Recent Submissions

  • Ly6G deficiency alters the dynamics of neutrophil recruitment and pathogen capture during Leishmania major skin infection.

    Kleinholz, Corinna L; Riek-Burchardt, Monika; Seiß, Elena A; Amore, Jonas; Gintschel, Patricia; Philipsen, Lars; Bousso, Philippe; Relja, Borna; Schraven, Burkhart; Handschuh, Juliane; et al. (NPG, 2021-07-23)
    Neutrophils represent one of the first immune cell types recruited to sites of infection, where they can control pathogens by phagocytosis and cytotoxic mechanisms. Intracellular pathogens such as Leishmania major can hijack neutrophils to establish an efficient infection. However the dynamic interactions of neutrophils with the pathogen and other cells at the site of the infection are incompletely understood. Here, we have investigated the role of Ly6G, a homolog of the human CD177 protein, which has been shown to interact with cell adhesion molecules, and serves as a bona fide marker for neutrophils in mice. We show that Ly6G deficiency decreases the initial infection rate of neutrophils recruited to the site of infection. Although the uptake of L. major by subsequently recruited monocytes was tightly linked with the concomitant uptake of neutrophil material, this process was not altered by Ly6G deficiency of the neutrophils. Instead, we observed by intravital 2-photon microscopy that Ly6G-deficient neutrophils entered the site of infection with delayed initial recruitment kinetics. Thus, we conclude that by promoting neutrophils' ability to efficiently enter the site of infection, Ly6G contributes to the early engagement of intracellular pathogens by the immune system.
  • Serum Concentration of the Phytohormone Abscisic Acid Is Associated With Immune-Regulatory Mediators and Is a Potential Biomarker of Disease Severity in Chronic Obstructive Pulmonary Disease.

    Hoang, Quynh Trang Mi; Nguyen, Van Kinh; Oberacher, Herbert; Fuchs, Dietmar; Hernandez-Vargas, Esteban A; Borucki, Katrin; Waldburg, Nadine; Wippermann, Jens; Schreiber, Jens; Bruder, Dunja; et al. (Frontiers, 2021-06-08)
    COPD and asthma are two distinct but sometimes overlapping diseases exhibiting varying degrees and types of inflammation on different stages of the disease. Although several biomarkers are defined to estimate the inflammatory endotype and stages in these diseases, there is still a need for new markers and potential therapeutic targets. We investigated the levels of a phytohormone, abscisic acid (ABA) and its receptor, LANCL2, in COPD patients and asthmatics. In addition, PPAR-γ that is activated by ABA in a ligand-binding domain-independent manner was also included in the study. In this study, we correlated ABA with COPD-propagating factors to define the possible role of ABA, in terms of immune regulation, inflammation, and disease stages. We collected blood from 101 COPD patients, 52 asthmatics, and 57 controls. Bronchoscopy was performed on five COPD patients and 29 controls. We employed (i) liquid chromatography-tandem mass spectrometry and HPLC to determine the ABA and indoleamine 2,3-dioxygenase levels, respectively; (ii) real-time PCR to quantify the gene expression of LANCL2 and PPAR-γ; (iii) Flow cytometry to quantify adipocytokines; and (iv) immunoturbidimetry and ELISA to measure CRP and cytokines, respectively. Finally, a multinomial regression model was used to predict the probability of using ABA as a biomarker. Blood ABA levels were significantly reduced in COPD patients and asthmatics compared to age- and gender-matched normal controls. However, PPAR-γ was elevated in COPD patients. Intriguingly, ABA was positively correlated with immune-regulatory factors and was negatively correlated with inflammatory markers, in COPD. Of note, ABA was increased in advanced COPD stages. We thereby conclude that ABA might be involved in regulation of COPD pathogenesis and might be regarded as a potential biomarker for COPD stages.
  • Post-injury immunosuppression and secondary infections are caused by an AIM2 inflammasome-driven signaling cascade.

    Roth, Stefan; Cao, Jiayu; Singh, Vikramjeet; Tiedt, Steffen; Hundeshagen, Gabriel; Li, Ting; Boehme, Julia D; Chauhan, Dhruv; Zhu, Jie; Ricci, Alessio; et al. (Elsevier (Cell Press), 2021-03-04)
    Loss of lymphocytes, particularly T cell apoptosis, is a central pathological event after severe tissue injury that is associated with increased susceptibility for life-threatening infections. The precise immunological mechanisms leading to T cell death after acute injury are largely unknown. Here, we identified a monocyte-T cell interaction driving bystander cell death of T cells in ischemic stroke and burn injury. Specifically, we found that stroke induced a FasL-expressing monocyte population, which led to extrinsic T cell apoptosis. This phenomenon was driven by AIM2 inflammasome-dependent interleukin-1β (IL-1β) secretion after sensing cell-free DNA. Pharmacological inhibition of this pathway improved T cell survival and reduced post-stroke bacterial infections. As such, this study describes inflammasome-dependent monocyte activation as a previously unstudied cause of T cell death after injury and challenges the current paradigms of post-injury lymphopenia.
  • Influenza A virus-induced thymus atrophy differentially affects dynamics of conventional and regulatory T-cell development in mice.

    Elfaki, Yassin; Robert, Philippe A; Binz, Christoph; Falk, Christine S; Bruder, Dunja; Prinz, Immo; Floess, Stefan; Meyer-Hermann, Michael; Huehn, Jochen; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany.; BRICS, Braunschweiger Zentrum für Systembiologie, Rebenring 56,38106 Braunschweig, Germany. (Wiley-VCH, 2021-02-26)
    Foxp3+ Treg cells, which are crucial for maintenance of self-tolerance, mainly develop within the thymus, where they arise from CD25+ Foxp3- or CD25- Foxp3+ Treg cell precursors. Although it is known that infections can cause transient thymic involution, the impact of infection-induced thymus atrophy on thymic Treg (tTreg) cell development is unknown. Here, we infected mice with influenza A virus (IAV) and studied thymocyte population dynamics post infection. IAV infection caused a massive, but transient thymic involution, dominated by a loss of CD4+ CD8+ double-positive (DP) thymocytes, which was accompanied by a significant increase in the frequency of CD25+ Foxp3+ tTreg cells. Differential apoptosis susceptibility could be experimentally excluded as a reason for the relative tTreg cell increase, and mathematical modeling suggested that enhanced tTreg cell generation cannot explain the increased frequency of tTreg cells. Yet, an increased death of DP thymocytes and augmented exit of single-positive (SP) thymocytes was suggested to be causative. Interestingly, IAV-induced thymus atrophy resulted in a significantly reduced T-cell receptor (TCR) repertoire diversity of newly produced tTreg cells. Taken together, IAV-induced thymus atrophy is substantially altering the dynamics of major thymocyte populations, finally resulting in a relative increase of tTreg cells with an altered TCR repertoire.
  • Immunity to TBEV Related Flaviviruses with Reduced Pathogenicity Protects Mice from Disease but Not from TBEV Entry into the CNS.

    Petry, Monique; Palus, Martin; Leitzen, Eva; Mitterreiter, Johanna Gracia; Huang, Bei; Kröger, Andrea; Verjans, Georges M G M; Baumgärtner, Wolfgang; Rimmelzwaan, Guus F; Růžek, Daniel; et al. (MDPI, 2021-02-26)
    Tick-borne encephalitis virus (TBEV) is a leading cause of vector-borne viral encephalitis with expanding endemic regions across Europe. In this study we tested in mice the efficacy of preinfection with a closely related low-virulent flavivirus, Langat virus (LGTV strain TP21), or a naturally avirulent TBEV strain (TBEV-280) in providing protection against lethal infection with the highly virulent TBEV strain (referred to as TBEV-Hypr). We show that prior infection with TP21 or TBEV-280 is efficient in protecting mice from lethal TBEV-Hypr challenge. Histopathological analysis of brains from nonimmunized mice revealed neuronal TBEV infection and necrosis. Neuroinflammation, gliosis, and neuronal necrosis was however also observed in some of the TP21 and TBEV-280 preinfected mice although at reduced frequency as compared to the nonimmunized TBEV-Hypr infected mice. qPCR detected the presence of viral RNA in the CNS of both TP21 and TBEV-280 immunized mice after TBEV-Hypr challenge, but significantly reduced compared to mock-immunized mice. Our results indicate that although TBEV-Hypr infection is effectively controlled in the periphery upon immunization with low-virulent LGTV or naturally avirulent TBEV 280, it may still enter the CNS of these animals. These findings contribute to our understanding of causes for vaccine failure in individuals vaccinated with TBE vaccines.
  • Chemical Conjugation of a Purified DEC-205-Directed Antibody with Full-Length Protein for Targeting Mouse Dendritic Cells In Vitro and In Vivo.

    Volckmar, Julia; Knop, Laura; Hirsch, Tatjana; Frentzel, Sarah; Erck, Christian; van Ham, Marco; Stegemann-Koniszewski, Sabine; Bruder, Dunja; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (MyJove Corporation, 2021-02-05)
    Targeted antigen delivery to cross-presenting dendritic cells (DC) in vivo efficiently induces T effector cell responses and displays a valuable approach in vaccine design. Antigen is delivered to DC via antibodies specific for endocytosis receptors such as DEC-205 that induce uptake, processing, and MHC class I- and II-presentation. Efficient and reliable conjugation of the desired antigen to a suitable antibody is a critical step in DC targeting and among other factors depends on the format of the antigen. Chemical conjugation of full-length protein to purified antibodies is one possible strategy. In the past, we have successfully established cross-linking of the model antigen ovalbumin (OVA) and a DEC-205-specific IgG2a antibody (αDEC-205) for in vivo DC targeting studies in mice. The first step of the protocol is the purification of the antibody from the supernatant of the NLDC (non-lymphoid dendritic cells)-145 hybridoma by affinity chromatography. The purified antibody is activated for chemical conjugation by sulfo-SMCC (sulfosuccinimidyl 4-[N-maleimidomethyl] cyclohexane-1-carboxylate) while at the same time the sulfhydryl-groups of the OVA protein are exposed through incubation with TCEP-HCl (tris (2-carboxyethyl) phosphine hydrochloride). Excess TCEP-HCl and sulfo-SMCC are removed and the antigen is mixed with the activated antibody for overnight coupling. The resulting αDEC-205/OVA conjugate is concentrated and freed from unbound OVA. Successful conjugation of OVA to αDEC-205 is verified by western blot analysis and enzyme-linked immunosorbent assay (ELISA). We have successfully used chemically crosslinked αDEC-205/OVA to induce cytotoxic T cell responses in the liver and to compare different adjuvants for their potential in inducing humoral and cellular immunity following in vivo targeting of DEC-205+ DC. Beyond that, such chemically coupled antibody/antigen conjugates offer valuable tools for the efficient induction of vaccine responses to tumor antigens and have been proven to be superior to classical immunization approaches regarding the prevention and therapy of various types of tumors.
  • Resolved Influenza A Virus Infection Has Extended Effects on Lung Homeostasis and Attenuates Allergic Airway Inflammation in a Mouse Model.

    Wu, Qingyu; Jorde, Ilka; Kershaw, Olivia; Jeron, Andreas; Bruder, Dunja; Schreiber, Jens; Stegemann-Koniszewski, Sabine; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (MDPI, 2020-11-27)
    Allergic airway inflammation (AAI) involves T helper cell type 2 (Th2) and pro-inflammatory responses to aeroallergens and many predisposing factors remain elusive. Influenza A virus (IAV) is a major human pathogen that causes acute respiratory infections and induces specific immune responses essential for viral clearance and resolution of the infection. Beyond acute infection, IAV has been shown to persistently affect lung homeostasis and respiratory immunity. Here we asked how resolved IAV infection affects subsequently induced AAI. Mice infected with a sublethal dose of IAV were sensitized and challenged in an ovalbumin mediated mouse model for AAI after resolution of the acute viral infection. Histological changes, respiratory leukocytes, cytokines and airway hyperreactivity were analyzed in resolved IAV infection alone and in AAI with and without previous IAV infection. More than five weeks after infection, we detected persistent pneumonia with increased activated CD4+ and CD8+ lymphocytes as well as dendritic cells and MHCII expressing macrophages in the lung. Resolved IAV infection significantly affected subsequently induced AAI on different levels including morphological changes, respiratory leukocytes and lymphocytes as well as the pro-inflammatory cytokine responses, which was clearly diminished. We conclude that IAV has exceptional persisting effects on respiratory immunity with substantial consequences for subsequently induced AAI.
  • Early career members at the ers lung science conference 2020: Metabolic alterations in lung ageing and disease

    Ogger, Patricia P.; Silva, Johnatas Dutra; Aghapour, Mahyar; Persson, Irma Mahmutovic; Tulen, Christy; Jurkowska, Renata; Ubags, Niki D.; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (European Respiratory Society, 2020-09-01)
    Every year, the European Respiratory Society (ERS) organises the Lung Science Conference (LSC) in Estoril, Portugal, to discuss basic and translational science. The topic of the LSC 2020 was “Metabolic alterations in lung ageing and disease”. In addition to an outstanding scientific programme, the LSC provides excellent opportunities for career development and inclusion of Early Career Members (ECMs). All scientific and poster sessions are chaired by an ECM who is paired with a senior faculty member to allow ECMs to become acquainted with session chairing. In addition, 40 travel bursaries are made available to abstract authors and all bursary recipients are invited to take part in a mentorship lunch. Moreover, there is a session organised by the Early Career Members Committee (ECMC) dedicated to career development. Here, we describe the scientific highlights of LSC 2020 for those who could not attend. The ERS presents several awards at the LSC and here we will highlight all winners of the LSC 2020 awards. The five highest ranked abstracts from ECMs are presented during the Young investigator session. Patricia Ogger (UK) was presented with the William MacNee Award for the best presentation in this session. Several abstracts were selected for programmed oral presentations and Renata Jurkowska (UK) was presented with the inaugural Geoffrey Laurent Award for the best oral presentation. Moreover, the organisers presented eight Distinguished Poster awards to Anne-Sophie Lamort (Germany), Julia Frankenberg Garcia (UK), Johnatas Silva (UK), Pauline Esteves (France), Claudio Bussi (UK), Elodie Picard (France), Felix Ritzmann (Germany) and Alen Faiz (Australia) for their excellent contributions during the poster session.
  • Respiratory Influenza A Virus Infection Triggers Local and Systemic Natural Killer Cell Activation Toll-Like Receptor 7.

    Stegemann-Koniszewski, Sabine; Behrens, Sarah; Boehme, Julia D; Hochnadel, Inga; Riese, Peggy; Guzmán, Carlos A; Kröger, Andrea; Schreiber, Jens; Gunzer, Matthias; Bruder, Dunja; et al. (Frontiers, 2018-02-13)
    The innate immune system senses influenza A virus (IAV) through different pathogen-recognition receptors including Toll-like receptor 7 (TLR7). Downstream of viral recognition natural killer (NK) cells are activated as part of the anti-IAV immune response. Despite the known decisive role of TLR7 for NK cell activation by therapeutic immunostimulatory RNAs, the contribution of TLR7 to the NK cell response following IAV infection has not been addressed. We have analyzed lung cytokine responses as well as the activation, interferon (IFN)-γ production, and cytotoxicity of lung and splenic NK cells following sublethal respiratory IAV infection in wild-type and TLR7ko mice. Early airway IFN-γ levels as well as the induction of lung NK cell CD69 expression and IFN-γ production in response to IAV infection were significantly attenuated in TLR7-deficient hosts. Strikingly, respiratory IAV infection also primed splenic NK cells for IFN-γ production, degranulation, and target cell lysis, all of which were fully dependent on TLR7. At the same time, lung type I IFN levels were significantly reduced in TLR7ko mice early following IAV infection, displaying a potential upstream mechanism of the attenuated NK cell activation observed. Taken together, our data clearly demonstrate a specific role for TLR7 signaling in local and systemic NK cell activation following respiratory IAV infection despite the presence of redundant innate IAV-recognition pathways.
  • PD-1/PD-L1 pathway inhibition to restore effector functions in exhausted CD8+ T cells: chances, limitations and potential risks

    Veluswamy, Priya; Bruder, Dunja; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (AME Publishing Company, 2018-04)
    T cell exhaustion is a well-known mechanism involved in escape of degenerated cells or certain pathogens from CD8+ T cell-mediated immune surveillance, ultimately resulting in tumor development and chronic infections, respectively. Next to activated T cells, exhausted CD8+ T cells typically express high levels of the programmed cell death-1 (PD-1) receptor. While interaction of PD-1 with its ligand programmed death-ligand 1 (PD-L1) on hemotopoietic and non-hemotopoietic cells is important for the re-establishment of homeostasis following immune activation, PD-1/PD-L1 interaction represents a major drawback in certain other disease settings such as cancer or chronic viral infections. Here PD-1 signalling in T cells prevents efficient anti-tumor or anti-viral immune responses. Thus, therapeutic interference with the PD-1/PD-L1 pathway represents a promising approach for releasing exhausted CD8+ T cells from PD-1-dependent suppression and reactivation of effector functions. However, recent reports have highlighted unexpected outcomes of PD-1/PD-L1 pathway inhibition in the context of chronic infections. We provide here a comprehensive overview of the recent discoveries made in the context of PD-1/PD-L1 checkpoint inhibition that are considered relevant with respect to the targeted reactivation of effector functions in exhausted CD8+ T cells. We briefly discuss the impact of PD-1 signalling on the expression of certain transcription factors, on epigenetic modifications affecting chromatin accessibility, on cellular metabolism and the expression of certain cytokine receptors involved in immune homeostasis. These newly uncovered facts should be carefully considered before further development of therapies targeting the PD-1/PD-L1 pathway that are aiming at the restoration of pathogen-specific and anti-tumor CD8+ T cell effector functions in order to prevent adverse side effects. © 2018, Translational Cancer Research.
  • ADAP Promotes Degranulation and Migration of NK Cells Primed During vivo Listeria monocytogenes Infection in Mice.

    Böning, Martha A L; Trittel, Stephanie; Riese, Peggy; van Ham, Marco; Heyner, Maxi; Voss, Martin; Parzmair, Gerald P; Klawonn, Frank; Jeron, Andreas; Guzman, Carlos A; et al. (Frontiers, 2019-01-01)
    The adhesion and degranulation-promoting adaptor protein (ADAP) serves as a multifunctional scaffold and is involved in the formation of immune signaling complexes. To date only limited and moreover conflicting data exist regarding the role of ADAP in NK cells. To extend existing knowledge we investigated ADAP-dependency of NK cells in the context of in vivo infection with the intracellular pathogen Listeria monocytogenes (Lm). Ex vivo analysis of infection-primed NK cells revealed impaired cytotoxic capacity in NK cells lacking ADAP as indicated by reduced CD107a surface expression and inefficient perforin production. However, ADAP-deficiency had no global effect on NK cell morphology or intracellular distribution of CD107a-containing vesicles. Proteomic definition of ADAPko and wild type NK cells did not uncover obvious differences in protein composition during the steady state and moreover, similar early response patterns were induced in NK cells upon infection independent of the genotype. In line with protein network analyses that suggested an altered migration phenotype in naïve ADAPko NK cells, in vitro migration assays uncovered significantly reduced migration of both naïve as well as infection-primed ADAPko NK cells compared to wild type NK cells. Notably, this migration defect was associated with a significantly reduced expression of the integrin CD11a on the surface of splenic ADAP-deficient NK cells 1 day post-Lm infection. We propose that ADAP-dependent alterations in integrin expression might account at least in part for the fact that during in vivo infection significantly lower numbers of ADAPko NK cells accumulate in the spleen i.e., the site of infection. In conclusion, we show here that during systemic Lm infection in mice ADAP is essential for efficient cytotoxic capacity and migration of NK cells.
  • Mucosal CD8+ T cell responses induced by an MCMV based vaccine vector confer protection against influenza challenge.

    Zheng, Xiaoyan; Oduro, Jennifer D; Boehme, Julia D; Borkner, Lisa; Ebensen, Thomas; Heise, Ulrike; Gereke, Marcus; Pils, Marina C; Krmpotic, Astrid; Guzmán, Carlos A; et al. (PLOS, 2019-09-01)
    Cytomegalovirus (CMV) is a ubiquitous β-herpesvirus that establishes life-long latent infection in a high percentage of the population worldwide. CMV induces the strongest and most durable CD8+ T cell response known in human clinical medicine. Due to its unique properties, the virus represents a promising candidate vaccine vector for the induction of persistent cellular immunity. To take advantage of this, we constructed a recombinant murine CMV (MCMV) expressing an MHC-I restricted epitope from influenza A virus (IAV) H1N1 within the immediate early 2 (ie2) gene. Only mice that were immunized intranasally (i.n.) were capable of controlling IAV infection, despite the greater potency of the intraperitoneally (i.p.) vaccination in inducing a systemic IAV-specific CD8+ T cell response. The protective capacity of the i.n. immunization was associated with its ability to induce IAV-specific tissue-resident memory CD8+ T (CD8TRM) cells in the lungs. Our data demonstrate that the protective effect exerted by the i.n. immunization was critically mediated by antigen-specific CD8+ T cells. CD8TRM cells promoted the induction of IFNγ and chemokines that facilitate the recruitment of antigen-specific CD8+ T cells to the lungs. Overall, our results showed that locally applied MCMV vectors could induce mucosal immunity at sites of entry, providing superior immune protection against respiratory infections.
  • Longitudinal proliferation mapping in vivo reveals NADPH oxidase-mediated dampening of Staphylococcus aureus growth rates within neutrophils.

    Seiß, Elena A; Krone, Anna; Formaglio, Pauline; Goldmann, Oliver; Engelmann, Susanne; Schraven, Burkhart; Medina, Eva; Müller, Andreas J; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (Nature publishing group, 2019-04-05)
    Upon the onset of inflammatory responses, bacterial pathogens are confronted with altered tissue microenvironments which can critically impact on their metabolic activity and growth. Changes in these parameters have however remained difficult to analyze over time, which would be critical to dissect the interplay between the host immune response and pathogen physiology. Here, we established an in vivo biosensor for measuring the growth rates of Staphylococcus aureus (S. aureus) on a single cell-level over days in an ongoing cutaneous infection. Using intravital 2-photon imaging and quantitative fluorescence microscopy, we show that upon neutrophil recruitment to the infection site and bacterial uptake, non-lethal dampening of S. aureus proliferation occurred. This inhibition was supported by NADPH oxidase activity. Therefore, reactive oxygen production contributes to pathogen containment within neutrophils not only by killing S. aureus, but also by restricting the growth rate of the bacterium.
  • Filamin A Phosphorylation at Serine 2152 by the Serine/Threonine Kinase Ndr2 Controls TCR-Induced LFA-1 Activation in T Cells.

    Waldt, Natalie; Seifert, Anke; Demiray, Yunus Emre; Devroe, Eric; Turk, Benjamin E; Reichardt, Peter; Mix, Charlie; Reinhold, Annegret; Freund, Christian; Müller, Andreas J; et al. (Frontiers, 2018-01-01)
    The integrin LFA-1 (CD11a/CD18) plays a critical role in the interaction of T cells with antigen presenting cells (APCs) to promote lymphocyte differentiation and proliferation. This integrin can be present either in a closed or in an open active conformation and its activation upon T-cell receptor (TCR) stimulation is a critical step to allow interaction with APCs. In this study we demonstrate that the serine/threonine kinase Ndr2 is critically involved in the initiation of TCR-mediated LFA-1 activation (open conformation) in T cells. Ndr2 itself becomes activated upon TCR stimulation and phosphorylates the intracellular integrin binding partner Filamin A (FLNa) at serine 2152. This phosphorylation promotes the dissociation of FLNa from LFA-1, allowing for a subsequent association of Talin and Kindlin-3 which both stabilize the open conformation of LFA-1. Our data suggest that Ndr2 activation is a crucial step to initiate TCR-mediated LFA-1 activation in T cells.
  • Characterization of mice with a platelet-specific deletion of the adapter molecule ADAP.

    Rudolph, Jochen Michael; Guttek, Karina; Weitz, Gabriele; Meinke, Clara Antonia; Kliche, Stefanie; Reinhold, Dirk; Schraven, Burkhart; Reinhold, Annegret; HZI, Helmholtz Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig Germany. (ASM, 2019-03-04)
    The adhesion and degranulation-promoting adapter protein (ADAP) is expressed in T cells, NK cells, myeloid cells, and platelets. The involvement of ADAP in the regulation of receptor-mediated inside-out signaling leading to integrin activation is well characterized, especially in T cells and in platelets. Due to the fact that animal studies using conventional knock-out mice are limited by the overlapping effects of the different ADAP-expressing cells, we generated conditional ADAP knock-out mice (ADAPfl/fl PF4-Cretg). We observed that loss of ADAP restricted to the megakaryocytic lineage has no impact on other hematopoietic cells even after stimulation conditions. ADAPfl/fl PF4-Cretg mice showed thrombocytopenia in combination with reduced plasma levels of PF4 and TGF-β1. In vitro, platelets from these mice revealed reduced P-selectin expression, lower TGF-β1 release, diminished integrin αIIbβ3 activation and decreased fibrinogen binding after stimulation with podoplanin, the ligand of the C-type lectin-like receptor-2 (CLEC-2). Furthermore, loss of ADAP was associated with impaired CLEC-2-mediated activation of PLCγ2 and Erk1/2. Induction of experimental autoimmune encephalomyelitis (EAE) in mice lacking ADAP expression in platelets caused a more severe disease. In vivo administration of TGF-β1 early after T cell transfer improved EAE severity in mice with loss of ADAP restricted to platelets. Our results reveal a regulatory function of ADAP in platelets in vitro and during autoimmune disease EAE in vivo.
  • Memantine potentiates cytarabine-induced cell death of acute leukemia correlating with inhibition of K1.3 potassium channels, AKT and ERK1/2 signaling.

    Lowinus, Theresa; Heidel, Florian H; Bose, Tanima; Nimmagadda, Subbaiah Chary; Schnöder, Tina; Cammann, Clemens; Schmitz, Ingo; Seifert, Ulrike; Fischer, Thomas; Schraven, Burkhart; et al. (BMC, 2019-01-16)
    Treatment of acute leukemia is challenging and long-lasting remissions are difficult to induce. Innovative therapy approaches aim to complement standard chemotherapy to improve drug efficacy and decrease toxicity. Promising new therapeutic targets in cancer therapy include voltage-gated K We analyzed acute lymphoid (Jurkat, CEM) and myeloid (HL-60, Molm-13, OCI-AML-3) leukemia cell lines and patients' acute leukemic blasts after treatment with either drug alone or the combination of cytarabine and memantine. Patch-clamp analysis was performed to evaluate inhibition of K Our study demonstrates that memantine inhibits K Our study underlines inhibition of K
  • CD11c-expressing Ly6C+CCR2+ monocytes constitute a reservoir for efficient Leishmania proliferation and cell-to-cell transmission.

    Heyde, Sandrina; Philipsen, Lars; Formaglio, Pauline; Fu, Yan; Baars, Iris; Höbbel, Guido; Kleinholz, Corinna L; Seiß, Elena A; Stettin, Juliane; Gintschel, Patricia; et al. (2018-10-01)
    The virulence of intracellular pathogens such as Leishmania major (L. major) relies largely on their ability to undergo cycles of replication within phagocytes, release, and uptake into new host cells. While all these steps are critical for successful establishment of infection, neither the cellular niche of efficient proliferation, nor the spread to new host cells have been characterized in vivo. Here, using a biosensor for measuring pathogen proliferation in the living tissue, we found that monocyte-derived Ly6C+CCR2+ phagocytes expressing CD11c constituted the main cell type harboring rapidly proliferating L. major in the ongoing infection. Synchronization of host cell recruitment and intravital 2-photon imaging showed that these high proliferating parasites preferentially underwent cell-to-cell spread. However, newly recruited host cells were infected irrespectively of their cell type or maturation state. We propose that among these cells, CD11c-expressing monocytes are most permissive for pathogen proliferation, and thus mainly fuel the cycle of intracellular proliferation and cell-to-cell transfer during the acute infection. Thus, besides the well-described function for priming and activating T cell effector functions against L. major, CD11c-expressing monocyte-derived cells provide a reservoir for rapidly proliferating parasites that disseminate at the site of infection.
  • CD11c-expressing Ly6C+CCR2+ monocytes constitute a reservoir for efficient Leishmania proliferation and cell-to-cell transmission.

    Heyde, Sandrina; Philipsen, Lars; Formaglio, Pauline; Fu, Yan; Baars, Iris; Höbbel, Guido; Kleinholz, Corinna L; Seiß, Elena A; Stettin, Juliane; Gintschel, Patricia; et al. (2018-10-01)
    The virulence of intracellular pathogens such as Leishmania major (L. major) relies largely on their ability to undergo cycles of replication within phagocytes, release, and uptake into new host cells. While all these steps are critical for successful establishment of infection, neither the cellular niche of efficient proliferation, nor the spread to new host cells have been characterized in vivo. Here, using a biosensor for measuring pathogen proliferation in the living tissue, we found that monocyte-derived Ly6C+CCR2+ phagocytes expressing CD11c constituted the main cell type harboring rapidly proliferating L. major in the ongoing infection. Synchronization of host cell recruitment and intravital 2-photon imaging showed that these high proliferating parasites preferentially underwent cell-to-cell spread. However, newly recruited host cells were infected irrespectively of their cell type or maturation state. We propose that among these cells, CD11c-expressing monocytes are most permissive for pathogen proliferation, and thus mainly fuel the cycle of intracellular proliferation and cell-to-cell transfer during the acute infection. Thus, besides the well-described function for priming and activating T cell effector functions against L. major, CD11c-expressing monocyte-derived cells provide a reservoir for rapidly proliferating parasites that disseminate at the site of infection.
  • JAK2-V617F promotes venous thrombosis through β1/β2 integrin activation.

    Edelmann, Bärbel; Gupta, Nibedita; Schnoeder, Tina M; Oelschlegel, Anja M; Shahzad, Khurrum; Goldschmidt, Jürgen; Philipsen, Lars; Weinert, Soenke; Ghosh, Aniket; Saalfeld, Felix C; et al. (2018-10-01)
    JAK2-V617F-positive chronic myeloproliferative neoplasia (CMN) commonly displays dysfunction of integrins and adhesion molecules expressed on platelets, erythrocytes, and leukocytes. However, the mechanism by which the 2 major leukocyte integrin chains, β1 and β2, may contribute to CMN pathophysiology remained unclear. β1 (α4β1; VLA-4) and β2 (αLβ2; LFA-1) integrins are essential regulators for attachment of leukocytes to endothelial cells. We here showed enhanced adhesion of granulocytes from mice with JAK2-V617F knockin (JAK2+/VF mice) to vascular cell adhesion molecule 1- (VCAM1-) and intercellular adhesion molecule 1-coated (ICAM1-coated) surfaces. Soluble VCAM1 and ICAM1 ligand binding assays revealed increased affinity of β1 and β2 integrins for their respective ligands. For β1 integrins, this correlated with a structural change from the low- to the high-affinity conformation induced by JAK2-V617F. JAK2-V617F triggered constitutive activation of the integrin inside-out signaling molecule Rap1, resulting in translocation toward the cell membrane. Employing a venous thrombosis model, we demonstrated that neutralizing anti-VLA-4 and anti-β2 integrin antibodies suppress pathologic thrombosis as observed in JAK2+/VF mice. In addition, aberrant homing of JAK2+/VF leukocytes to the spleen was inhibited by neutralizing anti-β2 antibodies and by pharmacologic inhibition of Rap1. Thus, our findings identified cross-talk between JAK2-V617F and integrin activation promoting pathologic thrombosis and abnormal trafficking of leukocytes to the spleen.
  • Beware the intruder: Real time observation of infiltrated neutrophils and neutrophil-Microglia interaction during stroke in vivo.

    Neumann, Jens; Henneberg, Sophie; von Kenne, Susanne; Nolte, Niklas; Müller, Andreas J; Schraven, Burkhart; Görtler, Michael W; Reymann, Klaus G; Gunzer, Matthias; Riek-Burchardt, Monika; et al. (2018)
    Inflammation plays an important role in the pathogenesis of ischemic stroke including an acute and prolonged inflammatory process. The role of neutrophil granulocytes as first driver of the immune reaction from the blood site is under debate due to controversial findings. In bone marrow chimeric mice we were able to study the dynamics of tdTomato-expressing neutrophils and GFP-expressing microglia after photothrombosis using intravital two-photon microscopy. We demonstrate the infiltration of neutrophils into the brain parenchyma and confirm a long-lasting contact between neutrophils and microglia as well as an uptake of neutrophils by microglia clearing the brain from peripheral immune cells.

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