Publications by authors named "Hans-Gustaf Ljunggren"

176 Publications

SARS-CoV-2-specific humoral and cellular immunity persists through 9 months irrespective of COVID-19 severity at hospitalisation.

Clin Transl Immunology 2021 5;10(7):e1306. Epub 2021 Jul 5.

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

Objectives: Humoral and cellular immunity to SARS-CoV-2 following COVID-19 will likely contribute to protection from reinfection or severe disease. It is therefore important to characterise the initiation and persistence of adaptive immunity to SARS-CoV-2 amidst the ongoing pandemic.

Methods: Here, we conducted a longitudinal study on hospitalised moderate and severe COVID-19 patients from the acute phase of disease into convalescence at 5 and 9 months post-symptom onset. Utilising flow cytometry, serological assays as well as B cell and T cell FluoroSpot assays, we assessed the magnitude and specificity of humoral and cellular immune responses during and after human SARS-CoV-2 infection.

Results: During acute COVID-19, we observed an increase in germinal centre activity, a substantial expansion of antibody-secreting cells and the generation of SARS-CoV-2-neutralising antibodies. Despite gradually decreasing antibody levels, we show persistent, neutralising antibody titres as well as robust specific memory B cell responses and polyfunctional T cell responses at 5 and 9 months after symptom onset in both moderate and severe COVID-19 patients.

Conclusion: Our findings describe the initiation and, importantly, persistence of cellular and humoral SARS-CoV-2-specific immunological memory in hospitalised COVID-19 patients long after recovery, likely contributing towards protection against reinfection.
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http://dx.doi.org/10.1002/cti2.1306DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8256672PMC
July 2021

Natural killer cells in antiviral immunity.

Nat Rev Immunol 2021 Jun 11. Epub 2021 Jun 11.

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

Natural killer (NK) cells play an important role in innate immune responses to viral infections. Here, we review recent insights into the role of NK cells in viral infections, with particular emphasis on human studies. We first discuss NK cells in the context of acute viral infections, with flavivirus and influenza virus infections as examples. Questions related to activation of NK cells, homing to infected tissues and the role of tissue-resident NK cells in acute viral infections are also addressed. Next, we discuss NK cells in the context of chronic viral infections with hepatitis C virus and HIV-1. Also covered is the role of adaptive-like NK cell expansions as well as the appearance of CD56 NK cells in the course of chronic infection. Specific emphasis is then placed in viral infections in patients with primary immunodeficiencies affecting NK cells. Not least, studies in this area have revealed an important role for NK cells in controlling several herpesvirus infections. Finally, we address new data with respect to the activation of NK cells and NK cell function in humans infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) giving rise to coronavirus disease 2019 (COVID-19).
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http://dx.doi.org/10.1038/s41577-021-00558-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8194386PMC
June 2021

Expansions of adaptive-like NK cells with a tissue-resident phenotype in human lung and blood.

Proc Natl Acad Sci U S A 2021 Mar;118(11)

Center for Infectious Medicine, Department of Medicine Huddinge, Karolinska Institutet, 14152 Stockholm, Sweden.

Human adaptive-like "memory" CD56CD16 natural killer (NK) cells in peripheral blood from cytomegalovirus-seropositive individuals have been extensively investigated in recent years and are currently explored as a treatment strategy for hematological cancers. However, treatment of solid tumors remains limited due to insufficient NK cell tumor infiltration, and it is unknown whether large expansions of adaptive-like NK cells that are equipped for tissue residency and tumor homing exist in peripheral tissues. Here, we show that human lung and blood contains adaptive-like CD56CD16 NK cells with hallmarks of tissue residency, including expression of CD49a. Expansions of adaptive-like lung tissue-resident NK (trNK) cells were found to be present independently of adaptive-like CD56CD16 NK cells and to be hyperresponsive toward target cells. Together, our data demonstrate that phenotypically, functionally, and developmentally distinct subsets of adaptive-like NK cells exist in human lung and blood. Given their tissue-related character and hyperresponsiveness, human lung adaptive-like trNK cells might represent a suitable alternative for therapies targeting solid tumors.
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http://dx.doi.org/10.1073/pnas.2016580118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7980282PMC
March 2021

Activated Natural Killer Cells Hit Neurogenesis in the Aging Brain.

Neurosci Bull 2021 Jul 29;37(7):1072-1074. Epub 2021 Mar 29.

Center of Infectious Medicine, Department of Medicine Huddinge, Karolinska Institutet, 141 52, Stockholm, Sweden.

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http://dx.doi.org/10.1007/s12264-021-00654-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8006623PMC
July 2021

MAIT cell activation is associated with disease severity markers in acute hantavirus infection.

Cell Rep Med 2021 Mar 16;2(3):100220. Epub 2021 Mar 16.

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

Hantaviruses are zoonotic RNA viruses that cause severe acute disease in humans. Infected individuals have strong inflammatory responses that likely cause immunopathology. Here, we studied the response of mucosal-associated invariant T (MAIT) cells in peripheral blood of individuals with hemorrhagic fever with renal syndrome (HFRS) caused by Puumala orthohantavirus, a hantavirus endemic in Europe. We show that MAIT cell levels decrease in the blood during HFRS and that residual MAIT cells are highly activated. This activation correlates with HFRS severity markers. activation of MAIT cells by hantavirus-exposed antigen-presenting cells is dependent on type I interferons (IFNs) and independent of interleukin-18 (IL-18). These findings highlight the role of type I IFNs in virus-driven MAIT cell activation and suggest a potential role of MAIT cells in the disease pathogenesis of viral infections.
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http://dx.doi.org/10.1016/j.xcrm.2021.100220DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7974553PMC
March 2021

Major alterations in the mononuclear phagocyte landscape associated with COVID-19 severity.

Proc Natl Acad Sci U S A 2021 02;118(6)

Center for Infectious Medicine, Department of Medicine Huddinge, Karolinska Institutet, Karolinska University Hospital, 171 77 Stockholm, Sweden.

Dendritic cells (DCs) and monocytes are crucial mediators of innate and adaptive immune responses during viral infection, but misdirected responses by these cells may contribute to immunopathology. Here, we performed high-dimensional flow cytometry-analysis focusing on mononuclear phagocyte (MNP) lineages in SARS-CoV-2-infected patients with moderate and severe COVID-19. We provide a deep and comprehensive map of the MNP landscape in COVID-19. A redistribution of monocyte subsets toward intermediate monocytes and a general decrease in circulating DCs was observed in response to infection. Severe disease coincided with the appearance of monocytic myeloid-derived suppressor cell-like cells and a higher frequency of pre-DC2. Furthermore, phenotypic alterations in MNPs, and their late precursors, were cell-lineage-specific and associated either with the general response against SARS-CoV-2 or COVID-19 severity. This included an interferon-imprint in DC1s observed in all patients and a decreased expression of the coinhibitory molecule CD200R in pre-DCs, DC2s, and DC3 subsets of severely sick patients. Finally, unsupervised analysis revealed that the MNP profile, alone, pointed to a cluster of COVID-19 nonsurvivors. This study provides a reference for the MNP response to SARS-CoV-2 infection and unravels mononuclear phagocyte dysregulations associated with severe COVID-19.
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http://dx.doi.org/10.1073/pnas.2018587118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8017719PMC
February 2021

Innate lymphoid cell composition associates with COVID-19 disease severity.

Clin Transl Immunology 2020 14;9(12):e1224. Epub 2020 Dec 14.

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

Objectives: The role of innate lymphoid cells (ILCs) in coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is unknown. Understanding the immune response in COVID-19 could contribute to unravel the pathogenesis and identification of treatment targets. Here, we describe the phenotypic landscape of circulating ILCs in COVID-19 patients and identified ILC phenotypes correlated to serum biomarkers, clinical markers and laboratory parameters relevant in COVID-19.

Methods: Blood samples collected from moderately ( = 11) and severely ill ( = 12) COVID-19 patients, as well as healthy control donors ( = 16), were analysed with 18-parameter flow cytometry. Using supervised and unsupervised approaches, we examined the ILC activation status and homing profile. Clinical and laboratory parameters were obtained from all COVID-19 patients, and serum biomarkers were analysed with multiplex immunoassays.

Results: Innate lymphoid cells were largely depleted from the circulation of COVID-19 patients compared with healthy controls. Remaining circulating ILCs revealed decreased frequencies of ILC2 in severe COVID-19, with a concomitant decrease of ILC precursors (ILCp) in all patients, compared with controls. ILC2 and ILCp showed an activated phenotype with increased CD69 expression, whereas expression levels of the chemokine receptors CXCR3 and CCR4 were significantly altered in ILC2 and ILCp, and ILC1, respectively. The activated ILC profile of COVID-19 patients was associated with soluble inflammatory markers, while frequencies of ILC subsets were correlated with laboratory parameters that reflect the disease severity.

Conclusion: This study provides insights into the potential role of ILCs in immune responses against SARS-CoV-2, particularly linked to the severity of COVID-19.
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http://dx.doi.org/10.1002/cti2.1224DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7734472PMC
December 2020

Paths taken towards NK cell-mediated immunotherapy of human cancer-a personal reflection.

Scand J Immunol 2021 Jan 20;93(1):e12993. Epub 2020 Nov 20.

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

The discovery that NK cells are able to specifically recognize cells lacking the expression of self-MHC class I molecules provided the first insight into NK cell recognition of tumour cells. It started a flourishing field of NK cell research aimed at exploring the molecular nature of NK cell receptors involved in tumour cell recognition. While much of the important early work was conducted in murine experimental model systems, studies of human NK cells rapidly followed. Over the years, human NK cell research has swiftly progressed, aided by new detailed molecular information on human NK cell development, differentiation, molecular specificity, tissue heterogeneity and functional capacity. NK cells have also been studied in many different diseases aside from cancer, including viral diseases, autoimmunity, allergy and primary immunodeficiencies. These fields of research have all, indirectly or directly, provided further insights into NK cell-mediated recognition of target cells and paved the way for the development of NK cell-based immunotherapies for human cancer. Excitingly, NK cell-based immunotherapy now opens up for novel strategies aimed towards treating malignant diseases, either alone or in combination with other drugs. Reviewed here are some personal reflections of select contributions leading up to the current state-of-the-art in the field, with a particular emphasis on contributions from our own laboratory. This review is part of a series of articles on immunology in Scandinavia, published in conjunction with the 50th anniversary of the Scandinavian Society for Immunology.
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http://dx.doi.org/10.1111/sji.12993DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7816273PMC
January 2021

MAIT cell activation and dynamics associated with COVID-19 disease severity.

Sci Immunol 2020 09;5(51)

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

Severe COVID-19 is characterized by excessive inflammation of the lower airways. The balance of protective versus pathological immune responses in COVID-19 is incompletely understood. Mucosa-associated invariant T (MAIT) cells are antimicrobial T cells that recognize bacterial metabolites, and can also function as innate-like sensors and mediators of antiviral responses. Here, we investigated the MAIT cell compartment in COVID-19 patients with moderate and severe disease, as well as in convalescence. We show profound and preferential decline in MAIT cells in the circulation of patients with active disease paired with strong activation. Furthermore, transcriptomic analyses indicated significant MAIT cell enrichment and pro-inflammatory IL-17A bias in the airways. Unsupervised analysis identified MAIT cell CD69 and CXCR3 immunotypes associated with poor clinical outcome. MAIT cell levels normalized in the convalescent phase, consistent with dynamic recruitment to the tissues and later release back into the circulation when disease is resolved. These findings indicate that MAIT cells are engaged in the immune response against SARS-CoV-2 and suggest their possible involvement in COVID-19 immunopathogenesis.
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http://dx.doi.org/10.1126/sciimmunol.abe1670DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7857393PMC
September 2020

Robust T Cell Immunity in Convalescent Individuals with Asymptomatic or Mild COVID-19.

Cell 2020 10 14;183(1):158-168.e14. Epub 2020 Aug 14.

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

SARS-CoV-2-specific memory T cells will likely prove critical for long-term immune protection against COVID-19. Here, we systematically mapped the functional and phenotypic landscape of SARS-CoV-2-specific T cell responses in unexposed individuals, exposed family members, and individuals with acute or convalescent COVID-19. Acute-phase SARS-CoV-2-specific T cells displayed a highly activated cytotoxic phenotype that correlated with various clinical markers of disease severity, whereas convalescent-phase SARS-CoV-2-specific T cells were polyfunctional and displayed a stem-like memory phenotype. Importantly, SARS-CoV-2-specific T cells were detectable in antibody-seronegative exposed family members and convalescent individuals with a history of asymptomatic and mild COVID-19. Our collective dataset shows that SARS-CoV-2 elicits broadly directed and functionally replete memory T cell responses, suggesting that natural exposure or infection may prevent recurrent episodes of severe COVID-19.
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http://dx.doi.org/10.1016/j.cell.2020.08.017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7427556PMC
October 2020

Expansion of SARS-CoV-2-Specific Antibody-Secreting Cells and Generation of Neutralizing Antibodies in Hospitalized COVID-19 Patients.

J Immunol 2020 11 2;205(9):2437-2446. Epub 2020 Sep 2.

Center for Infectious Medicine, Department of Medicine Huddinge, Karolinska Institutet, 141 52 Stockholm, Sweden;

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), emerged in late 2019 and has since become a global pandemic. Pathogen-specific Abs are typically a major predictor of protective immunity, yet human B cell and Ab responses during COVID-19 are not fully understood. In this study, we analyzed Ab-secreting cell and Ab responses in 20 hospitalized COVID-19 patients. The patients exhibited typical symptoms of COVID-19 and presented with reduced lymphocyte numbers and increased T cell and B cell activation. Importantly, we detected an expansion of SARS-CoV-2 nucleocapsid protein-specific Ab-secreting cells in all 20 COVID-19 patients using a multicolor FluoroSpot Assay. Out of the 20 patients, 16 had developed SARS-CoV-2-neutralizing Abs by the time of inclusion in the study. SARS-CoV-2-specific IgA, IgG, and IgM Ab levels positively correlated with SARS-CoV-2-neutralizing Ab titers, suggesting that SARS-CoV-2-specific Ab levels may reflect the titers of neutralizing Abs in COVID-19 patients during the acute phase of infection. Last, we showed that IL-6 and C-reactive protein serum concentrations were higher in patients who were hospitalized for longer, supporting the recent observations that IL-6 and C-reactive protein could be used as markers for COVID-19 severity. Altogether, this study constitutes a detailed description of clinical and immunological parameters in 20 COVID-19 patients, with a focus on B cell and Ab responses, and describes tools to study immune responses to SARS-CoV-2 infection and vaccination.
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http://dx.doi.org/10.4049/jimmunol.2000717DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7576114PMC
November 2020

Natural killer cell immunotypes related to COVID-19 disease severity.

Sci Immunol 2020 08;5(50)

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

Understanding innate immune responses in COVID-19 is important to decipher mechanisms of host responses and interpret disease pathogenesis. Natural killer (NK) cells are innate effector lymphocytes that respond to acute viral infections but might also contribute to immunopathology. Using 28-color flow cytometry, we here reveal strong NK cell activation across distinct subsets in peripheral blood of COVID-19 patients. This pattern was mirrored in scRNA-seq signatures of NK cells in bronchoalveolar lavage from COVID-19 patients. Unsupervised high-dimensional analysis of peripheral blood NK cells furthermore identified distinct NK cell immunotypes that were linked to disease severity. Hallmarks of these immunotypes were high expression of perforin, NKG2C, and Ksp37, reflecting increased presence of adaptive NK cells in circulation of patients with severe disease. Finally, arming of CD56 NK cells was observed across COVID-19 disease states, driven by a defined protein-protein interaction network of inflammatory soluble factors. This study provides a detailed map of the NK cell activation landscape in COVID-19 disease.
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http://dx.doi.org/10.1126/sciimmunol.abd6832DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7665314PMC
August 2020

Outcome of COVID-19 in multiple myeloma patients in relation to treatment.

Eur J Haematol 2020 Dec 18;105(6):751-754. Epub 2020 Aug 18.

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

COVID-19 has emerged as a global pandemic. Cancer patients have been reported to be at higher risk for adverse outcome of COVID-19. Studies are ongoing to decipher the risk factors and risk groups among cancer patients as well as strategies to refine treatment approaches. Here, we report eight patients with multiple myeloma that underwent immunomodulatory therapies with daratumumab or lenalidomide-based combination treatments and one patient with smoldering multiple myeloma, all of which presented with symptomatic COVID-19. We report that patients that succumbed to COVID-19 presented with either progressive tumor disease under daratumumab treatment or were in remission under lenalidomide-dexamethasone treatment.
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http://dx.doi.org/10.1111/ejh.13502DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7436812PMC
December 2020

Hantavirus inhibits apoptosis by preventing mitochondrial membrane potential loss through up-regulation of the pro-survival factor BCL-2.

PLoS Pathog 2020 02 7;16(2):e1008297. Epub 2020 Feb 7.

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

Hantaviruses, zoonotic RNA viruses belonging to the order Bunyavirales, cause two severe acute diseases in humans, hemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome (HPS). Hantavirus-infected patients show strong cytotoxic lymphocyte responses and hyperinflammation; however, infected cells remain mostly intact. Hantaviruses were recently shown to inhibit apoptosis in infected cells. By inhibiting granzyme B- and TRAIL-mediated apoptosis, hantaviruses specifically and efficiently inhibit cytotoxic lymphocyte-mediated killing of infected cells. Hantaviruses also strongly inhibit apoptosis triggered intrinsically; i.e., initiated through intracellular activation pathways different from those used by cytotoxic lymphocytes. However, insights into the latter mechanisms are currently largely unknown. Here, we dissected the mechanism behind how hantavirus infection, represented by the HFRS-causing Hantaan virus and the HPS-causing Andes virus, results in resistance to staurosporine-induced apoptosis. Less active caspase-8 and caspase-9, and consequently less active caspase-3, was observed in infected compared to uninfected staurosporine-exposed cells. While staurosporine-exposed uninfected cells showed massive release of pro-apoptotic cytochrome C into the cytosol, this was not observed in infected cells. Further, hantaviruses prevented activation of BAX and mitochondrial outer membrane permeabilization (MOMP). In parallel, a significant increase in levels of the pro-survival factor BCL-2 was observed in hantavirus-infected cells. Importantly, direct inhibition of BCL-2 by the inhibitor ABT-737, as well as silencing of BCL-2 by siRNA, resulted in apoptosis in staurosporine-exposed hantavirus-infected cells. Overall, we here provide a tentative mechanism by which hantaviruses protect infected cells from intrinsic apoptosis at the mitochondrial level by inducing an increased expression of the pro-survival factor BCL-2, thereby preventing MOMPs and subsequent activation of caspases. The variety of mechanisms used by hantaviruses to ensure survival of infected cells likely contribute to the persistent infection in natural hosts and may play a role in immunopathogenesis of HFRS and HPS in humans.
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http://dx.doi.org/10.1371/journal.ppat.1008297DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7032725PMC
February 2020

Magnitude and Functional Profile of the Human CD4 T Cell Response throughout Primary Immunization with Tick-Borne Encephalitis Virus Vaccine.

J Immunol 2020 02 10;204(4):914-922. Epub 2020 Jan 10.

Center for Infectious Medicine, ANA Futura, Department of Medicine Huddinge, Karolinska Institutet, 141 52 Stockholm, Sweden;

Tick-borne encephalitis (TBE) is a viral infection of the CNS caused by TBE virus. With no specific treatment available, the only protection is a formalin-inactivated whole virus vaccine. Primary immunization with European TBE vaccines, as recommended by the manufacturers, consists of three vaccine doses administered within a 1-y period. Protection from vaccination is believed to be mediated by Abs, yet T cells may also have a protective role. We set out to characterize the human CD4 T cell response throughout primary TBE immunization. The responses were evaluated before vaccination and 1 mo after each vaccine dose. A heterogeneous magnitude of CD4 T cell-mediated memory responses was observed in regard to lymphoblast expansion and cytokine production (IFN-γ, IL-2, and TNF), with the highest median magnitude detected after the second dose of vaccine. Stimulation with an overlapping peptide library based on structural TBE virus proteins E and C revealed that CD4 T cells concomitantly producing IL-2 and TNF dominated the responses from vaccinees after each vaccine dose, whereas a control cohort of TBE patients responded mainly with all three cytokines. CD107a expression was not upregulated upon peptide stimulation in the vaccinees. However, CD154 (CD40L) expression on cytokine-positive memory CD4 T cells significantly increased after the second vaccine dose. Taken together, TBE vaccination induced CD4 T cell responses dominated by IL-2 and TNF production together with CD154 upregulation and a lower IFN-γ response compared with TBE patients. This response pattern was consistent after all three doses of TBE vaccine.
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http://dx.doi.org/10.4049/jimmunol.1901115DOI Listing
February 2020

Guidelines for the use of flow cytometry and cell sorting in immunological studies (second edition).

Eur J Immunol 2019 Oct;49(10):1457-1973

Flow Cytometry Laboratory, Institute of Molecular Toxicology and Pharmacology, Helmholtz Zentrum München, German Research Center for Environmental Health, München, Germany.

These guidelines are a consensus work of a considerable number of members of the immunology and flow cytometry community. They provide the theory and key practical aspects of flow cytometry enabling immunologists to avoid the common errors that often undermine immunological data. Notably, there are comprehensive sections of all major immune cell types with helpful Tables detailing phenotypes in murine and human cells. The latest flow cytometry techniques and applications are also described, featuring examples of the data that can be generated and, importantly, how the data can be analysed. Furthermore, there are sections detailing tips, tricks and pitfalls to avoid, all written and peer-reviewed by leading experts in the field, making this an essential research companion.
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http://dx.doi.org/10.1002/eji.201970107DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7350392PMC
October 2019

Terminal Effector CD8 T Cells Defined by an IKZF2IL-7R Transcriptional Signature Express FcγRIIIA, Expand in HIV Infection, and Mediate Potent HIV-Specific Antibody-Dependent Cellular Cytotoxicity.

J Immunol 2019 10 13;203(8):2210-2221. Epub 2019 Sep 13.

U.S. Military HIV Research Program, Walter Reed Army Institute of Research, Silver Spring, MD 20910;

HIV-1 infection expands large populations of late-stage differentiated CD8 T cells that may persist long after viral escape from TCR recognition. In this study, we investigated whether such CD8 T cell populations can perform unconventional innate-like antiviral effector functions. Chronic untreated HIV-1 infection was associated with elevated numbers of CD45RACD57 terminal effector CD8 T cells expressing FcγRIIIA (CD16). The FcγRIIIA CD8 T cells displayed a distinctive transcriptional profile between conventional CD8 T cells and NK cells, characterized by high levels of and low expression of This transcriptional profile translated into a distinct NKp80 IL-7Rα surface phenotype with high expression of the Helios transcription factor. Interestingly, the FcγRIIIA CD8 T cells mediated HIV-specific Ab-dependent cellular cytotoxicity (ADCC) activity at levels comparable with NK cells on a per cell basis. The FcγRIIIA CD8 T cells were highly activated in a manner that correlated positively with expansion of the CD8 T cell compartment and with plasma levels of soluble mediators of antiviral immunity and inflammation such as IP-10, TNF, IL-6, and TNFRII. The frequency of FcγRIIIA CD8 T cells persisted as patients initiated suppressive antiretroviral therapy, although their activation levels declined. These data indicate that terminally differentiated effector CD8 T cells acquire enhanced innate cell-like characteristics during chronic viral infection and suggest that HIV-specific ADCC is a function CD8 T cells use to target HIV-infected cells. Furthermore, as the FcγRIIIA CD8 T cells persist in treatment, they contribute significantly to the ADCC-capable effector cell pool in patients on antiretroviral therapy.
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http://dx.doi.org/10.4049/jimmunol.1900422DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6778306PMC
October 2019

NK cells are activated and primed for skin-homing during acute dengue virus infection in humans.

Nat Commun 2019 08 29;10(1):3897. Epub 2019 Aug 29.

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

Despite animal models showing that natural killer (NK) cells are important players in the early defense against many viral infections, the NK cell response is poorly understood in humans. Here we analyze the phenotype, temporal dynamics, regulation and trafficking of NK cells in a patient cohort with acute dengue virus infection. NK cells are robustly activated and proliferate during the first week after symptom debut. Increased IL-18 levels in plasma and in induced skin blisters of DENV-infected patients, as well as concomitant signaling downstream of the IL-18R, suggests an IL-18-dependent mechanism in driving the proliferative NK cell response. Responding NK cells have a less mature phenotype and a distinct chemokine-receptor imprint indicative of skin-homing. A corresponding NK cell subset can be localized to skin early during acute infection. These data provide evidence of an IL-18-driven NK cell proliferation and priming for skin-homing during an acute viral infection in humans.
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http://dx.doi.org/10.1038/s41467-019-11878-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6715742PMC
August 2019

Unique transcriptional and protein-expression signature in human lung tissue-resident NK cells.

Nat Commun 2019 08 26;10(1):3841. Epub 2019 Aug 26.

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

Human lung tissue-resident NK cells (trNK cells) are likely to play an important role in host responses towards viral infections, inflammatory conditions and cancer. However, detailed insights into these cells are still largely lacking. Here we show, using RNA sequencing and flow cytometry-based analyses, that subsets of human lung CD69CD16 NK cells display hallmarks of tissue-residency, including high expression of CD49a, CD103, and ZNF683, and reduced expression of SELL, S1PR5, and KLF2/3. CD49aCD16 NK cells are functionally competent, and produce IFN-γ, TNF, MIP-1β, and GM-CSF. After stimulation with IL-15, they upregulate perforin, granzyme B, and Ki67 to a similar degree as CD49aCD16 NK cells. Comparing datasets from trNK cells in human lung and bone marrow with tissue-resident memory CD8 T cells identifies core genes co-regulated either by tissue-residency, cell-type or location. Together, our data indicate that human lung trNK cells have distinct features, likely regulating their function in barrier immunity.
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http://dx.doi.org/10.1038/s41467-019-11632-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6710242PMC
August 2019

Hantavirus Inhibits TRAIL-Mediated Killing of Infected Cells by Downregulating Death Receptor 5.

Cell Rep 2019 08;28(8):2124-2139.e6

Center for Infectious Medicine, Department of Medicine Huddinge, Karolinska Institutet, Karolinska University Hospital, 141 86 Stockholm, Sweden. Electronic address:

Cytotoxic lymphocytes normally kill virus-infected cells by apoptosis induction. Cytotoxic granule-dependent apoptosis induction engages the intrinsic apoptosis pathway, whereas death receptor (DR)-dependent apoptosis triggers the extrinsic apoptosis pathway. Hantaviruses, single-stranded RNA viruses of the order Bunyavirales, induce strong cytotoxic lymphocyte responses in infected humans. Cytotoxic lymphocytes, however, are largely incapable of eradicating hantavirus-infected cells. Here, we show that the prototypic hantavirus, Hantaan virus (HTNV), induces TRAIL production but strongly inhibits TRAIL-mediated extrinsic apoptosis induction in infected cells by downregulating DR5 cell surface expression. Mechanistic analyses revealed that HTNV triggers both 26S proteasome-dependent degradation of DR5 through direct ubiquitination of DR5 and hampers DR5 transport to the cell surface. These results corroborate earlier findings, demonstrating that hantavirus also inhibits cytotoxic cell granule-dependent apoptosis induction. Together, these findings show that HTNV counteracts intrinsic and extrinsic apoptosis induction pathways, providing a defense mechanism utilized by hantaviruses to inhibit cytotoxic cell-mediated eradication of infected cells.
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http://dx.doi.org/10.1016/j.celrep.2019.07.066DOI Listing
August 2019

Influenza A Virus Infection Induces Hyperresponsiveness in Human Lung Tissue-Resident and Peripheral Blood NK Cells.

Front Immunol 2019 17;10:1116. Epub 2019 May 17.

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

NK cells in the human lung respond to influenza A virus- (IAV-) infected target cells. However, the detailed functional capacity of human lung and peripheral blood NK cells remains to be determined in IAV and other respiratory viral infections. Here, we investigated the effects of IAV infection on human lung and peripheral blood NK cells and following clinical infection. IAV infection of lung- and peripheral blood-derived mononuclear cells induced NK cell hyperresponsiveness to K562 target cells, including increased degranulation and cytokine production particularly in the CD56CD16 subset of NK cells. Furthermore, lung CD16 NK cells showed increased IAV-mediated but target cell-independent activation compared to CD16 lung NK cells or total NK cells in peripheral blood. IAV infection rendered peripheral blood NK cells responsive toward the normally NK cell-resistant lung epithelial cell line A549, indicating that NK cell activation during IAV infection could contribute to killing of surrounding non-infected epithelial cells. , peripheral blood CD56CD16 and CD56CD16 NK cells were primed during acute IAV infection, and a small subset of CD16CD49aCXCR3 NK cells could be identified, with CD49a and CXCR3 potentially promoting homing to and tissue-retention in the lung during acute infection. Together, we show that IAV respiratory viral infections prime otherwise hyporesponsive lung NK cells, indicating that both CD16 and CD16 NK cells including CD16CD49a tissue-resident NK cells could contribute to host immunity but possibly also tissue damage in clinical IAV infection.
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http://dx.doi.org/10.3389/fimmu.2019.01116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6534051PMC
September 2020

Chronic hepatitis delta virus infection leads to functional impairment and severe loss of MAIT cells.

J Hepatol 2019 08 14;71(2):301-312. Epub 2019 May 14.

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

Background & Aims: Hepatitis delta virus (HDV) infection is the most severe form of viral hepatitis. Although HDV-associated liver disease is considered immune-mediated, adaptive immune responses against HDV are weak. Thus, the role of several other cell-mediated mechanisms such as those driven by mucosa-associated invariant T (MAIT) cells, a group of innate-like T cells highly enriched in the human liver, has not been extensively studied in clinical HDV infection.

Methods: MAIT cells from a sizeable cohort of patients with chronic HDV were analyzed ex vivo and in vitro after stimulation. Results were compared with MAIT cells from hepatitis B virus (HBV) monoinfected patients and healthy controls.

Results: Circulating MAIT cells were dramatically decreased in the peripheral blood of HDV-infected patients. Signs of decline were also observed in the liver. In contrast, only a modest decrease of circulating MAIT cells was noted in HBV monoinfection. Unsupervised high-dimensional analysis of residual circulating MAIT cells in chronic HDV infection revealed the appearance of a compound phenotype of CD38PD-1CD28CD127PLZFEomesHelios cells indicative of activation. Corroborating these results, MAIT cells exhibited a functionally impaired responsiveness. In parallel to MAIT cell loss, HDV-infected patients exhibited signs of monocyte activation and increased levels of proinflammatory cytokines IL-12 and IL-18. In vitro, IL-12 and IL-18 induced an activated MAIT cell phenotype similar to the one observed ex vivo in HDV-infected patients. These cytokines also promoted MAIT cell death, suggesting that they may contribute to MAIT cell activation and subsequent loss during HDV infection.

Conclusions: These results suggest that chronic HDV infection engages the MAIT cell compartment causing activation, functional impairment, and subsequent progressive loss of MAIT cells as the HDV-associated liver disease progresses.

Lay Summary: Hepatitis delta virus (HDV) infection is the most severe form of viral hepatitis. We found that in patients with HDV, a subset of innate-like T cells called mucosa-associated invariant T cells (or MAIT cells), which are normally abundant in peripheral blood and the liver, are activated, functionally impaired and severely depleted.
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http://dx.doi.org/10.1016/j.jhep.2019.04.009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6642010PMC
August 2019

Breadth and Dynamics of HLA-A2- and HLA-B7-Restricted CD8 T Cell Responses against Nonstructural Viral Proteins in Acute Human Tick-Borne Encephalitis Virus Infection.

Immunohorizons 2018 07 2;2(6):172-184. Epub 2018 Jul 2.

Center for Infectious Medicine, Department of Medicine Huddinge, Karolinska Institutet, Karolinska University Hospital, 141 86 Stockholm, Sweden;

Tick-borne encephalitis virus (TBEV) is a leading cause of viral meningoencephalitis in many parts of Europe and eastwards in Asia, with high morbidity and often long-term neurologic sequelae. With no treatment available, studies of the immune response to TBEV are essential for the understanding of the immunopathogenesis of tick-borne encephalitis and for the development of therapeutics. We have previously demonstrated that CD8 T cell responses in peripheral blood in patients with acute TBEV peak at around 7 d after hospitalization in the neuroinvasive phase of the disease. In this study, we identified six novel TBEV HLA-A2- and HLA-B7-restricted epitopes, all derived from the nonstructural proteins of TBEV. This identification allowed for a comprehensive phenotypic and temporal analysis of the HLA-A2- and HLA-B7-restricted Ag-specific CD8 T cell response during the acute stages of human TBEV infection. HLA-A2- and HLA-B7-restricted TBEV epitope-specific effector cells predominantly displayed a CD45RACCR7CD27CD57 phenotype at day 7, which transitioned into separate distinct phenotypes for HLA-A2- and HLA-B7-restricted TBEV-specific CD8 T cells, respectively. At day 21, the most prevalent phenotype in the HLA-A2-restricted CD8 T cell populations was CD45RACCR7CD27CD57, whereas the HLA-B7-restricted CD8 T cell population was predominantly CD45RACCR7CD27CD57 Almost all TBEV epitope-specific CD8 T cells expressed α4 and β1 integrins at days 7 and 21, whereas the bulk CD8 T cells expressed lower integrin levels. Taken together, human TBEV infection elicits broad responses to multiple epitopes, predominantly derived from the nonstructural part of the virus, establishing distinct maturation patterns for HLA-A2- and HLA-B7-restricted TBEV epitope-specific CD8 T cells.
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http://dx.doi.org/10.4049/immunohorizons.1800029DOI Listing
July 2018

Serum Markers Associated with Severity and Outcome of Hantavirus Pulmonary Syndrome.

J Infect Dis 2019 05;219(11):1832-1840

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

Background: Hantavirus pulmonary syndrome (HPS) is caused by Andes virus (ANDV) and related hantaviruses in the Americas. Despite a fatality rate of 40%, the pathogenesis of HPS is poorly understood and factors associated with severity, fatality, and survival remain elusive.

Methods: Ninety-three ANDV-infected HPS patients, of whom 34 had a fatal outcome, were retrospectively studied. Serum levels of cytokines and other inflammation-associated markers were analyzed using multiplex immunoassay and enzyme-linked immunosorbent assay. Associations with disease severity, fatal outcome, and survival were identified using logistic regression.

Results: HPS patients exhibited increased serum levels of markers associated with inflammation, intestinal damage, and microbial translocation compared to controls. Patients with fatal outcome displayed higher levels of interleukin (IL) 6, IL-10, interferon-γ, soluble tumor necrosis factor-related apoptosis-inducing ligand, and intestinal fatty acid-binding protein (I-FABP) than survivors. Levels of complement factor 5/5a were higher in survivors compared with fatal cases. IL-6 and I-FABP, the latter a marker for intestinal damage, were by multivariate analyses identified as independent markers associated with disease severity (odds ratio [OR], 2.25; 95% confidence interval [CI], 1.01-5.01) and fatal outcome (OR, 1.64; 95% CI, 1.01-2.64), respectively.

Conclusions: HPS patients displayed a multifaceted, systemic inflammatory response, with IL-6 and I-FABP as independent markers of disease severity and fatality, respectively.
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http://dx.doi.org/10.1093/infdis/jiz005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6500549PMC
May 2019

Orthohantaviruses belonging to three phylogroups all inhibit apoptosis in infected target cells.

Sci Rep 2019 01 29;9(1):834. Epub 2019 Jan 29.

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

Orthohantaviruses, previously known as hantaviruses, are zoonotic viruses that can cause hantavirus pulmonary syndrome (HPS) and hemorrhagic fever with renal syndrome (HFRS) in humans. The HPS-causing Andes virus (ANDV) and the HFRS-causing Hantaan virus (HTNV) have anti-apoptotic effects. To investigate if this represents a general feature of orthohantaviruses, we analysed the capacity of six different orthohantaviruses - belonging to three distinct phylogroups and representing both pathogenic and non-pathogenic viruses - to inhibit apoptosis in infected cells. Primary human endothelial cells were infected with ANDV, HTNV, the HFRS-causing Puumala virus (PUUV) and Seoul virus, as well as the putative non-pathogenic Prospect Hill virus and Tula virus. Infected cells were then exposed to the apoptosis-inducing chemical staurosporine or to activated human NK cells exhibiting a high cytotoxic potential. Strikingly, all orthohantaviruses inhibited apoptosis in both settings. Moreover, we show that the nucleocapsid (N) protein from all examined orthohantaviruses are potential targets for caspase-3 and granzyme B. Recombinant N protein from ANDV, PUUV and the HFRS-causing Dobrava virus strongly inhibited granzyme B activity and also, to certain extent, caspase-3 activity. Taken together, this study demonstrates that six different orthohantaviruses inhibit apoptosis, suggesting this to be a general feature of orthohantaviruses likely serving as a mechanism of viral immune evasion.
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http://dx.doi.org/10.1038/s41598-018-37446-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6351540PMC
January 2019

Cell-Mediated Immune Responses and Immunopathogenesis of Human Tick-Borne Encephalitis Virus-Infection.

Front Immunol 2018 26;9:2174. Epub 2018 Sep 26.

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

Tick-borne encephalitis virus (TBEV) is a flavivirus that belongs to the family. TBEV is transmitted to humans primarily from infected ticks. The virus causes tick-borne encephalitis (TBE), an acute viral disease that affects the central nervous system (CNS). Infection can lead to acute neurological symptoms of significant severity due to meningitis or meningo(myelo)encephalitis. TBE can cause long-term suffering and has been recognized as an increasing public health problem. TBEV-affected areas currently include large parts of central and northern Europe as well as northern Asia. Infection with TBEV triggers a humoral as well as a cell-mediated immune response. In contrast to the well-characterized humoral antibody-mediated response, the cell-mediated immune responses elicited to natural TBEV-infection have been poorly characterized until recently. Here, we review recent progress in our understanding of the cell-mediated immune response to human TBEV-infection. A particular emphasis is devoted to studies of the response mediated by natural killer (NK) cells and CD8 T cells. The studies described include results revealing the temporal dynamics of the T cell- as well as NK cell-responses in relation to disease state and functional characterization of these cells. Additionally, we discuss specific immunopathological aspects of TBEV-infection in the CNS.
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http://dx.doi.org/10.3389/fimmu.2018.02174DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6168641PMC
October 2019

Proteome analysis of human CD56 NK cells reveals a homogeneous phenotype surprisingly similar to CD56 NK cells.

Eur J Immunol 2018 09 1;48(9):1456-1469. Epub 2018 Aug 1.

Cellular Proteomics, Helmholtz Centre for Infection Research, Braunschweig, Germany.

NK cells lacking CD56 (CD56 ) were first identified in chronic HIV-1 infection. However, CD56 NK cells also exist in healthy individuals, albeit in significantly lower numbers. Here, we provide an extensive proteomic characterisation of human CD56 peripheral blood NK cells of healthy donors and compare them to their CD56 and CD56 counterparts. Unbiased large-scale surface receptor profiling clustered CD56 cells as part of the main NK cell compartment and indicated an overall CD56 -like phenotype. Total proteome analyses of CD56 NK cells further confirmed their similarity with CD56 NK cells, and revealed a complete cytolytic inventory with high levels of perforin and granzyme H and M. In the present study, twelve proteins discriminated CD56 NK cells from CD56 NK cells with nine up-regulated and three down-regulated proteins in the CD56 NK cell population. Those proteins were functionally related to lytic granule composition and transport, interaction with the extracellular matrix, DNA transcription or repair, and proliferation. Corroborating these results, CD56 NK cells showed modest cytotoxicity, degranulation, and IFN-ɣ secretion as compared to CD56 NK cells. In conclusion, CD56 NK cells constitute functionally competent cells sharing many features of bona fide CD56 NK cells in healthy individuals, but with some distinct characteristics.
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http://dx.doi.org/10.1002/eji.201747450DOI Listing
September 2018

Chronic hepatitis C virus infection irreversibly impacts human natural killer cell repertoire diversity.

Nat Commun 2018 06 11;9(1):2275. Epub 2018 Jun 11.

Department of Medicine Huddinge, Center for Infectious Medicine, Karolinska Institutet, Karolinska University Hospital Huddinge, 14186, Stockholm, Sweden.

Diversity is a central requirement for the immune system's capacity to adequately clear a variety of different infections. As such, natural killer (NK) cells represent a highly diverse population of innate lymphocytes important in the early response against viruses. Yet, the extent to which a chronic pathogen affects NK cell diversity is largely unknown. Here we study NK cell functional diversification in chronic hepatitis C virus (HCV) infection. High-dimensional flow cytometer assays combined with stochastic neighbor embedding analysis reveal that chronic HCV infection induces functional imprinting on human NK cells that is largely irreversible and persists long after successful interventional clearance of the virus. Furthermore, HCV infection increases inter-individual, but decreases intra-individual, NK cell diversity. Taken together, our results provide insights into how the history of infections affects human NK cell diversity.
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http://dx.doi.org/10.1038/s41467-018-04685-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5995831PMC
June 2018
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