulated with RNA-containing ICs (RNA-ICs), the expression of the signaling lymphocyte activation molecule (SLAM) family receptors CD319 and CD229 on pDCs and CD319 on CD56(dim) NK cells was selectively increased. Upregulation of CD319 and CD229 on RNA-IC-stimulated pDCs was induced by NK cells or cytokines (e.g., GM-CSF, IL-3). IFN-α-producing pDCs displayed a higher expression of SLAM molecules compared with IFN-α⁻ pDCs. With regard to signaling downstream of SLAM receptors, pDCs expressed SHIP-1, SHP-1, SHP-2, and CSK but lacked SLAM-associated protein (SAP) and Ewing's sarcoma-activated transcript 2 (EAT2), indicating that these receptors may act as inhibitory receptors on pDCs. Furthermore, pDCs from patients with SLE had decreased expression of CD319 on pDCs and CD229 on CD56(dim) NK cells, but RNA-IC stimulation increased CD319 and CD229 expression. In conclusion, this study reveals that the expression of the SLAM receptors CD319 and CD229 is regulated on pDCs and NK cells by lupus ICs and that the expression of these receptors is specifically altered in SLE. These results, together with the observed genetic association between the SLAM locus and SLE, suggest a role for CD319 and CD229 in the SLE disease process.
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http://dx.doi.org/10.4049/jimmunol.1301022DOI Listing
September 2013

Familial hemophagocytic lymphohistiocytosis type 3 (FHL3) caused by deep intronic mutation and inversion in UNC13D.

Blood 2011 Nov 19;118(22):5783-93. Epub 2011 Sep 19.

Childhood Cancer Research Unit, Department of Women's and Children's Health, Karolinska Institutet, Karolinska University Hospital Solna, Stockholm, Sweden.

Familial hemophagocytic lymphohistiocytosis (FHL) is an autosomal recessive, often-fatal hyperinflammatory disorder. Mutations in PRF1, UNC13D, STX11, and STXBP2 are causative of FHL2, 3, 4, and 5, respectively. In a majority of suspected FHL patients from Northern Europe, sequencing of exons and splice sites of such genes required for lymphocyte cytotoxicity revealed no or only monoallelic UNC13D mutations. Here, in 21 patients, we describe 2 pathogenic, noncoding aberrations of UNC13D. The first is a point mutation localized in an evolutionarily conserved region of intron 1. This mutation selectively impairs UNC13D transcription in lymphocytes, abolishing Munc13-4 expression. The second is a 253-kb inversion straddling UNC13D, affecting the 3'-end of the transcript and likewise abolishing Munc13-4 expression. Carriership of the intron 1 mutation was found in patients across Europe, whereas carriership of the inversion was limited to Northern Europe. Notably, the latter aberration represents the first description of an autosomal recessive human disease caused by an inversion. These findings implicate an intronic sequence in cell-type specific expression of Munc13-4 and signify variations outside exons and splice sites as a common cause of FHL3. Based on these data, we propose a strategy for targeted sequencing of evolutionary conserved noncoding regions for the diagnosis of primary immunodeficiencies.
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http://dx.doi.org/10.1182/blood-2011-07-369090DOI Listing
November 2011

Molecular mechanisms of natural killer cell activation.

J Innate Immun 2011 29;3(3):216-26. Epub 2011 Mar 29.

Center for Infectious Medicine, Department of Medicine, Karolinska Institutet, Karolinska University Hospital Huddinge, Stockholm, Sweden.

With an array of activating and inhibitory receptors, natural killer (NK) cells can specifically eradicate infected and transformed cells. Target cell killing is achieved through directed release of lytic granules. Recognition of target cells also induces production of chemokines and cytokines that can coordinate immune responses. Upon contact with susceptible cells, a multiplicity of activating receptors can induce signals for adhesion. Engagement of the integrin leukocyte functional antigen-1 mediates firm adhesion, provides signals for granule polarization and orchestrates the structure of an immunological synapse that facilitates efficient target cell killing. Other activating receptors apart from leukocyte functional antigen-1 signal for lytic granule exocytosis, a process that requires overcoming a threshold for activation of phospholipase C-γ, which in turn induces STIM1- and ORAI1-dependent store-operated Ca²+ entry as well as exocytosis mediated by the SNARE-containing protein syntaxin-11 and regulators thereof. Cytokine and chemokine release follows a different secretory pathway which also requires phospholipase C-γ activation and store-operated Ca²+ entry. Recent studies of human NK cells have provided insights into a hierarchy of effector functions that result in graded responses by NK cell populations. Responses display cellular heterogeneity and are influenced by environmental cues. This review highlights recent knowledge gained on the molecular pathways for and regulation of NK cell activation.
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http://dx.doi.org/10.1159/000325265DOI Listing
August 2011

Sensitive and viable quantification of inside-out signals for LFA-1 activation in human cytotoxic lymphocytes by flow cytometry.

J Immunol Methods 2011 Mar 2;366(1-2):106-18. Epub 2011 Feb 2.

Center for Infectious Medicine, Department of Medicine, Karolinska Institute, Karolinska University Hospital Huddinge, Stockholm, Sweden.

An early step in immunosurveillance by cytotoxic lymphocytes is leukocyte functional antigen (LFA)-1-dependent adhesion to target cells, which is promoted by inside-out signals from receptors such as the T cell receptor and a variety of natural killer (NK) cell activating receptors. Inside-out signals induce a conformational change in LFA-1, resulting in an extension of the extracellular domain of the receptor. Here, we have evaluated several mAbs that specifically detect the extended conformation of LFA-1 and detail a protocol for flow cytometric quantification of β2-integrin activation in human peripheral blood cytotoxic T cells and NK cells in response to target cell recognition. By comparison to the markers of degranulation and chemokine synthesis, e.g. surface CD107a expression and intracellular MIP-1β expression, respectively, evaluation of LFA-1 conformational changes represent a sensitive and rapid parameter of primary cytotoxic lymphocyte activation that can facilitate isolation of viable cells. Potentially, combined with other read-outs of cytotoxic lymphocyte function, this technique is applicable to the assessment of various clinical conditions, including for the diagnosis of primary immunodeficiency syndromes and for evaluating the efficacy of tumor targeting by donor lymphocytes.
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http://dx.doi.org/10.1016/j.jim.2011.01.014DOI Listing
March 2011
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