Recent Submissions

  • Demarcated thresholds of tumor-specific CD8 T cells elicited by MCMV-based vaccine vectors provide robust correlates of protection.

    Beyranvand Nejad, Elham; Ratts, Robert B; Panagioti, Eleni; Meyer, Christine; Oduro, Jennifer D; Cicin-Sain, Luka; Früh, Klaus; van der Burg, Sjoerd H; Arens, Ramon; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (BMC, 2019-01-31)
    The capacity of cytomegalovirus (CMV) to elicit long-lasting strong T cell responses, and the ability to engineer the genome of this DNA virus positions CMV-based vaccine vectors highly suitable as a cancer vaccine platform. Defined immune thresholds for tumor protection and the factors affecting such thresholds have not well been investigated in cancer immunotherapy. We here determined using CMV as a vaccine platform whether critical thresholds of vaccine-specific T cell responses can be established that relate to tumor protection, and which factors control such thresholds. We generated CMV-based vaccine vectors expressing the E7 epitope and tested these in preclinical models of HPV16-induced cancer. Vaccination was applied via different doses and routes (intraperitoneal (IP), subcutaneous (SC) and intranasal (IN)). The magnitude, kinetics and phenotype of the circulating tumor-specific CD8 Immunization with CMV-based vaccines via the IP or SC route eliciting vaccine-induced CD8 This study highlight the effectiveness of CMV-based vaccine vectors, and shows that demarcated thresholds of vaccine-specific T cells could be defined that correlate to tumor protection. Together, these results may hold importance for cancer vaccine development to achieve high efficacy in vaccine recipients.
  • An endothelial cell line infected by Kaposi's sarcoma-associated herpes virus (KSHV) allows the investigation of Kaposi's sarcoma and the validation of novel viral inhibitors in vitro and in vivo.

    Dubich, Tatyana; Lieske, Anna; Santag, Susann; Beauclair, Guillaume; Rückert, Jessica; Herrmann, Jennifer; Gorges, Jan; Büsche, Guntram; Kazmaier, Uli; Hauser, Hansjörg; Stadler, Marc; Schulz, Thomas F; Wirth, Dagmar; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (2019-01-04)
    Kaposi's sarcoma-associated herpesvirus (KSHV) is the etiological agent of Kaposi's sarcoma (KS), a tumor of endothelial origin predominantly affecting immunosuppressed individuals. Up to date, vaccines and targeted therapies are not available. Screening and identification of anti-viral compounds are compromised by the lack of scalable cell culture systems reflecting properties of virus-transformed cells in patients. Further, the strict specificity of the virus for humans limits the development of in vivo models. In this study, we exploited a conditionally immortalized human endothelial cell line for establishment of in vitro 2D and 3D KSHV latency models and the generation of KS-like xenograft tumors in mice. Importantly, the invasive properties and tumor formation could be completely reverted by purging KSHV from the cells, confirming that tumor formation is dependent on the continued presence of KSHV, rather than being a consequence of irreversible transformation of the infected cells. Upon testing a library of 260 natural metabolites, we selected the compounds that induced viral loss or reduced the invasiveness of infected cells in 2D and 3D endothelial cell culture systems. The efficacy of selected compounds against KSHV-induced tumor formation was verified in the xenograft model. Together, this study shows that the combined use of anti-viral and anti-tumor assays based on the same cell line is predictive for tumor reduction in vivo and therefore allows faithful selection of novel drug candidates against Kaposi's sarcoma. KEY MESSAGES: Novel 2D, 3D, and xenograft mouse models mimic the consequences of KSHV infection. KSHV-induced tumorigenesis can be reverted upon purging the cells from the virus. A 3D invasiveness assay is predictive for tumor reduction in vivo. Chondramid B, epothilone B, and pretubulysin D diminish KS-like lesions in vivo.
  • Signatures of T and B Cell Development, Functional Responses and PD-1 Upregulation After HCMV Latent Infections and Reactivations in Nod.Rag.Gamma Mice Humanized With Cord Blood CD34 Cells.

    Theobald, Sebastian J; Khailaie, Sahamoddin; Meyer-Hermann, Michael; Volk, Valery; Olbrich, Henning; Danisch, Simon; Gerasch, Laura; Schneider, Andreas; Sinzger, Christian; Schaudien, Dirk; Lienenklaus, Stefan; Riese, Peggy; Guzman, Carlos A; Figueiredo, Constanca; von Kaisenberg, Constantin; Spineli, Loukia M; Glaesener, Stephanie; Meyer-Bahlburg, Almut; Ganser, Arnold; Schmitt, Michael; Mach, Michael; Messerle, Martin; Stripecke, Renata; BRICS, Braunschweiger Zentrum für Systembiologie, Rebenring 56,38106 Braunschweig, Germany.; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (Frontiers, 2018-01-01)
    uman cytomegalovirus (HCMV) latency is typically harmless but reactivation can be largely detrimental to immune compromised hosts. We modeled latency and reactivation using a traceable HCMV laboratory strain expressing the Gaussia luciferase reporter gene (HCMV/GLuc) in order to interrogate the viral modulatory effects on the human adaptive immunity. Humanized mice with long-term (more than 17 weeks) steady human T and B cell immune reconstitutions were infected with HCMV/GLuc and 7 weeks later were further treated with granulocyte-colony stimulating factor (G-CSF) to induce viral reactivations. Whole body bio-luminescence imaging analyses clearly differentiated mice with latent viral infections vs. reactivations. Foci of vigorous viral reactivations were detectable in liver, lymph nodes and salivary glands. The number of viral genome copies in various tissues increased upon reactivations and were detectable in sorted human CD14+, CD169+, and CD34+ cells. Compared with non-infected controls, mice after infections and reactivations showed higher thymopoiesis, systemic expansion of Th, CTL, Treg, and Tfh cells and functional antiviral T cell responses. Latent infections promoted vast development of memory CD4+ T cells while reactivations triggered a shift toward effector T cells expressing PD-1. Further, reactivations prompted a marked development of B cells, maturation of IgG+ plasma cells, and HCMV-specific antibody responses. Multivariate statistical methods were employed using T and B cell immune phenotypic profiles obtained with cells from several tissues of individual mice. The data was used to identify combinations of markers that could predict an HCMV infection vs. reactivation status. In spleen, but not in lymph nodes, higher frequencies of effector CD4+ T cells expressing PD-1 were among the factors most suited to distinguish HCMV reactivations from infections. These results suggest a shift from a T cell dominated immune response during latent infections toward an exhausted T cell phenotype and active humoral immune response upon reactivations. In sum, this novel in vivo humanized model combined with advanced analyses highlights a dynamic system clearly specifying the immunological spatial signatures of HCMV latency and reactivations. These signatures can be merged as predictive biomarker clusters that can be applied in the clinical translation of new therapies for the control of HCMV reactivation.
  • Cytochalasans Act as Inhibitors of Biofilm Formation of Staphylococcus Aureus.

    Yuyama, Kamila Tomoko; Wendt, Lucile; Surup, Frank; Kretz, Robin; Chepkirui, Clara; Wittstein, Kathrin; Boonlarppradab, Chollaratt; Wongkanoun, Sarunyou; Luangsa-Ard, Jennifer; Stadler, Marc; Abraham, Wolf-Rainer; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (MDPI, 2018-10-30)
    During the course of our ongoing work to discover new inhibitors of biofilm formation of Staphylococcus aureus from fungal sources, we observed biofilm inhibition by cytochalasans isolated from cultures of the ascomycete Hypoxylon fragiforme for the first time. Two new compounds were purified by a bioassay-guided fractionation procedure; their structures were elucidated subsequently by nuclear magnetic resonance (NMR) spectroscopy and high-resolution mass spectrometry (HR-MS). This unexpected finding prompted us to test further cytochalasans from other fungi and from commercial sources for comparison. Out of 21 cytochalasans, 13 showed significant inhibition of Staphylococcus aureus biofilm formation at subtoxic levels. These findings indicate the potential of cytochalasans as biofilm inhibitors for the first time, also because the minimum inhibitory concentrations (MIC) are independent of the anti-biofilm activities. However, cytochalasans are known to be inhibitors of actin, making some of them very toxic for eukaryotic cells. Since the chemical structures of the tested compounds were rather diverse, the inclusion of additional derivatives, as well as the evaluation of their selectivity against mammalian cells vs. the bacterium, will be necessary as next step in order to develop structure-activity relationships and identify the optimal candidates for development of an anti-biofilm agent. View Full-Text
  • Advances and Challenges of Biodegradable Implant Materials with a Focus on Magnesium-Alloys and Bacterial Infections

    Rahim, Muhammad; Ullah, Sami; Mueller, Peter; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (MDPI, 2018-07-10)
    Medical implants made of biodegradable materials could be advantageous for temporary applications, such as mechanical support during bone-healing or as vascular stents to keep blood vessels open. After completion of the healing process, the implant would disappear, avoiding long-term side effects or the need for surgical removal. Various corrodible metal alloys based on magnesium, iron or zinc have been proposed as sturdier and potentially less inflammatory alternatives to degradable organic polymers, in particular for load-bearing applications. Despite the recent introduction of magnesium-based screws, the remaining hurdles to routine clinical applications are still challenging. These include limitations such as mechanical material characteristics or unsuitable corrosion characteristics. In this article, the salient features and clinical prospects of currently-investigated biodegradable implant materials are summarized, with a main focus on magnesium alloys. A mechanism of action for the stimulation of bone growth due to the exertion of mechanical force by magnesium corrosion products is discussed. To explain divergent in vitro and in vivo effects of magnesium, a novel model for bacterial biofilm infections is proposed which predicts crucial consequences for antibacterial implant strategies.
  • Macrophage entrapped silica coated superparamagnetic iron oxide particles for controlled drug release in a 3D cancer model.

    Ullah, Sami; Seidel, Katja; Türkkan, Sibel; Warwas, Dawid Peter; Dubich, Tatyana; Rohde, Manfred; Hauser, Hansjörg; Behrens, Peter; Kirschning, Andreas; Köster, Mario; Wirth, Dagmar; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (2018-12-23)
    Targeted delivery of drugs is a major challenge in treatment of diverse diseases. Systemically administered drugs demand high doses and are accompanied by poor selectivity and side effects on non-target cells. Here, we introduce a new principle for targeted drug delivery. It is based on macrophages as transporters for nanoparticle-coupled drugs as well as controlled release of drugs by hyperthermia mediated disruption of the cargo cells and simultaneous deliberation of nanoparticle-linked drugs. Hyperthermia is induced by an alternating electromagnetic field (AMF) that induces heat from silica-coated superparamagnetic iron oxide nanoparticles (SPIONs). We show proof-of-principle of controlled release by the simultaneous disruption of the cargo cells and the controlled, AMF induced release of a toxin, which was covalently linked to silica-coated SPIONs via a thermo-sensitive linker. Cells that had not been loaded with SPIONs remain unaffected. Moreover, in a 3D co-culture model we demonstrate specific killing of associated tumour cells when employing a ratio as low as 1:40 (SPION-loaded macrophage: tumour cells). Overall, our results demonstrate that AMF induced drug release from macrophage-entrapped nanoparticles is tightly controlled and may be an attractive novel strategy for targeted drug release.
  • Influenza-Activated ILC1s Contribute to Antiviral Immunity Partially Influenced by Differential GITR Expression.

    Vashist, Neha; Trittel, Stephanie; Ebensen, Thomas; Chambers, Benedict J; Guzmán, Carlos A; Riese, Peggy; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (Frontiers, 2018-01-01)
    Innate lymphoid cells (ILCs) represent diversified subsets of effector cells as well as immune regulators of mucosal immunity and are classified into group 1 ILCs, group 2 ILCs, and group 3 ILCs. Group 1 ILCs encompass natural killer (NK) cells and non-NK ILCs (ILC1s) and mediate their functionality via the rapid production of IFN-γ and TNF-α. The current knowledge of ILC1s mainly associates them to inflammatory processes. Much less is known about their regulation during infection and their capacity to interact with cells of the adaptive immune system. The present study dissected the role of ILC1s during early influenza A virus infection, thereby revealing their impact on the antiviral response. Exploiting in vitro and in vivo H1N1 infection systems, a cross-talk of ILC1s with cells of the innate and the adaptive immunity was demonstrated, which contributes to anti-influenza immunity. A novel association of ILC1 functionality and the expression of the glucocorticoid-induced TNFR-related protein (GITR) was observed, which hints toward a so far undescribed role of GITR in regulating ILC1 responsiveness. Overexpression of GITR inhibits IFN-γ production by ILC1s, whereas partial reduction of GITR expression can reverse this effect, thereby regulating ILC1 functionality. These new insights into ILC1 biology define potential intervention targets to modulate the functional properties of ILC1s, thus contributing toward the development of new immune interventions against influenza.
  • De Novo Fatty Acid Synthesis During Mycobacterial Infection Is a Prerequisite for the Function of Highly Proliferative T Cells, But Not for Dendritic Cells or Macrophages.

    Stüve, Philipp; Minarrieta, Lucía; Erdmann, Hanna; Arnold-Schrauf, Catharina; Swallow, Maxine; Guderian, Melanie; Krull, Freyja; Hölscher, Alexandra; Ghorbani, Peyman; Behrends, Jochen; Abraham, Wolf-Rainer; Hölscher, Christoph; Sparwasser, Tim D; Berod, Luciana; TWINCORE, Zentrum für experimentelle und klinische Infektionsforschung GmbH,Feodor-Lynen Str. 7, 30625 Hannover, Germany. (Frontiers, 2018-01-01)
    Mycobacterium tuberculosis (Mtb), the causative agent of human tuberculosis, is able to efficiently manipulate the host immune system establishing chronic infection, yet the underlying mechanisms of immune evasion are not fully understood. Evidence suggests that this pathogen interferes with host cell lipid metabolism to ensure its persistence. Fatty acid metabolism is regulated by acetyl-CoA carboxylase (ACC) 1 and 2; both isoforms catalyze the conversion of acetyl-CoA into malonyl-CoA, but have distinct roles. ACC1 is located in the cytosol, where it regulates de novo fatty acid synthesis (FAS), while ACC2 is associated with the outer mitochondrial membrane, regulating fatty acid oxidation (FAO). In macrophages, mycobacteria induce metabolic changes that lead to the cytosolic accumulation of lipids. This reprogramming impairs macrophage activation and contributes to chronic infection. In dendritic cells (DCs), FAS has been suggested to underlie optimal cytokine production and antigen presentation, but little is known about the metabolic changes occurring in DCs upon mycobacterial infection and how they affect the outcome of the immune response. We therefore determined the role of fatty acid metabolism in myeloid cells and T cells during Mycobacterium bovis BCG or Mtb infection, using novel genetic mouse models that allow cell-specific deletion of ACC1 and ACC2 in DCs, macrophages, or T cells. Our results demonstrate that de novo FAS is induced in DCs and macrophages upon M. bovis BCG infection. However, ACC1 expression in DCs and macrophages is not required to control mycobacteria. Similarly, absence of ACC2 did not influence the ability of DCs and macrophages to cope with infection. Furthermore, deletion of ACC1 in DCs or macrophages had no effect on systemic pro-inflammatory cytokine production or T cell priming, suggesting that FAS is dispensable for an intact innate response against mycobacteria. In contrast, mice with a deletion of ACC1 specifically in T cells fail to generate efficient T helper 1 responses and succumb early to Mtb infection. In summary, our results reveal ACC1-dependent FAS as a crucial mechanism in T cells, but not DCs or macrophages, to fight against mycobacterial infection.
  • Pseudomonas-Specific NGS Assay Provides Insight Into Abundance and Dynamics of Species Including P. aeruginosa in a Cooling Tower.

    Pereira, Rui P A; Peplies, Jörg; Mushi, Douglas; Brettar, Ingrid; Höfle, Manfred G; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (Frontiers, 2018-01-01)
    Pseudomonas species are frequent inhabitants of freshwater environments and colonizers of water supply networks via bioadhesion and biofilm formation. P. aeruginosa is the species most commonly associated with human disease, causing a wide variety of infections with links to its presence in freshwater systems. Though several other Pseudomonas species are of ecological and public health importance, little knowledge exists regarding environmental abundances of these species. In the present study, an Illumina-based next-generation sequencing (NGS) approach using Pseudomonas-specific primers targeting the 16S rRNA gene was evaluated and applied to a set of freshwater samples from different environments including a cooling tower sampled monthly during 2 years. Our approach showed high in situ specificity and accuracy. NGS read counts revealed a precise quantification of P. aeruginosa and a good correlation with the absolute number of Pseudomonas genome copies in a validated genus-specific qPCR assay, demonstrating the ability of the NGS approach to determine both relative and absolute abundances of Pseudomonas species and P. aeruginosa. The characterization of Pseudomonas communities in cooling tower water allowed us to identify 43 phylotypes, with P. aeruginosa being the most abundant. A shift existed within each year from a community dominated by phylotypes belonging to P. fluorescens and P. oleovorans phylogenetic groups to a community where P. aeruginosa was highly abundant. Co-occurrence was observed between P. aeruginosa and other phylotypes of P. aeruginosa group as well as the potentially pathogenic species P. stutzeri, but not with phylotypes of the P. fluorescens group, indicating the need to further investigate the metabolic networks and ecological traits of Pseudomonas species. This study demonstrates the potential of deep sequencing as a valuable tool in environmental diagnostics and surveillance of health-related pathogens in freshwater environments
  • Human monocyte-derived macrophages inhibit HCMV spread independent of classical antiviral cytokines.

    Becker, Jennifer; Kinast, Volker; Döring, Marius; Lipps, Christoph; Duran, Veronica; Spanier, Julia; Tegtmeyer, Pia-Katharina; Wirth, Dagmar; Cicin-Sain, Luka; Alcamí, Antonio; Kalinke, Ulrich; TWINCORE, Zentrum für experimentelle und klinische Infektionsforschung GmbH,Feodor-Lynen Str. 7, 30625 Hannover, Germany.; HZI, Helmholtz Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig. (2018-01-01)
    Infection of healthy individuals with human cytomegalovirus (HCMV) is usually unnoticed and results in life-long latency, whereas HCMV reactivation as well as infection of newborns or immunocompromised patients can cause life-threatening disease. To better understand HCMV pathogenesis we studied mechanisms that restrict HCMV spread. We discovered that HCMV-infected cells can directly trigger plasmacytoid dendritic cells (pDC) to mount antiviral type I interferon (IFN-I) responses, even in the absence of cell-free virus. In contrast, monocyte-derived cells only expressed IFN-I when stimulated by cell-free HCMV, or upon encounter of HCMV-infected cells that already produced cell-free virus. Nevertheless, also in the absence of cell-free virus, i.e., upon co-culture of infected epithelial/endothelial cells and monocyte-derived macrophages (moMΦ) or dendritic cells (moDC), antiviral responses were induced that limited HCMV spread. The induction of this antiviral effect was dependent on cell-cell contact, whereas cell-free supernatants from co-culture experiments also inhibited virus spread, implying that soluble factors were critically needed. Interestingly, the antiviral effect was independent of IFN-γ, TNF-α, and IFN-I as indicated by cytokine inhibition experiments using neutralizing antibodies or the vaccinia virus-derived soluble IFN-I binding protein B18R, which traps human IFN-α and IFN-β. In conclusion, our results indicate that human macrophages and dendritic cells can limit HCMV spread by IFN-I dependent as well as independent mechanisms, whereas the latter ones might be particularly relevant for the restriction of HCMV transmission via cell-to-cell spread.
  • Rapid In Vivo Assessment of Adjuvant's Cytotoxic T Lymphocytes Generation Capabilities for Vaccine Development

    Lirussi, Darío; Ebensen, Thomas; Schulze, Kai; Reinhard, Elena; Trittel, Stephanie; Riese, Peggy; Prochnow, Blair; Guzmán, Carlos A.; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany.
    The assessment of modern sub-unit vaccines reveals that the generation of neutralizing antibodies is important but not sufficient for adjuvant selection. Therefore, adjuvants with both humoral and cellular immuno-stimulatory capabilities that are able to promote cytotoxic T lymphocytes (CTL) responses are urgently needed. Thus, faithful monitoring of adjuvant candidates that induce cross-priming and subsequently enhance CTL generation represents a crucial step in vaccine development. In here we present an application for a method that uses SIINFEKL-specific (OT-I) T cells to monitor the cross-presentation of the model antigen ovalbumin (OVA) in vivo in the presence of different adjuvant candidates. This method represents a rapid test to select adjuvants with the best cross-priming capabilities. The proliferation of CD8+ T cells is the most valuable indication of cross-priming and it is also regarded as a correlate of adjuvant-induced cross-presentation. This feature can be evaluated in different immune organs like lymph nodes and spleen. The extent of the CTL generation can also be monitored, thereby giving insights on the nature of a local (draining lymph node mainly) or a systemic response (distant lymph nodes and/or spleen). This technique further allows multiple modifications for testing drugs that can inhibit specific cross-presentation pathways and also offers the possibility to be used in different strains of conventional and genetically modified mice. In summary, the application that we present here will be useful for vaccine laboratories in industry or academia that develop or modify chemical adjuvants for vaccine research and development. © 2018, Journal of Visualized Experiments.
  • Biocompatible Coatings from Smart Biopolymer Nanoparticles for Enzymatically Induced Drug Release.

    Tolle, Christian; Riedel, Jan; Mikolai, Carina; Winkel, Andreas; Stiesch, Meike; Wirth, Dagmar; Menzel, Henning; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (2018-09-28)
    Nanoparticles can be used as a smart drug delivery system, when they release the drug only upon degradation by specific enzymes. A method to create such responsive materials is the formation of hydrogel nanoparticles, which have enzymatically degradable crosslinkers. Such hydrogel nanoparticles were prepared by ionotropic gelation sodium alginate with lysine-rich peptide sequences-either α-poly-L-lysine (PLL) or the aggrecanase-labile sequence KKKK-GRD-ARGSV↓NITEGE-DRG-KKKK. The nanoparticle suspensions obtained were analyzed by means of dynamic light scattering and nanoparticle tracking analysis. Degradation experiments carried out with the nanoparticles in suspension revealed enzyme-induced lability. Drugs present in the polymer solution during the ionotropic gelation can be encapsulated in the nanoparticles. Drug loading was investigated for interferon-β (IFN-β) as a model, using a bioluminescence assay with MX2Luc2 cells. The encapsulation efficiency for IFN-β was found to be approximately 25%. The nanoparticles suspension can be used to spray-coat titanium alloys (Ti-6Al-4V) as a common implant material. The coatings were proven by ellipsometry, reflection-absorption infrared spectroscopy, and X-ray photoelectron spectroscopy. An enzyme-responsive decrease in layer thickness is observed due to the degradation of the coatings. The Alg/peptide coatings were cytocompatible for human gingival fibroblasts (HGFIB), which was investigated by CellTiterBlue and lactate dehydrogenase (LDH) assay. However, HGFIBs showed poor adhesion and proliferation on the Alg/peptide coatings, but these could be improved by modification of the alginate with a RGD-peptide sequence. The smart drug release system presented can be further tailored to have the right release kinetics and cell adhesion properties.
  • Identification of a Predominantly Interferon-λ-Induced Transcriptional Profile in Murine Intestinal Epithelial Cells.

    Selvakumar, Tharini A; Bhushal, Sudeep; Kalinke, Ulrich; Wirth, Dagmar; Hauser, Hansjörg; Köster, Mario; Hornef, Mathias W; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2017-01-01)
    Type I (α and β) and type III (λ) interferons (IFNs) induce the expression of a large set of antiviral effector molecules
  • Large-scale production of megakaryocytes in microcarrier-supported stirred suspension bioreactors.

    Eicke, Dorothee; Baigger, Anja; Schulze, Kai; Latham, Sharissa L; Halloin, Caroline; Zweigerdt, Robert; Guzman, Carlos A; Blasczyk, Rainer; Figueiredo, Constança; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2018-07-05)
    Megakaryocytes (MKs) are the precursors of platelets (PLTs) and may be used for PLT production in vivo or in vitro, as well as a source for PLT-derived growth factors. Induced pluripotent stem cells represent an unlimited cell source for the in vitro production of MKs. This study aimed at developing an effective, xeno-free and scalable system to produce high numbers of MKs. In particular, microcarrier beads-assisted stirred bioreactors were evaluated as a means of improving MK yields. This method resulted in the production of 18.7 × 10
  • Type I interferon receptor signaling delays Kupffer cell replenishment during acute fulminant viral hepatitis.

    Borst, Katharina; Frenz, Theresa; Spanier, Julia; Tegtmeyer, Pia-Katharina; Chhatbar, Chintan; Skerra, Jennifer; Ghita, Luca; Namineni, Sukumar; Lienenklaus, Stefan; Köster, Mario; Heikenwaelder, Mathias; Sutter, Gerd; Kalinke, Ulrich; TWINCORE, Zentrum für experimentelle und klinischeInfektionsforschung GmbH, Feodor-Lynen-Str. 7, 30625 Hannover, Germany. (2017-12-21)
    Virus-induced fulminant hepatitis is a major cause of acute liver failure. During acute viral hepatitis the impact of type I interferon (IFN-I) on myeloid cells, including liver-resident Kupffer cells (KC), is only partially understood. Herein, we dissected the impact of locally induced IFN-I responses on myeloid cell function and hepatocytes during acute liver inflammation. Two different DNA-encoded viruses, vaccinia virus (VACV) and murine cytomegalovirus (MCMV), were studied. In vivo imaging was applied to visualize local IFN-β induction and IFN-I receptor (IFNAR) triggering in VACV-infected reporter mice. Furthermore, mice with a cell type-selective IFNAR ablation were analyzed to dissect the role of IFNAR signaling in myeloid cells and hepatocytes. Experiments with Cx3cr1 VACV infection induced local IFN-β responses, which lead to IFNAR signaling primarily within the liver. IFNAR triggering was needed to control the infection and prevent fulminant hepatitis. The severity of liver inflammation was independent of IFNAR triggering of hepatocytes, whereas IFNAR triggering of myeloid cells protected from excessive inflammation. Upon VACV or MCMV infection KC disappeared, whereas infiltrating monocytes differentiated to KC afterwards. During IFNAR triggering such replenished monocyte-derived KC comprised more IFNAR-deficient than -competent cells in mixed bone marrow chimeric mice, whereas after the decline of IFNAR triggering both subsets showed an even distribution. Upon VACV infection IFNAR triggering of myeloid cells, but not of hepatocytes, critically modulates acute viral hepatitis. During infection with DNA-encoded viruses IFNAR triggering of liver-infiltrating blood monocytes delays the development of monocyte-derived KC, pointing towards new therapeutic strategies for acute viral hepatitis.
  • Induction of CD4(+) and CD8(+) anti-tumor effector T cell responses by bacteria mediated tumor therapy.

    Stern, Christian; Kasnitz, Nadine; Kocijancic, Dino; Trittel, Stephanie; Riese, Peggy; Guzman, Carlos A; Leschner, Sara; Weiss, Siegfried; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2015-10-15)
    Facultative anaerobic bacteria like E. coli can colonize solid tumors often resulting in tumor growth retardation or even clearance. Little mechanistic knowledge is available for this phenomenon which is however crucial for optimization and further implementation in the clinic. Here, we show that intravenous injections with E. coli TOP10 can induce clearance of CT26 tumors in BALB/c mice. Importantly, re-challenging mice which had cleared tumors showed that clearance was due to a specific immune reaction. Accordingly, lymphopenic mice never showed tumor clearance after infection. Depletion experiments revealed that during induction phase, CD8(+) T cells are the sole effectors responsible for tumor clearance while in the memory phase CD8(+) and CD4(+) T cells were involved. This was confirmed by adoptive transfer. CD4(+) and CD8(+) T cells could reject newly set tumors while CD8(+) T cells could even reject established tumors. Detailed analysis of adoptively transferred CD4(+) T cells during tumor challenge revealed expression of granzyme B, FasL, TNF-α and IFN-γ in such T cells that might be involved in the anti-tumor activity. Our findings should pave the way for further optimization steps of this promising therapy.
  • Self-Amplifying Replicon RNA Delivery to Dendritic Cells by Cationic Lipids

    Englezou, Pavlos C.; Sapet, Cedric; Démoulins, Thomas; Milona, Panagiota; Ebensen, Thomas; Schulze, Kai; Guzman, Carlos-Alberto; Poulhes, Florent; Zelphati, Olivier; Ruggli, Nicolas; McCullough, Kenneth C.; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany.
  • Memory CD8 T cells support the maintenance of hematopoietic stem cells in the bone marrow

    Geerman, Sulima; Brasser, Giso; Bhushal, Sudeep; Salerno, Fiamma; Kragten, Natasja A.; Hoogenboezem, Mark; de Haan, Gerald; Wolkers, Monika C.; Pascutti, María Fernanda; Nolte, Martijn A.; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany.
  • The chromatin remodeling factor SPOC1 acts as a cellular restriction factor against human cytomegalovirus by repressing the major immediate-early promoter.

    Reichel, Anna; Stilp, Anne-Charlotte; Scherer, Myriam; Reuter, Nina; Lukassen, Sören; Kasmapour, Bahram; Schreiner, Sabrina; Cicin-Sain, Luka; Winterpacht, Andreas; Stamminger, Thomas; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2018-05-09)
    The cellular protein SPOC1 (survival time-associated PHD finger protein in ovarian cancer 1) acts as a regulator of chromatin structure and DNA damage response. It binds H3K4me2/3 containing chromatin and promotes DNA condensation by recruiting corepressors such as KAP-1 and H3K9 methyltransferases. Previous studies identified SPOC1 as a restriction factor against human adenovirus (HAdV) infection that is antagonized by E1B-55K/E4orf6-dependent proteasomal degradation. Here, we demonstrate that, in contrast to HAdV-infected cells, SPOC1 is transiently upregulated during the early phase of HCMV replication. We show that expression of the immediate-early protein 1 (IE1) is sufficient and necessary to induce SPOC1. Additionally, we discovered that during later stages of infection SPOC1 is downregulated in a GSK-3β-dependent manner. We provide evidence that SPOC1 overexpression severely impairs HCMV replication by repressing the initiation of viral immediate early (IE) gene expression. Consistently, we observed that SPOC1-depleted primary human fibroblasts displayed augmented initiation of viral IE gene expression. This occurs in a MOI-dependent manner, a defining hallmark of intrinsic immunity. Interestingly, repression requires the presence of high SPOC1 levels at the start of infection while a later upregulation had no negative impact suggesting distinct temporal roles of SPOC1 during the HCMV replicative cycle. Mechanistically, we observed a highly specific association of SPOC1 with the major immediate-early promoter (MIEP) strongly suggesting that SPOC1 inhibits HCMV replication by MIEP binding and subsequent recruitment of heterochromatin building factors. Thus, our data add SPOC1 as a novel factor to the endowment of a host cell to restrict cytomegalovirus infections.
  • The chromatin remodeling factor SPOC1 acts as a cellular restriction factor against human cytomegalovirus by repressing the major immediate-early promoter.

    Reichel, Anna; Stilp, Anne-Charlotte; Scherer, Myriam; Reuter, Nina; Lukassen, Sören; Kasmapour, Bahram; Schreiner, Sabrina; Cicin-Sain, Luka; Winterpacht, Andreas; Stamminger, Thomas; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2018-05-09)
    The cellular protein SPOC1 (survival time-associated PHD finger protein in ovarian cancer 1) acts as a regulator of chromatin structure and DNA damage response. It binds H3K4me2/3 containing chromatin and promotes DNA condensation by recruiting corepressors such as KAP-1 and H3K9 methyltransferases. Previous studies identified SPOC1 as a restriction factor against human adenovirus (HAdV) infection that is antagonized by E1B-55K/E4orf6-dependent proteasomal degradation. Here, we demonstrate that, in contrast to HAdV-infected cells, SPOC1 is transiently upregulated during the early phase of HCMV replication. We show that expression of the immediate-early protein 1 (IE1) is sufficient and necessary to induce SPOC1. Additionally, we discovered that during later stages of infection SPOC1 is downregulated in a GSK-3β-dependent manner. We provide evidence that SPOC1 overexpression severely impairs HCMV replication by repressing the initiation of viral immediate early (IE) gene expression. Consistently, we observed that SPOC1-depleted primary human fibroblasts displayed augmented initiation of viral IE gene expression. This occurs in a MOI-dependent manner, a defining hallmark of intrinsic immunity. Interestingly, repression requires the presence of high SPOC1 levels at the start of infection while a later upregulation had no negative impact suggesting distinct temporal roles of SPOC1 during the HCMV replicative cycle. Mechanistically, we observed a highly specific association of SPOC1 with the major immediate-early promoter (MIEP) strongly suggesting that SPOC1 inhibits HCMV replication by MIEP binding and subsequent recruitment of heterochromatin building factors. Thus, our data add SPOC1 as a novel factor to the endowment of a host cell to restrict cytomegalovirus infections. Accumulating evidence indicates that during millennia of co-evolution host cells have developed a sophisticated compilation of cellular factors to restrict cytomegalovirus infections. Defining this equipment is important to understand cellular barriers against viral infection and to develop strategies to utilize these factors for antiviral approaches. So far, constituents of PML nuclear bodies and the interferon gamma inducible protein 16 (IFI16) were known to mediate intrinsic immunity against HCMV. In this study, we identify the chromatin modulator SPOC1 as a novel restriction factor against HCMV. We show that pre-existing high SPOC1 protein levels mediate a silencing of HCMV gene expression via a specific association with an important viral -regulatory element, the major immediate-early promoter. Since SPOC1 expression varies between cell-types, this factor may play an important role in the tissue-specific defense against HCMV.

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